Comparison of different media for the identification and quantification ofAeromonas spp. in water

In order to assess the suitability of the Starch glutamate ampicillin penicillin-10C agar for the isolation ofAeromonas spp. from waters it was necessary to compare the properties of this medium with those of three others, Starch ampicillin agar, Ampicillin dextrin agar and m-Aeromonas medium, and to monitor different kinds of waters. A selection of forty eight samples were taken from moderately polluted river water, highly polluted river water, polluted sea water (littoral) and treatment & distribution water and monitored using these media. The results were similar with Ampicillin dextrin agar, m-Aeromonas medium and Starch glutamate ampicillin penicillin-10C, but the simplicity of composition and use and its selectivity recommends the last medium as the most adequate for the isolation ofAeromonas spp.Abbreviations ADA ampicillin dextrin agar - mA m-aeromonas medium - SA starch ampicillin agar - SGAP-10C starch glutamate ampicillin penicillin-10C     

Immunity to Aerosol Challenge in Guinea Pigs Immunized with Gamma-Irradiated Venezuelan Equine Encephalitis Vaccines

An improved selective medium, Tryptose-sulfite-cycloserine (TSC) agar, for the enumeration of Clostridium perfringens is described. It consists of the same basal medium as Shahidi-Ferguson-perfringens (SFP) agar, but with 400 mug of D-cycloserine per ml substituted for the kanamycin and polymyxin. Tolerance of C. perfringens for D-cycloserine, its production of lecithinase, and its ability to reduce sulfite were used as the basis for development of this medium. Comparisons were made between TSC and SFP agars for the recovery of vegetative cells of C. perfringens by using statistical methods. The results showed that TSC allowed virtually complete recovery of most of the C. perfringens strains while inhibiting practically all facultative anaerobes tested. SFP agar allowed a slightly higher rate of recovery of C. perfringens but was found to be much less selective.     

Agarose soy casein digest medium for replacement of blood agar for potency determinations of live Pasteurella vaccines

Blood agar, prepared with Trypticase (BBL Microbiology Systems, Cockeysville, Md.) soy agar and 5% defibrinated bovine blood, is used for testing the potency of live Pasteurella multocida and Pasteurella haemolytica vaccines, but its potential for variation makes it undesirable to use in a standard assay method. Tests done with RPMI 1640 and Trypticase soy medium indicated that the benefits obtained by adding defibrinated blood to the Trypticase soy agar medium were more likely due to neutralization of toxic components than to the presence of transferrin or iron as growth factors. Reduction of toxic components in the Trypticase soy agar medium was accomplished by replacing agar with agarose and by autoclaving glucose as a separate solution to produce the replacement medium. The replacement medium was prepared by autoclaving three separate solutions--Trypticase soy broth without glucose; glucose; and agarose--cooling to 55 degrees C, and mixing and then pouring the mixtures into petri dishes. The growth obtained with this medium as judged by determination of the number of CFU and the colony sizes of P. multocida or P. haemolytica was equal to or better than those obtained with blood agar.     

Recovery of Spores of Bacillus stearothermophilus from Thermal Injury

Bacillus stearothermophilus grown in nutrient broth produced a product which promoted recovery from thermal injury of its spores. This phenomenon was observed with nutrient agar as the plating medium but not with a medium composed of Trypticase, Phytone, dextrose and phosphate (TPDP). Recovery of injured spores was greatest in such a medium if it contained starch or charcoal. Trypticase soy agar and dextrose tryptone agar were markedly inferior to TPDP medium.     

Agarose soy casein digest medium for replacement of blood agar for potency determinations of live Pasteurella vaccines.

Blood agar, prepared with Trypticase (BBL Microbiology Systems, Cockeysville, Md.) soy agar and 5% defibrinated bovine blood, is used for testing the potency of live Pasteurella multocida and Pasteurella haemolytica vaccines, but its potential for variation makes it undesirable to use in a standard assay method. Tests done with RPMI 1640 and Trypticase soy medium indicated that the benefits obtained by adding defibrinated blood to the Trypticase soy agar medium were more likely due to neutralization of toxic components than to the presence of transferrin or iron as growth factors. Reduction of toxic components in the Trypticase soy agar medium was accomplished by replacing agar with agarose and by autoclaving glucose as a separate solution to produce the replacement medium. The replacement medium was prepared by autoclaving three separate solutions--Trypticase soy broth without glucose; glucose; and agarose--cooling to 55 degrees C, and mixing and then pouring the mixtures into petri dishes. The growth obtained with this medium as judged by determination of the number of CFU and the colony sizes of P. multocida or P. haemolytica was equal to or better than those obtained with blood agar.     

Evaluation of Chromocult coliform agar for the detection and enumeration of Enterobacteriaceae from faecal samples from healthy subjects

The purpose of this study was to examine the use of Chromocult agar medium for isolation and enumeration of Enterobacteriaceae from human faecal samples, to compare it to MacConkey agar and to evaluate its usefulness as a possible alternative selective medium in human faecal studies. The medium was shown to be effective in identifying Escherichia coli and coliforms in faeces without the need for extensive accompanying biochemical tests for confirmation of identity. A positive correlation (r=0.86) was found between the recovery of Enterobacteriaceae on the two media, and no significant difference (P>0.05) between overall mean bacterial counts for the whole study group or at different intervals of faecal collection were observed. Chromocult agar is an effective replacement for MacConkey agar in human faecal studies and has the advantage of differentiating E. coli from other coliforms.     

Evaluation of several selective media for recovery of Aeromonas hydrophila from polluted waters.

Eleven media were studied for their suitability in the selective isolation of Aeromonas hydrophila. Preliminary results showed that five of them (inositol-brilliant green-bile salts agar, bile salts-brilliant green agar, Rimler-Shotts agar, xylose-sodium deoxycholate-citrate agar, and dextrin-fuchsin-sulfite agar) allowed the growth of several microorganisms that are usually present in the same samples in which A. hydrophila is found. Six media (mA agar, modified Rimler-Shotts agar, DNase-toluidine blue-ampicillin agar, Pril-xylose-ampicillin agar, MacConkey-trehalose agar, and starch-bile salts agar) were selected for evaluation as recovery selective media on the basis of their efficiency in the isolation of A. hydrophila from natural water samples. mA agar showed the best recovery rate and also an acceptable specificity, but its selectivity was low. Another medium that can be considered is DNase-toluidine blue-ampicillin agar, which showed good accuracy, but its specificity was low.     

Evaluation of several selective media for recovery of Aeromonas hydrophila from polluted waters

Eleven media were studied for their suitability in the selective isolation of Aeromonas hydrophila. Preliminary results showed that five of them (inositol-brilliant green-bile salts agar, bile salts-brilliant green agar, Rimler-Shotts agar, xylose-sodium deoxycholate-citrate agar, and dextrin-fuchsin-sulfite agar) allowed the growth of several microorganisms that are usually present in the same samples in which A. hydrophila is found. Six media (mA agar, modified Rimler-Shotts agar, DNase-toluidine blue-ampicillin agar, Pril-xylose-ampicillin agar, MacConkey-trehalose agar, and starch-bile salts agar) were selected for evaluation as recovery selective media on the basis of their efficiency in the isolation of A. hydrophila from natural water samples. mA agar showed the best recovery rate and also an acceptable specificity, but its selectivity was low. Another medium that can be considered is DNase-toluidine blue-ampicillin agar, which showed good accuracy, but its specificity was low.     

Comparison of Selective Media for Isolation of Presumptive Group D Streptococci from Human Feces

Pfizer Selective Enterococcus (PSE) agar, a medium containing bile, sodium azide, and esculin, was evaluated for its sensitivity and selectivity for detection and enumeration of presumptive group D streptococci in human feces. SF broth and SF broth plus agar (1.5%), representing selective media in common use, were studied simultaneously. Presumptive group D streptococci were recovered on PSE agar from the feces of all 25 subjects. No growth was observed in 8% of specimens in SF broth. No gram-negative organisms were recovered in any medium. PSE agar has the advantages of selecting out Streptococcus bovis, earlier appearance of distinctive reactions, and lack of requirement for special incubation temperature.     

Evaluation of a pour-plate system with a rabbit plasma-bovine fibrinogen agar for the enumeration of Staphylococcus aureus in food

Investigations were carried out concerning the selectivity and productivity of rabbit plasma fibrinogen (RPF) agar according to Beckers et al. (H. J. Beckers, F. M. van Leusden, W. M. Hogeboom, and E. H. M. Delfgou-van Asch. 1980. De Ware(n)-Chemicus, 10: 125-130). Its selectivity was compared with pork plasma fibrinogen (PPF) medium according to Hauschild et al. (A. H. W. Hauschild, C. E. Park, and R. Hilsheimer. 1979. Can. J. Microbiol. 25: 1052-1057) and its productivity was compared with PPF medium and Baird-Parker's egg yolk tellurite glycine pyruvate (ETGP) agar. In total 139 samples of naturally contaminated foodstuffs were examined. RPF agar scored higher than ETGP agar; although only small (mean value of differences 0.09 log units), the differences were statistically significant. While no significant differences in sensitivity between RPF agar and PPF medium were encountered, RPF agar was statistically more selective than PPF medium. It is concluded that RPF agar is very suitable for the enumeration of Staphyloccus aureus in foods.     

New selective and differential medium for Vibrio cholerae and Vibrio vulnificus

Thiosulfate-citrate-bile salts-sucrose agar has been routinely used for the isolation of pathogenic vibrios, although its selectivity for Vibrio cholerae and Vibrio vulnificus is inadequate. Therefore, a new plating medium, cellobiose-polymyxin B-colistin agar, was developed for the isolation of these two species. Cellobiose-polymyxin B-colistin agar demonstrated a significant advantage over other media designed for the isolation or differentiation of vibrios: of both the 136 strains representing 19 Vibrio species and the marine isolates of the genera Pseudomonas, Flavobacterium, and Photobacterium, only V. vulnificus and V. cholerae were able to grow. Furthermore, the fermentation of cellobiose by V. vulnificus allowed for the easy differentiation of these two species. This medium offers significant potential as a selective and differential medium for these two pathogenic vibrios.     

New selective and differential medium for Vibrio cholerae and Vibrio vulnificus.

Thiosulfate-citrate-bile salts-sucrose agar has been routinely used for the isolation of pathogenic vibrios, although its selectivity for Vibrio cholerae and Vibrio vulnificus is inadequate. Therefore, a new plating medium, cellobiose-polymyxin B-colistin agar, was developed for the isolation of these two species. Cellobiose-polymyxin B-colistin agar demonstrated a significant advantage over other media designed for the isolation or differentiation of vibrios: of both the 136 strains representing 19 Vibrio species and the marine isolates of the genera Pseudomonas, Flavobacterium, and Photobacterium, only V. vulnificus and V. cholerae were able to grow. Furthermore, the fermentation of cellobiose by V. vulnificus allowed for the easy differentiation of these two species. This medium offers significant potential as a selective and differential medium for these two pathogenic vibrios.     

Evaluation of m-T7 agar as a fecal coliform medium.

m-T7 agar, designed to improve recoveries of injured total coliforms, was evaluated for its effectiveness as a fecal coliform medium. The time and temperature of preincubation were found to be crucial to the optimal recovery of fetal coliforms. Isolation rates for fecal coliforms on m-T7 agar from sewage effluents were the highest when plates were preincubated at 37 degrees C for 8 h before transfer to 44.5 degrees C for 12 h. The medium was found to produce consistently higher fecal coliform counts than all the other methods tested. Recoveries were 3.1 times greater than the standard m-FC method and 1.7 times greater than the two-layer enrichment, temperature acclimation procedure. Verification rates for fecal coliforms isolated on m-T7 agar averaged 89.0%, whereas verification rates for m-FC agar averaged only 82.8%. Both media isolated similar fecal coliform populations. The advantages of a single medium, highly effective for the isolation of both total and fecal coliforms, are discussed.     

Evaluation of m-T7 agar as a fecal coliform medium

m-T7 agar, designed to improve recoveries of injured total coliforms, was evaluated for its effectiveness as a fecal coliform medium. The time and temperature of preincubation were found to be crucial to the optimal recovery of fetal coliforms. Isolation rates for fecal coliforms on m-T7 agar from sewage effluents were the highest when plates were preincubated at 37 degrees C for 8 h before transfer to 44.5 degrees C for 12 h. The medium was found to produce consistently higher fecal coliform counts than all the other methods tested. Recoveries were 3.1 times greater than the standard m-FC method and 1.7 times greater than the two-layer enrichment, temperature acclimation procedure. Verification rates for fecal coliforms isolated on m-T7 agar averaged 89.0%, whereas verification rates for m-FC agar averaged only 82.8%. Both media isolated similar fecal coliform populations. The advantages of a single medium, highly effective for the isolation of both total and fecal coliforms, are discussed.     

Value of the K+ salt of carageenan as an agar substitute in routine bacteriological media

The K+ salt of carageenan has no distinct advantages as a gelling agent, but it compared favorably with agar in most of the media tested. The difficulty involved in the preparation of blood plates and the results obtained with this medium prohibit its complete acceptance as a substitute for agar in routine solid media. However, it could be a suitable substitute for agar in all other routine bacteriological media.     

Efficiency of Various Growth Media in Recovering Oral Bacterial Flora from Human Dental Plaque

MM10 sucrose blood agar (MM10 SB agar), N(2)C agar, Schaedler agar (SH agar), and mitis salivarius agar (MS agar) were tested for their ability to recover human dental plaque flora by a continuous anaerobic procedure and by a conventional anaerobic method. MM10 SB agar yielded higher recovery of bacteria from plaque samples as determined by the enumeration of colony-forming units (CFU). The CFU on N(2)C agar, SH agar, and MS agar were lower than MM10 SB agar when the continuous anaerobic procedure was used. The superior performance of MM10 SB agar was much more apparent when used for the cultivation of dental plaque by the conventional anaerobic method. Under these conditions the counts were consistently higher on MM10 SB agar as compared to the other media tested. However, the differential counts of Streptococcus sanguis and S. mutans from carious plaque samples were in general comparable on all culture media. Deletion of blood from MM10 SB agar did not lower counts. The elimination of dithiothreitol from this medium resulted in a significantly lower recovery of bacteria from the plaque samples when cultured by the conventional anaerobic method. The storage of MM10 SB agar for varying periods of time aerobic conditions did not seem to affect its performance. These findings suggest that MM10 SB agar is an ideal culture medium for the isolation, nonselective enumeration, and differential counts of bacteria present in normal and disease-associated plaques.     

USE of an ELECTRONIC HGMF INTERPRETER SYSTEM FOR ENUMERATION of NALIDIXIC ACID RESISTANT SALMONELLAE IN CHICKEN CAECA

An electronic counting system using hydrophobic grid-membrane filters (HGMF) and the HGMF Interpreter was evaluated for its usefulness in enumerating nalidixic acid resistant Salmonella in frozen chicken caeca. Salmonella recovery was equivalent on both Hektoen Enteric and EF-18 agars. However, the color of the Salmonella growth on EF-18 agar was more easily differentiated by the HGMF Interpreter electronic counting system. the study showed that a 4 h resuscitation on a nonselective medium was required in order to maximize the subsequent recovery on Hektoen Enteric agar, though not on EF-18 agar. Using the EF-18 agar as the Salmonella selective medium, a method was established that recorded counts of nalidixic acid resistant Salmonella rapidly and easily in electronic data files, for subsequent retrieval and manipulation.     

Value of the K+ salt of carageenan as an agar substitute in routine bacteriological media.

The K+ salt of carageenan has no distinct advantages as a gelling agent, but it compared favorably with agar in most of the media tested. The difficulty involved in the preparation of blood plates and the results obtained with this medium prohibit its complete acceptance as a substitute for agar in routine solid media. However, it could be a suitable substitute for agar in all other routine bacteriological media.     

Evaluation of Pseudosel Agar as an Aid in the Identification of Pseudomonas aeruginosa

Growth of Pseudomonas aeruginosa and thirty-five other species of gramnegative bacilli was observed on 0.03% cetrimide in heart infusion agar medium and Pseudosel agar (BBL). The 0.03% cetrimide agar was more selective for growth of P. aeruginosa than was Pseudosel agar; however, certain bacteria other than P. aeruginosa also grew on the former medium. Although Pseudosel agar was not a highly selective medium for P. aeruginosa, it was preferable to technicolor agar for detection of the pyocyanin and pyorubin pigments produced by P. aeruginosa.     

Lysine-mannitol-glycerol agar, a medium for the isolation of Salmonella spp., including S. typhi and atypical strains.

An agar medium for the isolation of Salmonella spp. is described. The medium, lysine-mannitol-glycerol agar, has features of both xylose-lysine-deoxycholate agar and mannitol-lysine-crystal violet-brilliant green agar, but glycerol is added for the differentiation of Salmonella and Citrobacter spp. The medium facilitates the detection of strains having atypical fermentation patterns, such as the lactose- or sucrose-positive salmonellae. The medium also detects Salmonella typhi after enrichment.