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Antimicrobial Susceptibility and Resistance Mechanisms in Mannheimia haemolytica Isolates from Sheep at Slaughter.

ABSTRACT: Mannheimia haemolytica is the main pathogen contributing to pneumonic pasteurellosis in sheep. The aim of this study was to investigate the antimicrobial resistance levels in M. haemolytica isolates from the lungs of slaughtered sheep and to examine the genetic resistance mechanisms involved. A total of 256 M. haemolytica isolates, 169 from lungs with pneumonic lesions and 87 from lungs without lesions, were analyzed by the disk diffusion method for 12 antimicrobials, and the whole genome of 14 isolates was sequenced to identify antimicrobial resistance determinants. Levels of phenotypic resistance ranged from <2% for 10 antimicrobials (amoxicillin, amoxicillin-clavulanic, ceftiofur, cefquinome, lincomycin/spectinomycin, gentamicin, erythromycin, florfenicol, enrofloxacin, and doxycycline) to 4.3% for tetracycline and 89.1% for tylosin. Six isolates carried tetH genes and four isolates carried, in addition, the strA and sul2 genes in putative plasmid sequences. No mutations associated with macrolide resistance were identified in 23 rDNA sequences, suggesting that the M. haemolytica phenotypic results for tylosin should be interpreted with care in the absence of well-established epidemiological and clinical breakpoints. The identification of strains phenotypically resistant to tetracycline and of several resistance genes, some of which were present in plasmids, highlights the need for continuous monitoring of susceptibility patterns in Pasteurellaceae isolates from livestock.

SUBMITTER: Alvarez J

PROVIDER: S-EPMC10294796 | biostudies-literature | 2023 Jun

REPOSITORIES: biostudies-literature

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Antimicrobial Susceptibility and Resistance Mechanisms in Mannheimia haemolytica Isolates from Sheep at Slaughter.

Alvarez Julio J Calderón Bernal Johan M JM Torre-Fuentes Laura L Hernández Marta M Jimenez Chris E Pinto CEP Domínguez Lucas L Fernández-Garayzábal José F JF Vela Ana I AI Cid Dolores D

Animals : an open access journal from MDPI 20230614 12

Mannheimia haemolytica is the main pathogen contributing to pneumonic pasteurellosis in sheep. The aim of this study was to investigate the antimicrobial resistance levels in M. haemolytica isolates from the lungs of slaughtered sheep and to examine the genetic resistance mechanisms involved. A total of 256 M. haemolytica isolates, 169 from lungs with pneumonic lesions and 87 from lungs without lesions, were analyzed by the disk diffusion method for 12 antimicrobials, and th ...[more]

PMID: 37370501

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Project description:BACKGROUND:Mannheimia haemolytica has been recognized as the principal cause of pneumonic pasteurellosis in sheep and goats. It is one of the important diseases of small ruminants in Ethiopia. While annual vaccination using a monovalent vaccine (inactivated Pasteurella multocida biotype A) is common, respiratory diseases are still reported in various parts of Ethiopia. This suggests the need for further investigation into the species and strains responsible for the disease, which is vital information for development of a multivalent vaccine. The objective of the current study was to isolate M. heamolytica associated with pneumonic cases of sheep in selected areas of Central Ethiopia, determine its role and the strains/genotypes of the bacterium circulating in the study area. RESULTS:Bacteriological analysis of nasal swab samples collected from a total of 76 pneumonic cases of sheep showed that M. haemolytica was isolated from 26 of them while B.trehalosi from two cases. Further molecular analyses of the isolates using M. haemolytica species-specific and M.haemolytica serotype-1 antigen specific PCR assays revealed, 26 of the isolates were identified as M. haemolytica of which 21 of them were M. haemolytica serotype-1. Both M. haemolytica and B.trehalosi isolates were not detected in a PCR assay targeting capsular biosynthesis gene (capA) of P.multocida despite the non-specific products observed in M. haemolytica isolates. Phylogenetic analysis of M. haemolytica isolates included in this study in comparison with the reference strains with respect to PHSSA and Rpt2 genes revealed that the Ethiopian M. haemolytica isolates constituted three distinct genotypes consistent with site of origin. CONCLUSION:The study indicated that M.haemolytica is commonly associated with cases of pneumonia in sheep in the study areas of central Ethiopia although the remaining other pathogens responsible for majority of the cases are yet to be determined. Molecular characterization revealed the existence of three genotypes of M. haemolytica circulating in the study areas consistent to the site of isolation. The findings suggest further extensive work to determine all pathogens associated with sheep pneumonia and the strain distribution of M. heamolytica to understand its molecular epidemiology at national level and design cost effective prevention and control methods.

Project description:BackgroundMannhemia haemolytica is a Gram-negative bacterium and the principal etiological agent associated with bovine respiratory disease complex. They transform from a benign commensal to a deadly pathogen, during stress such as viral infection and transportation to feedlots and cause acute pleuropneumonia commonly known as shipping fever. The U.S beef industry alone loses more than one billion dollars annually due to shipping fever. Despite its enormous economic importance there are no specific and accurate genetic markers, which will aid in understanding the pathogenesis and epidemiology of M. haemolytica at molecular level and assist in devising an effective control strategy.DescriptionDuring our comparative genomic sequence analysis of three Mannheimia haemolytica isolates, we identified a number of genes that are unique to each strain. These genes are "high value targets" for future studies that attempt to correlate the variable gene pool with phenotype. We also identified a number of high confidence single nucleotide polymorphisms (hcSNPs) spread throughout the genome and focused on non-synonymous SNPs in known virulence genes. These SNPs will be used to design new hcSNP arrays to study variation across strains, and will potentially aid in understanding gene regulation and the mode of action of various virulence factors.ConclusionsDuring our analysis we identified previously unknown possible type III secretion effector proteins, clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated sequences (Cas). The presence of CRISPR regions is indicative of likely co-evolution with an associated phage. If proven functional, the presence of a type III secretion system in M. haemolytica will help us re-evaluate our approach to study host-pathogen interactions. We also identified various adhesins containing immuno-dominant domains, which may interfere with host-innate immunity and which could potentially serve as effective vaccine candidates.

Project description:Mastitis affects both dairy and meat/wool sheep industries with losses due to reductions in milk quality and quantity, increased treatment costs and restricted lamb growth. Effective vaccines would be important tools for mastitis control. However, the development of vaccines against mastitis has proved challenging due to the failure to target protective immunity to the mammary gland. In order to target responses to the mammary gland, this study tested whether local administration directly into the gland through the teat canal or in the udder skin confers protection against an intramammary infection. In this study, we tested a vaccine that confers protection against respiratory disease caused by Mannheimia haemolytica to determine if it also protects against intramammary infection by the same organism. No evidence of protection was observed in animals that received a subcutaneous immunisation in the udder skin, however, intramammary immunisation provided almost complete protection against an experimental challenge administered 7 days post immunisation but not if the challenge was delivered 14 days post immunisation. To investigate further the nature of this variation in response, the somatic cell count and concentration of cytokines Interleukin-1β, Interleukin-10 and Interleukin-17A was determined in milk over the course of each study. Intramammary immunisation induced an inflammatory response within the mammary gland, characterised by increases in SCC and in the production of cytokines IL-1β, IL-10, and IL-17A. This response was similar to that observed in un-vaccinated control animals post challenge. The SCC and cytokine levels had returned to levels comparable with un-vaccinated controls prior to challenge at both 7 and 14 days post immunisation. The transient nature of the protective effect is consistent with the priming of an innate antibacterial response within the mammary gland which provides protection against challenge at 7 days but is diminished by 14 days post-vaccination. Further studies are planned to determine the nature of the innate immune mechanisms associated with the protective effect described here to determine whether it may be exploited to improve ruminant udder health.

Dataset Information

Antimicrobial Susceptibility and Resistance Mechanisms in Mannheimia haemolytica Isolates from Sheep at Slaughter.

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Antimicrobial Susceptibility and Resistance Mechanisms in <i>Mannheimia haemolytica</i> Isolates from Sheep at Slaughter.

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