Animal Microbiome

Advances in metagenomic sequencing have transformed our understanding of host-associated microbiota, revealing critical roles in immune regulation, nutrient metabolism, and gastrointestinal (GI) homeostasis. However, the gut microbiome of large non-model species, particularly elephants, remains poorly characterized. Captivity introduces dietary, environmental, and management-related perturbations that may disrupt microbial balance and contribute to GI dysfunction. Here, we performed a longitudinal analysis of the fecal microbiome in five captive Asian elephants (Elephas maximus) at the Oregon Zoo exhibiting chronic fecal abnormalities, including mucus and inconsistent stool formation. Over 14 weeks, weekly fecal samples were collected and compared with samples from clinically normal elephants housed at three other zoological institutions. Using 16S rRNA gene sequencing, we identified marked differences in microbial community composition between affected and control elephants. Dysbiosis in Oregon Zoo elephants was characterized by enrichment of Akkermansia muciniphila and multiple members of the order Clostridiales, including taxa previously associated with gastrointestinal disorders. Administration of a commercially available probiotic formulation was associated with transient improvement in fecal characteristics and pronounced shifts in microbial composition, including a significant post-treatment reduction in overall microbial diversity and decreased abundance of several taxa linked to GI abnormalities. Notably, probiotic strains themselves were not detected, suggesting indirect or short-lived functional effects rather than durable colonization. Together, these findings provide one of the first longitudinal characterizations of gut microbiome dysbiosis in captive Asian elephants and identify candidate microbial contributors to chronic GI dysfunction in captivity, with implications for husbandry, dietary management, and microbiome-informed interventions in megafauna. Additionally, our study underscores the potential, although limited and likely indirect, benefit of probiotics when treating GI disorders in monograstric megavertebrates.

Cattle harbor diverse bacterial communities across the gastrointestinal, respiratory, reproductive, mammary and other anatomical systems, but body-wide microbial biogeography remains poorly defined. To address this knowledge gap, we conducted a meta-analysis of publicly available bovine 16S rRNA gene amplicon sequencing data from 5,637 samples from 47 studies across six geographic regions and 27 anatomical sample types. Bacterial community structure differed significantly among sample types (PERMANOVA, R2 = 0.245, P = 0.0001), indicating spatial organization of bacterial communities across the bovine body, although study-level effects also contributed substantially to community variation. Communities were generally more similar within than between anatomical systems. Bacterial richness, diversity, taxonomic composition, and indicator taxa varied among sample types, with gastrointestinal, mammary-associated, ocular, and hoof microbiota exhibiting greater diversity than microbiota from liver, joint, and several reproductive samples. Distinct bacterial communities characterized the gastrointestinal, respiratory, reproductive, and mammary systems, as well as other anatomical sites. Despite these differences, several bacterial taxa were shared across multiple anatomical niches, particularly among male and female reproductive sites and among mammary-associated niches. This study provides a comprehensive body-wide characterization of bacterial biogeography in cattle and establishes a baseline for future studies of bovine microbial ecology and host-microbiome interactions.

Gut microbiome responses to anthropogenic disturbance vary across wildlife species, even within similarly disturbed landscapes. What drives this variation is unclear: whether it reflects anthropogenic exposure itself or broader ecological differences among hosts. We tested this using three macaque species with contrasting ecology, Bonnet, Rhesus, and Lion-tailed macaques, sampled across 12 sites in southern India spanning contrasting anthropogenic exposure, using 16S rRNA gene sequencing (n = 127) and shotgun metagenomics on a subset of samples. The two synurbanized species exhibited a similar magnitude of microbiome restructuring but differed in the taxa underlying these changes; no differentially abundant amplicon sequence variants were shared across all three species, indicating that shared anthropogenic exposure did not produce uniform microbial responses across hosts. The specialist Lion-tailed macaque showed a more extensive response, characterized by reduced diversity and phylogenetically structured compositional change. The Bonnet macaque showed greater microbial similarity with the Rhesus macaque than with the Lion-tailed macaque during sympatric co-occurrence. Despite taxonomic divergence, functional pathway architecture was broadly conserved across species and habitats, with selective shifts in pathways including vitamin B6 biosynthesis and fermentation. Together, these findings show that microbiome responses to anthropogenic environments are jointly shaped by ecological context and host ecology, with host differences in diet, habitat use, and ecological history influencing the magnitude and nature of microbial restructuring. These findings show that taxonomic diversity and functional potential respond as partially decoupled axes under anthropogenic pressure, with implications for assessing microbiome resilience across ecologically heterogeneous wildlife.

The gut microbiota is crucial for immune development and overall health in chickens. In commercial production, birds routinely receive multiple vaccines during early life. While individual vaccines are known to affect microbial composition, the impact of complex, multi-vaccine programs, as used in the poultry industry, is not well understood. This longitudinal study examined the impact of multiple live and inactivated vaccines, given at commercially relevant times from an early age, on gut microbial diversity and composition in layer chickens. We characterised microbiota profiles using 16S rRNA gene sequencing at pre- and post-vaccination timepoints across different vaccine groups. Overall, microbial diversity remained stable across most vaccines, indicating strong resilience of the gut microbiota to repeated immunological interventions. Differential abundance analyses identified changes in selected bacterial taxa following vaccination, with responses varying among vaccine groups. Notably, these changes were not sustained, as the gut microbial community returned to a stable state after the vaccination schedule. These findings underscore the robustness of the chicken gut ecosystem and lay a foundation for future research into microbiome-vaccine interactions and their implications for poultry health, immunity, and production efficiency.

The respiratory microbiome plays important roles in poultry health, immune regulation, and pathogen resistance, yet its development throughout the broiler production cycle remains poorly understood. This study investigated temporal changes in the tracheal microbiome of broiler chickens across production phases. Tracheal samples were collected during the starter (day 12), grower (day 21), early finisher (day 26), and late finisher (day 35) phases and analyzed using 16S ribosomal RNA gene sequencing. Tracheal microbial richness, diversity, community structure, and taxonomic composition changed significantly across broiler production stages, including starter, grower, early finisher, and late finisher feeding phases. Alpha diversity increased progressively throughout production, with significant increases in richness, diversity, and phylogenetic diversity during later stages. Beta diversity analysis revealed distinct microbial communities associated with each production phase, with starter-phase samples clearly separated from later phases. Taxonomic profiling showed dominance of Proteobacteria during the starter and grower phases, with enrichment of Methylobacterium-Methylorubrum and Pseudomonas during the starter phase and of Escherichia-Shigella during the grower phase. In contrast, the finisher phases exhibited reduced Proteobacteria abundance and increased Firmicutes and Actinobacteriota, including Lactobacillus, Ligilactobacillus, Faecalibacterium, Streptococcus, Staphylococcus, Romboutsia, and Corynebacterium. Overall, the tracheal microbiome underwent progressive maturation, shifting from a Proteobacteria-dominated community to a more diverse, complex, Firmicutes-rich ecosystem. These findings provide new insights into the development of the respiratory microbiome in broiler chickens and may support strategies to improve poultry respiratory health. Because dietary transitions occurred concurrently with age progression, the observed microbiome shifts should be interpreted as production-stage-associated changes rather than diet-specific effects.

Host genetic markers often fail to resolve regional origins in highly connected or panmictic marine species. The Japanese jack mackerel, Trachurus japonicus, is a commercially important fishery species around Japan that shows little or no detectable population structure. Here, we used nationwide multi-omics profiling to compare host genomic variation and gut microbiome composition in wild T. japonicus collected from coastal regions across Japan. We generated MIG-seq data for 43 individuals and 16S rRNA gene profiles for 24 individuals; after quality filtering, 19 individuals remained for matched host-microbiome comparison. Genome-wide host SNP analyses showed weak or absent geographic population structure, consistent with previous evidence of panmixia in Japanese waters. In contrast, gut microbiome composition showed geographic structuring based on Bray-Curtis dissimilarity and PERMANOVA, and this pattern was not explained by proximity to river mouths or host-related variables. Locality- or individual-associated bacterial lineages contributed to the observed differences in the microbiome, while chloroplast-associated and Cyanobacteria-assigned ASVs suggested recent dietary or environmental input. These results indicate that gut microbiome can show regional biological variation not apparent from host genetic markers alone. Our study provides a proof-of-concept example of integrating host genomics and gut microbiome profiling to evaluate regional characteristics and origins in highly connected marine animals.

Industrialization has reshaped human gut microbiomes, but its effects on other human-associated mammals remain poorly understood. Domestic dogs provide an informative comparative system because they have shared human environments and food systems for millennia yet retain distinct host biology. However, most canine microbiome studies have focused on industrialized companion animals, limiting our understanding of the ecological range of the domestic dog gut microbiome. We analyzed fecal 16S rRNA gene profiles from 261 dogs sampled across Nepal, Thailand, the United Arab Emirates, and the United States, spanning forager, agrarian, pastoralist, urban, and industrialized lifestyles; 257 dogs remained after excluding recent antibiotic exposure. Lifestyle was the strongest measured correlate of canine gut microbiome composition, and this structure persisted in restricted analyses of mature, non-shelter dogs sampled from temperate climate regions. Industrialized dogs differed from non-industrialized dogs through directional genus-level turnover, restructuring of VANISH- and BloSSUM-like microbial guilds, and shifts in predicted functional potential. Non-industrialized dogs were not microbiologically uniform: pastoralist dogs carried non-industrialized microbiome profiles but diverged from a simple forager-to-industrialized continuum. Cross-species comparisons with humans sampled across matched lifestyle categories showed parallel lifestyle-associated restructuring in both hosts, but host species remained the dominant axis of variation and the genera responding to industrialization were largely host-specific. These findings expand the ecological baseline for the domestic dog gut microbiome and identify industrialization as a major axis of microbiome restructuring in a long-term human-associated mammal. More broadly, they show that shared lifestyle transitions can impose parallel ecological pressures across host species without overriding host-specific community assembly.

The aim of this study was to evaluate the impact of two sustainable dietary protein sources on the structure and composition of the gut microbiota in European sea bass (Dicentrarchus labrax) juveniles. These protein sources were incorporated to the aquafeeds containing (a) Lupinus albus meal, treated with either exogenous enzymes (Solid state hydrolysis-SSH) or fermented with Saccharomyces cerevisiae (Solid state fermentation, SSF) and (b) Lagocephalus sceleratus meal. In the first case (a), the control aquafeed simulated a standard commercial diet, containing soybean meal whereas in the rest of the diets soybean meal was partially or totally replaced by hydrolysed or fermented Lupin meal. In the second case (b) the fish were fed Lagocephalus sceleratus unprocessed fishmeal as well as treated at different temperatures to deactivate tetrodotoxin (TTX). A control diet with 30% commercial fish meal was also fed as a reference diet. Both diets in all inclusion levels did not cause any significant gut microbiota change, suggesting their neutral role in this aspect. However, the gut bacterial communities of the fish fed with 12.5% lupin meal inclusion, had increased amino acid biosynthetic pathways suggesting a beneficial effect.

Probiotics are widely used in food animal production to support gut health and immune function, but the indicators of probiotic efficacy and the conditions under which they translate to host benefit remain unclear. Microbiota composition is the most accessible data supporting probiotic effects, yet whether compositional change reliably predicts host outcomes is not well understood. We investigated this question in broiler chickens fed two nutritionally similar basal diets, with or without a commercial probiotic. Microbiota composition was profiled across 6 gastrointestinal regions using 16S rRNA sequencing. To assess systemic functional effects, an in vitro assay building on prior observations of elevated circulating immune cell ATP in probiotic-fed animals was developed. In this assay, serum from each treatment group was applied to a chicken T-lymphocyte cell line before ATP quantitation. Basal diet was the primary driver of microbial community structure, with probiotic-induced compositional shifts observed predominantly in one diet context but minimally in the other. Despite this difference, serum from probiotic-supplemented animals increased T-lymphocyte ATP production across both diets, supporting prior findings and revealing a systemic immunometabolic response independent of broad microbiota restructuring. Functional predictions revealed enrichment of pathways related to mevalonate and carbohydrate metabolism in probiotic-supplemented birds within the more responsive diet context, driven largely by Lactobacillaceae family taxa. These findings demonstrate that basal diet modulates the detectability and nature of probiotic effects on the microbiota, but not the physiological host response. This disconnect has implications for how probiotic efficacy is evaluated and for microbiome targeted interventions across species. ImportanceProbiotics are used widely in food animal production to support gut health and immune function, yet predicting which probiotic preparations will produce meaningful effects remains a challenge. Microbiota composition, profiled by 16S rRNA sequencing, is the most accessible measure of probiotic activity, but it captures only one aspect of the host-microbe dynamic. These data demonstrate that probiotic-induced compositional changes vary substantially between basal diets, while the host immunometabolic response is consistent across diets, demonstrating that compositional readouts alone cannot reliably predict host outcomes. The findings have practical implications for how probiotic efficacy is evaluated and inform the broader effort to design microbiome targeted interventions across both veterinary and human contexts.

Anthropogenic pressures are increasingly reshaping wildlife habitats worldwide. These transformations reduce natural areas, but also create new ecological niches, food resources, and environmental stressors, with potential consequences for wildlife behavior, physiology, and morphology. These changes may affect the gut microbiome, a critical component of host health, yet such effects are often inconsistent across species, particularly in wild non-human primates, and remain poorly understood. Here, we investigated how the gut microbiome of chacma baboons (Papio ursinus ursinus), an ecologically flexible generalist, responds to an anthropization gradient. We analyzed 512 fecal samples collected from 33 wild troops across a broad range of anthropogenic environments in the Western Cape, South Africa. Using a multi-metric approach including the Human Footprint Index, land-use variables and dietary proxies derived from stable isotopes, we assessed gut microbial diversity and composition based on 16S rRNA gene (V4) sequencing. Human-altered environments characterized by high Human Footprint and built-up areas were associated with reduced microbial diversity, and compositional and functional shifts, including decline in fiber-degrading taxa and increase in bacteria associated with simple carbohydrate and dairy metabolism. In contrast, highly cultivated areas showed no diversity difference and distinct microbial assemblages, while dietary variation had weaker effects, primarily altering rare taxa. Our results demonstrate that different components of anthropogenic pressure exert contrasting effects on the baboon gut microbiome, reflecting multiple ecological pathways extending beyond diet alone. Microbiome shifts may have implications for host health, potentially increasing susceptibility to pathogens or inflammatory diseases, with consequences for wildlife populations.

The gut microbiota is a key player in energy balance, impacting both digestion efficiency and the production of metabolites involved in metabolism. Its composition is highly adaptable, especially in response to diet. Changes in human diet and lifestyle over time - from active, fibre-rich diets to sedentary habits with calorie-dense foods - have likely contributed to the rise in metabolic diseases. Rodent models are widely used to study the links between diet, microbiota and metabolism. However, they have important limitations (e.g. artificial environments, uniform diets and biological differences from humans) which can affect the translation of findings to humans. While mice and humans differ in their microbiota species, they do share some functional similarities. The grey mouse lemur (Microcebus murinus) has been proposed as a promising alternative model. This small primate experiences strong seasonal changes in food availability, leading to distinct physiological states (energy-saving in winter vs active in summer), even in captivity. It is increasingly recognized as a valuable model for biomedical research, supported by recent genomic and molecular advances. However, its gut microbiota has not yet been the subject of study. Consequently, the present study focuses on investigating the gut microbiota of the grey mouse lemur, with a particular emphasis on how these microbiota vary under different dietary regimens. The microbiota of animals fed the standard colony diet was dominated by Prevotella, Bifidobacterium, Megamonas, Streptococcus, Megasphaera and Lactococcus, showing an Prevotella driven enterosignature. We showed that switch from a classical control diet to 3 different diets resulted in change on microbiota composition that is associated with functional redundancy. The present work underline the interest of Microcebus murinus as model for diet and lifestyle studies in relationship with metabolic diseases.

Anthelmintic resistance in cyathostomins is escalating worldwide, threatening equine health and highlighting the need for sustainable, ecology based parasite control strategies. Chicory (Cichorium intybus, Puna II) has emerged as a promising antiparasitic forage, yet its broader effects on the equine holobiont, parasites, microbiota, and host physiology remain poorly understood. We conducted a 32 day longitudinal grazing trial in young horses to assess how chicory affects parasitological outcomes, gut microbial ecology, nemabiome composition, behaviour, and host physiological and immune responses. Twenty-six naturally infected Anglo-Arabian horses were monitored weekly, with 13 grazing a chicory-based sward and 13 grazing a permanent pasture. Clinical parameters, body weight, and serum biochemistry remained stable across treatments, indicating that chicory was well tolerated. Immune profiles showed limited variation, although IL 10 increased in chicory fed horses, suggesting subtle immune modulation. Behavioural observations revealed no signs of discomfort and indicated slightly enhanced social interactions in the chicory group. Chicory grazing produced a marked reduction in cyathostomin egg excretion, accompanied by species specific shifts in nemabiome composition. Several cyathostomin taxa, including Cylicocyclus ashworthi, C. leptostomus, and C. nassatus, declined in chicory fed horses, whereas certain Cylicostephanus spp increased, indicating differential sensitivity rather than uniform suppression. Concomitantly, chicory induced profound ecological changes in the gut microbiota, including reduced alpha diversity, increased beta dispersion, and destabilised individual microbial trajectories. Several bacterial lineages, particularly Oscillospiraceae, Clostridiaceae, Lachnospiraceae, and Bacteroidales, were differentially enriched, reflecting a functional reorganisation of the intestinal ecosystem. Together, these findings demonstrate that chicory reduces parasite fitness, reshapes nemabiome composition, and alters gut microbial ecology while maintaining host physiological stability. Chicory thus emerges as a promising ecological tool for parasite control, capable of modulating the equine holobiont in ways that complement and potentially reduce reliance on conventional anthelmintic strategies. However, because its effects on gut microbial ecology remain uncertain, and may include shifts resembling dysbiosis, future studies are needed to monitor microbial dynamics more closely and clarify the long term ecological consequences of chicory grazing.

A tightly regulated inflammatory response occurs during and following parturition; however, excessive or prolonged inflammation negatively affects herd health and productivity. The gut microbiota plays an important role in host immunity and metabolism and undergoes substantial changes during the transition period. However, the relationship between systemic inflammation, measured by serum acute-phase proteins, and gut microbial dynamics during early lactation remains poorly understood. We investigated fecal microbiota dynamics in relation to systemic inflammation in early postpartum dairy cows. Fecal and blood samples were collected from 71 Holstein cows on days 1 and 3 in milk (DIM). The V3-V4 region of the 16S rRNA gene was sequenced and microbial diversity, differential abundance, and microbial interaction networks were evaluated. Inflammatory status, defined by fibrinogen, haptoglobin, and their combined classification, was associated with significant alterations in fecal microbial composition during the immediate postpartum period, independent of body condition score, parity, DIM, and DNA extraction parameters. Differential abundance analyses revealed extensive taxonomic restructuring, while network analyses identified increased modularity, altered keystone taxa distribution, and greater compartmentalization of microbial interactions in animals with elevated inflammatory markers. Several taxa were consistently associated with inflammatory status across analytical approaches. Notably, Ruminococcaceae UCG-002 and Dielma were associated with inflammatory states, whereas Xylanibacter, Marvinbryantia, Akkermansia, and Oscillibacter were associated with non-inflammatory status. This study identifies an association between systemic inflammation and fecal microbiota composition in early transition dairy cows, providing a foundation for future microbiome-based biomarkers of inflammatory status. IMPORTANCESubclinical systemic inflammation during early postpartum can negatively affect dairy cow health and productivity, yet current monitoring relies on repeated blood sampling and transient inflammatory biomarkers. For the first time association of systemic inflammation, assessed using fibrinogen, haptoglobin, and their combined classification, with alterations in fecal microbial composition, microbial interaction networks, and keystone taxa distribution during the early postpartum period was established. Several bacterial taxa were consistently associated with either elevated or normal inflammatory states across differential abundance, network, and odds ratio analyses. Many of these taxa remain poorly characterized in dairy cattle, highlighting the need for future mechanistic studies. This study demonstrates that systemic inflammation during early postpartum is associated with measurable alterations in the fecal microbiota. This work provides a foundation for developing microbiome-based biomarkers for detecting and monitoring subclinical systemic inflammation in dairy cattle.

Animal microbiomes are shaped by both environmental exposure and host-associated filtering, but the relative importance of these processes remains poorly understood. Dung-associated insects provide an ideal model because they develop and feed in highly dynamic microbial environments. We investigated the gut microbiomes of six sympatric dung fly species of the genus Sepsis (Diptera: Sepsidae) and compared them with microbial communities in cow dung throughout a growing season in Switzerland. Using full-length 16S rRNA gene sequencing (PacBio), we characterized bacterial communities from 74 fly and 15 dung samples. Seasonal variation was the strongest predictor of microbiome composition, whereas host species exerted weaker effects that persisted after removing dung-associated taxa, indicating that gut communities are not merely passive reflections of environmental exposure. Only few gut microbiome reads were attributable to dung-associated taxa, and environmental overlap differed among fly species rather than season. A highly non-random core microbiome persisted across all six species: 36 bacterial genera (of 469) were shared by all hosts at [~]119-fold enrichment above random expectation and remained after removing dung-associated taxa. These findings support a two-layer model of microbiome assembly, in which seasonal environmental variation determines microbial availability while host-specific processes selectively retain a subset of taxa.

The gut microbiome is increasingly recognized as a pivotal modulator of animal behaviour, yet its influence on wild fauna remains largely unexplored. We investigated the correlational relationship between gut microbiota, behavioural phenotypes, and management practices in 26 captive endangered Iberian lynxs (Lynx pardinus) maintained within the ex-situ Iberian breeding program facilities, in two geographically distant stations in SW Spain. Behavioural observations were intensively recorded over two years, and three personality profiles emerged, i.e., (i) anomalous (with the highest frequencies for stereotypies), and (ii) sedentary and (iii) active (with the highest frequencies for sedentarism and for locomotion and surveillance, respectively). Fecal samples were analyzed for biweekly periods by 16S rRNA gene amplicon sequencing to profile bacterial composition and predicted functional pathways, and significant associations were found for each of the behavioural phenotypes. Both breeding station and local environment influenced gut microbial communities and personality profiles, underscoring the influence of management practices and local habitat in shaping the microbiome-behaviour nexus. Specific bacterial taxa and metabolic pathways were consistently associated with each behavioural phenotype, suggesting that microbial fecal signatures could serve as non invasive biomarkers for individual personality monitoring. This work constitutes the first comprehensive, multi layered examination of the interplay among behaviour, gut microbiota, and environmental factors in a large, wild carnivore. This integrative approach may help conservation programmes to optimize management decisions and improve reintroduction success

On their own, probiotics do not entirely eliminate pathogens during infection and disease. Instead, they often serve as adjuncts that rely on distinct mechanisms to reduce the presence of harmful bacteria. Our prior research indicates that an isolated Aeromonas dhakensis strain A603 kills pathogenic Vibrio strains through two antibacterial mechanisms: the type VI secretion system (T6SS) and phenazine (AdPhen). Here, we investigate A603 as a standalone probiotic for a shrimp disease model. This study shows that A603 prevents mortality from acute hepatopancreatic necrosis disease (AHPND) in shrimp caused by pathogenic Vibrio spp. that produce the PirAB toxin. AHPND infection alters the shrimp microbiota by increasing pathogen abundance and decreasing beneficial bacterial abundance prior to death. As both a prophylactic and treatment, A603 removes pathogenic Vibrio from shrimp and reverses such alterations in the microbiota using both T6SS and AdPhen. Collectively, our findings show that A603 antibacterial mechanisms prevent AHPND.

Environmental change exposes ecosystems, including host-associated microbiomes, to stressors that occur repeatedly and in sequence, yet it remains unclear whether prior stressor history conditions host-microbiome responses to later perturbation. We used adult zebrafish (Danio rerio) to test whether sequential exposure to antibiotics, heat stress, the intestinal nematode Pseudocapillaria tomentosa, or pairwise stressor combinations altered gut microbiome structure, intestinal host gene expression, and host health outcomes. Across eight exposure regimes, prior stressor history and parasite exposure were associated with gut microbiome composition, while increasing prior stressor history was associated with reduced gut microbial diversity and convergence in community composition. Host intestinal transcriptional responses to parasite exposure were historically contingent, with parasite-associated differential gene expression varying non-linearly across prior stressor histories. Cumulative mortality increased with prior stressor history, whereas infection prevalence among surviving hosts decreased. Integrating microbial abundance, host gene expression, mortality, and neutral-community modeling identified Cetobacterium, Culicoidibacter, Flavobacterium, and Shewanella as candidate host-linked taxa associated with host response and survival. Collectively, these findings indicate that prior environmental stressor history shapes vertebrate host-microbiome responses to future perturbation and highlight specific gut microbial members as potential biomarkers or functional targets for follow-up studies.

This study aims to evaluate the effectiveness of curcumin and penicillin-streptomycin as diluents on changes in the structure and diversity of the chicken semen microbiome during storage. Semen was collected from Kampung chickens (native to Indonesia) and divided into five treatment groups: diluted without antibiotics or curcumin (control), and diluted with additions of 10 micromolar, 20 micromolar, and 30 micromolar curcumin, and penicillin-streptomycin, respectively. The semen was stored at 5 C for 24 hours. The composition and diversity of the semen microbiome were analyzed using 16S full-length amplicon sequencing. Analysis of the top 10 species showed that Uncultured Saccharofermentans sp. and Porphyromonas somerae served as the most dominant and stable core microbiome across all treatments. Alpha diversity analysis showed that the addition of curcumin and penicillin-streptomycin reduced microbial richness (Observed, ChaO1, ACE, and Fisher) in a dose-dependent manner, yet maintained overall diversity (Shannon and Simpson), with the penicillin-streptomycin treatment resulting in the highest species evenness (InvSimpson). Beta diversity analysis revealed extreme separation of taxonomic abundance variance in the penicillin-streptomycin group, whereas the curcumin treatment exhibited a dose-dependent pattern of microbial abundance transition. Venn diagram analysis identified 415 OTUs as the core microbiome and confirmed that curcumin acts through selective filtering that stabilizes the ecosystem without triggering the proliferation of opportunistic taxa. Penicillin-streptomycin acts more rapidly and dominantly in suppressing/killing bacterial populations, however, the addition of curcumin is able to modulate the microbial ecosystem in a more balanced manner by suppressing the growth of harmful bacteria without compromising the integrity of the chicken semen environment.

Ruminants obtain nutrients through the microbial fermentation of plant material in the rumen. Xylanibacter ruminicola is a highly abundant bacterial species in the rumen. During fermentation, X. ruminicola utilizes diverse carbohydrates from plant materials to synthesize propionate, a volatile fatty acid providing energy to ruminants. However, variation in pasture quality (e.g, nutrient and fibre content) across seasons and host pregnancy status can alter the rumen microenvironment, potentially affecting microbial activity. Bacteria in culture display distinct methylation (a reversible epigenetic modification capable of gene regulation) changes in response to the growth environment. We hypothesized that the changes to the rumen environment would affect the DNA methylation patterns within the X. ruminicola genome. Rumen fluid from 37 female Brahman cattle (17 pregnant) were sampled across four seasons. DNA methylation profiles (N6-methyladenine, N4-methylcytosine, and 5-methylcytosine) of X. ruminicola across seasons (varying pasture quality) and pregnancy statuses characterized using Oxford Nanopore sequencing. After correcting for relative abundance, DNA methylation levels within the coding DNA sequences of several X. ruminicola genes differed between seasons and pregnancy status. Most of these genes were classified as ExbD/TolR family proteins and related to the protein transport process. Our study demonstrates that the DNA methylation profiles of rumen X. ruminicola genes vary with host environment factors. These results provide insight into the role of bacterial DNA methylation in mediating interactions between bacteria and their environments. Lay SummaryRuminants rely on rumen microbes to convert plants into nutrients. Xylanibacter ruminicola is a bacterial species in the rumen that produces nutrients for ruminants during plant fermentation. Changes in pasture quality and host pregnancy status can influence the activity of X. ruminicola, as reflected in the DNA methylation profile across its genome. DNA methylation is a reversible DNA modification that can affect gene activity and help bacterial adaptation to changing environments. Rumen fluid from 37 female Brahman cattle (17 pregnant) across four seasons were used to evaluate the pasture quality and host pregnancy effects on the DNA methylation profile of X. ruminicola. The relative abundances of X. ruminicola were influenced by pasture quality, but not by host pregnancy. However, host pregnancy status and changes in pasture quality influenced the DNA methylation signatures of X. ruminicola. Genes with DNA methylation changes were associated with the protein transport process. These findings suggest that the DNA methylation profiles of rumen X. ruminicola vary with host environment factors. Teaser TextThis study demonstrates that the DNA methylation profiles of rumen Xylanibacter ruminicola vary with host environment factors. These findings provide insight into the role of bacterial DNA methylation in mediating interactions between rumen bacteria and their environment.

Gelatinous zooplankton are abundant predators and prey in marine food webs, yet their role in helminth (parasitic worms) transmission remains poorly resolved. Here we combine a global synthesis of published records with new morphological and molecular observations from the Mediterranean, Red, Celtic, Baltic and North Seas to assess the ecological role of helminths associated with gelatinous zooplankton. We compiled 431 host-parasite association records from 89 sources, including 23 new records, and show that helminth occurrence and richness in gelatinous hosts are concentrated at temperate latitudes, contrasting with the classical latitudinal diversity gradient. Our sampling revealed markedly higher parasite prevalence, abundance and diversity in the Red Sea than in the Mediterranean, while no helminths were detected in gelatinous zooplankton sampled from the Baltic and North Seas. All helminths recorded were larval stages, indicating that gelatinous zooplankton function as key intermediate hosts in marine helminth life cycles. Molecular analyses identified cestodes, nematodes and digenean trematodes associated with cnidarians, ctenophores, and chaetognaths, including the first record of trematode larvae in a pelagic tunicate. Our findings challenge the view of gelatinous zooplankton as dead-end hosts and identify them as overlooked vectors that may shape marine parasite biogeography, food web connectivity and invasion dynamics.