Characterization of third-generation cephalosporin-resistant Escherichia coli from slaughter calves and fattening pigs: A pilot study for monitoring antimicrobial resistance by whole genome sequencing in Switzerland.

INTRODUCTION: Whole genome sequencing (WGS) was introduced into Swiss antimicrobial resistance monitoring in 2022 as an additional method to phenotypic antimicrobial susceptibility testing by broth microdilution to characterize presumptive third-generation cephalosporin-resistant (3GC-R) Escherichia coli. Caecal samples from Swiss slaughter calves and fattening pigs, as well as beef and pork meat from Swiss retail taken in 2021, were analyzed for the presence of 3GC-R E. coli according to European harmonized protocols. In 2021, 3GC-R E. coli was detected in 23,8 % of slaughter calves, 5,9 % of fattening pigs, and 0 % of meat. Comparative analysis of the antimicrobial resistance results obtained by phenotypic measurement and those obtained by the detection of corresponding underlying molecular mechanisms by WGS showed very high agreement (99 %). Resistance to third-generation cephalosporins (3GCs) was mainly associated with the presence of blaCTX-M-15 in E. coli isolates from calves and blaCTX-M-1 in E. coli isolates from pigs and mutations in the ampC-promoter (g.-42 C>T) in E. coli isolates from both animal species. Moreover, WGS data were used for phylogenetic analysis based on multi locus sequence types (MLST) and core genome MLST(cgMLST) revealing that 3GC-R E. coli isolated from Swiss slaughter calves and fattening pigs were genetically diverse. In this study, it was shown that using WGS alone to monitor antimicrobial resistance could detect trends in known molecular antimicrobial resistance mechanisms while also providing other valuable information about the isolates, such as genetic relatedness. However, only by combining phenotypic susceptibility testing and WGS early detection of previously unknown resistance mechanisms will be possible.

INTRODUCTION: Le séquençage du génome entier (Whole Genome Sequencing, WGS) a été introduit dans la surveillance suisse de la résistance aux antibiotiques en 2022 en tant que méthode supplémentaire aux tests phénotypiques de sensibilité aux antibiotiques pour caractériser les Escherichia coli résistants aux céphalosporines de troisième génération (3GC-R). Des échantillons de cæcum pris en 2021 à l'abattoir de veaux et de porcs suisses, ainsi que de viande de bœuf et de porc provenant de détaillants suisses ont été analysés pour détecter la présence d'E. coli 3GC-R conformément aux protocoles européens harmonisés. En 2021, les E. coli 3GC-R ont été détectés dans 23,8 % des veaux d'abattage, 5,9 % des porcs d'engraissement et 0 % dans la viande. Les résultats de résistance aux antibiotiques obtenus par mesure phénotypique et ceux obtenus par la détection des mécanismes moléculaires sous-jacents concordaient à 99 %. La résistance aux céphalosporines de troisième génération était principalement associée à la pré-sence de blaCTX-M-15 dans les isolats d'E. coli provenant de veaux et de blaCTX-M-1 dans les isolats d'E. coli provenant de porcs et à des mutations dans le promoteur ampC (g.-42 C>T) dans les isolats d'E. coli provenant des deux espèces animales. Les données WGS ont également été utilisées pour une analyse phylogénétique basée sur les types de séquences multilocus (MLST) et MLST du génome de base (cgMLST) révélant que les E. coli 3GC-R isolés des veaux et des porcs suisses étaient génétiquement divers. Dans cette étude, il a été démontré que l'utilisation du WGS seul pour surveiller la résistance aux antibiotiques pouvait détecter des tendances dans les mécanismes moléculaires connus de la résistance aux antibiotiques tout en fournissant d'autres informations précieuses sur les isolats, comme la parenté génétique. Cependant, ce n'est qu'en combinant les tests de sensibilité phénotypique avec le WGS que la détection pré-coce de mécanismes de résistance inconnus sera possible.

Prevalence and whole genome-based phylogenetic, virulence and antibiotic -resistance characteristics of nasal -Staphylococcus aureus in healthy Swiss horses.

INTRODUCTION: A total of 100 nasal swabs were collected from healthy horses in Switzerland between January 2020 and August 2020. The samples were taken from horses at 40 different stables in 12 different cantons and screened for both methicillin-resistant (MRSA) and methicillin-susceptible S. aureus (MSSA) using selective agar plates. S. aureus were tested for antibiotic susceptibility by measurement of the minimal inhibitory concentration (MIC) and for virulence factors, antibiotic resistance genes and phylogenetic characteristics using whole genome sequence analysis. Ten horses were found to be positive (10 %, CI: 95 %, 0,0552 - 0,1744) for S. aureus, and four of them harboured MRSA (4 %, CI: 95 %, CI: 1,5 % - 9 %). The MRSA were detected in horses from three different stables in the same region of one canton and MSSA were detected in horses from five different cantons. All the MRSA isolates were genetically related (ST398-t011-IVa), while the MSSA were diverse (ST1-t127/t398/t1508, ST816-t1294, ST133-t1403, ST30-t012). MRSA showed resistance to penicillin (blaZ), cefoxitin (mecA), trimethoprim (dfrK), gentamicin, kanamycin (aac(6')-Ie - aph(2'')-Ia), and tetracycline (tet(M)). MSSA were resistant to either none or one of the antibiotics tested like penicillin (blaZ) and erythromycin (erm(T)). Virulence genes were more abundant in MSSA than in MRSA. This study provides first insight into the prevalence and type of S. aureus in healthy Swiss horses and reveals a source of strains, which may cause infections in both horses and humans.

INTRODUCTION: Au total, 100 écouvillons nasaux ont été prélevés sur des chevaux sains en Suisse entre janvier 2020 et août 2020. Les échantillons ont été prélevés sur des chevaux de 40 écuries différentes dans 12 cantons différents et ont été soumis à un dépistage de S. aureus résistant à la méthicilline (MRSA) et de S. aureus sensible à la méthicilline (MSSA) à l'aide de plaques de gélose sélectives. Les S. aureus ont été testés pour leur sensibilité aux antibiotiques en mesurant la concentration minimale inhibitrice (CMI) et pour les facteurs de virulence, les gènes de résistance aux antibiotiques et les caractéristiques phylogénétiques en analysant la séquence du génome entier. Dix chevaux se sont révélés positifs (10 %, IC: 95 %, 0,0552 ­ 0,1744) pour S. aureus, et quatre d'entre eux étaient porteurs de MRSA (4 %, IC: 95 %, IC: 1,5 % ­ 9 %). Les MRSA ont été détectés chez des chevaux provenant de trois écuries différentes de la même région d'un canton et les MSSA ont été détectés chez des chevaux provenant de cinq cantons différents. Tous les isolats de MRSA étaient génétiquement apparentés (ST398-t011-IVa), tandis que les MSSA étaient divers (ST1-t127/t398/t1508, ST816-t1294, ST133-t1403, ST30-t012). Les MRSA étaient résistants à la pénicilline (blaZ), à la céfoxitine (mecA), au triméthoprime (dfrK), à la gentamicine, à la kanamycine (aac(6')-Ie ­ aph(2")-Ia) et à la tétracycline (tet(M)). Les MSSA étaient résistants à aucun ou à un des antibiotiques testés soit à la pénicilline (blaZ) ou à l'érythromycine (erm(T)). Les gènes de virulence étaient plus abondants chez les MSSA que chez les MRSA. Cette étude donne, pour la première fois, un aperçu de la prévalence et du type de S. aureus chez les chevaux suisses en bonne santé et révèle la présence de souches susceptibles de provoquer des infections chez les chevaux et les humains.

Antimicrobial susceptibility in E. coli and Pasteurellaceae at the beginning and at the end of the fattening process in veal calves: Comparing 'outdoor veal calf' and conventional operations.

Animal husbandry requires practical measures to limit antimicrobial resistance (AMR). Therefore, a novel management and housing concept for veal calf fattening was implemented on 19 intervention farms (IF) and evaluated regarding its effects on AMR in Escherichia (E.) coli, Pasteurella (P.) multocida and Mannheimia (M.) haemolytica in comparison with 19 conventional control farms (CF). Treatment intensity (-80%) and mortality (-50%) were significantly lower in IF than in CF, however, production parameters did not differ significantly between groups. Rectal and nasopharyngeal swabs were taken at the beginning and the end of the fattening period. Susceptibility testing by determination of the minimum inhibitory concentration was performed on 5420 isolates. The presence of AMR was described as prevalence of resistant isolates (%), by calculating the Antimicrobial Resistance Index (ARI: number of resistance of one isolate to single drugs/total number of drugs tested), by the occurrence of pansusceptible isolates (susceptible to all tested drugs, ARI=0), and by calculating the prevalence of multidrug (≥3) resistant isolates (MDR). Before slaughter, odds for carrying pansusceptible E. coli were higher in IF than in CF (+65%, p=0.022), whereas ARI was lower (-16%, p=0.003), and MDR isolates were less prevalent (-65%, p=0.001). For P. multocida, odds for carrying pansusceptible isolates were higher in IF before slaughter compared to CF (+990%, p=0.009). No differences between IF and CF were seen regarding the prevalence of pansuceptible M. haemolytica. These findings indicate that easy-to-implement measures to improve calf management can lead to a limitation of AMR in Swiss veal fattening farms.

Distribution, genetic heterogeneity, and antimicrobial susceptibility of Brachyspira pilosicoli in Swiss pig herds.

Brachyspira (B.) pilosicoli is a bacterium causing porcine intestinal spirochaetosis, a disease characterized by diarrhoea and depressed growth rates especially in nursery and fattening pigs. Knowledge of the epidemiology and antimicrobial susceptibility of this pathogen is limited. Therefore, the aim of this study was to analyse the distribution, genetic heterogeneity, and antimicrobial susceptibility of B. pilosicoli field isolates from Swiss pig farms. Faecal swabs of 693 animals originating from 156 herds were analysed for the presence of Brachyspira spp. using culture and polymerase chain reaction identification. Further characterisation was performed using multilocus sequence typing (MLST) and broth dilution antimicrobial susceptibility testing. With 52.6% positive herds, B. pilosicoli could be frequently isolated from herds with animals suffering from diarrhoea. In herds with animals without clinical signs of diarrhoea, detection was significantly less frequent with only 10.5% positive herds (p 0.001). Among 80 isolates used for typing, genetic heterogeneity was observed with 44 different sequence types (ST) which often differed from herd to herd. No predominant ST was observed. More than 73.0% of the 41 B. pilosicoli isolates analysed, showed minimal inhibitory concentration values above the wild type cut-off values for lincomycin, tylvalosin and/ or tylosin. For tiamulin, valnemulin and doxycycline, this was the case in 48.8%, 43.9% and 36.6%, respectively. In conclusion, a diverse population of B. pilosicoli exhibited decreased susceptibility to antimicrobials used against Brachyspira infections. Monitoring of resistance in Brachyspira spp. is highly recommended to support targeted use of antimicrobials in pigs.

Prevalence and characterization of -methicillin-resistant Macrococcus spp. in food producing animals and meat in Switzerland in 2019.

INTRODUCTION: The prevalence of methicillin-resistant Macrococcus spp. in calves and pigs at slaughterhouses and in retail beef and pork meat was determined using samples taken in 2019 within the framework of the national monitoring of methicillin-resistant Staphylococcus aureus in food producing animals in Switzerland. The isolates were submitted to antimicrobial susceptibility testing of 19 antibiotics and to molecular techniques (e.g. PCR, microarray, WGS) for the identification of resistance genes, elements containing the methicillin resistance genes mec and sequence type (ST). Methicillin-resistant Macrococcus spp. (M. caseolyticus (n=38), M. bohemicus (n=4) and Macrococcus spp. (n=2)) were isolated in 40 of 299 nasal swabs from calves representing a prevalence of 13,38 % (95 % CI, 9,98 % - 17,70 %), and in four of 303 nasal swabs from pigs [1,32 % (95 % CI, 0,36 % - 3,35 %)]. One of 311 samples of Swiss pork meat contained a Macrococcus sp. [0,32 % (95 % CI, 0,01 % - 1,78 %)], and four of 309 beef meat samples (260 domestic and 49 imported) contained M. caseolyticus [1,29 % (95 % CI, 0,35 % - 3,28 %)]. The M. caseolyticus strains belonged to diverse STs, with ST21 being the most common in both pigs and calves. The mecD gene was located on Macrococcus resistance island mecD (McRImecD) in 42 strains and on staphylococcal cassette chromosome mec (SCCmecD) in three strains, while mecB was found on plasmids in four strains. In addition to resistance to ß-lactams, the strains also exhibited resistance to tetracycline (n=17; tet(L), tet(K), tet(M)), streptomycin (n=13; str, ant(6)-Ia, rpsL mutation [K56R in ribosomal protein S12]), kanamycin (n=10; aac(6')-Ie - aph(2'')-Ia, aph(2')-Ib, aph(2')-Ic, ant(4')-Ia), clindamycin (n=9; erm(B), erm(45)), erythromycin (n=9; erm(B), msr(G), erm(45)), fusidic acid (n=9; fusC) and gentamicin (n=1; aac(6')-Ie - aph(2'')-Ia). This study represents the first national prevalence study of methicillin-resistant Macrococcus spp. in pigs, calves, pork and beef meat in Switzerland and revealed a reservoir of genetically diverse strains carrying several resistance traits.

INTRODUCTION: La prévalence des macrococques résistants à la méthicilline chez les veaux et les porcs à l'abattoir et dans la viande de bœuf et de porc au détail a été déterminée à partir d'échantillons prélevés en 2019 dans le cadre du monitoring national des Staphylococcus aureus résistants à la méthicilline chez les animaux de rente en Suisse. Les isolats ont été soumis à des tests de sensibilité à 19 antibiotiques et à des techniques moléculaires (par exemple PCR, microarray, WGS) pour l'identification des gènes de résistance, des éléments contenant les gènes mec responsible de la résistance à la méthicilline, ainsi que du type de séquence (ST). Des macocoques résistants à la méthicilline (M. caseolyticus (n=38), M. bohemicus (n=4) et Macrococcus spp. (n=2)) ont été isolés dans 40 des 299 écouvillons nasaux de veaux, ce qui représente une prévalence de 13,38 % (IC 95 %, 9,98 % ­ 17,70 %), et dans quatre des 303 écouvillons nasaux de porcs [1,32 % (IC 95 %, 0,36 % ­ 3,35 %)]. Un des 311 échantillons de viande de porc suisse contenait un Macrococcus sp. [0,32 % (IC 95 %, 0,01 % ­ 1,78 %)], et quatre des 309 échantillons de viande de bœuf (260 domestiques et 49 importés) contenaient M. caseolyticus [1,29 % (IC 95 %, 0,35 % ­ 3,28 %)]. Les souches de M. caseolyticus appartenaient à divers ST, le ST21 étant le plus fréquent chez les porcs et les veaux. Le gène mecD a été localisé sur des éléments du chromosome McRImecD dans 42 souches et SCCmecD dans trois souches, tandis que le gène mecB se trouvait sur des plasmides dans quatre souches. En plus de la résistance aux ß-lactamines, les souches étaient également résistantes à la tétracycline (n=17 ; tet(L), tet(K), tet(M)), à la streptomycine (n=13 ; str, ant(6)-Ia, mutation rpsL [K56R dans la protéine ribosomale S12]), à la kanamycine (n=10 ; aac(6')-Ie ­ aph(2'')-Ia, aph(2')-Ib, aph(2')-Ic, ant(4')-Ia), clindamycine (n=9 ; erm(B), erm(45)), érythromycine (n=9 ; erm(B), msr(G), erm(45)), acide fusidique (n=9 ; fusC) et gentamicine (n=1 ; aac(6')-Ie ­ aph(2'')-Ia). Cette étude est la première à déterminer prévalence des Macrococcus spp. résistants à la méthicilline chez les porcs, les veaux, la viande de porc et de bœuf en Suisse et a révélé un réservoir de souches génétiquement diverses et portant plusieurs traits de résistance.

Effects of the novel concept 'outdoor veal calf' on antimicrobial use, mortality and weight gain in Switzerland.

The aim of the intervention study 'outdoor veal calf' was to evaluate a novel concept for calf fattening which aimed at reducing antimicrobial use without compromising animal health. Management practices such as commingling of calves from multiple birth farms, crowding, and suboptimal barn climate are responsible for high antimicrobial use and mortality in the veal calf population. The risk of selecting bacteria resistant to antimicrobials and of economic losses is accordingly elevated. The 'outdoor veal calf' concept, implemented in nineteen intervention farms (IF), is based on three main measures: 1. purchased calves are transported directly from neighboring birth farms to the fattening facility instead of commingling calves in livestock dealer trucks; 2. each calf is vaccinated against pneumonia after arrival and completes a three-week quarantine in an individual hutch; and 3. the calves spend the rest of the fattening period in outdoor hutches in groups not exceeding 10 calves. The covered and bedded paddock and the group hutches provide shelter from cold weather and direct sunshine, constant access to fresh air is warranted. Nineteen conventional calf fattening operations of similar size served as controls (CF). Every farm was visited once a month for a one-year period, and data regarding animal health, treatments, and production parameters were collected. Treatment intensity was assessed by use of the defined daily dose method (TIDDD in days per animal year), and calf mortality and daily weight gain were recorded in both farm groups. Mean TIDDD was 5.3-fold lower in IF compared to CF (5.9 ±â€¯6.5 vs. 31.5 ±â€¯27.4 days per animal year; p < 0.001). Mortality was 2.1-fold lower in IF than in CF (3.1% ± 2.3 vs. 6.3 % ± 4.9; p = 0.020). Average daily gain did not differ between groups (1.29 ±â€¯0.17 kg/day in IF vs. 1.35 ±â€¯0.16 kg/day in CF; p = 0.244). A drastic reduction in antimicrobial use and mortality was achieved in the novel 'outdoor veal calf' system without compromising animal health. The principles of risk reduction used in designing the system can be used to improve management and animal health, decrease the need for antimicrobial treatments and thus selection pressure on bacteria in veal operations.

Impact of the early-life skin microbiota on the development of canine atopic dermatitis in a high-risk breed birth cohort.

Canine atopic dermatitis (CAD) is a prevalent inflammatory skin disease of dogs worldwide. Certain breeds such as the West Highland White Terriers (WHWT) are predisposed to suffer from CAD. Microbial dysbiosis is known to play a significant role in the pathogenesis of the disease, which is similar to its human counterpart, atopic dermatitis (AD). To date, no large cohort-study has been conducted in a predisposed dog breed to study the impact of the early-life microbiota on the development of CAD, as well as the possible implication of factors such as hygiene and access to the outdoors. In this study skin samples of 143 WHWT, including 109 puppies up to three weeks old and 34 parent dogs, from 17 breeders, were subjected to 16S rRNA gene and ITS2 amplicon sequencing to disclose the bacterial and fungal oral and skin microbiota, respectively. The oral samples served as a control group to confirm differences between haired and mucosal surfaces. The cutaneous microbiota differed between sample sites and age of the dogs. The season of sampling, geographical origin as well as hygiene status of the household and the access to the outdoors shaped the skin microbiota of the puppies significantly. However, we found that the individual early-life microbiota did not predispose for the later development of CAD.

Antibiotic and quaternary ammonium compound resistance in Escherichia coli from calves at the beginning of the -fattening period in Switzerland (2017).

INTRODUCTION: In the Swiss veal calf production, antimicrobials and disinfectants are used to control bacterial infectious diseases, leading to a risk of selecting for a resistant bacterial population. While the prevalence of antibiotic resistance in E. coli from calves has been monitored at slaughterhouses in Switzerland since 2006, the resistance situation of E. coli from young calves entering the fattening period is not known. A total of 100 calves entering the fattening period in 20 geographically distant farms in Switzerland were screened for the presence of E. coli using rectal swabs in 2017. Genetic diversity between isolates was determined using repetitive palindromic Polymerase Chain Reaction (rep-PCR) revealing a genetically diverse E. coli population. Susceptibility to 13 antibiotics and to alkyldimethylbenzylammonium (ADBAC) was determined by the measurement of the minimal inhibitory concentration. Antibiotic and quaternary ammonium compound (QAC) resistance genes were identified using microarray and Polymerase Chain Reaction (PCR). Sixty-four percent of the isolates were resistant to at least one antibiotic, and 52% also exhibited decreased susceptibility to ADBAC. Resistance to more than 3 antibiotics was found in 40% of the isolates. Isolates exhibited resistance to tetracycline (57%) associated with the presence of tet genes (tet(A), (B), (E), (G)), to sulfonamides (61%) (sul1, sul2, sul3), ampicillin (56%) (blaTEM-1), trimethoprim (32%) (dfrA), phenicols (31%) (catA1, cmlA1, floR), gentamicin (27%) (ant(2")-Ia, aac(3)-IVa, aac(3)-VIa), and cefotaxime (2%) (blaCTX-M-14 (ESBL)). Mutations in GyrA (S83L) and ParC (S80I) were found in the fluoroquinolone-resistant isolates (6%). All isolates were susceptible to colistin, tigecycline and meropenem. No association between the presence of decreased susceptibility to ADBAC and qac genes was observed. In conclusion, antibiotic and QAC resistant E. coli are present in the gastrointestinal tract of young calves at the beginning of the fattening period, emphasizing the need for appropriate and reduced use of antibiotics and QAC-containing disinfectants in order to limit further selection of these bacteria during the fattening period.

INTRODUCTION: Dans la production de veaux en Suisses, des antimicrobiens et des désinfectants sont utilisés pour contrôler les maladies infectieuses bactériennes, ce qui entraîne un risque de sélection d'une population bactérienne résistante. Si la prévalence de la résistance de E. coli aux antibiotiques chez les veaux est surveillée dans les abattoirs suisses depuis 2006, la situation de la résistance de E. coli chez les jeunes veaux au début de la période d'engraissement n'est pas connue. Un total de 100 veaux entrant dans la période d'engraissement dans 20 exploitations géographiquement éloignées de Suisse ont été testés en 2017 pour détecter la présence de E. Coli à l'aide de prélèvements rectaux. La diversité génétique entre les isolats a été déterminée à l'aide de la réaction de polymérase en chaîne répétitive palindrome (rep-PCR) révélant une population de E.coli génétiquement diversifié. La sensibilité à 13 antibiotiques et au chlorure d'alkyldiméthylbenzylammonium (ADBAC) a été déterminée par la mesure de la concentration inhibitrice minimale. Les gènes de résistance aux antibiotiques et aux composés d'ammonium quaternaire (QAC) ont été identifiés à l'aide d'une puce à ADN et de la réaction de polymérase en chaîne (PCR). Soixante-quatre pour cent des isolats étaient résistants à au moins un antibiotique et 52% présentaient également une diminution de la sensibilité à l'ADBAC. Une résistance à plus de 3 antibiotiques a été trouvée dans 40% des isolats. Les isolats présentaient une résistance à la tétracycline (57%) associée à la présence de gènes tet (tet (A), (B), (E), (G)), aux sulfonamides (61%) (sul1, sul2, sul3), à l'ampicilline (56%) (blaTEM-1), au triméthoprime (32%) (dfrA), aux phénicols (31%) (catA1, cmlA1, floR), à la gentamicine (27%) (ant(2'')-Ia, aac (3) -IVa, aac (3) -VIa) et à la céfotaxime (2%) (blaCTX-M-14 (BLSE)). Les isolats résistants aux fluoroquinolones (6%) présentaient des mutations dans GyrA (S83L) et ParC (S80I). Tous les isolats étaient sensibles à la colistine, à la tigécycline et au méropénème. Aucune association entre la présence d'une sensibilité diminuée à l'ADBAC et les gènes qac n'a été observée. En conclusion, des E. coli résistants aux antibiotiques et aux QAC sont présents dans le tractus gastro-intestinal des jeunes veaux au début de la période d'engraissement, ce qui souligne la nécessité d'un usage approprié et réduit d'antibiotiques et de désinfectants contenant un QAC afin de limiter la sélection ultérieure de ces bactéries au cours de la période d'engraissement.

Multidrug resistance and multivirulence plasmids in enterotoxigenic and hybrid Shiga toxin-producing/enterotoxigenic Escherichia coli isolated from diarrheic pigs in Switzerland.

Enterovirulent Escherichia coli infections cause significant losses in the pig industry. However, information about the structures of the virulence and multidrug resistance (MDR) plasmids harboured by these strains is sparse. In this study, we used whole-genome sequencing with PacBio and Illumina platforms to analyse the molecular features of the multidrug-resistant enterotoxigenic E. coli (ETEC) strain 14OD0056 and the multidrug-resistant hybrid Shiga toxin-producing/enterotoxigenic E. coli (STEC/ETEC) strain 15OD0495 isolated from diarrheic pigs in Switzerland. Strain 14OD0056 possessed three virulence plasmids similar to others previously found in ETEC strains, while 15OD0495 harboured a 119-kb multivirulence IncFII/IncX1 hybrid STEC/ETEC plasmid (p15ODTXV) that co-carried virulence genes of both ETEC and STEC pathotypes, confirming the key role of plasmids in the emergence of hybrid pathotypes. All resistance genes of 14OD0056 that conferred resistance to ampicillin (blaTEM-1b), gentamicin (aac(3)-IIa), kanamycin (aph(3')-Ia), sulfonamide (sul1 and sul2), streptomycin (aph(3″)-Ib, aph(6)-Id), tetracycline (tet(B)) and trimethoprim (dfrA1) were identified on a single 207-kb conjugative MDR plasmid of incompatibility group (Inc) IncHI1/IncFIA (p14ODMR). Strain 15OD0495 carried two antimicrobial resistance plasmids (p15ODAR and p15ODMR). The 99-kb IncI1 plasmid p15ODAR harboured only aminoglycoside resistance genes (aac(3)-IIa, aph(3″)-Ib, aph(6)-Id, aph(4)-Ia), whilst the 49-kb IncN MDR plasmid p15ODMR carried genes conferring resistance to ampicillin (blaTEM-1b), sulfonamide (sul2), streptomycin (aph(6)-Id), tetracycline (tet(A)) and trimethoprim (dfrA14). Filter mating assays showed that p14ODMR, p15ODMR and p15ODAR were conjugative at room temperature and 37°C. The co-localization of multiple resistance genes on MDR conjugative plasmids such as p14ODMR and p15ODMR poses the risk of simultaneous selection of several resistance traits during empirical treatment. Thus, preventive strategies and targeted therapy following antibiotic susceptibility testing should be encouraged to avoid further dissemination of such plasmids.

Acinetobacter in veterinary medicine, with an emphasis on Acinetobacter baumannii.

Acinetobacter spp. are aerobic, rod-shaped, Gram-negative bacteria belonging to the Moraxellaceae family of the class Gammaproteobacteria and are considered ubiquitous organisms. Among them, Acinetobacter baumannii is the most clinically significant species with an extraordinary ability to accumulate antimicrobial resistance and to survive in the hospital environment. Recent reports indicate that A. baumannii has also evolved into a veterinary nosocomial pathogen. Although Acinetobacter spp. can be identified to species level using matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF/MS) coupled with an updated database, molecular techniques are still necessary for genotyping and determination of clonal lineages. It appears that the majority of infections due to A. baumannii in veterinary medicine are nosocomial. Such isolates have been associated with several types of infection such as canine pyoderma, feline necrotizing fasciitis, urinary tract infection, equine thrombophlebitis and lower respiratory tract infection, foal sepsis, pneumonia in mink, and cutaneous lesions in hybrid falcons. Given the potential multidrug resistance of A. baumannii, treatment of diseased animals is often supportive and should preferably be based on in vitro antimicrobial susceptibility testing results. It should be noted that animal isolates show high genetic diversity and are in general distinct in their sequence types and resistance patterns from those found in humans. However, it cannot be excluded that animals may occasionally play a role as a reservoir of A. baumannii. Thus, it is of importance to implement infection control measures in veterinary hospitals to avoid nosocomial outbreaks with multidrug-resistant A. baumannii.

Antimicrobial susceptibility patterns of blood culture isolates from foals in Switzerland.

INTRODUCTION: We report blood culture results of 43 foals admitted to an equine hospital for medical or surgical disorders and determine minimal inhibitory concentrations (MIC) of different antibiotics. Eleven foals had a positive blood culture result despite prior administration of antibiotics in 10 of these animals. MIC values above EUCAST and/or CLSI breakpoints were identified in coagulase-negative staphylococci, methicillin-resistant Staphylococcus aureus (MRSA) and Enterococcus faecium. Gram-negative isolates were less frequently identified and did not appear to exhibit increased MIC values. This study shows that bloodstream infections in foals in Switzerland are caused by diverse bacteria including Gram-positive bacteria which exhibit resistance to several classes of antibiotics.

INTRODUCTION: Nous rapportons les résultats d'hémoculture de 43 poulains admis dans un hôpital équin pour des affections médicales ou chirurgicales et déterminons les concentrations minimales inhibitrices (CMI) de différents antibiotiques. Le résultat de l'hémoculture a été positif pour onze poulains malgré l'administration préalable d'antibiotiques à 10 de ces animaux. Des valeurs de CMI supérieures aux seuils EUCAST et/ou CLSI ont été identifiées chez des staphylocoques coagulase négative, chez Staphylococcus aureus résistant à la méthicilline (MRSA) et chez Enterococcus faecium. Les isolats Gram négatifs étaient moins fréquemment identifiés et ne semblaient pas présenter de valeurs de CMI augmentées. Cette étude montre que les infections sanguines des poulains en Suisse sont causées par diverses bactéries, notamment des bactéries Gram positif, qui résistent à plusieurs classes d'antibiotiques.

Short communication: Lack of intramammary niche recolonization during a sanitation program for the contagious mastitis pathogen Staphylococcus aureus genotype B.

In Switzerland, sanitation programs of dairy herds infected with the contagious mastitis pathogen Staphylococcus aureus genotype B (GTB) have been established for several years. In recent years, Streptococcus uberis and non-aureus staphylococci have emerged as the bacteria most frequently isolated from bovine milk samples. The latter cause subclinical mastitis, and some species are more persistent or pathogenic than others. The present study aimed to investigate the developments in the intramammary colonization spectrum of 5 dairy herds undergoing a sanitation program for Staph. aureus GTB. We collected single-quarter milk samples aseptically from all lactating cows at 3-mo intervals during the sanitation period; after classical bacteriological analysis, MALDI-TOF mass spectrometry was used to identify the isolates to the species level. Non-aureus staphylococci were found to be the bacterial group most frequently occurring on the selected farms, with Staphylococcus chromogenes and Staphylococcus xylosus being predominant. The present study demonstrated that GTB-infected cows treated with antibiotics lacked systematic recolonization with other bacteria during herd sanitation for the contagious Staph. aureus GTB.

Isolation of Brachyspira hyodysenteriae from a crow (Corvus corone) in close proximity to commercial pigs.

The aim of this study was to determine whether crows (Corvus corone) can harbour Brachyspira hyodysenteriae, the cause of swine dysentery, and whether the organism carried by crows is related to strains infecting pigs. B. hyodysenteriae was isolated from one crow in close proximity to two pig farms in Switzerland. This isolate, along with five isolates of B. hyodysenteriae from one of the farms, belonged to sequence type (ST) 66 using multilocus sequence typing. This finding suggests that crows are potential vectors of B. hyodysenteriae, but further studies will be necessary to clarify the role of crows in the epidemiology of this organism.

Bacterial, fungal, parasitological and pathological analyses of abortions in small ruminants from 2012-2016.

INTRODUCTION: Abortion in small ruminants presents a clinical and economic problem with legal implications regarding animal health and zoonotic risk by some of the abortive pathogens. Several bacteria, fungi and parasites can cause abortion, but cost-orientated routine diagnostics only cover the most relevant epizootic agents. To cover a broad-range of common as well as underdiagnosed abortifacients, we studied 41 ovine and 36 caprine abortions by Stamp's modification of the Ziehl-Neelsen stain, culture for classical and opportunistic abortive agents, real-time PCR for C. burnetii, C. abortus, pathogenic Leptospira spp., Toxoplasma gondii and Neospora caninum. When the dam's serum was available detection of antibodies against B. melitensis, C. burnetii, C. abortus and Leptospira spp. was performed. In 37 cases sufficient placental tissue was available for pathological and histopathological examination. From the 77 cases 11 (14.3%) were positive by staining whereas real-time PCR detected C. burnetii and C. abortus in 49.3% and 32.5% of the cases. Antibodies against C. abortus and Leptospira spp. (33.3 and 26.7%) were detected. In 23.4% a bacterial culturable pathogen was isolated. Fungal abortion was confirmed in 1.3% of cases. A single abortive agent was identified in 44.2% of the cases and in 31.2% multiple possible abortifacients were present. Our study shows that the highest clarification rate can only be achieved by a combination of methods and evidences the role that multi-infections play as cause of abortion.

INTRODUCTION: Les avortements représentent un problème à la fois clinique et économique avec des conséquences en matière d'épizooties et un risque de zoonose pour certains agents. Diverses bactéries, champignons et parasites peuvent causer des avortements mais le diagnostic de routine, orienté sur les coûts, se concentre sur les principaux agents épizootiques. Afin d'avoir une vision large sur les agents d'avortements les plus fréquents et sur ceux qui sont sous-diagnostiqués, nous avons examinés 41 avortements de moutons et 36 de chèvres au moyen d'une coloration de Ziehl-Neelsen modifiée selon Stamp, de cultures ciblant les agents d'avortements classiques et opportunistes, d'une PCR en temps réel ciblant C. burnetii, C. abortus, les leptospires pathogènes, Toxoplasma gondii et Neospora caninum. Lorsque du sérum de la mère était disponible, nous avons procédé à une recherche d'anticorps contre B. melitensis, C. burnetii, C. abortus et Leptospira spp. Dans 37 cas, on disposait d'assez de tissu placentaire pour des examens pathologiques. Sur les 77 cas, 11 (14.3%) étaient positifs à la coloration alors que la PCR en temps réel démontrait la présence de C. burnetii et de C. abortus dans 49.3% respectivement 32.5% des cas. On a trouvé des anticorps contre C. abortus und Leptospira spp. dans 33.3% respectivement 26.7% des cas. Dans 23.4% des cas, on a pu mettre en évidence des pathogènes bactériens cultivables. Un avortement mycotique a été confirmé dans 1.3% des cas. Dans 44.2% des cas, un seul agent abortif était présent et dans 31.2% des cas, on trouvait plusieurs agents potentiels. Notre étude indique que le plus haut taux de diagnostic ne peut être atteint qu'en combinant diverses méthodes et montre le rôle possible de multi infections dans l'origine des avortements.

Pathotyping and antibiotic resistance of porcine enterovirulent Escherichia coli strains from Switzerland (2014-2015).

INTRODUCTION: A total of 131 porcine E. coli were isolated in 2014 and 2015 from the gut of 115 pigs raised in Switzerland and suffering from diarrhea. The isolates were tested for antibiotic resistance, serotypes, virulence factors and genetic diversity. Serotypes were assigned by agglutination tests and virulence genes were identified by polymerase chain reaction (PCR). Antibiotic resistance profile was determined by the measurement of the MIC of 14 antibiotics and by the detection of the corresponding genes using microarray and PCR approaches. Genetic diversity was determined by repetitive palindromic PCR (rep- PCR) revealing a heterogenous population. Half of the E. coli isolates possessing virulence factors could not be assigned to any of the 19 serotypes tested, but contained toxins and adhesins similarly to the sero-typable E. coli isolates. The most prevalent E. coli serotypes found were K88ac (18%), O139:K82 (6%), O141:K85ac (5%), O108:K`V189` (5%), O119:K`V113` (3%) and O157:K`V17` (2%). The combination of toxins EAST-1, STb and LT-I and adhesin F4 characterizing ETEC was the most frequent. The shigatoxin Stx2e (STEC) and intimin Eae (EPEC) were also detected, but less frequently. Seventy percent of the isolates were resistant to at least one antibiotic and 29% were resistant to more than 3 antibiotics. Isolates exhibited resistance to tetracycline (50%) associated to resistance genes tet(A), tet(B) and tet(C), sulfamethoxazole (49%) [sul1, sul2 and sul3], trimethoprim (34%) [dfr], nalidixic acid (29%), ampicillin (26%) [blaTEM-1], gentamicin (17%) [aac(3) -IIc, aac(3) -IVa and aac(3) -VIa], chloramphenicol (17%) [catAI and catAIII], and ciprofloxacin (8%) [mutations in GyrA (S83L) and ParC (S80I)]. All isolates were susceptible to 3rd generation cephalosporins, carbapenems, colistin and tigecycline. Pathogenic E. coli isolates from pigs in Switzerland could frequently not be assigned to a known serotype even if they contained diarrhea-causing virulence factors. They also harbor resistance mechanisms conferring resistance to antibiotics which are commonly used in pig husbandry, except for colistin. A careful identification of the causative agent and antibiotic susceptibility testing is highly recommended for targeted therapy and prudent use of antibiotics.

INTRODUCTION: Dans la cadre de cette étude, on a isolé, durant les années 2014 et 2015, 131 souches d'E.coli provenant des intestins de porcs suisses souffrant de diarrhée. Ces souches ont été testées quant à leurs résistances aux antibiotiques; en outre on a déterminé leurs sérotypes, leurs facteurs de virulence et leur diversité génétique. L'attribution des isolats d'E.coli aux divers sérogroupes a été réalisée au moyen d'une séro-agglutination et les facteurs de virulence ont été déterminés par PCR. Les profils de résistance aux antibiotiques ont été déterminés par la mesure des concentrations inhibitrices minimales de 14 antibiotiques. Les gènes qui y étaient associés ont été identifiés par puces à ADN et PCR. La diversité génétique a été déterminée au moyen de PCR répétitives palindromiques (rep PCR). La moitié des isolats d'E.coli qui possédaient des facteurs de virulence n'ont pas pu être classés dans un des 19 sérotypes testés, bien qu'ils aient possédé les mêmes toxines et adhésines que les isolats qu'il a été possible de typiser. Les sérotypes les plus fréquemment trouvés étaient (18%), O139:K82 (6%), O141:K85ac (5%), O108:K`V189` (5%), O119:K`V113` (3%) et O157:K`V17` (2%). La combinaison des toxines EAST-1, STb et LT-I et de l'adhésine F4, qui est caractéristique pour les E.coli entérotoxiques (ETEC), a été le plus fréquemment trouvée. La shigatoxine Stx2e (STEC) et l'Intimin Eae (EPEC) étaient par contre plutôt rares. Les résultats des rep PCR montraient une population hétérogène. 70% des isolats présentaient une résistance face à au moins un antibiotique et 29% étaient résistants à plus de 3 antibiotiques. Les isolats montraient des résistances vis-à-vis de la tétracycline (50%) associée avec les gènes de résistance tet(A), tet(B) et tet(C), du sulfaméthoxazole (49%) [sul1, sul2 et sul3], du trimethoprime (34%) [dfr], de l'acide nalidixique (29%), de l'ampicilline (26%) [blaTEM-1], de la gentamicine (17%) [aac(3) -IIc, aac(3) -IVa et aac(3) -VIa], du chloramphenicol (17%) [catAI and catAIII] et de la ciprofloxacine (8%) [mutations dans GyrA (S83L) et ParC (S80I)]. Tous les isolats étaient sensibles aux céphalosporines de troisième génération, au carbapénèmes, à la colistine et à la tigecycline. Les E.coli pathogènes des porcs en Suisse n'ont souvent pas pu être attribués aux sérogroupes connus, bien qu'ils possèdent les facteurs de virulence causant la diarrhée. En outre on a découvert divers mécanismes de résistance contre des antibiotiques qui sont régulièrement utilisés dans la production porcine. En conséquence, une identification rigoureuse et soignée des germes responsables de maladie et un testage de la sensibilité avant traitement est de première importance.

Quarter- and cow-level risk factors for intramammary infection with coagulase-negative staphylococci species in Swiss dairy cows.

Bacteriological status, evaluation of udder symmetry, udder hygiene, and teat end scores of 92 dairy cows were assessed on 3 Swiss dairy farms in a longitudinal 1-yr study to determine risk factors for intramammary infection (IMI) with coagulase-negative staphylococci (CNS) species. Farm visits were performed monthly including sterile quarter milk sampling and udder evaluation of all lactating cows. Milk samples were evaluated for the presence of staphylococci using selective agar plates. Species identification was performed using MALDI-TOF mass spectrometry. Intramammary infection was defined as milk samples having ≥100 cfu per mL of milk according to culture results. Overall, 3,151 quarter samples were included in the statistical analysis. Staphylococcus chromogenes, Staphylococcus haemolyticus, Staphylococcus xylosus, and a Staphylococcus warneri-like species were the 4 most prevalent CNS species found. Hierarchical multivariable logistic regression models were built to evaluate risk factors for species-specific CNS IMI. Risk factors for Staph. chromogenes IMI were presence in herd B, the period from June 2014 to August 2014 and December 2014 to February 2015, and presence of udder edema. For Staph. haemolyticus, the relevant risk factor included coinfection with Staph. xylosus coinfection with other than the above-mentioned CNS species ("others") and the period from June 2014 to November 2014. Coinfection with Staph. haemolyticus and "others," the periods from June 2014 to August 2014 and December 2014 to February 2015, early phase of lactation (1-60 d in milk), and belonging to herd B were significantly associated with Staph. xylosus IMI. Mid and late lactation, coinfection with Staph. xylosus, and the period September 2014 to May 2015 were identified as significant risk factors for Staph. warneri-like IMI. For Staph. chromogenes, 60.6 and 26% of the variance was observed at the quarter and cow level, respectively, whereas for the other investigated species the highest variance was observed at the sample level. The predominant species within herds differed and was most pronounced for the Staph. warneri-like species.

[Bacterial resistance in bacteria isolated from the nasal cavity of Swiss dairy calves]. / Resistenzen bei Bakterienisolaten aus der Nase von Aufzuchtkälbern in Milchviehbetrieben.

INTRODUCTION: Fifty-six E. coli, 37 P. multocida und 8 M. haemolytica were isolated from 157 nasal swabs taken from calves in 52 dairy herds. The antibiotic susceptibility of the organisms was determined by measurement of the minimal inhibitory concentrations. Of the 56 E. coli isolates, 55.3% exhibited resistance to tetracyclines, 55.3% to sulfonamides, 39.3% to beta-lactams, 30.3% to aminoglycosides, 8.9% to fluorochinolones and 3.5% to 3rd generation cephalosporins. The 3rd generation cephalosporin- resistant isolates contained the extended spectrum-beta-lactamase gene blaCTX-M-14 and came from 2 farms where the milk of cows under antimicrobial treatment was fed to the calves and mastitis was treated with cefquinome as first line therapy. Of the 37 P. multocida isolates, 48.6% exhibited resistance to tetracyclines, 16.2% to beta-lactams, and 5.4% each to macrolides, aminoglycosides and sulfonamides. The 8 M. haemolytica isolates showed no resistances against the tested antibiotics.

INTRODUCTION: Dans 52 exploitations laitières, 157 écouvillons ont été prélevés des fosses nasales de veaux d'élevage, desquels ont été isolés 56 Escherichia coli, 37 Pasteurella multocida et 8 Mannheimia haemolytica. Les profils de résistance de ces germes ont été déterminés par la mesure de la concentration minimale inhibitrice. Pour les E. coli, 55.3% des souches isolées montraient une résistance aux tétracyclines, 55.3% aux sulfamidés, 39.3% aux béta-lactamines, 30.3% aux aminoglycosides, 8.9% aux fluoroquinolones et 3.5% aux céphalosporines de 3ème génération. Les souches résistantes aux céphalosporines de 3ème génération présentaient un gène de béta-lactamase à spectre étendu blaCTX-M-14 et provenaient de deux exploitations dans lesquelles le lait de vaches sous traitement antibiotique était distribué aux veaux et où le cefquinome était utilisé comme antibiotique de premier recours pour le traitement des mammites. Pour P. multocida, 48.6% des couches étaient résistantes aux tétracyclines, 16.2% aux béta-lactamines et 5.4% chacun aux macrolides, aux aminoglycosides et aux sulfamidés. Les 8 isolats de M. haemolytica étaient sensibles à tous les antibiotiques testés.

Methicillin-resistant Staphylococcus aureus isolated from dogs and cats in Switzerland.

INTRODUCTION: Twenty-two methicillin-resistant Staphylococcus aureus (MRSA) strains isolated from various infected locations in domestic cats and dogs between June 2008 and September 2014 were analyzed for their genotype, genetic fingerprint, virulence and antibiotic resistance profile. Eighteen strains belonged to the clonal complex (CC) 22 [ST22(MLST)-A(PFGE)-t032(spa)-IV(SCCmec) and ST22-A-t1214-IV], 2 strains to the livestock associated MRSA ST398-t011-IV and two were individual strains of ST5-t002-II and ST1-t001-IV. They contained virulence factors such as γ-hemolysins, ß-hemolysin converting phage genes, leukocidins and enterotoxins. Most widespread resistances were observed against ß-lactams, trimethoprim and fluoroquinolones, but single strains also exhibited resistance to macrolides, lincosamides, aminoglycosides, tetracycline, chloramphenicol and/or mupirocin. The predominant presence of CC22 MRSA strongly indicates clonal spread of a human associated lineage in Swiss companion animals. It is therefore of public health importance to maintain a low level of MRSA infections in animals to avoid uncontrolled dissemination of MRSA clones in humans and animals.

INTRODUCTION: Entre juin 2008 et septembre 2014, 220 souches de Staphylococcus aureus résistantes à la méthicilline (SARM) ont été isolées de divers foyers infectieux chez des chiens et des chats et leur génotype, leur virotype et leur profil de résistance aux antibiotiques ont été déterminés. Le complexe clonal (CC) 22 [ST22(MLST)-A(PFGE)-t032(spa)-IV(SCCmec) et ST22-A-t1214-IV] était le type de SARM principal, suivi par le CC398 (ST398-D-t011-IV) normalement associé aux animaux de rente ainsi que par quelques autres souche de SARM (ST5-C-t002-II, ST1-B-t001-IV). Les types de SARM de cette étude avaient un type de virulence relativement conservé, marqué par des gènes associés à la γ-hémolysine, des gènes d'entérotoxines, des gènes de virulence sur les phages convertissant la ß-hémolysin et des gènes de leucocidine. Les souches étaient en outre principalement résistantes au ß-lactame, au triméthoprime et aux fluoroquinolones, certaines d'entre elles présentant des résistances supplémentaires aux macrolides, aux lincosamides, aux aminoglycosides, aux tétracyclines, au chloramphénicol et/ou à la mupirocine. La fréquence des infections avec des SARM du complexe clonal CC22 indique clairement que ce clone associé à l'être humain peut se répandre facilement dans la population suisse d'animaux de compagnie. Pour réduire cette expansion de SARM chez les hommes et les animaux, il faut chercher à limiter le plus possible les infections dues aux SARM chez les animaux de compagnie.

Short communication: Role of Streptococcus pluranimalium in Mediterranean buffaloes (Bubalus bubalis) with different udder health statuses.

The aims of the current study were to describe presence and clinical role over time of Streptococcus pluranimalium isolated in milk samples of Mediterranean buffalo (MB). Two hundred composite milk samples originating from 40 primiparous MB were collected at 10, 30, 60, 90, and 150d in milk (DIM) and from 20 pluriparous MB at 77 to 120 DIM. Milk samples were used for analysis of somatic cell counts, bacteriological cultures, and identification (matrix-assisted laser desorption/ionization time-of-flight mass spectrometry). Nine of 200 (4.5%) samples of primiparous MB and 3 of 20 (15%) samples of pluriparous MB were positive for Strep. pluranimalium. The prevalence of the bacterium in primipari was 0% (0/40) at 10, 30, and 150 DIM, whereas it was 5 (2/40) and 17.5% (7/40) at 60 and 90 DIM, respectively. Eight primipari were positive only once, whereas 1 was positive at 2 different samplings. Mono-infection was not detected in any of the age categories or udder health status. Infections were transient in primipari. Clinical mastitis was observed in primipari once at 90 DIM, subclinical mastitis detected twice in the same animals at 60 and 90 DIM, and intramammary infections were diagnosed 1 and 5 times at 60 and 90 DIM in primipari, respectively, whereas 3 infections were diagnosed in pluripari. The clinical reflections demonstrate for the first time the presence of Strep. pluranimalium in MB and its association with different udder health status. Nevertheless, it cannot be excluded that the bacterium may simply follow a pattern of commensal or opportunistic behavior, taking advantage of a preexisting bacterial udder infection.