Clonal population structure and specific genotypes of multidrug-resistant Campylobacter coli from Turkeys

Commercial turkey flocks in North Carolina have been found to be colonized frequently with Campylobacter coli strains that are resistant to several antimicrobials (tetracycline, streptomycin, erythromycin, kanamycin, and ciprofloxacin/nalidixic acid). Such strains have been designated multidrug resistant (MDR). However, the population structure of MDR C. coli from turkeys remains poorly characterized. In this study, an analysis of multilocus sequence typing (MLST)-based sequence types (STs) of 59 MDR strains from turkeys revealed that the majority of these strains corresponded to one of 14 different STs, with three STs accounting for 41 (69%) of the strains. The major STs were turkey specific, and most (87%) of the strains with these STs were resistant to the entire panel of antibiotics mentioned above. Some (13%) of the strains with these STs were susceptible to just one or two of the antibiotics in this panel. Further subtyping using fla typing and pulsed-field gel electrophoresis with SmaI and KpnI revealed that the major MDR STs corresponded to strains of related but distinct subtypes, providing evidence for genomic diversification within these STs. These findings suggest that MDR strains of C. coli from turkeys have a clonal population structure characterized by the presence of a relatively small number of clonal groups that appear to be disseminated in the turkey production system. In addition, the observed correlation between STs and the MDR profiles of the microbes indicates that MLST-based typing holds potential for source-tracking applications specific to the animal source (turkeys) and the antimicrobial resistance profile (MDR status) of C. coli.     

Antimicrobial Susceptibility Profiles and Molecular Typing of Campylobacter jejuni and Campylobacter coli Isolates from Ducks in South Korea

Campylobacter is a food-borne zoonotic pathogen that causes human gastroenteritis worldwide. Campylobacter bacteria are commensal in the intestines of many food production animals, including ducks and chickens. The objective of the study was to determine the prevalence of Campylobacter species in domestic ducks, and the agar dilution method was used to determine resistance of the isolates to eight antibiotics. In addition, multilocus sequence typing (MLST) was performed to determine the sequence types (STs) of selected Campylobacter isolates. Between May and September 2012, 58 duck farms were analyzed, and 56 (96.6%) were positive for Campylobacter. Among the isolates, 82.1% were Campylobacter jejuni, 16.1% were C. coli, and one was unidentified by PCR. Of the 46 C. jejuni isolates, 87.0%, 10.9%, and 21.7% were resistant to ciprofloxacin, erythromycin, and azithromycin, respectively. Among the C. coli isolates, all 9 strains were resistant to ampicillin, and 77.8% and 33.3% were resistant to ciprofloxacin and azithromycin, respectively. The majority of the Campylobacter isolates were classified as multidrug resistant. Twenty-eight STs were identified, including 20 STs for C. jejuni and 8 STs for C. coli. The most common clonal complexes in C. jejuni were the ST-21 complex and the ST-45 complex, while the ST-828 complex predominated in C. coli. The majority of isolates were of STs noted in ducks and humans from earlier studies, along with seven STs previously associated only with human disease. These STs overlapped between duck and human isolates, indicating that Campylobacter isolates from ducks should be considered potential sources of human infection.     

Multilocus Sequence Typing and Virulence Profiles in Uropathogenic Escherichia coli Isolated from Cats in the United States

The population structure, virulence, and antimicrobial resistance of uropathogenic E. coli (UPEC) from cats are rarely characterized. The aim of this study was to compare and characterize the UPEC isolated from cats in four geographic regions of USA in terms of their multilocus sequence typing (MLST), virulence profiles, clinical signs, antimicrobial resistance and phylogenetic grouping. The results showed that a total of 74 E. coli isolates were typed to 40 sequence types with 10 being novel. The most frequent phylogenetic group was B2 (n = 57). The most frequent sequence types were ST73 (n = 12) and ST83 (n = 6), ST73 was represented by four multidrug resistant (MDR) and eight non-multidrug resistant (SDR) isolates, and ST83 were significantly more likely to exhibit no drug resistant (NDR) isolates carrying the highest number of virulence genes. Additionally, MDR isolates were more diverse, and followed by SDR and NDR isolates in regards to the distribution of the STs. afa/draBC was the most prevalent among the 29 virulence-associated genes. Linking virulence profile and antimicrobial resistance, the majority of virulence-associated genes tested were more prevalent in NDR isolates, and followed by SDR and MDR isolates. Twenty (50%) MLST types in this study have previously been associated with human isolates, suggesting that these STs are potentially zoonotic. Our data enhanced the understanding of E. coli population structure and virulence association from cats. The diverse and various combinations of virulence-associated genes implied that the infection control may be challenging.     

Organic Turkey Flocks: A Reservoir of Streptococcus gallolyticus subspecies gallolyticus

Streptococcus gallolyticus subspecies gallolyticus (S. gallolyticus) can colonise the gastrointestinal tract of humans and animals and is known to cause similar infections in both humans and animals. Data about the spread or prevalence in farm animals are missing. In this study, Trypton Soya Agar was modified to a selective medium enabling the isolation and quantification of S. gallolyticus from faecal samples. The bacterium was observed in 82 out of 91 faecal samples obtained from 18 different organic turkey flocks. The prevalence of shedding birds was estimated by the number of positive fresh droppings and reached up to 100% on most farms. Furthermore, for the first time S. gallolyticus was quantified in faeces from poultry flocks. The median of colony forming units (CFU) per gramme faeces was 3.6 x 10⁵CFU/g. Typing of one isolate from each positive faecal sample by multilocus sequence typing delivered 24 sequence types (STs). Most of the isolates belonged to the clonal complex CC58. The same STs of this complex were detected in up to six different flocks. Partly, these flocks were located in various regions and stocked with varying breeding lines. Regarding the biochemical profiles of the same STs from different farms, the results did not contradict a spread of specific STs in the organic turkey production. Moreover, checking the pubMLST database revealed that STs found in this study were also found in other animal species and in humans. The high detection rate and the number of S. gallolyticus in turkey faeces indicate that this bacterium probably belongs to the common microbiota of the gastrointestinal tract of turkeys from organic flocks. Furthermore, the findings of this study support the suggestion of a possible interspecies transmission.     

Antimicrobial Susceptibility Profiles and Strain Type Diversity of Campylobacter jejuni Isolates from Turkeys in Eastern North Carolina

Campylobacter jejuni is one of the most common bacterial causes of human gastroenteritis, and recent findings suggest that turkeys are an important reservoir for this organism. In this study, 80 C. jejuni isolates from eastern North Carolina were characterized for resistance to nine antimicrobials, and strain types were determined by fla typing, pulsed-field gel electrophoresis (PFGE) with SmaI and KpnI, and (for 41 isolates) multilocus sequence typing (MLST). PFGE analysis suggested that many of the isolates (37/40 [ca. 93%]) in a major genomic cluster had DNA that was partially methylated at SmaI sites. Furthermore, 12/40 (30%) of the isolates in this cluster were completely resistant to digestion by KpnI, suggesting methylation at KpnI sites. MLST of 41 isolates identified 10 sequence types (STs), of which 4 were new. Three STs (ST-1839, ST-2132 and the new ST-2934) were predominant and were detected among isolates from different farms. The majority of the isolates (74%) were resistant to three or more antimicrobials, and resistance to ciprofloxacin was common (64%), whereas resistance to the other drug of choice for treatment of human campylobacteriosis, erythromycin, was never encountered. Most (33/34) of the kanamycin-resistant isolates were also resistant to tetracycline; however, only ca. 50% of the tetracycline-resistant isolates were also kanamycin resistant. Isolates with certain antimicrobial resistance profiles had identical or closely related strain types. Overall, the findings suggest dissemination of certain clonal groups of C. jejuni isolates in the turkey production industry of this region.     

Phylogenetic Analysis Reveals Common Antimicrobial Resistant Campylobacter coli Population in Antimicrobial-Free (ABF) and Commercial Swine Systems

The objective of this study was to compare the population biology of antimicrobial resistant (AR) Campylobacter coli isolated from swine reared in the conventional and antimicrobial-free (ABF) swine production systems at farm, slaughter and environment. A total of 200 C. coli isolates selected from fecal, environmental, and carcass samples of ABF (n = 100) and conventional (n = 100) swine production systems were typed by multilocus sequence typing (MLST). Sequence data from seven housekeeping genes was analyzed for the identification of allelic profiles, sequence types (STs) and clonal complex determination. Phylogenetic trees were generated to establish the relationships between the genotyped isolates. A total of 51 STs were detected including two novel alleles (glnA 424 and glyA 464) and 14 novel STs reported for the first time. The majority of the C. coli isolates belonged to ST-854 (ABF: 31, conventional: 17), and were grouped in clonal complex ST-828 (ABF: 68%, conventional: 66%). The mean genetic diversity (H) for the ABF (0.3963+/−0.0806) and conventional (0.4655+/−0.0714) systems were similar. The index of association () for the ABF ( = 0.1513) and conventional ( = 0.0991) C. coli populations were close to linkage equilibrium, indicative of a freely recombining population. Identical STs were detected between the pigs and their environment both at farm and slaughter. A minimum spanning tree revealed the close clustering of C. coli STs that originated from swine and carcass with those from the environment. In conclusion, our study reveals a genotypic diverse C. coli population that shares a common ancestry in the conventional and ABF swine production systems. This could potentially explain the high prevalence of antimicrobial resistant C. coli in the ABF system in the absence of antimicrobial selection pressure.     

Evaluation of the Potential Antimicrobial Resistance Transfer from a Multi-Drug Resistant Escherichia coli to Salmonella in Dairy Calves

Previous research conducted in our laboratory found a significant prevalence of multi-drug resistant (MDR) Salmonella and MDR Escherichia coli (MDR EC) in dairy calves and suggests that the MDR EC population may be an important reservoir for resistance elements that could potentially transfer to Salmonella. Therefore, the objective of the current research was to determine if resistance transfers from MDR EC to susceptible strains of inoculated Salmonella. The experiment utilized Holstein calves (approximately 3 weeks old) naturally colonized with MDR EC and fecal culture negative for Salmonella. Fecal samples were collected for culture of Salmonella and MDR EC throughout the experiment following experimental inoculation with the susceptible Salmonella strains. Results initially suggested that resistance did transfer from the MDR E. coli to the inoculated strains of Salmonella, with these stains demonstrating resistance to multiple antibiotics following in vivo exposure to MDR EC. However, serogrouping and serotyping results from a portion of the Salmonella isolates recovered from the calves post-challenge, identified two new strains of Salmonella; therefore transfer of resistance was not demonstrated under these experimental conditions.     

Frequent Combination of Antimicrobial Multiresistance and Extraintestinal Pathogenicity in Escherichia coli Isolates from Urban Rats (Rattus norvegicus) in Berlin,Germany

Urban rats present a global public health concern as they are considered a reservoir and vector of zoonotic pathogens, including Escherichia coli. In view of the increasing emergence of antimicrobial resistant E. coli strains and the on-going discussion about environmental reservoirs, we intended to analyse whether urban rats might be a potential source of putatively zoonotic E. coli combining resistance and virulence. For that, we took fecal samples from 87 brown rats (Rattus norvegicus) and tested at least three E. coli colonies from each animal. Thirty two of these E. coli strains were pre-selected from a total of 211 non-duplicate isolates based on their phenotypic resistance to at least three antimicrobial classes, thus fulfilling the definition of multiresistance. As determined by multilocus sequence typing (MLST), these 32 strains belonged to 24 different sequence types (STs), indicating a high phylogenetic diversity. We identified STs, which frequently occur among extraintestinal pathogenic E. coli (ExPEC), such as STs 95, 131, 70, 428, and 127. Also, the detection of a number of typical virulence genes confirmed that the rats tested carried ExPEC-like strains. In particular, the finding of an Extended-spectrum beta-lactamase (ESBL)-producing strain which belongs to a highly virulent, so far mainly human- and avian-restricted ExPEC lineage (ST95), which expresses a serogroup linked with invasive strains (O18:NM:K1), and finally, which produces an ESBL-type frequently identified among human strains (CTX-M-9), pointed towards the important role, urban rats might play in the transmission of multiresistant and virulent E. coli strains. Indeed, using a chicken infection model, this strain showed a high in vivo pathogenicity. Imagining the high numbers of urban rats living worldwide, the way to the transmission of putatively zoonotic, multiresistant, and virulent strains might not be far ahead. The unforeseeable consequences of such an emerging public health threat need careful consideration in the future.     

DNA identification and characterization of Campylobacter jejuni and Campylobacter coli isolated from caecal samples of chickens in Grenada

Aims: To speciate Campylobacter strains from the caeca of chickens in Grenada using PCR and to evaluate DNA‐based typing methods for the characterization of these isolates. Methods and Results: Isolates were speciated with two multiplex PCR assays and were typed with flaA‐RFLP, pulsed‐field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Results confirmed that Campylobacter coli strains were more predominant than Campylobacter jejuni strains. From 56 isolates, 18 were misidentified using biochemical tests. PFGE typing gave the highest discriminatory power among the methods used (Simpson’s index of diversity, D = 0·9061). However, the combination of flaA‐RFLP, PFGE and MLST results gave the highest discrimination for subtyping of these isolates (D = 0·9857). A band position tolerance of 4% in Bio Numerics was the most appropriate for the analysis of this database. MLST profiles were generally concordant with PFGE and/or flaA‐RFLP types. Several isolates exhibited new MLST sequence types (STs), and 43 of the 49 Camp. coli strains belonged to the ST‐828 clonal complex. Conclusions: Campylobacter coli was the most prevalent species isolated from broilers and layers in Grenada, and a combination of restriction and sequence methods was most appropriate for the typing of Camp. coli isolates. Campylobacter coli STs clustered with described poultry‐associated Camp. coli STs by phylogenetic analysis. Significance and Impact of the Study: Further studies to understand the predominance of Camp. coli within Campylobacter spp. from chickens in Grenada may help elucidate the epidemiology of these pathogens in chickens.     

Effect of Conventional and Organic Production Practices on the Prevalence and Antimicrobial Resistance of Campylobacter spp. in Poultry

Intestinal tracts of broilers and turkeys from 10 conventional broiler farms and 10 conventional turkey farms, where antimicrobials were routinely used, and from 5 organic broiler farms and 5 organic turkey farms, where antimicrobials had never been used, were collected and cultured for Campylobacter species. A total of 694 Campylobacter isolates from the conventional and organic poultry operations were tested for antimicrobial resistance to nine antimicrobial agents by the agar dilution method. Although Campylobacter species were highly prevalent in both the conventional and organic poultry operations, the antimicrobial resistance rates were significantly different between the organic operations and the conventional operations. Less than 2% of Campylobacter strains isolated from organically raised poultry were resistant to fluoroquinolones, while 46% and 67% of Campylobacter isolates from conventionally raised broilers and conventionally raised turkeys, respectively, were resistant to these antimicrobials. In addition, a high frequency of resistance to erythromycin (80%), clindamycin (64%), kanamycin (76%), and ampicillin (31%) was observed among Campylobacter isolates from conventionally raised turkeys. None of the Campylobacter isolates obtained in this study was resistant to gentamicin, while a large number of the isolates from both conventional and organic poultry operations were resistant to tetracycline. Multidrug resistance was observed mainly among Campylobacter strains isolated from the conventional turkey operation (81%). Findings from this study clearly indicate the influence of conventional and organic poultry production practices on antimicrobial resistance of Campylobacter on poultry farms.     

Dissemination of Cephalosporin Resistance Genes between Escherichia coli Strains from Farm Animals and Humans by Specific Plasmid Lineages

Third-generation cephalosporins are a class of β-lactam antibiotics that are often used for the treatment of human infections caused by Gram-negative bacteria, especially Escherichia coli. Worryingly, the incidence of human infections caused by third-generation cephalosporin-resistant E. coli is increasing worldwide. Recent studies have suggested that these E. coli strains, and their antibiotic resistance genes, can spread from food-producing animals, via the food-chain, to humans. However, these studies used traditional typing methods, which may not have provided sufficient resolution to reliably assess the relatedness of these strains. We therefore used whole-genome sequencing (WGS) to study the relatedness of cephalosporin-resistant E. coli from humans, chicken meat, poultry and pigs. One strain collection included pairs of human and poultry-associated strains that had previously been considered to be identical based on Multi-Locus Sequence Typing, plasmid typing and antibiotic resistance gene sequencing. The second collection included isolates from farmers and their pigs. WGS analysis revealed considerable heterogeneity between human and poultry-associated isolates. The most closely related pairs of strains from both sources carried 1263 Single-Nucleotide Polymorphisms (SNPs) per Mbp core genome. In contrast, epidemiologically linked strains from humans and pigs differed by only 1.8 SNPs per Mbp core genome. WGS-based plasmid reconstructions revealed three distinct plasmid lineages (IncI1- and IncK-type) that carried cephalosporin resistance genes of the Extended-Spectrum Beta-Lactamase (ESBL)- and AmpC-types. The plasmid backbones within each lineage were virtually identical and were shared by genetically unrelated human and animal isolates. Plasmid reconstructions from short-read sequencing data were validated by long-read DNA sequencing for two strains. Our findings failed to demonstrate evidence for recent clonal transmission of cephalosporin-resistant E. coli strains from poultry to humans, as has been suggested based on traditional, low-resolution typing methods. Instead, our data suggest that cephalosporin resistance genes are mainly disseminated in animals and humans via distinct plasmids.     

Molecular Characterization of the Recently Emerged Poultry Pathogen Ornithobacterium rhinotracheale by Multilocus Sequence Typing

Ornithobacterium rhinotracheale (ORT) is an economically important bacterial pathogen of turkeys and chickens worldwide. Since its first detection, a variety of typing methods have been used to gain basic knowledge about the bacterial population structure, an issue that still needs to be addressed. Serological characterization revealed at least 18 different serotypes (A-R) with ORT of serotype A to be predominate among poultry. This study aimed to establish a multilocus sequence typing (MLST) scheme for ORT that could easily be used by other laboratories and allows for worldwide comparison of sequence data. For this purpose, 87 ORT strains from different poultry hosts, geographical origins, years of isolation and serotypes were included in the analysis to identify correlations. Fourteen different sequence types (ST) were found. The most common ST1 was identified in 40 ORT strains from turkeys and chickens on 4 continents and in 3 different European countries. Together with ST9, both STs represented over three quarters (77%) of ORT strains used in the MLST analysis and included strains of frequently cross-reacting ORT serotypes A, E and I. Nine STs were only represented by one ORT strain and might indicate possible avian host, disease or serotype-specific relationships. In contrast, discrepancies between serotype and phylogenetic relatedness were clearly demonstrated by ORT strains that belonged to identical serotypes but differed in their ST. The overall identified low genetic diversity among strains isolated from turkeys and chickens independent of host and geographical origins suggests that ORT has only recently been introduced into domestic poultry and dispersed worldwide.     

Frequency and Distribution of Tetracycline Resistance Genes in Genetically Diverse, Nonselected, and Nonclinical Escherichia coli Strains Isolated from Diverse Human and Animal Sources

Nonselected and natural populations of Escherichia coli from 12 animal sources and humans were examined for the presence and types of 14 tetracycline resistance determinants. Of 1,263 unique E. coli isolates from humans, pigs, chickens, turkeys, sheep, cows, goats, cats, dogs, horses, geese, ducks, and deer, 31% were highly resistant to tetracycline. More than 78, 47, and 41% of the E. coli isolates from pigs, chickens, and turkeys were resistant or highly resistant to tetracycline, respectively. Tetracycline MICs for 61, 29, and 29% of E. coli isolates from pig, chickens, and turkeys, respectively, were ≥233 μg/ml. Muliplex PCR analyses indicated that 97% of these strains contained at least 1 of 14 tetracycline resistance genes [tetA, tetB, tetC, tetD, tetE, tetG, tetK, tetL, tetM, tetO, tetS, tetA(P), tetQ, and tetX] examined. While the most common genes found in these isolates were tetB (63%) and tetA (35%), tetC, tetD, and tetM were also found. E. coli isolates from pigs and chickens were the only strains to have tetM. To our knowledge, this represents the first report of tetM in E. coli.     

Impact of Sodium Alginate Packaging Film Synthesized Using Syzygium cumini Seed Extract on Multi Drug Resistant Escherichia coli Isolated from Raw Buffalo Meat

A total of 50 Escherichia coli were isolated from buffalo meat and their antibiotic profiling was carried out. 90% E. coli isolates showed resistant to two or more classes of 21 commonly used antibiotics. Moreover, there was also variation in resistance/sensitivity behavior towards antibiotics. Highest resistance was found to be against methicillin (84%) in the isolates followed by vancomicin (70%), sulphadiazine (68%) and cefaclor (66%), whereas, resistance was less common for kanamycin (8%) and chloramphenicol (4%). ECMB1, ECMA2, ECMA8, ECMS9 and ECMA31 strains showed highest MDR pattern with presence of ^blaCTX-M, qnr S and qnr B resistant genes. ECMB1 strain was resistant to 14 antibiotics belonging to 7 different classes. Therefore, ECMB1 was selected for further studies. Sodium Alginate Film incorporated with 10, 20, and 30% ethanolic extract of Syzygium cumini (EESC) were formulated and characterized using state-of-art techniques. A dose-dependent antibacterial activity against E. coli ECMB1 was recorded by the films made from EESC (EESCF). The growth kinetics of E. coli strain ECMB1 showed 9% decrease in log CFU when it was cultured in 30% EESCF as compared to control cells after 12 h of growth. Present finding highlight the efficacy and possible use of EESCF as meat packaging film to prevent food spoilage caused by MDR bacteria.     

Assessment of three indigenous South African herbivores as potential reservoirs and vectors of antibiotic-resistant <Emphasis Type="Italic">Escherichia coli</Emphasis>

Due to limited data available on the presence of antibiotic-resistant (ABR) bacteria in faeces of wild herbivores in South Africa, this study analysed resistance patterns for Escherichia coli isolates from wildebeest, zebra and giraffe in addition to pet and farm pig faeces. Total and faecal coliforms and E. coli were quantified in faecal matter using a most probable number (MPN) guideline procedure. Antibiotic resistance profiles against 12 selected antibiotics representing seven classes were determined for 30 randomly selected E. coli isolates from each animal using the European Committee on Antimicrobial Susceptibility Testing (EUCAST) disk diffusion procedure. While log₁₀ MPN values per gram of animal faeces for total/faecal coliforms ranged from 4.51/4.11 to 5.70/5.50, the E. coli MPN values were in a range of 3.43–5.14. The proportion of ABR E. coli isolates ranged from 43% (giraffe) to 93% (zebra). About 47% of E. coli isolates from zebra faeces were categorized as multidrug-resistant (MDR), while for wildebeest and giraffe, no MDR isolates were detected. In comparison, 10% of E. coli isolates from pet pig and about 7% from farm pig faeces were categorized as MDR. Although most MDR isolates were resistant to at least one β-lactam antibiotic, only one MDR isolate from farm pig faeces was resistant to both norfloxacin and ciprofloxacin, the two fluoroquinolones tested. However, no resistance was detected to the tested carbapenems and tigecycline. The results of this study indicate that indigenous South African herbivores may serve as potential reservoirs and vectors for the dissemination of ABR E. coli strains.     

Diversity of Multi-Drug Resistant Avian Pathogenic Escherichia coli (APEC) Causing Outbreaks of Colibacillosis in Broilers during 2012 in Spain

Avian pathogenic Escherichia coli (APEC) are the major cause of colibacillosis in poultry production. In this study, a total of 22 E. coli isolated from colibacillosis field cases and 10 avian faecal E. coli (AFEC) were analysed. All strains were characterised phenotypically by susceptibility testing and molecular typing methods such as pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST). The presence of 29 virulence genes associated to APEC and human extraintestinal pathogenic E. coli (ExPEC) was also evaluated. For cephalosporin resistant isolates, cephalosporin resistance genes, plasmid location and replicon typing was assessed. Avian isolates belonged to 26 O:H serotypes and 24 sequence types. Out of 22 APEC isolates, 91% contained the virulence genes predictors of APEC; iutA, hlyF, iss, iroN and ompT. Of all strains, 34% were considered ExPEC. PFGE analysis demonstrated a high degree of genetic polymorphism. All strains were multi-resistant, including those isolated from healthy animals. Eleven strains were resistant to cephalosporins; six contained bla _CTX-M-14, two bla _SHV-12, two bla _CMY-2 and one bla _SHV-2. Two strains harboured qnrA, and two qnrA together with aac(6’)-Ib-cr. Additionally, the emergent clone O25b:H4-B2-ST131 was isolated from a healthy animal which harboured bla _CMY-2 and qnrS genes. Cephalosporin resistant genes were mainly associated to the presence of IncK replicons. This study demonstrates a very diverse population of multi-drug resistant E. coli containing a high number of virulent genes. The E. coli population among broilers is a reservoir of resistance and virulence-associated genes that could be transmitted into the community through the food chain. More epidemiological studies are necessary to identify clonal groups and resistance mechanisms with potential relevance to public health.     

Natural Transformation-Mediated Transfer of Erythromycin Resistance in Campylobacter coli Strains from Turkeys and Swine

Erythromycin resistance in Campylobacter coli from meat animals is frequently encountered and could represent a substantial barrier to antibiotic treatment of human infections. Erythromycin resistance in this organism has been associated with a point mutation (A2075G) in the 23S rRNA gene. However, the mechanisms responsible for possible dissemination of erythromycin resistance in C. coli remain poorly understood. In this study, we investigated transformation-mediated acquisition of erythromycin resistance by genotypically diverse C. coli strains from turkeys and swine, with total genomic DNA from erythromycin-resistant C. coli of either turkey or swine origin used as a donor. Overall, transformation to erythromycin resistance was significantly more frequent in C. coli strains from turkeys than in swine-derived strains (P < 0.01). The frequency of transformation to erythromycin resistance was 10⁻⁵ to 10⁻⁶ for turkey-derived strains but 10⁻⁷ or less for C. coli from swine. Transformants harbored the point mutation A2075G in the 23S rRNA gene, as did the erythromycin-resistant strains used as DNA donors. Erythromycin resistance was stable in transformants following serial transfers in the absence of the antibiotic, and most transformants had high MICs (>256 μg/ml), as did the C. coli donor strains. In contrast to the results obtained with transformation, spontaneous mutants had relatively low erythromycin MICs (32 to 64 μg/ml) and lacked the A2075G mutation in the 23S rRNA gene. These findings suggest that natural transformation has the potential to contribute to the dissemination of high-level resistance to erythromycin among C. coli strains colonizing meat animals.     

Feeding the Probiotic Enterococcus faecium Strain NCIMB 10415 to Piglets Specifically Reduces the Number of Escherichia coli Pathotypes That Adhere to the Gut Mucosa

Feed supplementation with the probiotic Enterococcus faecium for piglets has been found to reduce pathogenic gut microorganisms. Since Escherichia coli is among the most important pathogens in pig production, we performed comprehensive analyses to gain further insight into the influence of E. faecium NCIMB 10415 on porcine intestinal E. coli. A total of 1,436 E. coli strains were isolated from three intestinal habitats (mucosa, digesta, and feces) of probiotic-supplemented and nonsupplemented (control) piglets. E. coli bacteria were characterized via pulsed-field gel electrophoresis (PFGE) for clonal analysis. The high diversity of E. coli was reflected by 168 clones. Multilocus sequence typing (MLST) was used to determine the phylogenetic backgrounds, revealing 79 sequence types (STs). Pathotypes of E. coli were further defined using multiplex PCR for virulence-associated genes. While these analyses discerned only a few significant differences in the E. coli population between the feeding groups, analyses distinguishing clones that were uniquely isolated in either the probiotic group only, the control group only, or both groups (shared group) revealed clear effects at the habitat level. Interestingly, extraintestinal pathogenic E. coli (ExPEC)-typical clones adhering to the mucosa were significantly reduced in the probiotic group. Our data show a minor influence of E. faecium on the overall population of E. coli in healthy piglets. In contrast, this probiotic has a profound effect on mucosa-adherent E. coli. This finding further substantiates a specific effect of E. faecium strain NCIMB 10415 in piglets against pathogenic E. coli in the intestine. In addition, these data question the relevance of data based on sampling fecal E. coli only.