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Frontiers in Microbiology 2022The is a globally emerging foodborne and zoonotic pathogen that can cause diarrhea in humans. It is relatively homogenous and clearly distinguishes the group from other...
The is a globally emerging foodborne and zoonotic pathogen that can cause diarrhea in humans. It is relatively homogenous and clearly distinguishes the group from other . () is a heterogeneous species and little is known about its genomic characterization in China. This study aims to determine the genetic and plasmid features of based on whole-genome sequence (WGS). Average Nucleotide Identity (ANI) and DNA-DNA hybridization (DDH) were used for the species classification for 90 initially identified strains. One complete genome and 42 draft genomes were obtained by whole genome sequencing. The genomic characteristics were determined using various bioinformatics software. The genomes of the strains examined were estimated to vary from 1.81 to 2.28 Mb in length, with a G + C content of around 27%. ANI and DDH results indicated that 90 initially identified strains should be reclassified into four new species (ANI > 96% or DDH > 70%). Two clades (four subclades) were identified among 90 genomes with the phylogenetic analysis. The phylogenetic tree indicated these 90 genomes exhibited a high intra-species genomic diversity. No clustering was assorted with the host or geographic location among these genomes. Aminoglycoside resistance genes, such as , , , , and streptothricin resistance gene were detected in the chromosomes from a third of the Chinese strains. Virulence-related genes were identified in all the sequenced strains. A novel large multiple drug-resistant plasmid (named pCNAC48 with 161,992 bp in length) was identified in strain ICDCAC48. Two antibiotic-resistance islands were found in the plasmid with lengths of 7,950 and 25,137 bp and G + C content of 38.23 and 32.39%, respectively. The drug resistance genes and some transposable elements were cross-distributed among the islands in the plasmid. Antimicrobial susceptibility tests indicated these resistance genes in the plasmid were functional. Plasmid conjugation and curing experiments proved pCNAC48 was stable in strain ICDCAC48. It was the first identified multiple drug resistance plasmid in .
PubMed: 36212879
DOI: 10.3389/fmicb.2022.984450 -
Journal of Food Protection Jan 2021Arcobacter species are gram-negative rods that have been implicated in food- and waterborne illness. Although various cultural isolation methods have been proposed, the...
ABSTRACT
Arcobacter species are gram-negative rods that have been implicated in food- and waterborne illness. Although various cultural isolation methods have been proposed, the current procedures are unable to fully suppress the growth of background microbiota present in food samples, which inhibits Arcobacter isolation. The purpose of this study was to develop a selective enrichment broth and chromogenic plating medium to detect three Arcobacter species that have been recognized as emerging foodborne pathogens: Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii. The developed Nguyen-Restaino-Juárez (NRJ) Arcobacter detection system consists of a selective enrichment broth (NRJ-B) and a selective-differential plating medium (NRJ-M). The protocol of the detection method was determined by evaluating the growth of A. butzleri, A. cryaerophilus, and A. skirrowii under various temperatures (30, 35, and 42°C) and incubation (aerobic, microaerophilic, and anaerobic) conditions. Additionally, 47 Arcobacter strains and 39 non-Arcobacter strains were tested in inclusivity and exclusivity evaluations of NRJ-B and NRJ-M. Overall, the study determined that the optimal growth conditions of Arcobacter species using the NRJ Arcobacter detection system were aerobic incubation at 30°C. NRJ-B supported good growth of A. butzleri, A. cryaerophilus, and A. skirrowii while effectively suppressing the growth of non-Arcobacter strains after 48 h. Furthermore, NRJ-M yielded 97.8% inclusivity and 100.0% exclusivity using the tested strains and resulted in salmon-pigmented Arcobacter colonies (1.0 to 1.5 mm in diameter) after 72 h. The novel protocol is the first to develop a chromogenic plating medium for the isolation of Arcobacter species. This simple and accurate test method would greatly contribute to understanding the distribution of pathogenic Arcobacter species in food samples.
Topics: Agar; Arcobacter; Culture Media
PubMed: 33411930
DOI: 10.4315/JFP-20-245 -
Veterinary Medicine and Science Jul 2022Arcobacter spp. has been considered an emerging foodborne pathogen and a hazard to human health. The dairy chain has been isolated from different sources; nevertheless,...
BACKGROUND
Arcobacter spp. has been considered an emerging foodborne pathogen and a hazard to human health. The dairy chain has been isolated from different sources; nevertheless, data on Arcobacter occurrence in raw milk and dairy products in Iran are still scant.
OBJECTIVE
The present study investigates the prevalence, antimicrobial susceptibility and the presence of virulence genes of Arcobacters species isolated from milk and dairy products.
METHODS
Then, a total of 350 raw milk samples and 400 dairy product samples were collected from dairy supply centers in Isfahan, Iran. Presumptive Arcobacter strains were obtained by enriching samples in Oxoid Arcobacter enrichment broth (AEB) followed by the filtration of enrichment product through 0.45-μm pore size membrane filters laid onto non-selective blood at 30°C under microaerophilic conditions. Molecular identification of Arcobacter cryaerophilus and A. butzleri was performed by Polymerase chain reaction (PCR) amplification of the 16S rRNA gene, followed by sequencing. The disc diffusion method was used to determine the antimicrobial susceptibility of isolates. Targeted resistance and virulence genes were detected using multiplex PCR.
RESULTS
The results show a low recovery rate of Arcobacter spp. in milk. Arcobacters were found in all types of milk, except raw camel milk, but were absent from all dairy products. Arcobacter butzleri was the predominant species in raw milk. Detection of virulence genes shows that all virulence genes targeted were found among A. butzleri, and six (cadF, cj1349, irgA, mviN, pldA, tlyA) were found among A. cryaerophilus. All A. butzleri strains and some A. cryaerophilus strains isolated from milk were resistant to amoxicillin-clavulanic acid and tetracycline. All A. cryaerophilus isolates from milk were susceptible to gentamycin, streptomycin, erythromycin and ciprofloxacin. The distribution of resistance genes in Arcobacter strains in milk shows that all isolates carried tet(O) and bla genes.
CONCLUSIONS
In conclusion, the results indicate a low recovery rate of Arcobacter spp. in milk and milk products. However, a significant number of Arcobacter strains with putative virulence genes may be potential pathogens for humans and an overall increase in Arcobacter resistance to first-line antibiotics. These results highlight the need for regular surveillance of Arcobacter strains in milk and milk products in Iran.
Topics: Animals; Anti-Bacterial Agents; Arcobacter; Drug Resistance, Microbial; Genotype; Humans; Milk; Multiplex Polymerase Chain Reaction; Prevalence; RNA, Ribosomal, 16S; Virulence Factors
PubMed: 35426255
DOI: 10.1002/vms3.800 -
Antibiotics (Basel, Switzerland) Mar 2021spp. are emerging waterborne and foodborne zoonotic pathogens responsible for gastroenteritis in humans. In this work, we evaluated the occurrence and the antimicrobial...
spp. are emerging waterborne and foodborne zoonotic pathogens responsible for gastroenteritis in humans. In this work, we evaluated the occurrence and the antimicrobial resistance profile of isolates recovered from different aquatic sources. Besides, we searched for spp. in seaweeds and the corresponding seawater samples. Bacteriological and molecular methods applied to 100 samples led to the isolation of 28 isolates from 27 samples. The highest prevalence was detected in rivers followed by artificial ponds, streams, well waters, and spring waters. Seaweeds contained a higher percentage of than the corresponding seawater samples. The isolates were identified as (96.4%) and (3.6%). All the isolates showed a multi-drug resistance profile, being resistant to at least three different classes of antibiotics. Molecular analysis of genetic determinants responsible for tetracycline resistance in nine randomly chosen isolates revealed the presence of and/or This work confirms the occurrence and the continuous emergence of antibiotic-resistant strains in environmental samples; also, the presence of quinolone-resistant spp. in aquatic sources used for water supply and irrigation represents a potential risk for human health.
PubMed: 33802125
DOI: 10.3390/antibiotics10030288 -
Journal of Food Protection Mar 2023Arcobacters are emerging pathogens that have been underestimated due to a lack of a standardized isolation method. The aim of this research was to evaluate the ability...
Arcobacters are emerging pathogens that have been underestimated due to a lack of a standardized isolation method. The aim of this research was to evaluate the ability to isolate Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii using two Arcobacter-specific culture detection systems: (i) the Houf broth and modified charcoal cefoperazone deoxycholate agar supplemented with cefoperazone, amphotericin B, and teicoplanin (HB/mCCDA+CAT), and (ii) the Nguyen-Restaino-Juárez Arcobacter enrichment broth and chromogenic agar (NRJ-B/M). Both detection systems were evaluated for productivity ratio, sensitivity, and specificity. As a result, the productivity ratio for both plating agars were >90%, which indicates that the selective agents used in the two plating agars did not inhibit Arcobacter growth. Moreover, sensitivity evaluations using artificially inoculated retail ground poultry (n = 780) determined that both detection systems were able to isolate A. butlzeri with >95% sensitivity at the 0.1 and 1.0-2.0 CFU/g detection level. The sensitivity in A. cryaerophilus isolation was higher for NRJ-B/M (78.0% at 0.1 CFU/g; 95.1% at 1.0-2.0 CFU/g) when compared with HB/mCCDA+CAT (34.1% at 0.1 CFU/g; 51.2% at 1.0-2.0 CFU/g). Both detection systems resulted in <50% sensitivity when isolating A. skirrowii at 0.1 and 1.0-2.0 CFU/g; however, the sensitivity for NRJ-B/M was significantly higher than HB/mCCDA+CAT. At the detection level of 5.0 CFU/g, both detection systems were able to isolate A. skirrowii with 100% sensitivity. Specificity comparisons using uninoculated ground poultry samples (n = 40) indicated the growth of background microbiota were significantly inhibited or could be easily differentiated on NRJ-B/M (90.0%, specificity) when compared with HB/mCCDA+CAT (30.0%, specificity). Overall, these results show that the NRJ-B/M detection system is a more sensitive and specific detection system when isolating Arcobacter spp. from ground chicken.
Topics: Animals; Poultry; Arcobacter; Agar; Cefoperazone
PubMed: 36916562
DOI: 10.1016/j.jfp.2023.100057 -
Italian Journal of Food Safety Mar 2021Given that the number of foodborne illness outbreaks linked to the consumption of ready-to-eat vegetables has been widely documented and considering that data on the...
Given that the number of foodborne illness outbreaks linked to the consumption of ready-to-eat vegetables has been widely documented and considering that data on the occurrence of spp. in such foodstuffs are lacking, the aim of the present study was to evaluate the presence of spp. and the occurrence of virulence factors as well as to genotype spp. in ready-to-eat (RTE) vegetable samples, using cultural and biomolecular assays. spp. was detected in 16/110 (14.5%) samples, with being detected in 15/16 and in 1/16 isolates. PCRs aimed at the nine putative virulence genes demonstrated widespread distribution of such genes among and isolates. In addition, multilocus sequence type (MLST) analysis revealed a low genetic diversity within the arcobacters isolates. The results underline the need to develop an appropriate surveillance system based on biomolecular characterization for an integrated microbiological risk assessment of ready-toeat vegetables, and consequently of composite foods.
PubMed: 33907683
DOI: 10.4081/ijfs.2021.8585 -
Frontiers in Microbiology 2020(formerly ) is a globally emerging foodborne and zoonotic pathogen. However, little is known about the species' genomic features and diversity, antibiotic resistance...
(formerly ) is a globally emerging foodborne and zoonotic pathogen. However, little is known about the species' genomic features and diversity, antibiotic resistance and virulence. In this study, 27 strains from water poultry in Thuringia, Germany, were investigated using whole-genome sequencing. Four of these strains were sequenced using long- and short-read sequencing methods to obtain circularized genomes. The German strains belong to the cluster I. Cluster I genomes exhibited a high degree of genetic diversity in which variable sites comprised 9.1% of the core genome. The German strains formed three subgroups that contained 2, 6, and 9 strains, respectively. The genomic analysis of cluster I revealed variable presence of mobile elements and that 65% of the strains lack CRISPR systems. The four circularized genomes carried a ∼2 Mbp chromosome and a single megaplasmid (size 98.1-154.5 Kbp). The chromosome was densely packed with coding sequences (∼92%) and showed inversions and shifts in the gene blocks between different strains. Antimicrobial resistance was assessed using a gradient strip diffusion method and showed that all 27 strains were resistant to cefotaxime and susceptible to erythromycin, gentamicin, and ampicillin. Sixteen strains were also resistant to ciprofloxacin, whereas 23 were resistant to streptomycin. The genetic prediction of antibiotic resistance identified numerous efflux pumps similar to those found in . All strains harbored two beta-lactamase genes which may explain the cefotaxime resistance. A correlation between the A point mutation (Thr-85-Ile) and ciprofloxacin resistance was partially discovered in 15 out of 16 strains. virulence profiling showed a wide range of virulence factors including a full chemotaxis system and most of the flagellar genes. In contrast to , no urease cluster was found. This study provides new insights into the genomic variability of strains of cluster I. The different genetic makeup of these strains may contribute to the virulence of strains and the severity of the infections in humans.
PubMed: 32754133
DOI: 10.3389/fmicb.2020.01549 -
Journal of Food Protection Dec 2020Arcobacter is considered an emergent foodborne enteropathogen. Despite the high prevalence of this genus in poultry, the occurrence of Arcobacter spp. contamination in...
ABSTRACT
Arcobacter is considered an emergent foodborne enteropathogen. Despite the high prevalence of this genus in poultry, the occurrence of Arcobacter spp. contamination in Tunisia remains unclear. The objectives of this study were (i) to isolate Arcobacter species (A. butzleri and A. cryaerophilus) by the culture method from different species of raw poultry meat, (ii) to verify the isolates by multiplex PCR (m-PCR) assay and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), and (iii) to determine the antibiotic resistance profiles of the isolates. A total of 250 poultry product samples (149 chicken and 101 turkey) were collected from various supermarkets in Sfax. The samples consisted of breasts, wings, legs, and neck skins. The overall isolation frequency of Arcobacter spp. was 10.4%. Arcobacter spp. were found in 13.42% of the chicken samples and in 5.49% of the turkey samples. All the acquired isolates were subject to detailed confirmation with subsequent species classification using m-PCR and MALDI-TOF MS. A. butzleri was found in 22 samples (84.61%) and A. cryaerophilus in 4 samples (15.38%). Thus, m-PCR and MALDI-TOF MS were able to detect A. butzleri significantly better than the conventional method (χ2 = 49.1 and P < 0.001). Arcobacter was isolated from poultry in every season, at contamination levels of 30.76, 23.07, 19.23, and 26.92% in summer, spring, autumn, and winter, respectively. The disk diffusion method was used to determine the susceptibility of Arcobacter isolates to six antimicrobial drugs. All A. butzleri isolates (n = 24) were significantly resistant to erythromycin (P = 0.0015), ampicillin (P = 0.001), and ciprofloxacin (P = 0.05). All tested A. cryaerophilus strains (n = 4) were susceptible to ampicillin, gentamicin, and amoxicillin-clavulanic acid. Multidrug resistance was observed in 83% of the Arcobacter spp. isolates. Our study detected Arcobacter spp. in Tunisian poultry; because of their multidrug resistance, these species may constitute a public health problem.
Topics: Animals; Arcobacter; Drug Resistance, Microbial; Food Microbiology; Poultry; Tunisia
PubMed: 32634222
DOI: 10.4315/JFP-20-056 -
Frontiers in Microbiology 2022spp. is a globally emerging zoonotic and foodborne pathogen. However, little is known about its prevalence and antimicrobial resistance in China. To investigate the...
spp. is a globally emerging zoonotic and foodborne pathogen. However, little is known about its prevalence and antimicrobial resistance in China. To investigate the prevalence of spp. isolated from various sources, 396 samples were collected from human feces, chicken cecum, and food specimens including chicken meat, beef, pork, lettuce, and seafood. spp. was isolated by the membrane filtration method. For 92 strains, the agar dilution method and next-generation sequencing were used to investigate their antimicrobial resistance and to obtain whole genome data, respectively. The virulence factor database (VFDB) was queried to identify virulence genes. ResFinder and the Comprehensive Antibiotic Resistance Database (CARD) were used to predict resistance genes. A phylogenetic tree was constructed using the maximum likelihood (ML) method with core single-nucleotide polymorphisms (SNPs). We found that 27.5% of the samples ( = 109) were positive for spp., comprising (53.0%), (39.6%), and (7.4%). Chicken meat had the highest prevalence (81.2%), followed by seafood (51.9%), pork (43.3%), beef (36.7%), lettuce (35.5%), chicken cecum (8%), and human fecal samples (0%, 0/159). Antimicrobial susceptibility tests revealed that 51 and 40 strains were resistant to streptomycin (98.1, 70%), clindamycin (94.1, 90%), tetracycline (64.7, 52.5%), azithromycin (43.1%, 15%), nalidixic acid (33.4, 35%), and ciprofloxacin (31.3, 35%) but were susceptible to erythromycin, gentamicin, chloramphenicol, telithromycin, and clindamycin (≤10%). was sensitive to all experimental antibiotics. The virulence factors A, , and were carried by all spp. strains at 100%, and the following percentages were (95.7%), (23.9%), 2.2%, and (1.1%). Only one strain (F061-2G) carried a macrolide resistance gene (). One and one harbored resistance island gene clusters, which were isolated from pork and chicken. Phylogenetic tree analysis revealed that , and were separated from each other. To our knowledge, this is the first report of the isolation of spp. from vegetables and seafood in China. The resistance island gene cluster found in pork and chicken meat and the presence of virulence factors could be a potential risk to human health.
PubMed: 36532418
DOI: 10.3389/fmicb.2022.1004224 -
Applied and Environmental Microbiology Apr 2020Pathogenic bacteria in wastewater are generally considered to be efficiently removed in biological wastewater treatment plants. This understanding is almost solely based...
Pathogenic bacteria in wastewater are generally considered to be efficiently removed in biological wastewater treatment plants. This understanding is almost solely based on culture-based control measures, and here we show, by applying culture-independent methods, that the removal of species in the genus was less effective than for many other abundant genera in the influent wastewater. was one of the most abundant genera in influent wastewater at 14 municipal wastewater treatment plants and was also abundant in the "clean" effluent from all the plants, reaching up to 30% of all bacteria as analyzed by 16S rRNA gene amplicon sequencing. Metagenomic analyses, culturing, genome sequencing of isolates, and visualization by fluorescent hybridization (FISH) confirmed the presence of the human-pathogenic and in both influent and effluent. The main reason for the high relative abundance in the effluent was probably that cells, compared to those of other abundant genera in the influent, did not flocculate and attach well to the activated sludge flocs, leaving a relatively large fraction dispersed in the water phase. The study shows there is an urgent need for new standardized culture-independent measurements of pathogens in effluent wastewaters, e.g., amplicon sequencing, and an investigation of the problem on a global scale to quantify the risk for humans and livestock. The genus was unexpectedly abundant in the effluent from 14 Danish wastewater treatment plants treating municipal wastewater, and the species included the human-pathogenic and Recent studies have shown that is common in wastewater worldwide, so the study indicates that discharge of members of the genus may be a global problem, and further studies are needed to quantify the risk and potentially minimize the discharge. The study also shows that culture-based analyses are insufficient for proper effluent quality control, and new standardized culture-independent measurements of effluent quality encompassing most pathogens should be considered.
Topics: Arcobacter; Denmark; RNA, Bacterial; RNA, Ribosomal, 16S; Waste Disposal, Fluid; Wastewater
PubMed: 32111585
DOI: 10.1128/AEM.03044-19