No abstract

Rapid detection of carbapenem-resistant Acinetobacter baumannii strains is critical and will benefit patient care by optimizing antibiotic therapies and preventing outbreaks. Herein we describe the development and successful application of a mass spectrometry profile generated by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) that utilized the imipenem antibiotic for the detection of carbapenem resistance in a large series of A. baumannii clinical isolates from France and Algeria. A total of 106 A. baumannii strains including 63 well-characterized carbapenemase-producing and 43 non-carbapenemase-producing strains, as well as 43 control strains (7 carbapenem-resistant and 36 carbapenem-sensitive strains) were studied. After an incubation of bacteria with imipenem for up to 4 h, the mixture was centrifuged and the supernatant analyzed by MALDI-TOF MS. The presence and absence of peaks representing imipenem and its natural metabolite was analyzed. The result was interpreted as positive for carbapenemase production if the specific peak for imipenem at 300.0 m/z disappeared during the incubation time and if the peak of the natural metabolite at 254.0 m/z increased as measured by the area under the curves leading to a ratio between the peak for imipenem and its metabolite being <0.5. This assay, which was applied to the large series of A. baumannii clinical isolates, showed a sensitivity of 100.0% and a specificity of 100.0%. Our study is the first to demonstrate that this quick and simple assay can be used as a routine tool as a point-of-care method for the identification of A. baumannii carbapenemase-producers in an effort to prevent outbreaks and the spread of uncontrollable superbugs.

We herein report the isolation and characterization of 21 Gram-stain-negative strains cultivated from the oropharynx of healthy children in Israel and Switzerland. Initially described as small colony variants of Kingella kingae, phenotypic analysis, biochemical analysis, phylogenetic analysis based on sequencing of the partial 16S rRNA gene and five housekeeping genes (abcZ, adk, G6PD, groEL and recA), and whole genome sequencing and comparison between members of the genera Kingella and Neisseria provided evidence for assigning them to the genus Kingella. Cellular fatty acids included important amounts of C12 : 0, C14 : 0, C16 : 0 and C16 : 1n7. Digital DNA-DNA hybridization between the isolates Sch538T and K. kingae ATCC 23330T revealed relatedness of 19.9 %. Comparative analysis of 16S rRNA gene sequences available in GenBank allowed matches to strains isolated in the USA, suggesting a wider geographical distribution. A novel species named Kingella negevensis sp. nov. is proposed, as most strains have been isolated in the Negev, a desert region of southern Israel. The type strain is Sch538T (=CCUG 69806T=CSUR P957).

The main objective of this study was to decipher the molecular mechanism of resistance to carbapenems and aminoglycosides in a large series of 150 Acinetobacter baumannii clinical isolates collected from July 2012 to September 2013 in Egypt. We report for the first time the emergence of bla NDM-1 and the cooccurrence of 16S rRNA methylase armA with bla NDM-1 and bla OXA-23 in Egyptian hospitals. Multilocus sequence typing identified 27 distinct sequence types, 11 of which were novel. Carbapenem and aminoglycoside resistance in Acinetobacter baumannii is considered an emerging, serious public health problem (1-3). The aim of the present study was to decipher the molecular support of resistance to carbapenems and aminoglycosides among 150 nonrepetitive A. baumannii clinical isolates collected from different clinical samples from inpatients at MabaretEl-Alasafra hospital (Alexandria, Egypt), El-Demerdash hospital (Cairo, Egypt), and the National Cancer Institute (Cairo, Egypt) from July 2012 to September 2013. Identification was performed using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) (4) and confirmed via PCR amplification and sequencing of the intrinsic bla OXA-51 -like gene (5). Antimicrobial susceptibility was determined using a modified Kirby-Bauer disk diffusion method. MICs were determined using Vitek 2 and confirmed for imipenem and amikacin by using Etest strips. Carbapenemase activity was detected phenotypically using the modified Hodge test (MHT) (6) and the modified Carba NP test (7) and confirmed by MALDI-TOF Ultraflex I mass spectrometry (8), while metallo-␤-lactamase activity was detected using the imipenem-EDTA combined-disk test (IECT) (9). All primers and probes used in this study for PCR amplification and sequencing are given in Table S1 in the supplemental material. Genetic mapping of carbapenemases (bla NDM-1 and bla OXA-23 -like), aminoglycoside-modifying enzyme (AME)-encoding resistance genes, and armA was performed for 20 selected isolates. Genotyping was done by multilocus sequence typing (MLST) (10). All isolates revealed a high degree of multidrug resistance (MDR) as shown in Fig. S1 in the supplemental material. Interestingly, 30/150 (20%) isolates were falsely identified as amikacin susceptible by Vitek 2, while they were identified as resistant by Etest (Fig. S1) as already reported (11). Overall, 131 out of 150 isolates (87.3%) were resistant to imipenem, with MICs of Ͼ16 g/ml. Among them, 125 isolates (95.4%) were MHT positive, and 59/150 isolates (39.3%) were IECT positive. In Egypt, there have been few reports describing the emergence of the bla OXA-23 gene among A. baumannii isolates (12-14) and one report of an NDM-2 carbapenemase isolate from a patient transferred to Germany from an Egyptian hospital (15). Herein, we found that 115/ 150 (76.7%) isolates were bla OXA-23 -like positive and 150/150 (100%) were bla OXA51 -like positive. The bla NDM-1 gene was not previously identified as playing a major role in carbapenem resistance...

BackgroundThe aims of this study were to investigate Salmonella contamination in broiler chicken farms and slaughterhouses, to assess the antibiotic resistance profile in avian and human Salmonella isolates, and to evaluate the relationship between avian and human Extended Spectrum β-Lactamase (ESBL)-producing isolates. Salmonella was screened in different sample matrices collected at thirty-two chicken farms and five slaughterhouses. The human isolates were recovered from clinical specimens at the University Teaching Hospital of Constantine (UTH). All suspected colonies were confirmed by MALDI-TOF (Matrix Assisted Laser Desorption Ionization Time OF light) and serotyped. Susceptibility testing against 13 antibiotics including, amoxicillin/clavulanic acid, ticarcillin, cefoxitin, cefotaxime, aztreonam, imipenem, ertapenem, gentamicin, amikacin, ciprofloxacin, colistin, trimethoprim/sulfamethoxazole and fosfomycin, was performed using the disk diffusion method on Mueller-Hinton agar. ESBL-production was screened by the double-disk synergy test and confirmed by molecular characterization using PCR (polymerase chain reaction) amplification and sequencing of ESBL encoding genes. Clonality of the avian and human strains was performed using the Multi Locus Sequencing Typing method (MLST).ResultsForty-five isolated avian Salmonella strains and 37 human collected ones were studied. Five S. enterica serotypes were found in avian isolates (mainly Kentucky) and 9 from human ones (essentially Infantis). 51.11% and 26.6% of the avian isolates were resistant to ciprofloxacin and cefotaxime, respectively, whereas human isolates were less resistant to these antibiotics (13.5% to ciprofloxacin and 16.2% to cefotaxime). Eighteen (12 avian and 6 human) strains were found to produce ESBLs, which were identified as bla CTX-M-1 (n = 12), bla CTX-M-15 (n = 5) and bla TEM group (n = 8). Interestingly, seven of the ESBL-producing strains (5 avian and 2 human) were of the same ST (ST15) and clustered together, suggesting a common origin.ConclusionThe results of the combined phenotypic and genotypic analysis found in this study suggest a close relationship between human and avian strains and support the hypothesis that poultry production may play a role in the spread of multidrug-resistant Salmonella in the human community within the study region.Electronic supplementary materialThe online version of this article (doi:10.1186/s12917-017-1050-3) contains supplementary material, which is available to authorized users.

is an important invasive pathogen in early childhood. The organism elaborates an RTX toxin presumably restricted to this species. Consequently, real-time quantitative PCR (qPCR) assays targeting the RTX locus have been developed in recent years and are gaining increasing use for the molecular diagnosis of infections. However, the present study shows that, a species newly identified in young children, harbors an identical RTX locus, raising the question of whether can be misidentified as by clinical microbiology laboratories. comparison of sp. RTX and genes and studies provided evidence that targeting the and genes could not differentiate between strains of and, whereas targeting the gene could. This prompted the design of a highly specific and sensitive qPCR assay targeting (). Ninety-nine culture-negative osteoarticular specimens from 99 children younger than 4 years of age were tested with a conventional 16S rRNA gene-based broad-range PCR assay and -specific, -specific (), and qPCR assays. Forty-two specimens were positive, including 41 that were also positive and 1 (the remaining one) that was positive. Thus, this study discloses an invasive infection caused by in humans and demonstrates that targeting the RTX locus cannot be used for the formal diagnosis of infections. These findings stress the need for further studies on the epidemiology of asymptomatic carriage and invasive infections caused by in humans.

Biochemical tests have been previously developed to identify carbapenemase-producing Enterobacteriaceae, Pseudomonas spp. (Carba NP test) and Acinetobacter spp. (CarbAcineto NP test). We evaluated a modified Carba NP test to detect carbapenemase production in Enterobacteriaceae, Pseudomonas and Acinetobacter species using a single protocol with rapid results and found good reliability and speed.

No abstract