Use of a selective medium and a membrane filter method for isolation ofCampylobacter species from Spanish paediatric patients

A study was conducted to assess the value of a combination of two culture methods for isolation ofCampylobacter spp. from Spanish children. Seven hundred twenty-nine diarrhoea] stool specimens from 599 patients were examined forCampylobacter spp. by culturing them on charcoal cefoperazone deoxycholate agar and on blood agar with a membrane filter. One hundred sixteenCampylobacter strains were isolated from a total of 108 specimens; 75 (64.6%) wereCampylobacter jejuni, 32 (27.5%) wereCampylobacter coli, 8 (6.8%) were non-typeable, and one (0.9%) wasCampylobacter upsaliensis. Campylobacters were isolated from 99 positive samples using charcoal cefoperazone deoxycholate agar alone. The filtration technique alone yielded only 86 positive samples. Seven specimens yielded differentCampylobacter spp. with different media. The only catalase-negative strain was recovered using the filter method. The combination of the selective medium with the filter method increased the isolation rate ofCampylobacter strains by 14.1%. Isolation rates of campylobacters using the filter method were similar to those reported in European studies, in which a similar frequency ofCampylobacter upsaliensis was observed. The addition of a filter method for routine laboratory isolation of campylobacters should be considered in selected age groups (in children <10 years of age) or in areas where catalase-negative or weakly-positiveCampylobacter strains may be of epidemiological significance.     

Detection of Campylobacter Species and Arcobacter butzleri in Stool Samples by Use of Real-Time Multiplex PCR

The presence of Campylobacter (or Campylobacter-like) species in stools from patients suspected of infectious gastroenteritis (n = 493) was investigated using real-time PCR for detection of Arcobacter butzleri (hsp60 gene), Campylobacter coli (ceuE gene), Campylobacter jejuni (mapA), five acknowledged pathogenic Campylobacter spp. (C16S_Lund assay), and the Campylobacter genus (C16S_LvI assay). In total, 71.4% of the samples were positive for Campylobacter DNA (n = 352) by a Campylobacter genus-specific (C16S_LvI) assay. A total of 23 samples (4.7%) were positive in the C16S_Lund assay, used for detection of C. jejuni, C. coli, C. lari, C. upsaliensis, and C. hyointestinalis. Subsequent identification of these samples yielded detection frequencies (DF) of 4.1% (C. jejuni), 0.4% (C. coli), and 0.4% (C. upsaliensis). The DF of A. butzleri was 0.4%. Interestingly, sequencing of a subgroup (n = 46) of C16S_LvI PCR-positive samples resulted in a considerable number of Campylobacter concisus-positive samples (n = 20). PCR-positive findings with the C16S_Lund and C. jejuni/C. coli-specific assays were associated with more serious clinical symptoms (diarrhea and blood). Threshold cycle (C_T) values of C. jejuni/C. coli PCR-positive samples were comparable to those of the C16S_Lund PCR (P = 0.21). C_T values for both assays were significantly lower than those of the C16S_LvI assay (P < 0.001 and P < 0.00001, respectively). In conclusion, this study demonstrated that in combination, the C. jejuni/C coli-specific assays and the C16S_Lund assay are both useful for routine screening purposes. Furthermore, the DF of the emerging pathogen C. concisus was at least similar to the DF of C. jejuni.     

Rapid detection ofCampylobacter jejuni andCampylobacter coli isolated from clinical specimens using the polymerase chain reaction

SeventeenCampylobacter strains isolated from 16 children hospitalised with acute diarrhea were analysed by in vitro enzymatic amplification using two sets of oligonucleotide primers specific forCampylobacter jejuni andCampylobacter coli, respectively. Thirteen strains (76 %) were identified asCampylobacter jejuni and four strains (24 %) asCampylobacter coli. Subsequent bacteriological identification confirmed the identity of the same 13Campylobacter jejuni strains and the 4Campylobacter coli strains. Thus, these PCR methods enabled rapid and specific detection of all theCampylobacter jejuni andCampylobacter coli strains without any false-positive or false-negative results.     

Antibiotic susceptibility pattern and genotyping of campylobacter species isolated from children suffering from gastroenteritis

Purpose: To study the prevalence and the antimicrobial resistance of campylobacter species isolated from children suffering from gastroenteritis. Materials and Methods: A total of 125 stool samples were collected from children with gastroenteritis. The identification of isolates was performed with conventional methods as well as with molecular methods based on 16SrRNA species-specific gene amplification by PCR and product analysis. Resistance pattern of the isolated strains was determined using agar dilution method. Results: Conventional methods including sodium hippurate hydrolysis revealed that 12 (9.6%) samples were positive for Campylobacter species. Ten out of 12 Campylobacter spp. were identified as Campylobacter jejuni and 2 as Campylobacter coli but PCR assay revealed that five samples only were positive for Campylobacter and all were C. jejuni. Antimicrobial susceptibility to 10 antimicrobials was performed and all isolates (five isolates of C. jejuni) were susceptible to chloramphenicol, gentamicin and amikacin but all were resistant to ceftriaxone. Conclusion: PCR assay method allows reliable detection of C. jejuni. C. jejuni was the most prevalent Campylobacter species. Gentamicin, amikacin and chloramphenicol were the most effective antibiotic.     

Survival of Campylobacter jejuni and Campylobacter coli water isolates in lake and well water

The role of water for transmission of Campylobacter jejuni and C. coli to humans might be underestimated, as factors important for bacterial viability in water are largely unknown. We have studied water survival of seven C. jejuni and eight C. coli isolates originally isolated from Swedish waters, together with selected reference strains, over eight days at 4 °C in the dark in untreated water collected from a local lake and a private well. To study seasonality, lake water samples were collected during spring and autumn. Samples for culturable bacterial counts were taken on days 2, 4, 6, and 8 and compared to the start inoculum. For C. jejuni, a significantly better survival was observed in autumn than in spring lake water. Furthermore, C. jejuni had a significantly better survival than C. coli in autumn lake and well water samples; the rate of culturability loss was almost double for C. coli in autumn lake water. These findings contribute to a better understanding on the seasonality of waterborne Campylobacter infections and the general predominance of C. jejuni.     

Accuracy of the API Campy system,the Vitek 2 Neisseria–Haemophilus card and matrix-assisted laser desorption ionization time-of-flight mass spectrometry for the identification of Campylobacter and related organisms

Biochemical identification of Campylobacter and related organisms is not always specific, and may lead to diagnostic errors. The API Campy, the Vitek 2 system and matrix-assisted desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) are commercially available methods that are routinely used for the identification of these microorganisms. In the present study, we used 224 clinical isolates and ten reference strains previously identified by multiple PCR assays, whole cell protein profiling and either DNA–DNA hybridization or sequencing analysis to compare the reliability of these three methods for the identification of Campylobacter and related pathogens. The API Campy accurately identified 94.4% of Campylobacter jejuni ssp. jejuni and 73.8% of Campylobacter coli, but failed to correctly identify 52.3% of other Epsilobacteria. The Vitek 2 Neisseria–Haemophilus card correctly identified most C. jejuni ssp. jejuni (89.6%) and C. coli (87.7%) strains, which account for the majority of campylobacterioses reported in humans, but it failed in the identification of all of the other species. Despite a good identification rate for both C. jejuni ssp. jejuni and C. coli, both methods showed poor sensitivity in the identification of related organisms, and additional tests were frequently needed. In contrast to API Campy and Vitek, MALDI-TOF MS correctly identified 100% of C. coli and C. jejuni strains tested. With an overall sensitivity of 98.3% and a short response time, this technology appears to be a reliable and promising method for the routine identification of Campylobacter and other Epsilobacteria.     

Identification of the Enteropathogens Campylobacter jejuni and Campylobacter coli Based on the cadF Virulence Gene and Its Product

Campylobacter jejuni and Campylobacter coli are common causes of gastroenteritis in humans. Infection with C. jejuni or C. coli is commonly acquired by eating undercooked chicken. The goal of this study was to develop specific detection assays for C. jejuni and C. coli isolates based on the cadF virulence gene and its product. The cadF gene from C. jejuni and C. coli encodes a 37-kDa outer membrane protein that promotes the binding of these pathogens to intestinal epithelial cells. A fragment of approximately 400 bp was amplified from 38 of 40 (95%) C. jejuni isolates and 5 of 6 (83.3%) C. coli isolates with primers designed to amplify an internal fragment of the cadF gene. PCR was then used to amplify Campylobacter DNA from store-bought chickens. A 400-bp band was amplified from 26 of the 27 chicken carcasses tested by the PCR-based assay. The CadF protein was detected in every C. jejuni and C. coli isolate tested, as judged by immunoblot analysis with a rabbit anti-C. jejuni 37-kDa serum. In addition, methanol-fixed samples of whole-cell C. jejuni and C. coli were detected with the rabbit anti-37-kDa serum by using an indirect-immunofluorescence microscopy assay. These findings indicate that the cadF gene and its product are conserved among C. jejuni and C. coli isolates and that a PCR assay based on the cadF gene may be useful for the detection of Campylobacter organisms in food products.     

Evaluation of a New Commercial Immunoassay >for Rapid Detection of Campylobacter jejuni in Stool Samples

 In order to evaluate a new commercial enzyme immunoassay (ProspecT Campylobacter Microplate Assay; Alexon-Trend, USA) for the detection of Campylobacter jejuni and Campylobacter coli in stool samples, 30 faecal specimens known to be culture-positive for Campylobacter jejuni were tested with the new assay. The detection limit was approximately 3×10⁶/ml in faecal suspensions. The sensitivity relative to culture was 80% (24/30). All of the 24 positive samples, except for one, remained positive after being stored at –20  °C for 60 days. The specificity of the test was 100%. Interestingly, 6 of 11 additional Campylobacter jejuni culture-positive samples that had been obtained from patients with Guillain-Barré syndrome and stored at –20  °C for periods of up to 5 years tested positive in the assay. The performance of the assay indicates that it has potential value for use in future early intervention studies.     

Detection by PCR of eight groups of enteric pathogens in 4,627 faecal samples: re-examination of the English case-control Infectious Intestinal Disease Study (1993–1996)

The English case-control Infectious Intestinal Disease Study (1993–1996) failed to detect an enteric pathogen or toxin in 49% of cases of gastroenteritis. In the present study, polymerase chain reaction (PCR) assays were applied to DNA and cDNA generated from 4,627 faecal samples from cases and controls archived during the original study for the detection of norovirus, rotavirus, sapovirus, Campylobacter spp., Salmonella spp., enteroaggregative Escherichia coli, Cryptosporidium spp., and Giardia spp. The percentage of archived samples from cases and from controls in which at least one agent (or toxin) was detected increased from 53% in the original study to 75% and from 19 to 42%, respectively, after the application of PCR assays. Among cases, the following percentages of enteric pathogens were detected: norovirus 36%, rotavirus A 31%, sapovirus 4%, Salmonella spp. 6%, Campylobacter jejuni 13%, Campylobacter coli 2%, other Campylobacter spp. 8%, enteroaggregative E. coli 6%, Giardia spp. 2%, and Cryptosporidium spp. 2%. The present study provides additional insight into the aetiology of infectious intestinal disease in England and highlights the occurrence of viral infections in cases as well as in asymptomatic individuals. Other notable findings include the frequent presence of Campylobacter spp. other than C. jejuni or C. coli, the high frequency of multiple agents in 41% of cases and in 13% of controls, and the variation in the aetiology and rate of infection found for different age groups. The results demonstrate the greater sensitivity of PCR-based methods compared to current conventional methods.     

Detection of Campylobacter in Stool and Determination of Significance by Culture,Enzyme Immunoassay,and PCR in Developing Countries

Campylobacter is a common bacterial enteropathogen that can be detected in stool by culture, enzyme immunoassay (EIA), or PCR. We compared culture for C. jejuni/C. coli, EIA (ProSpecT), and duplex PCR to distinguish Campylobacter jejuni/C. coli and non-jejuni/coli Campylobacter on 432 diarrheal and matched control stool samples from infants in a multisite longitudinal study of enteric infections in Tanzania, Bangladesh, and Peru. The sensitivity and specificity of culture were 8.5% and 97.6%, respectively, compared with the results of EIA and 8.7% and 98.0%, respectively, compared with the results of PCR for C. jejuni/C. coli. Most (71.6%) EIA-positive samples were positive by PCR for C. jejuni/C. coli, but 27.6% were positive for non-jejuni/coli Campylobacter species. Sequencing of 16S rRNA from 53 of these non-jejuni/coli Campylobacter samples showed that it most closely matched the 16S rRNA of C. hyointestinalis subsp. lawsonii (56%), C. troglodytis (33%), C. upsaliensis (7.7%), and C. jejuni/C. coli (2.6%). Campylobacter-negative stool spiked with each of the above-mentioned Campylobacter species revealed reactivity with EIA. PCR detection of Campylobacter species was strongly associated with diarrhea in Peru (odds ratio [OR] = 3.66, P < 0.001) but not in Tanzania (OR = 1.56, P = 0.24) or Bangladesh (OR = 1.13, P = 0.75). According to PCR, Campylobacter jejuni/C. coli infections represented less than half of all infections with Campylobacter species. In sum, in infants in developing country settings, the ProSpecT EIA and PCR for Campylobacter reveal extremely high rates of positivity. We propose the use of PCR because it retains high sensitivity, can ascertain burden, and can distinguish between Campylobacter infections at the species level.     

Phenotypic and genotypic differentiation of Campylobacter spp. isolated from Austrian broiler farms: A comparison

An identification scheme based on restriction fragment length polymorphism of polymerase chain reaction products (PCR-RFLP) was developed to differentiate isolates of the genera Campylobacter, Arcobacter and Helicobacter. Based on the 16S rRNA gene of these genera, PCR amplified a 1216-bp fragment. The amplicons were digested with the restriction enzymes RsaI and EcoRV. Additional differentiation was obtained using a PCR-assay based on the hippuricase gene. Genotyping was performed on several reference strains from the National Collection of Typing Culture (NCTC), London, and on 130 field isolates. In parallel, a phenotypic differentiation was performed, in order to compare the results. In 119 cases (91.5%) the results obtained from the genotypic characterization were concordant with those from phenotypic testing. Co-infections with Campylobacter jejuni and Campylobacter coli in two samples and seven hippurate-negative C. jejuni-strains were identified by the genotypic method. Furthermore, PCR-RFLP assays identified an atypical isolate as Campylobacter fetus/hyointestinalis.     

Re-thinking the chicken–Campylobacter jejuni interaction: a review

Chickens are recognized as an imperative source of thermophilic Campylobacter spp., carrying this microorganism in high numbers in their intestinal tract. For a long time, Campylobacter jejuni has been considered as a commensal microorganism which colonizes its primary host rather than infecting it, in the absence of any obvious clinical signs. However, recent studies question this and argue for a deeper understanding of the host–bacteria interaction. Following oral uptake, it was demonstrated that C. jejuni interacts intimately with the gut epithelium and influences cellular functions of the host, with consequences on nutrient absorption. The immune reaction of the host which was revealed in some studies confirmed the infectious nature of C. jejuni. In agreement with this, an increased expression of pro-inflammatory cytokine genes was noticed. The ability to induce intestinal damage and to modulate the barrier function of the intestinal epithelia has further consequences on gut integrity, as it facilitates the paracellular passage of C. jejuni into the underlying tissues and it supports the translocation of luminal bacteria such as Escherichia coli to internal organs. This is associated with an alteration of the gut microbiota as infected birds have a significantly lower abundance of E. coli in different parts of the intestine. Some studies found that the gut microbiota influences the infection and translocation of C. jejuni in chickens in various ways. The effects of C. jejuni on the intestinal function of chickens already indicate a possible interference with bird performance and welfare, which was confirmed in some experimental studies. Furthermore, it could be demonstrated that a Campylobacter infection has an influence on the movement pattern of broiler flocks, supporting experimental studies. The intense interaction of C. jejuni with the chicken supports its role as an infectious agent instead of simply colonizing the gut. Most of the findings about the impact of Campylobacter on chickens are derived from studies using different Campylobacter isolates, a specific type of bird and varying experimental design. However, experimental studies demonstrate an influence of the aforementioned parameters on the outcome of a certain trial, arguing for improved standardization. This review summarizes the actual knowledge of the host–pathogen interaction of C. jejuni in chickens, emphasizing that there are still major gaps despite recently gained knowledge. Resolving the cascade from oral uptake to dissemination in the organism is crucial to fully elucidating the interaction of C. jejuni with the chicken host and to assess the clinical and economic implications with possible consequences on preventive interventions.     

Comparison of two selective media and a membrane filter technique for isolation ofCampylobacter species from diarrhoeal stools

Diarrhoeal stool specimens from 415 patients were examined forCampylobacter spp. by culture on charcoal cefoperazone deoxycholate agar (CCDA), Skirrow medium and Columbia blood agar overlaid with a 0.65 µm pore size membrane filter. Forty-eightCampylobacter strains were isolated from 45 (10.8 %) specimens by all media; 44 wereCampylobacter jejuni (91.7 %), three wereCampylobacter coli (6.3 %) and one wasCampylobacter hyointestinalis (2.0 %). The percentages ofCampylobacter-positive specimens isolated on Skirrow medium, CCDA and the membrane filter were 62, 82 and 95 %, respectively. The recovery of moreCampylobacter spp. from the same stool sample was achieved by the membrane filter method only. The highest isolation rate (100 %) was observed when culture on CCDA and the membrane filter method were combined.     

Drinking water as the source of Campylobacter coli infection in grandparent heavy breeders

The aim of the present study was the molecular identification of a common source of infection of Campylobacter coli in two grandparent breeder farms. Campylobacter jejuni and C. coli were isolated from well water and cloacal swabs from grandparent chickens. Colonies were genotyped using restriction fragment length polymorphism-flaA gene, pulsed field gel electrophoresis and multi-locus sequence typing. The same genotype of C. coli was found in both farms and in the well from which drinking water was supplied to the farms. The well water was epidemiologically linked as the source of C. coli infection. The molecular identification for epidemiological source-tracking of C. coli in breeder farms could aid in combating the colonization of this pathogen and therefore to reduce their incidence in human campylobacteriosis.     

Real-time multiplex PCR for simultaneous detection of Campylobacter jejuni, Salmonella, Shigella and Yersinia species in fecal samples

Diarrheal diseases due to notifiable bacterial infections require rapid diagnosis of the causative pathogens. To facilitate detection, a real-time multiplex PCR was developed that identifies common diarrhea-causing bacteria in fecal samples. On the basis of published sequence data, sets of primers and probes were designed that were specific for Campylobacter jejuni, Salmonella, Shigella/enteroinvasive Escherichia coli EIEC, and Yersinia species, suitable for use in a one-tube PCR assay. The assay was assessed using a list of 137 well-defined intestinal bacterial strains or isolates. Furthermore, 393 routine clinical stool samples were analyzed, and the results of real-time multiplex PCR were compared with those obtained by established microbiological methods. The PCR yielded results within 3 h including DNA purification. No false-positive signals or cross-reactions were observed. The analytical sensitivity was 10³ cfu mL⁻¹ for Campylobacter jejuni, 10⁴ cfu mL⁻¹ for Salmonella, and 10⁵ cfu mL⁻¹ for Shigella/EIEC and Yersinia, respectively. Compared with culture, PCR detected 79 out of 81 Campylobacter jejuni (97.5%), 71 out of 74 Salmonella (96%), 8 out of 8 Shigella (100%), and 10 out of 10 Yersinia-positive (100%) clinical samples. In culture-negative samples (n = 192), PCR additionally detected 2 Shigella, 1 Salmonella, and 5 Campylobacter jejuni infections. Thus, the new real-time multiplex PCR provides reliable results within a short time and might be useful as an additional diagnostic tool whenever time is important in the diagnosis of enteropathogenic bacteria.     

Emergence of resistance to erythromycin and fluoroquinolones in thermotolerantCampylobacter strains isolated from feces 1987–1991

During the period 1987 to 1991 a retrospective study was performed to determine the resistance of thermotolerantCampylobacter species isolated from feces to erythromycin and fluoroquinolones. Of the 672 strains studied, 614 (91.3 %) were identified asCampylobacter jejuni and 58 (8.7 %) asCampylobacter coli. During the study period the rate of resistance ofCampylobacter jejuni to erythromycin remained relatively stable (0.9–3.5 %), while resistance ofCampylobacter coli to erythromycin emerged later (1989) with much higher rates (14.8–33 %). Overall, 11.8 % and 10.7 % ofCampylobacter jejuni strains isolated after 1987 were resistant to nalidixic acid and ciprofloxacin respectively, resistance increasing from 2.3 % in 1988 to 32 % in 1991. In 1991 the first strains ofCampylobacter coli with resistance to these fluoroquinolones were detected (rates 29 % and 26 % respectively). Of the strains resistant to nalidixic acid, only 10.9 % were susceptible to ciprofloxacin.     

Detection of Campylobacter species in foods by indirect competitive ELISA using hen and rabbit antibodies

The competitive enzyme immunoassays for detection of Campylobacter jejuni, C. coli and C. fetus subsp. fetus have been developed. Rabbit and hen immunoglobulins were prepared for these purposes. The working conditions of ELISAs, such as the concentrations of immunoreactants, incubation temperatures and time, and the composition of the substrate have been established. The detection limits were in the range 5.0 10⁴–3.2 10⁶ cfu/ml. The application of chemiluminescent substrates did not result in any significant improvement of the assay's detectability and sensitivity. Prepared antibodies showed rather high specificity and cross-reactivity profiles, and both rabbit and hen immunoglobulins were similar. Only IgY to C. jejuni cross-reacted with seven strains of C. jejuni and two other Campylobacter spp.

A limited number of naturally and artificially contaminated food samples were tested. The results obtained by means of an enzyme immunoassay were compared with those obtained from PCR or commercially available Singlepath® Campylobacter GLISA-Rapid Test. Poultry products were naturally contaminated with Campylobacters. The wild species were identified as C. jejuni and C. coli.     

Prevalence of thermotolerant Campylobacter in pheasants (Phasianus colchicus)

The present study was undertaken with the aim to evaluate the prevalence of thermotolerant Campylobacter spp. in living pheasants in Italy. To achieve this goal, a total of 240 living pheasants, equally shared between female and male birds, were examined. Thermotolerant Campylobacter spp. was isolated in 104 out of 204 (43.3%) living pheasants analysed. Campylobacter coli (100%) and Campylobacter jejuni (13.5%) were identified by polymerase chain reaction. Adult pheasants showed a significantly higher prevalence value (P < 0.05) than younger pheasants.     

Thermophilic Campylobacter spp. in Danish broiler production: A cross-sectional survey and a retrospective analysis of risk factors for occurrence in broiler flocks

In order to elucidate the rate of thermophilic Campylobacter spp. carriage in Danish broiler production and to identify risk factors for occurrence of campylobacter in broiler flocks, a total of 88 randomly selected broiler flocks were tested for campylobacter infection, and a subsequent study of risk factors based on a questionnaire was conducted. The sample material comprised cloacal swabs from live birds before slaughter, and neck skin samples from carcasses at the end of the processing line. A total of 52% of the flocks were found Campylobacter spp.-positive before slaughter. At the end of processing, 24% of the flocks were positive. The species distribution was 87% Campylobacter jejuni, 8% Campylobacter coli and 5% Campylobacter lari. The following parameters were identified as significant risk factors: lack of a hygiene barrier (odds ratio (OR) = 3.1, 1.1 < OR < 9.3), presence of animals in the vicinity of the broiler house on farms with a missing hygiene barrier (OR = 7.0, 1.6 < OR < 33.9), livestock other than chickens on farms with a missing hygiene barrier (OR = 7.6, 1.4 < OR < 44.9), dividing the flock into batches for staggered slaughter (OR = 6.8, 1.2 < OR < 49.3), a down period of less than 14 days (OR = 5.0, 1.2 < OR < 22.6), and feeding purchased wheat rather than home-grown wheat (OR = 3.1, 1.0 <OR < 9.9). The presence of a hygiene barrier was found to be the single most important biosecurity measure for production of campylobacter-free broilers.     

Molecular characterization and genotypic antimicrobial resistance analysis of Campylobacter jejuni and Campylobacter coli isolated from broiler flocks in northern Italy

Genetic variability and genotypic antimicrobial resistance (AMR) of Campylobacter jejuni and Campylobacter coli from commercial broiler farms were investigated in this study. Campylobacter isolates were genetically characterized by random amplified polymorphic DNA (RAPD)-polymerase chain reaction (PCR) and flaA-SVR and flaB-SVR sequence-based typing. Eight RAPD types were identified in C. jejuni and three in C. coli, while 16 fla profiles were detected among all isolates. Further, 13 flaA-SVR and 13 flaB-SVR alleles were identified. Both typing methods detected a high level of genetic diversity, but fla-SVR typing showed a higher discriminatory power. Indeed, Simpson's index of fla typing (D=0.920) was higher than that of RAPD typing (D=0.814). Moreover, the association of flaA-SVR and flaB-SVR sequence analysis showed a higher discriminatory power compared with the sequence analysis of single loci. Isolates were also analysed by the mismatch amplification mutation assay PCR test and the detection of cmeB gene to determine the occurrence of genetic determinants of AMR to macrolides and fluoroquinolones and multidrug resistance. The A2074C and A2075G mutations in the 23S rRNA gene, the C257T mutation in the gyrA gene, and the cmeB gene were higher in C. coli (19.0%, 67.0%, 100.0% and 100.0%, respectively) than in C. jejuni (0.0%, 3.1%, 48.3% and 48.3%, respectively). This study showed a high degree of genetic diversity and a high prevalence of genetic determinants of macrolide resistance, fluoroquinolone resistance and multidrug resistance among C. jejuni and C. coli isolates from Italian commercial broiler farms.