Molecular characterization of carbapenem and ceftazidime-avibactam-resistant Enterobacterales and horizontal spread of bla NDM-5 gene at a Lebanese medical center.

Introduction: In the battle against multidrug-resistant bacterial infections, ceftazidime- avibactam (CZA) stands as a pivotal defense, particularly against carbapenemresistant (CR) Gram-negative pathogens. However, the rise in resistance against this drug poses a significant threat to its effectiveness, highlighting the critical need for in-depth studies about its resistance mechanisms. Methods: This research focuses on the genomic characterization of CR- and CZA-resistant Escherichia coli (n=26) and Klebsiella pneumoniae (n=34) strains, harboring the blaNDM and/or blaOXA-48-like genes, at a major Lebanese tertiary care medical center, using whole genome sequencing (WGS). Results: Our findings revealed a notable prevalence of blaNDM in all K. pneumoniae strains isolates, with 27 of these also harboring blaOXA-48. On the other hand, E. coli strains predominantly carried the blaNDM-5 gene. Whole genome sequencing (WGS) identified a predominance of ST383 among K. pneumoniae strains, which possessed a multi-replicon IncFIB-IncHI1B plasmid harboring the blaNDM-5. Additionally, various Inc group plasmids in K. pneumoniae across multiple sequence types were found to carry the blaNDM. Similarly, diverse STs of E. coli were observed to carry blaNDM-5 on different plasmids. Discussion: The study underscores NDM carbapenemases as a paramount resistance mechanism in Lebanon,jeopardizing critical last-resort treatments. It also illuminates the role of varied sequence types and mobile genetic elements in the spread of NDM resistance,stressing the urgent need for strategies to mitigate this threat, especially in nosocomial infections.

Molecular Characteristics of Colistin Resistance in Acinetobacter baumannii and the Activity of Antimicrobial Combination Therapy in a Tertiary Care Medical Center in Lebanon.

(1) Background: Infections with pan-drug-resistant (PDR) bacteria, such as A. baumannii, are becoming increasingly common, especially in healthcare facilities. In this study, we selected 15 colistin-resistant clinical A. baumannii isolates from a hospital in Beirut, Lebanon, to test combination therapies and determine their sequence types (STs) and the mechanism of colistin resistance using whole-genome sequencing (WGS). (2) Methods: Antimicrobial susceptibility testing via broth microdilution against 12 antimicrobials from different classes and growth rate assays were performed. A checkerboard assay was conducted on PDR isolates using six different antimicrobials, each in combination with colistin. Genomic DNA was extracted from all isolates and subjected to WGS. (3) Results: All isolates were resistant to all tested antimicrobials with the one exception that was susceptible to gentamicin. Combining colistin with either meropenem, ceftolozane-tazobactam, or teicoplanin showed synergistic activity. Sequencing data revealed that 67% of the isolates belonged to Pasteur ST2 and 33% to ST187. Furthermore, these isolates harbored a number of resistance genes, including blaOXA-23. Mutations in the pmrC gene were behind colistin resistance. (4) Conclusions: With the rise in antimicrobial resistance and the absence of novel antimicrobial production, alternative treatments must be found. The combination therapy results from this study suggest treatment options for PDR ST2 A. baumannii-infected patients.

Molecular characterization and differential effects of levofloxacin and ciprofloxacin on the potential for developing quinolone resistance among clinical Pseudomonas aeruginosa isolates.

Background: Fluoroquinolones are some of the most used antimicrobial agents for the treatment of Pseudomonas aeruginosa. This study aimed at exploring the differential activity of ciprofloxacin and levofloxacin on the selection of resistance among P. aeruginosa isolates at our medical center. Methods: 233 P. aeruginosa clinical isolates were included in this study. Antimicrobial susceptibility testing (AST) was done using disk diffusion and broth microdilution assays. Random Amplification of Polymorphic DNA (RAPD) was done to determine the genetic relatedness between the isolates. Induction of resistance against ciprofloxacin and levofloxacin was done on 19 isolates. Fitness cost assay was done on the 38 induced mutants and their parental isolates. Finally, whole genome sequencing was done on 16 induced mutants and their 8 parental isolates. Results: AST results showed that aztreonam had the highest non-susceptibility. RAPD results identified 18 clusters. The 19 P. aeruginosa isolates that were induced against ciprofloxacin and levofloxacin yielded MICs ranging between 16 and 256 µg/mL. Levofloxacin required fewer passages in 10 isolates and the same number of passages in 9 isolates as compared to ciprofloxacin to reach their breakpoints. Fitness cost results showed that 12 and 10 induced mutants against ciprofloxacin and levofloxacin, respectively, had higher fitness cost when compared to their parental isolates. Whole genome sequencing results showed that resistance to ciprofloxacin and levofloxacin in sequenced mutants were mainly associated with alterations in gyrA, gyrB and parC genes. Conclusion: Understanding resistance patterns and risk factors associated with infections is crucial to decrease the emerging threat of antimicrobial resistance.

If You're Not Confused, You're Not Paying Attention: Ochrobactrum Is Not Brucella.

Bacteria of the genus Brucella are facultative intracellular parasites that cause brucellosis, a severe animal and human disease. Recently, a group of taxonomists merged the brucellae with the primarily free-living, phylogenetically related Ochrobactrum spp. in the genus Brucella. This change, founded only on global genomic analysis and the fortuitous isolation of some opportunistic Ochrobactrum spp. from medically compromised patients, has been automatically included in culture collections and databases. We argue that clinical and environmental microbiologists should not accept this nomenclature, and we advise against its use because (i) it was presented without in-depth phylogenetic analyses and did not consider alternative taxonomic solutions; (ii) it was launched without the input of experts in brucellosis or Ochrobactrum; (iii) it applies a non-consensus genus concept that disregards taxonomically relevant differences in structure, physiology, population structure, core-pangenome assemblies, genome structure, genomic traits, clinical features, treatment, prevention, diagnosis, genus description rules, and, above all, pathogenicity; and (iv) placing these two bacterial groups in the same genus creates risks for veterinarians, medical doctors, clinical laboratories, health authorities, and legislators who deal with brucellosis, a disease that is particularly relevant in low- and middle-income countries. Based on all this information, we urge microbiologists, bacterial collections, genomic databases, journals, and public health boards to keep the Brucella and Ochrobactrum genera separate to avoid further bewilderment and harm.

Prevalence of intestinal parasites during pre- and post-COVID-19 pandemic at a tertiary care center in Lebanon.

INTRODUCTION: Intestinal parasitic infections (IPIs) are a major medical and public health problem, especially in developing countries. This study aimed at comparing the prevalence and types of IPI during pre- and post-COVID-19 pandemics, and with data reported in Lebanon a decade ago. METHODOLOGY: Stool specimen results from a total of 4,451 and 4,158 patients were examined using the concentration method during the pre-covid (2017-2018) and post-covid (2020-2021) pandemic periods, respectively. Demographic information related to patient's age and gender was recorded. RESULTS: The overall positive detected parasites among the total tested in these two periods were 589 (13.2%) and 310 (7.5%), respectively. The protozoa accounted for most parasites (e.g., Blastocystis hominis, Entamoeba coli (E. coli), Entamoeba histolytica, and Giardia lamblia). Only B. hominis and E. coli showed significant differences; B. hominis was more prevalent in the post-covid period (33.5%) whereas E. coli in the pre-covid phase (44.5%). Among gender, E. histolytica was higher in males during the post-covid period (13.3% vs. 6.3%). Regarding age, adults (between 26 and 55 years) had the highest prevalence, with a noticeable decrease among the elderly in the post-covid time. Compared to the previous decade, the prevalence of B. hominis and E. coli remained higher, and that of E. histolytica and G. lamblia was almost the same. CONCLUSIONS: These findings indicate an overall reduction in the prevalence of IPI during the post-covid period, though IPIs persistence remains high. This highlights the need for enhancing public health awareness efforts to improve hygiene and sanitation to reduce parasitic prevalence in Lebanon.

The under investigated facet of the COVID-19 pandemic: Molecular analysis of secondary bacterial infections at a COVID dedicated intensive care unit within a tertiary care center in Lebanon.

Background: The coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread worldwide. Secondary bacterial infections are associated with unfavorable outcomes in respiratory viral infections. This study aimed at determining the prevalence of secondary bacterial infections in COVID-19 patients admitted at a tertiary medical center in Lebanon. Methodology: From May till November, 2020, a total of 26 Gram-negative isolates were recovered from 16 patients during the course of their COVID-19 infection with Escherichia coli being the most prevalent. The isolates were assessed for their antimicrobial susceptibility by broth microdilution against 19 antimicrobial agents from different classes. Whole genome sequencing of 13 isolates allowed the mining of antimicrobial resistance (AMR) determinants as well as mobile genetic elements and sequence types (ST). Finally, broth microdilution with three different efflux pump inhibitors [theobromine, conessine and PheArg-ß-naphthylamide (PAßN)] was done. Results: Antimicrobial susceptibility testing showed that out of the 26 Gram-negative isolates, 1 (4%) was extensively drug resistant and 14 (54%) were multi-drug resistant (MDR). Whole genome sequencing results revealed a plethora of AMR determinants among the 13 sequenced isolates. Moreover, the 9 Enterobacterales and 4 Pseudomonas aeruginosa sequenced isolates belonged to 9 and 2 different ST, respectively. Using a variety of efflux pump inhibitors we demonstrated that only PAßN had a significant effect when combined with levofloxacin, and the latter regained its activity against two P. aeruginosa isolates. Conclusion: The identification of carbapenem and colistin resistant Gram-negative bacilli causing secondary bacterial infections in critical patients diagnosed with COVID-19 should be of high concern. Additionally, it is crucial to monitor and track AMR, post-COVID pandemic, in order to better understand the effect of this disease on AMR exacerbation.

Genomic surveillance of SARS-CoV-2 in COVID-19 vaccinated healthcare workers in Lebanon.

BACKGROUND: The emergence of SARS-CoV-2 variants including the Delta and Omicron along with waning of vaccine-induced immunity over time contributed to increased rates of breakthrough infection specifically among healthcare workers (HCWs). SARS-CoV-2 genomic surveillance is an important tool for timely detection and characterization of circulating variants as well as monitoring the emergence of new strains. Our study is the first national SARS-CoV-2 genomic surveillance among HCWs in Lebanon. METHODS: We collected 250 nasopharyngeal swabs from HCWs across Lebanon between December 2021 and January 2022. Data on the date of positive PCR, vaccination status, specific occupation, and hospitalization status of participants were collected. Extracted viral RNA from nasopharyngeal swabs was converted to cDNA, library prepped using the coronaHIT method, followed by whole genome sequencing on the Illumina NextSeq 500 platform. RESULTS: A total of 133 (57.1%) samples belonging to the Omicron (BA.1.1) sub-lineage were identified, as well as 44 (18.9%) samples belonging to the BA.1 sub-lineage, 28 (12%) belonging to the BA.2 sub-lineage, and only 15 (6.6%) samples belonging to the Delta variant sub-lineage B.1.617.2. These results show that Lebanon followed the global trend in terms of circulating SARS-CoV-2 variants with Delta rapidly replaced by the Omicron variant. CONCLUSION: This study underscores the importance of continuous genomic surveillance programs in Lebanon for the timely detection and characterization of circulating variants. The latter is critical to guide public health policy making and to timely implement public health interventions.

Whole Genome-Based Characterization of Multidrug Resistant Enterobacter and Klebsiella aerogenes Isolates from Lebanon.

Enterobacter spp. and Klebsiella aerogenes are rod-shaped Gram-negative opportunistic pathogens. This study aimed at the molecular and genomic characterization of multidrug resistant Enterobacter spp. and K. aerogenes isolates recovered from hospitalized patients in a tertiary care hospital in Lebanon. A total of 59 Enterobacter spp. clinical isolates consisting of 41 carbapenem-resistant and 18 susceptible by Etest were included in this study. Genotypic identification through whole-genome sequencing (WGS) was performed and confirmed in silico. Resistance and plasmid profiles were studied using ResFinder4.0 and Plasmid-Finder2.1. Multilocus sequence typing (MLST) was used to determine the isolates' clonality. Using the average nucleotide identity (ANI) we identified and confirmed that 47 (80%) isolates were E. hormaechei, 11 (18%) were Klebsiella aerogenes and 1 (2%) was an E. cloacae. Carbapenem-resistance was detected among 41 isolates all showing an MIC90 of ≥ 32 µg/mL for ertapenem, imipenem, and meropenem. blaNDM-1 (58.5%), blaACT-16 (54%), and blaOXA-1 (54%) were the most common detected ß-lactamases, while blaCTX-M-15 (68%) was the main detected extended-spectrum ß-lactamase (ESBL) encoding gene. Chromosomal ampC, carbapenemase encoding genes, and porin modifications were among the detected carbapenem resistance determinants. The carbapenemase encoding genes were linked to three well-defined plasmid Inc groups, IncFII/IncFIB, IncX3, and IncL. MLST typing revealed the diversity within the studied isolates, with ST114 being the most common among the studied E. hormaechei.: The spread of carbapenem-resistant isolates in clinical settings in Lebanon is a serious challenge. Screening and continuous monitoring through WGS analysis could effectively limit the dissemination of drug-resistant isolates in hospitalized patients. IMPORTANCE Drug resistance is an increasing global public health threat that involves most disease-causing organisms and antimicrobial drugs. Drug-resistant organisms spread in health care settings, and resistance to multiple drugs is common. Our study demonstrated the mechanisms leading to resistance against the last resort antimicrobial agents among members of the Enterobacteriaceae family. The spread of carbapenem-resistant bacteria in clinical settings is a serious challenge. Screening and continuous monitoring could effectively limit the dissemination of drug-resistant isolates in hospitalized patients.

The impact of vaccination on the burden of invasive pneumococcal disease from a nationwide surveillance program in Lebanon: an unexpected increase in mortality driven by non-vaccine serotypes.

BACKGROUND: The impact of pneumococcal conjugate vaccines (PCVs) on the burden of invasive pneumococcal disease (IPD) and serotype distribution was examined across age groups from data collected by the Lebanese Inter-Hospital Pneumococcal Surveillance Program. METHODS: Between 2005 and 2020, 593 invasive Streptococcus pneumoniae isolates were collected from 79 hospitals throughout Lebanon. Serotypes and antimicrobial resistance (AMR) profiles were identified, and trends compared over 3 eras: PCV7, post-PCV7/ pre-PCV13, and PCV13 eras. RESULTS: The prevalence of PCV7 serotypes decreased significantly from 43.6% in the PCV7 era to 17.8% during the PCV13 era (p<0.001). PCV13-only serotypes remained stable in the PCV13 compared to the post-PCV7 eras, especially serotypes 1 and 3, whereas non-vaccine types (NVT) increased throughout the study period, especially 24 and 16F. The mortality rate increased substantially from 12.5% (PCV7 era) to 24.8% (PCV13 era). A significant decrease in AMR was observed across the three study eras. CONCLUSION: PCVs substantially impacted IPD and AMR in vaccinated and unvaccinated populations despite an increase in mortality driven by NVT. Broadening the recommendation of vaccination to include older age-groups, using higher valency vaccines, and implementing stringent antimicrobial stewardship are likely to further impact the burden of IPD.

Genomic Characterization of Mutli-Drug Resistant Pseudomonas aeruginosa Clinical Isolates: Evaluation and Determination of Ceftolozane/Tazobactam Activity and Resistance Mechanisms.

Resistance to ceftolozane/tazobactam (C/T) in Pseudomonas aeruginosa is a health concern. In this study, we conducted a whole-genome-based molecular characterization to correlate resistance patterns and ß-lactamases with C/T resistance among multi-drug resistant P. aeruginosa clinical isolates. Resistance profiles for 25 P. aeruginosa clinical isolates were examined using disk diffusion assay. Minimal inhibitory concentrations (MIC) for C/T were determined by broth microdilution. Whole-genome sequencing was used to check for antimicrobial resistance determinants and reveal their genetic context. The clonal relatedness was evaluated using MLST, PFGE, and serotyping. All the isolates were resistant to C/T. At least two ß-lactamases were detected in each with the blaOXA-4, blaOXA-10, blaOXA-50, and blaOXA-395 being the most common. blaIMP-15, blaNDM-1, or blaVIM-2, metallo-ß-lactamases, were associated with C/T MIC >256 µg/mL. Eight AmpC variants were identified, and PDC-3 was the most common. We also determined the clonal relatedness of the isolates and showed that they grouped into 11 sequence types (STs) some corresponding to widespread clonal complexes (ST111, ST233, and ST357). C/T resistance was likely driven by the acquired OXA ß-lactamases such as OXA-10, and OXA-50, ESBLs GES-1, GES-15, and VEB-1, and metallo- ß-lactamases IMP-15, NDM-1, and VIM-2. Collectively, our results revealed C/T resistance determinants and patterns in multi-drug resistant P. aeruginosa clinical isolates. Surveillance programs should be implemented and maintained to better track and define resistance mechanisms and how they accumulate and interact.

Wide Variability in the Sensitivity and Specificity of Rotavirus Immunoassay Diagnostic Kits in Practice.

INTRODUCTION: Most hospitals rely on rapid antigen-detection kits for the diagnosis of rotavirus infection. Several small studies reviewed the sensitivity and specificity of some of these kits. These studies showed discrepancy in results obtained for sensitivity and specificity that varied according to the type of kit used, area of study, and type of test used as standard for diagnosis of rotavirus infection. The objective of the study is to determine the sensitivity and specificity of five commonly used rotavirus immunoassay kits in comparison to RT-PCR as standard. METHODOLOGY: Stool samples (N = 1,414) collected from children under 5 years of age hospitalized with gastroenteritis were tested for rotavirus by immunoassay kits and RT-PCR in a prospective hospital-based surveillance study conducted at 7 centers in Lebanon. Concordance and discrepancy between the two methods was used to calculate sensitivity and specificity, using RT-PCR as the "gold standard". RESULTS: The sensitivity and specificity were respectively 95.08% and 86.62% for the SD Bioline® (Standard Diagnostics, Inc, South Korea) kit calculated on 645 samples, 65.86% and 45.90% for the VIROTECT® (Trinity Biotech, Ireland) kit calculated on 327 samples, 83.9% and 64.2% for the Rota-Strip (C-1001) (Coris Bioconcept, Belgium) calculated on 95 samples, 52.3% and 10.9% for the Acon® (Acon Laboratories, Inc, California, USA) kit calculated on 122 samples, 68.1% and 20% for the VIKIA® Rota-Adéno (Biomerieux, France) kit calculated on 32 samples. CONCLUSION: A wide discrepancy was detected between the calculated and advertised sensitivity and specificity for most of the kits.

COVID-19 and Candida duobushaemulonii superinfection: A case report.

INTRODUCTION: Critically ill COVID-19 patients are at high risk for nosocomial bacterial and fungal infections due to several predisposing factors such as intensive care unit stay, mechanical ventilation, and broad-spectrum antibiotics. Data regarding multidrug resistant (MDR) Candida species in COVID-19 patients is scarce, and nonexistent regarding Candida duobushaemulonii superinfections. CASE DESCRIPTION: A 34-year-old male presented to our institution with acute respiratory distress syndrome (ARDS) due to COVID-19 infection and developed Candida duobushaemulonii fungemia after multiple courses of antibiotics and prolonged mechanical ventilation. He died after recurrent pneumothorax led to respiratory failure and cardiac arrest. DISCUSSION: Bacterial and fungal infections are common complications of viral pneumonia in critically ill patients. Data regarding these infections in COVID-19 patients has been poorly studied with only a few cases reporting secondary infection, mostly without identifying specific pathogens. Prolonged hospital stays, invasive interventions (central venous catheter, mechanical ventilation), and the use of broad-spectrum antibiotics in COVID-19 infections could carry a high risk of bacterial and/or fungal superinfections. CONCLUSION: Strategies to improve outcome in COVID-19 ICU patients should include early recognition of candidemia and appropriate antifungal therapy.

An unequivocal superbug: PDR Klebsiella pneumoniae with an arsenal of resistance and virulence factor genes.

INTRODUCTION: Infections caused by extensively-drug resistant (XDR) and pan-drug resistant (PDR) Klebsiella pneumoniae represent an emerging threat due to the high associated mortality. This study aimed to characterize two carbapenem resistant K. pneumoniae strains from the same patient, the first being PDR (referred to as IMP 1078b) and the second being XDR (referred to IMP 1078s) isolated from the same patient. METHODOLOGY: Antimicrobial susceptibility testing was done for the 2 K. pneumoniae isolates, followed by carbapenem/ß-lactamase inhibitor combination assay, and fitness cost against cefepime and meropenem. Then, whole-genome sequence analysis was performed to decipher the molecular mechanisms behind the high level of resistance recorded in both isolates. Finally, qRT-PCR was done for ß-lactam resistant genes. RESULTS: This is the first report about a K. pneumoniae isolate harboring 47 antimicrobial resistance genes and having type IV pilli (Yersinia) and the fimbrial adherence determinant Stb (Salmonella) as virulence factors. Further analysis on both isolates are discussed within the article. CONCLUSION: The co-existence of a high number of antimicrobial resistant (AMR) genes and virulence factor genes may lead to a life threatening invasive and untreatable infection.

The Emergence of Invasive Streptococcus pneumoniae Serotype 24F in Lebanon: Complete Genome Sequencing Reveals High Virulence and Antimicrobial Resistance Characteristics.

BACKGROUND: Invasive pneumococcal disease (IPD) remains a global health problem. IPD incidence has significantly decreased by the use of pneumococcal conjugate vaccines (PCV). Nevertheless, non-PCV serotypes remain a matter of concern. Eight Streptococcus pneumoniae serotype 24F isolates, belonging to a non-PCV serotype, were detected through the Lebanese Inter-Hospital Pneumococcal Surveillance Program. The aim of the study is to characterize phenotypic and genomic features of the 24F isolates in Lebanon. METHODS: WGS using long reads sequencing (PacBio) was performed to produce complete circular genomes and to determine clonality, antimicrobial resistance and virulence determinants. RESULTS: The sequencing results yielded eight closed circular genomes. Three multilocus sequence typing (MLST) types were identified (ST11618, ST14184, ST15253). Both MLST and WGS analyses revealed that these isolates from Lebanon were genetically homogenous belonging to clonal complex CC230 and clustered closely with isolates originating from Canada, United States of America, United Kingdom and Iceland. Their penicillin binding protein profiles correlated with both ß-lactam susceptibility patterns and MLST types. Moreover, the isolates harbored the macrolide and tetracycline resistance genes and showed a similar virulence gene profile. To our knowledge, this study represents the first report of complete phenotypic and genomic characterization of the emerging Streptococcus pneumoniae, serotype 24F, in the Middle East and North Africa region.

Predominance of Candida Glabrata among Non-albicans Candida Species in a 16-Year Study of Candidemia at a Tertiary Care Center in Lebanon.

BACKGROUND: Candidemia is associated with a high mortality rate, and its incidence is increasing worldwide with a rise in non-albicans candidemia (NAC). Epidemiologic data from Arab countries are scarce and there are no data from Lebanon; Methods: This is a single-center retrospective chart review of patients with candidemia in a tertiary care center in Lebanon from 2004 to 2019. We extracted data on patient characteristics, isolated Candida species antifungal susceptibility, management and outcomes; Results: We included 170 cases of candidemia. NAC was more common than albicans candidemia (64.7% vs. 35.3%). C. glabrata was the most common non-albicans species (37 isolates) followed by C. tropicalis (14). Recent use of antifungals was a risk factor for NAC (OR = 2.8, p = 0.01), while the presence of a central venous catheter was protective (OR = 0.41, p = 0.02). Fluconazole resistance was 12.5% in C. albicans and 21.5% in non-albicans spp. Mortality at 30 days was 55.5%, with no difference between NAC and albicans candidemia. It was higher in older and critically ill patients but lower in patients whose central venous catheter was removed after detecting fungemia; Conclusions: Candidemia is associated with high mortality in Lebanon, with a predominance of NAC and high prevalence of C. glabrata.

Molecular Characterization of Candida auris Isolates at a Major Tertiary Care Center in Lebanon.

BACKGROUND: The globally emerging Candida auris pathogens poses heavy burden to the healthcare system. Their molecular analyses assist in understanding their epidemiology, dissemination, treatment, and control. This study was warranted to describe the genomic features and drug resistance profiles using whole genome sequencing (WGS) among C. auris isolates from Lebanon. METHODS: A total of 28 C. auris clinical isolates, from different hospital units, were phenotypically identified by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) and tested for antifungal resistance using Vitek-2 system and E test. The complete genomes were determined by WGS using long reads sequencing (PacBio) to reveal the clade distribution and antifungal resistance genes. RESULTS: Candida auris revealed uniform resistance to fluconazole and amphotericin B, with full susceptibility to echinocandins. Among key resistance genes studied, only two mutations were detected: Y132F in ERG11 gene and a novel mutation, D709E, found in CDR1 gene encoding for an ABC efflux pump. Phylogenetically, C. auris genomes belonged to South Asian clade I and showed limited genetic diversity, suggesting person to person transmission. CONCLUSION: This characterization of C. auris isolates from Lebanon revealed the exclusivity of clade I lineage together with uniform resistance to fluconazole and amphotericin B. The control of such highly resistant pathogen necessitates an appropriate and rapid recovery and identification to contain spread and outbreaks.

Nationwide proficiency assessment of bacterial identification and antimicrobial susceptibility testing among 110 laboratories in Lebanon.

INTRODUCTION: Proficiency testing (PT) is one of the most valuable and important activities for the Clinical Microbiology Laboratories (CML) to enroll in to ensure the accuracy and reliability of results. This first time conducted nationwide study was warranted to assess the PT performance activity among CML in Lebanon. METHODOLOGY: Four training and PT activities were organized for 110 nationwide laboratories involved in providing clinical microbiology services. In each PT activity, five different bacterial species were distributed to each laboratory to provide identification (ID) and antimicrobial susceptibility testing (AMST) according to prior discussions and guidelines. RESULTS: The percentages of labs that correctly identified the bacterial species and performed the relevant AMST to it, respectively, were as follows: S. aureus, (100% and 67.8%); Enterococcus faecalis (71% and 82%); Listeria monocytogenes (75% and 61%); Streptococcus agalactiae (86% and 71%); Corynebacterium amycolatum (7% and 33 %); Pseudomonas aeruginosa, (93 % and 53.4%); Klebsiella pneumoniae, (97% and 67.7%); Salmonella typhi ESBL (87 % and 66%); Enterobacter aerogenes (89% and 59%) and Stenotrophomonas maltophilia (84 % and 65%). The resistant types for the species were specified by labs as carbapenem resistant (CR) K. pneumoniae in 78 %, CR E. aerogenes in 34 %, MRSA in 83 %, and VRE in 80.5%. CONCLUSIONS: The wide variation as well as the overall humble scoring of accurate results reflects the dire need for the MOPH to establish and maintain a PT activity program, and entrust the reference laboratory to provide continuing education and training sessions.

Epidemiology and clinical characteristics of viral infections in hospitalized children and adolescents with cancer in Lebanon.

BACKGROUND: Viral infections in children and adolescents with malignancy are commonly encountered and have a significant impact on morbidity and mortality. Studies and epidemiological data regarding viral infections in children with cancer in developing countries are lacking. This retrospective cohort study aims to assess the burden of viral infections in children and adolescents with cancer, by assessing prevalence, risk factors, as well as morbidity and mortality of common viruses over a period of 8 years. METHODS AND FINDINGS: Medical records of cancer patients treated at the Children Cancer Center of Lebanon were reviewed and 155 participants under the age of 21 were identified with at least one documented viral infection during the period from July 2009 to November 2017. This subset included 136 participants with active malignancy and 19 participants with a history of cancer who underwent hematopoietic stem cell transplantation [HSCT] and were in remission; the latter group was analyzed separately. Information regarding participant characteristics, hospital course, and complications were obtained. Associations between viral infections and certain factors were assessed. In the cohort, 64% were male, 81% were Lebanese. In participants with active malignancy, 90% received chemotherapy in the 6 months preceding the viral infection episode, 11% received radiotherapy. 51% of participants were neutropenic at the time of viral detection, and 77% were lymphopenic. 17% experienced a bacterial co-infection, and 3 experienced a viral co-infection. Among 162 viral infection episodes, clinically diagnosed skin infections, mainly herpes simplex virus type 1 and varicella-zoster virus, were the most common [44% of cases]. These were followed by laboratory-proven systemic herpes infections: cytomegalovirus [14%] and Epstein-Barr virus [6%]. Respiratory viruses: influenza and respiratory syncytial virus, accounted for 9% and 4%, respectively, whereas rotavirus represented 11% and BK virus represented 3% of cases. Acute lymphocytic leukemia was the most prevalent neoplasia [57%]. Fever was the most common presenting symptom [55%] and febrile neutropenia was the reason for admission in 24% of cases. The mean length of stay was significantly longer in participants with cytomegalovirus infections and significantly lower in rotavirus infection. Admission to the ICU occurred in 9%, complications in 8%, and mortality in 5%. Participants with viral infections post-HSCT were noted to have a significantly longer length of hospital stay compared to non-HSCT participants, with no other significant differences in clinical course and outcome. The study was limited by its retrospective nature and by the late introduction and underuse of multiplex PCR panels, which may have led to underdiagnosis of viral infections. CONCLUSIONS: Viral infections were prevalent in our sample of cancer patients and may have contributed to morbidity and mortality. Newly available viral diagnostics are likely to vastly increase the number and scope of detectable viral infections in this population. Prospective studies using multiplex PCR technology with systematic testing of patients will be more helpful in defining the burden of viral infections. Furthermore, efforts at antimicrobial stewardship would benefit from the identification of viral causes of infection and limit the unnecessary use of antibiotics in the pediatric cancer population.

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.

Activity of ceftolozane/tazobactam against commonly encountered antimicrobial resistant Gram-negative bacteria in Lebanon.

INTRODUCTION: In view of the continuous rise in Gram-negative bacterial resistance and limited treatment options, Ceftolozane/tazobactam (C/T) is a newly introduced antimicrobial agent in Lebanon for its demonstrated activity against resistant Gram-negative bacteria. However, in vitro data is not available about its activity against commonly isolated bacteria in this country. METHODOLOGY: The analysis included clinical isolates, multidrug-resistant (MDR) and extended-spectrum Beta-lactamases (ESBLs), representing 124 Escherichia coli, 75 Klebsiella pneumoniae and 100 Pseudomonas aeruginosa, identified using the MALDI-TOF. The minimum inhibitory concentration (MIC) for C/T was determined by the Etest (Liofilchem, Roseto degli Abruzzi, Italy). In addition, the disk diffusion (DD) test was used to determine the activity of C/T and of the antimicrobials routinely used to test for such pathogens. RESULTS: The C/T activity against the ESBL producers E. coli and K. pneumoniae isolates were similar (MIC90 value of 1 and 1.5 µg/mL, respectively; susceptibility of 100% and 96%, respectively). However, the activity of C/T against the E. coli and K. pneumoniae MDR isolates was much lower (MIC90 value of 256 and 96 µg/mL, respectively; susceptibility of 54% for each). The C/T MIC90 value for the non-MDR P. aeruginosa isolates was 3 µg/mL and ≥ 256 µg/mL for the MDR P. aeruginosa isolates (susceptibility of 96% vs 42% respectively). Overall, the C/T activities show comparable or higher susceptibility to the routinely used antimicrobials. CONCLUSION: The high in vitro activity of C/T points out its value as a possible alternative to the antimicrobials currently used for treatment of infections caused by such pathogens and would help in minimizing toxicity and bacterial resistance.