agar and Bacteroides-Infections

We aimed to identify the enterotoxigenic Bacteroides fragilis (ETBF) and bft subtypes among patients with diarrhea. In addition, we assessed whether DNA gyrase subunit B (gyrB) and neuraminidase (nanH) genes are useful determinants for identification of B. fragilis compared to 16S rRNA sequencing as a reference method.. The 530 fecal specimens were cultured on BBE agar. The colonies which supposed to be a member of B. fragilis group were subjected to 16S rRNA gene sequencing and PCR assays targeting the Bacteroides fragilis group (BFG), gyrB and nanH. The B. fragilis toxin (bft) gene and its subtype was detected by PCR. The specificity of PCR assays was calculated considering the 16S rRNA gene sequencing as the reference method.. A total of 111 Gram-negative anaerobic coccobacilli were isolated from 530 fecal specimens using BBE agar. Of the 111 isolates, 100 (90.09%) were assumed to be a member of Bacteroides fragilis group as they yielded an amplicon through PCR using the group-specific primers (Bfra-F/g-Bfra-R). However, only 28 isolates out of 100 were definitively identified as species of Bacteroides using16S rRNA gene sequencing; of which 15 isolates were B. fragilis and the remaining 13 isolates were identified as B. thetaiotaomicron (n = 6), Parabacteroides distasonis (n = 3), B. vulgatus (Phocaeicola vulgatus) (n = 1), B. ovatus (n = 1), B. congonensis (n = 1) and B. nordii (n = 1). Among the 15 isolates of B. fragilis, 4 were found to be ETBF. Compared to the reference method, the specificity and accuracy of the PCR targeting gyrB gene (64.7% and 65%) was higher than of nanH (36.4% and 46%, respectively.. This study demonstrated that more than one-fourth of B. fragilis isolates harbored bft gene and less than 1% of patients with diarrhea harbored ETBF. The slight agreement between the PCR assays -already used for identification of B. fragilis which targeting gyrB or nanH - and 16S rRNA gene sequencing as the reference method was noted.

Topics: Agar; Bacterial Infections; Bacteroides fragilis; Bacteroides Infections; Diarrhea; Humans; Neuraminidase; RNA, Ribosomal, 16S

To compare the performance of agar dilution and broth microdilution by commercial and in-house prepared plates for the Bacteroides fragilis group. The cost analysis was performed to demonstrate that in-house prepared BMD plates were a suitable alternative to agar dilution given the high cost and low feasibility of incorporating commercial BMD plates in routine, particularly in the tertiary care institutes of many low- and middle-income countries.. Thirty B. fragilis group isolates were tested against six antibiotics, frequently used as empirical therapy for anaerobic infections including metronidazole, clindamycin, imipenem, piperacillin-tazobactam, cefoxitin, and chloramphenicol. The running consumable expenditure for all methodologies was calculated.. The results demonstrated essential and categorical agreement of >90% for all antibiotics except cefoxitin, which showed <90% categorical agreement. No major or very major errors were observed. We observed a high agreement and strong concordance for MIC values between both methods and inter-rate reliability of >0.9 by Cohen's kappa analysis, indicating almost perfect agreement between both methods using either of the plates. In contrast to agar dilution, a 20.5 fold cost reduction was seen in BMD using in-house plates and a 5.8 fold reduction using commercial plates to test a single isolate. However, when testing 30 isolates concurrently the cost significantly increased for commercial BMD plates by 8.4 folds, and only 1.03 fold cost reduction was seen with in-house BMD plates.. BMD gives comparable results to agar dilution and can be considered a method of choice to test a small number of samples. The technique is an economical option when plates are standardized in-house and could be employed for susceptibility testing of the B. fragilis group.

Topics: Agar; Anti-Bacterial Agents; Bacteroides fragilis; Bacteroides Infections; Clindamycin; Humans; Imipenem; Metronidazole; Microbial Sensitivity Tests

There have been an increasing number of reports describing the acquisition of antimicrobial resistance by Bacteroides fragilis including the occurrence of strains with resistance to multiple antimicrobials that are relied upon for treatment of infections. The aim of this study was to design a chromogenic selective medium for isolation of B. fragilis that could be adapted for specific isolation of antimicrobial-resistant strains. Bacteroides chromogenic agar (BCA) was the result of this endeavour and allowed growth of Bacteroides spp. as black colonies and the efficient inhibition of almost all other genera tested. The medium also allowed some differentiation of B. fragilis from other members of the B. fragilis group. When compared with an adaptation of Bacteroides bile-esculin agar (BBE) for the isolation of B. fragilis from 100 stool samples, 30 isolates of B. fragilis were recovered on BCA compared with 19 isolates recovered on BBE (P = 0.022). When supplemented with meropenem (4 μg/ml) or metronidazole (2 μg/ml), BCA could be used to select for the growth of B. fragilis isolates with resistance to these agents. We conclude that BCA is a useful research tool for surveillance studies to assess the prevalence of B. fragilis and, in particular, the occurrence of antimicrobial-resistant strains.

Topics: Agar; Anti-Bacterial Agents; Bacteroides fragilis; Bacteroides Infections; Chromogenic Compounds; Drug Resistance, Multiple, Bacterial; Esculin; Feces; Humans; Microbial Sensitivity Tests

With the emergence of antibiotic resistance among Bacteroides fragilis group isolates the need of susceptibility testing in routine laboratories is increasing. The aims of the present study were to evaluate the disk diffusion method for susceptibility testing in case of different clinical isolates of Bacteroides spp by comparing zone diameter results with MICs obtained earlier during an Europe-wide antibiotic susceptibility surveillance, and to propose zone diameter breakpoints, which correlate for the EUCAST MIC breakpoints. We tested 381 clinical isolates of the B. fragilis group to amoxicillin/clavulanic acid, cefoxitin, clindamycin, imipenem, metronidazole, moxifloxacin, piperacillin/tazobactam, tigecycline by agar dilution method previously. The inhibition zones of the same antibiotics including meropenem disc were determined by the disc diffusion on Brucella blood agar supplemented with haemin and vitamin K1. Plates were incubated at 37 °C in an anaerobic atmosphere for 24 h. The zone diameters were read at 100% inhibition. In case of discrepant results MICs were determined by gradient test and compared with the inhibition zones on the same plate. We found a good agreement between the inhibition zone diameters and the MICs for imipenem, metronidazole, moxifloxacin and tigecyclin. The inhibition zone diameters of meropenem also separated clearly the isolates, which can be considered wild-type isolates. In case of amoxicillin/clavulanic acid and piperacillin/tazobactam intermediate and susceptible isolates according to the MIC determination, overlap during the zone diameter determination. Isolates with an inhibition zone <23 mm for amoxicillin/clavulanic acid and <25 mm for piperacillin/tazobactam should be retested by a MIC determination method. The 10 μg clindamycin disc clearly separated the resistant and the susceptible population of B. fragilis group strains. In the case of cefoxitin only resistant population could be separated with an inhibition zone <17 mm, intermediate and susceptible isolates overlap. In conclusion, we suggest that disk diffusion can be an option for susceptibility testing of B. fragilis group isolates for most relevant antibiotics in routine laboratories.

Topics: Agar; Anaerobiosis; Anti-Bacterial Agents; Bacteroides fragilis; Bacteroides Infections; Culture Media; Europe; Humans; Microbial Sensitivity Tests; Temperature

A new medium selective for Bacteroides gracilis was developed. The medium is tryptic soy agar (Difco Laboratories, Detroit, Mich.) containing nalidixic acid, teicoplanin, sodium formate, sodium fumarate, and potassium nitrate. All 18 strains of B. gracilis tested grew with only minimal inhibition. Most of the other 214 organisms tested, including most Bacteroides species, other anaerobes, and a substantial number of facultative anaerobes, were significantly inhibited by the medium. In a diagnostic study of 49 clinical specimens (28 patients with intra-abdominal infection, mostly gangrenous or perforated appendicitis), four strains of B. gracilis were isolated (from 4 different patients) on B. gracilis selective agar but were not detected on standard media.

Topics: Agar; Anti-Bacterial Agents; Appendicitis; Bacteroides; Bacteroides Infections; Caseins; Culture Media; Formates; Fumarates; Gangrene; Glycopeptides; Humans; Nalidixic Acid; Nitrates; Potassium Compounds; Protein Hydrolysates; Saliva; Teicoplanin

The development of a pure Bacteroides fragilis infection in mice is described. The infection produces large subcutaneous abscesses at the site of injection which can be observed grossly within 7 days after injection. The infection was initiated by infection of pure cultures grown in semisolid agar medium. Similar infections were also produced with pure cultures of B. distasonis, B. ovatus, B. thetaiotaomicron, and B. vulgatus. However, a distinct deoxyribonucleic acid homology group, formerly classified as B. thetaiotaomicron, did not produce abscesses in any of the mice tested.

Topics: Agar; Animals; Bacteroides; Bacteroides fragilis; Bacteroides Infections; Base Sequence; Disease Models, Animal; DNA, Bacterial; Male; Mice

Metronidazole was found to be active against Bacteroides fragilis strains isolated from human lesions. The minimal inhibitory concentrations (MIC) were from 0.16 to 2.5 mug/ml and the minimal bactericidal concentrations (MBC) were from 0.16 to 2.5 mug/ml; usually the MIC and MBC figures were equivalent. These levels are easily attainable in the serum following normal therapeutic doses. The drug is not toxic and side effects are rare and it would therefore seem highly suitable for treating Bacteroides infections and also may be considered prophylactically in certain situations that are described.

Topics: Agar; Animals; Bacteroides; Bacteroides Infections; Blood; Culture Media; Methods; Metronidazole; Mice; Microbial Sensitivity Tests; Rabbits

Topics: Agar; Bacteriocins; Bacteriological Techniques; Bacteroides; Bacteroides Infections; Blood; Culture Media

Topics: Agar; Anti-Bacterial Agents; Bacteroides; Bacteroides Infections; Diffusion; Drug Resistance, Microbial; Humans; Microbial Sensitivity Tests; Species Specificity