Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers

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Saud" }, { "@type": "Person", "name": "Ahmed Emad Abood" }, { "@type": "Person", "name": "Maysam Abdulrahman Ghazi" } ], "publisher": { "@type": "Organization", "name": "F1000Research", "logo": { "@type": "ImageObject", "url": "https://f1000research.com/img/AMP/F1000Research_image.png", "height": 480, "width": 60 } }, "image": { "@type": "ImageObject", "url": "https://f1000research.com/img/AMP/F1000Research_image.png", "height": 1200, "width": 150 }, "description": " Background Ruminal impaction is a serious digestive disorder in sheep, leading to systemic metabolic acidosis and, as a consequence, widespread physiological disturbances. Thus, the objective of this study was to estimate biochemical, electrolyte and oxidative parameters in a local Iraqi sheep that was suffering from ruminal impaction. Methods A case control study was performed in 20 adult female sheep (Awassi sheep breeder) (10 ruminal impaction and 10 control) diagnosed with in Fallujah, Iraq, based on clinical signs and confirmed by laboratory findings of hyperkalaemia. The control group consisted of sheep of age, sex and weight consistent with clinically normal sheep. At the end of the experiment, blood samples were collected, and serum was analyzed for electrolytes, biochemical and oxidative markers using established laboratory methods. Results The impacted animals showed significant increases (P≤0.05) in alanine aminotransferase (ALT), aspartate Aminotransferase (AST), glucose, urea, creatinine, potassium, and phosphate and significant decreases (P≤0.05) in albumin, cholesterol, sodium, chloride, and calcium levels, where serum level of magnesium did not show any significant difference between the groups. In addition, oxidative stress was present as there was a significant decrease in reduced glutathione (GSH) level and superoxide dismutase (SOD) activity, and a significant increase in malondialdehyde (MDA) level. Background These findings indicate a pathogenesis cascade whereby ingestion of poorly digested food causes ruminal stasis, systemic acidosis, liver dysfunction, renal insufficiency, electrolyte disturbances, oxidative stress and oxidative stress. Using a single biomarker may underestimate the true severity of the disease, while combining biochemical, electrolyte and oxidative measurements provides a more complete picture. 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F1000Research 2026, 15 :110 ( https://doi.org/10.12688/f1000research.174402.2 ) NOTE: If applicable, it is important to ensure the information in square brackets after the title is included in all citations of this article. Close Copy Citation Details Export Export Citation Sciwheel EndNote Ref. Manager Bibtex ProCite Sente EXPORT Select a format first Track Share ▬ ✚ Research Article Revised Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] Sabea Khamees Abed https://orcid.org/0009-0005-6639-0384 1 , Mustafa A. Saud 1 , Ahmed Emad Abood 1 , Maysam Abdulrahman Ghazi https://orcid.org/0000-0002-8534-726X 1 Sabea Khamees Abed https://orcid.org/0009-0005-6639-0384 1 , Mustafa A. Saud 1 , Ahmed Emad Abood 1 , Maysam Abdulrahman Ghazi https://orcid.org/0000-0002-8534-726X 1 PUBLISHED 31 Mar 2026 Author details Author details 1 College of Veterinary Medicine, University of Fallujah, Al-Fallujah, Al Anbar Governorate, Iraq Sabea Khamees Abed Roles: Conceptualization, Formal Analysis, Investigation, Methodology, Project Administration, Supervision, Writing – Original Draft Preparation Mustafa A. Saud Roles: Data Curation, Formal Analysis, Investigation, Visualization, Writing – Review & Editing Ahmed Emad Abood Roles: Data Curation, Investigation, Resources, Validation, Writing – Review & Editing Maysam Abdulrahman Ghazi Roles: Formal Analysis, Software, Validation, Visualization, Writing – Review & Editing OPEN PEER REVIEW DETAILS REVIEWER STATUS This article is included in the Fallujah Multidisciplinary Science and Innovation gateway. Abstract Background Ruminal impaction is a serious digestive disorder in sheep, leading to systemic metabolic acidosis and, as a consequence, widespread physiological disturbances. Thus, the objective of this study was to estimate biochemical, electrolyte and oxidative parameters in a local Iraqi sheep that was suffering from ruminal impaction. Methods A case control study was performed in 20 adult female sheep (Awassi sheep breeder) (10 ruminal impaction and 10 control) diagnosed with in Fallujah, Iraq, based on clinical signs and confirmed by laboratory findings of hyperkalaemia. The control group consisted of sheep of age, sex and weight consistent with clinically normal sheep. At the end of the experiment, blood samples were collected, and serum was analyzed for electrolytes, biochemical and oxidative markers using established laboratory methods. Results The impacted animals showed significant increases (P≤0.05) in alanine aminotransferase (ALT), aspartate Aminotransferase (AST), glucose, urea, creatinine, potassium, and phosphate and significant decreases (P≤0.05) in albumin, cholesterol, sodium, chloride, and calcium levels, where serum level of magnesium did not show any significant difference between the groups. In addition, oxidative stress was present as there was a significant decrease in reduced glutathione (GSH) level and superoxide dismutase (SOD) activity, and a significant increase in malondialdehyde (MDA) level. Background These findings indicate a pathogenesis cascade whereby ingestion of poorly digested food causes ruminal stasis, systemic acidosis, liver dysfunction, renal insufficiency, electrolyte disturbances, oxidative stress and oxidative stress. Using a single biomarker may underestimate the true severity of the disease, while combining biochemical, electrolyte and oxidative measurements provides a more complete picture. READ ALL READ LESS Keywords Keywords: ruminal impaction; sheep; metabolic acidosis; oxidative stress Corresponding Author(s) Sabea Khamees Abed ( [email protected] ) Mustafa A. Saud ( [email protected] ) Close Corresponding authors: Sabea Khamees Abed, Mustafa A. Saud Competing interests: No competing interests were disclosed. Grant information: The author(s) declared that no grants were involved in supporting this work. Copyright: © 2026 Khamees Abed S et al . This is an open access article distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. How to cite: Khamees Abed S, A. Saud M, Emad Abood A and Abdulrahman Ghazi M. Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.12688/f1000research.174402.2 ) First published: 23 Jan 2026, 15 :110 ( https://doi.org/10.12688/f1000research.174402.1 ) Latest published: 31 Mar 2026, 15 :110 ( https://doi.org/10.12688/f1000research.174402.2 ) Revised Amendments from Version 1 This revised version of the manuscript presents several significant improvements over the previously published version. The study design and case characterization have been clarified, with detailed information added regarding the age, physiological status, and clinical condition of the animals. It is explicitly stated that ruminal impaction cases were naturally occurring, and control animals were clinically healthy with biochemical values within physiological reference ranges. The statistical analysis has been strengthened: data normality was assessed using the Shapiro–Wilk test, independent samples t-tests were applied, and the false discovery rate (FDR) was controlled using the Benjamini–Hochberg procedure. Tables have been updated to report accurate p-values and correct units for phosphate. In the discussion, causal interpretations have been removed, and mechanistic explanations are now presented as hypotheses supported by prior literature. Minor clarifications were also added, such as confirming that blood samples were analyzed individually and addressing inferred metabolic acidosis based on established pathophysiology. Collectively, these revisions enhance the methodological rigor, transparency, and scientific clarity of the study, ensuring that the results are presented accurately and reliably. This revised version of the manuscript presents several significant improvements over the previously published version. The study design and case characterization have been clarified, with detailed information added regarding the age, physiological status, and clinical condition of the animals. It is explicitly stated that ruminal impaction cases were naturally occurring, and control animals were clinically healthy with biochemical values within physiological reference ranges. The statistical analysis has been strengthened: data normality was assessed using the Shapiro–Wilk test, independent samples t-tests were applied, and the false discovery rate (FDR) was controlled using the Benjamini–Hochberg procedure. Tables have been updated to report accurate p-values and correct units for phosphate. In the discussion, causal interpretations have been removed, and mechanistic explanations are now presented as hypotheses supported by prior literature. Minor clarifications were also added, such as confirming that blood samples were analyzed individually and addressing inferred metabolic acidosis based on established pathophysiology. Collectively, these revisions enhance the methodological rigor, transparency, and scientific clarity of the study, ensuring that the results are presented accurately and reliably. See the authors' detailed response to the review by Şevket Evci See the authors' detailed response to the review by Jinxin Liu READ REVIEWER RESPONSES Introduction Ruminal impaction is a severe digestive condition in ruminants caused by excessive accumulation and hardening of ingesta in the rumen, leading to distention and functional stasis. In sheep, this is generally related to the ingestion of poor-quality, fibrous, or indigestible feed, leading to short-chain fatty acid (SCFA) generation, which can exceed absorption, and ruminal pH becomes depressed, especially in periods of nutritional stress or inadequate management ( Elmeligy et al. , 2025 ). In addition to its mechanical effects, ruminal impaction induces a cascade of systemic disorders, including diarrhea, severe dehydration, electrolyte disturbances, acid-base imbalance, and endotoxemia, which all compromise animal health and welfare and contribute to significant economic losses in small ruminant production systems ( Constable et al. , 2016 ; Voulgarakis et al. , 2023 ). The pathophysiological effects begin with interrupting normal ruminal motility and fermentation of the microflora. Alterations in the ruminal contents disrupt the normal symbiotic relationship between the host and its resident microflora, leading to an imbalance within the microbial community. This dysbiosis promotes the overgrowth of pathogenic species, releasing endotoxins into the digestive environment. At the same time, reduced food intake and fluid retention in the gastrointestinal tract lead to severe dehydration and haemoconcentration due to diarrhea ( Smith, 2014 ; Kumar et al. , 2019 ). The resulting hypovolaemia due to osmotic pressure impaired peripheral circulation and renal perfusion, predisposing the animal to pre-existing azotemia and altering electrolyte homeostasis, e.g., decreased concentration of strong cations (Lactic acid) or increased concentration of weak cations ( Molitoris, 2022 ). The central cause of the clinical defect in animals is biochemical and electrolyte disturbances ( Gał ę ska et al. , 2022 ). Reduced renal filtration rate increases urea nitrogen (BUN) and creatinine blood levels, which reflect impaired renal function. Furthermore, decreased salivary secretion, usually the primary source of sodium bicarbonate and phosphate, contributes to systemic acidosis and electrolyte disturbances, particularly hyponatraemia, hyperkalaemia and hypochloremia ( Elnady et al. , 2019 ). These ionic imbalances impair neuromuscular function and exacerbate systemic dysfunction, increasing the clinical impact due to an increase in the anion gap due to lactate acting as a non-measurable anion and lowering the measurable anion ( Hernández et al. , 2014 ). Oxidative stress is increasingly recognized as a key factor in ruminant gastrointestinal pathogenesis. Systemic inflammatory reactions and endotoxin release are associated with ruminal stasis, in particular lipopolysaccharides, which are released from the pathogenic species, stimulating the immune system and increasing the production of reactive oxygen species (ROS) ( Ayemele et al ., 2021 ). When antioxidant protection is impaired, an imbalance leads to oxidative stress, lipoprotein peroxidation, and further cell damage ( Celi, 2011 ; Khamees et al ., 2024 ). Increased lipid peroxidation, represented by malondialdehyde (MDA), has been observed in acute rumen acidosis and associated digestive disorders, highlighting the role of oxidative damage in disease progression ( Zeng et al ., 2023 ). Although the recognition of ruminal impaction is increasing, comprehensive assessments of serum biochemical, electrolyte, and oxidative stress profiles in sheep with naturally occurring ruminal impaction are still uncommon. Systematic characterization of these parameters is necessary to improve diagnostic accuracy and guide treatment of fluid overload, electrolyte replacement, and antioxidant therapy. This is one of the first studies to integrate haematobiochemical, electrolyte, and oxidative markers into evaluating ruminal impaction in sheep. This approach should improve our understanding of its pathophysiology and help to develop more effective treatments. Materials and methods The study involved clinically examined Awassi sheep aged approximately 2-3 years. All animals were non-pregnant and non-lactating females and were maintained under identical care and feeding conditions (10 in the impaction group and 10 in the control group). This group included animals with natural clinical cases of ruminal impaction, admitted to a veterinary clinic and diagnosed based on clinical examination and rumen condition data in Fallujah, Iraq, based on clinical signs of anorexia, dehydration, diarrhoea, foamy ruminal contents, and pale mucous membranes ( Constable et al. , 2016 ). The control group comprised age, sex, and weight-matched clinically normal animals whose biochemical values were within the physiological reference range reported for sheep. The venous blood samples were taken from each animal using sterile tubing containing lithium heparin, centrifuged at 3000 x g for 15 min, and the plasma was pooled and frozen at -20°C for analysis. Blood sampling, were performed in accordance with American Veterinary Medical Association (AVMA) guidelines ( Underwood and Raymond, 2013 ). Plasma biochemical parameters (albumin, cholesterol, urea, creatinine, alanine aminotransferase (ALT), and aspartate aminotransferase (AST) were measured using commercial colorimetric kits (Agape Diagnostics, Switzerland; Sam Diagnostics, United Arab Emirates; Biolab SAS, France). Electrolyte concentrations (sodium, potassium, chloride, calcium, magnesium, phosphate) were determined using an automatic chemical analyzer (Smart 150, Sam Diagnostic, UAE) and the corresponding commercial reagents. Oxidative stress biomarkers (superoxide dismutase (SOD), malondialdehyde (MDA), and reduced glutathione (GSH)) were quantified using ELISA kits (Biolab SAS, France). Statistical analysis was carried out with SPSS (version 21.0; IBM). Data are presented as mean ± standard error. Significance was assessed using the Shapiro-Wilk test. Differences between the occlusion and control groups were analyzed using an independent two-sample t-test. To reduce the risk of type I errors associated with multiple comparisons, the false-positive rate (FDR) was tested using the Benjamini-Hochberg test, and the significance level was set at P ≤ 0.05 ( Larsen et al. , 1973 ). Results 1. Effect of ruminal impaction on blood biochemical parameters in sheep. Table 1 , showed that most serum biochemical parameters were significantly higher in the impaction group (P ≤ 0.05), while albumin and cholesterol were significantly lower (P ≤ 0.05). 2. Effect of ruminal impaction on blood electrolytes. The result showed potassium and phosphate levels in serum were significantly increased ( Table 2 ) due to metabolic acidosis (Impaction), while sodium, calcium, and chloride levels were significantly decreased (P ≤ 0.05). Magnesium concentrations, on the other hand, did not change (P ≤ 0.05). 3. Effect of ruminal impaction on oxidative markers in the blood serum of sheep. The impaction group’s ( Table 3 ) plasma antioxidant markers showed significant declines, with GSH and SOD activity significantly lower than controls (P ≤ 0.05). In contrast, there was a significant increase in MDA levels in the afflicted sheep (P ≤ 0.05). Table 1. The effect of ruminal impaction on blood biochemical (alanine aminotransferase (ALT), aspartate Aminotransferase (AST), glucose, cholesterol, albumin, urea, and creatinine) values in local Iraqi sheep. Parameters Control group Impaction group P-value ALT (IU/l) 27 ± 1.7 B 34.6 ± 1.4 A 0.00037 AST (IU/l) 30.6 ± 1.5 B 37.5 ± 1.2 A 0.00015 Glucose (mg/dl) 72.6 ± 1.9 B 100.8 ± 2.1A 8.5*10 −12 Cholesterol (mg/dl) 86.4 ± 1.7 A 64.8 ± 2 B 2.19*10 −12 Albumin (g/dl) 5 ± 0.18A 3.34 ± 0.3 B 1.52*10 −9 Urea (mg/dl) 26.6 ± 1.3 B 44.4 ± 2.2 A 5.27*10 −10 Creatinine (mg/dl) 1.08 ± 0.09 B 2.2 ± 0.07 A 5.71*10 −17 Table 2. Effect of ruminal impaction blood electrolyte (sodium, potassium chloride, calcium, phosphate, and magnesium) values in local Iraqi sheep. Parameters Control group Impaction group P-value Na (mmol/L) 143.4 ± 3.2 A 122.6 ± 2.9 B 1.4*10 −7 K (mmol/L) 4.42 ± 0.19 B 6.94 ± 0.46 A 1.62*10 −8 Cl (mmol/L) 102 ± 2.9 A 90.4 ± 2.7 B 0.0001 Ca (mg/dl) 20 ± 0.9 A 16.7 ± 1.2 B 0.0002 Phosphate (mmol/L) 2.5 ± 0.14 B 10.5 ± 0.6 A 8.7*10 −16 Mg (mg/dl) 2.23 ± 0.09 A 2.22 ± 0.1 A 0.01 Table 3. The ruminal impaction effects on oxidative markers (reduced glutathione (GSH), superoxide dismutase (SOD), and malondialdehyde (MDA)) in the blood of local Iraqi sheep. Parameters Control group Impaction group P-value GSH (μmol/l) 9.2 ± 1.1 A 4.2 ± 1 B 0.056 SOD (U/ml) 19. 2± 1.3 A 10.8 ± 1.1 B 4.56*10 −5 MDA (μmol/l) 9.11 ± 0.91 B 14.15 ± 0.9 A 0.003 Discussion Although metabolic acidosis has been reported in ruminal impaction, direct acid-base parameters such as blood pH, bicarbonate concentration or blood gas analysis have not been measured in this study. Therefore, the current findings should be interpreted as biochemical changes possibly related to acid-base imbalance and not as a confirmed diagnosis of metabolic acidosis. So that, this study shows a coordinated pattern of biochemical, electrolyte, and oxidative markers in metabolic acidosis secondary to ruminal impaction in sheep. Elevated serum ALT and AST levels ( Table 1 ) indicate liver damage, most likely may indicated by lactic acid and other toxic fermentation products, and may indicate liver abscesses, pulmonary or renal damage ( Manosalva et al. , 2022 ), and are consistent with previous reports of elevated liver enzymes and decreased hepatic protein synthesis in sheep with acute ruminant acidosis ( Alkabi et al ., 2019 ; Elmeligy et al ., 2025 ). In addition, the hyperglycemia observed in the impaction group may contribute to elevate lactic acid, which seems to reflect a systemic stress response, as metabolic disturbance related with acidosis stimulates the release of catecholamine and cortisol, stimulating gluconeogenesis ( Zhang et al ., 2019 ). Similar glycaemic events have been documented in ruminants under dietary and acidotic stress ( Wang et al ., 2025 ). In addition to these changes, decreased serum albumin and cholesterol indicate impaired hepatic synthesis. On the other hand, hypoalbuminemia reduces plasma osmotic pressure and buffer capacity. In contrast, hypocholesterolaemia indicates impaired lipid metabolism under excessive stress on the liver may be due to the accumulation of D. lactate, supporting the apparent findings of elevated liver enzymes. These observations are consistent with previous findings of decreased total protein and albumin in ruminal acidosis and highlight the magnitude of liver dysfunction ( Zhang et al. , 2023 ). Furthermore, increased lactate production impairs the kidney’s ability to convert lactate into glucose ( DeFronzo et al. , 2012 ), adding to elevated urea and creatinine levels, which indicate renal involvement, most likely may be due to a decrease in glomerular filtration rate and reduced renal perfusion, both of which are exacerbated by dehydration and metabolic disturbance associated with acidosis. Comparable elevations in these parameters have been reported in sheep affected by ruminal acidosis, reinforcing the view that renal dysfunction is a frequent complication of the condition ( Zheng et al. , 2024 ). Electrolyte disturbances characterized by metabolic disturbance associated with acidosis ( Matyukhin et al. , 2020 ) have been clearly observed, with increases in serum potassium and phosphate levels, and decreases in sodium and calcium ( Table 2 ). These ionic shifts are attributed mainly to intracellular buffering of excess hydrogen ions, which causes the efflux of potassium to the extracellular space. When acidosis develops in cells, Na + -K + pump (ATPase pump) activity is consistent with reduced and the pH and phosphorus levels are altered, interfering with calcium binding and intestinal absorption. The resulting hypocalcemia may be associated with D-lactate accumulation in the intestinal mucosa, where a mucosal injury may further reduce calcium absorption. These results are consistent with this explanation; Kumar et al. (2019) reported a comparable decrease in serum calcium levels in ruminant cows with rumen acidosis. These changes have important physiological consequences, particularly for cardiac function and neuromuscular excitability. Oxidative stress is a significant factor in the pathophysiology of ruminant acidosis. The reduction in glutathione (GSH) and superoxide dismutase (SOD) and the increase in malondialdehyde (MDA) ( Table 3 ) indicate a weakened antioxidant protection. These amendments align with the Kirbaş et al. (2014) , which reported that acute ruminal acidosis promotes the production of excess reactive oxygen species (ROS), depleting the antioxidant capacity of the cells and speeding up lipid peroxidation. Excessive ROS production may be due to microbial dysbiosis, impaired ruminal fermentation, and systemic inflammatory reactions, exacerbating oxidative damage ( Lian et al. , 2024 ). Clinically, oxidative stress may impair organ function and neuromuscular performance, which underlines the importance of monitoring antioxidant markers in addition to biochemical and electrolyte parameters. Conclusion These findings may support a multifactorial metabolic disturbance in which ingestion of poorly digested food causes ruminal stasis, systemic acidosis, hepatic dysfunction, renal impairment, electrolyte disturbances, oxidative stress, and electrolyte disturbances. Relying on a single biomarker may underestimate the actual severity of the disease, while a combination of biochemical, electrolyte, and oxidative measurements provides a more comprehensive picture. In clinical practice, early detection using multiple markers is crucial, and early correction of potassium and calcium imbalances may be paramount for preventing complications. In addition, supportive treatment targeting liver, renal, and oxidative function is required to reduce systemic damage and to improve overall results. Ethical considerations All experimental procedures were conducted per the guidelines approved by the Scientific Research Ethics Committee of the Faculty of Veterinary Medicine, University of Fallujah (Approval No. 4; 16 September 2025). Data availability Underlying data Zenodo: Dataset for the ruminal impaction in sheep (2025). https://doi.org/10.5281/zenodo.18075931 .( Abed et al ., 2025 ). Arrive checklist : https://doi.org/10.5281/zenodo.18075931 ( Abed et al ., 2025 ). Data are available under the terms of the Creative Commons Zero “No rights reserved” data waiver (CC0 1.0 Public domain dedication). Acknowledgments The authors thank the study staff as well as the study participants. References Abed SK, Saud MA, Abood AE, et al. : Dataset for the ruminal impaction in sheep (2025). Zenodo. 2025. 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Comments on this article Comments (0) Version 2 VERSION 2 PUBLISHED 23 Jan 2026 ADD YOUR COMMENT Comment Author details Author details 1 College of Veterinary Medicine, University of Fallujah, Al-Fallujah, Al Anbar Governorate, Iraq Sabea Khamees Abed Roles: Conceptualization, Formal Analysis, Investigation, Methodology, Project Administration, Supervision, Writing – Original Draft Preparation Mustafa A. Saud Roles: Data Curation, Formal Analysis, Investigation, Visualization, Writing – Review & Editing Ahmed Emad Abood Roles: Data Curation, Investigation, Resources, Validation, Writing – Review & Editing Maysam Abdulrahman Ghazi Roles: Formal Analysis, Software, Validation, Visualization, Writing – Review & Editing Competing interests No competing interests were disclosed. Grant information The author(s) declared that no grants were involved in supporting this work. Article Versions (2) version 2 Revised Published: 31 Mar 2026, 15:110 https://doi.org/10.12688/f1000research.174402.2 version 1 Published: 23 Jan 2026, 15:110 https://doi.org/10.12688/f1000research.174402.1 Copyright © 2026 Khamees Abed S et al . This is an open access article distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Download Export To Sciwheel Bibtex EndNote ProCite Ref. Manager (RIS) Sente metrics Views Downloads F1000Research - - PubMed Central info_outline Data from PMC are received and updated monthly. - - Citations open_in_new 0 open_in_new 0 open_in_new SEE MORE DETAILS CITE how to cite this article Khamees Abed S, A. Saud M, Emad Abood A and Abdulrahman Ghazi M. Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.12688/f1000research.174402.2 ) NOTE: If applicable, it is important to ensure the information in square brackets after the title is included in all citations of this article. COPY CITATION DETAILS track receive updates on this article Track an article to receive email alerts on any updates to this article. TRACK THIS ARTICLE Share Open Peer Review Current Reviewer Status: ? Key to Reviewer Statuses VIEW HIDE Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions Version 2 VERSION 2 PUBLISHED 31 Mar 2026 Revised Views 0 Cite How to cite this report: Al-kurdy MJ. Reviewer Report For: Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.5256/f1000research.197497.r472403 ) The direct URL for this report is: https://f1000research.com/articles/15-110/v2#referee-response-472403 NOTE: it is important to ensure the information in square brackets after the title is included in this citation. Close Copy Citation Details Reviewer Report 21 Apr 2026 Masar Jabbar Al-kurdy , AL-Furat AL- Awsat Technical University, Najaf, Iraq Approved VIEWS 0 https://doi.org/10.5256/f1000research.197497.r472403 Multiple testing: The study compares 13 parameters (ALT, AST, glucose, cholesterol, albumin, urea, creatinine, Na, K, Cl, Ca, phosphate, Mg, plus 3 oxidative markers – effectively 16 tests) using independent t‑tests at α = 0.05. the ... Continue reading READ ALL Multiple testing: The study compares 13 parameters (ALT, AST, glucose, cholesterol, albumin, urea, creatinine, Na, K, Cl, Ca, phosphate, Mg, plus 3 oxidative markers – effectively 16 tests) using independent t‑tests at α = 0.05. the family‑wise error rate is ~56% (1 − 0.95^16). Many significant findings could be false positives. -Reporting of LSD: Tables 1–3 present “LSD” (least significant difference) values, which are not standard for independent t‑tests. This suggests a potential confusion with post‑hoc tests from ANOVA. The statistical section should describe exactly which test was used (t‑test or ANOVA) and report actual p‑values or adjusted thresholds, not LSD. -Normality testing: Only stated as “checked for normality” without naming the test (Shapiro‑Wilk?) or reporting results. Phosphate Units – Erroneous (Major) * Table 2 reports Phosphate (U/L). Phosphate concentration in serum is measured in mmol/L or mg/dL – not in enzymatic units (U/L). The control value (2.5 U/L) and impaction value (10.5 U/L) are biologically implausible for phosphate. This appears to be a typographical or transcription error. Required action: Verify the correct units (likely mg/dL or mmol/L) and correct the table. Also verify the numerical values – a four‑fold increase (2.5 to 10.5) is extreme but possible; however, the units must be correct. Is the work clearly and accurately presented and does it cite the current literature? Yes Is the study design appropriate and is the work technically sound? Yes Are sufficient details of methods and analysis provided to allow replication by others? Yes If applicable, is the statistical analysis and its interpretation appropriate? Partly Are all the source data underlying the results available to ensure full reproducibility? No source data required Are the conclusions drawn adequately supported by the results? Yes Competing Interests: No competing interests were disclosed. Reviewer Expertise: endocrine disorder, nanoparticles, I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. Close READ LESS CITE CITE HOW TO CITE THIS REPORT Al-kurdy MJ. Reviewer Report For: Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.5256/f1000research.197497.r472403 ) The direct URL for this report is: https://f1000research.com/articles/15-110/v2#referee-response-472403 NOTE: it is important to ensure the information in square brackets after the title is included in all citations of this article. COPY CITATION DETAILS Report a concern Respond or Comment COMMENT ON THIS REPORT Version 1 VERSION 1 PUBLISHED 23 Jan 2026 Views 0 Cite How to cite this report: Liu J. Reviewer Report For: Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.5256/f1000research.192297.r460234 ) The direct URL for this report is: https://f1000research.com/articles/15-110/v1#referee-response-460234 NOTE: it is important to ensure the information in square brackets after the title is included in this citation. Close Copy Citation Details Reviewer Report 28 Feb 2026 Jinxin Liu , Laboratory of Gastrointestinal Microbiology, Jiangsu Key Laboratory of Gastrointestinal Nutrition and Animal Health, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, China Approved with Reservations VIEWS 0 https://doi.org/10.5256/f1000research.192297.r460234 This manuscript investigates biochemical, electrolyte, and oxidative stress alterations in local Iraqi Awassi sheep affected by ruminal impaction using a case–control design. The study addresses a clinically relevant condition in small ruminant production and attempts to integrate metabolic, hepatic, renal, ... Continue reading READ ALL This manuscript investigates biochemical, electrolyte, and oxidative stress alterations in local Iraqi Awassi sheep affected by ruminal impaction using a case–control design. The study addresses a clinically relevant condition in small ruminant production and attempts to integrate metabolic, hepatic, renal, electrolyte, and oxidative parameters into a unified pathophysiological framework. The inclusion of oxidative stress markers alongside routine biochemical indices adds value and broadens the diagnostic perspective. However, while the study is clinically meaningful, several methodological and interpretational aspects require clarification. The sample characterization lacks sufficient detail, statistical methodology is overly simplified given the number of comparisons performed, and some mechanistic explanations in the discussion appear speculative relative to the data presented. Addressing these issues would improve scientific rigor and reproducibility while strengthening the translational value of the findings. Major Comments 1. Study design and case definition require clearer characterization Although the study is described as a case–control design (page 3), several key elements remain unclear: The age range, reproductive status (parity), and physiological stage of the sheep are not specified. These factors can significantly influence biochemical and oxidative parameters. It is not clearly stated whether ruminal impaction was naturally occurring or experimentally induced. The manuscript implies natural cases, but this should be explicitly clarified. Hyperkalaemia is described as confirmatory for diagnosis; however, potassium elevation is also one of the outcome variables. This raises concerns regarding circular reasoning in case definition. The authors should provide a more detailed clinical characterization of cases and clearly define diagnostic criteria independent of the primary outcome measures. 2. Statistical approach and multiple comparisons The study compares numerous biochemical, electrolyte, and oxidative markers using independent t-tests with a significance threshold of P < 0.05. Given the number of parameters assessed (ALT, AST, glucose, cholesterol, albumin, urea, creatinine, six electrolytes, and three oxidative markers), the risk of type I error is substantial. The authors should clarify: Whether assumptions of normality and variance homogeneity were formally tested. Whether any correction for multiple comparisons was considered. Why LSD values are reported in tables despite using independent t-tests. A more robust statistical framework or at least justification of the current approach would enhance credibility. 3. Interpretation of causality and mechanistic claims The discussion proposes a pathophysiological cascade linking ruminal impaction to systemic acidosis, hepatic dysfunction, renal impairment, electrolyte imbalance, and oxidative stress (pages 5–6). While the observed associations are plausible, the study design does not allow causal inference. Several mechanistic explanations (e.g., D-lactate–mediated hepatic injury, Na⁺/K⁺-ATPase suppression, lactate-driven renal gluconeogenesis impairment) are presented as explanatory conclusions but were not directly measured in this study. The discussion should be reframed to clearly distinguish between observed associations and hypothesized mechanisms based on prior literature. Minor Comments Pooling of blood samples (page 3) The methods state that pooled venous blood was collected and plasma was pooled before freezing. It is unclear whether pooling occurred within individuals or across individuals. If samples were pooled across animals, this would invalidate statistical comparisons. This must be clarified. Reference to metabolic acidosis The manuscript repeatedly refers to “metabolic acidosis,” yet no direct measurements of blood pH, bicarbonate, base excess, or blood gas analysis are provided. The authors should clarify how metabolic acidosis was confirmed. Phosphate units (Table 2) Phosphate is reported in U/L, which appears inconsistent with standard electrolyte reporting (typically mmol/L or mg/dL). Units should be verified. Is the work clearly and accurately presented and does it cite the current literature? Yes Is the study design appropriate and is the work technically sound? Yes Are sufficient details of methods and analysis provided to allow replication by others? Yes If applicable, is the statistical analysis and its interpretation appropriate? Yes Are all the source data underlying the results available to ensure full reproducibility? No source data required Are the conclusions drawn adequately supported by the results? Yes Competing Interests: No competing interests were disclosed. Reviewer Expertise: ruminant nutrition/gut microbiota I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above. Close READ LESS CITE CITE HOW TO CITE THIS REPORT Liu J. Reviewer Report For: Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.5256/f1000research.192297.r460234 ) The direct URL for this report is: https://f1000research.com/articles/15-110/v1#referee-response-460234 NOTE: it is important to ensure the information in square brackets after the title is included in all citations of this article. COPY CITATION DETAILS Report a concern Author Response 31 Mar 2026 Sabea Abed , College of Veterinary Medicine, University of Fallujah, Al-Fallujah, Iraq 31 Mar 2026 Author Response Dear Reviewer, Dr. Jinxin Liu We sincerely thank you for your careful evaluation of our manuscript and for your constructive and insightful comments. Your suggestions were extremely valuable and have ... Continue reading Dear Reviewer, Dr. Jinxin Liu We sincerely thank you for your careful evaluation of our manuscript and for your constructive and insightful comments. Your suggestions were extremely valuable and have significantly improved the clarity, scientific rigor, and overall quality of the manuscript. All comments have been carefully considered, and the manuscript has been revised accordingly. Our detailed responses to each comment are provided below. Comment 1: Study design and case definition require clearer characterization Thank you for this important observation. The manuscript has been revised to provide additional information regarding the clinical characterization of the animals, including age range and physiological status. We have also clarified that ruminal impaction cases were naturally occurring. Furthermore, the diagnostic criteria have been revised to ensure that potassium levels were not used as a confirmatory diagnostic parameter, thereby avoiding potential circular reasoning. Comment 2: Statistical approach and multiple comparisons We appreciate this valuable suggestion. To strengthen the statistical rigor of the study, the statistical analysis section has been revised. Normality of the data was assessed using the Shapiro–Wilk test, and differences between groups were analyzed using an independent samples t-test. Additionally, to control for potential type I error due to multiple comparisons, the false discovery rate (FDR) was adjusted using the Benjamini–Hochberg procedure. The statistical analysis section has been updated accordingly, and the tables were revised to remove LSD values and report appropriate p-values. Comment 3: Interpretation of causality and mechanistic claims We thank the reviewer for this insightful comment. The discussion has been carefully revised to avoid causal interpretations. The mechanistic explanations are now presented as hypotheses supported by previous literature rather than direct conclusions derived from the current data. Minor Comment 1: Pooling of blood samples We apologize for the lack of clarity. The manuscript has been revised to clarify that blood samples were collected and processed individually for each animal. No pooling occurred across animals, and each sample was analyzed separately. Minor Comment 2: Reference to metabolic acidosis We appreciate this comment. The discussion has been revised to clarify that metabolic acidosis was inferred based on previously reported pathophysiological mechanisms associated with ruminal disorders, rather than being directly measured in this study. This limitation has now been explicitly acknowledged in the revised manuscript. Minor Comment 3: Phosphate units The units for phosphate have been carefully reviewed and corrected in the revised tables. Once again, we sincerely thank the reviewer for the valuable feedback, which has greatly improved the quality and clarity of our manuscript. Sincerely , Sabea Khamees Abed Dear Reviewer, Dr. Jinxin Liu We sincerely thank you for your careful evaluation of our manuscript and for your constructive and insightful comments. Your suggestions were extremely valuable and have significantly improved the clarity, scientific rigor, and overall quality of the manuscript. All comments have been carefully considered, and the manuscript has been revised accordingly. Our detailed responses to each comment are provided below. Comment 1: Study design and case definition require clearer characterization Thank you for this important observation. The manuscript has been revised to provide additional information regarding the clinical characterization of the animals, including age range and physiological status. We have also clarified that ruminal impaction cases were naturally occurring. Furthermore, the diagnostic criteria have been revised to ensure that potassium levels were not used as a confirmatory diagnostic parameter, thereby avoiding potential circular reasoning. Comment 2: Statistical approach and multiple comparisons We appreciate this valuable suggestion. To strengthen the statistical rigor of the study, the statistical analysis section has been revised. Normality of the data was assessed using the Shapiro–Wilk test, and differences between groups were analyzed using an independent samples t-test. Additionally, to control for potential type I error due to multiple comparisons, the false discovery rate (FDR) was adjusted using the Benjamini–Hochberg procedure. The statistical analysis section has been updated accordingly, and the tables were revised to remove LSD values and report appropriate p-values. Comment 3: Interpretation of causality and mechanistic claims We thank the reviewer for this insightful comment. The discussion has been carefully revised to avoid causal interpretations. The mechanistic explanations are now presented as hypotheses supported by previous literature rather than direct conclusions derived from the current data. Minor Comment 1: Pooling of blood samples We apologize for the lack of clarity. The manuscript has been revised to clarify that blood samples were collected and processed individually for each animal. No pooling occurred across animals, and each sample was analyzed separately. Minor Comment 2: Reference to metabolic acidosis We appreciate this comment. The discussion has been revised to clarify that metabolic acidosis was inferred based on previously reported pathophysiological mechanisms associated with ruminal disorders, rather than being directly measured in this study. This limitation has now been explicitly acknowledged in the revised manuscript. Minor Comment 3: Phosphate units The units for phosphate have been carefully reviewed and corrected in the revised tables. Once again, we sincerely thank the reviewer for the valuable feedback, which has greatly improved the quality and clarity of our manuscript. Sincerely , Sabea Khamees Abed Competing Interests: I declare that I have no competing interests. Close Report a concern Respond or Comment COMMENTS ON THIS REPORT Author Response 31 Mar 2026 Sabea Abed , College of Veterinary Medicine, University of Fallujah, Al-Fallujah, Iraq 31 Mar 2026 Author Response Dear Reviewer, Dr. Jinxin Liu We sincerely thank you for your careful evaluation of our manuscript and for your constructive and insightful comments. Your suggestions were extremely valuable and have ... Continue reading Dear Reviewer, Dr. Jinxin Liu We sincerely thank you for your careful evaluation of our manuscript and for your constructive and insightful comments. Your suggestions were extremely valuable and have significantly improved the clarity, scientific rigor, and overall quality of the manuscript. All comments have been carefully considered, and the manuscript has been revised accordingly. Our detailed responses to each comment are provided below. Comment 1: Study design and case definition require clearer characterization Thank you for this important observation. The manuscript has been revised to provide additional information regarding the clinical characterization of the animals, including age range and physiological status. We have also clarified that ruminal impaction cases were naturally occurring. Furthermore, the diagnostic criteria have been revised to ensure that potassium levels were not used as a confirmatory diagnostic parameter, thereby avoiding potential circular reasoning. Comment 2: Statistical approach and multiple comparisons We appreciate this valuable suggestion. To strengthen the statistical rigor of the study, the statistical analysis section has been revised. Normality of the data was assessed using the Shapiro–Wilk test, and differences between groups were analyzed using an independent samples t-test. Additionally, to control for potential type I error due to multiple comparisons, the false discovery rate (FDR) was adjusted using the Benjamini–Hochberg procedure. The statistical analysis section has been updated accordingly, and the tables were revised to remove LSD values and report appropriate p-values. Comment 3: Interpretation of causality and mechanistic claims We thank the reviewer for this insightful comment. The discussion has been carefully revised to avoid causal interpretations. The mechanistic explanations are now presented as hypotheses supported by previous literature rather than direct conclusions derived from the current data. Minor Comment 1: Pooling of blood samples We apologize for the lack of clarity. The manuscript has been revised to clarify that blood samples were collected and processed individually for each animal. No pooling occurred across animals, and each sample was analyzed separately. Minor Comment 2: Reference to metabolic acidosis We appreciate this comment. The discussion has been revised to clarify that metabolic acidosis was inferred based on previously reported pathophysiological mechanisms associated with ruminal disorders, rather than being directly measured in this study. This limitation has now been explicitly acknowledged in the revised manuscript. Minor Comment 3: Phosphate units The units for phosphate have been carefully reviewed and corrected in the revised tables. Once again, we sincerely thank the reviewer for the valuable feedback, which has greatly improved the quality and clarity of our manuscript. Sincerely , Sabea Khamees Abed Dear Reviewer, Dr. Jinxin Liu We sincerely thank you for your careful evaluation of our manuscript and for your constructive and insightful comments. Your suggestions were extremely valuable and have significantly improved the clarity, scientific rigor, and overall quality of the manuscript. All comments have been carefully considered, and the manuscript has been revised accordingly. Our detailed responses to each comment are provided below. Comment 1: Study design and case definition require clearer characterization Thank you for this important observation. The manuscript has been revised to provide additional information regarding the clinical characterization of the animals, including age range and physiological status. We have also clarified that ruminal impaction cases were naturally occurring. Furthermore, the diagnostic criteria have been revised to ensure that potassium levels were not used as a confirmatory diagnostic parameter, thereby avoiding potential circular reasoning. Comment 2: Statistical approach and multiple comparisons We appreciate this valuable suggestion. To strengthen the statistical rigor of the study, the statistical analysis section has been revised. Normality of the data was assessed using the Shapiro–Wilk test, and differences between groups were analyzed using an independent samples t-test. Additionally, to control for potential type I error due to multiple comparisons, the false discovery rate (FDR) was adjusted using the Benjamini–Hochberg procedure. The statistical analysis section has been updated accordingly, and the tables were revised to remove LSD values and report appropriate p-values. Comment 3: Interpretation of causality and mechanistic claims We thank the reviewer for this insightful comment. The discussion has been carefully revised to avoid causal interpretations. The mechanistic explanations are now presented as hypotheses supported by previous literature rather than direct conclusions derived from the current data. Minor Comment 1: Pooling of blood samples We apologize for the lack of clarity. The manuscript has been revised to clarify that blood samples were collected and processed individually for each animal. No pooling occurred across animals, and each sample was analyzed separately. Minor Comment 2: Reference to metabolic acidosis We appreciate this comment. The discussion has been revised to clarify that metabolic acidosis was inferred based on previously reported pathophysiological mechanisms associated with ruminal disorders, rather than being directly measured in this study. This limitation has now been explicitly acknowledged in the revised manuscript. Minor Comment 3: Phosphate units The units for phosphate have been carefully reviewed and corrected in the revised tables. Once again, we sincerely thank the reviewer for the valuable feedback, which has greatly improved the quality and clarity of our manuscript. Sincerely , Sabea Khamees Abed Competing Interests: I declare that I have no competing interests. Close Report a concern COMMENT ON THIS REPORT Views 0 Cite How to cite this report: Evci Ş. Reviewer Report For: Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.5256/f1000research.192297.r453401 ) The direct URL for this report is: https://f1000research.com/articles/15-110/v1#referee-response-453401 NOTE: it is important to ensure the information in square brackets after the title is included in this citation. Close Copy Citation Details Reviewer Report 09 Feb 2026 Şevket Evci , Kırıkkale University, Kırıkkale, Turkey Approved with Reservations VIEWS 0 https://doi.org/10.5256/f1000research.192297.r453401 The materials section of the article lacks sufficient information regarding the sheep's age, number of births, and whether the impaction was experimentally induced. Furthermore, the normal range for blood values ​​is not specified, and comparisons are made using values ​​from ... Continue reading READ ALL The materials section of the article lacks sufficient information regarding the sheep's age, number of births, and whether the impaction was experimentally induced. Furthermore, the normal range for blood values ​​is not specified, and comparisons are made using values ​​from a control group. Were the blood values ​​of the control group normal? Is the work clearly and accurately presented and does it cite the current literature? Yes Is the study design appropriate and is the work technically sound? Yes Are sufficient details of methods and analysis provided to allow replication by others? Partly If applicable, is the statistical analysis and its interpretation appropriate? Partly Are all the source data underlying the results available to ensure full reproducibility? Yes Are the conclusions drawn adequately supported by the results? Yes Competing Interests: No competing interests were disclosed. Reviewer Expertise: Nutritional disease of animals I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above. Close READ LESS CITE CITE HOW TO CITE THIS REPORT Evci Ş. Reviewer Report For: Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.5256/f1000research.192297.r453401 ) The direct URL for this report is: https://f1000research.com/articles/15-110/v1#referee-response-453401 NOTE: it is important to ensure the information in square brackets after the title is included in all citations of this article. COPY CITATION DETAILS Report a concern Author Response 31 Mar 2026 Sabea Abed , College of Veterinary Medicine, University of Fallujah, Al-Fallujah, Iraq 31 Mar 2026 Author Response Dear Reviewer, Dr. Şevket Evci , We sincerely appreciate the time and effort you devoted to evaluating our manuscript. Your constructive and insightful comments have been extremely valuable in improving the ... Continue reading Dear Reviewer, Dr. Şevket Evci , We sincerely appreciate the time and effort you devoted to evaluating our manuscript. Your constructive and insightful comments have been extremely valuable in improving the clarity, methodological rigor, and overall quality of the study. We have carefully considered all of your suggestions and revised the manuscript accordingly. Detailed responses to each of your comments are provided below. Additional information regarding the animals has been added to the Materials section, including age range and physiological status. The ruminal impaction cases were naturally occurring clinical cases and were not experimentally induced. Furthermore, the control group consisted of clinically healthy animals, and their biochemical values fell within the physiological reference ranges reported for sheep. Once again, we sincerely thank the reviewer for the valuable feedback, which has greatly improved the quality and clarity of our manuscript. Sincerely , Sabea Khamees Abed Dear Reviewer, Dr. Şevket Evci , We sincerely appreciate the time and effort you devoted to evaluating our manuscript. Your constructive and insightful comments have been extremely valuable in improving the clarity, methodological rigor, and overall quality of the study. We have carefully considered all of your suggestions and revised the manuscript accordingly. Detailed responses to each of your comments are provided below. Additional information regarding the animals has been added to the Materials section, including age range and physiological status. The ruminal impaction cases were naturally occurring clinical cases and were not experimentally induced. Furthermore, the control group consisted of clinically healthy animals, and their biochemical values fell within the physiological reference ranges reported for sheep. Once again, we sincerely thank the reviewer for the valuable feedback, which has greatly improved the quality and clarity of our manuscript. Sincerely , Sabea Khamees Abed Competing Interests: I declare that I have no competing interests. Close Report a concern Respond or Comment COMMENTS ON THIS REPORT Author Response 31 Mar 2026 Sabea Abed , College of Veterinary Medicine, University of Fallujah, Al-Fallujah, Iraq 31 Mar 2026 Author Response Dear Reviewer, Dr. Şevket Evci , We sincerely appreciate the time and effort you devoted to evaluating our manuscript. Your constructive and insightful comments have been extremely valuable in improving the ... Continue reading Dear Reviewer, Dr. Şevket Evci , We sincerely appreciate the time and effort you devoted to evaluating our manuscript. Your constructive and insightful comments have been extremely valuable in improving the clarity, methodological rigor, and overall quality of the study. We have carefully considered all of your suggestions and revised the manuscript accordingly. Detailed responses to each of your comments are provided below. Additional information regarding the animals has been added to the Materials section, including age range and physiological status. The ruminal impaction cases were naturally occurring clinical cases and were not experimentally induced. Furthermore, the control group consisted of clinically healthy animals, and their biochemical values fell within the physiological reference ranges reported for sheep. Once again, we sincerely thank the reviewer for the valuable feedback, which has greatly improved the quality and clarity of our manuscript. Sincerely , Sabea Khamees Abed Dear Reviewer, Dr. Şevket Evci , We sincerely appreciate the time and effort you devoted to evaluating our manuscript. Your constructive and insightful comments have been extremely valuable in improving the clarity, methodological rigor, and overall quality of the study. We have carefully considered all of your suggestions and revised the manuscript accordingly. Detailed responses to each of your comments are provided below. Additional information regarding the animals has been added to the Materials section, including age range and physiological status. The ruminal impaction cases were naturally occurring clinical cases and were not experimentally induced. Furthermore, the control group consisted of clinically healthy animals, and their biochemical values fell within the physiological reference ranges reported for sheep. Once again, we sincerely thank the reviewer for the valuable feedback, which has greatly improved the quality and clarity of our manuscript. Sincerely , Sabea Khamees Abed Competing Interests: I declare that I have no competing interests. Close Report a concern COMMENT ON THIS REPORT Comments on this article Comments (0) Version 2 VERSION 2 PUBLISHED 23 Jan 2026 ADD YOUR COMMENT Comment keyboard_arrow_left keyboard_arrow_right Open Peer Review Reviewer Status info_outline Alongside their report, reviewers assign a status to the article: Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions Reviewer Reports Invited Reviewers 1 2 3 Version 2 (revision) 31 Mar 26 read Version 1 23 Jan 26 read read Şevket Evci , Kırıkkale University, Kırıkkale, Turkey Jinxin Liu , Nanjing Agricultural University, Nanjing, China Masar Jabbar Al-kurdy , AL-Furat AL- Awsat Technical University, Najaf, Iraq Comments on this article All Comments (0) Add a comment Sign up for content alerts Sign Up You are now signed up to receive this alert Browse by related subjects keyboard_arrow_left Back to all reports Reviewer Report 0 Views copyright © 2026 Al-kurdy M. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 21 Apr 2026 | for Version 2 Masar Jabbar Al-kurdy , AL-Furat AL- Awsat Technical University, Najaf, Iraq 0 Views copyright © 2026 Al-kurdy M. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. format_quote Cite this report speaker_notes Responses (0) Approved info_outline Alongside their report, reviewers assign a status to the article: Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions Multiple testing: The study compares 13 parameters (ALT, AST, glucose, cholesterol, albumin, urea, creatinine, Na, K, Cl, Ca, phosphate, Mg, plus 3 oxidative markers – effectively 16 tests) using independent t‑tests at α = 0.05. the family‑wise error rate is ~56% (1 − 0.95^16). Many significant findings could be false positives. -Reporting of LSD: Tables 1–3 present “LSD” (least significant difference) values, which are not standard for independent t‑tests. This suggests a potential confusion with post‑hoc tests from ANOVA. The statistical section should describe exactly which test was used (t‑test or ANOVA) and report actual p‑values or adjusted thresholds, not LSD. -Normality testing: Only stated as “checked for normality” without naming the test (Shapiro‑Wilk?) or reporting results. Phosphate Units – Erroneous (Major) * Table 2 reports Phosphate (U/L). Phosphate concentration in serum is measured in mmol/L or mg/dL – not in enzymatic units (U/L). The control value (2.5 U/L) and impaction value (10.5 U/L) are biologically implausible for phosphate. This appears to be a typographical or transcription error. Required action: Verify the correct units (likely mg/dL or mmol/L) and correct the table. Also verify the numerical values – a four‑fold increase (2.5 to 10.5) is extreme but possible; however, the units must be correct. Is the work clearly and accurately presented and does it cite the current literature? Yes Is the study design appropriate and is the work technically sound? Yes Are sufficient details of methods and analysis provided to allow replication by others? Yes If applicable, is the statistical analysis and its interpretation appropriate? Partly Are all the source data underlying the results available to ensure full reproducibility? No source data required Are the conclusions drawn adequately supported by the results? Yes Competing Interests No competing interests were disclosed. Reviewer Expertise endocrine disorder, nanoparticles, I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. reply Respond to this report Responses (0) Al-kurdy MJ. Peer Review Report For: Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.5256/f1000research.197497.r472403) NOTE: it is important to ensure the information in square brackets after the title is included in this citation. The direct URL for this report is: https://f1000research.com/articles/15-110/v2#referee-response-472403 keyboard_arrow_left Back to all reports Reviewer Report 0 Views copyright © 2026 Liu J. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 28 Feb 2026 | for Version 1 Jinxin Liu , Laboratory of Gastrointestinal Microbiology, Jiangsu Key Laboratory of Gastrointestinal Nutrition and Animal Health, College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, China 0 Views copyright © 2026 Liu J. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. format_quote Cite this report speaker_notes Responses (1) Approved With Reservations info_outline Alongside their report, reviewers assign a status to the article: Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions This manuscript investigates biochemical, electrolyte, and oxidative stress alterations in local Iraqi Awassi sheep affected by ruminal impaction using a case–control design. The study addresses a clinically relevant condition in small ruminant production and attempts to integrate metabolic, hepatic, renal, electrolyte, and oxidative parameters into a unified pathophysiological framework. The inclusion of oxidative stress markers alongside routine biochemical indices adds value and broadens the diagnostic perspective. However, while the study is clinically meaningful, several methodological and interpretational aspects require clarification. The sample characterization lacks sufficient detail, statistical methodology is overly simplified given the number of comparisons performed, and some mechanistic explanations in the discussion appear speculative relative to the data presented. Addressing these issues would improve scientific rigor and reproducibility while strengthening the translational value of the findings. Major Comments 1. Study design and case definition require clearer characterization Although the study is described as a case–control design (page 3), several key elements remain unclear: The age range, reproductive status (parity), and physiological stage of the sheep are not specified. These factors can significantly influence biochemical and oxidative parameters. It is not clearly stated whether ruminal impaction was naturally occurring or experimentally induced. The manuscript implies natural cases, but this should be explicitly clarified. Hyperkalaemia is described as confirmatory for diagnosis; however, potassium elevation is also one of the outcome variables. This raises concerns regarding circular reasoning in case definition. The authors should provide a more detailed clinical characterization of cases and clearly define diagnostic criteria independent of the primary outcome measures. 2. Statistical approach and multiple comparisons The study compares numerous biochemical, electrolyte, and oxidative markers using independent t-tests with a significance threshold of P < 0.05. Given the number of parameters assessed (ALT, AST, glucose, cholesterol, albumin, urea, creatinine, six electrolytes, and three oxidative markers), the risk of type I error is substantial. The authors should clarify: Whether assumptions of normality and variance homogeneity were formally tested. Whether any correction for multiple comparisons was considered. Why LSD values are reported in tables despite using independent t-tests. A more robust statistical framework or at least justification of the current approach would enhance credibility. 3. Interpretation of causality and mechanistic claims The discussion proposes a pathophysiological cascade linking ruminal impaction to systemic acidosis, hepatic dysfunction, renal impairment, electrolyte imbalance, and oxidative stress (pages 5–6). While the observed associations are plausible, the study design does not allow causal inference. Several mechanistic explanations (e.g., D-lactate–mediated hepatic injury, Na⁺/K⁺-ATPase suppression, lactate-driven renal gluconeogenesis impairment) are presented as explanatory conclusions but were not directly measured in this study. The discussion should be reframed to clearly distinguish between observed associations and hypothesized mechanisms based on prior literature. Minor Comments Pooling of blood samples (page 3) The methods state that pooled venous blood was collected and plasma was pooled before freezing. It is unclear whether pooling occurred within individuals or across individuals. If samples were pooled across animals, this would invalidate statistical comparisons. This must be clarified. Reference to metabolic acidosis The manuscript repeatedly refers to “metabolic acidosis,” yet no direct measurements of blood pH, bicarbonate, base excess, or blood gas analysis are provided. The authors should clarify how metabolic acidosis was confirmed. Phosphate units (Table 2) Phosphate is reported in U/L, which appears inconsistent with standard electrolyte reporting (typically mmol/L or mg/dL). Units should be verified. Is the work clearly and accurately presented and does it cite the current literature? Yes Is the study design appropriate and is the work technically sound? Yes Are sufficient details of methods and analysis provided to allow replication by others? Yes If applicable, is the statistical analysis and its interpretation appropriate? Yes Are all the source data underlying the results available to ensure full reproducibility? No source data required Are the conclusions drawn adequately supported by the results? Yes Competing Interests No competing interests were disclosed. Reviewer Expertise ruminant nutrition/gut microbiota I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above. reply Respond to this report Responses (1) Author Response 31 Mar 2026 Sabea Abed, College of Veterinary Medicine, University of Fallujah, Al-Fallujah, Iraq Dear Reviewer, Dr. Jinxin Liu We sincerely thank you for your careful evaluation of our manuscript and for your constructive and insightful comments. Your suggestions were extremely valuable and have significantly improved the clarity, scientific rigor, and overall quality of the manuscript. All comments have been carefully considered, and the manuscript has been revised accordingly. Our detailed responses to each comment are provided below. Comment 1: Study design and case definition require clearer characterization Thank you for this important observation. The manuscript has been revised to provide additional information regarding the clinical characterization of the animals, including age range and physiological status. We have also clarified that ruminal impaction cases were naturally occurring. Furthermore, the diagnostic criteria have been revised to ensure that potassium levels were not used as a confirmatory diagnostic parameter, thereby avoiding potential circular reasoning. Comment 2: Statistical approach and multiple comparisons We appreciate this valuable suggestion. To strengthen the statistical rigor of the study, the statistical analysis section has been revised. Normality of the data was assessed using the Shapiro–Wilk test, and differences between groups were analyzed using an independent samples t-test. Additionally, to control for potential type I error due to multiple comparisons, the false discovery rate (FDR) was adjusted using the Benjamini–Hochberg procedure. The statistical analysis section has been updated accordingly, and the tables were revised to remove LSD values and report appropriate p-values. Comment 3: Interpretation of causality and mechanistic claims We thank the reviewer for this insightful comment. The discussion has been carefully revised to avoid causal interpretations. The mechanistic explanations are now presented as hypotheses supported by previous literature rather than direct conclusions derived from the current data. Minor Comment 1: Pooling of blood samples We apologize for the lack of clarity. The manuscript has been revised to clarify that blood samples were collected and processed individually for each animal. No pooling occurred across animals, and each sample was analyzed separately. Minor Comment 2: Reference to metabolic acidosis We appreciate this comment. The discussion has been revised to clarify that metabolic acidosis was inferred based on previously reported pathophysiological mechanisms associated with ruminal disorders, rather than being directly measured in this study. This limitation has now been explicitly acknowledged in the revised manuscript. Minor Comment 3: Phosphate units The units for phosphate have been carefully reviewed and corrected in the revised tables. Once again, we sincerely thank the reviewer for the valuable feedback, which has greatly improved the quality and clarity of our manuscript. Sincerely , Sabea Khamees Abed View more View less Competing Interests I declare that I have no competing interests. reply Respond Report a concern Liu J. Peer Review Report For: Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.5256/f1000research.192297.r460234) NOTE: it is important to ensure the information in square brackets after the title is included in this citation. The direct URL for this report is: https://f1000research.com/articles/15-110/v1#referee-response-460234 keyboard_arrow_left Back to all reports Reviewer Report 0 Views copyright © 2026 Evci Ş. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. 09 Feb 2026 | for Version 1 Şevket Evci , Kırıkkale University, Kırıkkale, Turkey 0 Views copyright © 2026 Evci Ş. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License , which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. format_quote Cite this report speaker_notes Responses (1) Approved With Reservations info_outline Alongside their report, reviewers assign a status to the article: Approved The paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved Fundamental flaws in the paper seriously undermine the findings and conclusions The materials section of the article lacks sufficient information regarding the sheep's age, number of births, and whether the impaction was experimentally induced. Furthermore, the normal range for blood values ​​is not specified, and comparisons are made using values ​​from a control group. Were the blood values ​​of the control group normal? Is the work clearly and accurately presented and does it cite the current literature? Yes Is the study design appropriate and is the work technically sound? Yes Are sufficient details of methods and analysis provided to allow replication by others? Partly If applicable, is the statistical analysis and its interpretation appropriate? Partly Are all the source data underlying the results available to ensure full reproducibility? Yes Are the conclusions drawn adequately supported by the results? Yes Competing Interests No competing interests were disclosed. Reviewer Expertise Nutritional disease of animals I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above. reply Respond to this report Responses (1) Author Response 31 Mar 2026 Sabea Abed, College of Veterinary Medicine, University of Fallujah, Al-Fallujah, Iraq Dear Reviewer, Dr. Şevket Evci , We sincerely appreciate the time and effort you devoted to evaluating our manuscript. Your constructive and insightful comments have been extremely valuable in improving the clarity, methodological rigor, and overall quality of the study. We have carefully considered all of your suggestions and revised the manuscript accordingly. Detailed responses to each of your comments are provided below. Additional information regarding the animals has been added to the Materials section, including age range and physiological status. The ruminal impaction cases were naturally occurring clinical cases and were not experimentally induced. Furthermore, the control group consisted of clinically healthy animals, and their biochemical values fell within the physiological reference ranges reported for sheep. Once again, we sincerely thank the reviewer for the valuable feedback, which has greatly improved the quality and clarity of our manuscript. Sincerely , Sabea Khamees Abed View more View less Competing Interests I declare that I have no competing interests. reply Respond Report a concern Evci Ş. Peer Review Report For: Physiological Alterations in Local Iraqi Sheep Affected by Ruminal Impaction: Insights from Biochemical, Electrolyte, and Oxidative Markers [version 2; peer review: 1 approved, 2 approved with reservations] . F1000Research 2026, 15 :110 ( https://doi.org/10.5256/f1000research.192297.r453401) NOTE: it is important to ensure the information in square brackets after the title is included in this citation. The direct URL for this report is: https://f1000research.com/articles/15-110/v1#referee-response-453401 Alongside their report, reviewers assign a status to the article: Approved - the paper is scientifically sound in its current form and only minor, if any, improvements are suggested Approved with reservations - A number of small changes, sometimes more significant revisions are required to address specific details and improve the papers academic merit. Not approved - fundamental flaws in the paper seriously undermine the findings and conclusions Adjust parameters to alter display View on desktop for interactive features Includes Interactive Elements View on desktop for interactive features Competing Interests Policy Provide sufficient details of any financial or non-financial competing interests to enable users to assess whether your comments might lead a reasonable person to question your impartiality. 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