Evaluation of oral fluids for surveillance of foodborne and zoonotic pathogens in pig farms.

The use of oral fluid (OF) to detect zoonotic pathogens in pigs has been only scarcely assessed. We evaluated OF as a potential specimen for detection by culture of methicillin-resistant Staphylococcus aureus (MRSA) and Yersinia enterocolitica, and the detection of antibodies against Salmonella spp. and hepatitis E virus (HEV) using commercial ELISAs. Samples from 33 pig farms were collected at the beginning and end of the fattening period. Results of the OF samples were compared with the results of serum samples and nasal swabs from individual pigs and pen floor fecal samples, using the Cohen kappa (κ) and the McNemar test. For Salmonella spp. antibodies, OF samples were negative, although the corresponding serum samples were positive. The detection of HEV antibodies in sera and OF had agreement at the first sampling, and poor and significant agreement at the second sampling (κ = 0.185, McNemar p = 0.238; κ = 0.088, McNemar p < 0.001). At both sampling times, the detection of MRSA in nasal swabs and OF showed agreement (κ = 0.466, McNemar p = 0.077; κ = 0.603, McNemar p = 1); agreement was seen for the detection of Y. enterocolitica in fecal and OF samples (κ = 0.012, McNemar p = 0.868; κ = 0.082, McNemar p = 0.061, respectively). According to the McNemar test, the use of pen-based OFs is more feasible for the detection of MRSA and Y. enterocolitica by culture than is detection of antibodies by commercial ELISA.

Prevalence and molecular characterization of C. pecorum detected in Swiss fattening pigs.

Chlamydia (C.) pecorum, an obligate intracellular bacterial species commonly found in ruminants, can also occur in pigs. However, its significance as a potential porcine pathogen, or commensal, is still unclear. In a previous study (Hoffmann et al. 2015), mixed infections of C. suis and C. pecorum were detected in 14 Swiss fattening pig farms. Using these samples, we aimed to investigate the infection dynamics of C. suis and C. pecorum mixed infections in these farms. In addition, we analyzed the genetic diversity of Swiss porcine C. pecorum strains in relation to globally circulating strains. In total, 1284 conjunctival and rectal swabs from 391 pigs, collected at the beginning and end of the fattening period, were tested during the course of this study. We determined the bacterial loads of C. suis and C. pecorum using species-specific real-time PCR (qPCR) and compared these results to already existing DNA-microarray and Chlamydiaceae qPCR data. Overall, C. suis and Chlamydiaceae copy numbers decreased in the course of the fattening period, whereas C. pecorum copy numbers increased. No association was found between clinical signs (conjunctivitis, lameness and diarrhea) and the bacterial loads. Preventive antibiotic treatment at the beginning of the fattening period significantly lowered the chlamydial load and outdoor access was associated with higher loads. Proximity to the nearest ruminants correlated with increased C. pecorum loads, indicating that C. pecorum could be transmitted from ruminants to pigs. Multi-locus sequence typing (MLST) and major outer membrane protein (ompA) genotyping revealed two novel sequence types (STs) (301, 302) and seven unique ompA genotypes (1-7) that appear to form a specific clade separate from other European C. pecorum strains.

Selective Pressure Promotes Tetracycline Resistance of Chlamydia Suis in Fattening Pigs.

In pigs, Chlamydia suis has been associated with respiratory disease, diarrhea and conjunctivitis, but there is a high rate of inapparent C. suis infection found in the gastrointestinal tract of pigs. Tetracycline resistance in C. suis has been described in the USA, Italy, Switzerland, Belgium, Cyprus and Israel. Tetracyclines are commonly used in pig production due to their broad-spectrum activity and relatively low cost. The aim of this study was to isolate clinical C. suis samples in cell culture and to evaluate their antibiotic susceptibility in vitro under consideration of antibiotic treatment on herd level. Swab samples (n = 158) identified as C. suis originating from 24 farms were further processed for isolation, which was successful in 71% of attempts with a significantly higher success rate from fecal swabs compared to conjunctival swabs. The farms were divided into three treatment groups: A) farms without antibiotic treatment, B) farms with prophylactic oral antibiotic treatment of the whole herd consisting of trimethoprime, sulfadimidin and sulfathiazole (TSS), or C) farms giving herd treatment with chlortetracycline with or without tylosin and sulfadimidin (CTS). 59 isolates and their corresponding clinical samples were selected and tested for the presence or absence of the tetracycline resistance class C gene [tet(C)] by conventional PCR and isolates were further investigated for their antibiotic susceptibility in vitro. The phenotype of the investigated isolates was either classified as tetracycline sensitive (Minimum inhibitory concentration [MIC] < 2 µg/ml), intermediate (2 µg/ml ≤ MIC < 4 µg/ml) or resistant (MIC ≥ 4 µg/ml). Results of groups and individual pigs were correlated with antibiotic treatment and time of sampling (beginning/end of the fattening period). We found clear evidence for selective pressure as absence of antibiotics led to isolation of only tetracycline sensitive or intermediate strains whereas tetracycline treatment resulted in a greater number of tetracycline resistant isolates.

Molecular characterization and virus neutralization patterns of severe, non-epizootic forms of feline calicivirus infections resembling virulent systemic disease in cats in Switzerland and in Liechtenstein.

Feline calicivirus (FCV) infections are associated with oral ulceration, chronic stomatitis and a limping syndrome. Epizootic outbreaks of virulent systemic disease (VSD) have been reported in the USA and Europe. Here, the molecular characterization and neutralization patterns of FCV isolates from cases of severe, non-epizootic infection associated with skin ulceration and edema are presented. Samples from eleven symptomatic cats, four in-contact cats and 27 cats with no contact with symptomatic cats were collected and tested for FCV, feline herpesvirus-1 (FHV-1), feline leukemia virus (FeLV) and feline immunodeficiency virus (FIV). Phylogenetic analyses based on the capsid (VP1) gene of FCV and virus neutralization with antisera raised against four FCV vaccine strains were performed. Nine kittens and two adult cats in two shelters and two veterinary clinics in four geographically distinct locations in Switzerland and Liechtenstein were affected. The cats showed fever, tongue and skin ulceration, head and paw edema, and occasionally jaundice, generalized edema and dyspnea. All symptomatic cats tested FCV-positive but were negative for FHV-1, FeLV and FIV, with the exception of one FIV-positive kitten. All kittens of one litter and both adult cats died. The disease did not spread to cats in the environment. Cats in the environment displayed phylogenetically distinct, but related, FCV strains. Virus neutralization patterns suggested that some cases might have been potentially prevented by vaccination with the optimal vaccine strain. In conclusion, clinicians should be aware of severe, non-epizootic forms of FCV infections with initial clinical presentations similar to VSD.

Prevalence of Chlamydial Infections in Fattening Pigs and Their Influencing Factors.

Chlamydial infections in pigs are associated with respiratory disease, diarrhea, conjunctivitis and other pathologies. The aim of this study was to define the prevalence of Chlamydiaceae in Swiss fattening pigs by applying sensitive and specific detection methods and to correlate prior antibiotic treatment and farm related factors with differences in prevalence. Conjunctival and fecal swabs were collected from 636 pigs in 29 Swiss fattening pig farms with and without antibiotic treatment, at the beginning and the end of the fattening period. The swabs were screened by real-time PCR for Chlamydiaceae. For the chlamydial detection and species-identification, a DNA-microarray analysis was performed. All farms were positive for Chlamydiaceae with 94.3 and 92.0% prevalence in fecal swabs as well as 45.9 and 32.6% in conjunctival swabs at the first and second time points, respectively. Antibiotic treatment could not clear the infection on herd level. Potential contact with wild boars was a significant risk factor, while hygiene criteria did not influence chlamydial prevalence. A correlation of chlamydial positivity to diarrhea, but not to conjunctivitis was evident. Chlamydia suis was the predominant species. Mixed infections with C. suis and C. pecorum were common, with a substantial increase in C. pecorum positivity at the end of the fattening period, and this finding was associated with ruminant contact. C. abortus was detected in one conjunctival swab. In this study, C. suis inhabited the intestinal tract of nearly all examined pigs, implying a long-term infection. C. pecorum was also common and might be transmitted to pigs by ruminants.