Veterinary and Agrochemical Research Center

Belgium

Veterinary and Agrochemical Research Center

Belgium

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Nemeghaire S.,Ghent University | Vanderhaeghen W.,Ghent University | Argudin M.A.,Veterinary and Agrochemical Research Center | Haesebrouck F.,Ghent University | Butaye P.,Ghent University
The Journal of antimicrobial chemotherapy | Year: 2014

OBJECTIVES: This study aimed at assessing the epidemiology and genetic diversity of methicillin-resistant Staphylococcus sciuri (MRSS) from different farm animal species.METHODS: Nasal swabs were collected from 200 pigs, 100 dairy cows, 100 beef cows, 150 veal calves and 200 broilers. Colonies were isolated on selective media containing cefoxitin and the mecA gene was detected by PCR. Antimicrobial resistance was determined by broth microdilution. The genetic diversity was assessed by PFGE and resistance and virulence genes were detected by microarray analysis.RESULTS: The total MRSS prevalence at the animal level was estimated at 9.5%, varying from ∼10% in veal (13.3%), broilers (12.5%) and dairy cows (10.0%) to 6.5% in pigs and 3.0% in beef cows. mecA was detected in all isolates. SCCmec elements of type III and non-typeable ones were seen most frequently. More than 90% of isolates were non-wild-type (NWT) for gentamicin, penicillin, tiamulin, clindamycin and quinupristin/dalfopristin. The frequency of NWT isolates for fusidic acid and trimethoprim ranged between 78% and 87%. PFGE analysis allowed distinction between two major clusters. Most isolates tested by microarray carried erm and tet genes. Virulence genes were also detected, including an isa gene encoding an immune-evasion factor and the hsdS2 gene encoding a site-specific deoxyribonuclease.CONCLUSIONS: This study shows that multiresistant MRSS is carried by different farm animal species. Although some animals shared the same strain, PFGE showed different patterns, indicating high diversity among the MRSS isolates recovered. The absence of clusters associated with a certain animal species suggests low host specificity. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.


Jamrozy D.M.,Animal Health and Veterinary Laboratories Agency | Jamrozy D.M.,Kingston University | Coldham N.G.,Animal Health and Veterinary Laboratories Agency | Butaye P.,Ghent University | And 2 more authors.
Journal of Antimicrobial Chemotherapy | Year: 2014

Objectives: Previously described methicillin-resistant Staphylococcus aureus (MRSA) ST398 strains revealed a high frequency of phenotypic resistance to spectinomycin. However, only a few were found to carry the spc resistance determinant. The aim of this study was to identify the genetic mechanism of spectinomycin resistance among spc-negative MRSA ST398 strains. Methods: Nine spectinomycin-resistant, but spc-negative, MRSA ST398 strains were analysed. The strains were screened for carriage of the spw gene and tested for the presence of transferrable spectinomycin resistance. Plasmid DNAwas isolated from all strains and used in transformation assays. The plasmid identified as mediating resistance to spectinomycin was fully sequenced. The function of the novel spectinomycin resistance gene was confirmed by restriction digest inactivation and its distribution was determined using a PCR assay. Results: A single MRSA ST398 strain was spw positive. The remaining strains carried a plasmid that mediated resistance to spectinomycin. Sequence analysis of a single plasmid, termed pDJ91S, revealed that it was 3928 bp in size and contained three open reading frames: a novel spectinomycin resistance gene, designated spd, as well as a repN gene and a rec gene. The XmnI digest inactivation of the spd gene resulted in a 4-fold decrease in spectinomycin MIC. The spd gene was detected in seven other spectinomycin-resistant MRSA ST398 strains that carried a plasmid comparable in size to pDJ91S. Conclusions: A novel gene, designated spd, that confers resistance to spectinomycin has been identified on a small plasmid in MRSA ST398. © Crown copyright 2014.


Jamrozy D.M.,Animal Health and Veterinary Laboratories Agency | Jamrozy D.M.,Kingston University | Fielder M.D.,Kingston University | Butaye P.,Veterinary and Agrochemical Research Center | And 2 more authors.
PLoS ONE | Year: 2012

The high prevalence of methicillin-resistant Staphylococcus aureus (MRSA) ST398 among pigs in certain European countries and North America and its occurrence in other animal species raises a question concerning the molecular mechanisms mediating the success of this lineage. In this study a panel of S. aureus strains belonging to sequence type (ST) 5 (n = 4), ST8 (n = 5), ST15 (n = 5), ST22 (n = 8), clonal complex (CC) 30 (n = 8), CC97 (n = 8), CC130 (n = 4), CC151 (n = 4) and ST398 (n = 18) were screened by DNA microarray and PCR for the carriage of virulence and antimicrobial resistance genes. Isolates belonging to the same sequence type/clonal complex (ST/CC) were found to share similar virulence gene profiles. The ST398 lineage displayed the lowest content of virulence genes, which consisted mainly of genes detected among the majority or all of the analysed lineages. All MRSA ST398 isolates lacked accessory virulence genes that were detected in other ST/CC. In contrast to virulence genotype, the antimicrobial resistance genes profiles varied between isolates belonging to the same ST/CC and profile similarities could be observed for isolates from different lineages. MRSA ST398 isolates in particular displayed significant diversity and high content of antimicrobial resistance genes. This was comparable with certain MRSA belonging to other sequence types particularly the equine MRSA ST8. The apparent lack of significant virulence genes among MRSA ST398 strains, demonstrates that the lineage features a unique genetic background but no ST398-specific virulence markers could be identified. © 2012 Jamrozy et al.


Nemeghaire S.,Veterinary and Agrochemical Research Center | Argudin M.A.,Veterinary and Agrochemical Research Center | Haesebrouck F.,Ghent University | Butaye P.,Veterinary and Agrochemical Research Center | Butaye P.,Ghent University
Veterinary Microbiology | Year: 2014

Staphylococcus sciuri is commonly found on the skin of animals and humans as well as in the environment. However, little is known on its prevalence, resistance and epidemiology. Therefore, we investigated the prevalence of methicillin resistant S. sciuri (MRSS) strains in poultry, as they may represent a reservoir of resistance genes for other strains. In 2011, 281 poultry farms were sampled by taking nasal swabs of 20 animals. The swabs were pooled and MRSS were selectively isolated. Genus and methicillin resistance were determined by PCR and species identification was performed using transfer RNA-intergenic spacer analysis. MRSS were further characterised by SCC. mec typing, PFGE, microarray and susceptibility testing. Eighty-seven MRSS were isolated resulting in an estimated prevalence of 31.0%. The prevalence in broilers did not significantly differ from that in layers. Most isolates harboured a non-typeable SCC. mec and a little less than 40% carried SCC. mec type III. Isolates from broiler farms carried mostly the SCC. mec type III, while isolates from layer farms carried mostly the non-typeable SCC. mec cassette. The 87 isolates generated 47 different SmaI-PFGE profiles that grouped in two main clusters corresponding to the two farm types. All isolates were resistant to fusidic acid, tiamulin and gentamicin and were sensitive to rifampicin and vancomycin. Isolates selected for microarray analysis carried a broad range of antimicrobial resistance and virulence genes. This study showed that MRSS is carried by healthy chickens at the same level in both broilers and layers. They represent a large reservoir for resistance and virulence genes. Strains from layers and broilers represent different clusters. © 2014 Elsevier B.V.


Wieland B.,Lane College | Dhollander S.,Animal Health and Welfare Unit | Salman M.,Colorado State University | Koenen F.,Veterinary and Agrochemical Research Center
Preventive Veterinary Medicine | Year: 2011

In the absence of data, qualitative risk assessment frameworks have proved useful to assess risks associated with animal health diseases. As part of a scientific opinion for the European Commission (EC) on African Swine Fever (ASF), a working group of the European Food Safety Authority (EFSA) assessed the risk of ASF remaining endemic in Trans Caucasus Countries (TCC) and the Russian Federation (RF) and the risk of ASF becoming endemic in the EU if disease were introduced. The aim was to develop a tool to evaluate how current control or preventive measures mitigate the risk of spread and giving decision makers the means to review how strengthening of surveillance and control measures would mitigate the risk of disease spread. Based on a generic model outlining disease introduction, spread and endemicity in a region, the impact of risk mitigation measures on spread of disease was assessed for specific risk questions. The resulting hierarchical models consisted of key steps containing several sub-steps. For each step of the risk pathways risk estimates were determined by the expert group based on existing data or through expert opinion elicitation. Risk estimates were combined using two different combination matrices, one to combine estimates of independent steps and one to combine conditional probabilities. The qualitative risk assessment indicated a moderate risk that ASF will remain endemic in current affected areas in the TCC and RF and a high risk of spread to currently unaffected areas. If introduced into the EU, ASF is likely to be controlled effectively in the production sector with high or limited biosecurity. In the free range production sector, however, there is a moderate risk of ASF becoming endemic due to wild boar contact, non-compliance with animal movement bans, and difficult access to all individual pigs upon implementation of control measures. This study demonstrated the advantages of a systematic framework to assist an expert panel to carry out a risk assessment as it helped experts to disassociate steps in the risk pathway and to overcome preconceived notions of final risk estimates. The approach presented here shows how a qualitative risk assessment framework can address animal diseases with complexity in their spread and control measures and how transparency of the resulting estimates was achieved. © 2011 Elsevier B.V.


Boland C.,Veterinary and Agrochemical Research Center | Bertrand S.,Scientific Institute of Public Health | Mattheus W.,Scientific Institute of Public Health | Dierick K.,Scientific Institute of Public Health | Wattiau P.,Veterinary and Agrochemical Research Center
Veterinary Microbiology | Year: 2014

To assess the distribution of Salmonella 4,[5]:i:- subtypes in the Belgian food chain and compare it to the subtypes associated with human infections, a molecular assessment was initiated. Two hundred fifty-three Salmonella isolates serotyped as 4,[5]:i:- during the period 2008-2011 in Belgium and originating from animal productions, food or human clinical samples were analysed by a specific duplex PCR. One hundred ninety-four isolates (76.7%) fit the profile of a S. Typhimurium monophasic variant as defined by the European Food Safety Authority. The other isolates possessed but did not express the phase II flagellin gene (23.3%). Multiple Locus Variable Number of Tandem Repeats Analysis (MLVA) revealed many but closely related profiles in the fljB-negative S. Typhimurium monophasic variant isolates. Some MLVA types were associated with both human and animal isolates but no unique source of human contamination could be demonstrated. © 2013 Elsevier B.V.


Wattiau P.,Veterinary and Agrochemical Research Center | Boland C.,Veterinary and Agrochemical Research Center | Bertrand S.,Scientific Institute of Public Health
Applied and Environmental Microbiology | Year: 2011

For more than 80 years, subtyping of Salmonella enterica has been routinely performed by serotyping, a method in which surface antigens are identified based on agglutination reactions with specific antibodies. The serotyping scheme, which is continuously updated as new serovars are discovered, has generated over time a data set of the utmost significance, allowing long-term epidemiological surveillance of Salmonella in the food chain and in public health control. Conceptually, serotyping provides no information regarding the phyletic relationships inside the different Salmonella enterica subspecies. In epidemiological investigations, identification and tracking of salmonellosis outbreaks require the use of methods that can fingerprint the causative strains at a taxonomic level far more specific than the one achieved by serotyping. During the last 2 decades, alternative methods that could successfully identify the serovar of a given strain by probing its DNA have emerged, and molecular biology-based methods have been made available to address phylogeny and fingerprinting issues. At the same time, accredited diagnostics have become increasingly generalized, imposing stringent methodological requirements in terms of traceability and measurability. In these new contexts, the hand-crafted character of classical serotyping is being challenged, although it is widely accepted that classification into serovars should be maintained. This review summarizes and discusses modern typing methods, with a particular focus on those having potential as alternatives for classical serotyping or for subtyping Salmonella strains at a deeper level. © 2011, American Society for Microbiology.


Caij A.B.,Veterinary and Agrochemical Research Center | Tignon M.,Veterinary and Agrochemical Research Center
Transboundary and emerging diseases | Year: 2014

In January 2010, the United Kingdom notified cases of equine infectious anaemia (EIA) in two horses introduced from Belgium. The animals came from one assembly centre in Romania and had transited through Belgium with 16 other horses. Nine of them, bought by a Belgian horse breeder, were investigated in Belgium and revealed one additional EIA-positive animal. Afterwards, the Belgian Federal Agency for the Safety of the Food Chain (FASFC) organized a serological EIA survey of the horses introduced into Belgium from Romania between 2007 and 2009. Among the 95 horses identified, six additional serological positive cases were found that had been introduced into Belgium in 2008 (n = 4) and in 2009 (n = 2). The survey was extended to the horses in contact with the positive cases, but all contact animals were negative, indicating the absence of transmission. Virological examination performed on tissue samples collected from two seropositive animals demonstrated the presence of viral DNA of EIA virus. Phylogenetic analysis based on the sequences of EIA virus gag gene clustered the Belgian isolates with Romanian strains isolated in 2009. The presumption of a common Belgian origin could be rejected. © 2012 Blackwell Verlag GmbH.


Van Borm S.,Veterinary and Agrochemical Research Center | Rosseel T.,Veterinary and Agrochemical Research Center | Steensels M.,Veterinary and Agrochemical Research Center | Van den Berg T.,Veterinary and Agrochemical Research Center | Lambrecht B.,Veterinary and Agrochemical Research Center
Virus Research | Year: 2013

We used next generation sequencing on random amplified viral nucleic acids to determine the genome sequence of 11 pigeon paramyxovirus type 1 (PPMV-1) isolates from Belgium (period 1998-2011). The PPMV-1 deep sequence data allowed identification of sequence variability in multiple PPMV-1 isolates, including one STOP codon in the Matrix gene which was present in 15% of the viral population of one isolate. Notably, mutations that were previously associated with pathogenicity in chickens were identified as minor sequence variants in one parent laboratory strain. A phylogenetic analysis of the consensus PPMV-1 genome sequences was performed. In addition to providing nearly complete paramyxovirus genome sequences, our sequence-independent approach identified the presence of pigeon circovirus (PiCV) sequences in four of these viral stocks. Real-time quantitative RT-PCR analysis specific for PMV-1 and PiCV showed that these contaminations were present in seven viral stocks consisting of allantoic fluids and was occasionally also detected in stocks passaged in embryonated chicken eggs. Phylogenetic analysis of the PiCV consensus genome sequences showed a circulation of PiCV covering the full genetic diversity of known PiCV.This study shows the value of novel sequence independent technologies for access to sequence information for the control of reference virus stocks and other biological materials, as co-infecting viruses or sequence variants from the original sample may persist in the stocks without being identified by the routine virus-specific diagnostic tools. The exact role of PiCV in pigeon disease - in particular Newcastle disease - and its potential interference with PPMV-1 diagnostics remains to be investigated. © 2012 Elsevier B.V.


Vandendriessche S.,Free University of Colombia | Vandendriessche S.,Veterinary and Agrochemical Research Center | Kadlec K.,Institute of Farm Animal Genetics | Schwarz S.,Institute of Farm Animal Genetics | Denis O.,Free University of Colombia
Journal of Antimicrobial Chemotherapy | Year: 2011

Objectives: Methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-susceptible S. aureus (MSSA), collected from 109 Belgian acute-care hospitals during a national survey in 2008, were investigated for macrolide-lincosamide (ML) resistance with particular emphasis on the analysis of erm(T)-carrying isolates. Methods: In total, 314 MRSA and 212 MSSA were collected and characterized by spa typing. The SCCmec type of MRSA was determined. Resistance to ML antibiotics was detected by agar dilution and resistant strains were screened by PCR for erm(A), erm(C) and msr(A). Five ML-resistant MSSA isolates, negative by PCR for the aforementioned genes, were further characterized. Results: Half of all MRSA isolates (n=157; 50.0%) were resistant to erythromycin and harboured the gene erm(A) (n=112), erm(C) (n=41), erm(A) + erm(C) (n=3) or msr(A) (n=1). The erm(A) gene was mainly present in MRSA spa-CC002-ST5-SCCmec II and spa-CC008-ST8-SCCmec IV (where CC stands for clonal complex and ST stands for sequence type); the distribution of erm(C) was more diverse. Thirty-five of the 40 erythromycin-resistant MSSA (18.9%) carried the gene erm(A) (n=17), erm(C) (n=9) or msr(A) (n=9). The remaining five MSSA were ST398-t571 isolates, which exhibited closely related ApaI PFGE patterns, harboured the gene erm(T) in the chromosomal DNA and did not exhibit additional resistances. These isolates were from severe infections in patients, of whom four had no contact and one had only indirect contact with livestock via a family member working in animal husbandry. Conclusions: The ML-streptogramin B ('MLS B') resistance genes erm(A) or erm(C) were detected in the majority of ML-resistant MRSA and MSSA isolates. The erm(T) gene was identified in MSSA ST398 isolates from five independent patients who lacked direct contact with livestock. © The Author 2011. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved.

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