Prevalence of Extended-Spectrum β-Lactamase-Resistant Genes in Escherichia coli Isolates from Central China during 2016–2019

SIMPLE SUMMARY: β-lactam antibiotics are commonly used for the treatment of severe infection in both animals and humans, and the resistance of E. coli to third-generation cephalosporins is becoming a worldwide problem. Our results revealed 407 E. coli strains isolated from central China exhibited strong resistance to first- to fourth-generation cephalosporins and monobactam antibiotics, and piperacillin/tazobactam was the most effective drug. Phenotypically, 63.88% of the isolates were positive for ESBL production and the isolation rates kept growing. Genetic characterization identified CTX-M as the most prevalent type and bla(TEM) + bla(CTX-M) as the most common ESBL genotype combination. Furthermore, a novel multiplex real-time PCR method for detecting the three most common ESBL genes was developed as a complementary rapid-screening test for antimicrobial resistance genes. These results confirm the cephalosporins resistance urgency in the central Chinese poultry sector and suggest the continuous monitoring and timely detection of ESBL-producing isolates is vitally important in establishing appropriate antimicrobial therapies and blocking their transmission. ABSTRACT: The emergence and dissemination of Escherichia coli (E. coli) strains that produce extended-spectrum beta-lactamases (ESBLs) represents a major public health threat. The present study was designed to evaluate the prevalence and characteristics of ESBL-producing Escherichia coli isolates from chickens in central China during 2016–2019. A total of 407 E. coli strains isolated from 581 chicken swabs were identified conventionally and analyzed for various cephalosporin susceptibility by disk-diffusion assay. ESBL-producing strains were screened using the double=disk synergy test and ESBL-encoding genes were carried out by PCR/sequencing. A total of 402 E. coli isolates exhibited strong resistance to first- to fourth-generation cephalosporins and monobactam antibiotics, especially cefazolin (60.69%), cefuroxime (54.05%), cefepime (35.14%), ceftriaxone (54.30%), and aztreonam (40.29%). Piperacillin/tazobactam (1.72%) was the most effective drug against the strains, but the resistance rates increased each year. Among the isolates, 262 were identified as ESBL producers and the isolation rates for the ESBL producers increased from 63.37% to 67.35% over the four years. CTX-M (97.33%) was the most prevalent type, followed by TEM (76.72%) and SHV (3.05%). The most common ESBL genotype combination was bla(TEM) + bla(CTX-M) (74.46%), in which the frequency of carriers increased steadily, followed by bla(CTX-M) + bla(SHV) (3.05%). In addition, the most predominant specific CTX-M subtypes were CTX-M-55 (48.47%) and CTX-M-1 (17.94%), followed by CTX-M-14 (11.01%), CTX-M-15 (8.02%), CTX-M-9 (6.11%), CTX-M-65 (4.58%), and CTX-M-3 (1.15%). Moreover, a novel multiplex qPCR assay was developed to detect bla(CTX-M), bla(TEM), and bla(SHV), with limits of detection of 2.06 × 10(1) copies/μL, 1.10 × 10(1) copies/μL, and 1.86 × 10(1) copies/μL, respectively, and no cross-reactivity with other ESBL genes and avian pathogens. The assays exhibited 100% sensitivity and specificities of 85%, 100%, and 100% for bla(CTX-M), bla(TEM), and bla(SHV), respectively. In conclusion, our findings indicated that ESBL-producing E.coli strains isolated from chickens in central China were highly resistant to cephalosporins and frequently harbored diversity in ESBL-encoding genes. These isolates can pose a significant public health risk. The novel multiplex qPCR method developed in this study may be a useful tool for molecular epidemiology and surveillance studies of ESBL genes.