A post-mortem study of respiratory disease in small mustelids in south-west England

14 páginas, 3 tablas, 11 figuras.--This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.Background: Stoat (Mustela erminea) and weasel (Mustela nivalis) populations in south-west England are declining whilst polecats (Mustela putorius), absent for over a century, are increasing. Little is known about the health status of these species nationally. This study aimed at investigating respiratory disease in specimens found dead in south-west England. Results: Trauma caused by road traffic, predator attack or being trapped was the predominant cause of death in 42 stoats, 31 weasels and 20 polecats; most were in good physical condition. Skrjabingylus nasicola was present in all species (weasels 37 %, polecats 39 %, stoats 41 %) and infected animals showed no evidence of loss of body condition. Even in carcases stored frozen L larvae were frequently alive and highly motile. Angiostrongylus vasorum infection was diagnosed in two stoats and one weasel: in stoats infections were patent and the lung lesions were likely of clinical significance. These are believed to be the first records of A. vasorum in small mustelids. Pleuritis and pyothorax was seen in two polecats, in one case due to a migrating grass awn. Histological examination of lungs showed granulomata in stoats (38 %), weasels (52 %) and polecats (50 %). Spherules consistent with Emmonsia spp. adiaspores were present in the granulomata of stoats (60 %), weasels (36 %) and polecats (29 %). Adiaspore diameter in all three species was similar (means: stoats 39 μm, weasels 30 μm, polecats 36 μm); these are markedly smaller than that normally recorded for E. crescens. Although they lie within the accepted range for spores of Emmonsia parva this arid-zone species is not found in Britain, thus raising a question over the identity of the fungus. Cases showing numerous granulomata but few or no adiaspores were Ziehl-Neelsen-stain negative for acid-fast bacilli and IHC negative for Mycobacterium spp. However, in some cases PCR analyses revealed mycobacteria, including Mycobacterium kumamotonense and Mycobacterium avium Complex. One stoat had numerous unidentified small organisms present centrally within granulomata. Conclusions: Stoats, weasels and polecats in south-west England share several respiratory diseases, often of high prevalence, but the pathology would appear insufficient to impact on the health status of the populations and other ultimate causes of death should be investigated when examining these species.The authors gratefully acknowledge the histological support given by Trevor Whitbread, Judith Hargreaves, Richard Fox, Lucy Oldroyd, Malcolm Silkstone, Sonja Rivers and Michelle Woodman at Abbey Veterinary Services. They also thank Nicholas Davison, Beverley Rule and Philip Booth, AHVLA Truro, Mark Wessels, Finn Pathologists, Luke Roberts and Eric Morgan, Bristol University, Marc Artois, Campus Vétérinaire de Lyon. Becki Lawson, Fieke Molenaar, Tamsyn Stephenson, Zoe Greatorex and Jane Simpson at Wildlife Veterinary Investigation Centre assisted with post-mortem-examinations. David Groves, Kate Stokes, Derek Lord and Cornwall Mammal Group and Cornwall Wildlife Trust members and staff, James Williams, Somerset Otter Group, and David Couper, Royal Society for the Prevention of Cruelty to Animals helped with carcase submissions. Andrew Borman, Mycology Reference Laboratory South West Health Protection Agency kindly commented on draft manuscripts. Eileen Harris and Rodney Bray at Natural History Museum are thanked for advice on parasites. Those parts of this study performed at AHVLA were funded under the Diseases of Wildlife Scheme and those performed at the Moredun Research Institute were funded by the Scottish Government Rural and Environment Science and Analytical Services Division. J. Benavides is supported by a “Ramón y Cajal” contract of the Spanish Ministry of Economy and Competitiveness. None of the authors received funding from other outside sources for this work.Peer Reviewe

Phylogenomic exploration of the relationships between strains of Mycobacterium avium subspecies paratuberculosis.

BACKGROUND: Mycobacterium avium subspecies paratuberculosis (Map) is an infectious enteric pathogen that causes Johne's disease in livestock. Determining genetic diversity is prerequisite to understanding the epidemiology and biology of Map. We performed the first whole genome sequencing (WGS) of 141 global Map isolates that encompass the main molecular strain types currently reported. We investigated the phylogeny of the Map strains, the diversity of the genome and the limitations of commonly used genotyping methods. RESULTS: Single nucleotide polymorphism (SNP) and phylogenetic analyses confirmed two major lineages concordant with the former Type S and Type C designations. The Type I and Type III strain groups are subtypes of Type S, and Type B strains are a subtype of Type C and not restricted to Bison species. We found that the genome-wide SNPs detected provided greater resolution between isolates than currently employed genotyping methods. Furthermore, the SNP used for IS1311 typing is not informative, as it is likely to have occurred after Type S and C strains diverged and does not assign all strains to the correct lineage. Mycobacterial Interspersed Repetitive Unit-Variable Number Tandem Repeat (MIRU-VNTR) differentiates Type S from Type C but provides limited resolution between isolates within these lineages and the polymorphisms detected do not necessarily accurately reflect the phylogenetic relationships between strains. WGS of passaged strains and coalescent analysis of the collection revealed a very high level of genetic stability, with the substitution rate estimated to be less than 0.5 SNPs per genome per year. CONCLUSIONS: This study clarifies the phylogenetic relationships between the previously described Map strain groups, and highlights the limitations of current genotyping techniques. Map isolates exhibit restricted genetic diversity and a substitution rate consistent with a monomorphic pathogen. WGS provides the ultimate level of resolution for differentiation between strains. However, WGS alone will not be sufficient for tracing and tracking Map infections, yet importantly it can provide a phylogenetic context for affirming epidemiological connections

Inter- and Intra-subtype genotypic differences that differentiate <it>Mycobacterium avium</it> subspecies <it>paratuberculosis</it> strains

<p>Abstract</p> <p>Background</p> <p><it>Mycobacterium avium</it> subspecies <it>paratuberculosis</it> (Map) is the aetiological agent of Johne’s disease or paratuberculosis and is included within the <it>Mycobacterium avium</it> complex (MAC). Map strains are of two major types often referred to as ‘Sheep’ or ‘S-type’ and ‘Cattle’ or ‘C-type’. With the advent of more discriminatory typing techniques it has been possible to further classify the S-type strains into two groups referred to as Type I and Type III. This study was undertaken to genotype a large panel of S-type small ruminant isolates from different hosts and geographical origins and to compare them with a large panel of well documented C-type isolates to assess the genetic diversity of these strain types. Methods used included Mycobacterial Interspersed Repetitive Units - Variable-Number Tandem Repeat analysis (MIRU-VNTR), analysis of Large Sequence Polymorphisms by PCR (LSP analysis), Single Nucleotide Polymorphism (SNP) analysis of <it>gyr</it> genes, Pulsed-Field Gel Electrophoresis (PFGE) and Restriction Fragment Length Polymorphism analysis coupled with hybridization to IS<it>900</it> (IS<it>900</it>-RFLP) analysis.</p> <p>Results</p> <p>The presence of LSP^A4 and absence of LSP^A20 was confirmed in all 24 Map S-type strains analysed. SNPs within the gyr genes divided the S-type strains into types I and III. Twenty four PFGE multiplex profiles and eleven different IS<it>900</it>-RFLP profiles were identified among the S-type isolates, some of them not previously published. Both PFGE and IS<it>900</it>-RFLP segregated the S-type strains into types I and III and the results concurred with those of the <it>gyr</it> SNP analysis. Nine MIRU-VNTR genotypes were identified in these isolates. MIRU-VNTR analysis differentiated Map strains from other members of <it>Mycobacterium avium</it> Complex, and Map S-type from C-type but not type I from III. Pigmented Map isolates were found of type I or III.</p> <p>Conclusion</p> <p>This is the largest panel of S-type strains investigated to date. The S-type strains could be further divided into two subtypes, I and III by some of the typing techniques (IS<it>900</it>-RFLP, PFGE and SNP analysis of the <it>gyr</it> genes). MIRU-VNTR did not divide the strains into the subtypes I and III but did detect genetic differences between isolates within each of the subtypes. Pigmentation is not exclusively associated with type I strains.</p