The aim of the study was to determine the prevalence of New Delhi metallo-β lactamase-1 (NDM-1) producing Enterobacteriaceae in Kuwait over a one year period. Consecutive Enterobacteriaceae isolates with reduced susceptibility to carbapenems were collected from four government hospitals in Kuwait from January–December 2014. Their susceptibility to 18 antibiotics was performed by determining the minimum inhibitory concentration. Isolates resistant to carbapenems were tested by PCR for resistant genes. Finger printing of the positive isolates was done by DiversiLab^®. Clinical data of patients harboring NDM-1 positive isolates were analyzed. A total of 764 clinically significant Enterobacteriaceae isolates were studied. Of these, 61 (8%) were carbapenem-resistant. Twenty one out of these 61 (34.4%) were NDM-1-producers. All patients positive for NDM-1-carrying bacteria were hospitalized. About half were females (11/21 [52.3%]), average age was 53.3 years and the majority were Kuwaitis (14/21 [66.6%]). Six patients (28.5%) gave a history of travel or healthcare contact in an endemic area. Mortality rate was relatively high (28.6%). The predominant organism was Klebsiella pneumoniae (14 [66.6%]) followed by E. coli (4 [19%]). All NDM-1-positive isolates were resistant to meropenem, ertapenem, cefotaxime, cefoxitin and ampicillin, while 95.2% were resistant to imipenem, cefepime, and piperacillin-tazobactam. They were multidrug resistant including resistance to tigecycline, but 90% remained susceptible to colistin. About two-thirds of isolates (61.9%) co-produced-extended spectrum β-lactamases. During the study period, an outbreak of NDM-1 positive K. pneumoniae occurred in one hospital involving 3 patients confirmed by DiversiLab^® analysis. In conclusion, NDM-1-producing Enterobacteriaceae is a growing healthcare problem with increasing prevalence in Kuwait, especially in hospitalized patients, leaving few therapeutic options. A high prevalence of NDM-1 necessitates the implementation of strict infection control to prevent the spread of these organisms.

Health care-associated pneumonia due to multidrug-resistant organisms represents a major therapeutic challenge. Unfortunately, treatment is dependent on empirical therapy, which often leads to improper and inadequate antimicrobial therapy. A rapid multiplex PCR-based Unyvero pneumonia application (UPA) assay that assists in timely decision-making has recently become available. In this study, we evaluated the performance of UPA in detecting etiological pathogens and resistance markers in patients with nosocomial pneumonia (NP). The impact of this assay on the management of severe nosocomial pneumonia was also assessed. Appropriate specimens were processed by UPA according to the manufacturer's protocol in parallel with conventional culture methods. Of the 56 patients recruited into the study, 49 (87.5%) were evaluable. Of these, 27 (55.1%) and 4 (8.2%) harbored multiple bacteria by the PCR assay and conventional culture, respectively. A single pathogen was detected in 8 (16.3%) and 4 (8.2%) patients, respectively. Thirteen different genes were detected from 38 patients, including the ermB gene (40.8%), the bla _OXA-51 -like gene (28.6%), the sul1 (28.6%) and int1 (20.4%) integrase genes, and the mecA and bla _CTX-M genes (12.3% each). The time from sample testing to results was 4 h versus 48 to 96 h by UPA and culture, respectively. Initial empirical treatment was changed within 5 to 6 h in 33 (67.3%) patients based on the availability of UPA results. Thirty (62.2%) of the patients improved clinically. A total of 3 (6.1%) patients died, mainly from their comorbidities. These data demonstrate the potential of a multiplex PCR-based assay for accurate and timely detection of etiological agents of NP, multidrug-resistant (MDR) organisms, and resistance markers, which can guide clinicians in making early antibiotic adjustments.

Matrix-assisted laser desorption-ionization time-of-flight mass spectrometry (MALDI-TOF MS) recently became available for the identification of bacteria in routine diagnostic laboratories. It is rapid and cost-effective and likely to replace phenotypic identification. This study was undertaken to compare two MALDI-TOF MS-based, Bruker Microflex MS (BMS) and VITEK MS (VMS) systems, for identification (ID) of clinically significant bacterial isolates. Clinically relevant broad diversity of bacterial isolates obtained during a 6-consecutive months of routine laboratory processing of clinical specimens were subjected to ID by the BMS and VMS in parallel with Vitek 2, a conventional phenotypic system (CPS). For the BMS, the isolates were tested in duplicates directly and after pretreatment. Identification was provided with accompanying scores according to manufacturers’ instructions. With VMS, single deposits of the same sets of isolates were tested in duplicates directly on MALDI-plate. Results were interpreted according to the manufacturer’s protocols. Discrepant results were resolved by 16S rRNA gene amplification and sequencing. A total of 806 pathogens comprising 507 Gram-negative bacilli (GNB), 16 Gram-negative cocci (GNC), 267 Gram-positive cocci (GPC), and 16 Gram-positive bacilli (GPB) were tested. BMS and VMS correctly identified isolates to genus and species levels (ID 97.3% and 93.2%, and 99.8% and 99.0%, respectively). Both systems as well as the CPS correctly identified the majority of the species in the family Enterobacteriaceae, Pseudomonas spp., and Acinetobacter baumannii. Turnaround time for identification by BMS and VMS was <20 min compared with 24-48 h by the CPS. VMS performed slightly better than BMS with GPC ID, especially the Streptococcus spp. Some S. mitis isolates were identified as S. pneumoniae by BMS. BMS and VMS were rapid and proved to be consistently accurate for producing bacterial identification in a fraction of time it takes for identification by CPS.

To prospectively evaluate the performance of two matrix-assisted laser desorption/ionization time-of-flight mass spectrometry systems (MALDI-TOF MS) for the identification of clinically significant yeast isolates compared to the VITEK 2 system. One hundred and eighty-eight consecutive yeast isolates were analyzed by Bruker Biotyper and VITEK MS. The results were compared with the conventional VITEK 2 yeast identification system. Discrepant results were resolved by direct sequencing of rDNA. Accurate identification by VITEK 2, VITEK MS, and Bruker Biotyper MS was 94.1% (177/188), 93.0% (175/188), and 92.6% (174/188), respectively. Three isolates were not identified by VITEK MS, while nine Candida orthopsilosis were misidentified as Candida parapsilosis, as this species is not present in its database. Eleven isolates were not identified or were wrongly identified by Bruker Biotyper and although another 14 were correctly identified, the score was unreliable at <1.7. The overall accuracy of rapid MALDI-TOF MS systems was essentially comparable to that of the conventional VITEK 2 yeast identification system. However, future expansion of the databases may further improve the outcome and accuracy of identification of yeast species.

The purpose of this study was to identify the mechanisms leading to carbapenem resistance among multidrug-resistant Enterobacteriaceae isolates recovered from hospitalized patients with nosocomial infections in Mubarak Al Kabeer Hospital, Kuwait. Fourteen carbapenem-resistant Enterobacteriaceae isolates were obtained from inpatients in different wards and intensive care units between April 2009 and February 2011. Antibiotic susceptibilities were determined using the E-test method. Genes encoding β-lactamases were characterized by specific PCR amplification, sequencing and conjugation assays. All isolates were identified as metallo-β-lactamase (MBL) producers using phenotypic and molecular methods. Eleven of the 14 isolates produced VIM-4 (six Klebsiella pneumoniae, three Escherichia coli, one Enterobacter cloacae and one Klebsiella oxytoca). Three K. pneumoniae isolates produced the MBL NDM-1 and co-produced the plasmid-encoded AmpC CMY-4. The VIM-4-producing isolates co-produced extended-spectrum β-lactamases including CTX-M-15 and some SHV derivatives. The VIM-4 gene was not transferable by conjugation studies of six selected strains. We demonstrated here the emergence of VIM-4- and NDM-1-producing isolates in the largest teaching hospital in Kuwait.