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Frontiers in Veterinary Science 2022The threat of bovine respiratory disease (BRD) for cattle operations is exacerbated by increasing prevalence of antimicrobial resistance (AMR) in , a leading cause of...
The threat of bovine respiratory disease (BRD) for cattle operations is exacerbated by increasing prevalence of antimicrobial resistance (AMR) in , a leading cause of BRD. Characterization of AMR in by culture and susceptibility testing is complicated by uncertainty regarding the number of colonies that must be selected to accurately characterize AMR phenotypes (antibiograms) and genotypes in a culture. The study objective was to assess phenotypic and genotypic diversity of isolates on nasopharyngeal swabs (NPS) from 28 cattle at risk for BRD or with BRD. NPS were swabbed onto five consecutive blood agar plates; after incubation up to 20 colonies were selected per plate (up to 100 colonies per NPS). Phenotype was determined by measuring minimum inhibitory concentrations (MIC) for 11 antimicrobials and classifying isolates as resistant or not. Genotype was indirectly determined by matrix-assisted laser desorption/ionization time of flight mass spectroscopy (MALDI-TOF MS). NPS from 11 of 28 cattle yielded at least one isolate; median (range) of isolates per NPS was 48 (1-94). NPS from seven cattle yielded one phenotype, 3 NPS yielded two, and 1 NPS yielded three; however, within a sample all phenotypic differences were due to only one MIC dilution. On each NPS all isolated were the same genotype; genotype 1 was isolated from three NPS and genotype two was isolated from eight. Diversity of on bovine NPS was limited, suggesting that selection of few colonies might adequately identify relevant phenotypes and genotypes.
PubMed: 35647109
DOI: 10.3389/fvets.2022.883389 -
Veterinary Medicine (Auckland, N.Z.) 2023Pneumonic pasteurellosis is a respiratory system disease of sheep caused by , and responsible for the low productivity and economic loss resulting from death and...
Isolation and Molecular Detection of and from Clinically Pneumonic Pasteurellosis Cases of Bonga Sheep Breed and Their Antibiotic Susceptibility Tests in Selected Areas of Southwest Ethiopian Peoples Regional State, Ethiopia.
BACKGROUND
Pneumonic pasteurellosis is a respiratory system disease of sheep caused by , and responsible for the low productivity and economic loss resulting from death and treatment costs. This study was conducted to isolate and molecularly detect causative agents and antibiotic susceptibility tests from a nasal swab sample of the Bonga sheep breed that was suspected to have pneumonic pasteurellosis in selected areas of Southwest Ethiopian Peoples Regional State.
METHODS
A cross-sectional study design was used along with purposive sampling of nasal swab samples from sheep that were brought to veterinary clinics during the study period. Bacterial isolation and phenotypic characterization were carried out using microbiological and biochemical tests that followed standard microbiological techniques. To molecularly confirm the isolates, and , species-specific PCR primer genes were used. Using the disc diffusion method, molecularly confirmed isolates were subjected to an in vitro antibiotic susceptibility test.
RESULTS
The 85 samples that were scrutinized had an overall isolation rate of 31.76%, whereas the isolates of and had species compositions of 40.7% and 59.25%, respectively. Overall, 12.5% of the and 18.18% of the species were verified from phenotypical isolates using the species-specific PCR primer genes and , respectively. An in vitro antibiotic susceptibility test was carried out on all four PCR-confirmed isolates for seven commonly used antibiotics used to treat ovine pasteurellosis in the study area. It was found that both bacterial species were resistant to chloramphenicol and penicillin G.
CONCLUSION
Using phenotypic and molecular diagnostic techniques, the results of our current inquiry revealed that and are the causative agents of ovine pneumonic pasteurellosis in the study area.
PubMed: 38162823
DOI: 10.2147/VMRR.S435932 -
BMC Microbiology Aug 2020Mannheimia haemolytica strains isolated from North American cattle have been classified into two genotypes (1 and 2). Although members of both genotypes have been... (Comparative Study)
Comparative Study
BACKGROUND
Mannheimia haemolytica strains isolated from North American cattle have been classified into two genotypes (1 and 2). Although members of both genotypes have been isolated from the upper and lower respiratory tracts of cattle with or without bovine respiratory disease (BRD), genotype 2 strains are much more frequently isolated from diseased lungs than genotype 1 strains. The mechanisms behind the increased association of genotype 2 M. haemolytica with BRD are not fully understood. To address that, and to search for interventions against genotype 2 M. haemolytica, complete, closed chromosome assemblies for 35 genotype 1 and 34 genotype 2 strains were generated and compared. Searches were conducted for the pan genome, core genes shared between the genotypes, and for genes specific to either genotype. Additionally, genes encoding outer membrane proteins (OMPs) specific to genotype 2 M. haemolytica were identified, and the diversity of their protein isoforms was characterized with predominantly unassembled, short-read genomic sequences for up to 1075 additional strains.
RESULTS
The pan genome of the 69 sequenced M. haemolytica strains consisted of 3111 genes, of which 1880 comprised a shared core between the genotypes. A core of 112 and 179 genes or gene variants were specific to genotype 1 and 2, respectively. Seven genes encoding predicted OMPs; a peptidase S6, a ligand-gated channel, an autotransporter outer membrane beta-barrel domain-containing protein (AOMB-BD-CP), a porin, and three different trimeric autotransporter adhesins were specific to genotype 2 as their genotype 1 homologs were either pseudogenes, or not detected. The AOMB-BD-CP gene, however, appeared to be truncated across all examined genotype 2 strains and to likely encode dysfunctional protein. Homologous gene sequences from additional M. haemolytica strains confirmed the specificity of the remaining six genotype 2 OMP genes and revealed they encoded low isoform diversity at the population level.
CONCLUSION
Genotype 2 M. haemolytica possess genes encoding conserved OMPs not found intact in more commensally prone genotype 1 strains. Some of the genotype 2 specific genes identified in this study are likely to have important biological roles in the pathogenicity of genotype 2 M. haemolytica, which is the primary bacterial cause of BRD.
Topics: Animals; Bacterial Outer Membrane Proteins; Cattle; Cattle Diseases; Chromosomes, Bacterial; Genotype; Mannheimia haemolytica; Mutation; Phylogeny; Respiratory Tract Infections; Whole Genome Sequencing
PubMed: 32787780
DOI: 10.1186/s12866-020-01932-2 -
Microorganisms Oct 2022Bovine Respiratory Disease (BRD) is a multifactorial condition affecting cattle worldwide resulting in high rates of morbidity and mortality. The disease can be...
Bovine Respiratory Disease (BRD) is a multifactorial condition affecting cattle worldwide resulting in high rates of morbidity and mortality. The disease can be triggered by Bovine Herpesvirus-1 (BoHV-1) infection, stress, and the subsequent proliferation and lung colonization by commensal bacteria such as , ultimately inducing severe pneumonic inflammation. Due to its polymicrobial nature, the study of BRD microbes requires co-infection models. While several past studies have mostly focused on the effects of co-infection on host gene expression, we focused on the relationship between BRD pathogens during co-infection, specifically on effect on BoHV-1 replication. This study shows that negatively impacts BoHV-1 replication in a dose-dependent manner in different in vitro models. The negative effect was observed at very low bacterial doses while increasing the viral dose counteracted this effect. Viral suppression was also dependent on the time at which each microbe was introduced to the cell culture. While acidification of the culture medium did not grossly affect cell viability, it significantly inhibited viral replication. We conclude that and BoHV-1 interaction is dose and time-sensitive, wherein proliferation induces significant viral suppression when the viral replication program is not fully established.
PubMed: 36363750
DOI: 10.3390/microorganisms10112158 -
Veterinary Research Mar 2020Mannheimia haemolytica serotype A2 is the principal cause of pneumonic mannheimiosis in ovine and caprine livestock; this disease is a consequence of immune suppression...
Mannheimia haemolytica serotype A2 is the principal cause of pneumonic mannheimiosis in ovine and caprine livestock; this disease is a consequence of immune suppression caused by stress and associated viruses and is responsible for significant economic losses in farm production worldwide. Gram-negative bacteria such as M. haemolytica produce outer membrane (OM)-derived spherical structures named outer membrane vesicles (OMVs) that contain leukotoxin and other biologically active virulence factors. In the present study, the relationship between M. haemolytica A2 and bovine lactoferrin (BLf) was studied. BLf is an 80 kDa glycoprotein that possesses bacteriostatic and bactericidal properties and is part of the mammalian innate immune system. Apo-BLf (iron-free) showed a bactericidal effect against M. haemolytica A2, with an observed minimal inhibitory concentration (MIC) of 16 µM. Sublethal doses (2-8 µM) of apo-BLf increased the release of OMVs, which were quantified by flow cytometry. Apo-BLf modified the normal structure of the OM and OMVs, as observed through transmission electron microscopy. Apo-BLf also induced lipopolysaccharide (LPS) release from bacteria, disrupting OM permeability and functionality, as measured by silver staining and SDS and polymyxin B cell permeability assays. Western blot results showed that apo-BLf increased the secretion of leukotoxin in M. haemolytica A2 culture supernatants, possibly through its iron-chelating activity. In contrast, holo-BLf (with iron) did not have this effect, possibly due to differences in the tertiary structure between these proteins. In summary, apo-BLf affected the levels of several M. haemolytica virulence factors and could be evaluated for use in animals as an adjuvant in the treatment of ovine mannheimiosis.
Topics: Animals; Anti-Bacterial Agents; Exotoxins; Lactoferrin; Mannheimia haemolytica; Pasteurellosis, Pneumonic; Sheep; Sheep Diseases
PubMed: 32138772
DOI: 10.1186/s13567-020-00759-z -
Genome Feb 2022Common bacterial causes of bovine respiratory disease (BRD) include , , and . Within , two major genotypes are commonly found in cattle (1 and 2); however, genotype 2...
Common bacterial causes of bovine respiratory disease (BRD) include , , and . Within , two major genotypes are commonly found in cattle (1 and 2); however, genotype 2 strains are isolated from diseased lungs much more frequently than genotype 1 strains. Outer membrane proteins (OMPs) of , , and genotype 2 may be important factors for acquired host immunity. The predicted OMP differences between genotypes 1 and 2 have been previously identified. In this study, we expanded the focus to include bovine-isolated strain genomes representing all three species and the two genotypes. Reported here are the core genomes unique to each of them, core genomes shared between some or all combinations of the three species and two genotypes, and predicted OMPs within these core genomes. The OMPs identified in this study are potential candidates for further studies and the development of interventions against BRD.
Topics: Animals; Bacterial Outer Membrane Proteins; Cattle; Genotype; Mannheimia haemolytica; Pasteurella multocida
PubMed: 34348051
DOI: 10.1139/gen-2021-0038 -
Journal of Veterinary Diagnostic... Mar 2022We developed a rapid insulated isothermal PCR (iiPCR) assay for on-site detection of using a primer and probe set targeting the superoxide dismutase (A) gene. Our iiPCR...
We developed a rapid insulated isothermal PCR (iiPCR) assay for on-site detection of using a primer and probe set targeting the superoxide dismutase (A) gene. Our iiPCR assay detected clinical isolates successfully and produced negative results on other bovine or ovine respiratory pathogens, including , , , , and spp., indicating that the PCR reactions were specific. Additionally, our iiPCR assay detected as few as 21 copies of genomic DNA and 17.2 cfu/mL of bacterial culture, which was 10 and 100 times more sensitive than conventional PCR, respectively. Our iiPCR assay can be performed on a portable device in a total of 58 min and may be a useful tool for the detection of in bovine and ovine respiratory disease in the field.
Topics: Animals; Cattle; Cattle Diseases; Mannheimia haemolytica; Polymerase Chain Reaction; Sheep; Sheep Diseases
PubMed: 35139720
DOI: 10.1177/10406387211068447 -
Veterinary Microbiology Apr 2019Bovine respiratory disease (BRD) is economically significant, and influenza D virus (IDV) is commonly identified in cattle with BRD. Mannheimia haemolytica (MHA) is an...
Bovine respiratory disease (BRD) is economically significant, and influenza D virus (IDV) is commonly identified in cattle with BRD. Mannheimia haemolytica (MHA) is an opportunistic bacterial contributor to BRD; surveillance data suggest that MHA and IDV co-infection occurs in cattle. The objective of this study was to evaluate the synergistic pathogenesis in cattle co-infected with IDV and MHA. Sixteen dairy calves were randomly assigned to four groups of four calves. The IDV + MHA + group received D/bovine/C00046 N/Mississippi/2014 (D/46 N) intranasally at 0 days post-inoculation (DPI) and Mannheimia haemolytica D153 (MHA D153) intratracheally at 5 DPI. The IDV + MHA- group received only D/46 N at 0 DPI; the IDV-MHA + group received only MHA D153 at 5 DPI; and the IDV-MHA- group received neither agent. Clinical scores were calculated twice daily. At 10 DPI, IDV + MHA+, IDV-MHA+, and IDV-MHA- calves were euthanized and evaluated for pathologic lesions. The IDV + groups seroconverted to IDV by 10 DPI. Clinical scores were higher in IDV + groups than IDV- groups on 2-5 DPI (p = 0.001). After MHA challenge on 5 DPI, clinical scores (6-10 DPI) were slightly lower in IDV+MHA+ group than IDV-MHA+ group (p < 0.05) but not significantly different between MHA+ groups and MHA- groups. The average gross pathology score was higher for IDV-MHA+ group than groups IDV-MHA- and IDV+MHA+; however, no significant differences were identified among groups. Under the conditions of this study, infection with IDV before MHA enhance neither clinical disease nor lung pathology, relative to calves infected with MHA alone.
Topics: Animals; Cattle; Cattle Diseases; Coinfection; Lung; Male; Mannheimia haemolytica; Orthomyxoviridae Infections; Pasteurellaceae Infections; Respiratory Tract Infections; Seroconversion; Thogotovirus
PubMed: 30955818
DOI: 10.1016/j.vetmic.2019.03.027 -
Animal Microbiome Aug 2022Bovine respiratory disease (BRD) is caused by interactions among host, environment, and pathogens. One standard method for antemortem pathogen identification in cattle...
BACKGROUND
Bovine respiratory disease (BRD) is caused by interactions among host, environment, and pathogens. One standard method for antemortem pathogen identification in cattle with BRD is deep-guarded nasopharyngeal swabbing, which is challenging, costly, and waste generating. The objective was to compare the ability to recover Mannheimia haemolytica and compare microbial community structure using 29.5 inch (74.9 cm) deep-guarded nasopharyngeal swabs, 16 inch (40.6 cm) unguarded proctology swabs, or 6 inch (15.2 cm) unguarded nasal swabs when characterized using culture, real time-qPCR, and 16S rRNA gene sequencing. Samples for aerobic culture, qPCR, and 16S rRNA gene sequencing were collected from the upper respiratory tract of cattle 2 weeks after feedlot arrival.
RESULTS
There was high concordance of culture and qPCR results for all swab types (results for 77% and 81% of sampled animals completely across all 3 swab types for culture and qPCR respectively). Microbial communities were highly similar among samples collected with different swab types, and differences identified relative to treatment for BRD were also similar. Positive qPCR results for M. haemolytica were highly concordant (81% agreed completely), but samples collected by deep-guarded swabbing had lower amounts of Mh DNA identified (Kruskal-Wallis analysis of variance on ranks, P < 0.05; Dunn-test for pairwise comparison with Benjamini-Hochberg correction, P < 0.05) and lower frequency of positive compared to nasal and proctology swabs (McNemar's Chi-square test, P < 0.05).
CONCLUSIONS
Though differences existed among different types of swabs collected from individual cattle, nasal swabs and proctology swabs offer comparable results to deep-guarded nasopharyngeal swabs when identifying and characterizing M. haemolytica by culture, 16S rRNA gene sequencing, and qPCR.
PubMed: 35964128
DOI: 10.1186/s42523-022-00197-6 -
Biomaterials Apr 2022Emerging antimicrobial resistance in infections asks for novel intervention strategies. Galacto-oligosaccharides (GOS) might be attractive alternatives to antibiotics...
Emerging antimicrobial resistance in infections asks for novel intervention strategies. Galacto-oligosaccharides (GOS) might be attractive alternatives to antibiotics due to their anti-inflammatory and anti-adhesive properties. Mannheimia haemolytica is one of the major Pasteurellaceae associated with bovine lung infections. Using M. haemolytica, we demonstrated that GOS have the capacity to reduce bacterial viability and can be used as adjuvant to improve antibiotic efficacy. Using M. haemolytica-treated primary bronchial epithelial cells (PBECs) of calves, we identified the anti-adhesive and anti-invasive activities of GOS. The observed inhibition of cytokine/chemokine release and the prevention of airway epithelial barrier dysfunction in M. haemolytica-treated PBECs by GOS might be related to the downregulation of "toll-like receptor 4/nuclear factor-κB" pathway and the anti-invasive and anti-adhesive properties of GOS. Particularly, GOS lowered lipopolysaccharides- but not flagellin-induced cytokine/chemokine release in calf and human airway epithelial cells. Finally, we performed in vivo experiments in calves and demonstrated for the first time that intranasal application of GOS can relieve lung infections/inflammation and lower M. haemolytica positivity in the lungs without affecting clinical performance. These findings not only shed light on the anti-inflammatory mechanisms of GOS during lung infections, but GOS might also be a promising anti-bacterial agent for preventing (lung) infections.
Topics: Animals; Anti-Bacterial Agents; Bacteria; Cattle; Humans; Lung; Mannheimia haemolytica; Oligosaccharides; Pneumonia
PubMed: 35286857
DOI: 10.1016/j.biomaterials.2022.121461