Molecular epidemiological study of extended spectrum beta-lactamase (ESBL) producing bacteria from a hospital within Saudi Arabia
Student thesis: Doctoral Thesis
Extended spectrum beta-lactamase (ESBL) producing bacteria are the cause of widespread infections worldwide, in both hospital and community settings. The presence of ESBLs prevent treatment of the infection with many β-lactam antibiotics. Highly mobile genetic element such as plasmids carrying ESBL genes often also harbour resistance genes to other classes of antimicrobials. To date, susceptibility generally remains high to the carbapenem antibiotics and to the polymyxin colistin and the glycylcycline tigecycline. However rigorous antibiotic stewardship programmes on national and international level will be crucial in maintaining the status of these drugs. This study provides epidemiological and molecular information regarding ESBL-producing bacteria both-in- patients and out-patients at the Saudi Aramco Dhahran Health Center, an acute care facility in the Eastern Province of Saudi Arabia, over four years. Two sampling cohorts (2010-2011 and 2012-2013 gave a total of 385 ESBL-producing isolates dominated by Enterobacteriaceae, mainly from urine cultures, followed by blood cultures. Most patients were from the out-patient setting. The majority of strains were Escherichia coli, followed by K. pneumoniae. Other isolates included members of Enterobacter, Citrobacter, Morganella, Serratia and Proteus families. K. pneumoniae became relatively more common in 2012 compared to 2010. Overall, female gender was identified as a risk factor for ESBL infection. Older age (>60 years) was also a risk factor, particularly for males. Susceptibility of infections to nine β-lactams was tested, namely ampicillin/sulbactam, ceftazidime, ceftriaxone, cefazolin, cefepime, cefoxitin and the carbapenems imipenem, meropenem, and ertapenem. There were high levels of resistance in both 2010 and 2012 for ampicillin/sulbactam, cefazolin, ceftriaxone and ceftazidime. While high percentages of both E. coli and K. pneumoniae were sensitive to cefoxitin in both 2010 and 2012, both E. cloacae and E. aerogenes isolates were uniformly resistant to this drug. For cefepime, sensitivity fell sharply in 2012, attributable to equivalent drops among E. coli and K. pneumoniae. There was consistently high sensitivity in 2010 and 2012 to the carbapenems. Seven non--lactam antibiotics were tested, namely nitrofurantoin, amikacin, ciprofloxacin, trimethoprim/sulfamethoxazole, gentamicin, colistin and tigecycline. There was 100% sensitivity to colistin and tigecycline in both years. For ciprofloxacin, trimethoprim/sulfamethoxazole and gentamicin, there was low sensitivity in 2010 and 2012. For the aminoglycoside antibiotic amikacin, 98.4% of isolates were sensitive in 2010; this had fallen significantly to 69.3% in 2012, attributable to decreased sensitivity of E. coli isolates. ESBL-positive E. coli were relatively sensitive to nitrofurantoin, but the other bacterial species had low sensitivity. Overall, these results suggest a worrying trend for reduced sensitivity to a range of antibiotics of different classes. Multiplex PCR for the resistance determinants SHV, TEM and CTX-M showed that the CTX-M determinant was most prevalent, either alone or in combination with TEM and/or SHV, in all study years. Prevalence rose from 49.2% of isolates in 2010, to 84.7% in 2012. While the second most common genotype in 2010 was TEM-only, this had fallen to 5.4% in 2012, reflecting drops in both E. coli and most strikingly K. pneumoniae isolates. CTX-M in combination with other resistance determinants became more prevalent in 2012. This was particularly apparent for K. pneumoniae. Microarray MDR analysis on a sample set of 159 isolates in 2012 revealed that the most prevalent CTX-M subtype, was type 15 (61.6%), consistent with worldwide experience, followed by CTX-M9 (8.2%). MDR analysis revealed that OXA-48 was present in three out of the 159 isolates, while there was no expression of KPC or NDM. The three OXA-48 isolates were, however, sensitive to the carbapenems. In fact, there was uniformly high sensitivity to the carbapenems and to colistin and tigecycline. There was relatively high prevalence of sensitivity to cefoxitin among CTX-M-expressing isolates. Amikacin also emerged as a possible choice for infections expressing CTX-M type 15 in combination with TEM and/or SHV, for CTX-M-9 isolates, and for SHV-only isolates. Nitrofurantoin could also be cautiously considered for isolates expressing CTX-M-1 type 15- only or CTX-M-9, and for CTX-M-1 type 15 in combination with TEM isolates, but not with CTX-M –type 15+TEM+SHV isolates. There was 90.9% prevalence of gentamicin sensitivity for CTX-M-9 isolates. Genotyping of E. coli and K. pneumoniae isolates by ERIC-PCR and by PFGE identified groups with a potentially clonal origin, with the PFGE technique having the greater discriminatory power.
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