Biodegradation of High-Molecular-Weight PAHs in Polluted Mangrove Sediments Using Indigenous Microflora and Exogenous Strains Rhodococcus erythropolis and Bacillus subtilis

Biodegradation of High-Molecular-Weight PAHs in Polluted Mangrove Sediments Using Indigenous Microflora and Exogenous Strains Rhodococcus erythropolis and Bacillus subtilis | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Biodegradation of High-Molecular-Weight PAHs in Polluted Mangrove Sediments Using Indigenous Microflora and Exogenous Strains Rhodococcus erythropolis and Bacillus subtilis Firmin Semboung Lang, Laurette Ngo Nkot This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8520850/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 23 Feb, 2026 Read the published version in Biodegradation → Version 1 posted 15 You are reading this latest preprint version Abstract Mangroves are highly productive coastal ecosystems, yet they are heavily exposed to pollution by polycyclic aromatic hydrocarbons (PAHs), particularly high-molecular-weight compounds such as pyrene and fluoranthene. These recalcitrant contaminants exhibit high toxicity and persist in the environment, threatening microbial biodiversity and the ecological stability of sediments. This study aims to assess the biodegradation potential of pyrene and fluoranthene in contaminated mangrove sediments by comparing the efficiency of the indigenous microflora with that of exogenous strains (Rhodococcus erythropolis and Bacillus subtilis), and to optimize degradation through bioaugmentation and biostimulation using nutrient amendments. Sediments were artificially spiked with 10,000 mg·kg⁻¹ of pyrene or fluoranthene. Microcosms under sterile and non-sterile conditions were established to evaluate, over five weeks, the biodegradation performance of the endogenous bacterial consortium, exogenous strains, biostimulation (compost), and their combinations. The results revealed significant PAH degradation by the indigenous microflora (45–52% after five weeks). Exogenous strains enhanced degradation rates, reaching 58% for B. subtilis and 63% for R. erythropolis . The combined application of bioaugmentation and biostimulation yielded the highest degradation levels, with 75% for the endogenous consortium and up to 78% for R. erythropolis . Statistical analyses confirmed that these differences were significant compared with sterile and non-sterile controls. The synergistic exploitation of indigenous microflora and exogenous strains, combined with nutrient amendments, constitutes an effective strategy for the bioremediation of mangrove sediments contaminated with high-molecular-weight PAHs. These findings provide a robust foundation for developing pollution-control technologies adapted to tropical coastal ecosystems. Biodegradation Fluoranthene Hydrocarbons Mangroves Microorganisms Pollution Pyrene Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 1. Introduction Mangroves, composed mainly of woody plants, are among the most productive marine ecosystems, distributed along tropical and subtropical coastlines (Kathiresan and Bingham, 2001 ; Semboung et al. 2014 ). Situated at the interface of marine, freshwater, and terrestrial environments, they play a major ecological role by providing diverse ecosystem services: protection against coastal erosion, nursery grounds for numerous aquatic species, and the capacity to degrade certain contaminants (Alongi, 2002 ; Duke et al. 2007 ). Their functioning is strongly influenced by factors such as salinity, organic matter content, and tidal fluctuations, which promote remarkable biological diversity. Mangroves are therefore considered hotspots of microbial biodiversity (Thatoi et al. 2013 ), with microorganisms playing a central role in biogeochemical cycles and ecosystem stability. However, due to high exposure to anthropogenic activities, these environments are increasingly contaminated by polycyclic aromatic hydrocarbons (PAHs) (Zhao et al. 2012 ; Souza et al. 2018 ). This pollution represents a major threat to the ecological stability of mangroves (Brito et al. 2009 ; dos Santos et al. 2011 ; Andreote et al. 2012 ). Petroleum hydrocarbons and their derivatives have toxic effects on flora and fauna, with impacts ranging from acute to chronic. Low-molecular-weight aromatic compounds generally cause acute toxicity, whereas PAHs, due to their high recalcitrance, low solubility, and elevated toxicity, lead to particularly concerning chronic contamination (Mukherjee and Chattopadhyay 2017 ; Juhasz and Naidu 2000 ; Kanaly and Harayama 2010 ; Crampon et al. 2014 ). To date, most studies have focused on the biodegradation of low-molecular-weight PAHs, which are more accessible and more readily degradable (Muckian et al. 2007 ; de Menezes et al. 2012 ; Jiang et al. 2017 ; Bacosa and Inoue 2015 ). However, some studies have also explored the degradation of high-molecular-weight PAHs and the responses of microbial communities to these compounds (Tauler et al. 2016 ; Zhu et al. 2016 ). Bioremediation has emerged as a promising strategy to mitigate the effects of petroleum hydrocarbons in contaminated ecosystems, owing to its efficiency, low cost, and environmental compatibility. It relies primarily on the activity of microorganisms capable of degrading these pollutants, while interacting with other members of the microbial community (Fuentes et al. 2016 ). Hydrocarbon biodegradation has been investigated in various contaminated environments, including soils (Sun et al. 2015 ), oceans (Sauret et al. 2014 ), and mangroves (Jiang et al. 2013 ; Muangchinda et al. 2013 ). Although several studies have demonstrated the ability of indigenous bacteria from mangrove sediments to degrade hydrocarbons, including PAHs (Brito et al. 2009 ; Li et al. 2009 ; Tian et al., 2008 ), additional information is needed on their biodegradation rates and removal efficiencies to optimize bioremediation technologies. Several hydrocarbonoclastic bacterial genera such as Rhodococcus , Acinetobacter , Pseudomonas , Alcanivorax , and Sphingomonas have been isolated from polluted mangrove sediments (Brito et al. 2006 ; Rocha et al. 2013 ; Yu et al. 2005 ). Moreover, advances in high-throughput sequencing technologies (Illumina MiSeq, Illumina HiSeq, 454 GS FLX) have enabled a deeper understanding of microbial community structures and dynamics in various contaminated environments, including soils (Abbasian et al. 2016 ), Arctic soils (Tan et al. 2013 ), marine sediments (Mason et al. 2014 ), and mangrove sediments (Andreote et al. 2012 ; dos Santos et al. 2011 ). In the present study, sediments were artificially contaminated with two representative high-molecular-weight PAHs—pyrene and fluoranthene—commonly used in microcosm experiments to assess microbial responses to hydrocarbon pollution (Sauret et al. 2014 ; Schurig et al. 2014 ; Schwarz et al. 2017 ). The main objective of this work is to evaluate the biodegradation potential of pyrene and fluoranthene in contaminated mangrove sediments by comparing the endogenous microflora with exogenous strains ( Rhodococcus erythropolis and Bacillus subtilis ). Specifically, the aims are to (i) characterize the competent indigenous microflora capable of degrading these compounds, (ii) assess the degradation performance of the exogenous strains under controlled microcosm conditions, and (iii) optimize biodegradation through bioaugmentation and microbial stimulation using nutrient amendments (nitrogen, phosphorus, compost). This study will contribute to expanding knowledge on the impact of PAHs on microbial communities in mangrove sediments and on developing bioremediation approaches adapted to these sensitive ecosystems. 2. Materials and Methods 2.1 Materials Mangrove sediments The mangrove sediments used in this study were collected from the Wouri estuary mangroves in Cameroon, a heavily industrialized zone. Solid and liquid wastes from various industries are discharged into the mangroves without prior treatment. Sediments were collected at depths between 20 and 40 cm. This area harbors abundant microflora and is likely to contain pollutants. Microorganisms Two categories of microorganisms were used: indigenous microorganisms and exogenous microorganisms. Indigenous microorganisms were used as a bacterial consortium after isolation, identification, and characterization. Exogenous microorganisms consisted of two pure strains, Rhodococcus erythropolis and Bacillus subtilis . R. erythropolis was obtained from the Bioindustries Laboratory of Gembloux Agro-Bio Tech, University of Liège, and the B. subtilis strain was obtained from the Biology Laboratory of the University of Mons, Belgium. Polycyclic aromatic hydrocarbons : Pyrene and Fluoranthene PAHs are non-polar, hydrophobic compounds that do not ionize; consequently, their solubility in water is very low. In this study, two PAHs—pyrene (Pyr) and fluoranthene (Flt)—were used (purity > 97%, supplied by UCB, Belgium). Pyrene contains four benzene rings, with a molecular mass of 202.26 and a logKow of 5.18. Fluoranthene also contains four benzene rings, with a molecular mass of 178.24 and a logKow of 4.18. 2.2 Methods 2.2.1 Artificial contamination of sediments with pyrene and fluoranthene Sediments were artificially contaminated following the method described by Tam et al. (2008), with slight modifications. In a flask, 10,000 mg·kg⁻¹ dry weight of each PAH (pyrene and fluoranthene) was dissolved in 50 mL acetone and mixed with 200 g of mangrove sediments. The mixture was then combined with 1 kg of mangrove sediment and homogenized with a plastic spatula. The flask was left open for two days to allow acetone evaporation and PAH adsorption into the sediments. The resulting sediment concentration was approximately 10,000 mg·kg⁻¹ dry weight. Then, 250 mL of artificial seawater (salinity 10‰) was added to moisten the mixture. 2.2.2 Isolation of competent microflora and production of the indigenous bacterial consortium Isolation was performed following Weekers et al. (1999) with modifications. One gram of polluted sediment was placed in 9 mL sterile peptone water and heated at 80 °C for 15 min to select thermoresistant microflora. Serial dilutions were plated on selective mineral medium and incubated at 35 °C for 72 h. Colonies were suspended (10⁴ CFU·mL⁻¹) in 100 mL sterile rich liquid medium containing 10 g·L⁻¹ diesel (preculture 1; 30 °C, shaking, 72 h). This preculture inoculated 500 mL of identical medium (preculture 2), incubated under the same conditions. Preculture 2 was transferred into a 5-L bioreactor containing 4 L rich liquid medium + 10 g·L⁻¹ diesel. Growth was monitored by OD₆₀₀ over 96 h. Biomass was recovered by centrifugation (3500 rpm, 15 min) and stored at 0 °C with 2.5% (v/v) glycerol and 10% (v/v) maltodextrin as cryoprotectants. CFU·g⁻¹ and biomass yield were determined. This selection procedure was chosen because it enriches hydrocarbonoclastic and stress-resistant bacterial populations—typical traits of effective hydrocarbon degraders in mangrove ecosystems. Media compositions : Peptone water (g·L⁻¹): casein peptone 10, NaCl 5, Na₂HPO₄ 9, K₂HPO₄ 1.5; pH 7.0 ± 0.2 Rich medium (g·L⁻¹): glucose 20, casein peptone 10, yeast extract 10, agar 15; pH 7.0 ± 0.2 Selective mineral medium (mg·L⁻¹): (NH₄)₂SO₄ 1000, K₂HPO₄ 800, KH₂PO₄ 200, MgSO₄.7H₂O 200, CaCl₂.2H₂O 100, FeSO₄.7H₂O 12, MnSO₄.7H₂O 3, ZnSO₄.7H₂O 3, CoSO₄.7H₂O 1, (NH₄)₆Mo₇O₂₄.4H₂O 1, NaCl 4, agar 15 g, diesel 10 ppm; pH 7.0 ± 0.2. Diesel served as the sole carbon source. 2.2.3 Characterization and identification of the bacterial consortium using API 20E Characterization included: Gram staining : observation after crystal violet (1 min), Lugol iodine (1 min), ethanol decolorization (20–50 s) and fuchsin counterstain (1 min). Violet cells = Gram-positive; pink cells = Gram-negative. Catalase test : culture in exponential phase mixed with H₂O₂. Bubble formation (O₂) = positive (aerobic bacteria). Oxidase test : colony applied to filter paper impregnated with oxidase reagent. Purple coloration = positive. API 20E identification : a pure culture was suspended in physiological saline (0.5 mL), introduced into the API 20E strip, incubated at 37 °C for 24 h. After addition of the auxiliary reagent, enzymatic reactions were interpreted using the API 20E reading table to determine the biochemical profile and bacterial species. The combination of Gram staining, catalase/oxidase tests, and API 20E biochemical profiling was selected because it covers both fundamental morphological traits and the key metabolic capabilities involved in hydrocarbon degradation (oxidative metabolism, organic acid production, nitrogen metabolism). 2.2.4 Assessment of the biodegradation potential of Pyrene and Fluoranthene 2.2.4.1 Experimental setup Sterile conditions (Sterile Sediments “SS”) The sterile-condition experiment assessed the ability of the indigenous bacterial consortium to degrade pyrene and fluoranthene and compared its degradation rates with those of the exogenous strains R. erythropolis and B. subtilis . Two separate experiments were conducted: one with pyrene and one with fluoranthene. In a set of three flasks, 1000 g of mangrove sediments previously polluted with pyrene at 10,000 mg·kg⁻¹ dry weight were placed. Then, 500 mL seawater was added to keep sediments moist. All three flasks were autoclaved to ensure sterilization. After sterilization, the flasks were placed on the laboratory bench for a 5-week follow-up. These three flasks constituted the control (T0), as autoclaving eliminated all microbial life capable of degrading hydrocarbons. A second set of three flasks (T1) was prepared identically, with the addition of the indigenous bacterial consortium at 10⁷ CFU·g⁻¹ dry weight. A third set (T2) received Bacillus subtilis at 10⁷ CFU·g⁻¹. A fourth set (T3) received Rhodococcus erythropolis at 10⁷ CFU·g⁻¹. The same design was applied to fluoranthene-polluted sediments. Non-sterile conditions (Non-Sterile Sediments “NSS”) The non-sterile experiment evaluated various biodegradation strategies—natural attenuation, biostimulation, bioaugmentation, and combined biostimulation + bioaugmentation—using mangrove sediments polluted with either pyrene or fluoranthene. For natural attenuation (T1), relying solely on native degrading microorganisms, 3000 g of pyrene-polluted mangrove sediments were placed into three PVC microcosm jars kept in a greenhouse. For biostimulation (T2), compost was added at 30% of the sediment mass to stimulate microbial activity. For bioaugmentation, three sets were prepared: T3: indigenous bacterial consortium at 10⁷ CFU·g⁻¹ ; T4: Bacillus subtilis at 10⁷ CFU·g⁻¹ ; T5: Rhodococcus erythropolis at 10⁷ CFU·g⁻¹. Combined biostimulation and bioaugmentation consisted of: T6: compost + indigenous consortium ; T7: compost + Bacillus subtilis ; T8: compost + Rhodococcus erythropolis. T0 represented sterile sediments polluted with pyrene without microbial addition. The same design was applied to fluoranthene-polluted sediments. 2.2.4.1 pH monitoring under sterile and non-sterile conditions Throughout the experiment, pH was measured weekly using a HANNA HI 207 pH-meter. pH monitoring provides information on alkalinity and acidity, as pH is a limiting factor in biodegradation. 2.2.4.2 Monitoring of competent microflora under sterile conditions Enumeration of competent microflora in sterile sediments was performed by serial dilution and plating on Petri dishes. One gram of sediment was collected weekly from each flask and placed in a test tube containing 9 mL peptone water. Spreading was performed on rich solid medium. Each dilution was plated in triplicate. After 3 days of incubation at 30 °C, colony-forming units (CFUs) were counted and the mean value for each dilution determined. 2.2.4.3 Determination of total hydrocarbons under sterile and non-sterile conditions PAHs were extracted from the sediments every 7 days over a period of 5 weeks. The extraction procedure followed high-performance liquid chromatography (HPLC) analysis using an Agilent 1100 Series system, as described by Wannoussa et al. (2015), with slight modifications. One gram of wet sediment sample was placed in 18 mL glass tubes, and an equivalent weight of anhydrous sulfate along with 10 mL of hexane were added. The mixture was homogenized by vortexing and placed in an ultrasonic bath for 1 h. The tubes were then placed on a horizontal shaker protected from light for 16 h, and the organic phase was transferred into 15 mL Falcon tubes. After centrifugation and transfer into a new tube, the residue was extracted again with 10 mL of hexane and transferred into a glass flask. Hexane was evaporated using a rotary evaporator (BUCHI ROTAVAPOR R-200) heated to 55 °C, and the dry extract was reconstituted in 10 mL of methanol. Subsequently, 1 mL of this solution was placed in an HPLC vial, sealed with a septum, and crimped. Samples were then sent for HPLC analysis. HPLC analyses were performed using an Agilent 1100 Series system equipped with a C18 column (LiChroCART® 250–4.6 HPLC cartridge Purospher® STAR RP-18 endcapped 5 μm, Merck, Germany) maintained at 30 °C. 2.2.5 Statistical analyses Mean values were compared using analysis of variance (ANOVA) with a significance level of p ≤ 0.05. Differences between the various hydrocarbon reduction rates were also evaluated using the same test. ANOVA was used to assess differences between initial and final total petroleum hydrocarbon (TPH) concentrations in the treated flasks compared with the control flasks. All statistical analyses were performed using MINITAB 17® software (French version). 3. Results The present study aimed to evaluate the biodegradation potential of pyrene and fluoranthene present in contaminated mangrove sediments, using microorganisms including the indigenous microflora, represented here by the consortium, and exogenous strains such as Rhodococcus erythropolis and Bacillus subtilis . The results are presented in three sections: the first focuses on the characterization of the consortium, the second on experiments conducted under sterile conditions, and the third on experiments carried out under non-sterile conditions. 3.1. Characterization and identification of bacterial isolates 3.1.1 Total and Hydrocarbon-Degrading Microflora Enumeration of bacterial populations revealed that the polluted mangrove sediments of the Wouri estuary contained a total microflora estimated at 3 × 10⁸ CFU·g⁻¹ of sediment. Within this community, the hydrocarbon-degrading (competent) microflora represented approximately 1.5 × 10⁵ CFU·g⁻¹ of sediment. This indicates that only a small fraction of the indigenous bacterial population is directly involved in hydrocarbon degradation. 3.1.2 Microscopic Characteristics Microscopic examination of the bacterial isolates provided information on their motility, cell size, morphology, and cellular arrangement. These observations constitute preliminary criteria for the differentiation of the strains included in the starter consortium. Microscopic examination revealed that the bacterial isolates exhibited diverse morphotypes, including cocci, bacilli, and coccobacilli. All isolates were found to be motile, showed an isolated growth arrangement, and tested Gram-positive (Table 2 ). 3.1.3 Biochemical characteristics and API 20E identification tests Biochemical analyses performed with API 20E galleries provided insights into the metabolic profiles of the consortium isolates. These tests highlighted their energetic, carbohydrate, and protein metabolism, and enabled preliminary identification at the presumptive taxonomic level (Table 1 ). Table 1 Biochemical characteristics of the bacterial isolates Reactions / Traits Bacterial isolate 1 Bacterial isolate 2 Baterial isolate 3 Bacterial isolate 4 Energy metabolism Oxidase + + - + Catalase + - - + Respiratory type Aerobic Anaerobic Anaerobic Aerobic Carbohydrate metabolism TSI (Glucose) - + + + Lactose / Sucrose + + + + H₂S production - - - - Gelatinase (GEL) + + + + Arabinose (ARA) - - - - ONPG (β-galactosidase) - - - - Citrate utilization + - - - Motility + + + + Mannitol - + - + Voges-Proskauer (VP) + + - + Methyl Red (MR) - - - - Protein metabolism ADH (Arginine dihydrolase) - - - - LDC (Lysine decarboxylase) - - - - ODC (Ornithine decarboxylase) - - - - Urease - - - - Indole - - - - TDA (Tryptophan deaminase) - - - - Presumptive identification of isolates Bacterial isolate 1 (Oxidase +, Catalase +, Aerobic, Citrate +, VP +, Motile) → consistent with Bacillus sp. Bacterial isolate 2 (Oxidase +, VP +, Mannitol +, Anaerobic) → possible Enterococcus sp. or related Gram-positive bacteria. Bacterial isolate 3 (Oxidase –, Anaerobic, Glucose +, Motile) → may correspond to Clostridium sp. or related anaerobes. Bacterial isolate 4 (Oxidase +, Catalase +, Aerobic, Mannitol +, VP +, Motile) → compatible with Pseudomonas sp. Table 2 Biochemical and morphological characterization of bacterial isolates recovered from hydrocarbon-contaminated mangrove sediments Isolate Morphology (shape) Motility Arrangement Gram Probable bacterial genus Reported isolation in hydrocarbon-polluted mangroves Bacterial isolate 1 Coccus Motile Single + Bacillus sp. Bacillus spp. frequently reported as hydrocarbon-degraders in mangrove sediments (Das et al. 2014; Dias et al. 2012) Bacterial isolate 2 Coccus Motile Single + Enterococcus sp. (or related Gram-positive bacteria) Enterococcus spp. isolated from contaminated mangroves (Taylor et al. 2012; Kathiresan and Bingham 2001 ) Bacterial isolate 3 Bacillus Motile Single + Clostridium sp. (anaerobe) Clostridium spp. associated with anaerobic hydrocarbon degradation in mangroves (Peixoto et al. 2011; Reddy and Dhanasekaran 2016) Bacterial isolate 4 Coccobacillus Motile Single + Pseudomonas sp. Pseudomonas spp. widely documented as hydrocarbon-degrading bacteria in mangroves (Sathe et al. 2014; Serrano et al. 2021) 3.2 Experiments under sterile conditions 3.2.1 pH evolution over 5 weeks pH evolution in sterile sediments contaminated with pyrene The pH changes during the 5-week incubation indicate a progressive acidification of the sediments in all microbial treatments, unlike the T0 control (pH = 6.07 → 6.67), which remained stable or showed a slight increase. This acidification reflects the production of organic acids resulting from pyrene biodegradation. Among the treatments, Rhodococcus erythropolis (T3: pH 6.23 → 4.53) and especially its combination with compost (T6: pH 6.00 → 3.60) exhibited the most pronounced pH decreases, reaching 4.53 and 3.60, respectively, indicating high metabolic activity and enhanced pyrene degradation. In contrast, Bacillus subtilis (T2: pH 6.87 → 5.64) and its association with compost (T5: pH 6.58 → 5.03) showed a more moderate acidification. The positive effect of compost was particularly notable for the bacterial consortium (T4: pH 6.10 → 4.93), which outperformed the consortium alone (T1: 6.33 → 5.27). Overall, treatment T6 appears to be the most effective for pyrene degradation in sterile sediments. pH evolution in sterile sediments contaminated with fluoranthene The pH evolution during fluoranthene biodegradation shows a progressive decrease for all microbial treatments (T1–T6), whereas the T0 control (pH 6.67 → 6.87) remained stable around 6.7. This acidification results from the production of organic acids, reflecting metabolic activity associated with fluoranthene degradation. Treatments involving R. erythropolis (T3 and T6) exhibited the most significant pH decreases, reaching 4.73 and 3.81, respectively, indicating a strong fluoranthene biodegradation capacity. Compost addition notably enhanced microbial activity, particularly for R. erythropolis , making T6 (pH 6.12 → 3.81) the most effective treatment. In contrast, B. subtilis and the bacterial consortium induced moderate acidification, reflecting a lower degree of fluoranthene degradation. 3.2.2 Microflora dynamics in the flasks Microbial growth during pyrene biodegradation in sterile sediments Figure 3 illustrates microbial growth in sterile sediments contaminated with pyrene across different treatments (T0: sterile sediments, no inoculum; T1–T3: sediments + pyrene + bacterial inoculum; T4–T6: sediments + pyrene + compost + inoculum). The aim was to evaluate the ability of different inocula (consortium, B. subtilis , R. erythropolis ) with or without compost to colonize and potentially degrade pyrene. In the T0 negative control, bacterial populations ranged from 0 to 1.6 × 10⁻¹ CFU·g⁻¹ dry weight, with no significant microbial development, as expected in sterile conditions. The slight final increase likely reflects minor environmental contamination. Compost strongly stimulated microbial growth in sterile pyrene-contaminated sediments. R. erythropolis was the most adapted species, while B. subtilis performed moderately well. Surprisingly, the bacterial consortium was the least effective, likely due to poor adaptation to pyrene, internal competition among strains, or nutrient limitations in sterile sediment. Treatment T6 (Compost + R. erythropolis ) was the most effective, combining nutrient supplementation via compost with a hydrocarbon-specialist bacterium, making it the optimal candidate for pyrene bioremediation in sterile or nutrient-poor sediments. Microbial growth during fluoranthene biodegradation in sterile sediments Similarly, Fig. 4 shows microbial growth (CFU or cell counts) over 5 weeks across different treatments: T0: control (no inoculum), T1–T3: without compost, T4–T6: with compost; inocula: consortium, B. subtilis , R. erythropolis . Fluoranthene is slightly less toxic than pyrene for many bacteria, explaining some observed differences. In T0 (sterile sediments + fluoranthene), competent microflora ranged from 0 → 10² cells, consistent with the absence of growth in sterile conditions. Minor increases (~ 10²) reflect negligible contamination. For T1 (fluoranthene + bacterial consortium), the population declined from 2.6 × 10⁷ to 4.2 × 10⁶ by the end, reflecting poor adaptation in sterile sediments. For T2 ( B. subtilis ), populations increased from 2.5 × 10⁷ → 6 × 10⁸ cells, performing better than on pyrene. T3 ( R. erythropolis ) showed strong growth from 2.3 × 10⁷ → 9.6 × 10⁸ CFU·g⁻¹, confirming its powerful HAP-degrading capacity. Compost slightly improved consortium performance in T4 (2.7 × 10⁷ → 6 × 10⁷ CFU·g⁻¹). T5 (Compost + B. subtilis ) showed significant growth (2.7 × 10⁷ → 2.3 × 10⁹ CFU·g⁻¹), and T6 (Compost + R. erythropolis ) exhibited the highest growth (2.9 × 10⁷ → 3.6 × 10⁹ CFU·g⁻¹). Overall, fluoranthene was less inhibitory than pyrene. All treatments except T1 exhibited substantial growth. Compost acted as a nutrient amendment (C, N, P), providing microhabitats, stabilizing pH, and enhancing HAP bioavailability, leading to 3–10 fold microbial growth increases. R. erythropolis , a hydrophobic Gram-positive bacterium capable of producing biosurfactants, efficiently degraded four-ring HAPs such as fluoranthene, confirming its key role in bioremediation. 3.2.3 Pyrene and fluoranthene degradation rates in sterile mangrove sediments Pyrene degradation rates Figure 5 shows that pyrene degradation percentages significantly increased over 5 weeks in all treatments. The sterile control (T0) exhibited very low values (1.13 ± 0.72% at week 1 to 2.36 ± 1.83% at week 5), confirming the absence of biological activity and indicating negligible abiotic oxidation. Natural attenuation (T1) in non-sterile sediments showed degradation rates from 6.19 ± 1.28% to 36.54 ± 1.96%, demonstrating the capacity of indigenous microflora to metabolize pyrene. Compost biostimulation (T2) enhanced activity further, reaching 42.55 ± 5.89%. Bioaugmentation with the bacterial consortium (T3: 50.38 ± 5.5%), B. subtilis (T4: 44.18 ± 3.5%), and R. erythropolis (T5: 57.59 ± 1.65%) improved degradation, particularly for R. erythropolis , known for HAP degradation via specific metabolic pathways (e.g., catechol dioxygenases). The bacterial degradation gradient was: Rhodococcus > Consortium > B. subtilis . Combined bioaugmentation + biostimulation treatments were most effective: T6 (Compost + Consortium) 70.13 ± 3.31%, T7 (Compost + B. subtilis ) 62.33 ± 7.29%, and T8 (Compost + R. erythropolis ) 85.58 ± 5.26%, showing strong synergy between nutrient enrichment and inoculation with a specialized strain. Fluoranthene degradation rates Figure 6 presents fluoranthene degradation (%) in sterile sediments over 5 weeks. The T0 control exhibited very low abiotic degradation (3.68 ± 2.08%), indicating low bioavailability without microbial activity, typical for four-ring HAPs. In T1 (consortium), degradation increased from 7.38 ± 3.72% to 37.77 ± 2.09%, reflecting the consortium's bioaugmentation effect. Exogenous bacteria performed better: T2 ( B. subtilis ) 11.84 ± 3.37% → 48.03 ± 6.53%, T3 ( R. erythropolis ) 19.18 ± 2.5% → 60.57 ± 4.5%, demonstrating that isolated strains degraded fluoranthene more efficiently than the consortium. R. erythropolis ’s effectiveness is due to dioxygenases, resistance to toxic compounds, and ability to use HAPs as carbon sources. Compost biostimulation improved degradation in T4 (14.54 ± 1.34% → 50.28 ± 2.25%), T5 (19.06 ± 1.36% → 58.21 ± 4.9%), and T6 (23.30 ± 1.52% → 74.65 ± 5.13%), acting as a nutrient source (C, N, P), enhancing fluoranthene bioavailability and stimulating microbial growth. The degradation efficiency ranking after five weeks was: T0 < T1 < T2 < T4 < T5 < T3 B. subtilis > Consortium. Compost addition further amplified degradation by improving bioavailability, providing nutrients and organic matter, stimulating degradation enzyme expression, and enhancing microbial growth. Combined bioaugmentation + biostimulation (T6) was optimal. 3.2.4 Mass balance of pyrene and fluoranthene degradation in sterile and non-sterile sediments Tables 3 and 4 present the mass balance of pyrene and fluoranthene in sterile and non-sterile sediments after five weeks under different bioremediation strategies. Across all treatments, the low losses in the sterile control (T0) – below 4% for both HAPs – indicate that abiotic processes (volatilization, irreversible adsorption, photodegradation) contributed minimally. Therefore, losses observed in other treatments can be primarily attributed to microbial activity. In sterile sediments (Table 3 ), the introduction of exogenous microorganisms significantly increased HAP degradation compared with the control. Among the tested strains, R. erythropolis performed best, achieving 45.07% and 60.58% losses for pyrene and fluoranthene, respectively, in the absence of compost (T3). This is consistent with its known capacity to metabolize high-molecular-weight hydrocarbons via broad-spectrum dioxygenases and effective biosurfactant production. B. subtilis and the bacterial consortium also enhanced degradation, albeit at lower levels (28–48% depending on the compound). Compost addition generally boosted exogenous bacterial activity, particularly for R. erythropolis , which reached 59.56% and 74.66% losses for pyrene and fluoranthene, respectively (T6), indicating a biostimulation effect attributable to nutrient supply, improved HAP bioavailability, and concomitant stimulation of microbial growth. Table 3 : Mass balance of PAHs amount remained (mg.kg -1 ) and percentages in each fraction to total PAHs (pyrene and fluoranthene) added (input) in sterile sediments after 5 weeks of degradation Different biodegradation strategies Mangrove Sterile sediments Fate of PAHs Pyrene Fluoranthene Amounts (mg.kg -1 ) % Input Amounts (mg.kg -1 ) % Input Sterile Sediments (T0) Input In sediment phase Losses 10,088.33 ± 62.51 9,850 ± 130 338.33 ± 187.1 97.63 3.83 ±1.83 p 10,036 ± 58.6 9,666.66 ± 152.75 369.33 ± 211.34 96.32 3.68 ± 2.08 st Sterile Sediments + Bacterial Consortium (T1) Input In sediment phase Losses 10,161.33 ± 133.83 7233.33 ± 152.75 2,927.66 ± 25.42 71.18 28.8 ± 0.58 fg 10,070.33 ± 35.47 6,266.54 ± 208.16 3,803.66 ± 213.91 62.23 37.77 ± 2.1 ghi Sterile Sediments + B. subtilis (T2) Input In sediment phase Losses 10117.67 ± 64.3 6300 ± 300 3817.66 ± 162.69 62.27 37.73 ± 2.5 cd 10,134 ± 74.11 5,266.66 ± 665.83 4,867.33 ± 656.9 51.97 48.03 ± 6.53 cde Sterile Sediments + R. erythropolis (T3) Input In sediment phase Losses 10073.66 ± 45.5 5533.33 ± 208.2 4540.33 ± 162.7 54.93 45.07 ± 1.82 b 10,146 ± 113.33 4000 ± 500 6,146 ± 387.1 39.42 60.58 ± 4.5 b Sterile Sediments + Compost + Bacterial Consortium (T4) Input In sediment phase Losses 10,102 ± 132.04 6,683.33 ± 160.72 3,418.33 ± 288.56 66.15 33.85 ± 2.41 de 10,258.66 ± 264.2 5,100 ± 360.55 5,158.66 ± 116.21 48.71 51.39 ± 2.25 cd Sterile Sediments + Compost + B. subtilis (T5) Input In sediment phase Losses 10,066.33 ± 170 5,583.33 ± 550.75 4,483 ± 529.96 55.47 44.53 ± 5.24 b 10,130.66 ± 160.05 4,233.33 ± 550.75 5,897.33 ± 461.86 41.78 58.22 ± 4.99 b Sterile Sediments + Compost + R. erythropolis (T6) Input In sediment phase Losses 10,099 ± 227.51 4,083.33 ± 381.88 6,015.66 ± 335.92 40.44 59.56 ± 3.43 a 10,127 ± 163.64 2,566.66 ± 5.3.32 7,560.33 ± 567.72 25.34 74.66 ± 5.13 a In non-sterile sediments (Table 4 ), the presence of indigenous microflora significantly enhanced HAP degradation, even in the absence of external inputs, as demonstrated by natural attenuation (T1) with losses ranging from 30 to 36%. Compost addition alone (biostimulation) further improved this process, achieving losses of 40–42%, indicating a positive response of the native communities to nutrient enrichment. Bioaugmentation with isolated strains led to additional improvements, with R. erythropolis remaining the most effective strain (57.59% for pyrene and 73.91% for fluoranthene). The highest levels of dissipation were achieved when bioaugmentation was combined with biostimulation. In particular, the treatment combining R. erythropolis and compost (T8) resulted in near-complete HAP removal, with 85.58% loss for pyrene and 94.16% for fluoranthene. These results demonstrate a synergistic effect between the introduced bacteria, the indigenous microflora, and compost, leading to enhanced degradation capacity, likely through metabolic complementarity, improved HAP solubilization, and intensified microbial functional diversity. Overall, the results clearly show that HAP biodegradation in sediments strongly depends on the availability of specialized microorganisms and the organic enrichment of the substrate. Although fluoranthene is more recalcitrant than pyrene, it was degraded more extensively under combined treatments, indicating that increased bioavailability via compost and microbial cooperation is critical for the breakdown of highly hydrophobic compounds. The most effective strategy is unequivocally the bioaugmentation–biostimulation combination using R. erythropolis and compost in non-sterile sediments. This approach maximizes degradation capabilities by simultaneously leveraging the strengths of the indigenous microflora and exogenous high-metabolic-potential strains. These findings demonstrate that the simultaneous integration of bioaugmentation and biostimulation constitutes a robust and highly effective approach for remediating sediments contaminated with high-molecular-weight HAPs. This strategy therefore holds considerable potential for in situ bioremediation applications in coastal and port ecosystems, where the persistence of HAPs represents a major environmental concern. Table 4 : Mass balance of PAHs amount remained (mg.kg -1 ) and percentages in each fraction to total PAHs (pyrene and fluoranthene) added (input) in nonsterile sediments after 5 weeks of degradation under different techniques Biodegradation Techniques Differents biodegradation strategies Mangrove non sterile sediments Fate of PAHs Pyrene Fluoranthene Amounts (mg.kg -1 ) % Input Amounts (mg.kg -1 ) % Input Sterile control Sterile Sediments (T0) Input In sediment phase Losses 10,088.33 ± 62.51 9,850 ± 130 238.33 ± 187.1 97.64 2.36 ± 1.84 yz 10,036.1 ± 58.46 9,666.66 ± 152.75 369.43 ± 211.21 96.32 3.68 ± 2.07 tuv Natural attenuation Non sterile sediments (T1) Input In sediment phase Losses 10,137.67 ± 105.66 6,433.33 ± 152.75 3,704.33 ± 232.3 63.46 36.54 ± 1.96 jkl 10,180.64 ± 134.78 7033.33 ± 450.92 3,147.3 ± 428.09 69.09 30.91 ± 4.25 hijkl Biostimulation Non sterile sediments + Compost (T2) Input In sediment phase Losses 10,095.66 ± 199.13 5,800 ± 700 4,295.66 ± 522.92 57.45 42.54 ± 5.9 hij 10,175.86 ± 109.15 6100 ± 360.55 4,075.86 ± 287.02 59.95 40.05 ± 3.08 fg Bioaugmentation Non sterile sediments + Bacterial consortium (T3) Input In sediment phase Losses 10,143.66 ± 291.56 5,033.33 ± 642.91 5,110.33 ± 487.45 49.62 50.38 ± 5.5 fg 10,189.43 ± 145.41 4,766.66 ± 251.66 5,423.16 ± 345.24 46.78 53.22 ± 2.82 cd Non sterile sediments + B. subtilis (T4) Input In sediment phase Losses 10,151.33 ± 147 5,666.66 ± 288.67 4,484.66 ± 442.03 55.82 44.18 ± 3.58 ghi 10,225.45 ± 232.53 4366.66 ± 585.94 5858.78 ± 775.69 42.7 57.3 ± 6.41 cd Non sterile sediments + R. erythropolis (T5) Input In sediment phase Losses 10,138.33 ± 220.47 4,300 ± 200 5,838.33 ± 199 42.41 57.59 ± 1.65 de 10,221.1± 239.49 2,666.66 ± 663.76 7554.44 ± 524.79 26.09 73.91 ± 6.81 b Bioaugmentation and Biostimulation Non sterile sediments + Compost + Bacterial consortium (T6) Input In sediment phase Losses 10,210.66 ± 207.2 3,050 ± 278.38 7,160 ± 484.85 29.87 70.13 ± 3.31 b 10,354.86 ± 192.53 3,100 ± 529.15 7,254.86 ± 421.36 29.94 70.06 ± 4.76 b Non sterile sediments + Compost + B. subtilis (T7) Input In sediment phase Losses 10,176.66 ± 373.66 3,833.33 ± 702.37 6,343.33 ± 819.03 37.67 62.33 ± 7.3 cd 10,326.51 ± 359.92 2,566.66 ± 513.16 7,759.84 ± 245.93 24.86 75.14 ± 4.24 b Non sterile sediments + Compost + R. erythropolis (T8) Input In sediment phase Losses 10,174.33 ± 206.29 1,466.66 ± 550.75 8,707 ± 515.8 14.42 85.58 ± 5.26 a 10,336.78 ± 410.15 603.33 ± 355.01 9733.45 ± 308.31 5.84 94.16 ± 3.28 a 4. Discussion The results obtained in this study clearly demonstrate the potential for pyrene and fluoranthene biodegradation in contaminated mangrove sediments by mobilizing both the indigenous microflora and specialized exogenous strains such as Rhodococcus erythropolis and Bacillus subtilis . Overall, the observed performances align with microbiological mechanisms previously reported in the literature regarding polycyclic aromatic hydrocarbon (PAH) degradation in coastal and estuarine environments. Composition and potential of the indigenous mangrove microflora The total microflora observed in Wouri sediments (3 × 10⁸ CFU.g⁻¹) is comparable to values reported in other highly anthropized mangrove ecosystems, where microbial loads range from 10⁶ to 10⁹ CFU.g⁻¹ (Kathiresan and Bingham 2001 ; Thatoi et al. 2013 ). The low proportion of microorganisms directly competent for hydrocarbon degradation (≈ 10⁵ CFU.g⁻¹) is also consistent with observations by dos Santos et al. ( 2011 ), who noted that only 0.01–1% of the total mangrove microflora actively participates in PAH mineralization. The identified genera ( Bacillus, Enterococcus, Clostridium, Pseudomonas ) are regularly reported in hydrocarbon-contaminated sediments. Pseudomonas spp. and Bacillus spp. are particularly recognized as robust PAH degraders due to their tolerance to toxic stress, motility, and biosurfactant production (Das and Chandran 2011 ; dos Santos et al. 2021). The presence of Clostridium sp. may facilitate anaerobic degradation pathways, which are particularly relevant in the reducing conditions of mangrove sediments (Li et al. 2015 ). pH dynamics and metabolic activity during biodegradation The pH decrease observed in most treatments, especially with R. erythropolis and the compost + bacterial combinations, represents a well-recognized indicator of metabolic activity associated with HAP degradation. Organic acid production is an expected byproduct of hydrocarbon dioxygenation and catabolic pathways (Atlas and Hazen 2011 ; Cerniglia 1993 ). This acidification is particularly pronounced for four-ring HAPs (pyrene and fluoranthene), whose degradation requires intense enzymatic activity, notably aromatic mono- and dioxygenases (Seo et al. 2009 ). The amplifying role of compost has also been documented: it provides essential nutrients (N, P), increases sediment porosity, and enhances HAP bioavailability by stimulating desorption from solid particles (Sayara and Sánchez 2020 ; Silva-Castro et al. 2016 ). These mechanisms explain the more pronounced pH reductions observed in the combined treatments. Microbial growth and aptitude of exogenous strains Growth profiles confirm the metabolic superiority of R. erythropolis in HAP-contaminated environments. This species is known for its exceptional capacity to degrade complex hydrocarbons, notably through biosurfactant production, which enhances HAP solubility (Franzetti et al. 2010 ), strong hydrophobic surface properties facilitating adhesion to insoluble compounds (Boonchan et al. 2000 ), and a broad enzymatic arsenal including dioxygenases, monooxygenases, and dehydrogenases (Larkin et al. 2005 ). The exponential growth achieved in the compost + R. erythropolis treatment (up to 3.6 × 10⁹ CFU.g⁻¹) is consistent with previous studies showing that organic substrate addition strongly enhances rhodococci proliferation under polluted conditions (Pacwa-Płociniczak et al. 2016 ). Conversely, the low performance of the bacterial consortium may result from negative competitive interactions or poor prior adaptation to HAPs, factors previously reported in non-optimized multi-strain bioaugmentations (Nzila 2016 ). Pyrene and fluoranthene biodegradation performance Pyrene degradation The degradation rates clearly demonstrate the combined efficacy of bioaugmentation and biostimulation. The best performances were observed with R. erythropolis (≈ 57.6 ± 1.65%) and, notably, with the compost + R. erythropolis combination (≈ 85.6 ± 5.26%). Such levels are consistent with other studies using hydrocarbonoclastic Actinobacteria for pyrene degradation (Kim et al. 2018 ; Zhang et al. 2021). The role of the indigenous microflora (≈ 36.5 ± 1.96%) further confirms that mangroves, as hydrocarbon-exposed environments, naturally harbor communities capable of PAH mineralization (Boopathy, 2000 ; Yu et al. 2005 ). Fluoranthene degradation Fluoranthene degradation rates followed similar trends to those of pyrene, with maximum efficiency observed for the compost + R. erythropolis treatment (≈ 74.6 ± 5.13%). This aligns with studies demonstrating that rhodococci can effectively degrade four-ring HAPs due to their high physiological tolerance (Weissenfels et al. 1990 ). The observed hierarchy ( Rhodococcus > Bacillus subtilis > consortium > control) also matches numerous comparative studies on PAH bioaugmentation (Haritash and Kaushik 2009 ; Juhasz and Naidu 2000 ). Bioaugmentation + Biostimulation synergy: an optimal strategy Overall, the results confirm that the most effective strategy relies on the combination of highly competent specialized strains and appropriate nutrient additions (compost). This synergy is widely reported as the most effective approach for heavily polluted matrices, including coastal sediments (Azubuike et al. 2016; Villaverde et al. 2019 ). Compost serves as an energy and nutrient source, stimulating microbial growth; a structuring agent, improving sediment aeration; and a surfactant agent, increasing HAP bioavailability. Consequently, it promotes both microbial colonization and the expression of catabolic pathways involved in PAH degradation. Ecological implications for mangrove restoration The findings provide critical insights for mangrove bioremediation, ecosystems recognized for their vulnerability to petroleum pollutants (Alongi 2020 ). The high performance observed for R. erythropolis combined with compost suggests that a strategy based on stimulating the local microflora, adding adapted exogenous strains, and improving edaphic conditions could significantly accelerate sediment decontamination and reduce the ecotoxicological risks associated with HAPs. These results align with studies demonstrating microbial resilience in mangroves and their high potential for assisted restoration (dos Santos et al. 2021; Tavares et al. 2021 ). 5. Conclusion This study evaluated the biodegradation potential of high-molecular-weight polycyclic aromatic hydrocarbons (PAHs), represented by pyrene and fluoranthene, in polluted mangrove sediments from the Wouri estuary in Cameroon. The results confirm that the indigenous sediment microflora possesses an intrinsic capacity to degrade these recalcitrant pollutants, which can be enhanced through the introduction of exogenous strains such as Rhodococcus erythropolis and Bacillus subtilis . Experiments under sterile and non-sterile conditions demonstrated that biodegradation can be optimized through different strategies, including bioaugmentation, biostimulation, and their combination. The presence of nutrient amendments (nitrogen, phosphorus, compost) also contributes to stimulating microbial activity and improving degradation rates. Additionally, monitoring pH and microbial density provided insights into limiting factors and favorable conditions for HAP degradation in these sediments. Overall, this research demonstrates that the synergistic exploitation of indigenous microflora and exogenous strains constitutes an effective strategy for the bioremediation of contaminated mangrove sediments. These findings provide new knowledge on the role of microorganisms in coastal ecosystem depollution and open avenues for developing bioremediation technologies adapted to tropical mangroves, thereby contributing to biodiversity conservation and ecological restoration of these sensitive environments. Declarations Author Contribution Both authors contributed equally to the conceptualization of the study, including the definition of the overall aim and specific objectives. They jointly developed the methodology and experimental design, conducted the data analysis, and interpreted the results. Both authors reviewed, revised, and approved the final version of the manuscript prior to submission. 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1","display":"","copyAsset":false,"role":"figure","size":43903,"visible":true,"origin":"","legend":"\u003cp\u003epH evolution during pyrene biodegradation\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-8520850/v1/79a305aa3d7d68be3617d8db.png"},{"id":100363706,"identity":"5391740d-b930-4858-bc28-eaa4d05c450d","added_by":"auto","created_at":"2026-01-16 07:51:18","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":41175,"visible":true,"origin":"","legend":"\u003cp\u003epH evolution during fluoranthene biodegradation\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-8520850/v1/c5010bdd1b9e03ccea32d372.png"},{"id":100064234,"identity":"a4efae6f-50a1-4bdb-aded-4356aac70c4e","added_by":"auto","created_at":"2026-01-12 15:13:58","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":39072,"visible":true,"origin":"","legend":"\u003cp\u003eGrowth of competent microflora in sterile mangrove sediments contaminated with pyrene\u003c/p\u003e","description":"","filename":"3.png","url":"https://assets-eu.researchsquare.com/files/rs-8520850/v1/8e9448c60d13cea4803c1eda.png"},{"id":100064239,"identity":"e48dba74-5069-49b6-8cca-c4cf005bb4fa","added_by":"auto","created_at":"2026-01-12 15:13:59","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":45348,"visible":true,"origin":"","legend":"\u003cp\u003eGrowth of competent microflora in sterile mangrove sediments contaminated with fluoranthene\u003c/p\u003e","description":"","filename":"4.png","url":"https://assets-eu.researchsquare.com/files/rs-8520850/v1/3fdff8b6245ffb176e5062b7.png"},{"id":100364107,"identity":"abb3ad60-4604-43c6-bd71-5dcddd1ab5e1","added_by":"auto","created_at":"2026-01-16 07:52:37","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":26629,"visible":true,"origin":"","legend":"\u003cp\u003ePyrene degradation rates in non-sterile sediments over the experimental period\u003c/p\u003e","description":"","filename":"5.png","url":"https://assets-eu.researchsquare.com/files/rs-8520850/v1/0598f36ad5cab19233a39c03.png"},{"id":100064237,"identity":"32bc5cae-9411-4f83-aed7-5a5941d02f9e","added_by":"auto","created_at":"2026-01-12 15:13:59","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":26103,"visible":true,"origin":"","legend":"\u003cp\u003eFluoranthene degradation rates in non-sterile sediments over the experimental period\u003c/p\u003e","description":"","filename":"6.png","url":"https://assets-eu.researchsquare.com/files/rs-8520850/v1/72e946e1c701c760686992e4.png"},{"id":103765626,"identity":"55df4de4-4538-4413-8a70-8f56eee5db9c","added_by":"auto","created_at":"2026-03-02 16:06:02","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3409623,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8520850/v1/1772363d-3edf-49b7-a539-00e8947aae3b.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Biodegradation of High-Molecular-Weight PAHs in Polluted Mangrove Sediments Using Indigenous Microflora and Exogenous Strains Rhodococcus erythropolis and Bacillus subtilis","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003eMangroves, composed mainly of woody plants, are among the most productive marine ecosystems, distributed along tropical and subtropical coastlines (Kathiresan and Bingham, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2001\u003c/span\u003e; Semboung et al. \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). Situated at the interface of marine, freshwater, and terrestrial environments, they play a major ecological role by providing diverse ecosystem services: protection against coastal erosion, nursery grounds for numerous aquatic species, and the capacity to degrade certain contaminants (Alongi, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2002\u003c/span\u003e; Duke et al. \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2007\u003c/span\u003e). Their functioning is strongly influenced by factors such as salinity, organic matter content, and tidal fluctuations, which promote remarkable biological diversity. Mangroves are therefore considered hotspots of microbial biodiversity (Thatoi et al. \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2013\u003c/span\u003e), with microorganisms playing a central role in biogeochemical cycles and ecosystem stability.\u003c/p\u003e \u003cp\u003eHowever, due to high exposure to anthropogenic activities, these environments are increasingly contaminated by polycyclic aromatic hydrocarbons (PAHs) (Zhao et al. \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e2012\u003c/span\u003e; Souza et al. \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). This pollution represents a major threat to the ecological stability of mangroves (Brito et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; dos Santos et al. \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2011\u003c/span\u003e; Andreote et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). Petroleum hydrocarbons and their derivatives have toxic effects on flora and fauna, with impacts ranging from acute to chronic. Low-molecular-weight aromatic compounds generally cause acute toxicity, whereas PAHs, due to their high recalcitrance, low solubility, and elevated toxicity, lead to particularly concerning chronic contamination (Mukherjee and Chattopadhyay \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Juhasz and Naidu \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2000\u003c/span\u003e; Kanaly and Harayama \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2010\u003c/span\u003e; Crampon et al. \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). To date, most studies have focused on the biodegradation of low-molecular-weight PAHs, which are more accessible and more readily degradable (Muckian et al. \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; de Menezes et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2012\u003c/span\u003e; Jiang et al. \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2017\u003c/span\u003e; Bacosa and Inoue \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). However, some studies have also explored the degradation of high-molecular-weight PAHs and the responses of microbial communities to these compounds (Tauler et al. \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Zhu et al. \u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e2016\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eBioremediation has emerged as a promising strategy to mitigate the effects of petroleum hydrocarbons in contaminated ecosystems, owing to its efficiency, low cost, and environmental compatibility. It relies primarily on the activity of microorganisms capable of degrading these pollutants, while interacting with other members of the microbial community (Fuentes et al. \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Hydrocarbon biodegradation has been investigated in various contaminated environments, including soils (Sun et al. \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2015\u003c/span\u003e), oceans (Sauret et al. \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2014\u003c/span\u003e), and mangroves (Jiang et al. \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Muangchinda et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). Although several studies have demonstrated the ability of indigenous bacteria from mangrove sediments to degrade hydrocarbons, including PAHs (Brito et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; Li et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; Tian et al., \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e2008\u003c/span\u003e), additional information is needed on their biodegradation rates and removal efficiencies to optimize bioremediation technologies.\u003c/p\u003e \u003cp\u003eSeveral hydrocarbonoclastic bacterial genera such as \u003cem\u003eRhodococcus\u003c/em\u003e, \u003cem\u003eAcinetobacter\u003c/em\u003e, \u003cem\u003ePseudomonas\u003c/em\u003e, \u003cem\u003eAlcanivorax\u003c/em\u003e, and \u003cem\u003eSphingomonas\u003c/em\u003e have been isolated from polluted mangrove sediments (Brito et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2006\u003c/span\u003e; Rocha et al. \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e2013\u003c/span\u003e; Yu et al. \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2005\u003c/span\u003e). Moreover, advances in high-throughput sequencing technologies (Illumina MiSeq, Illumina HiSeq, 454 GS FLX) have enabled a deeper understanding of microbial community structures and dynamics in various contaminated environments, including soils (Abbasian et al. \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2016\u003c/span\u003e), Arctic soils (Tan et al. \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e2013\u003c/span\u003e), marine sediments (Mason et al. \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2014\u003c/span\u003e), and mangrove sediments (Andreote et al. \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2012\u003c/span\u003e; dos Santos et al. \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). In the present study, sediments were artificially contaminated with two representative high-molecular-weight PAHs\u0026mdash;pyrene and fluoranthene\u0026mdash;commonly used in microcosm experiments to assess microbial responses to hydrocarbon pollution (Sauret et al. \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Schurig et al. \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e2014\u003c/span\u003e; Schwarz et al. \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e2017\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe main objective of this work is to evaluate the biodegradation potential of pyrene and fluoranthene in contaminated mangrove sediments by comparing the endogenous microflora with exogenous strains (\u003cem\u003eRhodococcus erythropolis\u003c/em\u003e and \u003cem\u003eBacillus subtilis\u003c/em\u003e). Specifically, the aims are to (i) characterize the competent indigenous microflora capable of degrading these compounds, (ii) assess the degradation performance of the exogenous strains under controlled microcosm conditions, and (iii) optimize biodegradation through bioaugmentation and microbial stimulation using nutrient amendments (nitrogen, phosphorus, compost). This study will contribute to expanding knowledge on the impact of PAHs on microbial communities in mangrove sediments and on developing bioremediation approaches adapted to these sensitive ecosystems.\u003c/p\u003e"},{"header":"2. Materials and Methods","content":"\u003cp\u003e\u003cstrong\u003e2.1 Materials\u003c/strong\u003e\u003c/p\u003e\n\u003cul\u003e\n \u003cli\u003e\u003cstrong\u003eMangrove sediments\u003c/strong\u003e\u003c/li\u003e\n\u003c/ul\u003e\n\u003cp\u003eThe mangrove sediments used in this study were collected from the Wouri estuary mangroves in Cameroon, a heavily industrialized zone. Solid and liquid wastes from various industries are discharged into the mangroves without prior treatment. Sediments were collected at depths between 20 and 40 cm. This area harbors abundant microflora and is likely to contain pollutants.\u003c/p\u003e\n\u003cul\u003e\n \u003cli\u003e\u003cstrong\u003eMicroorganisms\u003c/strong\u003e\u003c/li\u003e\n\u003c/ul\u003e\n\u003cp\u003eTwo categories of microorganisms were used: indigenous microorganisms and exogenous microorganisms. Indigenous microorganisms were used as a bacterial consortium after isolation, identification, and characterization. Exogenous microorganisms consisted of two pure strains, \u003cem\u003eRhodococcus erythropolis\u003c/em\u003e and \u003cem\u003eBacillus subtilis\u003c/em\u003e.\u0026nbsp;\u003cem\u003eR. erythropolis\u003c/em\u003e was obtained from the Bioindustries Laboratory of Gembloux Agro-Bio Tech, University of Liège, and the \u003cem\u003eB. subtilis\u003c/em\u003e strain was obtained from the Biology Laboratory of the University of Mons, Belgium.\u003c/p\u003e\n\u003cul\u003e\n \u003cli\u003e\u003cstrong\u003ePolycyclic aromatic hydrocarbons : Pyrene and Fluoranthene\u003c/strong\u003e\u003c/li\u003e\n\u003c/ul\u003e\n\u003cp\u003ePAHs are non-polar, hydrophobic compounds that do not ionize; consequently, their solubility in water is very low. In this study, two PAHs—pyrene (Pyr) and fluoranthene (Flt)—were used (purity \u0026gt; 97%, supplied by UCB, Belgium). Pyrene contains four benzene rings, with a molecular mass of 202.26 and a logKow of 5.18. Fluoranthene also contains four benzene rings, with a molecular mass of 178.24 and a logKow of 4.18.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2 Methods\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.1 Artificial contamination of sediments with pyrene and fluoranthene\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eSediments were artificially contaminated following the method described by Tam et al. (2008), with slight modifications. In a flask, 10,000 mg·kg⁻¹ dry weight of each PAH (pyrene and fluoranthene) was dissolved in 50 mL acetone and mixed with 200 g of mangrove sediments. The mixture was then combined with 1 kg of mangrove sediment and homogenized with a plastic spatula. The flask was left open for two days to allow acetone evaporation and PAH adsorption into the sediments. The resulting sediment concentration was approximately 10,000 mg·kg⁻¹ dry weight. Then, 250 mL of artificial seawater (salinity 10‰) was added to moisten the mixture.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.2 Isolation of competent microflora and production of the indigenous bacterial consortium\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIsolation was performed following Weekers et al. (1999) with modifications. One gram of polluted sediment was placed in 9 mL sterile peptone water and heated at 80 °C for 15 min to select thermoresistant microflora. Serial dilutions were plated on selective mineral medium and incubated at 35 °C for 72 h. Colonies were suspended (10⁴ CFU·mL⁻¹) in 100 mL sterile rich liquid medium containing 10 g·L⁻¹ diesel (preculture 1; 30 °C, shaking, 72 h). This preculture inoculated 500 mL of identical medium (preculture 2), incubated under the same conditions. Preculture 2 was transferred into a 5-L bioreactor containing 4 L rich liquid medium + 10 g·L⁻¹ diesel. Growth was monitored by OD₆₀₀ over 96 h. Biomass was recovered by centrifugation (3500 rpm, 15 min) and stored at 0 °C with 2.5% (v/v) glycerol and 10% (v/v) maltodextrin as cryoprotectants. CFU·g⁻¹ and biomass yield were determined. This selection procedure was chosen because it enriches hydrocarbonoclastic and stress-resistant bacterial populations—typical traits of effective hydrocarbon degraders in mangrove ecosystems.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMedia compositions :\u003c/strong\u003e\u003c/p\u003e\n\u003cul\u003e\n \u003cli\u003e\u003cem\u003ePeptone water\u003c/em\u003e (g·L⁻¹): casein peptone 10, NaCl 5, Na₂HPO₄ 9, K₂HPO₄ 1.5; pH 7.0 ± 0.2\u003c/li\u003e\n \u003cli\u003e\u003cem\u003eRich medium\u003c/em\u003e (g·L⁻¹): glucose 20, casein peptone 10, yeast extract 10, agar 15; pH 7.0 ± 0.2\u0026nbsp;\u003c/li\u003e\n \u003cli\u003e\u003cem\u003eSelective mineral medium\u003c/em\u003e (mg·L⁻¹): (NH₄)₂SO₄ 1000, K₂HPO₄ 800, KH₂PO₄ 200, MgSO₄.7H₂O 200, CaCl₂.2H₂O 100, FeSO₄.7H₂O 12, MnSO₄.7H₂O 3, ZnSO₄.7H₂O 3, CoSO₄.7H₂O 1, (NH₄)₆Mo₇O₂₄.4H₂O 1, NaCl 4, agar 15 g, diesel 10 ppm; pH 7.0 ± 0.2. Diesel served as the sole carbon source.\u003c/li\u003e\n\u003c/ul\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.3 Characterization and identification of the bacterial consortium using API 20E\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eCharacterization included:\u003c/p\u003e\n\u003cul\u003e\n \u003cli\u003e\u003cstrong\u003eGram staining\u003c/strong\u003e: observation after crystal violet (1 min), Lugol iodine (1 min), ethanol decolorization (20–50 s) and fuchsin counterstain (1 min). Violet cells = Gram-positive; pink cells = Gram-negative.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eCatalase test\u003c/strong\u003e: culture in exponential phase mixed with H₂O₂. Bubble formation (O₂) = positive (aerobic bacteria).\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eOxidase test\u003c/strong\u003e: colony applied to filter paper impregnated with oxidase reagent. Purple coloration = positive.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eAPI 20E identification\u003c/strong\u003e: a pure culture was suspended in physiological saline (0.5 mL), introduced into the API 20E strip, incubated at 37 °C for 24 h. After addition of the auxiliary reagent, enzymatic reactions were interpreted using the API 20E reading table to determine the biochemical profile and bacterial species.\u003c/li\u003e\n\u003c/ul\u003e\n\u003cp\u003eThe combination of Gram staining, catalase/oxidase tests, and API 20E biochemical profiling was selected because it covers both fundamental morphological traits and the key metabolic capabilities involved in hydrocarbon degradation (oxidative metabolism, organic acid production, nitrogen metabolism).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.4 Assessment of the biodegradation potential of Pyrene and Fluoranthene\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.4.1 Experimental setup\u003c/strong\u003e\u003c/p\u003e\n\u003cul\u003e\n \u003cli\u003e\u003cstrong\u003eSterile conditions (Sterile Sediments “SS”)\u003c/strong\u003e\u003c/li\u003e\n\u003c/ul\u003e\n\u003cp\u003eThe sterile-condition experiment assessed the ability of the indigenous bacterial consortium to degrade pyrene and fluoranthene and compared its degradation rates with those of the exogenous strains \u003cem\u003eR. erythropolis\u003c/em\u003e and \u003cem\u003eB. subtilis\u003c/em\u003e. Two separate experiments were conducted: one with pyrene and one with fluoranthene.\u003c/p\u003e\n\u003cp\u003eIn a set of three flasks, 1000 g of mangrove sediments previously polluted with pyrene at 10,000 mg·kg⁻¹ dry weight were placed. Then, 500 mL seawater was added to keep sediments moist. All three flasks were autoclaved to ensure sterilization. After sterilization, the flasks were placed on the laboratory bench for a 5-week follow-up. These three flasks constituted the control (T0), as autoclaving eliminated all microbial life capable of degrading hydrocarbons. A second set of three flasks (T1) was prepared identically, with the addition of the indigenous bacterial consortium at 10⁷ CFU·g⁻¹ dry weight. A third set (T2) received \u003cem\u003eBacillus subtilis\u003c/em\u003e at 10⁷ CFU·g⁻¹. A fourth set (T3) received \u003cem\u003eRhodococcus erythropolis\u003c/em\u003e at 10⁷ CFU·g⁻¹. The same design was applied to fluoranthene-polluted sediments.\u003c/p\u003e\n\u003cul\u003e\n \u003cli\u003e\u003cstrong\u003eNon-sterile conditions (Non-Sterile Sediments “NSS”)\u003c/strong\u003e\u003c/li\u003e\n\u003c/ul\u003e\n\u003cp\u003eThe non-sterile experiment evaluated various biodegradation strategies—natural attenuation, biostimulation, bioaugmentation, and combined biostimulation + bioaugmentation—using mangrove sediments polluted with either pyrene or fluoranthene.\u003c/p\u003e\n\u003cp\u003eFor natural attenuation (T1), relying solely on native degrading microorganisms, 3000 g of pyrene-polluted mangrove sediments were placed into three PVC microcosm jars kept in a greenhouse.\u003c/p\u003e\n\u003cp\u003eFor biostimulation (T2), compost was added at 30% of the sediment mass to stimulate microbial activity. For bioaugmentation, three sets were prepared: T3: indigenous bacterial consortium at 10⁷ CFU·g⁻¹ ; T4: \u003cem\u003eBacillus subtilis\u003c/em\u003e at 10⁷ CFU·g⁻¹ ; T5: \u003cem\u003eRhodococcus erythropolis\u003c/em\u003e at 10⁷ CFU·g⁻¹. Combined biostimulation and bioaugmentation consisted of: T6: compost + indigenous consortium ; T7: compost + \u003cem\u003eBacillus subtilis\u0026nbsp;\u003c/em\u003e; T8: compost + \u003cem\u003eRhodococcus erythropolis.\u0026nbsp;\u003c/em\u003eT0 represented sterile sediments polluted with pyrene without microbial addition. The same design was applied to fluoranthene-polluted sediments.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.4.1 pH monitoring under sterile and non-sterile conditions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThroughout the experiment, pH was measured weekly using a HANNA HI 207 pH-meter. pH monitoring provides information on alkalinity and acidity, as pH is a limiting factor in biodegradation.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.4.2 Monitoring of competent microflora under sterile conditions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eEnumeration of competent microflora in sterile sediments was performed by serial dilution and plating on Petri dishes. One gram of sediment was collected weekly from each flask and placed in a test tube containing 9 mL peptone water. Spreading was performed on rich solid medium. Each dilution was plated in triplicate. After 3 days of incubation at 30 °C, colony-forming units (CFUs) were counted and the mean value for each dilution determined.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.4.3 Determination of total hydrocarbons under sterile and non-sterile conditions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePAHs were extracted from the sediments every 7 days over a period of 5 weeks. The extraction procedure followed high-performance liquid chromatography (HPLC) analysis using an Agilent 1100 Series system, as described by Wannoussa et al. (2015), with slight modifications. One gram of wet sediment sample was placed in 18 mL glass tubes, and an equivalent weight of anhydrous sulfate along with 10 mL of hexane were added. The mixture was homogenized by vortexing and placed in an ultrasonic bath for 1 h. The tubes were then placed on a horizontal shaker protected from light for 16 h, and the organic phase was transferred into 15 mL Falcon tubes. After centrifugation and transfer into a new tube, the residue was extracted again with 10 mL of hexane and transferred into a glass flask. Hexane was evaporated using a rotary evaporator (BUCHI ROTAVAPOR R-200) heated to 55 °C, and the dry extract was reconstituted in 10 mL of methanol. Subsequently, 1 mL of this solution was placed in an HPLC vial, sealed with a septum, and crimped. Samples were then sent for HPLC analysis. HPLC analyses were performed using an Agilent 1100 Series system equipped with a C18 column (LiChroCART® 250–4.6 HPLC cartridge Purospher® STAR RP-18 endcapped 5 μm, Merck, Germany) maintained at 30 °C.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.5 Statistical analyses\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMean values were compared using analysis of variance (ANOVA) with a significance level of p ≤ 0.05. Differences between the various hydrocarbon reduction rates were also evaluated using the same test. ANOVA was used to assess differences between initial and final total petroleum hydrocarbon (TPH) concentrations in the treated flasks compared with the control flasks. All statistical analyses were performed using MINITAB 17® software (French version).\u003c/p\u003e"},{"header":"3. Results","content":"\u003cp\u003eThe present study aimed to evaluate the biodegradation potential of pyrene and fluoranthene present in contaminated mangrove sediments, using microorganisms including the indigenous microflora, represented here by the consortium, and exogenous strains such as \u003cem\u003eRhodococcus erythropolis\u003c/em\u003e and \u003cem\u003eBacillus subtilis\u003c/em\u003e. The results are presented in three sections: the first focuses on the characterization of the consortium, the second on experiments conducted under sterile conditions, and the third on experiments carried out under non-sterile conditions.\u003c/p\u003e\n\u003cdiv id=\"Sec14\" class=\"Section2\"\u003e\n \u003ch2\u003e3.1. Characterization and identification of bacterial isolates\u003c/h2\u003e\n \u003cdiv id=\"Sec15\" class=\"Section3\"\u003e\n \u003ch2\u003e3.1.1 Total and Hydrocarbon-Degrading Microflora\u003c/h2\u003e\n \u003cp\u003eEnumeration of bacterial populations revealed that the polluted mangrove sediments of the Wouri estuary contained a total microflora estimated at 3 \u0026times; 10⁸ CFU\u0026middot;g⁻\u0026sup1; of sediment. Within this community, the hydrocarbon-degrading (competent) microflora represented approximately 1.5 \u0026times; 10⁵ CFU\u0026middot;g⁻\u0026sup1; of sediment. This indicates that only a small fraction of the indigenous bacterial population is directly involved in hydrocarbon degradation.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec16\" class=\"Section3\"\u003e\n \u003ch2\u003e3.1.2 Microscopic Characteristics\u003c/h2\u003e\n \u003cp\u003eMicroscopic examination of the bacterial isolates provided information on their motility, cell size, morphology, and cellular arrangement. These observations constitute preliminary criteria for the differentiation of the strains included in the starter consortium. Microscopic examination revealed that the bacterial isolates exhibited diverse morphotypes, including cocci, bacilli, and coccobacilli. All isolates were found to be motile, showed an isolated growth arrangement, and tested Gram-positive (Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec17\" class=\"Section3\"\u003e\n \u003ch2\u003e3.1.3 Biochemical characteristics and API 20E identification tests\u003c/h2\u003e\n \u003cp\u003eBiochemical analyses performed with API 20E galleries provided insights into the metabolic profiles of the consortium isolates. These tests highlighted their energetic, carbohydrate, and protein metabolism, and enabled preliminary identification at the presumptive taxonomic level (Table\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e\n \u003cdiv class=\"gridtable\"\u003e\u0026nbsp;\u003ctable id=\"Tab1\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eBiochemical characteristics of the bacterial isolates\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eReactions / Traits\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eBacterial isolate 1\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eBacterial isolate 2\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eBaterial isolate 3\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eBacterial isolate 4\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eEnergy metabolism\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003cth align=\"left\"\u003e\u0026nbsp;\u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eOxidase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCatalase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eRespiratory type\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAerobic\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnaerobic\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAnaerobic\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eAerobic\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eCarbohydrate metabolism\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTSI (Glucose)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eLactose / Sucrose\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eH₂S production\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGelatinase (GEL)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eArabinose (ARA)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eONPG (\u0026beta;-galactosidase)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCitrate utilization\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMotility\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMannitol\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eVoges-Proskauer (VP)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMethyl Red (MR)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eProtein metabolism\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003ctd align=\"left\"\u003e\u0026nbsp;\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eADH (Arginine dihydrolase)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eLDC (Lysine decarboxylase)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eODC (Ornithine decarboxylase)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eUrease\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eIndole\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eTDA (Tryptophan deaminase)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n \u003c/div\u003e\n \u003cul\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003ePresumptive identification of isolates\u003c/strong\u003e\u003c/p\u003e\n \u003c/li\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003eBacterial isolate 1\u003c/strong\u003e (Oxidase +, Catalase +, Aerobic, Citrate +, VP +, Motile) \u0026rarr; consistent with \u003cstrong\u003eBacillus sp.\u003c/strong\u003e\u003c/p\u003e\n \u003c/li\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003eBacterial isolate 2\u003c/strong\u003e (Oxidase +, VP +, Mannitol +, Anaerobic) \u0026rarr; possible \u003cstrong\u003eEnterococcus sp.\u003c/strong\u003e or related Gram-positive bacteria.\u003c/p\u003e\n \u003c/li\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003eBacterial isolate 3\u003c/strong\u003e (Oxidase \u0026ndash;, Anaerobic, Glucose +, Motile) \u0026rarr; may correspond to \u003cstrong\u003eClostridium sp.\u003c/strong\u003e or related anaerobes.\u003c/p\u003e\n \u003c/li\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003eBacterial isolate 4\u003c/strong\u003e (Oxidase +, Catalase +, Aerobic, Mannitol +, VP +, Motile) \u0026rarr; compatible with \u003cstrong\u003ePseudomonas sp.\u003c/strong\u003e\u003c/p\u003e\n \u003c/li\u003e\n \u003c/ul\u003e\n \u003cdiv class=\"gridtable\"\u003e\u0026nbsp;\u003ctable id=\"Tab2\" border=\"1\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eBiochemical and morphological characterization of bacterial isolates recovered from hydrocarbon-contaminated mangrove sediments\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eIsolate\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eMorphology (shape)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eMotility\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eArrangement\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eGram\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eProbable bacterial genus\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eReported isolation in hydrocarbon-polluted mangroves\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eBacterial isolate 1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCoccus\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMotile\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSingle\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eBacillus sp.\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eBacillus\u003c/em\u003e spp. frequently reported as hydrocarbon-degraders in mangrove sediments (Das et al. 2014; Dias et al. 2012)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eBacterial isolate 2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCoccus\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMotile\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSingle\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eEnterococcus sp.\u003c/em\u003e (or related Gram-positive bacteria)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eEnterococcus\u003c/em\u003e spp. isolated from contaminated mangroves (Taylor et al. 2012; Kathiresan and Bingham \u003cspan class=\"CitationRef\"\u003e2001\u003c/span\u003e)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eBacterial isolate 3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eBacillus\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMotile\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSingle\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eClostridium sp.\u003c/em\u003e (anaerobe)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003eClostridium\u003c/em\u003e spp. associated with anaerobic hydrocarbon degradation in mangroves (Peixoto et al. 2011; Reddy and Dhanasekaran 2016)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eBacterial isolate 4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eCoccobacillus\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eMotile\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eSingle\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e+\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003ePseudomonas sp.\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cem\u003ePseudomonas\u003c/em\u003e spp. widely documented as hydrocarbon-degrading bacteria in mangroves (Sathe et al. 2014; Serrano et al. 2021)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n \u003c/div\u003e\n \u003c/div\u003e\n\u003c/div\u003e\n\u003cdiv id=\"Sec18\" class=\"Section2\"\u003e\n \u003ch2\u003e3.2 Experiments under sterile conditions\u003c/h2\u003e\n \u003cdiv id=\"Sec19\" class=\"Section3\"\u003e\n \u003ch2\u003e3.2.1 pH evolution over 5 weeks\u003c/h2\u003e\n \u003cul\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003epH evolution in sterile sediments contaminated with pyrene\u003c/strong\u003e\u003c/p\u003e\n \u003c/li\u003e\n \u003c/ul\u003e\n \u003cp\u003eThe pH changes during the 5-week incubation indicate a progressive acidification of the sediments in all microbial treatments, unlike the T0 control (pH\u0026thinsp;=\u0026thinsp;6.07 \u0026rarr; 6.67), which remained stable or showed a slight increase. This acidification reflects the production of organic acids resulting from pyrene biodegradation. Among the treatments, \u003cem\u003eRhodococcus erythropolis\u003c/em\u003e (T3: pH 6.23 \u0026rarr; 4.53) and especially its combination with compost (T6: pH 6.00 \u0026rarr; 3.60) exhibited the most pronounced pH decreases, reaching 4.53 and 3.60, respectively, indicating high metabolic activity and enhanced pyrene degradation. In contrast, \u003cem\u003eBacillus subtilis\u003c/em\u003e (T2: pH 6.87 \u0026rarr; 5.64) and its association with compost (T5: pH 6.58 \u0026rarr; 5.03) showed a more moderate acidification. The positive effect of compost was particularly notable for the bacterial consortium (T4: pH 6.10 \u0026rarr; 4.93), which outperformed the consortium alone (T1: 6.33 \u0026rarr; 5.27). Overall, treatment T6 appears to be the most effective for pyrene degradation in sterile sediments.\u003c/p\u003e\n \u003cul\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003epH evolution in sterile sediments contaminated with fluoranthene\u003c/strong\u003e\u003c/p\u003e\n \u003c/li\u003e\n \u003c/ul\u003e\n \u003cp\u003eThe pH evolution during fluoranthene biodegradation shows a progressive decrease for all microbial treatments (T1\u0026ndash;T6), whereas the T0 control (pH 6.67 \u0026rarr; 6.87) remained stable around 6.7. This acidification results from the production of organic acids, reflecting metabolic activity associated with fluoranthene degradation. Treatments involving \u003cem\u003eR. erythropolis\u003c/em\u003e (T3 and T6) exhibited the most significant pH decreases, reaching 4.73 and 3.81, respectively, indicating a strong fluoranthene biodegradation capacity. Compost addition notably enhanced microbial activity, particularly for \u003cem\u003eR. erythropolis\u003c/em\u003e, making T6 (pH 6.12 \u0026rarr; 3.81) the most effective treatment. In contrast, \u003cem\u003eB. subtilis\u003c/em\u003e and the bacterial consortium induced moderate acidification, reflecting a lower degree of fluoranthene degradation.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec20\" class=\"Section3\"\u003e\n \u003ch2\u003e3.2.2 Microflora dynamics in the flasks\u003c/h2\u003e\n \u003cul\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003eMicrobial growth during pyrene biodegradation in sterile sediments\u003c/strong\u003e\u003c/p\u003e\n \u003c/li\u003e\n \u003c/ul\u003e\n \u003cp\u003eFigure \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e illustrates microbial growth in sterile sediments contaminated with pyrene across different treatments (T0: sterile sediments, no inoculum; T1\u0026ndash;T3: sediments\u0026thinsp;+\u0026thinsp;pyrene\u0026thinsp;+\u0026thinsp;bacterial inoculum; T4\u0026ndash;T6: sediments\u0026thinsp;+\u0026thinsp;pyrene\u0026thinsp;+\u0026thinsp;compost\u0026thinsp;+\u0026thinsp;inoculum). The aim was to evaluate the ability of different inocula (consortium, \u003cem\u003eB. subtilis\u003c/em\u003e, \u003cem\u003eR. erythropolis\u003c/em\u003e) with or without compost to colonize and potentially degrade pyrene. In the T0 negative control, bacterial populations ranged from 0 to 1.6 \u0026times; 10⁻\u0026sup1; CFU\u0026middot;g⁻\u0026sup1; dry weight, with no significant microbial development, as expected in sterile conditions. The slight final increase likely reflects minor environmental contamination. Compost strongly stimulated microbial growth in sterile pyrene-contaminated sediments. \u003cem\u003eR. erythropolis\u003c/em\u003e was the most adapted species, while \u003cem\u003eB. subtilis\u003c/em\u003e performed moderately well. Surprisingly, the bacterial consortium was the least effective, likely due to poor adaptation to pyrene, internal competition among strains, or nutrient limitations in sterile sediment. Treatment T6 (Compost\u0026thinsp;+\u0026thinsp;\u003cem\u003eR. erythropolis\u003c/em\u003e) was the most effective, combining nutrient supplementation via compost with a hydrocarbon-specialist bacterium, making it the optimal candidate for pyrene bioremediation in sterile or nutrient-poor sediments.\u003c/p\u003e\n \u003cul\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003eMicrobial growth during fluoranthene biodegradation in sterile sediments\u003c/strong\u003e\u003c/p\u003e\n \u003c/li\u003e\n \u003c/ul\u003e\n \u003cp\u003eSimilarly, Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e shows microbial growth (CFU or cell counts) over 5 weeks across different treatments: T0: control (no inoculum), T1\u0026ndash;T3: without compost, T4\u0026ndash;T6: with compost; inocula: consortium, \u003cem\u003eB. subtilis\u003c/em\u003e, \u003cem\u003eR. erythropolis\u003c/em\u003e. Fluoranthene is slightly less toxic than pyrene for many bacteria, explaining some observed differences. In T0 (sterile sediments\u0026thinsp;+\u0026thinsp;fluoranthene), competent microflora ranged from 0 \u0026rarr; 10\u0026sup2; cells, consistent with the absence of growth in sterile conditions. Minor increases (~\u0026thinsp;10\u0026sup2;) reflect negligible contamination.\u003c/p\u003e\n \u003cp\u003eFor T1 (fluoranthene\u0026thinsp;+\u0026thinsp;bacterial consortium), the population declined from 2.6 \u0026times; 10⁷ to 4.2 \u0026times; 10⁶ by the end, reflecting poor adaptation in sterile sediments. For T2 (\u003cem\u003eB. subtilis\u003c/em\u003e), populations increased from 2.5 \u0026times; 10⁷ \u0026rarr; 6 \u0026times; 10⁸ cells, performing better than on pyrene. T3 (\u003cem\u003eR. erythropolis\u003c/em\u003e) showed strong growth from 2.3 \u0026times; 10⁷ \u0026rarr; 9.6 \u0026times; 10⁸ CFU\u0026middot;g⁻\u0026sup1;, confirming its powerful HAP-degrading capacity. Compost slightly improved consortium performance in T4 (2.7 \u0026times; 10⁷ \u0026rarr; 6 \u0026times; 10⁷ CFU\u0026middot;g⁻\u0026sup1;). T5 (Compost\u0026thinsp;+\u0026thinsp;\u003cem\u003eB. subtilis\u003c/em\u003e) showed significant growth (2.7 \u0026times; 10⁷ \u0026rarr; 2.3 \u0026times; 10⁹ CFU\u0026middot;g⁻\u0026sup1;), and T6 (Compost\u0026thinsp;+\u0026thinsp;\u003cem\u003eR. erythropolis\u003c/em\u003e) exhibited the highest growth (2.9 \u0026times; 10⁷ \u0026rarr; 3.6 \u0026times; 10⁹ CFU\u0026middot;g⁻\u0026sup1;).\u003c/p\u003e\n \u003cp\u003eOverall, fluoranthene was less inhibitory than pyrene. All treatments except T1 exhibited substantial growth. Compost acted as a nutrient amendment (C, N, P), providing microhabitats, stabilizing pH, and enhancing HAP bioavailability, leading to 3\u0026ndash;10 fold microbial growth increases. \u003cem\u003eR. erythropolis\u003c/em\u003e, a hydrophobic Gram-positive bacterium capable of producing biosurfactants, efficiently degraded four-ring HAPs such as fluoranthene, confirming its key role in bioremediation.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec21\" class=\"Section3\"\u003e\n \u003ch2\u003e3.2.3 Pyrene and fluoranthene degradation rates in sterile mangrove sediments\u003c/h2\u003e\n \u003cul\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003ePyrene degradation rates\u003c/strong\u003e\u003c/p\u003e\n \u003c/li\u003e\n \u003c/ul\u003e\n \u003cp\u003eFigure \u003cspan class=\"InternalRef\"\u003e5\u003c/span\u003e shows that pyrene degradation percentages significantly increased over 5 weeks in all treatments. The sterile control (T0) exhibited very low values (1.13\u0026thinsp;\u0026plusmn;\u0026thinsp;0.72% at week 1 to 2.36\u0026thinsp;\u0026plusmn;\u0026thinsp;1.83% at week 5), confirming the absence of biological activity and indicating negligible abiotic oxidation. Natural attenuation (T1) in non-sterile sediments showed degradation rates from 6.19\u0026thinsp;\u0026plusmn;\u0026thinsp;1.28% to 36.54\u0026thinsp;\u0026plusmn;\u0026thinsp;1.96%, demonstrating the capacity of indigenous microflora to metabolize pyrene. Compost biostimulation (T2) enhanced activity further, reaching 42.55\u0026thinsp;\u0026plusmn;\u0026thinsp;5.89%. Bioaugmentation with the bacterial consortium (T3: 50.38\u0026thinsp;\u0026plusmn;\u0026thinsp;5.5%), \u003cem\u003eB. subtilis\u003c/em\u003e (T4: 44.18\u0026thinsp;\u0026plusmn;\u0026thinsp;3.5%), and \u003cem\u003eR. erythropolis\u003c/em\u003e (T5: 57.59\u0026thinsp;\u0026plusmn;\u0026thinsp;1.65%) improved degradation, particularly for \u003cem\u003eR. erythropolis\u003c/em\u003e, known for HAP degradation via specific metabolic pathways (e.g., catechol dioxygenases). The bacterial degradation gradient was: \u003cem\u003eRhodococcus\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;Consortium\u0026thinsp;\u0026gt;\u0026thinsp;\u003cem\u003eB. subtilis\u003c/em\u003e. Combined bioaugmentation\u0026thinsp;+\u0026thinsp;biostimulation treatments were most effective: T6 (Compost\u0026thinsp;+\u0026thinsp;Consortium) 70.13\u0026thinsp;\u0026plusmn;\u0026thinsp;3.31%, T7 (Compost\u0026thinsp;+\u0026thinsp;\u003cem\u003eB. subtilis\u003c/em\u003e) 62.33\u0026thinsp;\u0026plusmn;\u0026thinsp;7.29%, and T8 (Compost\u0026thinsp;+\u0026thinsp;\u003cem\u003eR. erythropolis\u003c/em\u003e) 85.58\u0026thinsp;\u0026plusmn;\u0026thinsp;5.26%, showing strong synergy between nutrient enrichment and inoculation with a specialized strain.\u003c/p\u003e\n \u003cul\u003e\n \u003cli\u003e\n \u003cp\u003e\u003cstrong\u003eFluoranthene degradation rates\u003c/strong\u003e\u003c/p\u003e\n \u003c/li\u003e\n \u003c/ul\u003e\n \u003cp\u003eFigure \u003cspan class=\"InternalRef\"\u003e6\u003c/span\u003e presents fluoranthene degradation (%) in sterile sediments over 5 weeks. The T0 control exhibited very low abiotic degradation (3.68\u0026thinsp;\u0026plusmn;\u0026thinsp;2.08%), indicating low bioavailability without microbial activity, typical for four-ring HAPs. In T1 (consortium), degradation increased from 7.38\u0026thinsp;\u0026plusmn;\u0026thinsp;3.72% to 37.77\u0026thinsp;\u0026plusmn;\u0026thinsp;2.09%, reflecting the consortium\u0026apos;s bioaugmentation effect. Exogenous bacteria performed better: T2 (\u003cem\u003eB. subtilis\u003c/em\u003e) 11.84\u0026thinsp;\u0026plusmn;\u0026thinsp;3.37% \u0026rarr; 48.03\u0026thinsp;\u0026plusmn;\u0026thinsp;6.53%, T3 (\u003cem\u003eR. erythropolis\u003c/em\u003e) 19.18\u0026thinsp;\u0026plusmn;\u0026thinsp;2.5% \u0026rarr; 60.57\u0026thinsp;\u0026plusmn;\u0026thinsp;4.5%, demonstrating that isolated strains degraded fluoranthene more efficiently than the consortium. \u003cem\u003eR. erythropolis\u003c/em\u003e\u0026rsquo;s effectiveness is due to dioxygenases, resistance to toxic compounds, and ability to use HAPs as carbon sources. Compost biostimulation improved degradation in T4 (14.54\u0026thinsp;\u0026plusmn;\u0026thinsp;1.34% \u0026rarr; 50.28\u0026thinsp;\u0026plusmn;\u0026thinsp;2.25%), T5 (19.06\u0026thinsp;\u0026plusmn;\u0026thinsp;1.36% \u0026rarr; 58.21\u0026thinsp;\u0026plusmn;\u0026thinsp;4.9%), and T6 (23.30\u0026thinsp;\u0026plusmn;\u0026thinsp;1.52% \u0026rarr; 74.65\u0026thinsp;\u0026plusmn;\u0026thinsp;5.13%), acting as a nutrient source (C, N, P), enhancing fluoranthene bioavailability and stimulating microbial growth. The degradation efficiency ranking after five weeks was: T0\u0026thinsp;\u0026lt;\u0026thinsp;T1\u0026thinsp;\u0026lt;\u0026thinsp;T2\u0026thinsp;\u0026lt;\u0026thinsp;T4\u0026thinsp;\u0026lt;\u0026thinsp;T5\u0026thinsp;\u0026lt;\u0026thinsp;T3\u0026thinsp;\u0026lt;\u0026thinsp;T6. Bioaugmentation alone (T1\u0026ndash;T3) significantly improved degradation, with individual strains outperforming the consortium: \u003cem\u003eR. erythropolis\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;\u003cem\u003eB. subtilis\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;Consortium. Compost addition further amplified degradation by improving bioavailability, providing nutrients and organic matter, stimulating degradation enzyme expression, and enhancing microbial growth. Combined bioaugmentation\u0026thinsp;+\u0026thinsp;biostimulation (T6) was optimal.\u003c/p\u003e\n \u003c/div\u003e\n \u003cdiv id=\"Sec22\" class=\"Section3\"\u003e\n \u003ch2\u003e3.2.4 Mass balance of pyrene and fluoranthene degradation in sterile and non-sterile sediments\u003c/h2\u003e\n \u003cp\u003eTables \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e and \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e present the mass balance of pyrene and fluoranthene in sterile and non-sterile sediments after five weeks under different bioremediation strategies. Across all treatments, the low losses in the sterile control (T0) \u0026ndash; below 4% for both HAPs \u0026ndash; indicate that abiotic processes (volatilization, irreversible adsorption, photodegradation) contributed minimally. Therefore, losses observed in other treatments can be primarily attributed to microbial activity.\u003c/p\u003e\n \u003cp\u003eIn sterile sediments (Table \u003cspan class=\"InternalRef\"\u003e3\u003c/span\u003e), the introduction of exogenous microorganisms significantly increased HAP degradation compared with the control. Among the tested strains, \u003cem\u003eR. erythropolis\u003c/em\u003e performed best, achieving 45.07% and 60.58% losses for pyrene and fluoranthene, respectively, in the absence of compost (T3). This is consistent with its known capacity to metabolize high-molecular-weight hydrocarbons via broad-spectrum dioxygenases and effective biosurfactant production. \u003cem\u003eB. subtilis\u003c/em\u003e and the bacterial consortium also enhanced degradation, albeit at lower levels (28\u0026ndash;48% depending on the compound). Compost addition generally boosted exogenous bacterial activity, particularly for \u003cem\u003eR. erythropolis\u003c/em\u003e, which reached 59.56% and 74.66% losses for pyrene and fluoranthene, respectively (T6), indicating a biostimulation effect attributable to nutrient supply, improved HAP bioavailability, and concomitant stimulation of microbial growth.\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eTable 3\u003c/strong\u003e : Mass balance of PAHs amount remained (mg.kg\u003csup\u003e-1\u003c/sup\u003e) and percentages in each fraction to total PAHs (pyrene and fluoranthene) added (input) in sterile sediments after 5 weeks of degradation\u0026nbsp;\u003c/p\u003e\n \u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"908\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 220px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDifferent biodegradation strategies\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"9\" valign=\"bottom\" style=\"width: 667px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eMangrove Sterile sediments\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 138px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eFate of PAHs\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"3\" valign=\"bottom\" style=\"width: 254px;\"\u003e\n \u003cp\u003e\u003cstrong\u003ePyrene\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"3\" valign=\"bottom\" style=\"width: 254px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eFluoranthene\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAmounts (mg.kg\u003csup\u003e-1\u003c/sup\u003e)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e% Input\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAmounts (mg.kg\u003csup\u003e-1\u003c/sup\u003e)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e% Input\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 220px;\"\u003e\n \u003cp\u003eSterile Sediments (T0)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 138px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,088.33 \u0026plusmn; 62.51\u003c/p\u003e\n \u003cp\u003e9,850 \u0026plusmn; 130\u003c/p\u003e\n \u003cp\u003e338.33 \u0026plusmn; 187.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e97.63\u003c/p\u003e\n \u003cp\u003e3.83 \u0026plusmn;1.83\u003csup\u003ep\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,036 \u0026plusmn; 58.6\u003c/p\u003e\n \u003cp\u003e9,666.66 \u0026plusmn; 152.75\u003c/p\u003e\n \u003cp\u003e369.33 \u0026plusmn; 211.34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e96.32\u003c/p\u003e\n \u003cp\u003e3.68 \u0026plusmn; 2.08\u003csup\u003est\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 220px;\"\u003e\n \u003cp\u003eSterile Sediments + Bacterial Consortium (T1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 138px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,161.33 \u0026plusmn; 133.83\u003c/p\u003e\n \u003cp\u003e7233.33 \u0026plusmn; 152.75\u003c/p\u003e\n \u003cp\u003e2,927.66 \u0026plusmn; 25.42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e71.18\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e28.8 \u0026plusmn; 0.58\u003csup\u003efg\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,070.33 \u0026plusmn; 35.47\u003c/p\u003e\n \u003cp\u003e6,266.54 \u0026plusmn; 208.16\u003c/p\u003e\n \u003cp\u003e3,803.66 \u0026plusmn; 213.91\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e62.23\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e37.77 \u0026plusmn; 2.1\u003csup\u003eghi\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 220px;\"\u003e\n \u003cp\u003eSterile Sediments + \u003cem\u003eB. subtilis\u003c/em\u003e (T2)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 138px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10117.67 \u0026plusmn; 64.3\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e6300 \u0026plusmn; 300\u003c/p\u003e\n \u003cp\u003e3817.66 \u0026plusmn; 162.69\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e62.27\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e37.73 \u0026plusmn; 2.5\u003csup\u003ecd\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,134 \u0026plusmn; 74.11\u003c/p\u003e\n \u003cp\u003e5,266.66 \u0026plusmn; 665.83\u003c/p\u003e\n \u003cp\u003e4,867.33 \u0026plusmn; 656.9\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e51.97\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e48.03 \u0026plusmn; 6.53\u003csup\u003ecde\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 220px;\"\u003e\n \u003cp\u003eSterile Sediments + \u003cem\u003eR. erythropolis\u003c/em\u003e (T3)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 138px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10073.66 \u0026plusmn; 45.5\u003c/p\u003e\n \u003cp\u003e5533.33 \u0026plusmn; 208.2\u003c/p\u003e\n \u003cp\u003e4540.33 \u0026plusmn; 162.7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e54.93\u003c/p\u003e\n \u003cp\u003e45.07 \u0026plusmn; 1.82\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,146 \u0026plusmn; 113.33\u003c/p\u003e\n \u003cp\u003e4000 \u0026plusmn; 500\u003c/p\u003e\n \u003cp\u003e6,146 \u0026plusmn; 387.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e39.42\u003c/p\u003e\n \u003cp\u003e60.58 \u0026plusmn; 4.5\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 220px;\"\u003e\n \u003cp\u003eSterile Sediments + Compost + Bacterial Consortium (T4)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 138px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,102 \u0026plusmn; 132.04\u003c/p\u003e\n \u003cp\u003e6,683.33 \u0026plusmn; 160.72\u003c/p\u003e\n \u003cp\u003e3,418.33 \u0026plusmn; 288.56\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e66.15\u003c/p\u003e\n \u003cp\u003e33.85 \u0026plusmn; 2.41\u003csup\u003ede\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,258.66 \u0026plusmn; 264.2\u003c/p\u003e\n \u003cp\u003e5,100 \u0026plusmn; 360.55\u003c/p\u003e\n \u003cp\u003e5,158.66 \u0026plusmn; 116.21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e48.71\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e51.39 \u0026plusmn; 2.25\u003csup\u003ecd\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 220px;\"\u003e\n \u003cp\u003eSterile Sediments + Compost + \u003cem\u003eB. subtilis\u003c/em\u003e (T5)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 138px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,066.33 \u0026plusmn; 170\u003c/p\u003e\n \u003cp\u003e5,583.33 \u0026plusmn; 550.75\u003c/p\u003e\n \u003cp\u003e4,483 \u0026plusmn; 529.96\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e55.47\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e44.53 \u0026plusmn; 5.24\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,130.66 \u0026plusmn; 160.05\u003c/p\u003e\n \u003cp\u003e4,233.33 \u0026plusmn; 550.75\u003c/p\u003e\n \u003cp\u003e5,897.33 \u0026plusmn; 461.86\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e41.78\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e58.22 \u0026plusmn; 4.99\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 220px;\"\u003e\n \u003cp\u003eSterile Sediments + Compost + \u003cem\u003eR. erythropolis\u003c/em\u003e (T6)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 138px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,099 \u0026plusmn; 227.51\u003c/p\u003e\n \u003cp\u003e4,083.33 \u0026plusmn; 381.88\u003c/p\u003e\n \u003cp\u003e6,015.66 \u0026plusmn; 335.92\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e40.44\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e59.56 \u0026plusmn; 3.43\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 150px;\"\u003e\n \u003cp\u003e10,127 \u0026plusmn; 163.64\u003c/p\u003e\n \u003cp\u003e2,566.66 \u0026plusmn; 5.3.32\u003c/p\u003e\n \u003cp\u003e7,560.33 \u0026plusmn; 567.72\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 93px;\"\u003e\n \u003cp\u003e25.34\u003c/p\u003e\n \u003cp\u003e74.66 \u0026plusmn; 5.13\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n \u003cp\u003e\u0026nbsp;In non-sterile sediments (Table \u003cspan class=\"InternalRef\"\u003e4\u003c/span\u003e), the presence of indigenous microflora significantly enhanced HAP degradation, even in the absence of external inputs, as demonstrated by natural attenuation (T1) with losses ranging from 30 to 36%. Compost addition alone (biostimulation) further improved this process, achieving losses of 40\u0026ndash;42%, indicating a positive response of the native communities to nutrient enrichment. Bioaugmentation with isolated strains led to additional improvements, with \u003cem\u003eR. erythropolis\u003c/em\u003e remaining the most effective strain (57.59% for pyrene and 73.91% for fluoranthene). The highest levels of dissipation were achieved when bioaugmentation was combined with biostimulation. In particular, the treatment combining \u003cem\u003eR. erythropolis\u003c/em\u003e and compost (T8) resulted in near-complete HAP removal, with 85.58% loss for pyrene and 94.16% for fluoranthene. These results demonstrate a synergistic effect between the introduced bacteria, the indigenous microflora, and compost, leading to enhanced degradation capacity, likely through metabolic complementarity, improved HAP solubilization, and intensified microbial functional diversity.\u003c/p\u003e\n \u003cp\u003eOverall, the results clearly show that HAP biodegradation in sediments strongly depends on the availability of specialized microorganisms and the organic enrichment of the substrate. Although fluoranthene is more recalcitrant than pyrene, it was degraded more extensively under combined treatments, indicating that increased bioavailability via compost and microbial cooperation is critical for the breakdown of highly hydrophobic compounds. The most effective strategy is unequivocally the bioaugmentation\u0026ndash;biostimulation combination using \u003cem\u003eR. erythropolis\u003c/em\u003e and compost in non-sterile sediments. This approach maximizes degradation capabilities by simultaneously leveraging the strengths of the indigenous microflora and exogenous high-metabolic-potential strains. These findings demonstrate that the simultaneous integration of bioaugmentation and biostimulation constitutes a robust and highly effective approach for remediating sediments contaminated with high-molecular-weight HAPs. This strategy therefore holds considerable potential for in situ bioremediation applications in coastal and port ecosystems, where the persistence of HAPs represents a major environmental concern.\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eTable 4\u003c/strong\u003e : Mass balance of PAHs amount remained (mg.kg\u003csup\u003e-1\u003c/sup\u003e) and percentages in each fraction to total PAHs (pyrene and fluoranthene) added (input) in nonsterile sediments after 5 weeks of degradation\u0026nbsp;under different techniques\u0026nbsp;\u003c/p\u003e\n \u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"898\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 101px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eBiodegradation Techniques\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 170px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eDifferents biodegradation strategies\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd colspan=\"3\" valign=\"top\" style=\"width: 170px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"5\" valign=\"bottom\" style=\"width: 434px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eMangrove non sterile sediments\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"2\" style=\"width: 112px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eFate of PAHs\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"3\" valign=\"bottom\" style=\"width: 235px;\"\u003e\n \u003cp\u003e\u003cstrong\u003ePyrene\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"bottom\" style=\"width: 235px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eFluoranthene\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAmounts (mg.kg\u003csup\u003e-1\u003c/sup\u003e)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 94px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e% Input\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e\u003cstrong\u003eAmounts (mg.kg\u003csup\u003e-1\u003c/sup\u003e)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 95px;\"\u003e\n \u003cp\u003e\u003cstrong\u003e% Input\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 101px;\"\u003e\n \u003cp\u003eSterile control\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 170px;\"\u003e\n \u003cp\u003eSterile Sediments (T0)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 112px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,088.33 \u0026plusmn; 62.51\u003c/p\u003e\n \u003cp\u003e9,850 \u0026plusmn; 130\u003c/p\u003e\n \u003cp\u003e238.33 \u0026plusmn; 187.1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 94px;\"\u003e\n \u003cp\u003e97.64\u003c/p\u003e\n \u003cp\u003e2.36 \u0026plusmn; 1.84\u003csup\u003eyz\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,036.1 \u0026plusmn; 58.46\u003c/p\u003e\n \u003cp\u003e9,666.66 \u0026plusmn; 152.75\u003c/p\u003e\n \u003cp\u003e369.43 \u0026plusmn; 211.21\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 95px;\"\u003e\n \u003cp\u003e96.32\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e3.68 \u0026plusmn; 2.07\u003csup\u003etuv\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 101px;\"\u003e\n \u003cp\u003eNatural attenuation\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 170px;\"\u003e\n \u003cp\u003eNon sterile sediments (T1)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 112px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,137.67 \u0026plusmn; 105.66\u003c/p\u003e\n \u003cp\u003e6,433.33 \u0026plusmn; 152.75\u003c/p\u003e\n \u003cp\u003e3,704.33 \u0026plusmn; 232.3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 94px;\"\u003e\n \u003cp\u003e63.46\u003c/p\u003e\n \u003cp\u003e36.54 \u0026plusmn; 1.96\u003csup\u003ejkl\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,180.64 \u0026plusmn; 134.78\u003c/p\u003e\n \u003cp\u003e7033.33 \u0026plusmn; 450.92\u003c/p\u003e\n \u003cp\u003e3,147.3 \u0026plusmn; 428.09\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 95px;\"\u003e\n \u003cp\u003e69.09\u003c/p\u003e\n \u003cp\u003e30.91 \u0026plusmn; 4.25\u003csup\u003ehijkl\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"3\" style=\"width: 101px;\"\u003e\n \u003cp\u003eBiostimulation\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 170px;\"\u003e\n \u003cp\u003eNon sterile sediments + Compost (T2)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 112px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,095.66 \u0026plusmn; 199.13\u003c/p\u003e\n \u003cp\u003e5,800 \u0026plusmn; 700\u003c/p\u003e\n \u003cp\u003e4,295.66 \u0026plusmn; 522.92\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 94px;\"\u003e\n \u003cp\u003e57.45\u003c/p\u003e\n \u003cp\u003e42.54 \u0026plusmn; 5.9\u003csup\u003ehij\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,175.86 \u0026plusmn; 109.15\u003c/p\u003e\n \u003cp\u003e6100 \u0026plusmn; 360.55\u003c/p\u003e\n \u003cp\u003e4,075.86 \u0026plusmn; 287.02\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 95px;\"\u003e\n \u003cp\u003e59.95\u0026nbsp;\u003c/p\u003e\n \u003cp\u003e40.05 \u0026plusmn; 3.08\u003csup\u003efg\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"9\" style=\"width: 101px;\"\u003e\n \u003cp\u003eBioaugmentation\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 170px;\"\u003e\n \u003cp\u003eNon sterile sediments + Bacterial consortium (T3)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 112px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,143.66 \u0026plusmn; 291.56\u003c/p\u003e\n \u003cp\u003e5,033.33 \u0026plusmn; 642.91\u003c/p\u003e\n \u003cp\u003e5,110.33 \u0026plusmn; 487.45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 94px;\"\u003e\n \u003cp\u003e49.62\u003c/p\u003e\n \u003cp\u003e50.38 \u0026plusmn; 5.5\u003csup\u003efg\u003c/sup\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,189.43 \u0026plusmn; 145.41\u003c/p\u003e\n \u003cp\u003e4,766.66 \u0026plusmn; 251.66\u003c/p\u003e\n \u003cp\u003e5,423.16 \u0026plusmn; 345.24\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 95px;\"\u003e\n \u003cp\u003e46.78\u003c/p\u003e\n \u003cp\u003e53.22 \u0026plusmn; 2.82\u003csup\u003ecd\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 170px;\"\u003e\n \u003cp\u003eNon sterile sediments + \u003cem\u003eB. subtilis\u003c/em\u003e (T4)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 112px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,151.33 \u0026plusmn; 147\u003c/p\u003e\n \u003cp\u003e5,666.66 \u0026plusmn; 288.67\u003c/p\u003e\n \u003cp\u003e4,484.66 \u0026plusmn; 442.03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 94px;\"\u003e\n \u003cp\u003e55.82\u003c/p\u003e\n \u003cp\u003e44.18 \u0026plusmn; 3.58\u003csup\u003eghi\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,225.45 \u0026plusmn; 232.53\u003c/p\u003e\n \u003cp\u003e4366.66 \u0026plusmn; 585.94\u003c/p\u003e\n \u003cp\u003e5858.78 \u0026plusmn; 775.69\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 95px;\"\u003e\n \u003cp\u003e42.7\u003c/p\u003e\n \u003cp\u003e57.3 \u0026plusmn; 6.41\u003csup\u003ecd\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 170px;\"\u003e\n \u003cp\u003eNon sterile sediments + \u003cem\u003eR. erythropolis\u003c/em\u003e (T5)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 112px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,138.33 \u0026plusmn; 220.47\u003c/p\u003e\n \u003cp\u003e4,300 \u0026plusmn; 200\u003c/p\u003e\n \u003cp\u003e5,838.33 \u0026plusmn; 199\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 94px;\"\u003e\n \u003cp\u003e42.41\u003c/p\u003e\n \u003cp\u003e57.59 \u0026plusmn; 1.65\u003csup\u003ede\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,221.1\u0026plusmn; 239.49\u003c/p\u003e\n \u003cp\u003e2,666.66 \u0026plusmn; 663.76\u003c/p\u003e\n \u003cp\u003e7554.44 \u0026plusmn; 524.79\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 95px;\"\u003e\n \u003cp\u003e26.09\u003c/p\u003e\n \u003cp\u003e73.91 \u0026plusmn; 6.81\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd rowspan=\"7\" style=\"width: 101px;\"\u003e\n \u003cp\u003eBioaugmentation and Biostimulation\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 170px;\"\u003e\n \u003cp\u003eNon sterile sediments + Compost + Bacterial consortium (T6)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 112px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,210.66 \u0026plusmn; 207.2\u003c/p\u003e\n \u003cp\u003e3,050 \u0026plusmn; 278.38\u003c/p\u003e\n \u003cp\u003e7,160 \u0026plusmn; 484.85\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 94px;\"\u003e\n \u003cp\u003e29.87\u003c/p\u003e\n \u003cp\u003e70.13 \u0026plusmn; 3.31\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,354.86 \u0026plusmn; 192.53\u003c/p\u003e\n \u003cp\u003e3,100 \u0026plusmn; 529.15\u003c/p\u003e\n \u003cp\u003e7,254.86 \u0026plusmn; 421.36\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 95px;\"\u003e\n \u003cp\u003e29.94\u003c/p\u003e\n \u003cp\u003e70.06 \u0026plusmn; 4.76\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" style=\"width: 170px;\"\u003e\n \u003cp\u003eNon sterile sediments + Compost + \u003cem\u003eB. subtilis\u003c/em\u003e (T7)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 112px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,176.66 \u0026plusmn; 373.66\u003c/p\u003e\n \u003cp\u003e3,833.33 \u0026plusmn; 702.37\u003c/p\u003e\n \u003cp\u003e6,343.33 \u0026plusmn; 819.03\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 94px;\"\u003e\n \u003cp\u003e37.67\u003c/p\u003e\n \u003cp\u003e62.33 \u0026plusmn; 7.3\u003csup\u003ecd\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,326.51 \u0026plusmn; 359.92\u003c/p\u003e\n \u003cp\u003e2,566.66 \u0026plusmn; 513.16\u003c/p\u003e\n \u003cp\u003e7,759.84 \u0026plusmn; 245.93\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd rowspan=\"3\" valign=\"bottom\" style=\"width: 95px;\"\u003e\n \u003cp\u003e24.86\u003c/p\u003e\n \u003cp\u003e75.14 \u0026plusmn; 4.24\u003csup\u003eb\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 12px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 170px;\"\u003e\n \u003cp\u003eNon sterile sediments + Compost + \u003cem\u003eR. erythropolis\u003c/em\u003e (T8)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 112px;\"\u003e\n \u003cp\u003eInput\u003c/p\u003e\n \u003cp\u003eIn sediment phase\u003c/p\u003e\n \u003cp\u003eLosses\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 11px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd colspan=\"2\" valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,174.33 \u0026plusmn; 206.29\u003c/p\u003e\n \u003cp\u003e1,466.66 \u0026plusmn; 550.75\u003c/p\u003e\n \u003cp\u003e8,707 \u0026plusmn; 515.8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 94px;\"\u003e\n \u003cp\u003e14.42\u003c/p\u003e\n \u003cp\u003e85.58 \u0026plusmn; 5.26\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 11px;\"\u003e\u003cbr\u003e\u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 141px;\"\u003e\n \u003cp\u003e10,336.78 \u0026plusmn; 410.15\u003c/p\u003e\n \u003cp\u003e603.33 \u0026plusmn; 355.01\u003c/p\u003e\n \u003cp\u003e9733.45 \u0026plusmn; 308.31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"bottom\" style=\"width: 95px;\"\u003e\n \u003cp\u003e5.84\u003c/p\u003e\n \u003cp\u003e94.16 \u0026plusmn; 3.28\u003csup\u003ea\u003c/sup\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n \u003cp\u003e\u003cbr\u003e\u003c/p\u003e\n \u003c/div\u003e\n\u003c/div\u003e"},{"header":"4. Discussion","content":"\u003cp\u003eThe results obtained in this study clearly demonstrate the potential for pyrene and fluoranthene biodegradation in contaminated mangrove sediments by mobilizing both the indigenous microflora and specialized exogenous strains such as \u003cem\u003eRhodococcus erythropolis\u003c/em\u003e and \u003cem\u003eBacillus subtilis\u003c/em\u003e. Overall, the observed performances align with microbiological mechanisms previously reported in the literature regarding polycyclic aromatic hydrocarbon (PAH) degradation in coastal and estuarine environments.\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eComposition and potential of the indigenous mangrove microflora\u003c/b\u003e \u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eThe total microflora observed in Wouri sediments (3 \u0026times; 10⁸ CFU.g⁻\u0026sup1;) is comparable to values reported in other highly anthropized mangrove ecosystems, where microbial loads range from 10⁶ to 10⁹ CFU.g⁻\u0026sup1; (Kathiresan and Bingham \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2001\u003c/span\u003e; Thatoi et al. \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2013\u003c/span\u003e). The low proportion of microorganisms directly competent for hydrocarbon degradation (\u0026asymp;\u0026thinsp;10⁵ CFU.g⁻\u0026sup1;) is also consistent with observations by dos Santos et al. (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2011\u003c/span\u003e), who noted that only 0.01\u0026ndash;1% of the total mangrove microflora actively participates in PAH mineralization.\u003c/p\u003e \u003cp\u003eThe identified genera (\u003cem\u003eBacillus, Enterococcus, Clostridium, Pseudomonas\u003c/em\u003e) are regularly reported in hydrocarbon-contaminated sediments. \u003cem\u003ePseudomonas spp.\u003c/em\u003e and \u003cem\u003eBacillus spp.\u003c/em\u003e are particularly recognized as robust PAH degraders due to their tolerance to toxic stress, motility, and biosurfactant production (Das and Chandran \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2011\u003c/span\u003e; dos Santos et al. 2021). The presence of \u003cem\u003eClostridium sp.\u003c/em\u003e may facilitate anaerobic degradation pathways, which are particularly relevant in the reducing conditions of mangrove sediments (Li et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e2015\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003epH dynamics and metabolic activity during biodegradation\u003c/b\u003e \u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eThe pH decrease observed in most treatments, especially with \u003cem\u003eR. erythropolis\u003c/em\u003e and the compost\u0026thinsp;+\u0026thinsp;bacterial combinations, represents a well-recognized indicator of metabolic activity associated with HAP degradation. Organic acid production is an expected byproduct of hydrocarbon dioxygenation and catabolic pathways (Atlas and Hazen \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2011\u003c/span\u003e; Cerniglia \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e1993\u003c/span\u003e). This acidification is particularly pronounced for four-ring HAPs (pyrene and fluoranthene), whose degradation requires intense enzymatic activity, notably aromatic mono- and dioxygenases (Seo et al. \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2009\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe amplifying role of compost has also been documented: it provides essential nutrients (N, P), increases sediment porosity, and enhances HAP bioavailability by stimulating desorption from solid particles (Sayara and S\u0026aacute;nchez \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2020\u003c/span\u003e; Silva-Castro et al. \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). These mechanisms explain the more pronounced pH reductions observed in the combined treatments.\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eMicrobial growth and aptitude of exogenous strains\u003c/b\u003e \u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eGrowth profiles confirm the metabolic superiority of \u003cem\u003eR. erythropolis\u003c/em\u003e in HAP-contaminated environments. This species is known for its exceptional capacity to degrade complex hydrocarbons, notably through biosurfactant production, which enhances HAP solubility (Franzetti et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2010\u003c/span\u003e), strong hydrophobic surface properties facilitating adhesion to insoluble compounds (Boonchan et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2000\u003c/span\u003e), and a broad enzymatic arsenal including dioxygenases, monooxygenases, and dehydrogenases (Larkin et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2005\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe exponential growth achieved in the compost\u0026thinsp;+\u0026thinsp;\u003cem\u003eR. erythropolis\u003c/em\u003e treatment (up to 3.6 \u0026times; 10⁹ CFU.g⁻\u0026sup1;) is consistent with previous studies showing that organic substrate addition strongly enhances rhodococci proliferation under polluted conditions (Pacwa-Płociniczak et al. \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Conversely, the low performance of the bacterial consortium may result from negative competitive interactions or poor prior adaptation to HAPs, factors previously reported in non-optimized multi-strain bioaugmentations (Nzila \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2016\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003ePyrene and fluoranthene biodegradation performance\u003c/b\u003e \u003c/p\u003e \u003c/li\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003ePyrene degradation\u003c/b\u003e \u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eThe degradation rates clearly demonstrate the combined efficacy of bioaugmentation and biostimulation. The best performances were observed with \u003cem\u003eR. erythropolis\u003c/em\u003e (\u0026asymp;\u0026thinsp;57.6\u0026thinsp;\u0026plusmn;\u0026thinsp;1.65%) and, notably, with the compost\u0026thinsp;+\u0026thinsp;\u003cem\u003eR. erythropolis\u003c/em\u003e combination (\u0026asymp;\u0026thinsp;85.6\u0026thinsp;\u0026plusmn;\u0026thinsp;5.26%). Such levels are consistent with other studies using hydrocarbonoclastic Actinobacteria for pyrene degradation (Kim et al. \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Zhang et al. 2021).\u003c/p\u003e \u003cp\u003eThe role of the indigenous microflora (\u0026asymp;\u0026thinsp;36.5\u0026thinsp;\u0026plusmn;\u0026thinsp;1.96%) further confirms that mangroves, as hydrocarbon-exposed environments, naturally harbor communities capable of PAH mineralization (Boopathy, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2000\u003c/span\u003e; Yu et al. \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2005\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eFluoranthene degradation\u003c/b\u003e \u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eFluoranthene degradation rates followed similar trends to those of pyrene, with maximum efficiency observed for the compost\u0026thinsp;+\u0026thinsp;\u003cem\u003eR. erythropolis\u003c/em\u003e treatment (\u0026asymp;\u0026thinsp;74.6\u0026thinsp;\u0026plusmn;\u0026thinsp;5.13%). This aligns with studies demonstrating that rhodococci can effectively degrade four-ring HAPs due to their high physiological tolerance (Weissenfels et al. \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e1990\u003c/span\u003e). The observed hierarchy (\u003cem\u003eRhodococcus\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;\u003cem\u003eBacillus subtilis\u003c/em\u003e\u0026thinsp;\u0026gt;\u0026thinsp;consortium\u0026thinsp;\u0026gt;\u0026thinsp;control) also matches numerous comparative studies on PAH bioaugmentation (Haritash and Kaushik \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; Juhasz and Naidu \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2000\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eBioaugmentation\u0026thinsp;+\u0026thinsp;Biostimulation synergy: an optimal strategy\u003c/b\u003e \u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eOverall, the results confirm that the most effective strategy relies on the combination of highly competent specialized strains and appropriate nutrient additions (compost). This synergy is widely reported as the most effective approach for heavily polluted matrices, including coastal sediments (Azubuike et al. 2016; Villaverde et al. \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e2019\u003c/span\u003e). Compost serves as an energy and nutrient source, stimulating microbial growth; a structuring agent, improving sediment aeration; and a surfactant agent, increasing HAP bioavailability. Consequently, it promotes both microbial colonization and the expression of catabolic pathways involved in PAH degradation.\u003c/p\u003e \u003cp\u003e \u003cul\u003e \u003cli\u003e \u003cp\u003e \u003cb\u003eEcological implications for mangrove restoration\u003c/b\u003e \u003c/p\u003e \u003c/li\u003e \u003c/ul\u003e \u003c/p\u003e \u003cp\u003eThe findings provide critical insights for mangrove bioremediation, ecosystems recognized for their vulnerability to petroleum pollutants (Alongi \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). The high performance observed for \u003cem\u003eR. erythropolis\u003c/em\u003e combined with compost suggests that a strategy based on stimulating the local microflora, adding adapted exogenous strains, and improving edaphic conditions could significantly accelerate sediment decontamination and reduce the ecotoxicological risks associated with HAPs. These results align with studies demonstrating microbial resilience in mangroves and their high potential for assisted restoration (dos Santos et al. 2021; Tavares et al. \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2021\u003c/span\u003e).\u003c/p\u003e"},{"header":"5. Conclusion","content":"\u003cp\u003eThis study evaluated the biodegradation potential of high-molecular-weight polycyclic aromatic hydrocarbons (PAHs), represented by pyrene and fluoranthene, in polluted mangrove sediments from the Wouri estuary in Cameroon. The results confirm that the indigenous sediment microflora possesses an intrinsic capacity to degrade these recalcitrant pollutants, which can be enhanced through the introduction of exogenous strains such as \u003cem\u003eRhodococcus erythropolis\u003c/em\u003e and \u003cem\u003eBacillus subtilis\u003c/em\u003e.\u003c/p\u003e \u003cp\u003eExperiments under sterile and non-sterile conditions demonstrated that biodegradation can be optimized through different strategies, including bioaugmentation, biostimulation, and their combination. The presence of nutrient amendments (nitrogen, phosphorus, compost) also contributes to stimulating microbial activity and improving degradation rates. Additionally, monitoring pH and microbial density provided insights into limiting factors and favorable conditions for HAP degradation in these sediments.\u003c/p\u003e \u003cp\u003eOverall, this research demonstrates that the synergistic exploitation of indigenous microflora and exogenous strains constitutes an effective strategy for the bioremediation of contaminated mangrove sediments. These findings provide new knowledge on the role of microorganisms in coastal ecosystem depollution and open avenues for developing bioremediation technologies adapted to tropical mangroves, thereby contributing to biodiversity conservation and ecological restoration of these sensitive environments.\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eBoth authors contributed equally to the conceptualization of the study, including the definition of the overall aim and specific objectives. They jointly developed the methodology and experimental design, conducted the data analysis, and interpreted the results. Both authors reviewed, revised, and approved the final version of the manuscript prior to submission.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003e\u003cstrong\u003eAbbasian F, Palanisami T, Megharaj M, Naidu R, Lockington R, Ramadass K,\u003c/strong\u003e (\u003cstrong\u003e2016)\u003c/strong\u003e Microbial diversity and hydrocarbon degrading gene capacity of a crude oil field soil as determined by metagenomics analysis. \u003cem\u003eBiotechnol. 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Bull\u003c/em\u003e. 57\u0026nbsp;: 707\u0026ndash;715.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eVillaverde J, L\u0026aacute;iz L, Lara-Moreno A, Gonz\u0026aacute;lez-Pimentel JL, \u0026amp; Morillo E (2019)\u003c/strong\u003e. \u003cem\u003eBioaugmentation of PAH-contaminated soils with novel specific degrader strains isolated \u0026nbsp;from a contaminated industrial site: Effect of hydroxypropyl-\u0026beta;-cyclodextrin as PAH \u0026nbsp; bioavailability enhancer\u003c/em\u003e.\u0026nbsp;\u003cem\u003eFrontiers in Microbiology\u003c/em\u003e, 10: 2588. https://doi.org/10.3389/fmicb.2019.02588.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eWeissenfels WD, Beyer M, \u0026amp; Klein J (1990)\u003c/strong\u003e Degradation of phenanthrene, fluorene and fluoranthene by pure bacterial cultures. \u003cem\u003eApplied Microbiology and Biotechnology\u003c/em\u003e, 32(4) : 479\u0026ndash;484.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eYu KSH, Wong AHY, Yau KWY, Wong YS, \u0026amp; Tam NFY (2005)\u003c/strong\u003e Natural attenuation, biostimulation and bioaugmentation on biodegradation of polycyclic aromatic hydrocarbons (PAHs) in mangrove sediments. \u003cem\u003eMarine Pollution Bulletin\u003c/em\u003e, 51(8‑12)\u0026nbsp;: 1071\u0026ndash;1077. https://doi.org/10.1016/j.marpolbul.2005.06.006.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eZhang L, Qiu X, Huang L, Xu J, Wang W, Li Z, et al, (2021d)\u003c/strong\u003e Microbial degradation of multiple PAHs by a microbial consortium and its application on contaminated wastewater. \u003cem\u003eJ. Hazard. Mater\u003c/em\u003e. 419 : 126524. 10.1016/j.jhazmat.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eZhao Z, Zhuang YX, Gu JD, (2012)\u003c/strong\u003e. Abundance, composition and vertical distribution of polycyclic aromatic hydrocarbons in sediments of the Mai Po inner deep bay of Hong Kong. \u003cem\u003eEcotoxicology\u003c/em\u003e 21 : 1734\u0026ndash;1742.\u003c/li\u003e\n \u003cli\u003e\u003cstrong\u003eZhu F, Storey S, Ashaari MM, Clipson N, Doyle E, (2016)\u003c/strong\u003e Benzo(a)pyrene degradation and microbial community responses in composted soil. \u003cem\u003eEnviron. Sci. Pollut. Res\u003c/em\u003e. \u003cem\u003e- Int\u003c/em\u003e. 24 : 5404\u0026ndash;5414.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"biodegradation","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"biod","sideBox":"Learn more about [Biodegradation](http://link.springer.com/journal/10532)","snPcode":"10532","submissionUrl":"https://submission.nature.com/new-submission/10532/3","title":"Biodegradation","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Biodegradation, Fluoranthene, Hydrocarbons, Mangroves, Microorganisms, Pollution, Pyrene","lastPublishedDoi":"10.21203/rs.3.rs-8520850/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8520850/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eMangroves are highly productive coastal ecosystems, yet they are heavily exposed to pollution by polycyclic aromatic hydrocarbons (PAHs), particularly high-molecular-weight compounds such as pyrene and fluoranthene. These recalcitrant contaminants exhibit high toxicity and persist in the environment, threatening microbial biodiversity and the ecological stability of sediments. This study aims to assess the biodegradation potential of pyrene and fluoranthene in contaminated mangrove sediments by comparing the efficiency of the indigenous microflora with that of exogenous strains (Rhodococcus erythropolis and Bacillus subtilis), and to optimize degradation through bioaugmentation and biostimulation using nutrient amendments. Sediments were artificially spiked with 10,000 mg\u0026middot;kg⁻\u0026sup1; of pyrene or fluoranthene. Microcosms under sterile and non-sterile conditions were established to evaluate, over five weeks, the biodegradation performance of the endogenous bacterial consortium, exogenous strains, biostimulation (compost), and their combinations. The results revealed significant PAH degradation by the indigenous microflora (45\u0026ndash;52% after five weeks). Exogenous strains enhanced degradation rates, reaching 58% for \u003cem\u003eB. subtilis\u003c/em\u003e and 63% for \u003cem\u003eR. erythropolis\u003c/em\u003e. The combined application of bioaugmentation and biostimulation yielded the highest degradation levels, with 75% for the endogenous consortium and up to 78% for \u003cem\u003eR. erythropolis\u003c/em\u003e. Statistical analyses confirmed that these differences were significant compared with sterile and non-sterile controls. The synergistic exploitation of indigenous microflora and exogenous strains, combined with nutrient amendments, constitutes an effective strategy for the bioremediation of mangrove sediments contaminated with high-molecular-weight PAHs. These findings provide a robust foundation for developing pollution-control technologies adapted to tropical coastal ecosystems.\u003c/p\u003e","manuscriptTitle":"Biodegradation of High-Molecular-Weight PAHs in Polluted Mangrove Sediments Using Indigenous Microflora and Exogenous Strains Rhodococcus erythropolis and Bacillus subtilis","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-01-12 15:13:53","doi":"10.21203/rs.3.rs-8520850/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-01-27T02:02:55+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-01-27T01:33:48+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-01-20T22:10:30+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-01-14T09:27:45+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-01-13T05:30:47+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"239456485633016598515749658045480921152","date":"2026-01-12T02:03:57+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"138982380374595419854828647075188267020","date":"2026-01-08T00:51:54+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"11058606510152138554282383602474371228","date":"2026-01-07T08:20:51+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"324467025219985332112054090590155950243","date":"2026-01-07T02:52:37+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"68684644374401556476823433955189391151","date":"2026-01-07T01:01:28+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"305686009413767896107790773055894746283","date":"2026-01-07T00:24:09+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-01-07T00:19:09+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-01-06T10:28:37+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-01-06T10:22:48+00:00","index":"","fulltext":""},{"type":"submitted","content":"Biodegradation","date":"2026-01-05T11:20:34+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"biodegradation","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"biod","sideBox":"Learn more about [Biodegradation](http://link.springer.com/journal/10532)","snPcode":"10532","submissionUrl":"https://submission.nature.com/new-submission/10532/3","title":"Biodegradation","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"c512991f-55c8-46a1-8187-68fd3ba42a06","owner":[],"postedDate":"January 12th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2026-03-02T16:02:56+00:00","versionOfRecord":{"articleIdentity":"rs-8520850","link":"https://doi.org/10.1007/s10532-026-10264-3","journal":{"identity":"biodegradation","isVorOnly":false,"title":"Biodegradation"},"publishedOn":"2026-02-23 15:59:15","publishedOnDateReadable":"February 23rd, 2026"},"versionCreatedAt":"2026-01-12 15:13:53","video":"","vorDoi":"10.1007/s10532-026-10264-3","vorDoiUrl":"https://doi.org/10.1007/s10532-026-10264-3","workflowStages":[]},"version":"v1","identity":"rs-8520850","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8520850","identity":"rs-8520850","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}