Life-Stage-Specific Survivorship and Mortality Aetiology of Gonimbrasia belina: The Interplay of Habitat Type, Seasons and Edaphic Factors

Life-Stage-Specific Survivorship and Mortality Aetiology of Gonimbrasia belina: The Interplay of Habitat Type, Seasons and Edaphic Factors | 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 Life-Stage-Specific Survivorship and Mortality Aetiology of Gonimbrasia belina: The Interplay of Habitat Type, Seasons and Edaphic Factors Fortunes Felix Matutu, Donald Mlambo, Angella Chichinye This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9058677/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 4 You are reading this latest preprint version Abstract Gonimbrasia belina (mopane worms) is a keystone species in the mopane woodland, yet its population dynamics remain unpredictable and poorly understood. This study aims to analyse the survivorship and mortality aetiology of G. belina by evaluating how habitat and seasonal variations interact to influence reproductive fitness, larval and pupal survivorship. The study used multi-factorial experiments and necropsy-based diagnostic frameworks across four sub-sites representing a combination of habitat and season, and the addition of five soil substrates for pupae. The greenhouse increased the overall survivorship (10%) of G. belina compared to natural mopane woodlands (1%). The greenhouse mitigated extrinsic mortality pressure from predation and abiotic stress, but it inadvertently created intrinsic pressures, especially disease and parasitoid-related mortality. The larval stage had the highest killing power, especially in the natural woodland, where predation is severe (65.6%). Pupae were influenced by seasons and soil substrate, with organic soils supporting high moth emergence in the wet season but with high mortality during the dry season (70%), while the addition of mulch to the substrate increased diapause rates. The study shows that natural woodlands under wet conditions serve as reproductive reservoirs where G. belina produces the best. Greenhouses protect against predation and weather, but they attract diseases and parasitoids. The conservation and domestication of G. belina require an integrated approach that preserves natural woodland to enhance productivity. Semi-domestication systems can be utilised as protective nurseries to mitigate predation and abiotic bottlenecks, but there is a need to develop bio-sanitation protocols to protect against pathogens and parasitoids. Clinical trial number: not applicable. Gonimbrasia belina Greenhouse Mortality aetiology Season Survivorship Woodland Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 1. Introduction Gonimbrasia belina , a keystone species in the mopane ( Colophospermum mopane) woodlands of southern Africa, is a major livelihood source for the rural folk while playing a vital role in nutrient recycling in the ecosystem (Bara et al., 2022 ; Kwiri et al., 2020 ; Nemadodzi et al., 2023 ; Glew et al., 1999 ). Despite this ecological and socio-economic importance, the species is characterised by unpredictable outbreaks, where certain seasons yield immense abundance while others result in localised near-extinction (Shen et al., 2023 ; Nethavhani et al., 2025 ). Extensive evidence suggests that the survivorship of G. belina is compromised by a suite of environmental stressors and biological threats that act across all phases of its life cycle (Sarangi et al., 2026 ; Ghazoul, 2006 ). Further anecdotal evidence suggests that even when moths successfully lay eggs, subsequent larval and pupal recruitment can be severely limited (Ditlhogo, 1996 ; Kwiri et al., 2020 ). Several studies have shown that deforestation, climate variability, and harvesting practices have led to lower G. belina abundance and distribution (Mataboge et al., 2016 ; Shen et al., 2023 ; Ndlovu et al., 2024 ). Furthermore, G. belina's presence and abundance are linked to weather elements, including rainfall, temperature and humidity, with extreme weather patterns negatively impacting its abundance and life-cycle recruitment (Sekonya, 2016; Togarepi et al., 2020 ). Studies by Ghazoul ( 2006 ) identified various biotic and abiotic factors influencing G. belina , including parasitoids, predation and diseases, while Dube (2013) identified the specific influence of temperature and humidity on its development. However, there remains a paucity of research examining the drivers of mortality at each life stage and their influence on ultimate survivorship. There are currently no comparative studies showing how these mortality agents vary across seasons, habitat types and soil substrates for specific cohorts. Beyond the habitat condition, G. belina faces various biotic mortality factors. Several studies have identified mortality factors, including diseases that are caused by pathogens such as viruses, flesh-eating bacteria and fungi (Ditlhogo, 1996 ; Ghazoul, 2006 ; Hope et al., 2009 ). Furthermore, G. belina is subject to several predation factors, which consume it at all stages of the life cycle, including insectivorous birds, lizards, jackals, bat-eared foxes, honey, warthogs, rats, monkeys, baboons, snakes and importantly, people (Shen et al., 2023 ; Hope et al., 2009 ; De Nagy Koves Hrabar, 2006 ). Additional parasites have been shown to drive mortality; the main parasites in G. belina and other lepidoptera are chalcids ( Mesocomys pulchriceps and Hockeria spp ) and ichneumonids ( Enicospilus antancarus ) (Ghazoul, 2006 ). However, current ecological models for G. belina do not adequately account for the nuanced interplay between habitat type and seasonal fluctuations, particularly in how these factors filter specific mortality agents across the life cycle. The subterranean pupal life stage represents a critical phase during which survival and developmental fate are heavily dictated by edaphic factors (Bangira et al., 2024 ; Matutu et al., 2025 ). Pupae spend a significant portion of their life cycle in the ground and have generally been found to pupate in crumbly soil substrates, including river sand, pit sands and organic soils (Mufandaedza et al., 2018 ; Wen et al., 2016 ). These soil substrates possess distinct physical and chemical properties and attract varying levels of biotic activity (Delgado and Gómez, 2024 ). Consequently, these substrates influence the aetiology and survivorship of the pupae differently. However, there remains a need to better understand how these differences are filtered through specific habits and seasonal changes to influence the aetiology of survival and mortality. In response to the volatility of wild G. belina populations, there is increasing interest in semi-domestication through greenhouse rearing (Ghazoul, 2006 ; Hope et al., 2009 ). However, efforts to semi-domesticate G. belina in such artificial habitats have not been adequately tested and compared with natural woodlands. It remains unclear if these controlled environments truly mitigate mortality pressures or inadvertently introduce new physiological stressors when compared to natural woodlands. Therefore, there is a need to understand variations in survivorship and mortality across habitats and seasonal dynamics. This study will adopt a comparative approach to identify the optimal conditions and the physiological and environmental bottlenecks or trade-offs of the species across various habitats. Given the evident influence of biological and environmental pressures on G. belina , there remains a significant gap in understanding how these pressures affect their population dynamics. Current research has primarily focused on broad aspects, such as habitat loss, climate variability, and harvesting systems, but it fails to explain how various mortality agents are filtered through the landscape and across seasons. The semi-domestication (rearing) of G. belina in greenhouses introduces new dynamics that need to be tested and compared with the natural woodland benchmarks. This lack of comparative, quantitative data creates a critical bottleneck, preventing the development of truly adaptive management strategies that can effectively respond to the erratic nature of G. belina outbreaks and the varying pressures across diverse landscapes. Therefore, this study aims to analyse the survivorship and life-stage-specific (eggs, larvae and pupae) mortality aetiology of G. belina by evaluating how habitat and seasonal variations interact to influence reproductive fitness, larval survivorship, pupal survivorship and developmental fate. Specifically, we employ necropsy-based etiological assessments to evaluate the contributions of abiotic stress, pathogens, parasites, and predation across habitats (greenhouse vs natural woodland) and seasons (dry and wet). The study addressed the following research questions: How do habitats (natural woodland vs greenhouse) and seasonal variation (dry and wet) interact to determine the life-stage-specific survivorship and mortality dynamics of G. belina ? How do habitats (natural woodland vs. greenhouse) and seasons (dry and wet) differentially influence the reproductive output and egg hatching efficiency of G. belina ? How does habitat type (natural woodland vs. greenhouse) and seasonal variation (dry vs. wet) interact to influence the filtering of abiotic factors, diseases, parasitism and predation as contributors to G. belina larval mortality? How do habitat type, season and soil substrates (River Sand, River Sand with mulch, Pit Sand, Pit Sand with mulch and Organic Soil) interact to influence the pupal mortality aetiology and developmental fate of G. belina pupae? 2. Materials and methods 2.1. Study area This study was done in the Gwanda district (20◦56’20"S, 29◦01’07"E), in southern Zimbabwe. The district is characterised as a semi-arid to arid area, with a mean annual temperature of 22°C and an average annual rainfall of 450 mm (Mambiravana, 2025 ; Baudron et al., 2024 ). It experiences a prolonged dry season from May to November, followed by a very short rainy season from December to March. The dominant vegetation is C. mopane (mopane woodland), interspersed with occasional Combretum and Acacia species (Kumalo and Manyani, 2023 ). The district's soils are predominantly sandy Kalahari soils, with pockets of organic soils found in areas of high biomass (Sharara et al., 2022 ). These conditions represent the primary ecological stronghold for G. belina , with the highest occurrence and harvest frequencies in Zimbabwe (Nunu et al., 2019 ; Bara et al., 2022 ). 2.2. Study species G. belina is a species of emperor moth under the family Saturniidae, order Lepidoptera, and phylum Arthropoda. This economically and ecologically significant insect is native to the semi-arid regions of southern Africa. Its life cycle encompasses four distinct stages: egg, larva, pupae and adult moth. The life cycle of G. belina begins with the emergence of moths, which lay eggs within 5 days (Dube and Dube, 2010 ). These eggs hatch after 7 days, and the resulting larvae moult four times (28 to 35 days) before burrowing underground. Once underground, the larvae moult a fifth time into pupae (Ditlhogo, 1996 ). Within 4 days of burrowing, larvae are fully pupated (Dube and Dube 2010 ). G. belina then enters diapause at this stage, remaining dormant until environmental conditions suit their emergence as moths (Netshanzhe, 2025). As a bivoltine insect, G. belina produces two generations annually, a trait shared by other saturniidae whose voltinism often varies by location (Ditlhogo, 1996 ; Mogomotsi et al., 2018 ). The first generation larvae occur between October and January (dry season to early wet season), while the second spans February to May (wet season) (Kwiri et al., 2020 ). The bivoltism characteristic of G. belina , combined with the greenhouse environment, means that successive generations develop under contrasting environmental regimes. These environmental disparities are likely to shape the patterns of survivorship and the specific drivers of mortality. 2.3. Study design and sampling We used a multi-factorial experimental design to evaluate how habitat and seasonal differences interact to influence reproductive fitness, larval survivorship, pupal survivorship and developmental fate. The study area was stratified by habitat (greenhouse and natural woodland-outside) and season (wet and dry), resulting in four subsites: Greenhouse wet, Outside wet, Greenhouse dry and Outside dry. Data collection was replicated across the 2022 and 2024 bivoltine generations to ensure a substantial analysis of seasonal and habitat-driven dynamics. The sites were selected based on the historical presence of G. belina in resettlement areas with low human disturbance. The controlled experiments were conducted within two purpose-built greenhouses, each measuring 15 m x 10 m and 5 m in height. These structures were covered with a 40% shade net to accurately simulate mature canopy cover. Concurrently, outside experiments were established in the adjacent natural mopane woodlands, situated within 10 meters of the greenhouses, ensuring similar macro-climatic conditions while representing natural habitat dynamics. For the egg stage assessment, a total of 200 freshly laid G. belina egg clutches were selected at each of the four experimental settings. Clutch size was recorded initially, with hatching success recorded after 10 days of incubation. The larval stage investigation began with the hatchling larvae from the egg stage assessment. These larvae were subsequently observed over a 28-day period. To prevent host tree defoliation and larval migration, each tree was restricted to a single representative clutch. All additional eggs were removed and relocated to other trees. Larval mortality was recorded at 7-day intervals, with each specimen undergoing a field post-mortem examination to determine the aetiology of mortality. A diagnostic framework based on observed physical markers and remains was consistently applied to determine the cause of larval mortality (Table 1 ). Diseases were identified by tissue liquefaction or mummification (Gouli et al, 2011 ) while parasitism was recorded upon the presence of external eggs or cuticle exit holes (Ghazoul, 2006 ). In the absence of biotic signs, but with larvae appearing shrivelled and brittle, mortality was attributed to abiotic factors (Speight et al, 1999 ; Sarangi et al., 2026 ), whereas missing or partially consumed larvae were classified as predation (Coelho et al., 2023 ). Any ambiguous cases that lacked clear diagnostic signs were excluded from the dataset. For the pupal stage assessment, mature larvae (stage 5) were collected and introduced into controlled pupation beds. This phase of the experiment was designed as a multi-factorial study to evaluate how habitat type, season and soil substrates (River Sand, River Sand with mulch, Pit Sand, Pit Sand with mulch, and Organic Soil) interact to influence the aetiology of mortality and developmental fate of the pupae. These treatments were replicated across 180 individual pupation beds, each measuring 1 m × 1 m × 0.5 m. In each bed, 100 mature G. belina larvae, at their pre-pupal stage, were introduced and allowed to burrow, pupate naturally and eventually emerge as moths. We monitored the population throughout the emergence season to record the number of moths emerging. At the conclusion of the emergence period, pupae were excavated to determine whether they were alive or dead. We classified and recorded the surviving pupae as being in diapause and performed necropsies on all dead specimens to identify the specific drivers of mortality (Table 1 ). For pupae with shattered fragments or a total absence, mortality was attributed to predation (Coelho et al., 2023 ). Parasitism was identified by the presence of parasitoids or holes, and by distinguishing solitary ichneumonid emergence from gregarious chalcid apertures (Ghazoul, 2006 ; Gouli et al., 2011 ). Disease was confirmed by the presence of fungal mummification, pungent odours, or tissue liquefaction under slight physical pressure (Rothman and Myers, 1995 ; Ghazoul, 2006 ). Table 1 Diagnostic criteria and physical indicators used to determine the aetiology of mortality in G. belina larvae and pupae during field surveys. Life Stage Observation Diagnostic Indicators Aetiology Larvae Withered Shrivelled, dry and brittle, no odour, no liquefaction Abiotic Tissue decay or liquefaction, discolouration or mummification Fragile/ruptured skin, Flaccid, shrunken body, milky/brownish liquid, fungal mycelia or foul odour Disease: bacteria, fungi or viruses External tags or cuticle holes Small white/dark eggs, Visible exit or entry holes on the larval skin Parasitoids Physical damage or absence Presence of larval remains, partially consumed; disappearance Predators Pupae Softened or tissue inside decay or mummification Dark spots/mouldy patches on cuticle; pungent odour, Mummified (hard) body with erupting fungal spores; ruptures easily under slight physical pressure. Disease Exit holes or hollowed Small holes in the pupal body, the shell contains a parasitoid Ichneumonids: Large (15-40mm); solitary; single large circular exit hole. Chalcids: Small (1-5mm); gregarious; multiple minute pin-hole exit apertures. Parasitoids Shattered or absence Fragmented chitinous shell remains in the soil; missing pupae Predators 2.4 Data analysis All statistical analyses were performed in R version 4.4.3 (R Core Team, 2025 ). All data sets underwent quality control to assess accuracy, identify outliers, and detect missing values. The Shapiro-Wilk test was utilised to assess the normality of continuous variables, while Levene’s test was employed to evaluate the homogeneity of variances. The life-stage-specific dynamics of G. belina were analysed by constructing age-specific life tables to quantify survivorship and mortality. Population data were first aggregated to calculate the mean number of individuals surviving at each developmental stage (Egg, Larvae and Pupae). Survivorship was determined by using the proportion of individuals surviving from one life stage to the next. Stage mortality rate was determined by the percentage of individuals dying during the specific stage. Killing Power was calculated as the difference between the logarithms of successive life stages. Survivorship curves were plotted on a logarithmic scale to compare population decline rates across habitats (Greenhouse vs. Outside) and seasons (Wet vs. Dry). The killing power was visualised through multi-panel, faceted plots to compare the relative killing power of different life stages across habitats. The reproductive performance of G. belina , including clutch size and hatching success, was analysed using linear and generalised linear modelling. A two-way Analysis of Variance (ANOVA) was used to test the main effects of the interaction between habitat and season on clutch size and hatched eggs. A Generalised Linear Modelling (GLM) with a binomial distribution and a logit link function was used to examine the overall efficiency of moth emergence relative to initial egg counts, and to assess how these were filtered by the interaction between habit and season. Post-hoc comparisons were performed using Estimated Marginal Means (EMMs), and a Compact Letter Display (CLD) was generated using Tukey’s Honestly Significant Difference (HSD) test to identify significant differences (p < 0.05) between specific treatment combinations, and the results were visualised using boxplots. The success of G. belina larval development and the relative impact of various mortality drivers were analysed through a combination of frequentist modelling and predictive risk mapping. An HSD post hoc was used to evaluate the interaction between habitat and season on pupation success. For the etiological analysis of larval death, a stacked bar graph and larval mortality risk factor heatmap were constructed by aggregating total mortality percentages by cause and habitat, visualised as a gradient-filled tile matrix. The developmental fate of G. belina pupae (emerged moth, diapause, or mortality) was analysed using a three-way ANOVA to determine the interactive effects of habitat, season and soil substrate. To identify specific differences between treatment combinations, post hoc pairwise comparisons were done using the HSD test (p < 0.05). These results were visualised as faceted boxplots, with compact letter displays indicating statistical significance within each fate category. To further investigate the specific drivers of pupal death, a post-mortem etiological analysis was performed using a multinomial logistic regression. The significance of habitat and soil substrate as predictors of these causes was assessed using Type II Likelihood Ratio Tests. The proportional contributions of each mortality factor were visualised through stacked bar plots to illustrate compositional shifts between habitats, while heat map matrices were used to visualise the intensity and diversity of mortality risks across soil substrates. 3. Results 3.1 Effects of Habitat and Season on G. belina Survivorship and Mortality The life cycle analysis of G. belina showed that survivorship (lx) and killing power (kx) are significantly influenced by the interaction of habitat (Greenhouse and Outside) and season (Wet and Dry). Generalised Linear Modelling (GLM) results for overall survival to the moth (adult) stage showed that the odds of G. belina surviving from egg to moth were significantly lower for individuals raised outside (natural woodland) than for those raised in the greenhouse (β = -1.52, p < 0.001). However, the wet season was a significant positive driver of survival (β = 1.14, p < 0.001). The Outside (natural woodland) habitat exhibited a survival disadvantage that was significantly mitigated during the wet season (β = 0.31, p < 0.001). Survivorship (lx) declined across all cohorts as they progressed from egg to moth, resulting in a low percentage of individuals reaching the moth stage (Fig. 1 A and 1 B). However, the probability of survival from egg to the moth stage was consistently higher in the greenhouse, at an average of 10%, whereas survival in the outside environment was severely depressed, between 1% and 3%. This disparity was mostly pronounced during the dry season, when populations on the outside (natural woodland) experienced the steepest decline, resulting in negligible recruitment to the moth stage. Analysis of killing power (kx) revealed the larval stage as the primary developmental bottleneck for individuals outside the greenhouse, as evidenced by a sharp spike in mortality (kx = 0.8) across seasons (Fig. 1 C and 1 D). While the wet season generally improved survivorship across both habitats, the greenhouse appeared to stabilise mortality rates across all life stages (Fig. 1 B and 1 D). Furthermore, the combination of external (natural woodland) habitats and dry-season conditions exerted high mortality across the egg, larval, and pupal stages simultaneously, suggesting that environmental exposure and dry weather conditions compound to severely limit population persistence. 3.2 Influence of Habitat Type and Season on G. belina Egg Clutch Size and Hatching Efficiency The analysis of egg clutch size and hatching outputs of G. belina showed that they were significantly influenced by the interaction between habitat and season (Fig. 2 ). While both habitats (greenhouse and natural woodland; p < 0.05) and season (wet and dry; p < 0.05) independently affected clutch size and larval hatching, the impact of season (wet or dry) was significantly moderated by the greenhouse habitat. During the dry season, the greenhouse habitat exhibited significantly larger clutch and larval counts than the outside (natural woodland) habitat, which was highly sensitive to seasonal variation. The transition from dry to wet season increased clutch size and hatching efficiency in the natural woodland, but remained constant in the greenhouse. The beneficial effect of the wet season was also disproportionately greater in the outside (natural woodlands) habitat than in the greenhouse, with outside-wet having greater hatching success than greenhouse-wet. In contrast, the outside (natural woodland) habitat exhibited a clear egg laying and hatching efficiency deficit during the dry season, which was largely mitigated during the wet season (Fig. 2 ). 3.3 Influence of Habitat Type and Seasons on G. belina Larval Mortality The survival of G. belina larvae to the pupae life stage was significantly influenced by both rearing habitat and seasonal timing. Larvae growing in the greenhouse had higher survival rates across both seasons compared to those reared in an outside (natural woodland) habitat (Fig. 3 A and 3 B). Within the greenhouse, survival was significantly higher during the wet season (median ≈ 57%) compared to the dry season (median ≈ 45%; p < 0.05) (Fig. 3 A). In contrast, the outside (natural woodland) habitat recorded a high mortality, with survival rates consistently suppressed. In both the wet and dry seasons, the outside habitat was heavily skewed towards total mortality, with only a few outliers showing high survival (Fig. 3 B). The differences in larval survival between habitats and seasons were driven by a distinct aetiology of mortality (Fig. 4 ). In the outside (natural woodland) habitat, predation was the primary driver of larval death, contributing 46.6% of mortality in the dry season and 65.6% in the wet season. The larval loss was further exacerbated by substantial abiotic (24.4%) and disease (16.1%) losses during the dry period. On the other hand, the greenhouse habitat successfully mitigated these external pressures by reducing predation (2.6–7.3%) and abiotic-related deaths (0–0.6%). However, mortality in the greenhouses was attributed to diseases with 46.7% in the dry season and 43.9% in the wet season (Fig. 4 C). Parasitism remained a relatively minor mortality factor across all treatments. 3.4 The influence of habitat type, season and soil substrate on G. belina pupae fate and mortality aetiology The fate of G. belina pupae was significantly influenced by the interaction of habitat, season and soil substrate (p < 0.05) (Fig. 5 ). Overall, adult moth emergence from pupae was maximised under wet-season conditions, particularly in the greenhouse environment, where emergence rates in organic and pit sand soil substrates ranged from 60–70%. In contrast, diapause was highly variable, peaking in the RS + M (river sand and mulch) substrate under greenhouse/wet conditions. The addition of mulch to soil substrates appeared to shift the pupal fate toward diapause rather than immediate moth emergence. Pupal mortality was most pronounced under dry-season treatments, with a marked increase observed in the outside (natural woodland) habitat. Pupae that were reared in the Outside-Dry-Organic (natural woodland in the dry season on organic soil substrate) habitat had the highest mortality rate at over 70% (p < 0.05). Conversely, the wet-season treatments across the greenhouse and outside (natural woodland) acted as a survival threshold, consistently suppressing mortality to below 30%. Even though the soil substrate influences pupal mortality, its effects are secondary to those of wet and seasonal conditions. Organic soil, which supported high emergence during the wet season, exhibited very high mortality during the dry season. A further post-mortem multinomial logistic regression analysis of G. belina pupal mortality aetiology revealed that both habitat (χ² = 55.79, p < 0.05) and soil substrate (χ² = 40.71, p < 0.05) significantly influenced the causes of pupal mortality (Fig. 6 ). In the greenhouse habitat, biotic factors were the major cause of morality with chalcids (parasites) being the main cause of death across all soil substrates with RS + M (river sand and mulch) accounting for 72.9% of the pupae mortality (Fig. 6 A and 6 C). Disease also contributed significantly to mortality in the greenhouse, peaking at 40.7% in RS (river sand) soil (Fig. 6 C). Disease was also a major mortality factor in the greenhouse, with its highest impact in the river sand at 40.7%; disease was lower in the outside (natural woodland) across all soil substrates. Chalcids were also the main cause of mortality in the outdoor environment across all substrates, accounting for 54.1% of mortality in pit sand and river sand substrates. However, the heat map shows a more diverse set of mortality causes in the natural woodland than in the greenhouse (Fig. 6 b and 6 D). The outside (natural woodland) environment had high odds of predation, which were nearly absent in the greenhouse, but reached 41.5% in organic soils and 40.5% in the pit sand and mulch treatment (Fig. 6 D). Ichneumonid parasites maintained a relatively consistent presence across both habitats, ranging from 7.5% to 20% (Fig. 6 C and 6 D). 4. Discussion The life-cycle analysis of G. belina revealed a precipitous decline in survivorship from egg to moth, a pattern fundamentally characteristic of a Type III survivorship curve (Kumaraswamy et al., 2023 ; Demetrius, 1978 ). This strategy is expected in G. belina , since it is typical of r-selected species that produce large numbers of offspring to offset high juvenile mortality rates (Pianka, 1970 ; Durrant, 2024 ). The high mortality of G. belina's early life stage is ecologically important for energy and nutrient flow in ecosystems, transferring resources from C. mopane trees (producers) to several predators such as insectivorous birds, reptiles and mammals, and, importantly, providing essential nutrition and livelihoods for people. The study highlights that the dry season in natural woodlands imposes a compounded mortality effect, with killing power remaining high across the egg, larval and pupal stages. The killing power (kx) analysis identified the larval stage as the primary demographic bottleneck, with a sharp spike in mortality indicating that the transition through the larval instars is the most precarious phase of the G. belina life cycle. The natural woodland exhibited a lower overall survival rate (1–3%) than the greenhouse (10%). In the natural woodland, G. belina face a wider range of mortality factors, whereas the greenhouse limits exposure by protecting it from predation, harvesting by people, and extreme weather events such as high temperatures, whirlwinds and storms (abiotic factors). In the dry season, extreme heat and desiccation stress likely impair G. belina's physiological fitness, making it less efficient at feeding and more likely to die (Mutamiswa et al., 2023 ). This is in line with previous studies by Dube et al. (2023), who found that temperatures above 35°C and relative humidity below 40% were associated with increased hatch failure and 60% mortality in G. belina . These findings suggest that the natural mopane woodland is a higher attrition habitat for G. belina to survive than the greenhouse, which acts as an essential safety net, sheltering them from harsh weather and predators that cause most deaths in nature. The protective conditions provided by the greenhouse could be replicated in the natural mopane woodland by increasing canopy cover and preserving primary mopane woodlands, which can naturally buffer larvae against weather stress and environmental exposure, thereby improving recruitment. The reproductive analysis and hatchability efficiency showed that greenhouse and wet-season conditions provide a favourable habitat for G. belina , with respect to clutch size and hatchability. However, the natural woodland habitat during the wet season provided optimal conditions for hatching success. This suggests that while the greenhouse may offer safety nets, it does not provide the optimal conditions for reproduction. Although the greenhouse provides a survival advantage, it may not reproduce the specific environmental cues required for optimal reproductive performance. This may be due to reduced light quality, limited airflow, or altered environmental cues. These factors may prevent the population from reaching the peak reproductive bursts seen in a thriving, rain-fed natural woodland. The results reveal that the larval life stage of G. belina is a critical demographic bottleneck. During the larval life stage, the aetiology of mortality shifts from extrinsic pressures (predation and abiotic factors) in the woodland to intrinsic drivers (diseases and parasitism) in the greenhouse. In natural woodland habitats, survival is severely suppressed by a combination of high-intensity predation (up to 65.6%) and abiotic stress (24.4%) during dry periods. Conversely, the greenhouse acts as a buffer, nearly eliminating predation and environmental exposure. These findings are consistent with the results of Ghanzol et al. (2006), who reported higher survival rates of G. belina larvae under shade houses and suggested that “larvae in the open probably died because of the high temperatures and poor food quality”. Despite the greenhouse's controlled advantages, the results also reveal that it triggers a compensatory mortality response through an escalation of diseases and increased parasitism, regardless of the season. Unlike natural woodlands, where fluctuating environmental conditions often suppress pathogen populations, the greenhouse environment likely provides a stable niche for pathogen proliferation. This results in elevated infection rates that persist year-round. This evidence suggests that optimising yields requires environment-specific interventions. Stringent biosecurity protocols are necessary for stabilising greenhouse production. Conversely, in natural woodlands, improving woodland canopy cover could shield G. belina from both environmental volatility and predation. The results for pupae fate and mortality aetiology show that habitat, season and soil substrate create a complex survival scenario and trade-offs, where mitigating one stressor creates another bottleneck. The wet season is important for the survival of G. belina across all habitats, as shown by mortality rates below 30%, while the dry season causes an increase in pupae mortality above 70%. Organic soil substrates are highly productive during the wet season, but they lack buffering capacity against the stressors of the long dry season. The addition of mulch to soil substrates seems to alter pupae's fate by shifting them towards diapause rather than emergence as moths. This suggests that while mulch may provide a physical buffer, it may also alter the micro-climatic cues necessary for synchronised moth emergence. The aetiology of G. belina pupae shifts with habitat, from diverse ecological pressures in the natural woodland to concentrated biotic threats in the greenhouse. The natural woodland had a more diverse set of mortality drivers, with high predation pressure, resulting in more than 40% of deaths in organic and pit sand soil substrates. On the other hand, the greenhouses had negligible predation pressure. However, this also triggered compensatory mortality from biotic intensification (diseases and parasites). In the greenhouse, chalcid parasites and disease emerged as the dominant causes of mortality, in contrast to the lower rate in the natural woodland. These differences highlight the importance of woodland and natural environments in maintaining the volatility that keeps pathogen and parasite populations in check. According to Ennos ( 2014 ), natural woodlands exposed to climate variability limit the performance of parasitic and pathogenic organisms, while removing ultraviolet light (UV) in greenhouses increases the prevalence of fungal pathogens, and stable greenhouse temperatures facilitate rapid transmission cycles (Raviv & Antignus, 2004 ). Practical Implications for G.belina Conservation and Semi-domestication The conservation of wild G. belina populations needs to focus on the management of mopane woodland, through the conservation of secondary woodland to improve canopy cover, and on the preservation of primary woodlands. A healthy canopy would act as a natural buffer, like a greenhouse, protecting the species at all life stages from environmental variability and inclement weather, while providing the optimal natural cues necessary for maximum hatching and moth emergence success, which greenhouse setups fail to replicate. Furthermore, mopane woodland must be protected from fragmentation into isolated, unconnected patches, as this would facilitate the rapid transmission of diseases and parasites while making G. belina more vulnerable to concentrated predation. For semi-domestication, the practical challenge lies in managing the high-risk, high-reward and trade-offs nature of the greenhouse habitat. While the greenhouse provides a survival safety net, it also triggers compensatory mortality through biotic intensification by parasitoids and pathogens. The shift in mortality aetiology in the greenhouse necessitates the development of rigorous sanitation and parasite-exclusion protocols throughout the life cycle. Furthermore, soil substrate management needs to be refined to ensure low mortality and high emergence rates. Organic soil is a preferred soil substrate for pupation by G. belina and shows higher emergence rates in the absence of parasitoids and disease. Thus, there is a need to develop sanitation protocols that reduce diseases and parasitism. The addition of mulch to soil substrates shifts pupal development to diapause and lesser emergence. The longer the pupae stay in diapause, the more likely they are to be exposed to mortality factors. Therefore, the use of mulch needs to be refined to avoid its likely effect on environmental and climatic cues for moth emergence, while maintaining the safety buffer it provides. 5. Conclusion This study reveals ecological trade-offs between natural mopane woodlands and semi-domesticated greenhouse environments by demonstrating how habitat, season and substrate interact to influence the life-stage-specific success of G. belina . The life-cycle analysis shows a steep decline in survivorship, with the larval life stage having the highest mortality, and the natural woodland showing higher attrition than the greenhouse. Even though the greenhouse acts as a survival safety net, it nonetheless creates a high-risk, high-reward scenario characterised by biotic intensification, which facilitates rapid pathogen transmission and concentrated parasitism in greenhouses. Wet-season conditions maximise moth emergence and reproductive output, whereas the dry season imposes a compounded mortality effect. Greenhouses were found to be effective in reducing predation and abiotic mortality, but they failed to replicate the optimal environmental cues of natural woodlands, resulting in lower hatching efficiency and moth emergence. Additionally, organic soil substrate supported the highest moth emergence during the wet season, but it did not buffer pupae during the dry season. The addition of mulch to soil substrates inadvertently shifts the developmental fate of pupae toward diapause and away from moth emergence. Ultimately, an integrated management strategy is needed that incorporates natural woodland conservation and sustainable harvesting of G. belina larvae with semi-domesticated systems. This can include utilising primary mopane woodlands as high-quality reproductive reservoirs while managing greenhouses as survival buffers. Declarations Author contributions Fortunes Felix Matutu (FM) developed the conceptual framework, designed the sampling protocols of this study and collected the data. FM, Donald Mlambo (DM) and Angela Chichinye (AC) participated in processing and interpretation, and FM wrote the paper with contributions from DM and AC. All authors have read and agreed to the published version of the manuscript. Funding This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors. Declaration of competing interests The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Clinical trial number : Not applicable Data availability Data will be made available on request. Acknowledgements The authors extend their gratitude to the Forestry Commission and its staff in Matabeleland South province for their support during data collection, including the provision of essential measuring instruments. References Bangira, C., Madimutsa, O., Manditsera, F. A., Gracian, B., Sithole, R., Mubaiwa, J., & Macheka, L. (2024). Influence of soil properties on pre-pupal burrowing, pupation and nutritional content of Gonimbrasia belina Westwood. International Journal of Tropical Insect Science , 44 (5), 2629-2635. https://doi.org/10.1007/s42690-024-01347-w Bara, G., Sithole, R., Macheka, L., 2022. 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Mopane Worms (Imbrasia Belina) Value Chain: Experiences of Rural Women on the Impact of Climate Change and Environmental Sustainability in Mangwe District. Gender and Behaviour , 22 (1), 22509-22521. https://hdl.handle.net/10520/ejc-genbeh_v22_n1_a32 Nemadodzi, L. E., Managa, G. M., & Prinsloo, G. (2023). The Use of Gonimbrasia belina (Westwood, 1849) and Cirina forda (Westwood, 1849) Caterpillars (Lepidoptera: Sarturniidae) as Food Sources and Income Generators in Africa. Foods , 12 (11), 2184. https://doi.org/10.3390/foods12112184 Nethavhani, Z., Veldtman, R., Nyamukondiwa, C., Versfeld, W., & van Asch, B. (2025). Multimarker genetic analyses of Gonimbrasia belina, the most harvested wild edible insect of mopane woodlands in Southern Africa, supports concerns over the sustainability of the species. Conservation Genetics , 26 (3), 545-559. https://doi.org/10.1007/s10592-025-01687-1 Netshanzhe, M. W., Swanepoel, C. M., Gardiner, A. J., & Swanepoel, L. H. (2025). Modelling the suitable habitat of Gonimbrasia belina , a communally exploited edible insect, in Southern African mopane ( Colophospermum mopane ) savannah. African Journal of Ecology , 63(2), e70029. https://doi.org/10.1111/aje.70029 Nunu, W. N., Ncube, B., Dube, O., Mpofu, C., Ndlovu, B., Dzinomwa, T., & Khumalo, N. (2019). Nutritional factors associated with distribution of Mopani Worms in Mopani woodlands in Tsholotsho and Gwanda Districts, Zimbabwe: A comparative survey. Scientific Reports , 9 (1), 17320. https://doi.org/10.1038/s41598-019-53923-7 Pianka, E. R. (1970). On r- and K-Selection. The American Naturalist. https://doi.org/10.1086/282697 R Core Team, 2025. R: A Language and Environment for Statistical Computing. RFoundation for Statistical Computing, Vienna. https://www.R-project.org/ Raviv, M., & Antignus, Y. (2004). UV radiation effects on pathogens and insect pests of greenhouse-grown crops. Photochemistry and photobiology, 79(3), 219–226. https://doi.org/10.1562/si-03-14.1 Rothman, L. D., & Myers, J. H. (1995). Debilitating Effects of Viral Diseases on Host Lepidoptera. Journal of Invertebrate Pathology , 67 (1), 1-10. https://doi.org/10.1006/jipa.1996.0001 Sarangi, S., Li, X., Guo, W., Tariq, K., Ullah, F., Guedes, R. N. C., ... & Lu, Y. (2026). Lepidopterans and abiotic stresses: Insights into adaptation and survival strategies. Entomologia Generalis . https://doi.org/10.1127/entomologia/2026/2241 Sharara, A., Shekede, M. D., Gwitira, I., Masocha, M., & Dube, T. (2022). Fine-scale multi-temporal and spatial analysis of agricultural drought in agro-ecological regions of Zimbabwe. Geomatics, Natural Hazards and Risk, 13(1), 1342–1365. https://doi.org/10.1080/19475705.2022.2072774 Shen, D. Y., Ferguson-Gow, H., Groner, V., Munyai, T. C., Slotow, R., & Pearson, R. G. (2023). Potential decline in the distribution and food provisioning services of the mopane worm (Gonimbrasia belina) in southern Africa. Frontiers of biogeography , 15 (2), e59408. https://doi.org/10.21425/F5FBG59408 Speight, M. R., Hunter, M. D., & Watt, A. D. (1999). Ecology of insects: concepts and applications (pp. ix+-350). https://www.cabidigitallibrary.org/doi/full/10.5555/19991111671 Togarepi, C., Nashidengo, E., & Siyambango, N. (2020). Effects of climatic variability and non-climatic factors on mopane worms'(Gonimbrasia belina) distribution and livelihood options in North Central Namibia. https://doi.org/10.5539/enrr.v10n2p14 Wen, Y., Jin, X., Zhu, C. et al. Effect of Substrate Type and Moisture on Pupation and Emergence of Heortia vitessoides (Lepidoptera: Crambidae): Choice and No-Choice Studies. J Insect Behav 29 , 473–489 (2016). https://doi.org/10.1007/s10905-016-9572-2 Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Reviewers invited by journal 16 May, 2026 Editor assigned by journal 13 Mar, 2026 Submission checks completed at journal 13 Mar, 2026 First submitted to journal 07 Mar, 2026 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-9058677","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":605571267,"identity":"2cfb8cdb-f131-4d5b-a54c-79435af0a109","order_by":0,"name":"Fortunes Felix Matutu","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA40lEQVRIiWNgGAWjYPACOQY+ZgbGB0AWDx9h1cwMDAcYjBnYmBmYDUBa2IjXwsDAJgHiE9Si237+mPTHNgM5NnYes8qvOXYybAzMDx/dwKPF7Ewym8TBNgNjNma2tNuy25KBDmMzNs7Bp+UAWMufxDZm5mO3JbcxA7XwsEnj1XL+MdiW+jZmxrZiyW31RGi5AXFYAhvQFsaP2w4To+WxscWZcwaGbcxsydKM247zAD1FwC/nEx/eqCgzkOfnP2P48ee2ant+9uaHj/FpQQHMPGCSWOUgwPiDFNWjYBSMglEwYgAAwwA+e417u5kAAAAASUVORK5CYII=","orcid":"","institution":"National University of Science and Technology","correspondingAuthor":true,"prefix":"","firstName":"Fortunes","middleName":"Felix","lastName":"Matutu","suffix":""},{"id":605571268,"identity":"56bd7aa0-a673-45c5-a108-fc7b9f4c3632","order_by":1,"name":"Donald Mlambo","email":"","orcid":"","institution":"National University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Donald","middleName":"","lastName":"Mlambo","suffix":""},{"id":605571269,"identity":"5f08aeb7-bc82-459d-9bed-942e9e4f0682","order_by":2,"name":"Angella Chichinye","email":"","orcid":"","institution":"National University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Angella","middleName":"","lastName":"Chichinye","suffix":""}],"badges":[],"createdAt":"2026-03-07 13:23:18","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-9058677/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-9058677/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":107130447,"identity":"a847a638-68c0-4026-9109-4cd5074091e3","added_by":"auto","created_at":"2026-04-17 07:01:07","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":138834,"visible":true,"origin":"","legend":"\u003cp\u003eLife-stage specific survivorship and mortality of \u003cem\u003eG. belina\u003c/em\u003e across habitats and seasons. (A, B) Survivorship (lx) curves on a logarithmic scale showing the decline from egg to moth life stages under different habitats (Greenhouse vs. Outside) and seasonal (Dry vs. Wet) conditions. (C, D) Killing Power (kx) representing stage-specific mortality intensity for the egg, larval and pupal stages.\u003c/p\u003e","description":"","filename":"image1.png","url":"https://assets-eu.researchsquare.com/files/rs-9058677/v1/49943bce4e97f742f87213ee.png"},{"id":107130448,"identity":"54caac7e-f3d5-4cdd-97d2-46f81b425bd6","added_by":"auto","created_at":"2026-04-17 07:01:07","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":41448,"visible":true,"origin":"","legend":"\u003cp\u003eReproductive output of \u003cem\u003eG. belina\u003c/em\u003e across different habitats and seasonal moisture levels. Data are presented as box-and-whisker plots showing total clutch size (grey) and the number of hatched larvae (green) in Greenhouse and Outside habitats during Dry and Wet seasons. Different lowercase letters (a, b, c, d) above the bars denote statistically significant differences between treatment groups (p \u0026lt; 0.05) based on Tukey’s HSD post-hoc test.\u003c/p\u003e","description":"","filename":"image2.png","url":"https://assets-eu.researchsquare.com/files/rs-9058677/v1/62222f61b0df565faa905aca.png"},{"id":107482034,"identity":"e50a8b35-1c4c-434f-a8d2-648e5464309c","added_by":"auto","created_at":"2026-04-22 02:21:34","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":57199,"visible":true,"origin":"","legend":"\u003cp\u003eSurvival of \u003cem\u003eG. belina\u003c/em\u003e larvae to the pupal stage across different habitats and seasons. (A) Larval survival percentage compared across dry and wet seasons within the Greenhouse and Outside habitats. (B) Comparison of larval survival between habitats categorised by wet and dry season. Different lowercase letters (a, b, c, d) above the bars denote statistically significant differences between treatment groups (p \u0026lt; 0.05) based on Tukey’s HSD post-hoc test.\u003c/p\u003e","description":"","filename":"image3.png","url":"https://assets-eu.researchsquare.com/files/rs-9058677/v1/161c4de01aa65d5eeb80919f.png"},{"id":107130450,"identity":"7eab4641-5732-41ba-9695-98ac8abffb95","added_by":"auto","created_at":"2026-04-17 07:01:07","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":78253,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003eG. belina\u003c/em\u003elarval mortality by cause, habitat, and season. (A) Stacked bar plots of causes of mortality by season. (B) Stacked mortality causes compared by habitat type. (C) Heat map showing the mean percentage of mortality attributed to each cause across habitat and season combinations. Significance letters within bar segments denote statistical differences between treatments for that specific mortality cause (p \u0026lt; 0.05).\u003c/p\u003e","description":"","filename":"image4.png","url":"https://assets-eu.researchsquare.com/files/rs-9058677/v1/93ce5f962d07bb0a800d3e59.png"},{"id":107130451,"identity":"a2421611-8953-49bf-b69e-1ea71d7c912a","added_by":"auto","created_at":"2026-04-17 07:01:07","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":123274,"visible":true,"origin":"","legend":"\u003cp\u003eThe fate of \u003cem\u003eG. belina\u003c/em\u003e pupae (%) across different soil substrates under Greenhouse and Outside (natural woodland) habitats with varying seasons (Dry and Wet). Colours represent pupal outcomes: Dead (red), Diapause (purple) and Emerged Moth (green). Different lowercase letters above boxes indicate statistically significant differences based on pairwise post hoc tests and adjusted to the equivalent of the Tukey’s HSD test within each fate category and treatment group (p \u0026lt; 0.05). RS stands for River Sand, and \"+M\" indicates the addition of mulch to the substrate.\u003c/p\u003e","description":"","filename":"image5.png","url":"https://assets-eu.researchsquare.com/files/rs-9058677/v1/9dd9a4022020266f451bfefe.png"},{"id":107130452,"identity":"e2115829-77b2-4e9c-8680-14ee0d743c64","added_by":"auto","created_at":"2026-04-17 07:01:07","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":97114,"visible":true,"origin":"","legend":"\u003cp\u003eProportional contribution of mortality factors to G. belina pupae across different soil substrates and habitats. Stacked bar plots (A, B) showing the percentage of mortality caused by Chalcids, Disease, Ichneumonids, Predators and other factors in Greenhouse (A) and Outside (natural woodland) (B) habitats. Different lowercase letters above boxes indicate statistically significant differences based on Tukey’s HSD test within each fate category and treatment group (p \u0026lt; 0.05). (C, D) Heat map visualisation of the same mortality data for the Greenhouse (C) and Outside (D) habitats. Cell values represent the mean percentage, and colour intensity reflects the magnitude of mortality. Soil substrates include Organic, Pit sand, Pitsand+M (Pit sand and mulch), RS (River sand), and RS+M (River sand and mulch).\u003c/p\u003e","description":"","filename":"image6.png","url":"https://assets-eu.researchsquare.com/files/rs-9058677/v1/a583bca0446f589c2d169cc4.png"},{"id":107705242,"identity":"7118dcd0-cd2d-44c7-8388-9c721cec5a28","added_by":"auto","created_at":"2026-04-24 09:10:02","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":608465,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-9058677/v1/3ae3e493-0c39-4a75-b83d-cf67f899fce9.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Life-Stage-Specific Survivorship and Mortality Aetiology of Gonimbrasia belina: The Interplay of Habitat Type, Seasons and Edaphic Factors","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003e \u003cem\u003eGonimbrasia belina\u003c/em\u003e, a keystone species in the mopane (\u003cem\u003eColophospermum mopane)\u003c/em\u003e woodlands of southern Africa, is a major livelihood source for the rural folk while playing a vital role in nutrient recycling in the ecosystem (Bara et al., \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2022\u003c/span\u003e; Kwiri et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2020\u003c/span\u003e; Nemadodzi et al., \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Glew et al., \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e1999\u003c/span\u003e). Despite this ecological and socio-economic importance, the species is characterised by unpredictable outbreaks, where certain seasons yield immense abundance while others result in localised near-extinction (Shen et al., \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Nethavhani et al., \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2025\u003c/span\u003e). Extensive evidence suggests that the survivorship of \u003cem\u003eG. belina\u003c/em\u003e is compromised by a suite of environmental stressors and biological threats that act across all phases of its life cycle (Sarangi et al., \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2026\u003c/span\u003e; Ghazoul, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2006\u003c/span\u003e). Further anecdotal evidence suggests that even when moths successfully lay eggs, subsequent larval and pupal recruitment can be severely limited (Ditlhogo, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e1996\u003c/span\u003e; Kwiri et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2020\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eSeveral studies have shown that deforestation, climate variability, and harvesting practices have led to lower \u003cem\u003eG. belina\u003c/em\u003e abundance and distribution (Mataboge et al., \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e2016\u003c/span\u003e; Shen et al., \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Ndlovu et al., \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). Furthermore, \u003cem\u003eG. belina's\u003c/em\u003e presence and abundance are linked to weather elements, including rainfall, temperature and humidity, with extreme weather patterns negatively impacting its abundance and life-cycle recruitment (Sekonya, 2016; Togarepi et al., \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). Studies by Ghazoul (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2006\u003c/span\u003e) identified various biotic and abiotic factors influencing \u003cem\u003eG. belina\u003c/em\u003e, including parasitoids, predation and diseases, while Dube (2013) identified the specific influence of temperature and humidity on its development. However, there remains a paucity of research examining the drivers of mortality at each life stage and their influence on ultimate survivorship. There are currently no comparative studies showing how these mortality agents vary across seasons, habitat types and soil substrates for specific cohorts.\u003c/p\u003e \u003cp\u003eBeyond the habitat condition, \u003cem\u003eG. belina\u003c/em\u003e faces various biotic mortality factors. Several studies have identified mortality factors, including diseases that are caused by pathogens such as viruses, flesh-eating bacteria and fungi (Ditlhogo, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e1996\u003c/span\u003e; Ghazoul, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2006\u003c/span\u003e; Hope et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). Furthermore, \u003cem\u003eG. belina\u003c/em\u003e is subject to several predation factors, which consume it at all stages of the life cycle, including insectivorous birds, lizards, jackals, bat-eared foxes, honey, warthogs, rats, monkeys, baboons, snakes and importantly, people (Shen et al., \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Hope et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; De Nagy Koves Hrabar, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2006\u003c/span\u003e). Additional parasites have been shown to drive mortality; the main parasites in \u003cem\u003eG. belina\u003c/em\u003e and other lepidoptera are chalcids (\u003cem\u003eMesocomys pulchriceps\u003c/em\u003e and \u003cem\u003eHockeria spp\u003c/em\u003e) and ichneumonids (\u003cem\u003eEnicospilus antancarus\u003c/em\u003e) (Ghazoul, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2006\u003c/span\u003e). However, current ecological models for \u003cem\u003eG. belina\u003c/em\u003e do not adequately account for the nuanced interplay between habitat type and seasonal fluctuations, particularly in how these factors filter specific mortality agents across the life cycle.\u003c/p\u003e \u003cp\u003eThe subterranean pupal life stage represents a critical phase during which survival and developmental fate are heavily dictated by edaphic factors (Bangira et al., \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e2024\u003c/span\u003e; Matutu et al., \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2025\u003c/span\u003e). Pupae spend a significant portion of their life cycle in the ground and have generally been found to pupate in crumbly soil substrates, including river sand, pit sands and organic soils (Mufandaedza et al., \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Wen et al., \u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). These soil substrates possess distinct physical and chemical properties and attract varying levels of biotic activity (Delgado and G\u0026oacute;mez, \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). Consequently, these substrates influence the aetiology and survivorship of the pupae differently. However, there remains a need to better understand how these differences are filtered through specific habits and seasonal changes to influence the aetiology of survival and mortality.\u003c/p\u003e \u003cp\u003eIn response to the volatility of wild \u003cem\u003eG. belina\u003c/em\u003e populations, there is increasing interest in semi-domestication through greenhouse rearing (Ghazoul, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2006\u003c/span\u003e; Hope et al., \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). However, efforts to semi-domesticate \u003cem\u003eG. belina\u003c/em\u003e in such artificial habitats have not been adequately tested and compared with natural woodlands. It remains unclear if these controlled environments truly mitigate mortality pressures or inadvertently introduce new physiological stressors when compared to natural woodlands. Therefore, there is a need to understand variations in survivorship and mortality across habitats and seasonal dynamics. This study will adopt a comparative approach to identify the optimal conditions and the physiological and environmental bottlenecks or trade-offs of the species across various habitats.\u003c/p\u003e \u003cp\u003eGiven the evident influence of biological and environmental pressures on \u003cem\u003eG. belina\u003c/em\u003e, there remains a significant gap in understanding how these pressures affect their population dynamics. Current research has primarily focused on broad aspects, such as habitat loss, climate variability, and harvesting systems, but it fails to explain how various mortality agents are filtered through the landscape and across seasons. The semi-domestication (rearing) of \u003cem\u003eG. belina\u003c/em\u003e in greenhouses introduces new dynamics that need to be tested and compared with the natural woodland benchmarks. This lack of comparative, quantitative data creates a critical bottleneck, preventing the development of truly adaptive management strategies that can effectively respond to the erratic nature of \u003cem\u003eG. belina\u003c/em\u003e outbreaks and the varying pressures across diverse landscapes. Therefore, this study aims to analyse the survivorship and life-stage-specific (eggs, larvae and pupae) mortality aetiology of \u003cem\u003eG. belina\u003c/em\u003e by evaluating how habitat and seasonal variations interact to influence reproductive fitness, larval survivorship, pupal survivorship and developmental fate. Specifically, we employ necropsy-based etiological assessments to evaluate the contributions of abiotic stress, pathogens, parasites, and predation across habitats (greenhouse vs natural woodland) and seasons (dry and wet). The study addressed the following research questions:\u003c/p\u003e \u003cp\u003e \u003col\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eHow do habitats (natural woodland vs greenhouse) and seasonal variation (dry and wet) interact to determine the life-stage-specific survivorship and mortality dynamics of \u003cem\u003eG. belina\u003c/em\u003e?\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eHow do habitats (natural woodland vs. greenhouse) and seasons (dry and wet) differentially influence the reproductive output and egg hatching efficiency of \u003cem\u003eG. belina\u003c/em\u003e?\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eHow does habitat type (natural woodland vs. greenhouse) and seasonal variation (dry vs. wet) interact to influence the filtering of abiotic factors, diseases, parasitism and predation as contributors to \u003cem\u003eG. belina\u003c/em\u003e larval mortality?\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003cspan\u003e \u003cli\u003e \u003cp\u003eHow do habitat type, season and soil substrates (River Sand, River Sand with mulch, Pit Sand, Pit Sand with mulch and Organic Soil) interact to influence the pupal mortality aetiology and developmental fate of \u003cem\u003eG. belina\u003c/em\u003e pupae?\u003c/p\u003e \u003c/li\u003e \u003c/span\u003e \u003c/ol\u003e \u003c/p\u003e"},{"header":"2. Materials and methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1. Study area\u003c/h2\u003e \u003cp\u003eThis study was done in the Gwanda district (20◦56\u0026rsquo;20\"S, 29◦01\u0026rsquo;07\"E), in southern Zimbabwe. The district is characterised as a semi-arid to arid area, with a mean annual temperature of 22\u0026deg;C and an average annual rainfall of 450 mm (Mambiravana, \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e2025\u003c/span\u003e; Baudron et al., \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). It experiences a prolonged dry season from May to November, followed by a very short rainy season from December to March. The dominant vegetation is \u003cem\u003eC. mopane\u003c/em\u003e (mopane woodland), interspersed with occasional \u003cem\u003eCombretum\u003c/em\u003e and \u003cem\u003eAcacia\u003c/em\u003e species (Kumalo and Manyani, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). The district's soils are predominantly sandy Kalahari soils, with pockets of organic soils found in areas of high biomass (Sharara et al., \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). These conditions represent the primary ecological stronghold for \u003cem\u003eG. belina\u003c/em\u003e, with the highest occurrence and harvest frequencies in Zimbabwe (Nunu et al., \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2019\u003c/span\u003e; Bara et al., \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2022\u003c/span\u003e).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2. Study species\u003c/h2\u003e \u003cp\u003e \u003cem\u003eG. belina\u003c/em\u003e is a species of emperor moth under the family Saturniidae, order Lepidoptera, and phylum Arthropoda. This economically and ecologically significant insect is native to the semi-arid regions of southern Africa. Its life cycle encompasses four distinct stages: egg, larva, pupae and adult moth. The life cycle of \u003cem\u003eG. belina\u003c/em\u003e begins with the emergence of moths, which lay eggs within 5 days (Dube and Dube, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). These eggs hatch after 7 days, and the resulting larvae moult four times (28 to 35 days) before burrowing underground. Once underground, the larvae moult a fifth time into pupae (Ditlhogo, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e1996\u003c/span\u003e). Within 4 days of burrowing, larvae are fully pupated (Dube and Dube \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). \u003cem\u003eG. belina\u003c/em\u003e then enters diapause at this stage, remaining dormant until environmental conditions suit their emergence as moths (Netshanzhe, 2025). As a bivoltine insect, \u003cem\u003eG. belina\u003c/em\u003e produces two generations annually, a trait shared by other saturniidae whose voltinism often varies by location (Ditlhogo, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e1996\u003c/span\u003e; Mogomotsi et al., \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2018\u003c/span\u003e). The first generation larvae occur between October and January (dry season to early wet season), while the second spans February to May (wet season) (Kwiri et al., \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2020\u003c/span\u003e). The bivoltism characteristic of \u003cem\u003eG. belina\u003c/em\u003e, combined with the greenhouse environment, means that successive generations develop under contrasting environmental regimes. These environmental disparities are likely to shape the patterns of survivorship and the specific drivers of mortality.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3. Study design and sampling\u003c/h2\u003e \u003cp\u003eWe used a multi-factorial experimental design to evaluate how habitat and seasonal differences interact to influence reproductive fitness, larval survivorship, pupal survivorship and developmental fate. The study area was stratified by habitat (greenhouse and natural woodland-outside) and season (wet and dry), resulting in four subsites: Greenhouse wet, Outside wet, Greenhouse dry and Outside dry. Data collection was replicated across the 2022 and 2024 bivoltine generations to ensure a substantial analysis of seasonal and habitat-driven dynamics. The sites were selected based on the historical presence of \u003cem\u003eG. belina\u003c/em\u003e in resettlement areas with low human disturbance. The controlled experiments were conducted within two purpose-built greenhouses, each measuring 15 m x 10 m and 5 m in height. These structures were covered with a 40% shade net to accurately simulate mature canopy cover. Concurrently, outside experiments were established in the adjacent natural mopane woodlands, situated within 10 meters of the greenhouses, ensuring similar macro-climatic conditions while representing natural habitat dynamics.\u003c/p\u003e \u003cp\u003eFor the egg stage assessment, a total of 200 freshly laid \u003cem\u003eG. belina\u003c/em\u003e egg clutches were selected at each of the four experimental settings. Clutch size was recorded initially, with hatching success recorded after 10 days of incubation. The larval stage investigation began with the hatchling larvae from the egg stage assessment. These larvae were subsequently observed over a 28-day period. To prevent host tree defoliation and larval migration, each tree was restricted to a single representative clutch. All additional eggs were removed and relocated to other trees. Larval mortality was recorded at 7-day intervals, with each specimen undergoing a field post-mortem examination to determine the aetiology of mortality. A diagnostic framework based on observed physical markers and remains was consistently applied to determine the cause of larval mortality (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Diseases were identified by tissue liquefaction or mummification (Gouli et al, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2011\u003c/span\u003e) while parasitism was recorded upon the presence of external eggs or cuticle exit holes (Ghazoul, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2006\u003c/span\u003e). In the absence of biotic signs, but with larvae appearing shrivelled and brittle, mortality was attributed to abiotic factors (Speight et al, \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e1999\u003c/span\u003e; Sarangi et al., \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e2026\u003c/span\u003e), whereas missing or partially consumed larvae were classified as predation (Coelho et al., \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Any ambiguous cases that lacked clear diagnostic signs were excluded from the dataset.\u003c/p\u003e \u003cp\u003eFor the pupal stage assessment, mature larvae (stage 5) were collected and introduced into controlled pupation beds. This phase of the experiment was designed as a multi-factorial study to evaluate how habitat type, season and soil substrates (River Sand, River Sand with mulch, Pit Sand, Pit Sand with mulch, and Organic Soil) interact to influence the aetiology of mortality and developmental fate of the pupae. These treatments were replicated across 180 individual pupation beds, each measuring 1 m \u0026times; 1 m \u0026times; 0.5 m. In each bed, 100 mature \u003cem\u003eG. belina\u003c/em\u003e larvae, at their pre-pupal stage, were introduced and allowed to burrow, pupate naturally and eventually emerge as moths. We monitored the population throughout the emergence season to record the number of moths emerging. At the conclusion of the emergence period, pupae were excavated to determine whether they were alive or dead. We classified and recorded the surviving pupae as being in diapause and performed necropsies on all dead specimens to identify the specific drivers of mortality (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). For pupae with shattered fragments or a total absence, mortality was attributed to predation (Coelho et al., \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Parasitism was identified by the presence of parasitoids or holes, and by distinguishing solitary ichneumonid emergence from gregarious chalcid apertures (Ghazoul, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2006\u003c/span\u003e; Gouli et al., \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). Disease was confirmed by the presence of fungal mummification, pungent odours, or tissue liquefaction under slight physical pressure (Rothman and Myers, \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e1995\u003c/span\u003e; Ghazoul, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2006\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eDiagnostic criteria and physical indicators used to determine the aetiology of mortality in \u003cem\u003eG. belina\u003c/em\u003e larvae and pupae during field surveys.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eLife Stage\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eObservation\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eDiagnostic Indicators\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eAetiology\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003eLarvae\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eWithered\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eShrivelled, dry and brittle, no odour, no liquefaction\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eAbiotic\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTissue decay or liquefaction, discolouration or mummification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eFragile/ruptured skin, Flaccid, shrunken body, milky/brownish liquid, fungal mycelia or foul odour\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eDisease: bacteria, fungi or viruses\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eExternal tags or cuticle holes\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmall white/dark eggs, Visible exit or entry holes on the larval skin\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eParasitoids\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePhysical damage or absence\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePresence of larval remains, partially consumed; disappearance\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePredators\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\" morerows=\"2\" rowspan=\"3\"\u003e \u003cp\u003ePupae\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSoftened or tissue inside decay or mummification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eDark spots/mouldy patches on cuticle; pungent odour, Mummified (hard) body with erupting fungal spores; ruptures easily under slight physical pressure.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eDisease\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eExit holes or hollowed\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmall holes in the pupal body, the shell contains a parasitoid\u003c/p\u003e \u003cp\u003eIchneumonids: Large (15-40mm); solitary; single large circular exit hole.\u003c/p\u003e \u003cp\u003eChalcids: Small (1-5mm); gregarious; multiple minute pin-hole exit apertures.\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003eParasitoids\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eShattered or absence\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eFragmented chitinous shell remains in the soil; missing pupae\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ePredators\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003e2.4 Data analysis\u003c/h2\u003e \u003cp\u003eAll statistical analyses were performed in R version 4.4.3 (R Core Team, \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2025\u003c/span\u003e). All data sets underwent quality control to assess accuracy, identify outliers, and detect missing values. The Shapiro-Wilk test was utilised to assess the normality of continuous variables, while Levene\u0026rsquo;s test was employed to evaluate the homogeneity of variances. The life-stage-specific dynamics of \u003cem\u003eG. belina\u003c/em\u003e were analysed by constructing age-specific life tables to quantify survivorship and mortality. Population data were first aggregated to calculate the mean number of individuals surviving at each developmental stage (Egg, Larvae and Pupae). Survivorship was determined by using the proportion of individuals surviving from one life stage to the next. Stage mortality rate was determined by the percentage of individuals dying during the specific stage. Killing Power was calculated as the difference between the logarithms of successive life stages. Survivorship curves were plotted on a logarithmic scale to compare population decline rates across habitats (Greenhouse vs. Outside) and seasons (Wet vs. Dry). The killing power was visualised through multi-panel, faceted plots to compare the relative killing power of different life stages across habitats.\u003c/p\u003e \u003cp\u003eThe reproductive performance of \u003cem\u003eG. belina\u003c/em\u003e, including clutch size and hatching success, was analysed using linear and generalised linear modelling. A two-way Analysis of Variance (ANOVA) was used to test the main effects of the interaction between habitat and season on clutch size and hatched eggs. A Generalised Linear Modelling (GLM) with a binomial distribution and a logit link function was used to examine the overall efficiency of moth emergence relative to initial egg counts, and to assess how these were filtered by the interaction between habit and season. Post-hoc comparisons were performed using Estimated Marginal Means (EMMs), and a Compact Letter Display (CLD) was generated using Tukey\u0026rsquo;s Honestly Significant Difference (HSD) test to identify significant differences (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) between specific treatment combinations, and the results were visualised using boxplots. The success of \u003cem\u003eG. belina\u003c/em\u003e larval development and the relative impact of various mortality drivers were analysed through a combination of frequentist modelling and predictive risk mapping. An HSD post hoc was used to evaluate the interaction between habitat and season on pupation success. For the etiological analysis of larval death, a stacked bar graph and larval mortality risk factor heatmap were constructed by aggregating total mortality percentages by cause and habitat, visualised as a gradient-filled tile matrix.\u003c/p\u003e \u003cp\u003eThe developmental fate of \u003cem\u003eG. belina\u003c/em\u003e pupae (emerged moth, diapause, or mortality) was analysed using a three-way ANOVA to determine the interactive effects of habitat, season and soil substrate. To identify specific differences between treatment combinations, post hoc pairwise comparisons were done using the HSD test (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). These results were visualised as faceted boxplots, with compact letter displays indicating statistical significance within each fate category. To further investigate the specific drivers of pupal death, a post-mortem etiological analysis was performed using a multinomial logistic regression. The significance of habitat and soil substrate as predictors of these causes was assessed using Type II Likelihood Ratio Tests. The proportional contributions of each mortality factor were visualised through stacked bar plots to illustrate compositional shifts between habitats, while heat map matrices were used to visualise the intensity and diversity of mortality risks across soil substrates.\u003c/p\u003e \u003c/div\u003e"},{"header":"3. Results","content":"\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003e3.1 Effects of Habitat and Season on G. belina Survivorship and Mortality\u003c/h2\u003e \u003cp\u003eThe life cycle analysis of \u003cem\u003eG. belina\u003c/em\u003e showed that survivorship (lx) and killing power (kx) are significantly influenced by the interaction of habitat (Greenhouse and Outside) and season (Wet and Dry). Generalised Linear Modelling (GLM) results for overall survival to the moth (adult) stage showed that the odds of \u003cem\u003eG. belina\u003c/em\u003e surviving from egg to moth were significantly lower for individuals raised outside (natural woodland) than for those raised in the greenhouse (β = -1.52, p\u0026thinsp;\u0026lt;\u0026thinsp;0.001). However, the wet season was a significant positive driver of survival (β\u0026thinsp;=\u0026thinsp;1.14, p\u0026thinsp;\u0026lt;\u0026thinsp;0.001). The Outside (natural woodland) habitat exhibited a survival disadvantage that was significantly mitigated during the wet season (β\u0026thinsp;=\u0026thinsp;0.31, p\u0026thinsp;\u0026lt;\u0026thinsp;0.001).\u003c/p\u003e \u003cp\u003eSurvivorship (lx) declined across all cohorts as they progressed from egg to moth, resulting in a low percentage of individuals reaching the moth stage (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA and \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB). However, the probability of survival from egg to the moth stage was consistently higher in the greenhouse, at an average of 10%, whereas survival in the outside environment was severely depressed, between 1% and 3%. This disparity was mostly pronounced during the dry season, when populations on the outside (natural woodland) experienced the steepest decline, resulting in negligible recruitment to the moth stage.\u003c/p\u003e \u003cp\u003eAnalysis of killing power (kx) revealed the larval stage as the primary developmental bottleneck for individuals outside the greenhouse, as evidenced by a sharp spike in mortality (kx\u0026thinsp;=\u0026thinsp;0.8) across seasons (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eC and \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eD). While the wet season generally improved survivorship across both habitats, the greenhouse appeared to stabilise mortality rates across all life stages (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB and \u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eD). Furthermore, the combination of external (natural woodland) habitats and dry-season conditions exerted high mortality across the egg, larval, and pupal stages simultaneously, suggesting that environmental exposure and dry weather conditions compound to severely limit population persistence.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec9\" class=\"Section2\"\u003e \u003ch2\u003e3.2 Influence of Habitat Type and Season on G. belina Egg Clutch Size and Hatching Efficiency\u003c/h2\u003e \u003cp\u003eThe analysis of egg clutch size and hatching outputs of \u003cem\u003eG. belina\u003c/em\u003e showed that they were significantly influenced by the interaction between habitat and season (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). While both habitats (greenhouse and natural woodland; p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) and season (wet and dry; p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) independently affected clutch size and larval hatching, the impact of season (wet or dry) was significantly moderated by the greenhouse habitat. During the dry season, the greenhouse habitat exhibited significantly larger clutch and larval counts than the outside (natural woodland) habitat, which was highly sensitive to seasonal variation. The transition from dry to wet season increased clutch size and hatching efficiency in the natural woodland, but remained constant in the greenhouse. The beneficial effect of the wet season was also disproportionately greater in the outside (natural woodlands) habitat than in the greenhouse, with outside-wet having greater hatching success than greenhouse-wet. In contrast, the outside (natural woodland) habitat exhibited a clear egg laying and hatching efficiency deficit during the dry season, which was largely mitigated during the wet season (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec10\" class=\"Section2\"\u003e \u003ch2\u003e3.3 Influence of Habitat Type and Seasons on G. belina Larval Mortality\u003c/h2\u003e \u003cp\u003eThe survival of \u003cem\u003eG. belina\u003c/em\u003e larvae to the pupae life stage was significantly influenced by both rearing habitat and seasonal timing. Larvae growing in the greenhouse had higher survival rates across both seasons compared to those reared in an outside (natural woodland) habitat (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA and \u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB). Within the greenhouse, survival was significantly higher during the wet season (median\u0026thinsp;\u0026asymp;\u0026thinsp;57%) compared to the dry season (median\u0026thinsp;\u0026asymp;\u0026thinsp;45%; p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA). In contrast, the outside (natural woodland) habitat recorded a high mortality, with survival rates consistently suppressed. In both the wet and dry seasons, the outside habitat was heavily skewed towards total mortality, with only a few outliers showing high survival (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB).\u003c/p\u003e \u003cp\u003eThe differences in larval survival between habitats and seasons were driven by a distinct aetiology of mortality (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e). In the outside (natural woodland) habitat, predation was the primary driver of larval death, contributing 46.6% of mortality in the dry season and 65.6% in the wet season. The larval loss was further exacerbated by substantial abiotic (24.4%) and disease (16.1%) losses during the dry period. On the other hand, the greenhouse habitat successfully mitigated these external pressures by reducing predation (2.6\u0026ndash;7.3%) and abiotic-related deaths (0\u0026ndash;0.6%). However, mortality in the greenhouses was attributed to diseases with 46.7% in the dry season and 43.9% in the wet season (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eC). Parasitism remained a relatively minor mortality factor across all treatments.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003cem\u003e3.4 The influence of habitat type, season and soil substrate on G. belina pupae fate and mortality aetiology\u003c/em\u003e \u003c/p\u003e \u003cp\u003eThe fate of \u003cem\u003eG. belina\u003c/em\u003e pupae was significantly influenced by the interaction of habitat, season and soil substrate (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e). Overall, adult moth emergence from pupae was maximised under wet-season conditions, particularly in the greenhouse environment, where emergence rates in organic and pit sand soil substrates ranged from 60\u0026ndash;70%. In contrast, diapause was highly variable, peaking in the RS\u0026thinsp;+\u0026thinsp;M (river sand and mulch) substrate under greenhouse/wet conditions. The addition of mulch to soil substrates appeared to shift the pupal fate toward diapause rather than immediate moth emergence. Pupal mortality was most pronounced under dry-season treatments, with a marked increase observed in the outside (natural woodland) habitat. Pupae that were reared in the Outside-Dry-Organic (natural woodland in the dry season on organic soil substrate) habitat had the highest mortality rate at over 70% (p\u0026thinsp;\u0026lt;\u0026thinsp;0.05). Conversely, the wet-season treatments across the greenhouse and outside (natural woodland) acted as a survival threshold, consistently suppressing mortality to below 30%. Even though the soil substrate influences pupal mortality, its effects are secondary to those of wet and seasonal conditions. Organic soil, which supported high emergence during the wet season, exhibited very high mortality during the dry season.\u003c/p\u003e \u003cp\u003eA further post-mortem multinomial logistic regression analysis of \u003cem\u003eG. belina\u003c/em\u003e pupal mortality aetiology revealed that both habitat (χ\u0026sup2; = 55.79, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) and soil substrate (χ\u0026sup2; = 40.71, p\u0026thinsp;\u0026lt;\u0026thinsp;0.05) significantly influenced the causes of pupal mortality (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e). In the greenhouse habitat, biotic factors were the major cause of morality with chalcids (parasites) being the main cause of death across all soil substrates with RS\u0026thinsp;+\u0026thinsp;M (river sand and mulch) accounting for 72.9% of the pupae mortality (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eA and \u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eC). Disease also contributed significantly to mortality in the greenhouse, peaking at 40.7% in RS (river sand) soil (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eC). Disease was also a major mortality factor in the greenhouse, with its highest impact in the river sand at 40.7%; disease was lower in the outside (natural woodland) across all soil substrates. Chalcids were also the main cause of mortality in the outdoor environment across all substrates, accounting for 54.1% of mortality in pit sand and river sand substrates. However, the heat map shows a more diverse set of mortality causes in the natural woodland than in the greenhouse (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eb and \u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eD). The outside (natural woodland) environment had high odds of predation, which were nearly absent in the greenhouse, but reached 41.5% in organic soils and 40.5% in the pit sand and mulch treatment (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eD). Ichneumonid parasites maintained a relatively consistent presence across both habitats, ranging from 7.5% to 20% (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eC and \u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eD).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"4. Discussion","content":"\u003cp\u003eThe life-cycle analysis of \u003cem\u003eG. belina\u003c/em\u003e revealed a precipitous decline in survivorship from egg to moth, a pattern fundamentally characteristic of a Type III survivorship curve (Kumaraswamy et al., \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e2023\u003c/span\u003e; Demetrius, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e1978\u003c/span\u003e). This strategy is expected in \u003cem\u003eG. belina\u003c/em\u003e, since it is typical of r-selected species that produce large numbers of offspring to offset high juvenile mortality rates (Pianka, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e1970\u003c/span\u003e; Durrant, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). The high mortality of \u003cem\u003eG. belina's\u003c/em\u003e early life stage is ecologically important for energy and nutrient flow in ecosystems, transferring resources from \u003cem\u003eC. mopane\u003c/em\u003e trees (producers) to several predators such as insectivorous birds, reptiles and mammals, and, importantly, providing essential nutrition and livelihoods for people.\u003c/p\u003e \u003cp\u003eThe study highlights that the dry season in natural woodlands imposes a compounded mortality effect, with killing power remaining high across the egg, larval and pupal stages. The killing power (kx) analysis identified the larval stage as the primary demographic bottleneck, with a sharp spike in mortality indicating that the transition through the larval instars is the most precarious phase of the \u003cem\u003eG. belina\u003c/em\u003e life cycle. The natural woodland exhibited a lower overall survival rate (1\u0026ndash;3%) than the greenhouse (10%). In the natural woodland, \u003cem\u003eG. belina\u003c/em\u003e face a wider range of mortality factors, whereas the greenhouse limits exposure by protecting it from predation, harvesting by people, and extreme weather events such as high temperatures, whirlwinds and storms (abiotic factors). In the dry season, extreme heat and desiccation stress likely impair \u003cem\u003eG. belina's\u003c/em\u003e physiological fitness, making it less efficient at feeding and more likely to die (Mutamiswa et al., \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). This is in line with previous studies by Dube et al. (2023), who found that temperatures above 35\u0026deg;C and relative humidity below 40% were associated with increased hatch failure and 60% mortality in \u003cem\u003eG. belina\u003c/em\u003e. These findings suggest that the natural mopane woodland is a higher attrition habitat for \u003cem\u003eG. belina\u003c/em\u003e to survive than the greenhouse, which acts as an essential safety net, sheltering them from harsh weather and predators that cause most deaths in nature. The protective conditions provided by the greenhouse could be replicated in the natural mopane woodland by increasing canopy cover and preserving primary mopane woodlands, which can naturally buffer larvae against weather stress and environmental exposure, thereby improving recruitment.\u003c/p\u003e \u003cp\u003eThe reproductive analysis and hatchability efficiency showed that greenhouse and wet-season conditions provide a favourable habitat for \u003cem\u003eG. belina\u003c/em\u003e, with respect to clutch size and hatchability. However, the natural woodland habitat during the wet season provided optimal conditions for hatching success. This suggests that while the greenhouse may offer safety nets, it does not provide the optimal conditions for reproduction. Although the greenhouse provides a survival advantage, it may not reproduce the specific environmental cues required for optimal reproductive performance. This may be due to reduced light quality, limited airflow, or altered environmental cues. These factors may prevent the population from reaching the peak reproductive bursts seen in a thriving, rain-fed natural woodland.\u003c/p\u003e \u003cp\u003eThe results reveal that the larval life stage of \u003cem\u003eG. belina\u003c/em\u003e is a critical demographic bottleneck. During the larval life stage, the aetiology of mortality shifts from extrinsic pressures (predation and abiotic factors) in the woodland to intrinsic drivers (diseases and parasitism) in the greenhouse. In natural woodland habitats, survival is severely suppressed by a combination of high-intensity predation (up to 65.6%) and abiotic stress (24.4%) during dry periods. Conversely, the greenhouse acts as a buffer, nearly eliminating predation and environmental exposure. These findings are consistent with the results of Ghanzol et al. (2006), who reported higher survival rates of \u003cem\u003eG. belina\u003c/em\u003e larvae under shade houses and suggested that \u0026ldquo;larvae in the open probably died because of the high temperatures and poor food quality\u0026rdquo;. Despite the greenhouse's controlled advantages, the results also reveal that it triggers a compensatory mortality response through an escalation of diseases and increased parasitism, regardless of the season. Unlike natural woodlands, where fluctuating environmental conditions often suppress pathogen populations, the greenhouse environment likely provides a stable niche for pathogen proliferation. This results in elevated infection rates that persist year-round. This evidence suggests that optimising yields requires environment-specific interventions. Stringent biosecurity protocols are necessary for stabilising greenhouse production. Conversely, in natural woodlands, improving woodland canopy cover could shield \u003cem\u003eG. belina\u003c/em\u003e from both environmental volatility and predation.\u003c/p\u003e \u003cp\u003eThe results for pupae fate and mortality aetiology show that habitat, season and soil substrate create a complex survival scenario and trade-offs, where mitigating one stressor creates another bottleneck. The wet season is important for the survival of \u003cem\u003eG. belina\u003c/em\u003e across all habitats, as shown by mortality rates below 30%, while the dry season causes an increase in pupae mortality above 70%. Organic soil substrates are highly productive during the wet season, but they lack buffering capacity against the stressors of the long dry season. The addition of mulch to soil substrates seems to alter pupae's fate by shifting them towards diapause rather than emergence as moths. This suggests that while mulch may provide a physical buffer, it may also alter the micro-climatic cues necessary for synchronised moth emergence.\u003c/p\u003e \u003cp\u003eThe aetiology of \u003cem\u003eG. belina\u003c/em\u003e pupae shifts with habitat, from diverse ecological pressures in the natural woodland to concentrated biotic threats in the greenhouse. The natural woodland had a more diverse set of mortality drivers, with high predation pressure, resulting in more than 40% of deaths in organic and pit sand soil substrates. On the other hand, the greenhouses had negligible predation pressure. However, this also triggered compensatory mortality from biotic intensification (diseases and parasites). In the greenhouse, chalcid parasites and disease emerged as the dominant causes of mortality, in contrast to the lower rate in the natural woodland. These differences highlight the importance of woodland and natural environments in maintaining the volatility that keeps pathogen and parasite populations in check. According to Ennos (\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e2014\u003c/span\u003e), natural woodlands exposed to climate variability limit the performance of parasitic and pathogenic organisms, while removing ultraviolet light (UV) in greenhouses increases the prevalence of fungal pathogens, and stable greenhouse temperatures facilitate rapid transmission cycles (Raviv \u0026amp; Antignus, \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2004\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cb\u003ePractical Implications for\u003c/b\u003e \u003cb\u003eG.belina\u003c/b\u003e \u003cb\u003eConservation and Semi-domestication\u003c/b\u003e\u003c/p\u003e \u003cp\u003eThe conservation of wild \u003cem\u003eG. belina\u003c/em\u003e populations needs to focus on the management of mopane woodland, through the conservation of secondary woodland to improve canopy cover, and on the preservation of primary woodlands. A healthy canopy would act as a natural buffer, like a greenhouse, protecting the species at all life stages from environmental variability and inclement weather, while providing the optimal natural cues necessary for maximum hatching and moth emergence success, which greenhouse setups fail to replicate. Furthermore, mopane woodland must be protected from fragmentation into isolated, unconnected patches, as this would facilitate the rapid transmission of diseases and parasites while making \u003cem\u003eG. belina\u003c/em\u003e more vulnerable to concentrated predation.\u003c/p\u003e \u003cp\u003eFor semi-domestication, the practical challenge lies in managing the high-risk, high-reward and trade-offs nature of the greenhouse habitat. While the greenhouse provides a survival safety net, it also triggers compensatory mortality through biotic intensification by parasitoids and pathogens. The shift in mortality aetiology in the greenhouse necessitates the development of rigorous sanitation and parasite-exclusion protocols throughout the life cycle. Furthermore, soil substrate management needs to be refined to ensure low mortality and high emergence rates. Organic soil is a preferred soil substrate for pupation by \u003cem\u003eG. belina\u003c/em\u003e and shows higher emergence rates in the absence of parasitoids and disease. Thus, there is a need to develop sanitation protocols that reduce diseases and parasitism. The addition of mulch to soil substrates shifts pupal development to diapause and lesser emergence. The longer the pupae stay in diapause, the more likely they are to be exposed to mortality factors. Therefore, the use of mulch needs to be refined to avoid its likely effect on environmental and climatic cues for moth emergence, while maintaining the safety buffer it provides.\u003c/p\u003e"},{"header":"5. Conclusion","content":"\u003cp\u003eThis study reveals ecological trade-offs between natural mopane woodlands and semi-domesticated greenhouse environments by demonstrating how habitat, season and substrate interact to influence the life-stage-specific success of \u003cem\u003eG. belina\u003c/em\u003e. The life-cycle analysis shows a steep decline in survivorship, with the larval life stage having the highest mortality, and the natural woodland showing higher attrition than the greenhouse. Even though the greenhouse acts as a survival safety net, it nonetheless creates a high-risk, high-reward scenario characterised by biotic intensification, which facilitates rapid pathogen transmission and concentrated parasitism in greenhouses. Wet-season conditions maximise moth emergence and reproductive output, whereas the dry season imposes a compounded mortality effect. Greenhouses were found to be effective in reducing predation and abiotic mortality, but they failed to replicate the optimal environmental cues of natural woodlands, resulting in lower hatching efficiency and moth emergence. Additionally, organic soil substrate supported the highest moth emergence during the wet season, but it did not buffer pupae during the dry season. The addition of mulch to soil substrates inadvertently shifts the developmental fate of pupae toward diapause and away from moth emergence. Ultimately, an integrated management strategy is needed that incorporates natural woodland conservation and sustainable harvesting of \u003cem\u003eG. belina\u003c/em\u003e larvae with semi-domesticated systems. This can include utilising primary mopane woodlands as high-quality reproductive reservoirs while managing greenhouses as survival buffers.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAuthor contributions\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFortunes Felix Matutu (FM) developed the conceptual framework, designed the sampling protocols of this study and collected the data. FM, Donald Mlambo (DM) and Angela Chichinye (AC) participated in processing and interpretation, and FM wrote the paper with contributions from DM and AC. All authors have read and agreed to the published version of the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDeclaration of competing interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eClinical trial number\u003c/strong\u003e:\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eData will be made available on request.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors extend their gratitude to the Forestry Commission and its staff in Matabeleland South province for their support during data collection, including the provision of essential measuring instruments.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eBangira, C., Madimutsa, O., Manditsera, F. 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Ecology of insects: concepts and applications (pp. ix+-350). https://www.cabidigitallibrary.org/doi/full/10.5555/19991111671\u003c/li\u003e\n\u003cli\u003eTogarepi, C., Nashidengo, E., \u0026amp; Siyambango, N. (2020). Effects of climatic variability and non-climatic factors on mopane worms\u0026apos;(Gonimbrasia belina) distribution and livelihood options in North Central Namibia. https://doi.org/10.5539/enrr.v10n2p14\u003c/li\u003e\n\u003cli\u003eWen, Y., Jin, X., Zhu, C. \u003cem\u003eet al.\u003c/em\u003e Effect of Substrate Type and Moisture on Pupation and Emergence of \u003cem\u003eHeortia vitessoides\u003c/em\u003e (Lepidoptera: Crambidae): Choice and No-Choice Studies. \u003cem\u003eJ Insect Behav\u003c/em\u003e \u003cstrong\u003e29\u003c/strong\u003e, 473\u0026ndash;489 (2016). https://doi.org/10.1007/s10905-016-9572-2 \u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"international-journal-of-tropical-insect-science","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jtis","sideBox":"Learn more about [International Journal of Tropical Insect Science](http://link.springer.com/journal/42690)","snPcode":"42690","submissionUrl":"https://www.editorialmanager.com/jtis/default2.aspx","title":"International Journal of Tropical Insect Science","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Gonimbrasia belina, Greenhouse, Mortality aetiology, Season, Survivorship, Woodland","lastPublishedDoi":"10.21203/rs.3.rs-9058677/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-9058677/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cem\u003eGonimbrasia belina\u003c/em\u003e (mopane worms) is a keystone species in the mopane woodland, yet its population dynamics remain unpredictable and poorly understood. This study aims to analyse the survivorship and mortality aetiology of \u003cem\u003eG. belina\u003c/em\u003e by evaluating how habitat and seasonal variations interact to influence reproductive fitness, larval and pupal survivorship. The study used multi-factorial experiments and necropsy-based diagnostic frameworks across four sub-sites representing a combination of habitat and season, and the addition of five soil substrates for pupae. The greenhouse increased the overall survivorship (10%) of \u003cem\u003eG. belina\u003c/em\u003e compared to natural mopane woodlands (1%). The greenhouse mitigated extrinsic mortality pressure from predation and abiotic stress, but it inadvertently created intrinsic pressures, especially disease and parasitoid-related mortality. The larval stage had the highest killing power, especially in the natural woodland, where predation is severe (65.6%). Pupae were influenced by seasons and soil substrate, with organic soils supporting high moth emergence in the wet season but with high mortality during the dry season (70%), while the addition of mulch to the substrate increased diapause rates. The study shows that natural woodlands under wet conditions serve as reproductive reservoirs where \u003cem\u003eG. belina\u003c/em\u003e produces the best. Greenhouses protect against predation and weather, but they attract diseases and parasitoids. The conservation and domestication of \u003cem\u003eG. belina\u003c/em\u003e require an integrated approach that preserves natural woodland to enhance productivity. Semi-domestication systems can be utilised as protective nurseries to mitigate predation and abiotic bottlenecks, but there is a need to develop bio-sanitation protocols to protect against pathogens and parasitoids.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eClinical trial number:\u003c/strong\u003e not applicable.\u003c/p\u003e","manuscriptTitle":"Life-Stage-Specific Survivorship and Mortality Aetiology of Gonimbrasia belina: The Interplay of Habitat Type, Seasons and Edaphic Factors","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-04-17 07:01:03","doi":"10.21203/rs.3.rs-9058677/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"reviewersInvited","content":"","date":"2026-05-16T19:15:33+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-03-13T09:49:50+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-03-13T09:49:45+00:00","index":"","fulltext":""},{"type":"submitted","content":"International Journal of Tropical Insect Science","date":"2026-03-07T13:17:17+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"international-journal-of-tropical-insect-science","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jtis","sideBox":"Learn more about [International Journal of Tropical Insect Science](http://link.springer.com/journal/42690)","snPcode":"42690","submissionUrl":"https://www.editorialmanager.com/jtis/default2.aspx","title":"International Journal of Tropical Insect Science","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"3d8602e9-dac2-448a-8adb-d1a1d08e1cc2","owner":[],"postedDate":"April 17th, 2026","published":true,"recentEditorialEvents":[{"type":"reviewersInvited","content":"20","date":"2026-05-16T19:15:33+00:00","index":"","fulltext":""}],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-05-16T19:23:13+00:00","versionOfRecord":[],"versionCreatedAt":"2026-04-17 07:01:03","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-9058677","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-9058677","identity":"rs-9058677","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}