Combating antimicrobial resistance using antimicrobial combination therapy and ß-lactamase inhibitors.

INTRODUCTION: The range of antimicrobial agents used to treat bacterial infections is becoming limited with the constant increase in antimicrobial resistance (AMR). Several genetic factors underlie AMR, including ß-lactamase-encoding genes such as blaCTXM-15 that confers resistance to third-generation cephalosporins, and blaOXA-48, blaNDM-1, and blaKPC-2 that confer resistance to carbapenems. Remaining treatment approaches for such resistant infections include antimicrobial combination therapy and the use of ß-lactamase inhibitors. This study assesses the molecular effects of such treatment approaches on antimicrobial resistant Enterobacteriaceae clinical isolates in vitro and in vivo. METHODOLOGY: Nine clinical Enterobacteriaceae isolates were included in the study. One harboring blaCTXM-15, one harboring blaOXA-48, one harboring blaKPC-2, two harboring blaNDM-1 and blaCTXM-15, and four harboring blaOXA-48 and blaCTXM-15. Minimal inhibitory concentrations were determined for carbapenems with ß-lactamase inhibitors: avibactam, Ca-EDTA, and relebactam. Synergism between antibiotic combinations was determined by double disc diffusion when using colistin with several antibiotics. In vitro and in vivo gene expression levels were done on these combinations with and without inhibitors. RESULTS: The use of meropenem, imipenem, and ertapenem with the selected ß-lactamase inhibitors restored isolate susceptibility in 100%, 87.5%, and 25% of the cases, respectively. Antimicrobial synergism was mostly detected between colistin and meropenem, fosfomycin, or tigecycline. Survival studies revealed the survival of most mice receiving antimicrobial combination therapy with inhibitors as compared to the controls. Overall gene expression levels of resistance genes were variable depending on treatment. CONCLUSIONS: The threat of antibiotic resistant bacterial infections remains viable; however, different approaches to therapy are available.

Genotyping of Mycobacterium tuberculosis in Lebanon using a novel rapid spoligotyping multiplex luminex method.

INTRODUCTION: Incidence of Tuberculosis (TB) in Lebanon, according to the WHO, is estimated to be 35 cases per 100,000 people. However, data about the genotypes of circulating Mycobacterium tuberculosis isolates (MTB) in this country is lacking. This study aims to reveal the genotypes of TB isolates recovered from patients in Lebanon. METHODOLOGY: Fifty M. tuberculosis isolates from patients in Lebanon were recovered and identified at the reference TB center of the Ministry of Public Health. All isolates were heat killed and subjected to DNA extraction. Spoligotyping method (TB-Spol, Beamedex, France) was used to identify the presence of 43 spacers via a multi-analyte profiling system (Luminex, Bio-Rad). Generated patterns were assigned to families using the SITVIT2 international database of the Pasteur Institute of Guadeloupe. RESULTS: The spoligotyping of the 50 MTB isolates revealed 13 lineages, one being novel. The most frequent shared-types (SIT) identified lineage was the Ural (34%), followed by the Central Asian lineage (10%) and a single isolate (2%) belonging to the rare Manu-Ancestor SIT523 lineage, associated with a highly virulent XDR MTB phenotype. The rest of the SIT isolates (18%) were equally distributed along 9 different lineages. The 13th non-SIT lineage is a novel one constituting 36% of the total isolates. CONCLUSION:  The application of Spoligotyping Multiplex Luminex method is a novel, discriminatory and rapid method to use for genotyping of MTB isolates employing the multi-spacer analysis system. Our study showed genomic diversification of MTB isolates from Lebanon.

Antimicrobial agents active against carbapenem-resistant Escherichia coli and Klebsiella pneumoniae isolates in Lebanon.

INTRODUCTION: It is not yet clear which antimicrobial agents should be used to treat the ominously increasing infections with carbapenem-resistant (CR) bacteria. We therefore investigated the activity of different antimicrobial agents against CR Escherichia coli and Klebsiella pneumoniae in Lebanon. METHODOLOGY: This retrospective study assessed the minimum inhibitory concentrations (MICs) of three carbapenems (by Etest), as well as the in vitro activity of eight other antimicrobials (by disk diffusion) against CR E. coli (n = 300) and K. pneumoniae (n = 232) isolates recovered at a major University Medical Center in Lebanon. RESULTS: Higher percentages of isolates showing carbapenem MICs of ≤ 8 µg/mL were noted among the CR E. coli compared to the CR K. pneumoniae for ertapenem (48% vs 27%), imipenem (74 % vs 58%) and meropenem (82% vs 63%). Among the eight other antimicrobials, activity was generally higher when the MICs for the three carbapenems were ≤ 8 µg/mL. Regardless of the MIC level of the three carbapenems, very low susceptibility rates (≤ 33%) were noted for ciprofloxacin, trimethoprim-sulfamethoxazole and aztreonam against both E. coli and K. pneumoniae isolates. With Amikacin, higher susceptibility rates were seen against E. coli isolates (81%-97%) than against K. pneumoniae isolates (55%-86%), also reflecting higher activity than gentamicin (44%-54%). The best activity (66%-100%) was observed for tigecycline, colistin and fosfomycin against both CR species. CONCLUSIONS: Based on the in vitro findings in this study, the combination of a carbapenem showing an MIC of ≤ 8 µg/mL together with an active colistin, tigecycline, or fosfomycin, would offer a promising treatment option for patients infected with CR E. coli or K. pneumoniae.

Candida profiles and antifungal resistance evolution over a decade in Lebanon.

INTRODUCTION: Infection with and antifungal resistance of Candida species have been on the rise globally. Relevant data on these pathogens are relatively few in our region, including Lebanon, thus warranting this study. METHODOLOGY: This retrospective study of Candida spp. profiles and their in vitro antifungal susceptibility was based on analysis requests for 186 Candida non-albicans and 61 C. albicans during three periods (2005-2007, 2009-2011, and 2012-2014) over the span of the last 10 years at the American University of Beirut Medical Center (AUBMC), a major tertiary care center in Lebanon. Identification of Candida was done using the API 20C AUX system, and the E-test was used to determine the minimum inhibitory concentrations (MICs) of antifungal agents. RESULTS: Among the 1,300-1,500 Candida isolates recovered yearly, C. albicans rates decreased from 86% in 2005 to around 60% in 2014. Simultaneously, the non-albicans rates increased from 14% in 2005 to around 40% in 2014, revealing 11 species, the most frequent of which were C. tropicalis, C. glabrata, and C. parapsilosis. All these demonstrated high resistance (35%-79%) against itraconazole, but remained uniformly susceptible (100%) to amphotericin B. Though C. albicans and the other species maintained high susceptibility against fluconazole and voriconazole, their MIC90 showed an elevated trend over time, and C. glabrata had the highest resistance rates. CONCLUSIONS: The observed rise in resistance among Candida spp. in Lebanon mandates the need for close surveillance and monitoring of antifungal drug resistance for both epidemiologic and treatment purposes.