Poultry Science

IntroductionAntimicrobial Resistance (AMR) presents a critical public health challenge, particularly in smallholder broiler farming, where antibiotics are often used preventively in the absence of effective biosecurity measures. ObjectiveThis study investigates the adoption of biosecurity practices as a sustainable alternative to antibiotics through Participatory Systems Mapping and Experimental Games. MethodsA participatory mixed-methods study was conducted in southern Bangladesh (September 2024-June 2025). Causal Loop Diagrams (CLDs) were co-created with farmers, dealers, and veterinary officers. Ten broiler farmers from single village were selected via purposive and snowball sampling. Experimental games simulated four production cycles where farmers chose Option A (biosecurity, adopters) or Option B (antibiotics, non-adopters) after several interactive trainings. Key metrics including biosecurity compliance (0-12 scale), mortality, FCR, antibiotic use, outbreak history, and economic outcomes were recorded. ResultsCLD analysis revealed a reinforcing loop of increased antibiotic reliance driven by fear of mortality, and balancing loops involving training, biosecurity practices, and consumer incentives to reduce use. Five farmers chose Option A, and both groups remained stable until Round 4. Adopters had flock sizes of 800-2000 birds (non-adopters, 600-1000; mean for both = 1000), were younger, and more educated compared to non-adopters. At baseline, both groups had similar biosecurity scores (0). Adopters had higher mean outbreaks (2 vs. 1.4), mortality (5.6 vs. 4.2), antibiotic use (3.6 vs. 3), and FCR (1.8 vs. 1.6) compared to non-adopters. By Round 4, adopters improved biosecurity scores by 125%, eliminated outbreaks, reduced mortality by 52.6%, stopped antibiotic use, improved FCR by 13.3%, and gained 71.72% profit per bird compared to non-adopters. Non-adopters, influenced by adopters, increased biosecurity scores by 25%, reducing outbreaks, mortality, antibiotic use, and FCR. Adopters also increased direct sales to consumers, yielding a 10%-16% profit gain per bird each round. ConclusionThis study highlights the successful adoption of biosecurity practices by farmers, replacing antibiotics and improving production outcomes. Farmer-driven adoption of these practices fosters long-term sustainability and supports a healthier planet within the One Health framework.

Restlessness is a symptom of chronic boredom in humans and a behavioural phenomenon anecdotally described as a concern in bulls raised for fattening purposes, but it has so far not been addressed in research. The two studies presented in this paper aimed to gain first insights into restlessness in bulls. We operationally defined restlessness by the number of transitions between behaviours in a given time period, and quantified restlessness in bulls of different weight classes (300, 400, 500 kg) on farms keeping bulls on fully-slatted floors (n=8, Study 1) as well as across three different husbandry systems (fully-slatted floor (FS, n=4), straw-based (SB, n=4) and organic pasture (OP, n=3), Study 2). All farms were visited twice, and the behaviour of different individuals was continuously recorded for 15 minutes each between 9 a.m. and 5 p.m. (Study 1) and for 8 minutes each between 6 a.m. and 10 p.m. (Study 2). The effects of weight class and husbandry system were analysed using generalised linear mixed-effects models, and we ran a sequence analysis to cluster observations by the sequence, frequency, and duration of bulls behaviours in Study 1. Bulls kept in fully-slatted floor systems in Study 1 changed their behaviour on average 48.3 times per 10 minutes, with high variability both within and across farms. Weight class did not have a statistically supported effect on the number of transitions, and the sequence analysis revealed four clusters that differed in sequence and in the number of transitions. In Study 2, OP bulls showed fewer transitions than SB and FS bulls (X22 = 23.6, p < 0.001), while SB and FS bulls did not differ. While SB pens were more structured and offered more space per animal, both SB and FS systems can be characterised by monotony, which may explain the similar level of restlessness in both systems. Alternatively, or in addition, the high feeding intensity in SB and FS systems may have caused the higher number of transitions compared to the OP system, potentially elicited by subacute ruminal acidosis and/or laminitis and the resulting pain. However, these explanations are speculative and require systematic disentanglement in future studies. This study provides initial insights into restlessness in bulls and lays the groundwork for future research to identify the causes underlying restlessness and investigate its association with bull welfare.

Enteric methane emissions from ruminant livestock contribute to global warming, creating an urgent need for effective mitigation strategies that do not compromise animal productivity and welfare. Methanogenic archaea within the rumen microbiome drive enteric methane emissions. However, large-scale rumen-fluid sampling in commercial production systems is impractical, due to its invasive nature and the associated logistical challenges. This study hypothesised that rumination enables the capture of rumen microbial signals within the oral cavity and using oral microbiome profiles to provide a practical, non-invasive alternative method for proxy methane phenotyping in commercial production systems. To test the hypothesis, we estimated the oral microbiability, defined as the proportion of phenotypic variance in methane emissions explained by oral microbiome variation. Samples were collected from 209 animals across two trials in Queensland, Australia. Oral microbiome samples were obtained from all animals, with paired rumen samples in one trial, and methane emissions were measured using either the sulphur hexafluoride (SF6) tracer technique or the GreenFeed system. Microbial features were characterised using taxonomic and functional annotations, and microbiability was estimated using mixed linear models incorporating microbiome-based relationship matrices. Although the small sample size limited strong conclusions, the oral microbiability estimates reported in this study were comparable to those derived from rumen samples. Functional microbial profiles generally explained a greater proportion of methane variation than taxonomic profiles, suggesting that microbial function is more closely linked to methane production than community composition alone. However, these differences were not statistically significant due to large standard errors. These findings suggest that oral microbiome sampling potentially provides a practical, minimally invasive, scalable proxy method for methane emissions of individual cattle in grazing systems, where direct methane gas measurements are labour-intensive and difficult to implement. Integrating oral microbiome profiles in the existing breeding model with the host genetics, weight and environmental factors could provide a promising pathway for enabling selection for low emissions and advancing reduced emissions livestock farming under real-world production conditions. Lay summaryCattle produce methane as part of their normal digestion and this contributes to climate change. Reducing methane emission in grazing livestock systems is therefore important. However, measuring methane from individual grazing animals is difficult, costly, and often impractical under commercial conditions. The rumen microbiome has been used as a proxy for estimating methane emissions, but collecting rumen samples is invasive and impractical for large-scale use. Because rumination transfers material from the rumen to the mouth, we investigated whether microbes found in cattle mouths could also be used to estimate how much methane an individual animal produced. We suggest that mouth-swab sampling method can be an alternative to rumen fluid sampling because it was less invasive, relatively quick and practically applicable in commercial conditions. Importantly, the microbiome explained a meaningful proportion of the between-animal variation for methane emission. This suggests that collection of mouth swabs is a potentially scalable alternative proxy method to identify cattle that naturally produce less methane. Overall, our findings support the potential use of oral ruminant microbial information to improve breeding and management strategies aimed at reducing methane emissions while maintaining productive livestock systems. Teaser TextThis study demonstrates that collecting oral swabs from the mouths of grazing beef cattle could provide a scalable method to estimate individual methane emissions in commercial production systems, offering a practical alternative to invasive rumen sampling and complex gas measurement systems. These findings support the development of scalable breeding and management strategies for methane mitigation in large-scale livestock production systems.

Reported avocado dietary fiber (DF) content and composition are inconsistently reported, particularly during ripening. Thus, this study aimed to characterize the amount and type of DF in Hass avocados and evaluate DF changes during ripening. Unripe (day 0), ripe (day 5), and overripe (day 12) Hass avocados were freeze-dried and defatted. DF was analyzed using non-starch polysaccharide (NSP) enzymatic-chemical methods. Per 100g of as-is avocado, unripe contained 3.96g total DF, ripe 3.68g, and overripe 3.26g. In ripe avocados, DF comprised 43% soluble (SDF) and 57% insoluble dietary fiber (IDF). SDF consisted primarily of rhamnogalacturonan-1 and arabinan pectins, while IDF was predominantly cellulose (32%), hemicelluloses (23%), and lignin (2%). Total DF decreased with ripening, with pectin undergoing solubilization and depolymerization, while cellulose and hemicelluloses remained stable. These findings are important as dietary fibers differentially influence intestinal microbial fermentation and health benefits.

Managing post-weaning diarrhoea (PWD) in piglets is difficult due to limits on antibiotics and zinc. Chitosan is emerging as a potential feed additive. We analysed a chito-oligosaccharide hydrochloride (COS-HCl), a low molecular weight (LMW) chitosan, and a medium molecular weight (MMW) chitosan, and assessed their effects on growth, faecal consistency, microbiota, and potential interference with enterotoxigenic Escherichia coli (ETEC). The three chitosans were characterised using {superscript 1}H-NMR, SEC-RI-MS, and SEC-RI-MALLS. COS-HCl had an Mw of 0.824 kDa; LMW and MMW showed Mw ranges of 14.4 kDa (0.3-30 kDa) and 116 kDa (15-600 kDa). Degrees of acetylation were 9.5%, 6.5%, and 15%. Two 42-day field studies evaluated average daily gain (ADG), faecal consistency, and microbiota. In the first trial, COS-HCl at 0.025-0.1% did not significantly affect ADG (-33 to - 12 g/d). In the second, LMW and MMW at 0.01% did not significantly change ADG (-7 and +3 g/d). Faecal consistency, ETEC shedding, and microbiota composition were similar to controls. An enzymatic HPLC-MS method enabled quantification of MMW chitosan in premix. Our results highlight the importance of advanced chitosan characterisation for precision nutrition and suggest that a threshold dosemay be needed to benefit growth and gut health in PWD management. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/714014v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@19c9e23org.highwire.dtl.DTLVardef@152461aorg.highwire.dtl.DTLVardef@7886e0org.highwire.dtl.DTLVardef@df0d9b_HPS_FORMAT_FIGEXP M_FIG C_FIG

Sentinel behaviour occurs when individuals use raised positions to scan for predators while the rest of the group forages. Here, we investigated whether a colonial cooperatively breeding species that forages in large groups, the sociable weaver, Philetairus socius, displays sentinel behaviour. This behaviour has been reported in species with similar ecology, behaviour and foraging habits, (e.g. ground foraging in open habitats where aerial predators are common) and, hence, we expected that it could occur in sociable weavers. On the other hand, sentinel behaviour appears to be less common in species that live in very large groups. We used an experimental set-up consisting of an artificial feeding station and perches to assess occurrence of sentinel related behaviours: (i) perching events > 30s on an elevated position, (ii) head-movements and (iii) alarm calling. Birds were seldom observed perching while others fed, and those that did, perched for periods that were too short to be considered as sentinel behaviour (less than 5s on average). Our results suggest that this behaviour is uncommon or even absent in sociable weavers. We discuss whether other factors such as foraging in very large groups, or interspecific foraging associations might make sentinel behaviour less important in this species.

BackgroundThis study investigated rumen microbiome reconstitution and methane (CH4) emissions following a complete exchange of rumen contents between low- and high-CH4-yielding Norwegian Red dairy cows. Twenty cows were screened for CH4 yield, and two low and two high emitters were selected for rumen cannulation and content swap. Total rumen contents were swapped after complete evacuation and washing of both the rumen and omasum. Rumen samples were collected twice in weeks -1, 1, 3, and 7 for fermentation analysis, metagenomics, and metaproteomics, and at week 8 CH4 production was measured. ResultsPrior to the swap, low and high emitters produced 21.2 {+/-} 0.7 and 26.3 {+/-} 1.4 g CH4/kg dry-matter intake (DMI), respectively. Eight weeks after swap, CH4 yields were 12.7 {+/-} 0.3 and 28.9 {+/-} 0.3 g CH4/kg DMI, respectively, showing that the CH4 phenotype of each cow was maintained. Analysis of metagenome-derived 16S rRNA gene sequences showed that low emitters gradually re-established their original microbial community, whereas high emitters retained donor-like microbiota. Metaproteomic mapping suggested higher expression of Prevotella-associated succinate-propionate pathway enzymes in low emitters at week 7, though these differences were modest. ConclusionThese findings suggest that host factors influence CH4 output and microbial reconstitution, with low emitters restoring their native microbiome while high emitters retained a donor-associated community yet continued to emit high CH4. Results should be interpreted with caution given the small sample size (n = 2 per phenotype) and require confirmation in larger studies. ImportanceReducing enteric methane from cattle requires understanding whether the rumen microbiome or the host animal is the primary driver of methane output. We exchanged the entire rumen contents between low- and high-methane-yielding dairy cows and measured methane production alongside metagenomic and metaproteomic profiling over two months. Despite receiving each others microbiomes, each cows methane phenotype persisted--low emitters stayed low and high emitters stayed high. Microbiome reconstitution was asymmetric: low emitters restored their original microbial community, while high emitters retained the donor microbiota. Methanogen communities did not differ between phenotypes, pointing to host-level rather than microbial-level control of methane yield. These pilot findings suggest that breeding for favorable host traits may be essential for lasting methane reduction, and that microbiome transfer alone is unlikely to shift an animals methane phenotype. Larger studies are needed to confirm these observations.

Enterohemorrhagic Escherichia coli O157:H7 (EHEC) is an important zoonotic pathogen that disrupts intestinal epithelial barrier integrity and induces excessive inflammatory responses, thereby leading to impaired growth performance and intestinal injury. EHEC is also an important cause Hemolytic Uremic Syndrome (HUS) in children and older adults. In pig production, chitosan is considered a promising alternative to antibiotics due to its bioadhesive and antimicrobial properties, but the effects and underlying mechanisms of chitosan (COS) under pathogenic challenge remain to be elucidated. One hundred and eight pigs were randomly divided into three treatments: an unchallenged control group (CON), an EHEC-challenged control group (ECON), and an EHEC-challenged group supplemented with 100 mg/kg COS (ECOS). Results show that EHEC challenge increased the feed conversion ratio (FCR), increased inflammatory cytokine levels, disrupted intestinal morphology, and downregulated tight junction and nutrient transporter gene expression (P<0.05). Dietary COS supplementation significantly improved average daily gain (ADG) and FCR during day 6-14 (P<0.05). Moreover, COS reduced fecal shedding of total E. coli (P = 0.085) and EHEC, attenuated systemic inflammation by decreasing serum TNF- and IL-6 levels, and enhanced humoral immunity as indicated by increased IgA and IgM concentrations (P<0.05). Importantly, COS alleviated EHEC-induced intestinal injury by restoring villus height and villus-to-crypt ratio, with enhanced mucosal digestive enzyme activities, and upregulated expression of tight junction proteins (ZO-1 and occludin) and nutrient transporters (SGLT-1 and PEPT1) (P<0.05). In conclusion, these findings indicate that dietary COS improves growth performance in EHEC-challenged weaned pigs, with enhanced intestinal barrier integrity and nutrient transport capacity.

Mycoplasma bovis was first detected in cattle in New Zealand in 2017, prompting an eradication programme that incorporated extensive surveillance and a test-and-cull policy. Genome sequence data and phylodynamic models were used to inform decision making throughout the eradication programme. Isolates from 697 cattle on 126 farms were collected and sequenced between July 2017 and December 2023. Phylodynamic models were used to estimate the time of most recent common ancestor, the effective reproduction number (Reff) and effective population size, and long-range and local between-farm transmission dynamics. The analysis revealed the dramatic impact of movement restrictions and culling up to early 2020, with a sharp reduction in the Reff to less than 1 in 2018/9 and the extinction of two of three major lineages in 2020. This was followed by three-years of residual infection in farms in the South Island, associated with persistent infection of a large feedlot farm and nearby farms. The comprehensive dataset of genomic and epidemiological data provided a unique opportunity to study the dynamics of a country-wide outbreak of a single-host pathogen from first detection to potential eradication, underlining the utility of integrated genomic surveillance during an outbreak response. Author summaryThe economically important cattle pathogen, Mycoplasma bovis, was first detected in New Zealand in 2017. This led to a large-scale, successful control programme aimed at eradication of the pathogen. The decision to undertake an eradication programme was informed by initial analyses of whole genome sequences from isolates collected as part of the surveillance programme. The analysis showed that the bacteria had entered New Zealand relatively recently and was unlikely to be widespread. Over the subsequent years, genome sequencing and modelling of transmission dynamics informed important policy decisions made by the New Zealand Government and the cattle industry, and helped to monitor progress of the eradication programme. The impact of the detection, movement control and culling programme was profound, with sharp reductions in transmission between 2018 and 2020. This was followed by a long tail of localised infection in the South Island, involving transmission from a large feedlot farm. Provisional eradication was achieved after depopulation of this feedlot. This analysis highlights the role of genomic surveillance and modelling to inform decision making during an infectious disease outbreak.

Iron is an essential nutrient for all types of organisms, including insects and the microbes that infect them. We predicted that insects fed an iron-supplemented diet would accumulate more iron in their hemolymph, and, because infectious microbes acquire iron from their hosts, that this extra iron would increase the severity of bacterial infections. To test this hypothesis, we studied the effects of dietary iron supplementation on infection outcomes in Manduca sexta (tobacco hornworm). Larvae were fed an artificial diet, with or without antibiotics, or the same diets supplemented with 10 mM iron. Control and iron-treated larvae were inoculated with non-pathogenic Escherichia coli or the entomopathogenic Enterococcus faecalis, and bacterial load and larval survival were measured. We found that dietary iron supplementation increased the iron content of hemolymph by approximately 20 fold; however, contrary to our prediction, this increase in iron did not result in an increase in the bacterial load of either E. coli or E. faecalis. The effect of iron supplementation on survival was more complicated. As expected, for larvae inoculated with nonpathogenic E. coli, iron supplementation had no effect. For larvae inoculated with E. faecalis, the effect of iron supplementation depended on whether antibiotics were present in the diet. Without antibiotics, iron supplementation prolonged larval survival; with antibiotics, iron supplementation decreased larval survival. The results of this study do not support the hypothesis that dietary iron supplementation increases infection severity in M. sexta. Instead, the results support the viewpoint that the relationship between dietary iron and infection outcome is complex.

This study was aimed at characterizing the physicochemical analysis of stingless bees honey (SBH) in the Wonchi district, Southwest Shewa Zone, Ethiopia. In this study, a total of 30 stingless bees honey samples were collected from Damu Dagele, Fite Wato, and Warabu Messe sites from underground soils through an excavation of natural nests. Physicochemical characterization of properties and proximate analysis of the honey were performed. The result showed a total mean of 20.12{+/-}1.14% moisture content, 8.62{+/-}2.73 meq./kg free acidity, 1.8{+/-}0.52 mS/cm electrical conductivity, 3.39{+/-}0.32 pH, 40.52{+/-}6.61 mg/kg HMF, 0.83{+/-}0.33% ash, 0.56{+/-}0.25% protein, 0.56{+/-}0.24% fat, and 0.59{+/-}0.23% WISC for physicochemical properties of stingless bees honey. Among sugar profiles of SBH, fructose constituted the highest proportion at 18.87 g per 100 g (53.87%), while sucrose exhibited the lowest concentration at 5 g per 100 g (14.33%). The result showed that the highest constituted mean of mineral composition was observed with potassium (K) of 16.64{+/-}0.257 mg/kg, while magnesium (Mg) showed the lowest concentration of 3.48{+/-}0.17 mg/kg. A substantial correlation was observed between K and Mg, with a correlation coefficient of 0.72 and 0.72, and similarly between K and Calcium (Ca); the correlation was highly significant, exhibiting a correlation coefficient of 0.65. Furthermore, the correlation between fatty and other physicochemical and proximate analyses showed very insignificant correlations. In general, this study showed that the SBH produced in the current study area has good physicochemical properties and moisture and contains high-quality honey, which may help its traditional medicinal uses. The findings of the study further suggests the potentiality of the area for quality honey, and to easily locate priority areas for stingless bee conservation, further detailed studies of other stingless species honey medicinal values are recommended.

Functional dairy products are increasingly recognized for their ability to provide both essential nutrition and additional health benefits. This study aimed to develop and evaluate a synbiotic yogurt enriched with Lactobacillus plantarum as a probiotic and Stevia rebaudiana extract (1% w/v) as a prebiotic source. Thirteen lactic acid bacteria (LAB) strains were isolated from fermented dairy and vegetable samples and evaluated for probiotic potential through tests for acid and bile tolerance, hydrophobicity, aggregation abilities, and pathogen co-aggregation. Isolate PG1 (Lactobacillus plantarum) demonstrated the highest prebiotic growth stimulation index (49%) in the presence of stevia extract and was selected for yogurt formulation. Yogurt samples were prepared and stored at 4{degrees}C for 10 days. Physicochemical properties (pH, titratable acidity, and protein content), microbiological viability, total phenolic and flavonoid content, antioxidant activity (DPPH assay), and sensory attributes were monitored. The synbiotic yogurt (St-Y) showed enhanced functional properties, with a total phenolic content of 16.67 {micro}g GAE/g, a flavonoid content of 6.28 {micro}g QE/g, and 57.84% antioxidant activity. Additionally, it showed improved protein content and superior sensory scores compared to control samples. These findings suggest that S. rebaudiana fortified probiotic yogurt can serve as a nutritious, antioxidant-rich, and sensory-acceptable functional dairy product.

Environmental factors affect crop growth and development thus their consideration across sites and years become essential for genotypic evaluation. Genomic selection (GS) has been broadly implemented to accelerate breeding cycles by skipping field evaluations thus allowing early identification of outperforming genotypes. In this study, 7,740 phenotypic records corresponding to 516 Miscanthus sacchariflorus genotypes evaluated in five locations across three years were considered for analysis. Additionally, environmental data on six weather covariates was implemented to characterize similarities between locations. Different sets of locations of variable sizes were used for model calibration based on two cross-validations (CV00 and CV0) schemes leaving out one location at a time. Predictive ability across locations of the best model varied between 0.45 and 0.90 for both schemes. These results were compared to associate predictive ability in function of weather patterns between training and testing sets to allow models calibration optimization. We found it is feasible to optimize resource allocation by considering environmentally correlated sets. In most cases, the information from only one and, at most, two locations were enough to deliver better results than using all four locations, reducing training sets by up to 75%. The results obtained shed light on helping breeders make informed decisions considering weather data when designing evaluations.

RNA viruses are common in tephritid fruit flies including the Queensland fruit fly, Australias most significant horticultural pest. For many their transmission, tissue tropism and load across host development remain unexplored. Yet these factors are important for host biology, ecology and pest management. We investigated Bactrocera tryoni orbivirus (OV), Bactrocera tryoni xinmovirus (XV), Bactrocera tryoni toti-like virus (TLV) and Bactrocera tryoni iflavirus species 2 (IVsp.2) that commonly coinfect B. tryoni laboratory populations. OV and XV transmission was vertical within and on eggs, while TLV transmission was vertical within eggs. IVsp.2 was not detected in eggs but was present in adults; however, IVsp.2 was horizontally transmitted, with viral load increasing with cohabitation time with infected flies. Horizontal transmission was not observed for the other viruses. OV had a similar load across all tissues, while XV was consistently more abundant in ovaries. TLV had a high viral load in the brain whereas IVsp.2 was abundant in the thorax, foregut and midgut. Besides differences in eggs, the viruses were detected in all other developmental stages, but viral load patterns differed: viral load remained constant for TLV, fluctuated for OV and XV, and was low in pre-adult stages and high in adults for IVsp.2. Our findings demonstrate distinct transmission strategies and tissue tropism among the viruses, providing new insights into their epidemiology and role in host biology. Furthermore, contrary to prevailing views that viruses are generally horizontally transmitted, most known RNA viruses of B. tryoni are vertically transmitted affecting the evolution of host-virus interactions.

Gastrointestinal disorder caused by the ingestion of (oo)cysts of Cryptosporidium and Giardia is one of the major health problems in developing countries. Fruits and vegetables that are usually consumed unpeeled, poorly washed and or cooked and are the major modes of transmission. Frequent large-scale screening of the food samples is necessary to prevent outbreaks but screening of vegetables for such microbes is limited in Nepal. In this study, we used a smartphone microscopy system to study prevalence and quantification of (oo)cysts of Cryptosporidium and Giardia in 651 vegetable samples collected from nine major vegetable collection sites across Nepal. The overall prevalence rate of vegetable samples was 37.5% with at least with one of the parasites. We found that 23.2% samples were contaminated with Giardia and 33.3% samples were contaminated with Cryptosporidium. Among eight vegetable types, the prevalence rate was lowest in carrot (20%) and highest in spinach (48%). The prevalence rate of vegetable samples at different sites ranged from 13% in Dhading to 61% in Dhangadi. The contamination rate was 28% for winter, 43% for summer and 33% for monsoon seasons in samples collected from Kathmandu. These vegetables should be considered as a potential source of parasitic contamination in people. These vegetables can cause infection if consumed poorly washed and or cooked, posing a potential source of parasitic contamination in people.

We assessed whether pigs provide consolation, referring to targeted affiliation that attenuates a partners stress, under experimental conditions that manipulated exposure to stressed partners. Using a within-subject design, 74 pigs were tested in three contexts: a helping task in which group members could observe and help a trapped focal pig to return to the group, a direct-reunion, in which group members were naive to the experience of a separated focal pig until reunion, and an undisturbed control. We measured affiliative and non-affiliative interactions, anxiety behaviours and changes in salivary cortisol. Only the helping context satisfied most consolation criteria: there were selective increases in unidirectional affiliative contacts from the observer to the focal pig, non-affiliative interactions remained at baseline, and focal pigs showed fewer anxiety behaviours. In contrast, direct-reunions triggered increases in affiliative and non-affiliative interactions and higher anxiety. Cortisol increased during both direct-reunions and helping, but its level was not linked to affiliation. Results add to growing evidence for consolation behaviour in pigs and suggest best practices for reintegrating pigs into groups. Graded reintroductions that allow observers to assess the emotional state of targets may promote social buffering, whereas abrupt regrouping may trigger more generalized arousal or personal distress.

The caliciviruses include important human and animal pathogens such as norovirus, sapovirus and feline calicivirus. Viral reverse genetics is performed to understand the fundamental biology of these viruses, as well as a potential route to generate live-attenuated vaccines. Calicivirus reverse genetics systems have typically relied on either on the production of in vitro-transcribed RNA or plasmid-based rescue either from a mammalian promoter, or through supplementing with helper enzymes through means of a helper virus. Here, we present a novel system integrating vaccinia capping enzymes D1R and D12L encoded on plasmids as part of a system for Murine Norovirus (MNV) reverse genetics. Addition of D1R, D12L and T7 RNA polymerase-expressing plasmids increases the viral titres of rescued MNV in both BSR-T7 cells and transgenic BSR-T7CD300LF cells, and viral polyprotein abundance. When the murine norovirus receptor is expressed in BSR-T7CD300LFcells, viral titres increased 100-1000-fold compared over standard BSR-T7 cells. This system offers a robust, high-throughput means of assessing viral mutants.

It has been known for decades that bacteriophages encode tRNA genes, but their function and the factors contributing to their acquisition and retention are unclear. Although tRNAs are found in a variety of phages infecting a variety of bacteria, many large-scale computational studies investigating tRNA acquisition and retention in phages are specific to Mycobacterium phages; however, these findings may not be representative of other phages or bacteria. This work uses a broader sampling of phages and hosts to investigate the relationships between codon usage bias, infection cycle, and tRNA gene numbers in phage genomes. We analyzed 154 phages infecting 7 host genera, including Gram-negative (Escherichia, Shigella, Salmonella) and Gram-positive (Bacillus, Lactobacillus, Staphylococcus, Mycobacterium) bacteria. Phages included temperate and virulent representatives, plus a range of tRNA numbers and morphologies. All phages and hosts were analyzed using four metrics: GC content, Effective Number of Codons, Relative Synonymous Codon Usage, and tRNA Adaptation Index. On a global scale, virulent phages with many tRNA genes show greater differences in codon usage and codon adaptation compared to their respective hosts. Gram-negative bacteria and their phages generally exhibit greater differences in codon usage compared to Gram-positive bacteria and their phages. Phages infecting Gram-negative hosts also tend to encode more tRNA genes. In nearly all genus-level comparisons, Mycobacterium phages were different from any other host and from global patterns. This suggests previous computational studies performed in Mycobacterium phages are likely not applicable on a global scale or to phages infecting other host genera. AUTHOR SUMMARYBacteriophages, or phages, are viruses infecting bacteria. They are abundant in all environments, yet how they interact with their bacterial hosts is still not well-understood. Like other viruses, phages must rely on the host translational components to replicate and form new phage particles; and similarly to other parasites, phages have genomes that differ significantly from their hosts in terms of composition. In this work, we explore the relationship between phage lifestyle, number of tRNA genes encoded, and genome differences from the host using a variety of phages and their associated hosts. Phages can be either virulent (do not integrate into the host genome) or temperate (capable of integrating into the host genome), with differences from the host genome more pronounced in virulent phages. There are many phages that also carry tRNA genes, and having higher numbers of tRNAs is associated with larger differences from the host genome. The findings here indicate that virulent phages carrying large numbers of tRNAs diverge the most from host genome composition.

Numerical abilities are widespread in the animal kingdom and are not exclusive to humans. Domestic chicks (Gallus gallus) have been shown to discriminate numerosities spontaneously, but prior research has focused exclusively on the visual modality. Whether chicks can discriminate numerical information in the auditory domain remains unknown, despite evidence that they can perceive other auditory features such as tone and rhythm. In this study, we investigated spontaneous numerical discrimination in the auditory modality in naive domestic chicks. In Experiment 1, newly-hatched chicks were tested for their ability to discriminate between two auditory sequences differing in numerosity (4 vs. 12 identical sounds), with and without controlling for continuous variables such as duration and total sound amount. Experiment 2 examined chicks filial imprinting responses to familiar or unfamiliar numerosities. Experiment 3 controlled for potential spontaneous preferences for a single longer sound versus a shorter one. Our results showed a preference for the 12-sound sequence only when duration and total sound amount were not matched. When these continuous variables were controlled, no spontaneous numerical preference emerged. Experiment 2 revealed an overall preference for the 12-sound sequence regardless of imprinting conditions, while Experiment 3 confirmed that chicks do not have an inherent preference for longer sounds. These findings suggest that chicks are sensitive to overall magnitude in the auditory domain but do not spontaneously discriminate numerical differences when other continuous variables are held constant. Future studies will explore how specific stimulus features, such as heterogeneity of sounds, influence these preferences.

Capturing and handling wild animals is essential for ecological and evolutionary research, yet their effects on physiology, behaviour, and reproductive success remain poorly understood. We investigated short- and longer-term consequences of a capture-handling-restraint protocol in wild great tits (Parus major) over three breeding seasons. To assess short-term responses, we measured circulating corticosterone, a metabolic hormone that responds to unpredictable challenges, and automatically recorded provisioning behaviour. We also explored whether environmental and individual traits were related to provisioning latency (i.e., time to resume provisioning after capture). To evaluate longer-term effects, we monitored provisioning in the days following capture and related it to reproductive success (fledgling number and body condition). We predicted that longer handling would increase stress-induced corticosterone and provisioning latency, that these variables would be positively correlated, and that higher corticosterone and longer latencies would be associated with lower reproductive success. After capture, great tits showed elevated corticosterone and delayed provisioning. Contrary to our predictions, handling duration was negatively associated with stress-induced corticosterone in males (but not females) and did not affect provisioning latency. Provisioning latency was unrelated to corticosterone, environmental, or individual variables. Following capture, parents resumed provisioning, and short-term responses had little influence on reproductive success. We show that parental behaviour and physiology are affected by capture restraint protocols on the short term, but offspring condition and survival are not. However, these results should be interpreted cautiously, as our study lacks an uncaptured control group. Our findings highlight that evaluating welfare impacts requires rigorous study design incorporating both immediate and longer-term behavioural and fitness effects.