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Food Microbiology Aug 2016Considering that several recent cases of human gastroenteritis have been associated with species from the Arcobacter genus, and that few data are currently available...
Considering that several recent cases of human gastroenteritis have been associated with species from the Arcobacter genus, and that few data are currently available about the occurrence of this genus in Italian shellfish, the aim of the present study was to evaluate the occurrence of Arcobacter spp. and the presence of virulence-associated genes. The approach consisted of cultural and biomolecular (multiplex-PCR and 16S-RFLP) methods identifying isolates, followed by PCR assays aimed at the cadF, ciaB, cjl349, irgA, hecA putative virulence genes. Arcobacter spp. was detected in 16/70 (22.8%) shellfish samples. Specifically, Arcobacter spp. was highlighted in 10/42 (23.8%) mussel and in 6/28 (21.4%) clam samples. Subsequently, biomolecular assays revealed Arcobacter butzleri in 12/16 (75%) and Arcobacter cryaerophilus 1B in 4/16 (25%) isolates. PCRs aimed at the five putative virulence genes demonstrated widespread distribution of these genes among Arcobacter isolates and some differences from the results published by other authors. Our research provides more information regarding the health risks associated with the consumption of raw bivalve molluscs and underlines the need to implement an adequate control plan by performing intensive and continuous monitoring in order to guarantee human health.
Topics: Animals; Arcobacter; Bacterial Proteins; Bivalvia; Consumer Product Safety; Food Contamination; Humans; Polymerase Chain Reaction; Shellfish
PubMed: 27052698
DOI: 10.1016/j.fm.2015.12.010 -
The Journal of Antimicrobial... May 2016To evaluate the feasibility of different methods for susceptibility testing of human Arcobacter isolates, to assess susceptibility to antibiotics commonly used to treat...
OBJECTIVES
To evaluate the feasibility of different methods for susceptibility testing of human Arcobacter isolates, to assess susceptibility to antibiotics commonly used to treat diarrhoeal illness and to obtain MIC distribution data.
METHODS
One-hundred-and-six unique Arcobacter strains were collected during an epidemiological study on pathogens in gastroenteritis. Strains were identified by multiplex PCR and PCR-RFLP, and characterized by PFGE. Susceptibility to ampicillin, erythromycin, tetracycline, doxycycline, gentamicin and ciprofloxacin was determined using gradient strip and disc diffusion methodology. Optimal conditions for growth and incubation were tested. Azithromycin was tested with gradient strip diffusion on a subset of 40 strains. Sequence analysis of the quinolone resistance-determining region of gyrA was performed for a subset of 18 strains.
RESULTS
Based on gradient diffusion results, most Arcobacter strains were susceptible to gentamicin (99%) and tetracycline (89%). Erythromycin (78%), ciprofloxacin (72%) and doxycycline (76%) retained moderate activity against Arcobacter spp. Only 9% of the strains were susceptible to ampicillin. Most Arcobacter butzleri strains were susceptible to ciprofloxacin (87%), whereas half of the Arcobacter cryaerophilus isolates (51%) showed high-level resistance (MIC >32 mg/L). MIC50 values were comparable for both macrolide antibiotics. Ciprofloxacin-resistant strains possessed an identical mutation in gyrA. Overall, categorical agreement between gradient and disc diffusion results was 60%. Gradient diffusion showed superior readability.
CONCLUSIONS
Gradient diffusion methodology is preferred for routine susceptibility testing. Acquired resistance to fluoroquinolones was observed in A. cryaerophilus. Macrolides are not first-choice empirical antibiotics for Arcobacter infections. Tetracyclines can be suggested for treatment of documented Arcobacter-related gastrointestinal infections.
Topics: Anti-Bacterial Agents; Arcobacter; Belgium; Electrophoresis, Gel, Pulsed-Field; Epidemiologic Studies; Gastroenteritis; Gram-Negative Bacterial Infections; Humans; Microbial Sensitivity Tests; Polymerase Chain Reaction; Polymorphism, Restriction Fragment Length
PubMed: 26851610
DOI: 10.1093/jac/dkv483 -
Journal of Microbiological Methods Feb 2016As the pathogenicity of Arcobacter species might be associated with various virulence factors, this study was aimed to develop and optimize three single-tube multiplex...
As the pathogenicity of Arcobacter species might be associated with various virulence factors, this study was aimed to develop and optimize three single-tube multiplex PCR (mPCR) assays that can efficiently detect multiple virulence-associated genes (VAGs) in Arcobacter spp. including the Arcobacter butzleri, Arcobacter cryaerophilus and Arcobacter skirrowii, respectively. The recognized target virulence factors used in the study were fibronectin binding protein (cj1349), filamentous hemagglutinin (hecA), hemolysin activation protein (hecB), hemolysin (tlyA), integral membrane protein virulence factor (mviN), invasin (ciaB), outer membrane protein (irgA) and phospholipase (pldA). Identical results were obtained between singleplex PCR and mPCR assays and no cross- and/or non-specific amplification products were obtained when tested against other closely related bacterial species. The sensitivities of these three mPCR assays were ranging from 1ngμL(-1) to 100ngμL(-1) DNA. The developed assays with combinations of duplex or triplex PCR primer pairs of VAGs were further evaluated and validated by applying them to isolates of the A. butzleri, A. cryaerophilus and A. skirrowii recovered from fecal samples of human and animal origins. The findings revealed that the distribution of the ciaB (90%), mviN (70%), tlyA (50%) and pldA (45%) genes among these target species was significantly higher than the hecA (16%), hecB (10%) and each of irgA and cj1349 (6%) genes, respectively. The newly developed mPCR assays can be used as rapid technique and useful markers for the detection, prevalence and profiling of VAGs in the Arcobacter spp. Moreover, these assays can easily be performed with a high throughput to give a presumptive identification of the causal pathogen in epidemiological investigation of human infections.
Topics: Animals; Arcobacter; Bacterial Proteins; Bacterial Typing Techniques; Base Sequence; DNA Primers; DNA, Bacterial; Fibronectins; Humans; Multiplex Polymerase Chain Reaction; Sensitivity and Specificity; Virulence; Virulence Factors
PubMed: 26769558
DOI: 10.1016/j.mimet.2015.12.017 -
BioMed Research International 2015Incidence of 9 virulence-associated genes and genetic diversity was determined in 79 A. butzleri and 6 A. cryaerophilus isolates from pork, beef, and chicken meat. All...
Incidence of 9 virulence-associated genes and genetic diversity was determined in 79 A. butzleri and 6 A. cryaerophilus isolates from pork, beef, and chicken meat. All A. butzleri isolates harboured the tlyA gene, and most of them carried ciaB, mviN, pldA, cadF, and cj1349 genes. ciaB was found to occur with higher frequency in poultry if compared with pork (p = 0.0007), while irgA was more frequent in poultry than in beef (p = 0.007). All 6 A. cryaerophilus isolates harboured the ciaB gene, while mviN and tlyA were detected in 3 out of these isolates. Only one isolate carried the cadF gene. All beef-derived A. cryaerophilus isolates carried ciaB, mviN, and tlyA genes. A. cryaerophilus isolates from chicken meat harboured ciaB gene only. The pork-derived isolate harboured ciaB and cadF genes. Seventy-four genotypes were distinguished within 79 A. butzleri isolates. Nineteen from 21 isolates derived from beef and pork were found to be closely related to A. butzleri from chicken meat. Each of the 6 A. cryaerophilus isolates was found to have unique genotype. We demonstrated that closely related genotypes can spread within pork, beef, and chicken meat populations of A. butzleri but not A. cryaerophilus.
Topics: Animals; Arcobacter; Cattle; Chickens; Food Microbiology; Genetic Variation; Incidence; Meat; Poland; Swine; Virulence Factors
PubMed: 26539546
DOI: 10.1155/2015/956507 -
Epidemiologie, Mikrobiologie,... Sep 2015Detection of biofilm formation by microbial pathogens relevant to the food industry and comparison of biofilm formation under different conditions of culture.
OBJECTIVE
Detection of biofilm formation by microbial pathogens relevant to the food industry and comparison of biofilm formation under different conditions of culture.
MATERIAL AND METHODS
The following microorganisms were selected for the study: Staphylococcus aureus, Listeria innocua, Listeria ivanovii, Cronobacter sakazakii, Cronobacter muytjensii, Arcobacter butzleri, Arcobacter cryaerophilus, Campylobacter jejuni, and Campylobacter coli. To detect biofilm formation the microtiter plate assay, as described by Christensen and culture on stainless steel coupons were used.
RESULTS
The biofilm forming capacity was confirmed in all microorganisms tested, both on the microtiter plates and stainless steel coupons. Biofilm formation was influenced by the culture medium, material used, and culture duration as well as by the test microorganism. It was found that different species and strains of the same genus differ in biofilm formation. Differences were also found between the collection strains and isolates from the environment. Some bacteria tended to form biofilm more readily on the surface of the polyethylene microtiter plates and less readily on stainless steel coupons while others appeared to have an opposite tendency. Some pathogens were able to increase the planktonic cell density in the initial suspension even by three orders of magnitude within 72 hours while producing plenty of biofilm.
CONCLUSIONS
The study of biofilm formation by high risk pathogens is of utmost importance, not only to the food industry. From the obtained results, it is evident that bacterial biofilms form rapidly (within 24 hours in the present study). Due to their architecture, these biofilms are difficult to eradicate, and therefore, it is crucial to prevent biofilm formation.
Topics: Bacteria; Bacterial Adhesion; Bacterial Physiological Phenomena; Biofilms; Colony Count, Microbial; Food Microbiology
PubMed: 26448305
DOI: No ID Found -
Food Microbiology Dec 2015Arcobacter spp. are considered to be emerging food- and waterborne pathogens for both humans and animals. However, their virulence mechanisms are still poorly...
Arcobacter spp. are considered to be emerging food- and waterborne pathogens for both humans and animals. However, their virulence mechanisms are still poorly understood. In this study the presence of ten virulence genes (cadF, ciaB, cj1349, hecA, hecB, mviN, pldA, irgA, tlyA and iroE) was assessed in a set of 47 strains of Arcobacter butzleri, 10 of Arcobacter cryaerophilus and 1 Arcobacter skirrowii strain recovered from different food products (pork, chicken, beef, milk, clams and mussels). Overall, the genes cadF, ciaB, cj1349, mviN, pldA and tlyA were detected in all A. butzleri and A. skirrowii strains. Lower detection rates were observed for irgA, iroE, hecA and hecB. The genes hecB and iroE were detected neither in A. cryaerophilus nor in A. skirrowii. The genes hecA and irgA were not detected in A. skirrowii. It was noteworthy that the genes hecA and hecB were significantly (P < 0.05) highly detected in A. butzleri strains isolated from clams compared with strains isolated from milk and chicken. Therefore, our findings underline clams as a source of A. butzleri strains with high prevalence of putative virulence genes. This could be hazardous to human health, especially because these bivalves are usually consumed raw or undercooked.
Topics: Animals; Arcobacter; Bacterial Proteins; Bivalvia; Cattle; Chickens; Food Contamination; Food Microbiology; Meat; Milk; Shellfish; Swine; Virulence Factors
PubMed: 26338128
DOI: 10.1016/j.fm.2015.07.015 -
Journal of Dairy Science Oct 2015Ricotta cheese is a ready-to-eat product with properties (pH >6.0, aw >0.98-0.99) and moisture content (75-80%) that may pose a risk to public health due to postprocess...
Ricotta cheese is a ready-to-eat product with properties (pH >6.0, aw >0.98-0.99) and moisture content (75-80%) that may pose a risk to public health due to postprocess contamination by several bacterial pathogens, including Arcobacters. The objective of the study was to evaluate the behavior of Arcobacter butzleri and Arcobacter cryaerophilus in ricotta cheese during its shelf life assuming postprocessing contamination. Two types of ricotta cheese, artisanal water buffalo (WB) and industrial cow milk ricotta cheese, were experimentally contaminated with A. butzleri and A. cryaerophilus and the count was monitored at 2 different temperatures (6°C and 12°C) during shelf life of 5 d for WB cheese and 22 d for industrial ricotta cheese. In WB ricotta cheese the A. butzleri count remained stable during the 5 d of storage at 6°C, whereas a moderate but significant decrease was observed in A. cryaerophilus count. The counts of both species increased when WB ricotta cheese was stored at 12°C. In industrial ricotta cheese stored at 6°C, a significant reduction was observed both in A. butzleri and A. cryaerophilus counts during the 22-d storage period; at 12°C storage, a count increase was observed for both Arcobacter species up to d 14 of storage after which the log cfu/g count resulted constant until d 22 of storage. The ability of A. butzleri and A. cryaerophilus to survive at 6°C and to grow at 12°C in ricotta cheese has significant food safety implications.
Topics: Animals; Arcobacter; Buffaloes; Cattle; Cheese; Food Microbiology; Food Safety; Species Specificity; Temperature
PubMed: 26233450
DOI: 10.3168/jds.2015-9560 -
Applied and Environmental Microbiology Aug 2015Even though dairy cows are known carriers of Arcobacter species and raw or minimally processed foods are recognized as the main sources of human Arcobacter infections in...
Even though dairy cows are known carriers of Arcobacter species and raw or minimally processed foods are recognized as the main sources of human Arcobacter infections in industrialized countries, data on Arcobacter excretion patterns in cows and in milk are scant. This study aimed to identify potentially pathogenic Arcobacter species in a dairy herd and to investigate the routes of Arcobacter transmission among animals and the potential sources of cattle infection and milk contamination. A strategy of sampling the same 50 dairy animals, feed, water, and milk every month for a 10-month period, as well as the sampling of quarter milk, animal teats, the milking environment, and animals living on the farm (pigeons and cats), was used to evaluate, by pulsed-field gel electrophoresis (PFGE), the characteristic patterns in animals, their living environment, and the raw milk they produced. Of the 463 samples collected, 105 (22.6%) were positive for Arcobacter spp. by culture examination. All the matrices except quarter milk and pigeon gut samples were positive, with prevalences ranging from 15 to 83% depending on the sample. Only three Arcobacter species, Arcobacter cryaerophilus (54.2%), A. butzleri (34.2%), and A. skirrowii (32.3%), were detected. PFGE analysis of 370 isolates from positive samples provided strong evidence of Arcobacter circulation in the herd: cattle likely acquire the microorganisms by orofecal transmission, either by direct contact or from the environment, or both. Water appears to be a major source of animal infection. Raw milk produced by the farm and collected from a bulk tank was frequently contaminated (80%) by A. butzleri; our PFGE findings excluded primary contamination of milk, whereas teats and milking machine surfaces could be sources of Arcobacter milk contamination.
Topics: Animals; Animals, Domestic; Arcobacter; Carrier State; Cats; Cattle; Columbidae; DNA Fingerprinting; Electrophoresis, Gel, Pulsed-Field; Environmental Microbiology; Food Contamination; Gram-Negative Bacterial Infections; Humans; Milk; Molecular Typing
PubMed: 26002896
DOI: 10.1128/AEM.01035-15 -
Polish Journal of Veterinary Sciences 2015Arcobacter butzleri and A. cryaerophilus are considered potential foodborne pathogens. Consumption of Arcobacter-contaminated food is regarded the most likely source of...
Arcobacter butzleri and A. cryaerophilus are considered potential foodborne pathogens. Consumption of Arcobacter-contaminated food is regarded the most likely source of human poisoning. We investigated the prevalence and antimicrobial resistance of Arcobacter isolates in 210 retail meat samples. Seventy-nine A. butzleri and 6 A. cryaerophilus were isolated from pork, beef and chicken meat. Incidence ofA. butzleri was found to be the highest in chicken meat (83%). Less of A. butzleri was isolated from beef (16%) and pork (14%). Most of the A. butzleri isolates were resistant to β-lactams, like ampicillin (85%), amoxicillin with clavulonic acid (63%), cefotaxime (66%) and mac- rolides, i.e., erythromycin (62%). In contrast, all except one A. cryaerophilus isolates were susceptible to erythromycin. Tetracycline and aminoglycosides showed the highest efficacy against A. butzleri and A. cryaerophilus since almost 80% of their population was susceptible to these agents. All, except one A. cryaerophilus and the majority ofA. butzleri isolates (70%) were susceptible to fluoroquinolones. The incidence of multiresistant isolates was found in forty two (53%) A. butzleri, and one (16%) A. cryaerophilus isolates Eight A. butzleri isolates were resistant to all antimicrobials tested. These results indicate significant incidence of potential foodborne zoonotic agents, i.e. A. butzleri and A. cryaerophilus including multiresistant isolates in retail meat in Poland.
Topics: Animals; Anti-Bacterial Agents; Arcobacter; Cattle; Chickens; Commerce; Drug Resistance, Bacterial; Food Microbiology; Meat; Poland; Species Specificity; Swine
PubMed: 25928911
DOI: 10.1515/pjvs-2015-0008 -
Folia Microbiologica Nov 2015This study provides information on the occurrence of Arcobacter in several types of water and food products of animal origin in the Czech Republic. We processed 190...
This study provides information on the occurrence of Arcobacter in several types of water and food products of animal origin in the Czech Republic. We processed 190 samples using the modified method, and the occurrence of Arcobacter spp. was confirmed in 36.8 % of these. This total incidence consisted of Arcobacter butzleri (27.3 %), Arcobacter cryaerophilus (8.4 %) and Arcobacter skirrowii (1.1 %). We newly described the common presence of Arcobacter spp. in sewage water in the Czech Republic that is released into waterways after processing in water treatment plants (86.7 %). All the acquired isolates were subject to detailed confirmation with subsequent species classification using multiplex PCR and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). In this study, we used a modification of a method using passive filtration of an enriched sample, which could be suitable for the isolation of Arcobacter, especially in combination with Campylobacter selective agar chromogenic medium. Our studies have shown this agar to be quite suited to the isolation of Arcobacter and that it can be an appropriate instrument for accelerating culture diagnostics.
Topics: Arcobacter; Colony Count, Microbial; Culture Media; Czech Republic; Food Microbiology; Sewage
PubMed: 25912846
DOI: 10.1007/s12223-015-0395-x