Pig Isolates Research Articles

Genetic diversity of the ORF5 gene of porcine reproductive and respiratory syndrome virus isolates in China from 2006 to 2008

Since April 2006, swine herds have experienced the outbreaks of a highly pathogenic porcine reproductive and respiratory syndrome (PRRS) in China. To explore the possible mechanism of the emergence of the highly pathogenic PRRS and more fully understand the extent of genetic diversity of PRRSV in China, we analyzed the ORF5 gene sequences of 159 representative PRRSV isolates in 16 provinces from 2006 to 2008. Sequence and phylogenetic analyses showed that all these 159 isolates belonged to the North American genotype and were further divided into six subgenotypes; 140 of 159 isolates were closely related to the highly pathogenic PRRSV with 98.5–100% nucleotide and 98.3–100% amino acid sequence identities and belonged to Subgenotype I; and 3, 8, 4, 3, 1 of 159 isolates were part of Subgenotypes II–VI, respectively. Amino acid analysis of the GP5 protein revealed that all the isolates in Subgenotypes I–III were found to be highly variable in the primary neutralizing epitope; most of the isolates in Subgenotypes I and IV had more glycosylation sites than those in Subgenotypes II, III, V and VI; and 1, 5, and 9 unique amino acid mutations were observed in Subgenotypes I, IV and VI, respectively. In conclusion, our study provides the evidence of coexistence of six different subgenotype isolates in pigs in China from 2006 to 2008, and emphasizes the importance of reinforcing PRRSV surveillance, especially after the emergence of highly pathogenic PRRS in China.

Molecular diversity of porcine and human isolates ofPasteurella multocida

To examine the variability among Pasteurella multocida strains isolated from pigs (nasal, tonsil and lung specimens) and humans in France. The genetic diversity of 117 French isolates of P. multocida, obtained from pigs (n = 101) and humans (n = 16) and three reference strains, was evaluated by pulsed-field gel electrophoresis (PFGE) after macrorestriction with ApaI. Sixty-four patterns were detected. The genetic relationships revealed five clusters (Aa1, Aa2, Aa3, Ab and B). The pig isolates obtained from pneumonic lungs and nasal cavities were clustered in groups Ab and Aa1, respectively (P < 0.05). Up to four different PFGE patterns were detected in the same farm. Isolates producing dermonecrotic toxins were clustered only in group Aa1, suggesting that the toxigenic isolates were more genetically homogenous than the others. Conversely, cluster Aa3 was significantly associated with human isolates even if the human isolates are spread over most of the clusters. Pasteurella multocida strains were genetically diverse, but pig and human isolates were significantly clustered in distinct phylogenetic groups. The discrimination index was >0.95 in both populations of human and pig isolates. Therefore, ApaI-PFGE seems to be a useful tool for epidemiological tracing of P. multocida infections.

Characterization of sulphonamide-resistant Escherichia coli using comparison of sul2 gene sequences and multilocus sequence typing

The sul2 gene encodes sulphonamide resistance (Sul(R)) and is commonly found in Escherichia coli from different hosts. We typed E. coli isolates by multilocus sequence typing (MLST) and compared the results to sequence variation of sul2, in order to investigate the relation to host origin of pathogenic and commensal E. coli strains and to investigate whether transfer of sul2 into different genomic lineages has happened multiple times. Sixty-eight E. coli isolated in Denmark and Norway from different hosts and years were MLST typed and sul2 PCR products were sequenced and compared. PFGE was performed in a subset of isolates. All isolates were divided into 45 different sequence types (STs), with clonal complexes CC10, CC23, CC168, CC350 and CC69 being the most frequent. The sul2 gene from the majority of E. coli strains had only two point mutations, at positions 159 and 197, leading to a synonymous and a non-synonymous change, respectively. Five strains had extra single mutations. All poultry, poultry meat, and Danish human blood isolates had the same sul2 ST and some of these strains clustered under the same MLST STs, indicating that they shared habitats. Most PFGE profiles clustered according to source, but some included different sources. Sul(R) E. coli from different animals, food, human faeces and infections did not cluster according to their origin, suggesting that these habitats share E. coli and sul2 gene types. However, while pig isolates on one occasion clustered with urinary tract infection isolates, poultry isolates seemed more related to isolates from bloodstream infections in humans. Presence of mainly two types of the sul2 gene in both human and animal isolates, irrespective of date and geography, and the presence of both types in the same clonal lineages, suggest horizontal transfer of sul2.

Clostridium difficile: a new zoonotic agent?

Clostridium difficile is mainly considered a nosocomial pathogen associated with diarrhea and pseudomembranous colitis in hospitalized patients. Austrian hospitals reported 2761 cases of C. difficile infection (including 277 lethal outcomes) in 2007, compared with 777 cases (including 54 lethal outcomes) in 2003. The occurrence of community-acquired C. difficile infection is also increasingly reported. Recent studies have shown the occurrence of C. difficile in food and animals. The aim of the present study was to determine the occurrence of C. difficile in food and animals in Austria. Between March and July 2008, gut or fecal samples from 67 cows, 61 pigs and 59 broiler chickens were collected at Austrian abattoirs. Between February and April 2008 meat samples (51 beef [25 ground], 27 pork [17 ground] and 6 samples of chicken meat) were purchased at retail outlets. Of the 187 samples tested, eight yielded C. difficile: in cows 3/67 samples (4.5%) were positive, in pigs 2/61 (3.3%), in broiler chickens 3/59 (5%). Six of the eight isolates yielded toxigenic C. difficile (toxins A and B): 2/67 (3%) cow samples, 2/61 (3.3%) pig samples, 2/59 (3.4%) chicken samples. Genes for the binary toxin were detected in one of the two pig isolates, a PCR ribotype 126 strain. None of the 84 meat samples yielded C. difficile. The results of this Austrian study suggest that animal reservoirs are possible sources, via food, of human C. difficile infection.

Genetic Variability of Antigen B among Echinococcus granulosus Egyptian Isolates

Genetic polymorphisms of encoding antigen B2 gene (AgB2) in Echinococcus granulosus were studied using PCR-RFLP and DNA sequencing among 20 Egyptian isolates. Five isolates from different host origins (humans, camels, pigs, and sheep) were collected and used. All examined isolates of each host group gave very similar patterns of PCR-RFLP after restriction enzyme digestion with AluI, with the gene size of approximately 140 bp and 240 bp for sheep and human isolates, and approximately 150 bp and 250 bp for pig and camel isolates. No digestion pattern was obtained after incubation of all studied isolates with EcoRI. These results reveal high intra-group homogeneity. DNA sequence analysis highlighted that human infecting strain showed 100% identity with respect to sheep infecting isolate, 96% and 99% with pig and camel infecting isolates, respectively.

Evolution of theRhodococcus equi vapPathogenicity Island Seen through Comparison of Host-AssociatedvapAandvapBVirulence Plasmids

The pathogenic actinomycete Rhodococcus equi harbors different types of virulence plasmids associated with specific nonhuman hosts. We determined the complete DNA sequence of a vapB(+) plasmid, typically associated with pig isolates, and compared it with that of the horse-specific vapA(+) plasmid type. pVAPB1593, a circular 79,251-bp element, had the same housekeeping backbone as the vapA(+) plasmid but differed over an approximately 22-kb region. This variable region encompassed the vap pathogenicity island (PAI), was clearly subject to selective pressures different from those affecting the backbone, and showed major genetic rearrangements involving the vap genes. The pVAPB1593 PAI harbored five different vap genes (vapB and vapJ to -M, with vapK present in two copies), which encoded products differing by 24 to 84% in amino acid sequence from the six full-length vapA(+) plasmid-encoded Vap proteins, consistent with a role for the specific vap gene complement in R. equi host tropism. Sequence analyses, including interpolated variable-order motifs for detection of alien DNA and reconstruction of Vap family phylogenetic relationships, suggested that the vap PAI was acquired by an ancestor plasmid via lateral gene transfer, subsequently evolving by vap gene duplication and sequence diversification to give different (host-adapted) plasmids. The R. equi virulence plasmids belong to a new family of actinobacterial circular replicons characterized by an ancient conjugative backbone and a horizontally acquired niche-adaptive plasticity region.

Tentative identification of the species of Balantidium from ostriches ( Struthio camelus) as Balantidium coli-like by analysis of polymorphic DNA

The characteristics of Balantidium from ostriches ( Struthio camelus) are similar to those of Balantidium coli; however, the species Balantidium struthionis was proposed on the basis of the host species (ostriches) and the shape of the macronucleus (with a deep depression in one side). In the present work, we have performed morphological and genetic comparisons between isolates of Balantidium from ostriches and B. coli from pigs to determine the specific status of B. struthionis. The morphological characteristics of the trophozoites of Balantidium from ostriches were reviewed in 100 trophozoites from two isolates. The macronucleus’ shape of the ostrich Balantidium was highly variable, thus the use of this criterion for diagnostic purposes is not reliable. Besides, very few trophozoites showed a deep depression in their macronucleus and almost all the trophozoites conform to the description of B. coli. The complete sequence of the DNA coding for the 18s-rRNA–ITS1–5.8s-rRNA–ITS2 regions were obtained by PCR from five pig and five ostrich isolates. The sequences corresponding to the 18s and 5.8s-rRNA genes were identical for the ostrich and pig isolates. Two clearly different genotypes were found in the analysis of the ITS1 and ITS2 regions of the pig isolates; the genotype A was identified in all isolates, while the genotype B was found in only two of them. Their sequences show clear differences from that published corresponding to a B. coli gorilla isolate, which we will consider as a different genotype, C. In our opinion, these different B. coli genotypes reflect the genetic variability of this organism, but further studies would be necessary to determine if it could have practical importance. The polymorphism of the ITS regions have been also found in the ostrich isolates. The same genotypes A and B have been identified, although not as mixed infections. The morphological characteristics and the genetic results suggest that the species name B. struthionis is a synonym of B. coli; however, until experimental infections are carried out to determine if the parasite is transmissible between pigs and ostriches, it would be preferable to tentatively designate it as B. coli-like.

Serotypes of Pasteurella haemolytica and Pasteurella trehalosi Isolated from Farm Animals in Hungary

The biochemical and serological characteristics of 486 P. haemolytica and 31 P. trehalosi strains (517 in total) isolated from different lesions of cattle, sheep, goats, pigs and poultry were examined. A total of 476 P. haemolytica strains (97.9%) showed the characteristics typical of the former biotype A of P. haemolytica, while 10 isolates (2.1%), all from poultry, could not be biotyped. A total of 481 strains (93.0%) could be assigned to one of the 17 serotypes of P. haemolytica-P. trehalosi and 36 strains (7.0%) could not. The majority (83.6%) of the cattle isolates were serotypes A1 and A2. Among strains isolated from sheep all serotypes of P. haemolytica could be identified with the exception of A14, but serotypes A1, A2, A6, A8 and A5 were the most frequent. The overwhelming majority (94%) of the caprine isolates were A2, other serotypes occurred only sporadically. The pig isolates, which could be isolated only very rarely, represented different serotypes, while none of the 10 strains isolated from poultry could be biotyped or serotyped.

Comparison of genetic profiles of Campylobacter strains isolated from poultry, pig and Campylobacter human infections in Brittany, France

Five hundred eighty-two Campylobacter isolates (177 from humans, 319 from poultry and 86 from pig) collected in Brittany, France, in 2003 and 2004 were typed by pulsed-field gel electrophoresis. The number of human cases increased during the hot season, particularly for C. jejuni. Twelve genetic groups out of 27 contained human isolates collected over the two years. These groups had 21.3 and 17.0% of the isolates obtained in 2003 and 2004, respectively. In four cases, isolates from 2003 have the same Pulsed-field gel electrophoresis (PFGE) profile as isolates from 2004. Six PFGE profiles common to poultry and human isolates were identified. Poultry isolates were found in 47 clusters containing human isolates. Caeca from farms and slaughterhouses accounted for 66% of these isolates, with chicken legs obtained from supermarkets accounting for the other 34%. Pig isolates never clustered with poultry and human isolates. In conclusion, the analysis of the genetic profiles of Campylobacter resulting from human cases showed that there were few identical or genetically close isolates between the human cases declared in 2003 and those declared in 2004. This highlighted a great genetic diversity in the isolates and indicated that it should be difficult to bind the human infections with groups of Campylobacter isolates presenting particular genetic profiles. The Campylobacter isolates obtained from the two animal production systems had different genotypes, and isolates from pigs differed genetically from isolates obtained from humans. We found that 44.6% of human Campylobacter isolates were genetically related to genotypes found in poultry and a part of these campylobacteriosis are due to contact with poultry. This is not particularly surprising in Brittany, a farming area with many animal-rearing farms and slaughterhouses. This work highlights the implication of the poultry in the French human cases and that handling of poultry is also an important risk for Campylobacter infection in humans.

Characterization of TEM-, SHV- and AmpC-type β-lactamases from cephalosporin-resistant Enterobacteriaceae isolated from swine

Extended-spectrum β-lactamase (ESBL) and AmpC-producing Enterobacteriaceae are an increasing problem in human medicine and an emerging problem in the veterinary field. Our study, therefore, focused on assessing the prevalence of β-lactamases isolated from swine. Sixty-six Salmonella enterica serovar Typhimurium ( S. Typhimurium), 33 Salmonella enterica serovar Enteritidis ( S. Enteritidis), 26 Klebsiella pneumonia ( K. pneumoniae) and 130 Escherichia coli ( E. coli) pig isolates collected from 1999–2006 were screened for β-lactam resistance by the disk diffusion test (DDT) and micro-broth dilution. Among the isolates, five E. coli and five K. pneumoniae exhibited reduced susceptibility to the cephalosporins tested. PCR, plasmid profiling and Southern blot hybridization showed the presence of multiple β-lactamases in these isolates of animal origin. Hybridization patterns of the DHA-1 specific probe indicated that dissemination of DHA-1 related β-lactamases could be attributed to plasmids of one common size among the enteric microbes of animal origin. To the best of our knowledge, this study reports the first identification of SHV-28 and DHA-1 from microbes of animal origin.

Diagnostic testing of different stages of avian tuberculosis in naturally infected hens (Gallus domesticus) by the tuberculin skin and rapid agglutination tests, faecal and egg examinations

Avian tuberculosis (ATBC) is a significant cause of morbidity and mortality in birds in zoos and breeding establishments. The primary sources of &lt;I&gt;Mycobacterium avium&lt;/I&gt; subsp. &lt;I&gt;avium (MAA)&lt;/I&gt; of serotypes 1, 2 and 3 are infected animals (esp. birds), who shed &lt;I&gt;MAA&lt;/I&gt; in different ways and thus contaminate the environment. The first aim of this work was to compare the efficiency of the diagnostic methods that are routinely used for the diagnosis of ATBC (skin test, serology and culture of faeces and eggs) in naturally infected hens with different levels of infection. The second aim was to determine the excretion rate of &lt;I&gt;MAA&lt;/I&gt; in faeces and eggs. The tuberculin skin test gave a positive result in nine (42.9%) infected hens of which four (57.1%) and one (14.3%) were heavily and slightly infected hens, respectively. A positive serological response to &lt;I&gt;MAA&lt;/I&gt;-b antigen (water bird isolate of serotype 1) was observed in five (23.8%) and to &lt;I&gt;MAA&lt;/I&gt;-p antigen (pig isolate serotype 2) in seven (33.3%) hens. No correlation between serological and skin-test data was found. The results show that both techniques, serological and skin-test data are inadequate for the diagnosis of ATBC. In consecutively euthanized hens, with heavy infection and tuberculous lesions, serological positivity was significant (&lt;I&gt;P&lt;/I&gt; &lt; 0.05) in comparison with slightly infected hens lacking tuberculous lesions. Faecal culture detected &lt;I&gt;MAA&lt;/I&gt; in 50 (29.8%) of 168 samples collected for eight days before euthanasia. &lt;I&gt;MAA&lt;/I&gt; excretion in faeces was intermittent, but significantly (&lt;I&gt;P&lt;/I&gt; &lt; 0.01) higher in heavily infected hens. No mycobacteria were detected in any of the 43 examined eggs, which implies that the shedding of &lt;I&gt;MAA&lt;/I&gt;and/or transmission of ATBC through eggs may not be frequent events.

Salmonella Prevalence in Slaughtered Buffaloes and Pigs and Antimicrobial Susceptibility of Isolates in Vientiane, Lao People's Democratic Republic

This is the first report regarding isolation of Salmonella from cecum samples of buffaloes and pigs and characterization of the isolates in Laos. The organisms were isolated from 8% (4/50) of buffaloes and 76% (37/49) of pigs. In buffaloes, 3 animals harbored serotype 9,12: -:1,5, and 1 animal harbored both S. Derby and S. Javiana. In pigs, the most predominant serotypes were S. Derby (51%) followed by S. Anatum (45%), S. Weltevreden (15%) and S. Stanley (5%). The buffalo isolates were susceptible to the antimicrobials tested, whereas the pig isolates showed 10 resistance patterns to 1-5 antibiotics. Of the 59 pig isolates, the resistance rates to tetracycline, streptomycin, ampicillin, sulfamethoxazole-trimethoprim, chloramphenicol, amoxicillin-clavulanic acid and nalidixic acid were 24%, 22%, 14%, 5%, 2%, 2% and 2%, respectively. The results suggest that pigs and buffaloes harbor Salmonella, with a higher prevalence especially in pigs, and all the isolates showed sensitivity to cefotaxime, norfloxacin and ciprofloxacin.

Detection of hepatitis E virus in archived German wild boar serum samples

Hepatitis E is a rare human disease in Central Europe commonly imported from endemic regions. For autochthonous infections a zoonotic transmission from pigs, deer and wild boar is assumed. Using three different RT-PCR protocols, hepatitis E virus (HEV) RNA was detected in 10 out of 189 (5.3%) serum samples collected in 1995/1996 from wild boars in Germany. Sequence analysis indicates a close relationship with genotype 3 isolates of pigs and humans from the Netherlands and Japan. The results indicate that HEV is present in Germany since more than 10 years and that wild boar may function as a reservoir for HEV.

Genetic variability of Mycobacterium avium subsp. avium of pig isolates

The genetic diversity of 132 pig isolates of &lt;i&gt;Mycobacterium avium&lt;/i&gt; subsp. &lt;i&gt;avium&lt;/i&gt; (MAA) from the Czech Republic, the Slovak Republic and Slovenia was examined by IS&lt;i&gt;901&lt;/I&gt; restriction fragment length polymorphism (RFLP) analysis with restriction endonuclease &lt;i&gt;PvuII&lt;/i&gt;. A total of 18 RFLP types were detected. The occurrence frequency of respective RFLP types varied between respective pig farms, with the exception of one RFLP type F found in 21 (34.4%) of 61 farms and in 10 (55.6%) of 18 farms in the Czech Republic and Slovak Republic, respectively. Two different RFLP types were detected in 5 (33.3%) of the 15 studied farms, from which more than one isolate were examined. These results show the low variability of the &lt;i&gt;MAA&lt;/i&gt; isolates among the pig farms and the possibility of various sources of infection for pigs from infected farms.

Detection of Multiple Serotypes of Foot-and Mouth Disease Virus in Stored Isolates and the Implications for Control of the Disease in Kenya

The introduction of the antigen detection enzyme linked immunosorbent assay (ELISA) for routine serotyping at the foot-and-mouth disease laboratory, Embakasi, identified the existence of multiple serotype infections in some recent FMD outbreaks in Kenya. The study set out to reserotype using the ELISA some of the isolates from the collection at Embakasi previously serotypes by the complement fixation test (CFT) as single serotype isolates to identify any previous multiple serotype outbreaks in districts most affected by FMD. Seventy five (75%) foot-and-mouth diseases virus (FMDV) isolates stored at the laboratory were reserotyped. The isolates were obtained from the African buffalo (Syncerus caffer) eland (Taurotragus orynx), pigs and cattle during the period from 1971- to 2001. Serotypes O, A, SAT1 and SAT2 were identified from the cattle isolates. Serotypes O, SAT1 and SAT2 were found in buffalo isolates but not serotype A. Serotype SAT2 was identified in the eland isolate. The pig isolates were all serotype O. Five isolates (6.7%) had mixed infections of two serotypes each as follows; A and SAT 2(1), O and SAT 2(2) and SAT 1 and SAT2(2). The existence of multiple serotypes in infections among livestock and wildlife in Kenya presents challenges in FMD diagnosis requiring the use of more sensitive techniques. The inability of the CFT to identify the existence of an extra serotype in the sample could have led to ineffective, prolonged and costly control measures to contain the outbreaks. It is therefore desirable that the FMD laboratory at Embakasi applies more sensitive techniques such as ELISA and Polymerase Chain Reaction (PCR) for routine diagnosis. The Kenya Veterinarian Vol. 28 2005: pp. 20-23

Blastocystis sp. subtype 5: a possibly zoonotic genotype

Blastocystis has a widespread distribution in a variety of animals, which is a potential source of infection for humans. Previous studies show that Blastocystis sp. subtypes 1-4, 6, and 7 were composed of isolates from humans and animals, while Blastocystis sp. subtype 5 included only pig and cattle isolates. A more recent study on the basis of the SSU rDNA sequence has showed that a single Blastocystis isolate amplified directly from the faeces of a Thai human belongs to Blastocystis sp. subtype 5, but that study failed to cultivate this isolate. We report herein two human isolates from in vitro cultures belonging to Blastocystis sp. subtype 5 and one human isolate from in vitro culture containing two distinct genotypes of Blastocystis sp. subtypes 3 and 5 using PCR amplification with seven kinds of sequence-tagged site (STS) primers. Additionally, 16 Blastocystis isolates from pigs living in the same rural area with the three humans infected Blastocystis sp. subtype 5 were also genotyped by PCR with the STS primers, and all isolates from pigs and humans were compared by small-subunit ribosomal RNA (SSU rRNA) restriction-fragment-length polymorphism (RFLP) analyses using two restriction endonucleases (HinfI and RsaI). The results indicated that all of the isolates from pigs showed Blastocystis sp. subtype 5 and the RFLP patterns of all of the isolates from humans except for the mixed one were identical or quite similar to those of the 16 pig isolates with both HinfI and RsaI enzymes. These findings provide additional molecular-based evidence supporting the zoonotic potential of Blastocystis sp. subtype 5. This study also showed that Blastocystis sp. subtype 3 overgrew Blastocystis sp. subtype 5 in vitro.

An outbreak of pig rabies in Hunan province, China

The first dog-associated outbreak of rabies in swine in China (Hunan province) has been diagnosed and the related virus isolated. Sequence analysis showed that the pig isolate was a genotype 1 rabies virus with a very high nucleotide identity to local dog isolates.

An Easter outbreak of Salmonella Typhimurium DT 104A associated with traditional pork salami in Italy

Salmonella enterica is a common cause of gastrointestinal illness in Italy. S. Typhimurium accounts for approximately 40% of isolates, and most of these strains belong to the phage type DT104. We describe the investigation of an outbreak of S. Typhimurium DT104A, a subtype never observed before in Italy, which occurred in Rome during spring 2004.We conducted a matched case control study between 24 July and 9 September 2004. Controls were matched for age and area of residence. Each case had between one and four controls. Odds of exposure to potential risk factors and vehicles for the outbreak were compared between cases and controls. A multivariate analysis was conducted to estimate adjusted Odds Ratios.Sixty-three cases of S. Typhimurium DT 104A infection with onset between 1 April and 5 May 2004 were identified. Sixty-one were residents of Rome and two were residents of a neighbouring region. Twenty-six cases (43%) were enrolled in the study. Their median age was 7.5 years. Fourteen of 26 cases and 16 of 62 controls had eaten pork salami (OR= 25.5; 95% CI 1.6- 416.8). No food samples were available for testing. In northern Italy, two months prior to the outbreak, the veterinary surveillance system identified the first isolation of S. Typhimurium DT104A in a pig isolate. Both human and pig isolates showed indistinguishable PFGE patterns. It was not possible to trace the pig after the sample was taken at slaughter. The epidemiological evidence on the implication of pork salami in this outbreak suggests that pork products can also be a vehicle for salmonella in Italy and underlines the importance of good manufacturing practices for ready-to-eat foods. This investigation highlights the value of laboratory-based surveillance in identifying community-wide outbreaks of uncommon pathogens. It also underlines the need to improve surveillance timeliness, for promptly detecting outbreaks, undergoing field investigation, and implementing control measures. Moreover, our study shows the usefulness of integrated human and animal surveillance in tracing the possible source of infection.

P1810 Bulgarian Surveillance Tracking Antimicrobial Resistance –BulSTAR:bloodstream infections in 2005

Using the agar dilution method, antimicrobial susceptibility to human-use antibiotics was determined among Belgian faecal Salmonella isolates from healthy pigs and broiler chickens. Both epidemiological cut-off values and clinical breakpoints were applied for interpretation of the results. Cephalosporin-resistant isolates were examined for the presence of genes encoding CTX-M, SHV, TEM and CMY β-lactamases. All isolates with decreased quinolone susceptibility were screened for plasmid-borne genes qnr, qepA and aac(6′)-Ib-cr. In all, 368 Salmonella isolates were recovered from pigs and 452 from chickens. Clinical resistance to ciprofloxacin was absent in isolates of both host species, and was 1.9 and 13.1% to cefotaxime in pig and poultry isolates, respectively. Decreased susceptibility to cefotaxime amounted to 2.2 and 0.7%, whereas for ciprofloxacin this was 3.0 and 23.0% in pig and poultry isolates, respectively. Ciprofloxacin decreased susceptibility was limited to few serovars, mainly Paratyphi B. Multidrug resistance was markedly higher for pig isolates (39.7%) than for chicken isolates (17.3%). Sixty-six cefotaxime-resistant isolates, 59 from chickens and 7 from pigs, were phenotypically determined as ESBL/AmpC producers; predominantly Paratyphi B and Typhimurium serovars. BlaCTX-M (mostly blaCTXM-1, but also blaCTXM-2 and blaCTXM-9) and blaTEM-52 were the predominant ESBL genes. Only few isolates expressed SHV-12 or an AmpC enzyme (CMY-2). Isolates of four serovars carried qnr genes: Brandenburg and Llandof from pigs, both qnrS; Indiana and Paratyphi B from chickens with qnrB and qnrA. The latter isolate carried blaCTX-M-9 and was the only strain with a plasmid-borne quinolone resistance gene among the ESBL/AmpC producers. This Salmonella survey confirms that the ESBL/AmpC producers are particularly prevalent in chickens (12.8%), and much less in pigs (1.9%). A link between plasmid-borne quinolone resistance genes and ESBLs/AmpC was uncommon.

Multiple-locus variable-number tandem repeat analysis of Salmonella enterica subsp. enterica serovar Typhimurium: comparison of isolates from pigs, poultry and cases of human gastroenteritis

Pulsed-field gel electrophoresis (PFGE) and variable number tandem repeat (VNTR) profiles of 195 epidemiologically unrelated Salmonella Typhimurium strains isolated in 1997-2004 from pigs were analysed and the results compared to establish the discriminatory ability of each method. In order to investigate the epidemiology of S. Typhimurium from different populations, the VNTR profiles from pigs were compared with those obtained from 190 S. Typhimurium strains isolated from poultry and 186 strains isolated from human cases of gastroenteritis. A total of 195 strains of S. Typhimurium were tested by PFGE and VNTR. For PFGE, the restriction enzyme XbaI was used, and for VNTR, the number of repeats at five loci (STTR 9, 5, 6, 10pl and 3) were counted and assigned an allele number based on an established VNTR scheme. The results obtained showed improved discrimination of VNTR when compared with PFGE with 34 PFGE profiles identified compared with 96 different VNTR profiles for the pig isolates and 56 different VNTR types within the most common PFGE type. Within the three different populations, VNTR showed distinct subpopulations of VNTR type related not only to source, but also demonstrated common VNTR types within samples obtained from humans, poultry and pigs, especially in strains of phage type DT104. VNTR has taken the discrimination to a further level than that obtained through PFGE, and demonstrated an overlap in the genetic diversity of isolates tested across the three different populations, confirming previous suggestions that animals have an involvement in the dissemination of S. Typhimurium through the food chain. Salmonella Typhimurium remains an important concern as a food-borne zoonotic agent. The VNTR strategy described provides an accurate method of tracing strain dissemination, and adds a further level of discrimination to the PFGE type, providing potential benefits to epidemiological studies and the possibility of deciphering source attribution of cases.