Occurrence of genes encoding aminoglycoside-modifying enzymes in Escherichia coli isolates from chicken meat

ABSTRACT

1. The aim of the experiment was to determine the occurrence of genes encoding aminoglycoside-modifying enzymes (AMEs) in Escherichia coli isolates recovered from chicken meat.

2. Antibiotic sensitivity was tested using the disc diffusion test. AMEs and virulence profile were determined by PCR/sequencing.

3. Out of 195 meat samples collected, 185 (95%) isolates were identified as E. coli. Disc diffusion showed a resistance value of 22% (n = 42) for at least one of the antibiotic aminoglycosides (AGs) tested (tobramycin, gentamycin, amikacin and kanamycin). PCR screening showed the presence of three classes of AMEs, namely, aac(3)-II (12%), aac(6?)-Ib (7%) and aac(2?)-Ia (5%). Eight of the 42 isolates were positive for the stx₁ and sxt₂ genes and were defined as Shiga toxin-producing E coli., while the eae gene was positive in one strain. Among the 42 isolates, group A was the predominant phylogenetic identified (76%), followed by group D (21%). One isolate belonged to subgroup B2₃.

4. The results suggested that chicken meat could be an important reservoir of AMEs, and pose a potential risk by dissemination of resistance to humans through the food chain.     

微卫星DNA标记与鸡肉品质性状的相关分析

本研究测定了隐性白羽和仙居鸡参考家系F2代500羽个体的26个微卫星座位的多态性,分析微卫星标记与鸡肉品质性状之间的相关性,结果发现:位于2号染色体上的标记ADL212分别与pH值和嫩度显著相关;位于3号和5号染色体上的微卫星标记MCW 004和MCW 223与肉色显著相关;与胸肌失水率显著相关的微卫星标记分别为:位于2号染色体的MCW 264和ADL212以及5号染色体的MCW 223和9号染色体的ADL211。     

Antimicrobial resistance and virulence profiles of Enterococcus spp. isolated from horses in korea

Antimicrobial-resistant (AR) enterococci have emerged as leading nosocomial pathogens. Transmission of AR Enterococci from animals to humans has been demonstrated. However, there is limited information on the transmission of enterococci from horses to humans. To address this issue, we characterized 260 enterococci isolated from horse-associated samples in Korea in 2013 based on their AR profiles and virulence traits. AR profiling revealed an average ratio of AR enterococci of 23.8%. Seven isolates (2.7%) were multidrug-resistant Enterococcus faecalis. Most tetracycline-resistant enterococci harbored either tetM or tetL or both genes; genes conferring resistance to other antimicrobials were detected at low rates. Biofilm formation and gelatinase activity were observed in 51.1% and 47.7% of isolates, respectively; most were E. faecalis harboring the gelE gene. Evidence of transmission of AR enterococci between horses and their environments was provided by pulsed-field gel electrophoresis, and highlights the risk of AR enterococcus transmission to horse riders and handlers through close contact.     

Detection of a novel clone of Acinetobacter baumannii isolated from a dog with otitis externa

In this study, the isolation of Acinetobacter baumannii in a dog with clinical bilateral otitis externa is described. Moreover, to investigate the zoonotic potential of the isolate, microbiological examinations on the family members were performed. An A. baumanniistrain was isolated from nasal swab in one of the dog owners. The identity of bacterial strains, either from dog and owner, was confirmed by phenotypic and molecular typing (wgMLST). Furthermore, to assess the pathogenic potential of the isolates a deep characterization of virulence and antibiotic resistance genes was done by Whole Genome Sequencing (WGS). Finally, the susceptibility towards a wide panel of antimicrobials was investigated. In our knowledge, this is the first recorded case of A. baumanniiisolation from canine auricular swabs in Italy. And interestingly, this study underlines the possible spread of this microorganism from human to animal.     

Extraintestinal pathogenic Escherichia coli in poultry meat products on the Finnish retail market

Background

Extraintestinal pathogenic Escherichia coli bacteria (ExPEC) exist as commensals in the human intestines and can infect extraintestinal sites and cause septicemia. The transfer of ExPEC from poultry to humans and the role of poultry meat as a source of ExPEC in human disease have been discussed previously. The aim of the present study was to provide insight into the properties of ExPEC in poultry meat products on the Finnish retail market with special attention to their prevalence, virulence and phylogenetic profiles. Furthermore, the isolates were screened for possible ESBL producers and their resistance to nalidixic acid and ciprofloxacin was tested.

Methods

The presence of ExPEC in 219 marinated and non-marinated raw poultry meat products from retail shops has been analyzed. One E. coli strain per product was analyzed further for phylogenetic groups and possession of ten virulence genes associated with ExPEC bacteria (kpsMT K1, ibeA, astA, iss, irp2, papC, iucD, tsh, vat and cva/cv) using PCR methods. The E. coli strains were also screened phenotypically for the production of extended-spectrum β-lactamase (ESBL) and the susceptibility of 48 potential ExPEC isolates for nalidixic acid and ciprofloxacin was tested.

Results

E. coli was isolated from 207 (94.5%) of 219 poultry meat products. The most common phylogenetic groups were D (50.7%), A (37.7%), and B2 (7.7%). Based on virulence factor gene PCR, 23.2% of the strains were classified as ExPEC. Two ExPEC strains (1%) belonged to [O1] B2 svg+ (specific for virulent subgroup) group, which has been implicated in multiple forms of ExPEC disease. None of the ExPEC strains was resistant to ciprofloxacin or cephalosporins. One isolate (2.1%) showed resistance to nalidixic acid.

Conclusions

Potential ExPEC bacteria were found in 22% of marinated and non-marinated poultry meat products on the Finnish retail market and 0.9% were contaminated with E. coli [O1] B2 svg+ group. Marinades did not have an effect on the survival of ExPEC as strains from marinated and non-marinated meat products were equally often classified as ExPEC. Poultry meat products on the Finnish retail market may have zoonotic potential.     

鸡钙蛋白酶抑制蛋白基因多态性及其与肉质性状的相关性

为探讨钙蛋白酶抑制蛋白(Calpastatin,CAST)基因作为影响鸡肉质性状候选基因的可能性,本研究以8个优质鸡新品系为试验材料,采用PCR-SSCP方法对CAST基因的部分编码区进行单核苷酸多态性(SNPs)检测。结果发现了CAST基因1个多态性位点(G→T),使试验鸡群表现出AA、BB和AB这3种基因型。利用SAS(8.01)软件统计分析了3种基因型与优质鸡肉质性状的相关性,结果表明,CAST基因的多态性对鸡肌内脂肪含量、肌纤维密度和直径有显著的影响(P0.05),对其他肉质性状无显著影响(P0.05);推测CAST基因可能对鸡肉质性状具有很大的影响或与控制肉质性状的主基因连锁。     

Molecular and virulence characteristics of multi-drug resistant Salmonella Enteritidis strains isolated from poultry

Forty-six Salmonella enterica subspecies enterica serovar Enteritidis (S. Enteritidis) strains were isolated from chicken meat, faeces, and eggshells collected from hatcheries throughout Korea. The strains were examined for the presence of antimicrobial resistance and virulence genes. All 46 isolates were resistant to at least one of 21 antibiotics used in this study, 30 (65.2%) were resistant to three or more antimicrobials, and a single remarkable isolate was resistant to 15 antimicrobials. The isolates were primarily resistant to penicillins, sulfisoxazole, streptomycin, tetracycline and quinolones.The high rate of resistance in S. Enteritidis strains, sometimes to multiple drugs, may complicate future options for treating human infections. Nineteen of the 21 penicillin resistant isolates carried the bla_TEM gene, while one strain, resistant both to penicillins and ceftriaxone, carried the bla_CTX-M gene. Thirty-seven of the 45 sulfisoxazole resistant isolates carried sul2, and 23/24 streptomycin resistant isolates carried both strA and strB. All 10 tetracycline resistant isolates carried the tet(A) gene. Most isolates harboured both SPI-1 and SPI-2-associated genes, and the spv operon, which are known to be associated with human infections. The presence of these genes suggests that these strains could give rise to public health problems if dispersed in the general human population.     

The role of Acinetobacter baumannii as a nosocomial pathogen for dogs and cats in an intensive care unit

Acinetobacter baumannii is a nosocomial pathogen associated with high morbidity and mortality in humans. Whereas infections with strains of Acinetobacter species have been reported in various situations, the importance of A baumannii as a nosocomial pathogen in veterinary hospitals has not been studied so far. In this retrospective case series, we describe 17 dogs and 2 cats from which A baumannii had been isolated during a 21/2-year period. In 7 dogs, A baumannii induced systemic signs of illness, whereas 12 animals showed signs of local infection. In all animals with systemic infection, and in 2 with localized infection, A baumannii contributed to the death of the animal or contributed to euthanasia; the remaining 8 dogs and both cats recovered. Molecular typing of the isolates with restriction polymorphisms of ribosomal DNA provided evidence of nosocomial spread of this pathogen and for the presence of several strains of A baumannii in the hospital environment.     

Effects of different rearing systems on meat production traits and meat fiber microstructure of Beijing‐you chicken

Beijing‐you is a Chinese local chicken which is raised for both meat and eggs. In the present study, we detected the effects of different rearing systems on growth, slaughtering performances and meat quality of Beijing‐you chickens at 26–40 weeks of age. Six hundred Beijing‐you hens were randomly allocated into two groups at 16 weeks of age and raised in free range or battery cage systems. The body weight, slaughtering performance and meat quality were measured for each group at the ages of 26, 30, 35 and 40 weeks. Some of the traits were dramatically influenced by the two systems, although most of them did not show significant changes. For the meat fiber microstructure, we found that the diameter of thigh and breast muscle fiber in the free range group were significantly increased than in the cage group (P < 0.05) at 26 weeks of age. The ratio of fast muscle fiber in thigh muscle samples of the free range group was significantly reduced compared to that of cage group at both 35 (P < 0.01) and 40 (P < 0.01) weeks of age, indicating that the free range system could promote the transforming of fast muscle fiber to slow muscle fiber.     

禽蛋和禽肉中兽药残留检测技术研究进展

随着我国家禽产业的规模不断扩大,兽药残留问题也更加严峻。兽药残留对人体与环境危害重大。现对目前禽蛋和禽肉中常见的兽药残留种类及最大残留限量进行综述,探讨了(超)高效液相色谱法、(超)高效液相色谱-质谱法、气相色谱-质谱法和免疫学检测法、分子印迹技术、生物芯片技术、生物传感器检测法在禽蛋和禽肉中的应用价值,以期为兽药残留检测和监管提供依据。     

江苏某牛场生牛乳中肺炎克雷伯菌和鲍曼不动杆菌的分离鉴定、耐药性和毒力分析

肺炎克雷伯菌和鲍曼不动杆菌均是重要的条件致病菌,其具备的多重耐药性是造成医院内感染并难以治愈的主要原因。本试验从江苏某奶牛场隐性乳房炎奶牛的生牛乳样品中分离到3株肺炎克雷伯菌及4株鲍曼不动杆菌,通过耐药性检测发现其中部分菌株具有对万古霉素、克林霉素、头孢唑啉和呋喃妥因等抗生素的多重耐药性;小鼠毒力试验表明分离菌株具备较强的毒力。本试验为奶牛场预防和治疗该类病原感染提供参考依据,同时对耐药菌株可能引起的公共卫生安全问题提出警示。     

Characterisation of avian pathogenic Escherichia coli (APEC) associated with colisepticaemia compared to faecal isolates from healthy birds

A total of 114 avian pathogenic Escherichia coli (APEC) isolates were collected from cases of colisepticaemia occurring in broilers (77) and layers (37) within Ireland. In addition 45 strains isolated from faeces of healthy birds were included for comparison. All isolates were serogrouped, and examined for known virulence factors, mostly by PCR. The O78 serogroup represented 55 and 27% of broiler and layer colisepticaemic isolates respectively. All isolates were positive for curli fimbriae (crl, csg) and negative for afimbrial adhesin (afa). S-fimbrial (sfa) sequences were present in 8.8% of septicaemic isolates and 8.9% of healthy bird isolates. The majority of E. coli from cases of colisepticaemia (97.4%) and healthy bird (95.6%) isolates were positive for aerobactin (aer), and temperature sensitive haemagglutinin (tsh) was similarly detected in high numbers in 93.9 and 93.3%, respectively. In comparison to E. coli isolates from the faeces of healthy birds, a significantly higher percentage of isolates from septicaemic cases possessed Type 1 fimbriae (fimC) and increased serum survival (iss) gene sequences. Forty-seven (41.2%) isolates from septicaemic birds possessed P-fimbriae (pap) gene sequences, compared with only 15.6% from E. coli isolated from healthy birds. Haemolysin (hlyE) sequences were detected in 46.7% of isolates from healthy birds in comparison with 6.1% of septicaemic isolates. Sequences encoding colicin V (cvaC) were detected in 99.1% of septicaemic isolates and 82.2% of isolates from healthy birds. The K1 capsule was only present in two septicaemic isolates, both taken from layers. Motility was detected in 36.8% of E. coli isolated from cases of septicaemia, compared with 93.3% of isolates from healthy birds. These results demonstrate the presence of 11 virulence genes in E. coli isolated from cases of colisepticaemia within Ireland, and indicate the prevalence of iss and fimC.     

Frequency of virulence factors in Escherichia coli isolated from suckling pigs with diarrhoea in China

Escherichia coli-associated diarrhoea is an important disease adversely affecting the pig industry. This study was conducted to investigate the frequency of virulence factors expressed by E. coli strains isolated from suckling pigs with diarrhoea in China. A total of 381 E. coli strains, obtained from 290 faecal samples from pigs on 38 farms, were tested for fimbriae (K88, K99, 987P, F41, F18, F17), non-fimbrial adhesins (AIDA-I, paa, CS31A, eae, saa), enterotoxin (LT-I, LT-II, STa, STb, EAST1), Shiga toxin (Stx1, Stx2, Stx2e), pathogenicity islands (HPI, LEE), α-haemolysin (hlyA), afa8 gene cluster (afaD, afaE) and sepA genes by PCR. Out of the 381 isolates, 206 carried at least one virulence gene. Of the 206 virulence positive isolates, the virulence factor genes detected were EAST1 (n = 120), irp2 (n = 59), paa (n = 50), STb (n = 41), AIDA-I (n = 34), LT-I (n = 23), ler (n = 11), hlyA (n = 9), K88 (n = 8), eae (n = 8), STa (n = 7), sepA (n = 6), F18 (n = 5), afaD (n = 3), afaE (n = 3), K99 (n = 2) and Stx2e (n = 1), with most isolates carrying multiple virulence genes. These results demonstrate that relatively few isolates from the study population express K88, K99, LT-I or STa, but that EAST1 (58%), irp2 (29%), AIDA-I (16.5%), paa (24%) and STb (20%) are frequent virulence factors expressed by E. coli strains isolated from suckling pigs with diarrhoea in China.     

Population structure and virulence content of avian pathogenic Escherichia coli isolated from outbreaks in Sri Lanka

Avian pathogenic Escherichia coli (APEC) causes economically significant infections in poultry. The genetic diversity of APEC and phylogenetic relationships within and between APEC and other pathogenic E. coli are not yet well understood. We used multilocus sequence typing (MLST), PCR-based phylogrouping and virulence genotyping to analyse 75 avian E. coli strains, including 55 isolated from outbreaks of colisepticaemia and 20 from healthy chickens. Isolates were collected from 42 commercial layer and broiler chicken farms in Sri Lanka. MLST identified 61 sequence types (ST) with 44 being novel. The most frequent ST, ST48, was represented by only six isolates followed by ST117 with four isolates. Phylogenetic clusters based on MLST sequences were mostly comparable to phylogrouping by PCR and MLST further differentiated phylogroups B1 and D into two subgroups. Genotyping of 16 APEC associated virulence genes found that 27 of the clinical isolates and one isolate from a healthy chicken belonged to highly virulent genotype according to previously established classification schemes. We found that a combination of four genes, ompT, hlyF, iroN and papC, gave a comparable prediction to that of using five and nine genes by other studies. Four STs (ST10, ST48, ST117 and ST2016) contained APEC isolates from this study and human UPEC isolates reported by others, suggesting that these STs are potentially zoonotic. Our results enhanced the understanding of APEC population structure and virulence association.     

Molecular detection of Coxiella burnetii in raw meat intended for pet consumption

The discovery of antibodies against Coxiella burnetii in cattery‐confined breeding cats indicating prior or current exposure (Shapiro et al., 2015) prompted an investigation into possible sources of infection. One hypothesis was that raw meat diets containing reservoir species may provide a source of C. burnetii transmission. The aim of this pilot study was to determine whether C. burnetii DNA was present in raw meat sold exclusively for companion animal consumption. The sample population consisted of raw meat packages (n = 58) of primarily kangaroo origin, with three to four aliquots (50–120 mg) randomly selected from each package. Genomic DNA was extracted from whole tissue in each of these aliquots using a modified protocol. Three quantitative PCR assays were used for the detection of C. burnetii targeting the IS1111 gene, the heat shock operon htpAB and the C. burnetii outer membrane protein‐coding gene, com1. Coxiella burnetii DNA was detected in 25/58 samples (43%) using the IS1111, htpAB and/or com1 PCR assays and confirmed by DNA sequencing. All samples amplifying a product in the com1 assay also amplified a product in the htpAB and IS1111 assays. A total of 17/58 (29%) packets were positive with all three genes, 4/58 (7%) were positive with two genes (IS1111 and htpAB) and 4/58 (7%) were positive with the IS1111 gene only. Coxiella burnetii DNA was five times more likely to be found in offal than skeletal muscle meat samples. All meat samples in which C. burnetii DNA was found were from kangaroo tissues, while samples labelled as non‐kangaroo meat (n = 4) were negative. Multi‐locus variable number of tandem repeat analysis (MLVA) identified three different genotypes of C. burnetii that have all been identified previously from Australian human clinical Q fever cases. Further investigations are required to determine the potential role of certain raw meats in the transmission of C. burnetii to cats and humans.     

河西走廊地区犊牛腹泻致病性大肠杆菌分离鉴定、毒力因子及耐药性检测

[目的]为了分析河西走廊地区犊牛腹泻致病性大肠杆菌携带毒力基因和耐药性情况,[方法]2020—2021年在河西走廊地区采集患腹泻病犊牛的粪便、肛拭子及肝脏等病料组织279份,采用人工感染试验动物、PCR方法和K-B药敏纸片法分别检测犊牛腹泻性大肠杆菌致病性、毒力因子和耐药性。[结果]结果表明,分离得到了126株大肠杆菌,其中79株犊牛腹泻性大肠杆菌能引起小鼠死亡;分离的79株致病性大肠杆菌的毒力基因crl、irp2、fimH、papC、K88、K99、stx1、stx2检测率在40.5%～100%之间,其他毒力基因检测率在15.2%～34.2%之间;分离的79株致病性大肠杆菌对氨苄西林、阿莫西林、新霉素等8种药物的耐药率在49.4%～96.2%之间,对其他药物的耐药率在5.1%～32.9%之间。[结论]从河西走廊地区患腹泻病犊牛病料组织中分离得到79株致病性大肠杆菌,这些菌株携带多种毒力基因,对临床中常用的抗菌药物产生了耐药性。