Inflammation and immunity in stocker cattle are active areas of research, but much remains poorly defined. Evidence to date indicates stocker cattle undergo rapid immunological transitions during stressful events such as weaning, moving through marketing channels, transport, commingling, and nutritional adaptation. These events appear to activate neuroendocrine stress pathways and systemic inflammatory responses while simultaneously depressing protective functions, especially those mediated by innate immune cells. This leads to a short, high-risk period where viral exposure, bacterial colonization, and dysregulated host responses converge to influence bovine respiratory disease and other disease outcomes.

Mannheimia haemolytica is a pathobiont bacterium of the upper respiratory tract in cattle and is associated with acute bovine respiratory disease (BRD). Despite research into bacterial pathogens and their associations with BRD, little is known about how bacterial variant dynamics interplay with disease progression. Therefore, our objective was to explore the clonal diversity of M. haemolytica in beef cattle utilizing whole genome sequencing. Ninety-nine M. haemolytica isolates from a commingled cohort of 33 cattle sampled in a prior study were selected for analysis. Long-read whole-genome sequencing was conducted to evaluate antimicrobial resistance genes, virulence factors, plasmids, and single nucleotide polymorphisms. Clonal lineages were derived using multi-locus sequence typing (MLST), a recently developed genomic sequence variant (GSV) method, and a core SNP-based clustering analysis with the average linkage method. We identified no differences in pathogenic profiles across isolates. We identified a single sequence type (ST1) and a single GSV (GSV 2) utilizing the MLST and GSV methods, respectively. The core SNP-based average linkage clustering analysis produced different clonal lineage estimations depending on the core SNP cutoff values (2 to 18 clusters). All estimations were greater than the single sequence type and GSV suggested by the MLST and the GSV methods. BRD status did not correlate with clonal interpretations. Our results suggest that method selection can influence the clonal interpretation of M. haemolytica within a population but may not reflect pathogenic capability. Alternate methods that increase discriminatory power or transcriptional profiles may provide new insights and should be evaluated.IMPORTANCEBovine respiratory disease remains a major challenge for cattle health and production, yet the population dynamics of key bacterial pathogens remain poorly understood. Using long-read whole-genome sequencing, this study shows that clonal diversity estimates of Mannheimia haemolytica are highly dependent on the analytical method employed, despite uniform virulence gene content. These findings highlight the importance of method selection in genomic epidemiology and caution against overinterpreting clonal structure as a proxy for pathogenic potential.

Background Bovine respiratory disease (BRD) is a major cause of morbidity, mortality, impaired welfare, production loss, and antimicrobial use in cattle. These European Network for Optimization of Veterinary Antimicrobial Therapy (ENOVAT) guidelines provide evidence-based recommendations for antimicrobial and non-steroidal anti-inflammatory drug (NSAID) use in cattle with acute, undifferentiated BRD, balancing animal-level outcomes with antimicrobial stewardship. Methods Recommendations were developed using the GRADE approach and Evidence-to-Decision frameworks, following ENOVAT procedures and RIGHT reporting guidance. Evidence came from two prespecified systematic reviews and meta-analyses of randomized controlled trials evaluating individual treatment of clinical BRD: one comparing antimicrobial classes head-to-head and one evaluating NSAIDs as adjunctive treatment or monotherapy. Metaphylaxis and prophylaxis were excluded. Certainty of evidence was assessed with GRADE. Panel judgments incorporated stakeholder-derived thresholds for clinically meaningful differences in short-term therapeutic failure, defined as need for antimicrobial re-treatment, and a merged stewardship perspective based on WHO antimicrobial importance and EMA AMEG categories. Results Across antimicrobial comparisons, no class showed a clinically meaningful advantage in reducing short-term re-treatment sufficient to override stewardship considerations. The panel conditionally suggests penicillin G, oxytetracycline, or florfenicol as first-line empirical treatment, guided by local susceptibility data, herd treatment history, product authorization, withdrawal periods, and suspicion of Mycoplasmopsis bovis infection. Macrolides are conditionally suggested as second-line treatment, preferably guided by culture and susceptibility testing or documented treatment failure. Fluoroquinolones are recommended against as primary or secondary treatment, except in exceptional, justified circumstances consistent with regulations. Adding NSAIDs to antimicrobials did not meaningfully reduce antimicrobial re-treatment; the panel recommends against NSAID use for this specific purpose. Evidence was insufficient to recommend for or against NSAIDs for welfare-related outcomes or as antimicrobial substitutes. Conclusions These guidelines prioritize effective BRD treatment while reducing unnecessary use of higher-priority antimicrobials. Local adoption or adaptation using GRADE-ADOLOPMENT is recommended.

RT-qPCR can be employed to quantify target gene expression normalized to reference genes that are expected to remain stable across experimental conditions. However, previously reported reference genes may not be stably expressed across all experimental conditions and individuals. This research identified novel candidate reference genes from publicly available healthy bovine tissue transcriptomes and subsequently developed and validated primers for five candidate reference genes. Forty-one transcriptomes were obtained from the NCBI Gene Expression Omnibus (GEO) from apparently healthy Bos taurus samples: bone marrow, bronchial lymph nodes (LN), mesenteric LN, kidney, liver, lung, nasal epithelium, unspecified LN, spleen, thymus, and trachea. Bioinformatic analysis of each dataset was performed using FastQC, Trimmomatic, STAR, and R, with library normalization carried out using the Relative Log Expression (RLE) method and normalized gene counts converted to log2-CPM values. The values for each gene across the transcriptomes were ranked by coefficient of variation (CV). The top 200 genes ranked by CV were analyzed using RefFinder for inter-tissue stability. Five candidates were selected: DDX23, NMT1, CRNKL1, TMEM183A, and UBE2Q1. To validate stability, RT-qPCR was used to assess the stability of these genes in tissues from seven bovine respiratory syncytial virus-infected calves and two healthy calves, including lung, bronchus, trachea, nasal epithelium, tracheobronchial LN, mediastinal LN, thymus, spleen, and bone marrow. The averaged CT values were analyzed with RefFinder. Comprehensive stability values identified CRNKL1 as the most or second most stable gene in 8/10 tissues, with variation in ranking of each reference gene across all tissues. This study identified a bioinformatic workflow and candidate reference genes, with primers validated for five genes which demonstrated stability across various bovine tissues.

Background Non-steroidal anti-inflammatory drugs (NSAIDs) are widely used as ancillary therapy for bovine respiratory disease (BRD) alongside antimicrobials, and interest in NSAID monotherapy has grown amid antimicrobial stewardship and welfare concerns. We evaluated the clinical effectiveness of NSAIDs for BRD as adjunctive or sole therapy. Methods We conducted a systematic review and meta-analysis of randomized controlled trials (RCTs) in cattle with naturally occurring BRD. Databases searched were MEDLINE (Ovid), Embase (Ovid), CAB Abstracts (Ovid), Biological Abstracts (Ovid), Web of Science Core Collection, and Scopus (search: 22 Apr 2024; update: 12 Sep 2025). Primary outcomes were short-term treatment failure (need for re-treatment); secondary outcomes included relapse, mortality, adverse events, and performance where available. Random-effects pairwise meta-analyses estimated risk ratios (RRs) with 95% CIs. Risk of bias was assessed with a modified RoB 2.0 tool; certainty of evidence was appraised using GRADE. Registration/protocol: this review extends a prior ENOVAT BRD protocol; no separate registration was created. Funding: COST Action CA18217. Results Seventeen RCTs (22 comparisons; 4,909 animals) compared NSAID+antimicrobial versus antimicrobial alone. Adding an NSAID did not reduce re-treatment (RR 0.94, 95% CI 0.84–1.05; I 2 = 0%; moderate certainty). Subgroup (same vs different antimicrobial) and sensitivity analyses (handling of multi-arm trials; follow-up restricted to ≤14 or ≤ 10 days; risk-of-bias restrictions) did not change conclusions. Two RCTs in UK dairy calves compared NSAID monotherapy with antimicrobial monotherapy (RR 1.19, 95% CI 0.72–1.97; very low certainty). Re-treatment is an indirect outcome and may not capture analgesia, fever reduction, or growth effects. Conclusions Across RCTs, NSAID use did not confer a clinically relevant reduction in re-treatment when added to antimicrobials, and evidence is very uncertain for NSAID monotherapy versus antimicrobials. Future trials should include validated pain/welfare measures, clinical and ultrasonographic outcomes, and performance metrics, with pathogen identification to explore effect modification.

Bovine respiratory disease (BRD) is a leading cause of morbidity and antimicrobial use in calves. Laboratory diagnostics are limited by difficulty obtaining lower airway samples from live animals and the common presence of many BRD-associated bacteria in healthy calves, complicating culture interpretation. This study aimed to investigate the occurrence, clinical relevance, and antimicrobial resistance patterns of BRD-associated bacteria in calves from Norwegian fattening herds with enzootic pneumonia, and to evaluate agreement between upper and lower airway sampling sites. In a cross-sectional study, 88 calves from seven fattening herds were clinically scored and classified as healthy or diseased. Nasal swabs, nasopharyngeal swabs, bronchoalveolar lavage (BAL) fluid, and serum samples were collected for bacterial culture, antimicrobial susceptibility testing by disc diffusion, and serology. Pasteurella multocida and Mannheimia haemolytica were the most frequently detected bacteria across sampling sites, while Histophilus somni was less common. In BAL samples, P. multocida was the predominant bacterium identified and was significantly associated with clinical disease. Agreement between upper airway samples and BAL for Pasteurellaceae detection was slight to fair at the individual level. However, group-level isolation rates for P. multocida were similar across sampling sites in diseased calves. Most isolates were susceptible to common BRD antimicrobials, although penicillin-resistant M. haemolytica was detected in 18% of M. haemolytica-positive calves. Antibodies against Mycoplasmopsis bovis were not detected. In conclusion, P. multocida appears to be the bacterial pathogen most strongly associated with clinical disease in calves from Norwegian fattening herds with enzootic pneumonia. Upper airway sampling had limited value for individual diagnosis but may be useful for herd-level assessment when diseased calves are sampled. Antimicrobial resistance levels were generally low, although resistance was detected.

High-resolution genomic characterization is essential for understanding diversity, pathogenicity, and transmission dynamics of bacterial pathogens. Mannheimia haemolytica (Mh) is the most consequential bacterial agent associated with bovine respiratory disease (BRD) in cattle, as a leading cause of morbidity, mortality, and antimicrobial use. Historically, BRD pathogens, including Mh, have been studied using culture or PCR approaches that provided limited ability to characterize fine-scale genomic variation across communities. Here, we evaluated target-enriched (TE) shotgun sequencing, a culture-independent method capable of strain-level resolution within metagenomic data, for detecting and characterizing Mh in comparison with qPCR and 16S rRNA gene sequencing. Nasal swabs (10 individual and 2 composited DNA samples per pen) and environmental samples (three ropes hung on pen rails and three water bowl swabs per pen) were collected from four pens in each of five distinct cattle populations. DNA was extracted for TE sequencing to identify Mh at both species and genomic sequence variant (GSV) levels, and to characterize antimicrobial resistance genes across the bacterial communities. qPCR was performed to quantify Mh genome copies, and 16S rRNA gene sequencing was used to assess the broader respiratory microbiome. TE sequencing identified Mh in 100% of TE-tested samples and classified multiple GSVs in all but 3 of 121 samples. GSV profiles clustered within housing groups and varied across cattle populations, indicating structured strain-level diversity. In contrast, Mannheimia spp. were detected in only 47.7% of samples by 16S rRNA sequencing. These findings demonstrate that TE sequencing enables sensitive, strain-level characterization of Mh in cattle and environmental samples and reveals substantial within-population genomic diversity not captured by conventional approaches.IMPORTANCETarget-enriched shotgun sequencing enabled sensitive, strain-level detection of Mannheimia haemolytica (Mh), revealing multiple co-circulating genomic sequence variants (GSVs) within and among cattle groups. This demonstrates greater genetic variability of Mh populations in beef cattle than has been previously recognized. The clustering of GSVs within housing groups, together with the overlap between respiratory and environmental samples, is consistent with the hypothesis that contagious transmission contributes to Mh ecology. These results highlight the potential utility of composite nasal swab and environmental samples for future studies evaluating relationships between Mh genomic variation and disease risk.

Background/Objectives: Bovine respiratory disease (BRD) remains a major concern in cattle research, and the long-term effects of vaccination on health and immune responses are not well defined. This study compared gene expression in vaccinated (VAX) and unvaccinated (NOVAX) preweaned calves and subsequent BRD development during backgrounding. Methods: Whole blood was collected at four timepoints (TIME; T1-4; median age 107, 114, 183, and 230, respectively) from 73 bull calves enrolled in a blinded randomized controlled trial; VAX calves received a commercial attenuated multivalent viral vaccine at T1 and T3. Results: Whole-blood transcriptomics was used to quantify mRNA, identifying 5364 differentially expressed genes (DEGs) for TIME, 84 DEGs for vaccination (VAX), and 129 for BRD status using both glmmSeq and QLF testing (glmmSeq only DEGs: 11,068 TIME, 358 VAX, and 9241 BRD). VAX calves at T3 were clustered uniquely with the enrichment of pathways related to the cellular response to stress, neutrophil degranulation, and antigen processing and presentation compared to NOVAX cattle and VAX at other timepoints. Interferon pathways, natural killer cell responses, and neutrophil activity were generally absent across all timepoints, while antigen presentation pathways were persistently enriched. Regardless of vaccination or future BRD diagnosis, immunological development over time was indicated by DEGs related to adaptive immunity, lymphocyte development, and inflammatory resolution. At T4, cattle diagnosed with BRD during backgrounding had differential gene expression related to oxygen transport, hemoglobin function, and metabolic processes compared to cattle that remained healthy. Conclusions: This study provides insights into the possible genomic mechanisms underlying vaccine responses and preclinical BRD susceptibility in preweaned beef cattle.

Bovine respiratory disease (BRD) is a leading cause of illness and death in cattle and a major driver of antimicrobial use. This systematic review and meta-analysis assessed the clinical effectiveness of five commonly used antimicrobial classes (penicillins, tetracyclines, phenicols, macrolides, and fluoroquinolones) in the treatment of cattle with natural BRD. Cattle from all production systems (feedlot, dairy, slaughter, and stocker) were included, and metaphylaxis trials were excluded. Peer-reviewed trials were searched in several databases, and all reports were screened in duplicate for data extraction and risk of bias. The Grading of Recommendations Assessment, Development, and Evaluation (GRADE) methodology was employed to evaluate the certainty of evidence. Twenty-eight randomized controlled trials were included. We did not detect clinically meaningful differences between antimicrobial classes in reducing the need for re-treatment (a surrogate for short-term therapeutic failure). Certainty of evidence ranged from very low to moderate, and results should be interpreted cautiously. Subgroup estimates were broadly consistent for production systems (feedlot and non-feedlot) and for trials with Mycoplasma -positive animals; however, these subgroup analyses were small and imprecise and do not allow firm conclusions. Mortality, relapse, and growth outcomes were measured inconsistently and could not be reliably pooled. Overall, we did not identify any class with a clinically meaningful short-term advantage based on our prespecified thresholds. The results of this systematic review and meta-analysis will inform the BRD Antimicrobial Use Guidelines of the European Network for Optimization of Antimicrobial Therapy (ENOVAT).

Abstract Background Enzootic pneumonia in calves is associated with pathogens such as Pasteurellaceae bacteria and Mycoplasmopsis bovis . Some of these bacteria are commensals in healthy calves, complicating culture interpretation, discouraging laboratory diagnostic use, and limiting knowledge of their impact. This study aimed to investigate the occurrence and antimicrobial resistance pattern of respiratory pathogens in calves from dairy herds with enzootic pneumonia. Culture results between upper and lower airway sites in healthy and diseased calves were also compared to evaluate their diagnostic value. A cross-sectional study was conducted on 131 calves (72 healthy and 59 diseased) from nine Norwegian dairy herds. Nasal swabs (NS), nasopharyngeal swabs (NPS), and serum were obtained from each calf, and bronchoalveolar lavage (BAL) from 113 calves. Results At the calf level, Pasteurella multocida was present in 60%, Mannheimia haemolytica in 55%, and Histophilus somni in 3%. At the sampling site level, from NS, NPS, and BAL, P. multocida was detected in 47%, 45%, and 27%, and M. haemolytica in 46%, 37%, and 8%, respectively. H. somni was detected in ≤ 2% per site. P. multocida appeared as pure culture in 73% (22/30) of positive BAL cultures. Serum antibodies to M. bovis were not detected. Most Pasteurellaceae isolates were susceptible to penicillin and other common pneumonia antimicrobials (disc diffusion). The exceptions were penicillin-resistant M. haemolytica isolates in two calves. Logistic regression identified an association between dominant, abundant cultures of P. multocida from BAL and clinical disease. Agreement for P. multocida detection between NS and NPS cultures and BAL was fair (kappa). Positive predictive values of P. multocida culture results at different abundance levels in NS and NPS, relative to BAL, were at most 49%. Conclusion P. multocida was the predominant bacterium in lungs of calves from Norwegian dairy herds with enzootic pneumonia, and its abundance in the lungs may help differentiate infection from colonization in clinically diseased calves. M. haemolytica and H. somni appeared to have less clinical impact. Antimicrobial resistance appeared to be low. Upper airway cultures were inaccurate predictors of bacterial presence and abundance in the lungs and must be interpreted carefully alongside other diagnostic tools.

Bovine respiratory disease (BRD) remains the leading cause of morbidity and economic loss in North American cattle production. Cattle at high risk of BRD are often managed through antimicrobial metaphylaxis, although there are growing concerns regarding the use of antimicrobial drugs in food animals. To better understand host immunological responses to BRD and the effects of metaphylaxis, this study evaluated whole-blood gene expression patterns in high-risk stocker heifers using RNA sequencing (RNA-Seq). Eighty-four commercial heifers were randomly assigned to receive metaphylaxis with tulathromycin (META) or no metaphylaxis (NO META) upon arrival. Cattle were monitored over a 70-day period for clinical BRD, with whole blood collected from a subset of 60 heifers across multiple timepoints in both cattle that developed (TREAT) or failed to develop BRD (HEALTHY) for RNA-Seq. Principal component and differential gene expression analyses (FDR ≤ 0.05) were conducted to explore transcriptomic differences associated with BRD diagnosis over time, metaphylactic treatment, and disease severity. Minimal differences in the transcriptome were observed between META and NO META cattle at the time of BRD diagnosis, while comparisons between HEALTHY and TREAT cattle at onset of clinical disease revealed greater than 2000 differentially expressed genes (DEGs), with significant functional enrichment of immune-related pathways, including cytokine signaling, lymphocyte activation, and inflammatory responses. Notably, five genes (IL1R2, HP, S100A9, TLR4, and ALOX15) were consistently up- or downregulated in BRD cases, regardless of study group allocation. These observations align with previous studies, which support their candidacy as useful biomarkers for BRD detection. Despite the significant changes observed at the onset of clinical BRD, no differences were identified among these animals at the start of the study. This inability to predict future disease development or severity may have been affected by the low morbidity rate observed in this cohort. Expression of genes coding for heat shock proteins at subsequent treatments suggested that these may be useful markers of disease persistence or severity. These findings suggest that while metaphylaxis reduces BRD incidence, host transcriptomic profiles at disease onset offer improved insight into BRD pathophysiology.

Acute interstitial pneumonia (AIP) has been a recognized bovine respiratory disease for many decades, yet the sporadic occurrences of AIP in feedlot cattle are poorly understood. We searched 3 databases (PubMed, CABI Direct Library, AGRICOLA) for primary literature on North American feedlot cattle, with a focus on interstitial pneumonia. We comprehensively reviewed reports of clinical signs, pathology findings, etiologies, risk factors, and treatments associated with AIP in feedlot cattle and identified gaps in knowledge. We included 19 peer-reviewed studies from 1976 to 2023. In 6 articles, the most common clinical signs for diagnosing AIP were severe dyspnea, increased expiratory effort, and grunting while breathing. In the 17 articles that provided detailed descriptions of the pathology and diagnostic features of AIP, consistent postmortem examination findings were interlobular edema and emphysema, a checkerboard appearance, and diffusely overinflated lung lobes. The most frequent histopathology findings were hyaline membranes, type II pneumocyte hyperplasia, and obliterative bronchiolitis. Although some potential etiologies, such as bovine respiratory syncytial virus (BRSV) and 3-methylindole (3MI), have been evaluated, no definitive cause has been identified consistently. Risk factors significantly associated with AIP in multiple studies included heifers, clinical onset at >45 days on feed (DOF), death at >78 DOF, and summer; the exact mechanisms remain unclear. The absence of standardized diagnostic criteria, along with a lack of research into potential effective prevention or treatment, underscore the need for further study to clarify the causes, diagnosis, and treatment of AIP in feedlot cattle.

Characterizing microbial genetic sequences and key variants is critical for understanding pathogen ecology, transmission, and clinical impact. Yet, conventional metagenomic sequencing often yields too few on-target reads to move beyond species-level identification. We developed a target-enriched (TE) metagenomic workflow, including bait design, an optimized TE shotgun protocol, and the VARIANT++ pipeline, to recover and classify reads at a clustered genomic sequence-variant (GSV) level (see Graphical abstract). The computational component clusters reference genomes by average nucleotide identity, builds a GSV database, and integrates Kraken2, Themisto, and mSWEEP to increase call confidence while reducing false positives. Using Mannheimia haemolytica ( Mh ), the primary cause of bovine respiratory disease, we designed 114,375 DNA baits targeting sequences across 70 reference genomes. TE libraries from nasopharyngeal swabs of feedlot cattle achieved >250-fold increases in on-target Mh reads (∼2.5% of non-host reads on average) compared with conventional shotgun sequencing, despite using one-quarter the sequencing depth. This variant-level resolution revealed six GSVs; most samples contained at least two, indicating variant mixtures difficult to detect with culture- or shotgun-based surveys. Because the approach leverages available reference sequences, it can be reconfigured for other microbial targets. TE metagenomics paired with genome-similarity clustering provides a scalable approach to variant-level characterization from complex microbial populations. ![Graphical abstract][1] Graphical abstract Overview of the components in our three-part workflow. ### Competing Interest Statement The authors have declared no competing interest. All sequence reads are available through BioProject PRJNA1309097 at the NCBI’s Sequence Read Archive. The code and instructions for the bioinformatic and statistical analyses can be found at the GitHub repository ([https://github.com/Microbial-Ecology-Group/Manuscript-Mh\_TE\_validation][2]) and by the corresponding DOI: 10.5281/zenodo.16989755. Texas A&M University [1]: pending:yes [2]: https://github.com/Microbial-Ecology-Group/Manuscript-Mh_TE_validation

Inhalation of Rhodococcus equi causes severe pneumonia in humans and animals worldwide, most commonly affecting horse foals. The standard for preventing R. equi pneumonia in foals is transfusion of hyperimmune plasma, which is expensive and carries the risk of adverse effects. Our goal was to passively immunize foals against R. equi by nebulizing mRNA encoding an equine monoclonal antibody (mAb) against the virulence-associated protein A (VapA) directly into the lungs. VapA-specific memory B cells from an immunized horse were used to identify and select the sequence for an equine immunoglobulin (Ig)G1 mAb. In vitro-transcribed mRNA encoding this sequence expressed full-length, VapA-specific mAbs in vitro and safely and effectively produced intrapulmonary mAb in foals for at least 5 days following nebulization. These findings establish a platform to generate mRNA-encoded mAbs for immunotherapeutic and immunoprophylactic applications in horses and demonstrate the feasibility of delivering nebulized mRNA-mAb for intrapulmonary mAb expression in neonates.

Bovine trichomonosis is caused by the urogenital parasite Tritrichomonas foetus (T. foetus). In the United States, approved therapies are lacking, and management is limited to culling infected bulls. Preputial therapy with synthetic mRNA could lead to effective new treatments. We developed synthetic mRNA encoding bovine IgG1 against two epitopes of the T. foetus cell surface antigen TF1.17 and used the mRNA to transfect bovine cells in vitro. Transfected cells expressed membrane anchored or secreted versions of the antibodies with a NanoLuciferase (NanoLuc) reporter molecule fused to each light chain. Luminescence in cells and supernatants collected 24 and 48 h post-transfection confirmed the production of anti-TF1.17 and was significantly higher than in non-transfected controls (p < 0.05). Anti-TF1.17 bound to live parasites as indicated by significantly higher luminescence following treatment with 24 and 48 h post-transfection supernatants compared to transfection controls (p = 0.001). Treatment of T. foetus with concentrated anti-TF1.17 antibody decreased parasite viability. When T. foetus were added to mRNA transfected kidney cells 48 h post transfection, cytopathic effects of the parasites were reduced following 24 h of co-culture with cells producing anti-TF1.17 as compared to controls (p < 0.05). To our knowledge, this is the first use of mRNA transfection of bovine cells to induce the expression of antibodies that can bind to T. foetus, decrease their viability and their cytopathic effects on host cells. This work forms the basis for the development of novel mRNA-mediated approaches to treat or prevent bovine trichomonosis.

For years veterinarians have understood that stressful experi­ences increase disease in lightweight, recently transported and commingled cattle. Increased incidence of bovine respiratory disease (BRD) in high-risk cattle has traditionally been attrib­uted to endogenous cortisol, but in fact, increased cortisol is often not measured in cattle presumed to be stressed. While high-risk cattle are often seronegative to common respiratory viruses at receiving, field trials have confirmed they can mount significant humoral response to vaccination at arrival, indicat­ing they are not too immunosuppressed to produce antibodies. However, at-arrival vaccination of high-risk cattle is not reli­ably associated with improved health over the subsequent 60-90 days. Research indicates that stress actually increases the mag­nitude of inflammatory responses to infection or other stimuli. Taken together, these findings indicate that high BRD incidence in some high-risk cattle may be more attributable to excessive or prolonged inflammation than to immunosuppression. Re­cent studies demonstrated that high-risk cattle that nonetheless stay healthy have increased expression of genes related to pro­duction of specific pro-resolving mediators (SPM), which bring inflammatory responses to a close. Research is ongoing to de­termine whether SPM can be induced by certain management practices, genetic selection, or therapeutic or prophylactic in­terventions, to improve cattle health.

Bovine respiratory disease (BRD) remains a priority topic with­in livestock research. A leading topic of BRD-related research is vaccination of pre-weaned calves to mitigate the effects of BRD post-weaning. Comparison of gene expression pathways in vaccinated and unvaccinated calves that develop or resist BRD would help clarify factors that may result in benefits following vaccination.

Bovine respiratory disease (BRD) is a multifactorial disease complex resulting from the interaction of various pathologic, immune-mediated and management-driven factors. Specifi­cally, management decisions around vaccination against viral components of BRD, marketing through commercial sale sys­tems, and long-haul shipping of cattle modulate the bovine im­mune system and susceptibility to disease. Therefore, we inves­tigated these factors and their influence on immunomodulation via RNA sequencing methodology.

Global warming exacerbates heat stress in dairy cattle, impacting their productivity and welfare due to high metabolic heat production. Heat stress causes reduced feed intake, panting, and prolonged standing-like behavior in dairy cows, which are key welfare indicators of heat stress. While various mitigation strategies exist, their limitations highlight the urgent need for more efficient, cost-effective approaches to enhance animal welfare in response to thermal stress. This pilot study aimed to assess whether the topical application of menthol, a natural, FDA-approved-safe cooling agent obtained from the Mentha piperita plant, and a specific agonist of cold receptor Transient Receptor Potential Melastatin 8 (TRPM8), around the auricular vagal region of cattle’s ear could alleviate heat stress by stimulating the vagus nerve. Twelve lactating Holsteins were divided into three groups: negative control (NC, n=3), plain wax (PW, n=3), and menthol wax (MW, n=6) and kept in the heat stress condition for 14 days, with limited access to sprinklers. The PW and MW groups were topically applied with menthol wax (100mM) and olive-oil-based plain wax during the morning (5:30 AM) and afternoon (5:30 PM) feedings on both ears from day 0 to 14, respectively. The stress behaviors recorded included eating, lying, and standing, monitored over 14 hours on day 0, day 6th of the trial, and day 13th of the trial. Observations were conducted from 8:00 AM to 3:00 PM and from 8:00 PM to 3:00 AM following wax application to assess behavioral responses under stress conditions. Visual live observations on menthol sensing behavior were observed just after the menthol and plain wax application on the MW and PW group animals. Behavior phenotypes during the 14-hour interval were monitored per animal per group in recorded videos, and differences between groups were assessed using Linear Mixed Model (LMM). Interestingly, menthol application in the MW group caused ear drooping immediately, lasting up to 30 minutes. This response, absent in the PW group, suggests that cows perceived menthol stimulation. The total time spent eating reduced in the NC on day 13th, i.e., 54 min cow-1 day-1 compared to MW and PW group i.e. 93 min cow-1 day-1 and 72.3 min cow-1 day-1 respectively, although not significant (P = 0.59). Based on estimated feed intake, the MW group exhibited significantly higher daily feed intake compared to the combined NC and PW groups (51.64 ± 5.81 kg/cow vs 46.03 ± 7.49 kg/cow, P=0.03). The total time spent standing and lying showed no significant differences (P > 0.05) between the three groups during the trial. These findings suggest that topical menthol application has the potential to alleviate heat stress in dairy cows, but further blood and physiological parameter analysis is needed to explore its broader effects.

Although bovine respiratory disease (BRD) is frequent and cost­ly in beef cattle operations, diagnosis of the disease complex is inaccurate. Metaphylactic use of antimicrobials effectively re­duces the risk of BRD, but antimicrobial administration may in­duce resistance. Therefore, this project was designed to identify differences in levels of transcribed gene expression driven by clinical BRD, both with and without administration of metaphy­laxis at-arrival in high-risk stocker cattle. Using RNA sequenc­ing, we aimed to identify predictive gene expression pathways at-arrival in cattle that developed BRD or not, characterize host genomic patterns in response to clinical BRD, and describe dif­ferences in gene expression of clinically diseased cattle based on treatment success and time of treatment within the study.