Multilocus sequence typing of Campylobacter jejuni and Campylobacter coli to identify potential sources of colonization in commercial turkey farms.

Poultry are the main reservoir for thermophilic Campylobacter spp., which is the most common causative agent of human bacterial gastroenteritis. The epidemiology of Campylobacter in poultry, particularly in turkeys, is not completely understood. This study aimed at identifying potential sources and transmission routes of thermophilic Campylobacter spp. in commercial turkey farms. C. jejuni and C. coli isolates from breeders (n = 29, 20 C. jejuni and 9 C. coli) and their progeny (n = 51, 18 C. jejuni and 33 C. coli) reared in two different farms for three sequential production cycles were analysed by multilocus sequence typing (MLST). Strains (n = 88, 42 C. jejuni and 46 C. coli) isolated from environmental (i.e. anteroom and in-house overshoes), water (i.e. drinkers and water line), and pest (i.e. flies, Alphitobius diaperinus, and mice) sources were also examined. MLST of C. jejuni and C. coli isolates resulted in 13 and 12 different sequence types (STs) belonging to six and one previously-described clonal complexes (CCs), respectively. Three novel STs were identified. Genetic similarities were detected between isolates from fattening turkeys and the considered environmental, water, and pest sources, and with the breeders to a lesser extent. Source attribution analysis estimated that environmental and water sources accounted for most (∼75%) of fattening turkey isolates and were therefore identified as the most likely sources of flock colonization, followed by pests (∼20%) and breeders (∼5%). These sources may thus be targeted by control measures to mitigate the risk of Campylobacter colonization in commercial turkeys. RESEARCH HIGHLIGHTS High occurrence of C. jejuni and C. coli in commercial turkey flocks. High genetic diversity of C. jejuni and C. coli in commercial turkey flocks. Horizontal transmission responsible for Campylobacter colonization of commercial turkey flocks. Environmental and water sources involved in Campylobacter colonization of commercial turkey flocks. Strategies for prevention and control of Campylobacter colonization of commercial turkey flocks are needed.

Class 1 and class 2 integrons in avian pathogenic Escherichia coli from poultry in Italy.

The aim of this study was to investigate the occurrence of class 1 and 2 integrons in avian pathogenic Escherichia coli (APEC) from poultry in northern Italy. Strains were tested for phenotypic resistance to aminoglycosides and sulphonamides, and the association between the presence of integrons and the resistance to these antimicrobials was evaluated. A total of 299 isolates (158 from turkeys, 110 from broilers, and 31 from layer hens) were collected from 200 industrial farms. Antimicrobial susceptibility test by the disk diffusion method was performed in accordance with the Clinical and Laboratory Standards Institute (CLSI) guidelines. All strains were screened for the presence of class 1 and 2 integrons by PCR and sequencing. About 55% of APEC contained integrons (class 1, 49.8%; class 2, 10.4%). Different variants of the aadA (5 variants) and the dfrA (4 variants) genes, encoding for streptomycin and trimethoprim resistance respectively, were detected in integron-positive isolates. Less common gene cassettes, such as sat, estX, and orfF, were also identified. Fifteen and 4 gene cassette arrays were found among class 1 and 2 integrons, respectively. High levels of resistance were observed for triple sulphonamides (79.3%), streptomycin (67.2%), and sulfamethoxazole combined with trimethoprim (62.2%), whereas resistance against gentamycin (16.7%), kanamycin (14.7%), and apramycin 3.0%) was low. Integron positivity was significantly higher in isolates phenotypically resistant to aminoglycosides (63.6% vs. 37.8%, P<0.001) and sulfonamides (64.1% vs. 21.1%, P<0.001) than in susceptible ones. Integron-borne aminoglycoside and sulfonamide resistance in APEC represents a concern for the poultry industry in Italy, since they are among the most commonly used antimicrobials in poultry therapy.

Multilocus sequence typing of Campylobacter jejuni and Campylobacter coli from humans and chickens in North-Eastern Italy.

This paper reports the multilocus sequence typing (MLST) of 57 C. jejuni and C. coli isolates from humans and chickens in Italy and the identification of 17 new sequence types (STs). A high genetic diversity was detected among C. jejuni/C. coli and human/chicken isolates, with a predominance of clonal complexes CC21 and CC828. Although human STs were not the same as those found in chickens, 3 CCs overlapped between human and chicken isolates. Genotyping of Campylobacter strains by MLST should be encouraged in order to implement surveillance and con- trol of infection in humans and in animal reservoirs in Italy.

A longitudinal study on thermophilic Campylobacter spp. in commercial turkey flocks in northern Italy: occurrence and genetic diversity.

Poultry are recognized as a main reservoir of thermophilic campylobacters, but few studies have been carried out on commercial meat turkeys. This study was aimed at assessing the occurrence of thermophilic Campylobacter spp., their genetic diversity, and the trend of the infection during the whole production cycle of three turkey flocks from different farms in Northern Italy. Flocks were monitored from the time of housing 1-day-old poults to slaughter time by collecting samples (meconium and cloacal swabs) at weekly intervals up to the recovery of Campylobacter spp. and then twice a month. A conventional culture method and a multiplex PCR assay were used for Campylobacter detection and identification. A subset of isolates was genetically characterized by random amplified polymorphic DNA-PCR (RAPD-PCR) and flagellin gene A short variable region (flaA-SVR) sequencing. Although at different times, all flocks became colonized by Campylobacter jejuni or Campylobacter coli (or both) that persisted throughout the entire production cycle. Overall, nine RAPD types and 14 flaA-SVR types were detected with differences in their distribution among flocks and sampling times. Moreover, changes in the Campylobacter genotypes colonizing turkeys were observed over time within each flock. These findings suggest that Italian commercial turkeys might be widely colonized by different genotypes of C. jejuni and C. coli and also suggest that differences in the distribution and epidemiologic dynamics of these microorganisms might occur among flocks.

Antimicrobial resistance of Campylobacter jejuni and Campylobacter coli from poultry in Italy.

This study was aimed at assessing the antimicrobial resistance (AMR) of Campylobacter isolates from broilers and turkeys reared in industrial farms in Northern Italy, given the public health concern represented by resistant campylobacters in food-producing animals and the paucity of data about this topic in our country. Thirty-six Campylobacter jejuni and 24 Campylobacter coli isolated from broilers and 68 C. jejuni and 32 C. coli from turkeys were tested by disk diffusion for their susceptibility to apramycin, gentamicin, streptomycin, cephalothin, cefotaxime, ceftiofur, cefuroxime, ampicillin, amoxicillin+clavulanic acid, nalidixic acid, flumequine, enrofloxacin, ciprofloxacin, erythromycin, tilmicosin, tylosin, tiamulin, clindamycin, tetracycline, sulfamethoxazole+trimethoprim, chloramphenicol. Depending on the drug, breakpoints provided by Comité de l'antibiogramme de la Société Française de Microbiologie, Clinical and Laboratory Standards Institute, and the manufacturer were followed. All broiler strains and 92% turkey strains were multidrug resistant. Very high resistance rates were detected for quinolones, tetracycline, and sulfamethoxazole+trimethoprim, ranging from 65% to 100% in broilers and from 74% to 96% in turkeys. Prevalence of resistance was observed also against ampicillin (97% in broilers, 88% in turkeys) and at least three cephalosporins (93-100% in broilers, 100% in turkeys). Conversely, no isolates showed resistance to chloramphenicol and tiamulin. Susceptibility prevailed for amoxicillin+clavulanic acid and aminoglycosides in both poultry species, and for macrolides and clindamycin among turkey strains and among C. jejuni from broilers, whereas most C. coli strains from broilers (87.5%) were resistant. Other differences between C. jejuni and C. coli were observed markedly in broiler isolates, with the overall predominance of resistance in C. coli compared to C. jejuni. This study provides updates and novel data on the AMR of broiler and turkey campylobacters in Italy, revealing the occurrence of high resistance to several antimicrobials, especially key drugs for the treatment of human campylobacteriosis, representing a potential risk for public health.

High prevalence of oqxAB in Escherichia coli isolates from domestic and wild lagomorphs in Italy.

This study aimed to identify and characterize class 1 and 2 integrons and plasmid-mediated quinolones resistance (PMQR) genes in a collection of 113 multidrug resistance (MDR) Escherichia coli isolated from farm and wild lagomorphs between 2006 and 2008 in Northern Italy. Strains were examined for antimicrobial susceptibility by agar disk diffusion method and E-test for colistin (COL); integrons and gene cassettes content by real-time polymerase chain reaction (PCR) and DNA sequencing; PMQR genes by PCR and DNA sequencing; clonal relatedness by multilocus sequence typing; and plasmids by PCR-based replicon typing. Class 1 integrons were detected in 69 isolates (47 farm rabbits, 14 wild rabbits, and 8 wild hares). No class 2 integrons were found. Five different gene cassettes arrays were identified (aadA1, dfrA1-aadA1, orf in682-dhfrA5, orf in682-dfrA5-orfD ins21, and dfrA17-aadA5). Fifteen percent (17/113) of isolates carried oqxAB, no other PMQR determinants. All but one oqxAB-positive E. coli strains were recovered from farm rabbits. Seven out of 17 strains were associated with the predominant ST238 and carried from three to six different plasmid types, such as IncF, IncHI1, IncI1, IncN, IncP, IncX1, IncY, and ColE. COL resistance was identified in 6/113 strains (5.3%). This study provides new insights on the resistance phenotypes and the prevalence and dissemination of oqxAB in E. coli from farm and wild lagomorphs, suggesting that these animals may be reservoir of these genetic determinants in Italy and thus a potential source of PMQR E. coli for humans. PMQR mediated by oqxAB has not been detected in farm and wild lagomorphs before.