背景:弯曲杆菌属。根据世界卫生组织(WHO),是细菌性食源性胃肠炎的最常见原因,也是高度优先考虑的抗生素抗性细菌。欧洲对耐热弯曲杆菌的监测。并不能反映全球细菌种群中已经循环的耐药性的全球负担。
方法:我们系统地比较了全基因组测序与综合表型抗菌素敏感性,分析来自越南和德国的494株耐热弯曲杆菌家禽分离物。通过重复湿实验室和改进干实验室部分来检查任何差异。通过长期阅读的牛津纳米孔技术对选定的分离株进行了额外分析,导致封闭的染色体和质粒。
结果:总体而言,22个不同的抗性基因和基因变体(e。g.erm(B),aph(3')-IIIa,aph(2\'\')-如果,CatA,lnu(C),blaOXA,sat4)和三个不同基因(gyrA,23SrRNA,与AMR相关的rpsL)存在于弯曲杆菌分离物中。数据库中缺少两个AMR基因,一个与抗性错误相关。基于短读数据的生物信息学分析部分未能识别tet(O)和aadE,当基因作为重复或同源基因变体存在时。有趣的是,分离株还含有不同的决定因素,多余地赋予氯霉素抗性,庆大霉素,卡那霉素,林可霉素和链霉素。我们在四环素敏感菌株中发现了一个新的tet(W),窝点突变。此外,由于基因内poly-C束的变化,基于短读数据装配的分析受损,无法识别全长相变量aad9。无法鉴定来自德国的一种分离株的庆大霉素抗性的遗传决定子。GyrT86I,(氟)喹诺酮耐药的主要决定因素导致环丙沙星耐药的罕见非典型表型,但萘啶酸敏感性。长读测序预测AMR基因主要位于染色体上,很少在质粒上。来自长读和短读测序的预测,分别,经常不同。AMR基因通常组织在多药耐药岛(MDRI)中,部分位于转座酶基因附近,表明抗性决定子的主要动员是通过弯曲杆菌的自然转化和转座。
结论:这项研究的结果表明,存在频繁的抗性基因重复,马赛克,和突变导致基因变异和截短的弯曲杆菌菌株,在以前的研究中没有报道,从数据库中缺失。此外,有必要破译未知的抗性机制和耐药性在耐热弯曲杆菌属中的传播。这可能对全球食品安全构成挑战。
BACKGROUND: Campylobacter spp. is the most frequent cause of bacterial food-borne gastroenteritis and a high priority antibiotic resistant bacterium according to the World Health Organization (WHO). European monitoring of thermotolerant Campylobacter spp. does not reflect the global burden of resistances already circulating within the bacterial population worldwide.
METHODS: We systematically compared whole genome sequencing with comprehensive phenotypic antimicrobial susceptibility, analyzing 494 thermotolerant Campylobacter poultry isolates from Vietnam and Germany. Any discrepancy was checked by repeating the wet lab and improving the dry lab part. Selected isolates were additionally analyzed via long-read Oxford Nanopore technology, leading to closed chromosomes and plasmids.
RESULTS: Overall, 22 different resistance genes and gene variants (e. g. erm(B), aph(3\')-IIIa, aph(2\'\')-If, catA, lnu(C), blaOXA, sat4) and point mutations in three distinct genes (gyrA, 23S rRNA, rpsL) associated with AMR were present in the Campylobacter isolates. Two AMR genes were missing in the database and one falsely associated with resistance. Bioinformatic analysis based on short-read data partly failed to identify tet(O) and aadE, when the genes were present as duplicate or homologous gene variants. Intriguingly, isolates also contained different determinants, redundantly conferring resistance to chloramphenicol, gentamicin, kanamycin, lincomycin and streptomycin. We found a novel tet(W) in tetracycline sensitive strains, harboring point mutations. Furthermore, analysis based on assemblies from short-read data was impaired to identify full length phase variable aad9, due to variations of the poly-C tract within the gene. The genetic determinant responsible for gentamicin resistance of one isolate from Germany could not be identified. GyrT86I, presenting the main determinant for (fluoro-)quinolone resistance led to a rare atypical phenotype of ciprofloxacin resistance but nalidixic acid sensitivity. Long-read sequencing predicted AMR genes were mainly located on the chromosome, and rarely on plasmids. Predictions from long- and short-read sequencing, respectively, often differed. AMR genes were often organized in multidrug resistance islands (MDRI) and partially located in proximity to transposase genes, suggesting main mobilization of resistance determinants is via natural transformation and transposition in Campylobacter.
CONCLUSIONS: The results of this study suggest that there is frequent resistance gene duplication, mosaicism, and mutation leading to gene variation and truncation in Campylobacter strains that have not been reported in previous studies and are missing from databases. Furthermore, there is a need for deciphering yet unknown resistance mechanisms and resistance spread in thermotolerant Campylobacter spp. that may pose a challenge to global food safety.