Identification and molecular characterization of Escherichia coli bla_SHV genes in a Chinese teaching hospital

Highlights

• Four (0.8%) bla_SHV β-lactamase-encoding genes were identified in Escherichia coli

• A novel bla_SHV variant (bla_SHV-195) was reported

• Two bla_SHV-5 genes located in a 23 kb size of transferable plasmid, respectively

• Two wild strains (EC D2485 and EC D2487) which harbored bla_SHV-5 and formed one cluster in PFGE analysis were clonally related

Abstract

Escherichia coli (E. coli) commonly reside in human intestine and most E. coli strains are harmless, but some serotypes cause serious food poisoning. This study identified and molecularly characterized bla_SHV genes from 490 E. coli strains with multi-drug resistance in a hospital population. PCR and molecular cloning and southern blot were performed to assess functions and localizations of this resistant E. coli gene and the pulsed-field gel electrophoresis (PFGE) was utilized to demonstrate the clonal relatedness of the positive E. coli strains. The data showed that 4 of these 490 E. coli strains (4/499, 0.8%) carried bla_SHV genes that included EC D2485 (bla_SHV-5), EC D2487 (bla_SHV-5), EC D2684 (bla_SHV-11) and EC D2616 (bla_SHV-195, a novel bla_SHV). Analysis of bla_SHV open-reading frame showed that bla_SHV-5 had a high hydrolysis activity to the broad-spectrum penicillin (ampicillin or piperacillin), ceftazidime, ceftriaxone, cefotaxime and aztreonam. bla_SHV-195 and bla_SHV-11 had similar resistant characteristics with high hydrolysis activities to ampicillin and piperacillin, but low activities to cephalosporins. Moreover, the two bla_SHV-5 genes were located on a transferable plasmid (23 kb), whereas the other two bla_SHV variants (bla_SHV-11 and bla_SHV-195) seemed to be located in the chromosomal material. Both EC D2485 and EC D2487 clones isolated in 2010 had the same DNA finger printing profile and they might be the siblings of clonal dissemination. The data from the current study suggest that the novel bla_SHV and clonal dissemination may be developed, although bla_SHV genes were infrequently identified in this hospital population. The results of the work demonstrate the necessity for molecular surveillance in tracking bla_SHV-producing strains in large teaching hospital settings and emphasize the need for epidemiological monitoring.

Introduction

Enterobacteriaceae bacteria (Escherichia coli or Klebsiella pneumoniae, etc.) are the most common nosocomial pathogens in human being and can cause the purulent diseases, such as pneumonia, meningitis, bacteremia, or the urinary tract and intestinal infection, while others are the important pathogens in nosocomial infection (Abreu et al., 2011). For example, E. coli commonly reside peacefully in human intestine because most E. coli strains are harmless, but some serotypes cause serious food poisoning (Vogt and Dippold, 2005). Moreover, some non-virulent bacteria could obtain exgenous virulent genes and become pathogens to human beings. However, the resistant mechanism of Enterobacteriaceae bacteria is very complex and one of the main mechanism of drug resistance is their production of beta-lactamase, an enzyme to hydrolyze β-lactam antibiotics and lead to their deactivation (Jones and Wilson, 1982). Among the large family of the β-lactamases, the extended-spectrum β-lactamases (ESBLs), AmpC enzyme and carbapenemase are the most common and there are numerous variants existing in clinical isolates (Gutkind et al., 2013, Vasoo et al., 2015).

In recent decades, the global over- and misuse of antimicrobial agents has caused the rapid spread of multiple-risistance genes in Europe (Coque et al., 2008), Asia (Hawkey, 2008), and the USA (Bush, 2008). E. coli is an important cause of community- and health care-associated infection, and frequently associated with ESBL carriage (Vasoo et al., 2015). The ESBLs consist of three major genetic groups, including bla_TEM, bla_SHV and bla_CTX-M types (Chong et al., 2011). In the 1990s, bla_TEM or bla_SHV type of the ESBLs was generally found in Klebsiella pneumonia (K. pneumonia) and most isolates were from nosocomial infection spread (Yano et al., 2013), although there were reports about E. coli and other bacteria (Yezli et al., 2015). Nevertheless, currently available data indicate that the main ESBL identified in E. coli was bla_CTX-M (bla_CTX-M-15 in particular). But in K. pneumoniae, the predominant β-lactamases are bla_SHV (especially, bla_SHV-12, bla_SHV-5, and bla_SHV-1), bla_CTX-M (mainly bla_CTX-M-15) and bla_TEM (Yezli et al., 2015). In 1983, bla_SHV-1 gene was first discovered on the chromosome of K. pneumoniae in Germany and considered as the ancestor of bla_SHV-ESBLs (Bedenic et al., 2005). Antibiotics misuse in clinic and agriculture led rapid emergence of genotypes of bla_SHV ESBLs due to point mutations to bla_SHV variants (Gutkind et al., 2013). For example, E. coli strains isolated from clinical samples showed the characteristics of multidrug resistance and contained multiple antibiotics-resistant genes, especially those of β-lactamases, which are normally carried on the plasmids. The plasmid could transfer among the same and different species of bacteria to induce dissemination of drug resistance (Rice et al., 1990). The previous studies on β-lactamase genes have mainly focused on bla_SHVs in K. pneumonia (Gutmann et al., 1989, Nuesch-Inderbinen et al., 1997) and bla_SHV genes were globally disseminated by plasmids not only among genera of E. coli and K. pneumoniae, but also among other hospital-acquired or community infection microorganisms (Gutkind et al., 2013). Even though bla_SHV-type ESBLs were originally found in K. pneumonia; thus, case reports and outbreaks involving different Enterobacteriaceae and non-fermenting gram negative bacilli producing bla_SHV-ESBLs are well documented (Ben-Hamouda et al., 2004, Gundes et al., 2005, Mansour et al., 2009, Mazzariol et al., 2007, Naiemi et al., 2005, Palasubramaniam et al., 2009, Poirel et al., 2004).

One of the early reports of bla_SHV-2 enzymes was from China (Cheng and Chen, 1994); however, before a study of bla_CTX-M-13 and bla_CTX-M-14 from Guangzhou, China was published, there were very few reports of ESBL genotypes from China (Hawkey, 2008). To date, the ESBLs-producing E. coli and K. pneumonia in China become serious and bla_CTX-M ESBLs were among the most common genotypes (Gao et al., 2015, Liu et al., 2013a, Liu et al., 2009, Ma et al., 2012, Shi et al., 2015, Sun et al., 2010, Wang et al., 2003, Yu et al., 2007), but a few studies focused on bla_SHV. Thus, in this study, we identified and molecularly characterized bla_SHV genes in 490 E. coli strains isolated from a hospital in Wenzhou, China to access bla_SHV genotypes and their dissemination in this hospital population.