Spoligotyping of Mycobacterium tuberculosis isolates using Luminex®-based method in Lebanon.

INTRODUCTION: Data about the genotypes of circulating Mycobacterium tuberculosis isolates (MTB) in Lebanon are scarce. This study was undertaken to reveal the spoligotypes of MTB isolates recovered from patients in Lebanon. METHODOLOGY: MTB isolates from 49 patients living in Lebanon were recovered and identified. The samples were heat killed and subjected to DNA extraction. Spoligotyping was performed using microbeads from TB-SPOL Kit and the fluorescence intensity was measured using Luminex 200®. Generated patterns were assigned to families using the SITVIT2 international database of the Pasteur Institute of Guadeloupe and compared. RESULTS: The spoligotyping of the 49 MTB isolates revealed that 31 isolates belonged to Lineage 4 (Euro-American, 63.3%), 12 to Lineage 3 (East- African Indian, 24.5%), 3 to Lineage 2 (East Asian, 6%) and 2 were unknown. Over half of the genotypes (16 of 30) harbored SIT127 supposed to belong to the L4.5 sublineage. One isolate belonging to the rare Manu-Ancestor SIT523 was recovered for the first time in Lebanon, being associated with highly virulent extensively drug-resistant (XDR) MTB phenotype. CONCLUSION: The application of the Spoligotyping Multiplex Luminex® method is an efficient, discriminatory and rapid method to use for first-lane genotyping of MTB isolates. Though humble numbers were tested, this study is one of the first to describe the genomic diversity and epidemiology of MTB isolates of Lebanon, and suggests an increasing prevalence of SIT127 in the country.

Non-tuberculous mycobacteria profiles and their anti-mycobacterial resistance at a major medical center in Lebanon.

INTRODUCTION: Infection with non-tuberculosis mycobacteria (NTM) has been on the rise globally causing a wide spectrum of respiratory and extrapulmonary infections in humans. Studies on these pathogens from the Middle-East including Lebanon are scarce. METHODOLOGY: This retrospective study addresses the approach used for investigation, speciation and antimicrobial resistance (AMR) profiles of recovered NTM isolates from respiratory sources at a major tertiary care center in Lebanon during two periods (2003-2007 and 2013-2017). Processing of specimens, culture and differentiation of recovered NTM isolates from Mycobacterium tuberculosis were done in-house according to standard procedures. Upon request, speciation and AMR testing were performed using molecular and broth dilution methods, respectively, at Mayo Medical Laboratories (Rochester, Minnesota, USA). RESULTS: Among 108 NTM analyzed isolates, 8 species were revealed during the two periods: M. simiae (51% vs 61%), M. avium complex (MAC) (6 % vs 12%) M. fortuitum (12% vs 5%), M. gordonae (6% vs 5%), M. abscessus (6% vs 7%), M. immunogenum (12% vs 0%), M. szulgai (4% vs 0%) and M. peregrinum (0% vs 2%). M. simiae isolates showed high susceptibility (93%-96%) to amikacin and clarithromycin, but high resistance to rifampin, ethambutol, ciprofloxacin, rifabutin, linezolid, trimethoprim/sulfamethoxazole and moxifloxacin. MAC isolates were only susceptible to clarithromycin (86%). M. abscessus isolates were uniformly susceptible to amikacin (100%). CONCLUSION: The revelaed different NTM species, with predominance of M.simiae and various AMR profiles provide a current epidemiologic database and help guiding the selection of appropriate empirical therapy once the clinical relevance is established.

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.

TUBERCULOSIS BURDEN IN LEBANON: EVOLUTION AND CURRENT STATUS.

BACKGROUND: In the absence of surveillance data and consolidated information about tuberculosis (TB) and its drug resistance burden in Lebanon, this retrospective study was conducted to shed light on TB and its other relevant aspects over the last 15 years in this country. METHODS: To generate the TB data for this retrospective study, two main sources were used: 1) the records of patients in the National Tuberculosis Program (NTP); 2) the results of the Clinical Microbiology Laboratory (CML) at the American University of Beirut Medical Center (AUBMC). The TB data review pertained to its epidemiological aspect, implementation of the directly observed therapy strategy (DOTS) all over, the gender distribution, the impact of high risk groups (non-national population, Syrian refugees, patients with multi-drug resistance-TB [MDR-TB] and the inmate population) on the trend of TB in Lebanon between 1999 and 2013. Reviewed also are TB in children, extrapulmonary tuberculosis and the mycobacterium other than tuberculosis (MOTT). RESULTS: During the last 15 years, 7548 TB cases were diagnosed and evaluated at the NTP. After the decreasing of TB incidence from 13/100 000 population in 2001 to 9/100 000 in 2006, the incidence started to increase in 2007, reaching 20/100 000 in 2013, mostly due to increased cases among non-national population. Fluctuations in TB rates over the years were seen among children, inmates, MDR-TB, and HIV patients. MOTT isolates recovery rates also fluctuated during the study period, M. simiae being the most common. CONCLUSION: After the decreasing trends of TB incidence between 1999 and 2006, Lebanon has been experiencing an increasing incidence in tuberculosis population since 2007. This is mainly attributed to the dramatic increase of TB patients among non-nationals and the influx of Syrian refugees. The ongoing collaboration between the public and private sectors, improvements of the surveillance system and TB control are important factors for successful elimination of TB in this country.

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.

A reflection on bacterial resistance to antimicrobial agents at a major tertiary care center in Lebanon over a decade.

BACKGROUND: Antimicrobial resistance has been inflecting deleterious health and economic consequences locally and globally. This study addresses the patterns and trends of bacterial resistance to antimicrobial agents over a decade, at a major tertiary care center in Beirut. METHODS: Data on bacterial susceptibility patterns at the CAP accredited Clinical Microbiology Laboratory is analyzed from January 2000 to November 2011, along with related different studies conducted during this period. RESULTS: Increasing rates of ESBL-producing isolates were noted for Escherichia coli, Klebsiella pneumoniae, Salmonella spp. and Shigella spp. Resistance to carbapenems remains problematic in Acinetobacter spp, and Pseudomonas aeruginosa, and started emerging in E. coli and K. pneumoniae. Tigecycline and colistin maintained excellent activity against most ESBL and carbapenem resistant bacteria relevant to the treatment by these agents. Resistance to quinolones is being encountered in Streptococcus pneumoniae, Haemophilus influenzae, Salmonella spp. and Shigella spp. Methicillin resistant Staphylococcus aureus (MRSA), though remaining relatively high, showed decreasing trends of resistance, while vancomycin maintain uniform activity. Rare and sporadic vancomycin resistant strains in enterococci are encountered. Macrolide and clindamycin increasing rates of resistance is noted in S. pneumoniae, group A streptococci, S. aureus, viridans streptococci and some others. CONCLUSION: Physicians should be aware of the local epidemiology of antimicrobial resistance to properly guide the initial therapy. These resistance problems can be attributed to uncontrolled use of antimicrobial agents, thus, highlighting the need for antimicrobial stewardship to curb this threat.