An overview of the detection of bovine respiratory disease complex pathogens using immunohistochemistry: emerging trends and opportunities.

The bovine respiratory disease complex (BRDC) is caused by a variety of pathogens, as well as contributing environmental and host-related risk factors. BRDC is the costliest disease for feedlot cattle globally. Immunohistochemistry (IHC) is a valuable tool for enhancing our understanding of BRDC given its specificity, sensitivity, cost-effectiveness, and capacity to provide information on antigen localization and immune response. Emerging trends in IHC include the use of multiplex IHC for the detection of coinfections, the use of digital imaging and automation, improved detection systems using enhanced fluorescent dyes, and the integration of IHC with spatial transcriptomics. Overall, identifying biomarkers for early detection, utilizing high-throughput IHC for large-scale studies, developing standardized protocols and reagents, and integrating IHC with other technologies are some of the opportunities to enhance the accuracy and applicability of IHC. We summarize here the various techniques and protocols used in IHC and highlight their current and potential role in BRDC research.

Comparative Analysis of the Prevalence of Bovine Viral Diarrhea Virus in Cattle Populations Based on Detection Methods: A Systematic Review and Meta-Analysis.

Infectious diseases of cattle, including bovine viral diarrhea (BVD), pose a significant health threat to the global livestock industry. This study aimed to investigate the prevalence and risk factors associated with bovine viral diarrhea virus (BVDV) infections in cattle populations through a systematic review and meta-analysis. PubMed, Web of Science, and Scopus were systematically searched for relevant articles reporting the prevalence of and associated risk factors in studies published between 1 January 2000 and 3 February 2023. From a total of 5111 studies screened, 318 studies were included in the final analysis. BVDV prevalence in cattle populations was estimated using various detection methods. The analysis detected heterogeneity in prevalence, attributed to detection techniques and associated risk factors. Antibody detection methods exhibited a higher prevalence of 0.43, reflecting the cumulative effect of detecting both active and past infections. Antigen detection methods showed a prevalence of 0.05, which was lower than antibody methods. A prevalence of 0.08 was observed using nucleic acid detection methods. The health status of the examined cattle significantly influenced the prevalence of BVDV. Cattle with bovine respiratory disease complex (BRDC) exhibited higher antibody (prevalence of 0.67) and antigen (prevalence 0.23) levels compared to cattle with reproductive problems (prevalence 0.13) or diarrhea (prevalence 0.01). Nucleic acid detection methods demonstrated consistent rates across different health conditions. Age of cattle influenced prevalence, with higher rates in adults compared to calves. Risk factors related to breeding and reproduction, such as natural or extensive breeding and a history of abortion, were associated with increased prevalence. Coinfections with pathogens like bovine herpesvirus-1 or Neospora caninum were linked to higher BVDV prevalence. Management practices, such as commingling, introducing new cattle, and direct contact with neighboring farms, also influenced prevalence. Herd attributes, including larger herd size, and the presence of persistently infected cattle, were associated with higher prevalence. These findings indicated the importance of detection methods and risk factors in BVDV epidemiological studies.

Correction: Messele et al. Longitudinal Analysis of Antimicrobial Resistance among Enterococcus Species Isolated from Australian Beef Cattle Faeces at Feedlot Entry and Exit. Animals 2022, 12, 2690.

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Phylogeny, Virulence, and Antimicrobial Resistance Gene Profiles of Enterococcus faecium Isolated from Australian Feedlot Cattle and Their Significance to Public and Environmental Health.

The extent of similarity between E. faecium strains found in healthy feedlot beef cattle and those causing extraintestinal infections in humans is not yet fully understood. This study used whole-genome sequencing to analyse the antimicrobial resistance profile of E. faecium isolated from beef cattle (n = 59) at a single feedlot and compared them to previously reported Australian isolates obtained from pig (n = 60) and meat chicken caecal samples (n = 8), as well as human sepsis cases (n = 302). The E. faecium isolated from beef cattle and other food animal sources neither carried vanA/vanB responsible for vancomycin nor possessed gyrA/parC and liaR/liaS gene mutations associated with high-level fluoroquinolone and daptomycin resistance, respectively. A small proportion (7.6%) of human isolates clustered with beef cattle and pig isolates, including a few isolates belonging to the same sequence types ST22 (one beef cattle, one pig, and two human isolates), ST32 (eight beef cattle and one human isolate), and ST327 (two beef cattle and one human isolate), suggesting common origins. This provides further evidence that these clonal lineages may have broader host range but are unrelated to the typical hospital-adapted human strains belonging to clonal complex 17, significant proportions of which contain vanA/vanB and liaR/liaS. Additionally, none of the human isolates belonging to these STs contained resistance genes to WHO critically important antimicrobials. The results confirm that most E. faecium isolated from beef cattle in this study do not pose a significant risk for resistance to critically important antimicrobials and are not associated with current human septic infections.

Meta-Analysis on the Global Prevalence of Tetracycline Resistance in Escherichia coli Isolated from Beef Cattle.

Antimicrobial resistance (AMR) is an emerging global concern, with the widespread use of antimicrobials in One Health contributing significantly to this phenomenon. Among various antimicrobials, tetracyclines are extensively used in the beef cattle industry, potentially contributing to the development of resistance in bacterial populations. This meta-analysis aimed to examine the association between tetracycline use in beef cattle and the development of tetracycline resistance in Escherichia coli isolates. A comprehensive search was conducted using multiple databases to gather relevant observational studies evaluating tetracycline use and tetracycline resistance in Escherichia coli isolates from beef cattle. The rate of tetracycline resistance from each study served as the effect measure and was pooled using a random-effects model, considering possible disparities among studies. The meta-analysis of 14 prospective longitudinal studies resulted in a 0.31 prevalence of tetracycline resistance in Escherichia coli in non-intervention (no exposure), contrasting numerically elevated resistance rates in the intervention (exposed) groups of 0.53 and 0.39 in those receiving tetracyclines via feed or systemically, respectively. Despite the observed numerical differences, no statistically significant differences existed between intervention and non-intervention groups, challenging the conventional belief that antimicrobial use in livestock inherently leads to increased AMR. The findings of this study underscore the need for additional research to fully understand the complex relationship between antimicrobial use and AMR development. A considerable degree of heterogeneity across studies, potentially driven by variations in study design and diverse presentation of results, indicates the intricate and complex nature of AMR development. Further research with standardized methodologies might help elucidate the relationship between tetracycline use and resistance in Escherichia coli isolated from beef cattle.

Phylogenetic Analysis of Escherichia coli Isolated from Australian Feedlot Cattle in Comparison to Pig Faecal and Poultry/Human Extraintestinal Isolates.

The similarity of commensal Escherichia coli isolated from healthy cattle to antimicrobial-resistant bacteria causing extraintestinal infections in humans is not fully understood. In this study, we used a bioinformatics approach based on whole genome sequencing data to determine the genetic characteristics and phylogenetic relationships among faecal Escherichia coli isolates from beef cattle (n = 37) from a single feedlot in comparison to previously analysed pig faecal (n = 45), poultry extraintestinal (n = 19), and human extraintestinal E. coli isolates (n = 40) from three previous Australian studies. Most beef cattle and pig isolates belonged to E. coli phylogroups A and B1, whereas most avian and human isolates belonged to B2 and D, although a single human extraintestinal isolate belonged to phylogenetic group A and sequence type (ST) 10. The most common E. coli sequence types (STs) included ST10 for beef cattle, ST361 for pig, ST117 for poultry, and ST73 for human isolates. Extended-spectrum and AmpC ß-lactamase genes were identified in seven out of thirty-seven (18.9%) beef cattle isolates. The most common plasmid replicons identified were IncFIB (AP001918), followed by IncFII, Col156, and IncX1. The results confirm that feedlot cattle isolates examined in this study represent a reduced risk to human and environmental health with regard to being a source of antimicrobial-resistant E. coli of clinical importance.

Longitudinal Analysis of Antimicrobial Resistance among Enterococcus Species Isolated from Australian Beef Cattle Faeces at Feedlot Entry and Exit.

Enterococcus faecium are commensal bacteria inhabiting the gastrointestinal tract of animals and humans and an important cause of drug-resistant nosocomial infections. This longitudinal study aimed to determine whether changes in the antimicrobial resistance (AMR) phenotype and genotype occurred among Enterococcus spp. isolated from cattle rectal samples obtained at the entry to and exit from an Australian feedlot. The samples obtained at the feedlot induction yielded enterococci (104/150; 69.3%), speciated as E. hirae (90/104; 86.5%), E. faecium (9/104; 8.7%), E. mundtii (3/104; 2.9%), E. durans, and E. casseliflavus (1/104; 1.0% each). AMR was observed to lincomycin (63/104; 60.6%), daptomycin (26/104; 25.0%), nitrofurantoin (9/104; 8.7%), ciprofloxacin (7/104; 6.7%), tetracycline (5/104; 4.8%), tigecycline (4/104; 3.9%), and quinupristin/dalfopristin (3/104; 2.9%). From the rectal swab samples collected at the abattoir from the same animals (i.e., the feedlot exit), the enterococci recovery was significantly higher (144/150; 96.0%), with a marked shift in species distribution dominated by E. faecium (117/144; 81.3%). However, the prevalence of AMR to individual antimicrobials remained largely static between the entry and exit except for the increased resistance to nitrofurantoin (77/144; 53.5%) and quinupristin/dalfopristin (26/144; 18.1%). Overall, 13 AMR genes were observed among the 62 E. faecium isolates. These included aac(6')Ii, aac(6')-Iid, and ant(6)-Ia (aminoglycosides); eatAv, lnu(G), vat(E), msr(C), and erm(B) (macrolides, lincosamides, and streptogramins); efmA (fluoroquinolones); and tet(45), tet(L), tet(M), and tet(S) (tetracyclines). The results confirm the presence of fluoroquinolone- and streptogramin-resistant enterococci in cattle faeces at the feedlot entry in the absence of antimicrobial selection pressure. E. faecium, exhibiting increased nitrofurantoin resistance, became the dominant Enterococcus spp. during the feeding period.

Phenotypic and Genotypic Analysis of Antimicrobial Resistance in Escherichia coli Recovered from Feedlot Beef Cattle in Australia.

This study investigated the antimicrobial resistance (AMR) profile of fecal Escherichia coli isolates from beef cattle (n = 150) at entry and exit from an Australian feedlot. Sample plating on MacConkey agar and Brilliance ESBL agar differentiated generic from extended-spectrum ß-lactamase (ESBL)-producing E. coli, respectively. Resistance profiles were determined by minimum inhibitory concentration (MIC) testing and further analyzed by whole-genome sequencing (WGS). At entry, the prevalence of antimicrobial resistance to amoxicillin/clavulanic acid, ampicillin, streptomycin, and trimethoprim/sulfamethoxazole was very low (0.7%, each). At the exit, the resistance prevalence was moderate to tetracycline (17.8%) and low to ampicillin (5.4%), streptomycin (4.7%), and sulfisoxazole (3.9%). The most common AMR genes observed in phenotypically resistant isolates were tet(B) (43.2%), aph(3″)-Ib and aph(6)-Id (32.4%), blaTEM-1B, and sul2 (24.3%, each), which are responsible for resistance to tetracyclines, aminoglycosides, ß-lactams, and sulfonamides, respectively. The ESBL-producing E. coli were recovered from one sample (0.7%) obtained at entry and six samples (4.0%) at the exit. The ESBL-producing E. coli harbored blaTEM (29.7%), blaCTX m(13.5%), and blaCMY (5.4%). The resistance phenotypes were highly correlated with resistance genotypes (r ≥ 0.85: p < 0.05). This study demonstrated that E. coli isolated from feedlot beef cattle can harbour AMR genes, but the low incidence of medically important resistance reflected the prudent antimicrobial use in the Australian industry.

The proteome of human Fallopian tube lavages during the phase of embryo transit reveals candidate proteins for the optimization of preimplantation embryo culture.

STUDY QUESTION: Are there phase-specific changes in the early secretory (ES) phase human tubal lavage proteome that can inform and potentially optimize IVF culture media? SUMMARY ANSWER: The human tubal lavage proteome during the ES phase relative to the menstrual phase reveals substantial differential protein abundance in pathways such as glycolysis, redox homeostasis and activation of 14-3-3 zeta-mediated signaling. WHAT IS KNOWN ALREADY: The Fallopian tube is uniquely suited to the development of the preimplantation embryo as it transits the tube during the ES phase of the menstrual cycle. Euploid cleavage-stage embryo arrest may reflect incomplete recapitulation of in-vivo conditions by current media formulations. STUDY DESIGN, SIZE, DURATION: Proteome-wide analysis of distal tubal lavage specimens collected from 26 healthy women undergoing open microtubal anastomosis surgery from January 2013 to January 2018 was performed. Specimens were grouped by menstrual cycle phase in order to analyze phase-specific differences in protein abundance. For the murine embryo assay, single-cell embryos (N = 482) were collected from superovulated wild type C57BL/6 female mice and cultured in microdrops over 5 days for the assessment of blastocyst development. PARTICIPANTS/MATERIALS, SETTING, METHODS: Human tubal lavage specimens were processed for label-free mass spectrometry. Reported menstrual cycle day was confirmed by measuring serum hormones. Key protein targets in the ES phase were validated via immunoblot. The ES phase-specific increase in 14-3-3 zeta protein was confirmed via ELISA of conditioned media obtained from primary human Fallopian tube epithelial cell culture. A murine embryo assay was performed to investigate the impact of graduated concentrations of 14-3-3 zeta on the blastocyst development rate. MAIN RESULTS AND THE ROLE OF CHANCE: Comparison of the ES and menstrual phase human tubal lavage proteomes revealed 74 differentially expressed proteins with enrichment of pathways and biological processes involved in the regulation of carbohydrate metabolism, oxidative stress and cell survival. The adapter-regulator protein 14-3-3 zeta was among the most significantly increased in the ES phase. Supplementation of embryo culture media with 14-3-3 zeta at concentrations tested did not significantly improve the murine blastocyst development. LIMITATIONS, REASONS FOR CAUTION: Although select associations were recapitulated in the conditioned media from sex steroid exposed primary human tubal epithelial cells, cell culture represents an in-vitro approximation. Changes to embryo culture media, such as protein supplementation, must undergo rigorous preclinical safety testing prior to adoption for human use. WIDER IMPLICATIONS OF THE FINDINGS: This study represents the first description of the human Fallopian tube lavage proteome across the menstrual cycle, revealing a unique proteomic signature during the ES phase. Although supplementation of culture media with 14-3-3 zeta at appropriate concentrations showed no significant impact on the murine blastocyst development rate, other biologically plausible candidate proteins for individual or high throughput testing strategies are identified. STUDY FUNDING/COMPETING INTEREST(S): This work was funded in part by an Army Medical Department Advanced Medical Technology Initiative grant from the United States Army Medical Research and Materiel Command's Telemedicine and Advanced Technology Research Center. There are no competing interests. TRIAL REGISTRATION NUMBER: N/A.

Multiple isotope analyses of the pike tapeworm Triaenophorus nodulosus reveal peculiarities in consumer-diet discrimination patterns.

Previous studies of dietary isotope discrimination have led to the general expectation that a consumer will exhibit enriched stable isotope levels relative to its diet. Parasite-host systems are specific consumer-diet pairs in which the consumer (parasite) feeds exclusively on one dietary source: host tissue. However, the small numbers of studies previously carried out on isotopic discrimination in parasite-host (ΔXP-HT) systems have yielded controversial results, showing some parasites to be isotopically depleted relative to their food source, while others are enriched or in equilibrium with their hosts. Although the mechanism for these deviations from expectations remains to be understood, possible influences of specific feeding niche or selection for only a few nutritional components by the parasite are discussed. ΔXP-HT for multiple isotopes (δ13C, δ15N, δ34S) were measured in the pike tapeworm Triaenophorus nodulosus and two of its life-cycle fish hosts, perch Perca fluviatilis and pike Esox lucius, within which T. nodulosus occupies different feeding locations. Variability in the value of ΔXP-HT calculated for the parasite and its different hosts indicates an influence of feeding location on isotopic discrimination. In perch liver ΔXP-HT was relatively more negative for all three stable isotopes. In pike gut ΔXP-HT was more positive for δ13C, as expected in conventional consumer-diet systems. For parasites feeding on pike gut, however, the δ15N and δ34S isotope values were comparable with those of the host. We discuss potential causes of these deviations from expectations, including the effect of specific parasite feeding niches, and conclude that ΔXP-HT should be critically evaluated for trophic interactions between parasite and host before general patterns are assumed.

Differential expression of fibulin family proteins in the para-cervical weak zone and other areas of human fetal membranes.

OBJECTIVE: Human fetal membranes (FM) at term have been shown to contain a weak zone in the region overlying the cervix which exhibits characteristics of increased collagen remodeling and apoptosis. It has been hypothesized that the FM rupture initiation site is within this weak zone. Although the FM weak zone has been partially characterized, it is unclear what structural differences in the extracellular matrix result in its decreased rupture strength. A screen for differentially expressed proteins in the amnion of the weak zone versus other FM areas demonstrated that fibulin 1 was decreased. We investigated potential regional differences in all fibulin protein family members. METHODS: FM fibulins were localized by immunohistochemistry. Detected fibulins were screened by Western blot for differences in abundance in the amnion of the weak zone versus non-weak zone FM regions. Amnion epithelial and mesenchymal cells were also screened for fibulin production. RESULTS: Fibulins 1 and 5 were detected in the cytoplasm of and in a pericellular pattern surrounding all FM cells, and in a dense extracellular pattern in the amniotic compact zone. Fibulin 3 was detected within the cytoplasm of amnion epithelial and chorion trophoblast cells. Fibulins 2 and 4 were not detected. Fibulins 1, 3 and 5 demonstrated decreased abundance of 33%, 63% and 58% (all P<0.01) in amnion of the weak zone relative to other FM regions. Amnion cells produced all three detected fibulins. Furthermore, TNF inhibited amnion cell fibulin production in a dose dependent manner. CONCLUSION: Fibulins 1, 3 and 5 were localized coincident with major microfibrillar networks in amnion. Each showed decreased abundance in the amnion component of the FM weak zone. Amnion epithelial and mesenchymal cells produced all three fibulins and their abundance was inhibited by TNF. We speculate that the amnion microfibrillar layer undergoes significant remodeling with the development of the FM weak zone.