Multilaboratory comparison of growth characteristics for anaerobes, using 5 different agar media.

A multilaboratory study compared the growth of 30 fastidious anaerobes, using 5 different agar media: Wilkins-Chalgren (WC), WC with either whole or laked sheep blood, and Brucella supplemented with vitamin K(1) and hemin and either laked or whole sheep blood. The media were compared for quality and quantity of growth. Experiments were conducted either entirely in an anaerobic chamber or inoculated in ambient air with anaerobic incubation. The results showed that (1) any medium plus whole or laked blood was better than unsupplemented WC, (2) whole blood and laked blood additives gave similar results, (3) supplemented Brucella with whole or laked blood was superior to WC and WC with whole or laked blood, and (4) anaerobic and aerobic inoculation with anaerobic incubation gave similar results. Brucella agar supplemented with whole or laked blood supports the growth of fastidious anaerobic species better than the WC agars do.

Multilaboratory comparison of anaerobe susceptibility results using 3 different agar media.

A 5-laboratory study was performed that used the National Committee for Clinical Laboratory Standards (NCCLS) reference agar dilution method with 3 media formulations to determine whether the use of different media would affect minimum inhibitory concentration (MIC) results. Wilkins-Chalgren, Brucella-based blood agar (BRU), and Wilkins-Chalgren agar plus blood (WCB) and 6 antibiotics (clindamycin, cefoxitin, ceftizoxime, piperacillin, metronidazole, and trovafloxacin) were evaluated with 58 isolates. The MIC values were compared, and a significant correlation of >0.80 was demonstrated for all media and each antibiotic/organism group. The cumulative rate of errors for all antibiotics was 0.1%. These data indicate that a change in the NCCLS reference medium for testing of anaerobic bacteria susceptibility to either BRU or WCB will not affect the MIC results for the antibiotics and organisms evaluated.

Characterization of tetracycline and erythromycin resistance in Actinobacillus pleuropneumoniae.

Minimum inhibitory concentrations to tetracycline and erythromycin were determined for nineteen isolates of Actinobacillus pleuropneumoniae of Norwegian origin. The isolates were screened for rRNA methylase determinants (Erm genes) and for the tetracycline resistance Tet B determinant, using oligonucleotide probes, polymerase chain reaction and hybridization. Ten isolates (53%) carried the Erm C determinant, two isolates (10%) carried the Erm A determinant, four isolates (21%) carried both the Erm A and the Erm C determinants, and three isolates (16%) carried none of the Erm determinants examined. Eight isolates (45%) carried the Tet B determinant. Selected isolates were shown to transfer the Erm C and Erm A determinants at a frequency of 10(-7)-10(-9) per recipient cell. This is the first description of A. pleuropneumoniae carrying either Erm A, Erm C or/and Tet B determinants.

Mobile rRNA methylase genes coding for erythromycin resistance in Actinobacillus actinomycetemcomitans.

We found that 17 (68%) of 25 Actinobacillus actinomycetemcomitans isolated from periodontally diseased sites had MICs of erythromycin > or = 8 mg/L. The isolates were hybridized with five rRNA methylase genes and gene mobility was determined. Twenty-four (96%) of 25 isolates hybridized with one or more rRNA methylase genes. Representative isolates were able to transfer erythromycin resistance to Haemophilus influenzae and Enterococcus faecalis recipients. The transconjugants were shown to carry rRNA methylase genes by DNA probe hybridization and had MICs of erythromycin 32 to 256 mg/L.

Characterization of tetracycline and erythromycin resistance determinants in Treponema denticola.

Treponema denticola isolates were evaluated for the presence of known tetracycline and erythromycin resistance determinants by Southern blot hybridization of whole-cell DNA and PCR assays. We examined all isolates available, which included 12 clinical and 4 American Type Culture Collection isolates. Two isolates carried the Tet B determinant, five isolates carried both the Tet B and Erm F determinants, seven isolates carried the Erm F determinant, and two did not carry any of the Tet or Erm determinants tested. Both the Tet B and Erm F determinants appeared to be associated with the chromosome. Neither of the two T. denticola donors tested could transfer the Tet B determinant, but three of four T. denticola tested transferred the Erm F determinant to an Enterococcus faecalis recipient. This extends the host range of both the tetB and ermF genes into the genus Treponema.

Characterization of tetracycline resistance in Actinobacillus actinomycetemcomitans.

Comparison of susceptibility data for Actinobacillus actinomycetemcomitans has been difficult because of the lack of standard susceptibility testing conditions. In this study, minimum inhibitory concentration to tetracycline was evaluated by comparing different media, air conditions and incubation times. Ten of 22 (45%) A. actinomycetemcomitans isolated from periodontally diseased sites grew on media supplemented with 4 micrograms per ml of tetracycline, but minimum inhibitory concentrations ranged from 0.125 to 8 micrograms/ml depending on the media and condition used. The best results were obtained with brain heart infusion agar (Difco Laboratories, Detroit MI) incubated in 5% CO2 for 48 h. Eighteen (82%) of the A. actinomycetemcomitans isolates hybridized with the Tet B determinant. The Tet B determinant was transferable between A. actinomycetemcomitans isolates as well as a Haemophilus influenzae recipient and appears to be associated with conjugative plasmids.