Phenotypic Detection Of Extended Spectrum Beta Lactamases Producing Organism Among Godfrey Okoye University Students

1.1 Introduction

Extended-spectrum beta-lactamases (ESBL) are enzymes that confer resistance to most beta-lactam antibiotics, including penicillin, cephalosporin, and aztreonams (Bush and Jacoby, 2010). 

Extended-spectrum Beta(β)-lactamases (ESBLs) are a group which are mostly plasmid-mediated, diverse, complex and rapidly evolving enzymes that are posing a major therapeutic challenge today in the treatment of hospitalized and community-based patients. Infections due to ESBL producers range from uncomplicated urinary tract infections (UTI) to life-threatening sepsis. These enzymes share the ability to hydrolyze third-generation cephalosporin and aztreonam and yet, are inhibited by clavulanic acid. In addition, ESBL-producing organisms exhibit co-resistance to many other classes of antibiotics, resulting in limitation of therapeutic option. Because of inoculum effect and substrate specificity, their detection is also a major challenge (Deepthiet al 2010).

Numerous studies have barbed towards high incidence rate of UTI associated with Escherichia coli (E. coli) and antibiotic resistance. The emergence of Multi Drug Resistant (MDR) variant of E. colihas been accounted. MDR is defined as resistance to at least two antibiotics of different classes including aminoglycosides, chloramphenicol, tetracycline and/or erythromycin. MDR in many bacteria is due to the action of multi-drug efflux pumps and by the accumulation on Resistance (R) plasmids or transposons of genes with each coding for resistance to a specific agent. Nowadays, in UTIs, ESBL -expressing Gram-Negative Bacilli (ESBL-GNB) generally cause community-acquired infections. The resistance of Gram-negative bacteria is typically owed to plasmid mediated enzymes of ESBL. ESBL producing bacteria are typically associated with multi-drug resistance (MDR) and antibacterial choice is often complicated by multi-drug resistance (Prakash and Yadav, 2017).

1.2 CLASSIFICATION OF ESBL

There are two major classification systems for β-lactamases:

1.2.1 Molecular classification is based on the amino acid sequence and divides β-lactamases Ambler classes into A (serine penicillinases), C (cephalosporinases), and D (oxa-cillinases) enzymes which utilize serine for β-lactam hydrolysis and class B metalloenzymes which require divalent zinc ions for substrate hydrolysis (Bush and Jacoby, 2010).

1.2.2 Functional classification scheme was initially proposed by Bush in 1989 and then expanded in 1995. It takes into account substrate and inhibitor profiles in an attempt to group the enzymes in ways that can be correlated with their phenotype in clinical isolates (Bush and Jacoby, 2010).

1.3 DIVERSITYOF THE TYPES OF ESBL

1.3.1 TEM beta-lactamases 

The first plasmid-mediated beta-lactamase in gram-negative bacteria was discovered in Greece in the 1960s. It was named TEM after the patient from whom it was isolated (Temoniera).   Although TEM-type beta-lactamases are most often found in Escherichia coli and Klebsiellapneumoniae, they are also found in other species of Gram-negative bacteria with increasing frequency (Clark et al., 1990).

1.3.2 SHV beta-lactamases 

Sulfhydryl variable, (SHV) shares 68 percent of its amino acids with TEM and has a similar overall structure. The SHV beta-lactamase is most commonly found in Klebsiellapneumoniae and is responsible for up to 20% of the plasmid-mediated ampicillin resistance in this species. ESBLs in this family also have amino acid changes around the active site. More than 60 SHV varieties are known  (Chow et al., 2010).

1.3.3 CTX-M beta-lactamases 

Cefotaximase Munich (CTX-M), these enzymes were named for their greater activity against cefotaxime than other oxyimino-beta-lactam substrates (e.g., ceftazidime, ceftriaxone, or cefepime). Rather than arising by mutation, they represent examples of plasmid acquisition of beta-lactamase genes normally found on the chromosome of Kluyvera species, a group of rarely pathogenic commensal organisms. These enzymes are not very closely related to TEM or SHV beta-lactamases in that they show only approximately 40% identity with these two commonly isolated beta-lactamases. More than 80 CTX-M enzymes are currently known. Despite their name, a few are more active on ceftazidime than cefotaxime. They have mainly been found in strains of Salmonella entericaserovartyphimurium and E. coli, but have also been described in other species of Enterobacteriaceae(Chow et al., 2010).

1.3.4 OXA beta-lactamases 

The OXA-type β-lactamases are so named because of their oxacillin-hydrolyzing abilities. OXA beta-lactamases were long recognized as a less common but also plasmid-mediated beta-lactamase variety that could hydrolyze oxacillin and related anti-staphylococcal penicillin. These beta-lactamases differ from the TEM and SHV enzymes in that they belong to molecular class D and functional group 2d. The OXA-type beta-lactamases confer resistance to ampicillin and cephalothin and are characterized by their high hydrolytic activity against oxacillin and cloxacillin and the fact that they are poorly inhibited by clavulanic acid. Amino acid substitutions in OXA enzymes can also give the ESBL phenotype. While most ESBLs have been found in E. coli, Klebsiellapneumoniae, and other Enterobacteriaceae, the OXA-type ESBLs have been found mainly in Pneumoniaeaeruginosa. The OXA beta-lactamase family was originally created as a phenotypic rather than a genotypic group for a few beta-lactamases that had a specific hydrolysis profile. Therefore, there is as little as 20% sequence homology among some of the members of this family. However, recent additions to this family show some degree of homology to one or more of the existing members of the OXA beta-lactamase family. Some confer resistance predominantly to ceftazidime (Chow et al., 2010).

1.3.5 PER type

The PER-type ESBLs share only around 25–27% homology with known TEM- and SHV-type ESBLs. PER-1 β-lactamase efficiently hydrolyzes penicillin and cephalosporin and is susceptible to clavulanic acid inhibition. PER was first detected in Pseudomonas aeruginosa, and later in Salmonella entericaserovarTyphimurium and Acinetobacter isolates as well. In Turkey, as many as 46% of nosocomial isolates of Acinetobacter spp. and 11% of P. aeruginosa were found to produce PER, which shares 86% homology to PER-1, has been detected in S. entericaserovarTyphimurium, E. coli, K. pneumoniae, Proteus mirabilis, and Vibrio cholerae (Danish et al 2015).