agar and Enterobacteriaceae-Infections

Leclercia adecarboxylata is a bacteria closely related to Escherichia coli according to its biochemical characteristics and is commonly considered non-pathogenic although a growing number of publications classify it as an emerging pathogen. Fosfomycin resistance is a common trait for L. adecarboxylata encoded by fosA. To analyze genomic traits of sixteen L. adecarboxylata strains isolated from blood culture and a bottle of total parenteral nutrition.. Twenty-eight L. adecarboxylata strains isolated from blood culture and a bottle of total parenteral nutrition were identified biochemically with a Vitek ® automated system. The strains were phenotyped by their growth on Eosin Methylene Blue agar or MacConkey agar plates. Additionally, Pulsed field gel electrophoresis (PFGE) was performed to establish the clonal relationship. The genomic DNA of sixteen strains was obtained using a Qubit ® dsDNA HS Assay Kit and sequenced on an Illumina ® MiSeq instrument. Draft genomes were assembled using PROKKA and Rast. Assemblies were submitted to Resfinder and PathogenFinder from the Center for Genomic Epidemiology in order to find resistance genes and pathogenic potential. IslandViewer4 was also used to find Pathogenicity and Phage Islands. For identification of the fosA gene, manual curation and Clustal analysis was performed. A novel FosA variant was identified. Finally, phylogenetic analysis was performed using VAMPhyRE software and Mega X.. In this paper, we report the genomes of sixteen strains of Leclercia adecarboxylata causing an outbreak associated with parenteral nutrition in public hospitals in Mexico. The genomes were analyzed for genetic determinants of virulence and resistance. A high pathogenic potential (pathogenicity index 0.82) as well as multiple resistance genes including carbapenemics, colistin and efflux pumps were determined. Based on sequence analysis, a new variant of the fosA. Commensal strains of L. adecarboxylata may acquire genetic determinants that provide mechanisms of host damage and go unnoticed in clinical diagnosis. L. adecarboxylata can evolve in a variety of ways including the acquisition of resistance and virulence genes representing a therapeutic challenge in patient care.

Topics: Agar; Anti-Bacterial Agents; Disease Outbreaks; Enterobacteriaceae Infections; Escherichia coli; Genomics; Hospitals, Public; Humans; Mexico; Phylogeny

As carbapenemase-producing Enterobacterales are increasingly reported worldwide, their rapid detection is crucial to reduce their spread and prevent infections and outbreaks. Lateral flow immunoassays (LFIAs) have become major tools for the detection of carbapenemases. However, as for most commercially available assays, only the five main carbapenemases are targeted.. Here, we have developed and evaluated an LFIA prototype for the rapid and reliable detection of the increasingly identified GES-type β-lactamases.. The GES LFIA was validated on 103 well-characterized Gram-negative isolates expressing various β-lactamases grown on Mueller-Hinton (MH) agar, chromogenic, and chromogenic/selective media.. The limit of detection of the assay was 106 cfu per test with bacteria grown on MH agar plates. GES LFIA accurately detected GES-type β-lactamases irrespective of the culture media and the bacterial host. The GES LFIA was not able to distinguish between GES-ESBLs and GES-carbapenemases. Because GES enzymes are still rare, their detection as an ESBL or a carbapenemase remains important, especially because extensive use of carbapenems to treat ESBL infections may select for GES variants capable of hydrolysing carbapenems.. The GES LFIA is efficient, rapid and easy to implement in the routine workflow of a clinical microbiology laboratory for the confirmation of GES-type β-lactamases. Combining it with immunochromatographic assays targeting the five main carbapenemases (KPC, NDM, VIM, IMP and OXA-48) would improve the overall sensitivity for the most frequently encountered carbapenemases and ESBLs, especially in non-fermenters.

Topics: Agar; Bacteriological Techniques; beta-Lactamases; Carbapenems; Culture Media; Enterobacteriaceae Infections; Gram-Negative Bacteria; Humans; Immunoassay; Sensitivity and Specificity

The European Food Safety Authority (EFSA) advised to prioritize monitoring carbapenemase producing Enterobacteriaceae (CPE) in food producing animals. Therefore, this study evaluated the performance of different commercially available selective agars for the detection of CPE using spiked pig caecal and turkey meat samples and the proposed EFSA cultivation protocol. Eleven laboratories from nine countries received eight samples (four caecal and four meat samples). For each matrix, three samples contained approximately 100 CFU/g CPE, and one sample lacked CPE. After overnight enrichment in buffered peptone water, broths were spread upon Brilliance™ CRE Agar (1), CHROMID® CARBA (2), CHROMagar™ mSuperCARBA™ (3), Chromatic™ CRE (4), CHROMID® OXA-48 (5) and Chromatic™ OXA-48 (6). From plates with suspected growth, one to three colonies were selected for species identification, confirmation of carbapenem resistance and detection of carbapenemase encoding genes, by methods available at participating laboratories. Of the eleven participating laboratories, seven reported species identification, susceptibility tests and genotyping on isolates from all selective agar plates. Agars 2, 4 and 5 performed best, with 100% sensitivity. For agar 3, a sensitivity of 96% was recorded, while agar 1 and 6 performed with 75% and 43% sensitivity, respectively. More background flora was noticed for turkey meat samples than pig caecal samples. Based on this limited set of samples, most commercially available agars performed adequately. The results indicate, however, that OXA-48-like and non-OXA-48-like producers perform very differently, and one should consider which CPE strains are of interest to culture when choosing agar type.

Topics: Agar; Animals; Bacterial Proteins; Bacteriological Techniques; beta-Lactamases; Carbapenem-Resistant Enterobacteriaceae; Enterobacteriaceae Infections; Microbial Sensitivity Tests; Sensitivity and Specificity; Swine

Heteroresistance is the phenomenon wherein subpopulations of presumed isogenic bacteria show varied antibiotic susceptibilities, and the current gold standard for the determination of heteroresistance is population analysis profiling (PAP). However, when conducting PAP to confirm carbapenem heteroresistance in Enterobacteriaceae, the authors found some isolates that did not seem to be heteroresistant, despite meeting PAP criteria. This article elaborates on the validity of PAP for the determination of heteroresistance, especially among carbapenemase-producing Enterobacteriaceae (CPE). Bacterial cells that were originally non-viable on selective agar supplemented with a high concentration of meropenem were found to be occasionally viable, likely due to the hydrolysis of carbapenems by carbapenemases produced by dying cells, mimicking the emergence of subpopulations with enhanced resistance. As such, PAP for CPE is highly affected by carbapenemases produced by dying populations, and may not detect heterogeneity in carbapenem resistance appropriately among seemingly isogenic clones.

Topics: Agar; Anti-Bacterial Agents; Bacterial Proteins; beta-Lactamases; Carbapenem-Resistant Enterobacteriaceae; Carbapenems; Enterobacteriaceae Infections; Humans; Meropenem; Microbial Sensitivity Tests

Detecting carbapenemase-producing Enterobacteriaceae (CPE) has become increasingly difficult due to the emergence of diverse enzymes. The aim of the study was to evaluate an agar plate-based modified carbapenem inactivation method (p-mCIM) for detection of CPE. Stock strains and clinical isolates of CPE were used to evaluate the p-mCIM. The p-mCIM was performed as described for the mCIM, except that meropenem disks were placed on the lawn of test organisms on Mueller-Hinton agar (MHA) plates. Among 17 stock strains of CPE, six of eight KPC-2-like- and all six NDM-1-like carbapenemase-producing strains were positive by the p-mCIM without incubation in the carbapenem inactivation (CI) step. Among 380 CPE clinical isolates detected, 308 and 38 were KPC-2-like and NDM-1-like enzyme producers, respectively. The required incubation time in the CI step to show all isolates were positive by p-mCIM was 3 h for isolates with KPC-2-like enzyme and 1 h for isolates with metallo-β-lactamases. Twenty-eight of 30 isolates with OXA-48-like enzymes were p-mCIM positive. Sensitivities of both the p-mCIM and the mCIM (based on inhibition zone of ≤15 mm) for detection of CPE were 100%. All 70 ertapenem-nonsusceptible, but carbapenemase gene-negative isolates tested were both p-mCIM (based on inhibition zone of ≥21 mm) and mCIM negative. In conclusion, performance of the p-mCIM, which uses a lawn of bacterial colonies on MHA plate instead of a bacteria-suspended Tryptic soy broth tube in the CI step, is essentially identical to that of the CLSI-recommended mCIM in the detection of clinical isolates of Enterobacteriaceae producing carbapenemases including difficult to detect bla

Topics: Agar; Anti-Bacterial Agents; Bacterial Proteins; beta-Lactamases; Carbapenem-Resistant Enterobacteriaceae; Carbapenems; Enterobacteriaceae Infections; Humans; Meropenem; Microbial Sensitivity Tests; Sensitivity and Specificity

Topics: Agar; Bacterial Proteins; Bacteriological Techniques; beta-Lactamases; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Humans; Sensitivity and Specificity

To assess a cost-effective in-house selective plate formula for actively screening carbapenem-resistant Enterobacteriaceae (CRE).. The in-house formula included CHROMagar. The in-house plate had high sensitivity and specificity, particularly for Escherichia coli and the KESC group (Klebsiella spp., Enterobacter spp., Serratia marscescens, and Citrobacter spp.), and it may be widely applied as an alternative to other ready-to-use commercial plates.. The formula developed in the present study may facilitate the early detection and isolation of CRE and decrease transmission, particularly in low- and middle-income countries with a high rate of CRE colonization and limited access to ready-to-use commercial plates.

Topics: Agar; Anti-Bacterial Agents; Carbapenem-Resistant Enterobacteriaceae; Culture Media; Enterobacteriaceae Infections; Gastrointestinal Tract; Microbial Sensitivity Tests; Prevalence; Quality Control; Sensitivity and Specificity

CRE-JU is a novel selective agar for carbapenem-resistant Enterobacteriaceae that contains ceftazidime, cloxacillin, meropenem, and vancomycin. This study evaluated the ability of 63 carbapenem-resistant isolates and 53 non-carbapenem-resistant Enterobacteriaceae strains clinically isolated in Japan, Myanmar, Nepal, and Vietnam to grow on CRE-JU. CRE-JU showed 92.1% sensitivity and 100% specificity for detecting carbapenem-resistant Enterobacteriaceae compared with dug susceptibility profiles.

Topics: Agar; Anti-Bacterial Agents; Bacterial Proteins; beta-Lactamases; Carbapenem-Resistant Enterobacteriaceae; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Fermentation; Humans; Lactose; Microbial Sensitivity Tests; Sensitivity and Specificity

Topics: Agar; Bacteriological Techniques; Carbapenem-Resistant Enterobacteriaceae; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Humans; Pilot Projects; Sensitivity and Specificity

We aimed to validate quality assurance (QA) criteria for rectal surveillance cultures (RSCs) for extended-spectrum β-lactamase-producing Enterobacteriaceae. QA for RSCs were tested by observing the presence or absence of fecal soiling and by examining the growth of Enterobacteriaceae on MacConkey agar. Extended-spectrum β-lactamase-producing Enterobacteriaceae were detected in 136 out of 434 soiled swabs (31.3%) and in 61 out of 257 nonsoiled swabs (23.7%) (P = .036). Observation of fecal soiling on RSCs can serve as a simple QA criterion and prevent the reporting of false-negative results.

Topics: Agar; Anti-Bacterial Agents; beta-Lactamases; Carrier State; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Feces; Gene Expression; Humans; Prospective Studies; Quality Control; Rectum

To validate a combined disc method along with resazurin chromogenic agar for early screening and differentiation of Klebsiella pneumoniae carbapenemase, metallo-β-lactamase and OXA-48 carbapenemase-producing Enterobacteriaceae.. The combined disc test comprising of meropenem alone and with EDTA, phenylboronic acid or both EDTA and phenylboronic acid, and temocillin alone were evaluated with the resazurin chromogenic agar plate assay against a total of 86 molecularly confirmed Enterobacteriaceae clinical isolates (11 metallo-β-lactamases, eight Kl. pneumoniae carbapenemases, 11 OXA-48, 32 AmpC and 15 extended-spectrum-β-lactamase producers and nine co-producers of extended-spectrum-β-lactamase and AmpC). The inhibition zone diameters were measured and interpreted at 7 h for the presence of carbapenemase. All carbapenemase producers were phenotypically distinguished by this assay with 100% sensitivity and specificity.. This early phenotypic method is very simple, inexpensive, and reliable in the detection and differentiation of carbapenemase-producing Enterobacteriaceae. It could be exploited in any microbiological laboratory for diagnosis of these recalcitrant bacteria.. This assay poses excellent performance in discrimination of Kl. pneumoniae carbapenemase, metallo-β-lactamase and OXA-48 carbapenemases within 7 h, which is much faster than conventional disc diffusion methods. The rapid detection could help clinicians screen patients, control infection and provide epidemiological surveillance.

Topics: Agar; Anti-Bacterial Agents; Bacterial Proteins; beta-Lactam Resistance; beta-Lactamases; Enterobacteriaceae; Enterobacteriaceae Infections; Humans; Microbial Sensitivity Tests; Oxazines; Oxytocin; Xanthenes

This assessment describes the enteric colonization of German soldiers 8-12 weeks after returning from mostly but not exclusively subtropical or tropical deployment sites with third-generation cephalosporin-resistant Enterobacteriaceae, vancomycin-resistant enterococci (VRE), and methicillin-resistant Staphylococcus aureus (MRSA). Between 2007 and 2015, 828 stool samples from returning soldiers were enriched in nonselective broth and incubated on selective agars for Enterobacteriaceae expressing extended-spectrum beta-lactamases (ESBL), VRE and MRSA. Identification and resistance testing of suspicious colonies was performed using MALDI-TOF-MS, VITEK-II and agar diffusion gradient testing (bioMérieux, Marcy-l'Étoile, France). Isolates with suspicion of ESBL were characterized by ESBL/ampC disc-(ABCD)-testing and molecular approaches (PCR, Sanger sequencing). Among the returnees, E. coli with resistance against third-generation cephalosporins (37 ESBL, 1 ESBL + ampC, 1 uncertain mechanism) were found in 39 instances (4.7%). Associated quinolone resistance was found in 46.2% of these isolates. Beta-lactamases of the blaCTX-M group 1 predominated among the ESBL mechanisms, followed by the blaCTX-M group 9, and blaSHV. VRE of vanA-type was isolated from one returnee (0.12%). MRSA was not isolated at all. There was no clear trend regarding the distribution of resistant isolates during the assessment period. Compared with colonization with resistant bacteria described in civilians returning from the tropics, the colonization in returned soldiers is surprisingly low and stable. This finding, together with high colonization rates found in previous screenings on deployment, suggests a loss of colonization during the 8- to 12-week period between returning from the deployments and assessment.

Topics: Agar; Anti-Bacterial Agents; beta-Lactam Resistance; beta-Lactamases; Carrier State; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Feces; Female; Gene Expression; Germany; Humans; Male; Methicillin-Resistant Staphylococcus aureus; Military Personnel; Travel; Tropical Climate; Vancomycin-Resistant Enterococci

We compared the Remel Spectra CRE agar plate to CDC standard methodology for the isolation of carbapenem-resistant Enterobacteriaceae (CRE) from 300 rectal swab specimens obtained from patients residing in a long-term-care facility (LTCF). Multiplex PCR experiments were performed on isolates to identify specific Klebsiella pneumoniae carbapenemases (KPC) and additional β-lactamases. Of the 300 patients, 72 (24%) harbored CRE and were PCR positive for KPC enzymes. The Remel Spectra CRE plates detected KPC-type CRE in isolates from 70 of 72 patients (97.2%), while the CDC method detected CRE in 56 of 72 (77.8%). CRE identification results were available in 18 h compared to 36 h for the CDC method. Remel Spectra CRE agar plates can provide useful means for a fast and reliable method for detecting KPC-type CRE and for accelerated institution of appropriate infection control precautions.

Topics: Agar; Anti-Bacterial Agents; beta-Lactam Resistance; beta-Lactamases; Carbapenems; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Humans; Long-Term Care; Polymerase Chain Reaction; Rectum; Sensitivity and Specificity; Time Factors

The detection of enterobacteria with production of beta-lactamases of extended spectrum in selective chromogenic agar was analyzed The results ofdetection of beta-lactamases of extended spectrum was compared with "double disc" technique. The smears from mucous membrane of guttur and rectum from patients were analyzed in parallel on solid growth agar (Endo or Mac Conkey) and on selective agar CHROMagartm ESBL (CHROMagar France). The production of beta-lactamases of extended spectrum was confirmed using "double discs" technique. To exclude hyper-production of ampC beta-lactamases E-test was applied containing cefotetan and cefotetan with cloxacillin. The sampling consisted of 1552 samples from patients. The study permitted to isolate 1243 strains of enterobacteria on agar Endo or Mac Conkey and 409 strains of enterobacteria on selective agar CHROMagartm ESBL (Escherichia coli n = 226, Klebsiella pneumoniae n = 105, enterobacter spp. n = 35, Citrobacter spp. n = 21, others n = 22). The application of "double discs" technique confirmed production of beta-lactamases of extended spectrum in 386 (94%) out of 409 strains isolated on agar CHROMagartm ESBL. In 23 (6%) of strains no confirmation was established and hyper-production of ampC of beta-lactamases was established 15 out of total. Additionally, 8 were sensitive to cephalosporin of third generation. All enterobacteria isolated on agar Endo or Mac Conkey also were tested by "double discs" technique. Overall, 394 strains of enterobacteria with production of beta-lactamases of extended spectrum were obtained. On all agars (agar Endo or Mac Conkey and CHROMagartm ESBL)--263 (67%) strains; only on CHROMagartm ESBL--123 (31%) and only on agar Endo or Mac Conkey--8 (2%) (p < 0.0001). The sensitivity of selective agar CHROMagartm ESBL made up to 98% and specificity--97%. The resolution about detection of enterobacteria producing beta-lactamases of extended spectrum were submitted to clinic in 18-24 hours after arrival ofsamplesfrom patients in laboratory. The CHR OMagartm ESBL has higher sensitivity and specificity to detect enterobacteria with production of beta-lactamases of extended spectrum and can be applied in common laboratory practice.

Topics: Agar; Anti-Bacterial Agents; Bacterial Typing Techniques; beta-Lactamases; Cefotetan; Cephalosporins; Chromogenic Compounds; Cloxacillin; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Gene Expression; Humans; Microbial Sensitivity Tests; Sensitivity and Specificity

The adequate detection of carbapenemase-producing Enterobacteriaceae (CPE) is essential for adequate antibiotic therapy and for infection control purposes, especially in an outbreak setting. Selective agars play an important role in the detection of CPE. The Oxoid Brilliance™ CRE Agar (Thermo Fisher Scientific) was evaluated for the detection of CPE using 255 non-repetitive Enterobacteriaceae isolates, including 95 CPE (36 KPC, 4 KPC plus VIM, 4 NDM, 6 GIM, 20 VIM, and 25 OXA-48-producing isolates). The sensitivity of the CRE agar for the detection of CPE was 94 % (89/95), but differed per carbapenemase gene (100 % for KPC, NDM, and GIM, 90 % for VIM, and 84 % for OXA-48-producing isolates). The specificity of the CRE agar was 71 %, due to the growth of AmpC- and/or ESBL-producing isolates. The CRE agar is a sensitive tool for the detection of KPC and metallo-carbapenemase-producing Enterobacteriaceae, although the detection of OXA-48 producers is less optimal. The relatively low specificity requires confirmation of carbapenemase production for isolates recovered from the CRE agar.

Topics: Agar; Bacterial Proteins; Bacteriological Techniques; beta-Lactamases; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Humans; Sensitivity and Specificity

Extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae strains are frequent causative agents both in community-acquired infections and in nosocomial infections. The newly developed ChromID ESBL agar (bioMerieux, Marcy I'Etoile, France) is a chromogenic medium that helps rapid identification of ESBL-positive Enterobacteriaceae species from the clinical samples. The aim of this study was to evaluate the performance of ChromID ESBL agar in the rapid identification of ESBL-positive pathogens from the urine samples of the patients with urinary tract infections. A total of 672 urine samples (437 outpatients, 235 inpatients) were included in the study. All of the samples were inoculated simultaneously to 5% sheep blood agar, McConkey agar and ChromID ESBL agar media, and evaluated after incubation at 37°C for 18-24 hours. Gram-negative pathogens were tested for ESBL both by the standard combined double-disk diffusion (CDD) method using ceftazidime and cefotaxime disks and by doubledisk synergy (DDS) test. Among 672 urine cultures, 199 yielded microbial growth in routine media (sheep blood agar and/or McConkey agar), whereas 57 yielded bacterial growth in ChromID ESBL agar. When CDD method was accepted as the reference method according to Clinical and Laboratory Standards Institute (CLSI) recommendations, the sensitivity, specificity, positive and negative predictive values for ChromID ESBL agar for the detection of ESBL-positive bacteria in urinary tract infections were estimated as 97%, 92.9%, 89.1%, and 98.1%, respectively. Additionally, we also discovered that Chrom ID ESBL agar could detect vancomycin-resistant enterococci (VRE) as well as ESBL-positive bacteria, in our study. In order to investigate this observation we inoculated a total of 203 stock strains of Enterococcus spp. (118 vancomycin-sensitive, 85 vancomycin-resistant) to this medium. None of the vancomycinsensitive Enterococcus spp. did grow in ChromID ESBL medium, while 83 of the 85 resistant isolates (97.6%) did grow in the medium. As a result, it was concluded that ChromID ESBL agar medium was advantageous since it led to the growth of VRE and ESBL-positive Enterobacteriaceae isolates in different colors and helped in early identification of these two problematic bacteria. We thought that especially early detection of VRE will accelerate the establishment of necessary measures to prevent the nosocomial spread of this microorganism.

Topics: Agar; beta-Lactamases; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Enterococcus; Urinary Tract Infections; Urine; Vancomycin Resistance

The worldwide prevalence of extended-spectrum-beta-lactamase-producing ESBL-producing Enterobacteriaceae (ESBL-E) is increasing, making the need for optimized detection techniques more urgent. In this study we investigated the performance of two ESBL-E screening and two ESBL-E confirmation techniques. In accordance with the Dutch national guidelines (www.wip.nl), a collection of 642 highly resistant Enterobacteriaceae strains, as identified by Vitek2, was used to test the performances of two screening techniques (EbSA ESBL agar plate and ChromID ESBL agar plate) and of two confirmation techniques (MIC-strip ESBL and Vitek2 ESBL test panel). The individual test results were compared by using Etest, followed by a combination disk test if Etest results were inconclusive. Among group 1 isolates (Escherichia coli, Klebsiella spp., Proteus spp., Salmonella spp., and Shigella spp.) 291 (57.6%) were ESBL-E, versus 65 (47.4%) in group 2 (Enterobacter spp., Citrobacter spp., Morganella morganii, Serratia spp., and Providencia spp.). The sensitivities of all four tests for group 1 were comparable (EbSA, 96.6%; ChromID, 97.3%; MIC-strip, 99.6%; and Vitek2, 95.1%). The specificities of the EbSA and ChromID were the same (93.9%). However, the confirmation techniques produced many inconclusive test results, which reduces the applicability in routine laboratories. Only the two screening agar plates were validated for ESBL testing of group 2 microorganisms. They showed comparable sensitivities; however, the EbSA screening agar plate had a significantly higher specificity (78.6% versus 44.3%). In conclusion the screening agar plates performed better than the two confirmation techniques. The EbSA agar plate had the best overall performance.

Topics: Agar; Bacteriological Techniques; beta-Lactamases; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Humans; Mass Screening; Sensitivity and Specificity

The increased worldwide spread of carbapenem-resistant Enterobacteriaceae (CRE) emphasizes the need for a sensitive screening procedure to identify these microorganisms. Gastrointestinal carriers may serve as the reservoir for cross-transmission in the health care setting, and thus active surveillance is a key part in preventing the spread of such strains. Three agar-based methods for direct CRE detection from rectal swabs were compared: CHROMagar-KPC (Chrom); MacConkey agar with imipenem at 1 μg/ml (MacI); and MacConkey plates with imipenem, meropenem, and ertapenem disks (MacD). First, we compared the levels of detection (LODs) of 10 molecularly characterized carbapenemase-producing Enterobacteriaceae strains by the three methods. Second, we compared their performance in a surveillance study using rectal swabs (n = 139). The LODs of carbapenemase-producing Enterobacteriaceae strains were influenced by their MICs to carbapenems and were best for MacI, followed by Chrom. The MacD method was able to detect only the strains exhibiting MICs of ≥ 32 μg/ml to at least ertapenem. In the surveillance study, both Chrom and MacI had greater sensitivity (85%) than MacD (76%). However, MacI was the most specific method. In conclusion, MacI appears to be most appropriate medium for the detection of CRE in settings in which multiclonal CRE strains with various MICs to carbapenems are circulating.

Topics: Agar; Anti-Bacterial Agents; beta-Lactam Resistance; Carbapenems; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Feces; Humans; Mass Screening; Microbial Sensitivity Tests; Sensitivity and Specificity

Escherichia fergusonii is an emerging potentially zoonotic organism which has been recovered from a broad range of human and animal sources. Efforts to recover E. fergusonii from mixed flora hitherto however have been constrained by the lack of a suitable selective medium for its isolation. This paper reports for the first time the recovery of E. fergusonii from reindeer carcases in a wildlife park and the use of citrate adonitol agar to selectively screen for the presence of this organism in faecal samples from further animals in the park, and reindeer in their natural habitat in Norway.

Topics: Agar; Animals; Citrates; Culture Media; Diarrhea; Enterobacteriaceae Infections; Escherichia; Female; Male; Reindeer

Topics: Agar; Anti-Bacterial Agents; Culture Media; Drug Resistance, Bacterial; Enterobacter; Enterobacter aerogenes; Enterobacter cloacae; Enterobacteriaceae Infections; Humans; In Vitro Techniques; Microbial Sensitivity Tests; Minocycline; Tigecycline

The aim of this study was to evaluate the performance of Brilliance ESBL agar (OX; Oxoid, Basingstoke, United Kingdom), a novel chromogenic agar for the selective isolation and the presumptive identification of extended-spectrum-beta-lactamase (ESBL)-producing Enterobacteriaceae. A panel of 200 clinical Gram-negative Enterobacteriaceae and nonfermenting isolates with defined resistance mechanisms was inoculated onto OX and onto ChromID ESBL agar (BM; bioMérieux, Marcy l'Etoile, France) chromogenic medium in the first part of the study to evaluate the growth selectivity and chromogenic features of these two media. Of the 156 Enterobacteriaceae challenge isolates, 8 fully susceptible isolates were inhibited, all 98 ESBL producers were detected, and 50 isolates harboring other resistance mechanisms were recovered on both chromogenic agars. In the second phase, 528 clinical samples (including 344 fecal specimens) were plated onto OX, BM, and MacConkey agar with a ceftazidime disk (MCC) for the screening of ESBL-producing Enterobacteriaceae. Growth on at least one medium was observed with 144 (27%) of the clinical samples screened. A total of 182 isolates, including 109 (60%) of Enterobacteriaceae, were recovered and 70 of these (from 59 specimens) were confirmed as ESBL-producing isolates. The sensitivities of MCC, BM, and OX were 74.6%, 94.9%, and 94.9%, respectively. The specificities of MCC, BM, and OX by specimens reached 94.9%, 95.5%, and 95.7%, respectively, when only colored colonies were considered on the two selective chromogenic media. The high negative predictive value (99.3%) found for OX suggests that this medium may constitute an excellent screening tool for the rapid exclusion of patients not carrying ESBL producers.

Topics: Agar; Bacteriological Techniques; beta-Lactamases; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Humans; Mass Screening; Predictive Value of Tests; Sensitivity and Specificity

Topics: Agar; Bacteremia; Bacterial Proteins; Bacteriological Techniques; beta-Lactamases; Blood; Cephalosporin Resistance; Cephalosporins; Chromogenic Compounds; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Humans; Microbial Sensitivity Tests; R Factors; Sensitivity and Specificity

Enterobacter sakazakii is associated with neonatal infections and is occasionally present at low levels (<1 CFU/g) in powdered infant formula milk (IFM). It has been previously reported that some E. sakazakii strains do not grow in standard media for Enterobacteriaceae and coliform bacteria; therefore, a reliable method is needed for recovery of the organism. Three E. sakazakii enrichment broths-Enterobacteriaceae enrichment broth (EE), E. sakazakii selective broth (ESSB), and modified lauryl sulfate broth (mLST)-were compared with a novel broth designed for maximum recovery of E. sakazakii, E. sakazakii enrichment broth (ESE). One hundred seventy-seven strains (100%) grew in ESE, whereas between 2 and 6% of strains did not grow in EE, mLST, or ESSB. E. sakazakii possesses alpha-glucosidase activity, and a number of selective, chromogenic agars for E. sakazakii isolation based on this enzyme have been developed. E. sakazakii isolation agar produced fewer false-positive colonies than did Druggan-Forsythe-Iversen agar. However, the latter supported the growth of more E. sakazakii strains. It was also determined that 2% of E. sakazakii strains did not produce yellow pigmentation on tryptone soya agar at 25 degrees C, a characteristic frequently cited in the identification of E. sakazakii. The recovery of desiccated E. sakazakii (0.2 to 2000 CFU/25 g) from powdered IFM in the presence of a competing flora was determined with various enrichment broths and differential selective media. Current media designed for the isolation and presumptive identification of E. sakazakii do not support the growth of all currently known E. sakazakii phenotypes; therefore, improvements in the proposed methods are desirable.

Topics: Agar; Bacteriological Techniques; Colony Count, Microbial; Cronobacter sakazakii; Culture Media; Enterobacteriaceae Infections; Environmental Microbiology; Food Contamination; Humans; Infant; Infant Food; Infant Formula

Increasing antibiotic resistance in gram-negative bacteria has recently renewed interest in colistin as a therapeutic option. The increasing use of colistin necessitates the availability of rapid and reliable methods for colistin susceptibility testing. We compared seven methods of colistin susceptibility testing (disk diffusion, agar dilution on Mueller-Hinton [MH] and Isosensitest agar, Etest on MH and Isosensitest agar, broth microdilution, and VITEK 2) on 102 clinical isolates collected from patient materials during a selective digestive decontamination or selective oral decontamination trial in an intensive-care unit. Disk diffusion is an unreliable method to measure susceptibility to colistin. High error rates and low levels of reproducibility were observed in the disk diffusion test. The colistin Etest, agar dilution, and the VITEK 2 showed a high level of agreement with the broth microdilution reference method. Heteroresistance for colistin was observed in six Enterobacter cloacae isolates and in one Acinetobacter baumannii isolate. This is the first report of heteroresistance to colistin in E. cloacae isolates. Resistance to colistin in these isolates seemed to be induced upon exposure to colistin rather than being caused by stable mutations. Heteroresistant isolates could be detected in the broth microdilution, agar dilution, Etest, or disk diffusion test. The VITEK 2 displayed low sensitivity in the detection of heteroresistant subpopulations of E. cloacae. The VITEK 2 colistin susceptibility test can therefore be considered to be a reliable tool to determine susceptibility to colistin in isolates of genera that are known not to exhibit resistant subpopulations. In isolates of genera known to (occasionally) exhibit heteroresistance, an alternative susceptibility testing method capable of detecting heteroresistance should be used.

Topics: Acinetobacter baumannii; Acinetobacter Infections; Agar; Anti-Bacterial Agents; Bacterial Infections; Colistin; Cross Infection; Culture Media; Drug Resistance, Multiple, Bacterial; Enterobacter cloacae; Enterobacteriaceae Infections; Humans; Intensive Care Units; Microbial Sensitivity Tests; Polymyxin B

Lactobacilli produce many antimicrobial substances including organic acids, hydrogen peroxide and bacteriocins. Antagonistic activity of lactobacilli is an important factor in the protection of the vagina of fertile women against infection by certain pathogens. In the present study, we investigated 17 strains of lactobacilli, including 11 strains of vaginal origin. The aim was to investigate in more detail the antibacterial activity of lactobacilli and to attempt to assess substances responsible for inhibition. The investigated lactobacilli inhibited some strains of Escherichia coli, Serratia marcescens, Shigella boydii, Listeria monocytogenes, Listeria ivanovii, Listeria innocua and Staphylococcus aureus. We have provided evidence that inhibition is due mainly to organic acids and to a lesser extent, to bacteriocins.

Topics: Agar; Antibiosis; Bacteriocins; Carboxylic Acids; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Female; Gram-Positive Bacteria; Gram-Positive Bacterial Infections; Humans; Hydrogen Peroxide; Lactobacillus; Vagina

The aim of the study was to evaluate the chromogenic agar plate CPS ID2 (bioMérieux) and determine its cost-benefit ratio.. A total of 2,193 urinary sediments were processed. The urine culture was carried out in CPS ID2 agar and in cystine-lactose electrolyte deficient (CLED) agar, when needed. Identification of the microorganisms was performed following standard microbiologic procedures through biochemical tests prepared in our laboratory. The identification, from CPS ID2 agar, by direct detection in medium of four metabolic activities: beta-glucuronidase, beta-glucosidase, deaminase, and indol production, was performed following to manufacturer's instructions.. A total of 289 urine cultures were positive, 18 were negative and 34 were contaminated samples. The identification, directly performed from the colonies detected in CPS ID2 agar, was correct in 96% of 166 Escherichia coli, in 92% of 24 Proteus mirabilis and in 97% of 38 enterococci. CPS ID2 agar exhibited 94% and 100% sensitivity and specificity, respectively in E. coli identification, 92% and 100% in P. mirabilis and 97% and 99% in Enterococcus. The use of this new media, CPS ID2, in our laboratory, implies a budgetary increment. However, if commercial galleries are used for routine identification, the cost will be reduced using this new media.. The CPS ID2 agar allows the isolation and direct identification of the most frequent urinary tract pathogens: E. coli, P. mirabilis and Enterococcus in primary isolation medium. Using this medium, bacteriologists will be able to save time and reagents when identifying the most common uropathogens. Furthermore, the use of this medium would reduce costs in some laboratories.

Topics: Agar; Amino Acid Oxidoreductases; Bacterial Proteins; Bacteriological Techniques; beta-Glucosidase; Candida albicans; Candidiasis; Chromogenic Compounds; Cost-Benefit Analysis; Culture Media; Enterobacteriaceae; Enterobacteriaceae Infections; Evaluation Studies as Topic; Glucuronidase; Gram-Negative Bacteria; Gram-Negative Bacterial Infections; Gram-Positive Bacteria; Gram-Positive Bacterial Infections; Humans; Indoles; L-Amino Acid Oxidase; Urinary Tract Infections; Urine

From 1 February through 12 October 1990, 27 blood cultures processed at Shiprock Hospital were positive for Enterobacter cloacae; only 3 had been reported in the preceding 12 months. Twenty (74%) of the cultures were obtained from patients without clinical evidence of gram-negative septicemia. The increase in E. cloacae-positive blood cultures was temporally associated with the introduction of a new blood culturing system. To evaluate potential risk factors for an E. cloacae-positive blood culture (case-culture), we conducted a case-control study. Case-cultures were compared with 81 randomly selected cultures that were processed during the epidemic period and that were not positive for E. cloacae (controls). Because several factors suggested the possibility of pseudoinfection, we limited our analysis to the 20 blood cultures that appeared to be contaminants. Blood samples received in the laboratory during the midnight shift (5 of 20 [25%] versus 5 of 81 [6%]; odds ratio, 5.1; 95% confidence intervals, 1.01 to 24.6; P = 0.02) or present in the incubator with other E. cloacae-positive samples (17 of 20 [85%] versus 29 of 81 [36%]; odds ratio, 10.2, 95% confidence interval, 2.6 to 57.3; P < 0.001) were at increased risk for contamination. During mock experiments of the procedures for processing blood samples for culture, several breaks in aseptic technique and leakage from the blood culturing system were observed. Cultures of samples obtained from several environmental sites in the laboratory and the hand washings of two laboratory technicians grew E. cloacae. Plasmid and restriction enzyme analyses of E. cloacae isolates recovered from the patients' blood cultures, the two technicians' hand washings, and environmental sites in the laboratory indicated that all had identical plasmid profiles. Our findings suggest that the breaks in aseptic technique and the environmental contamination that occurred in association with the use of the new blood culturing system resulted in contamination of the blood cultures. This outbreak highlights the importance of routine environmental cleaning, periodic quality control assessments, and adherence to aseptic practices in clinical laboratories, particularly when new methods or equipment are introduced and/or new personnel are hired.

Topics: Agar; Bacteremia; Bacteriological Techniques; Case-Control Studies; Cluster Analysis; Disease Outbreaks; Enterobacter cloacae; Enterobacteriaceae Infections; Humans; Quality Control

The production of a precipitate by Serratia marcescens on Oxoid MacConkey agar has proven useful as a laboratory diagnostic test. This phenomenon is specific for Serratia within the Enterobacteriaceae, although precipitate production is also given by Acinetobacter anitratus and some Pseudomonas, Alcaligenes, and Aeromonas species. Precipitate production seems to be specific for certain batches of MacConkey agar, and is probably related to a specific property of some batches of bile salts.

Topics: Agar; Bile Acids and Salts; Diagnosis, Differential; Enterobacteriaceae Infections; Humans; Hydrogen-Ion Concentration; Serratia marcescens

A technique is described that allows presumptive identification of either Salmonella or Shigella organisms directly upon the original isolation plate, in this case, MacConkey agar. This was accomplished by applying a drop of specifically sensitized protein A-containing Staphylococcus aureus over a "suspected" colony or several colonies of organisms grown on MacConkey agar. The plate is tilted to and fro to allow mixing of the particles with specific antigen that ir readily solubilized from the colony and observing for agglutination of the sensitized particles by use of a dissecting microscope. The agglutination can frequently be seen within 15 s, increasing in intensity over a 2-min period. The polyvalent Salmonella antiserum was slower in developing strong agglutination (1.5 to 2 min) compared to particles sensitized with group-specific antisera (15 to 45 s). A high-titer antiserum was important for a test reagent to have the required sensitivity.

Topics: Agar; Agglutination Tests; Antigens, Bacterial; Bacterial Proteins; Bacteriological Techniques; Enterobacteriaceae Infections; Epitopes; Humans; Salmonella; Shigella; Staphylococcus aureus

Bile-esculin agar has been used for several years for the presumptive identification of group D streptococci. All members of the Enterobacteriaceae family will also grow on this medium, but only certain ones can hydrolyze esculin to 6,7-dihydroxycoumarin, which reacts with iron to produce a characteristic blackening of the medium. One thousand and six cultures from clinical specimens representing 20 genera were isolated and identified. Heavy inocula from fresh pure culture isolates on heart infusion agar were placed on bile-esculin agar slants and incubated at 35 C. The slants were examined at 4 h and again at 18 h for esculin hydrolysis. Shigella, Salmonella, Arizona, Proteus mirabilis, Proteus morganii, Providencia alcalifaciens, and Providencia stuartii all produced negative results. Klebsiella pneumoniae, Enterobacter aerogenes, Serratia marcescens, and Serratia rubidaea produced a positive reaction in 4 h. The other remaining eight genera exhibited varying results. The use of this medium in conjunction with triple sugar iron-lysine iron agar has been of great value in differentiating the Klebsiella-Enterobacter-Serratia group from other Enterobacteriaceae.

Topics: Agar; Bile; Enterobacteriaceae; Enterobacteriaceae Infections; Esculin; Evaluation Studies as Topic; Humans; Hydrolysis

Topics: Adolescent; Adult; Agar; Aged; Animals; Anti-Bacterial Agents; Bacteriological Techniques; Bile; Caniformia; Carrier State; Child; Child, Preschool; Citrates; Disease Reservoirs; Drug Resistance, Microbial; Enterobacteriaceae; Enterobacteriaceae Infections; Feces; Female; Gastroenteritis; Humans; Indoles; Infant; Male; Middle Aged; Rectum; Reptiles; Sepsis; Thailand; Zoonoses

A 10-minute test, utilizing a urease paper-reagent strip (PATHO-TEC), for differentiating Klebsiella and Enterobacter species is described. By using a heavy suspension of organisms and 50 C temperature for incubation, 93% of Klebsiella strains (186/200) were positive and 95% of Enterobacter strains (190/200) were negative with this testing system. The rapid nature of the test (10 min), the facility with which it can be carried out, and the ease with which the strips can be stored and handled may make this a useful aid for the clinical microbiologist.

Topics: Agar; Bacteriological Techniques; Diagnosis, Differential; Enterobacter; Enterobacteriaceae Infections; Humans; Indicators and Reagents; Klebsiella; Klebsiella Infections; Paper; Urease

Topics: Agar; Animals; Anti-Bacterial Agents; Culture Media; Drug Resistance, Microbial; Drug Synergism; Enterobacteriaceae; Enterobacteriaceae Infections; Escherichia coli Infections; Folic Acid Antagonists; Haemophilus Infections; Meningococcal Infections; Mice; Microbial Sensitivity Tests; Pneumococcal Infections; Pyrimidines; Streptococcal Infections; Sulfamethoxazole