Form of Selenium in Selenite Enrichment Media for Isolation of Salmonellae

Selenite-F and selenite-cystine media, commercially available for the routine isolation of salmonellae, were treated by anion exchange chromatography to separate the selenium from other components of the media. A chemical assay, based on an ascorbic acid reduction, showed that the selenium was all in the form of selenite.     

Evaluation of Culture Media for the Isolation of Salmonellae from Feces

Serial passage of the 64-2389 strain of type 3 parainfluenza virus in cercopithecus monkey kidney tissue cultures at low temperatures resulted in the selection of a variant which had a higher efficiency of plaque formation at 25 C than the parent line grown at 37 C. The cold variant, unlike the parent strain, plaqued readily at 25 C, and at 37 C it produced significantly larger plaques. Virus titers of the cold variant in hamster lungs were significantly lower and this was probably caused by the stimulation of interferon by the cold variant during the early phase of the infection. The cold variant, like the virus grown at 37 C, also induced the synthesis of interferon late in the infection. Hamsters responded to the intranasal inoculation of each virus line by the development of hemagglutinating-inhibiting antibodies in the sera.     

Evaluation of Media for Selective Isolation of Yeasts from Oral, Rectal, and Burn Wound Specimens

Six media were evaluated to determine their ability to isolate yeasts and inhibit bacteria. The media included the following: Snyder, Snyder tellurite, Sabouraud tellurite, Littman-gentamicin, molybdate, and Mycosel (BBL). Doses of mixed intestinal gram-negative bacilli and enterococci were most effectively inhibited by Snyder tellurite agar. Klebsiella pneumoniae was the most common bacterial contaminant of the other media. All six media were comparable in isolating yeasts while preventing the growth of the oral bacterial flora. The selection of a basal fungal growth medium for tellurite incorporation to inhibit bacteria but permit growth of yeasts was affected by pH. The bacteriostatic effect of tellurite was decreased with increasing pH of media while fungistatic action was increased. The arbitrary selection of Snyder and Littman agars to isolate yeast from burn wound cultures demonstrated the need to include a selective medium for these specimens. Blood, phenylethyl alcohol blood agar, and Columbia CN blood agar were all inadequate for isolating yeasts from burns. Growth of a variety of filamentous saprophytic and pathogenic dimorphic fungi grew adequately on four of five selective media tested.     

Isolation of Salmonellae by Selective Motility Systems

Salmonellae were isolated from naturally infected swine and human feces by means of selective migration through semisolid enrichment (SM) media in modified “U” tubes. Comparative studies showed that recovery of Salmonella by SM techniques was equal or superior to that of standard procedures employed in two routine diagnostic laboratories. Primary Salmonella isolations from SM enrichment were relatively free from normal fecal bacteria and often pure. The SM method required only 15% of the media and correspondingly less time and skill than the usual methods for isolating Salmonella.     

Isolation of Salmonellae from Foods Samples: IV. Comparison of Methods of Enrichment

A comparison of various methods of enhancing frequency of Salmonella isolations revealed that inoculation of a second enrichment broth, with culture from the first, was no improvement over the single direct enrichment method. It was inferior to centrifugation.

Selenite was observed to produce more positive isolations at 48 hr than at 24. No change occurred in tetrathionate. Reconstitution of dried albumen with water produced a significant increase in isolations over direct inoculation of enrichment broth in the case of tetrathionate but not selenite broth.

Pre-enrichment in lactose broth before inoculation of enrichment media was vastly superior to reconstitution in water for both enrichment broths. A comparison of results obtained using dulcitol, mannitol, lactose and carbohydrate-free purple broths in pre-enrichment indicated that the carbohydrate added was immaterial.

     

Comparison of Media for Direct Isolation and Transport of Shigellae from Fecal Specimens

Xylose-lysine-deoxycholate (XLD) agar, SS agar, and MacConkey agar for isolating shigellae from fecal specimens were compared. XLD agar was superior to both SS agar and MacConkey agar for isolating Shigella sonnei, and both XLD and SS agar were superior to MacConkey agar for isolating S. flexneri. Direct plating of the fecal specimens in the field resulted in a greater yield of shigellae as compared to transporting specimens to the laboratory either in holding media or enrichment broth. Buffered glycerol saline was superior to other transport media evaluated, yielding 83% of shigella isolates when plated within 48 hr as compared to direct plating. The combination of XLD agar and SS agar is recommended for direct isolation of shigellae, and, whenever possible, these solid media should be taken to the bedside and inoculated directly.     

Enrichment in Muller-Kauffmann's Broth and Rappaport's Broth from Buffered Peptone Water in the Isolation of Salmonellae from Minced Meat

Four hundred and ninetysix samples of minced meat were examined for the presence of salmonellae. The methods of isolation used in the study were: (a) the standardized direct technique of enrichment in Muller-Kauffmann's broth incubated at 43· for 24 and 48 h, described by Edel & Kampelmacher (1969), (b) secondary enrichment in Rappaport's broth from the former medium after 24 h incubation, and (c) pre-enrichments in buffered peptone water incubated at 37·, followed by enrichments in Muller-Kauffmann's and in Rappaport's broth. These last procedures resulted in the higher rates of isolation of salmonellae with a slight, but not statistically significant, superiority of the enrichment in Rappaport's medium.     

Comparison of Two Plating Media for the Isolation of Erysipelothrix rhusiopathiae from Enrichment Broth Culture

Sodium azide-crystal violet-agar and a modified blood-azide (MBA)-agar were used to isolate Erysipelothrix rhusiopathiae from swine tissues. There was no significant difference in the number of isolations. However, 78% of the isolants from modified blood-azide medium required only 24 hr of incubation, whereas all of the isolants obtained on sodium azide-crystal violet medium required 48 hr.     

Evaluation of Three Media for Selective Isolation of Gram-Positive Bacteria from Burn Wounds

Three media, phenylethyl alcohol blood agar, esculin-mannitol agar, and Columbia CN blood agar, were studied for the selective isolation of gram-positive bacteria from swab cultures of burn wounds.     

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.     

A Comparison of Various Selective Media, including a New Selective Medium for the Isolation of Brucellae from Milk

S ummary : A comparative trial of a new medium for brucella was made on samples of milk from individual cows and herd samples of milk. The new medium gave a higher isolation rate than any of the other 3 selective media tested. There was no evidence that the new medium inhibited any of the biotypes of Brucella abortus , especially biotype 2; it was the only medium suitable for isolating from samples of herd milk strains of biotype 2 which are sensitive to antibiotics and aniline dyes.     

Comparison of Four Agar Plating Media with and Without Added Novobiocin for Isolation of Salmonellae from Beef and Deboned Poultry Meat

Four plating media, Hektoen enteric (HE), xylose-lysine deoxycholate (XLD), tryptic soy-xylose-lysine (TSXL), and tryptic soy-brillant green (TSBG) agars with and without 10 mg of added novobiocin per ml, were evaluated for recovery of Salmonella from roast beef and deboned turkey. Colonies producing a reaction typical of H₂S-positive salmonellae (alkaline with black centers) were picked. On the media without novobiocin, from 109 determinations on 75 samples, number of salmonellae found and false-positives were, respectively: HE—13, 58; XLD—17, 18; TSXL—23, 0; TSBG—22, 7. When novobiocin was present the corresponding results were: HE—17, 24; XLD—21, 2; TSXL—23, 3; TSBG—20, 7. A total of 25 determinations were positive on one or more agars. False-positives on HE and XLD without novobiocin were predominantly Proteus, which were almost totally eliminated by addition of 10 mg of novobiocin per liter. If alkaline H₂S-negative colonies had been considered, many more false-positives would have been found on HE and XLD but not on TSBG or TSXL. Addition of novobiocin markedly improved isolations of salmonellae from XLD and HE and reduced the number of false-positives. Addition of novobiocin did not improve performance of TSXL and slightly impaired differentiation of salmonellae from Citrobacter on TSBG. XLD with novobiocin and TSXL are highly specific for H₂S-positive salmonellae, and the appearance of Salmonella-like colonies on these media can be considered a presumptive test for H₂S-positive salmonellae.     

Comparison of Methods for Isolation of Anaerobic Bacteria from Clinical Specimens

Five different anaerobic culture methods and several different media were compared for their ability to recover anaerobes from clinical specimens. Specimens were obtained from patients with documented infections, avoiding contamination with normal flora, and immediately placed in an anaerobic transporter. Each specimen was cultured by all methods and on all the various media. The comparative data indicate that anaerobic jars (GasPak and evacuation-replacement types) are just as effective in the recovery of clinically significant anaerobes as the more complex roll-tube and chamber methods employing prereduced media. Liquid media were disappointing as a "back-up" system but chopped-meat glucose was superior to two thioglycolate formulations. Growth of all anaerobes was poorer on selective media, but these media were very helpful in the workup of specimens containing mixed growth of anaerobic and facultative organisms. A variety of different anaerobes was isolated, but no very fastidious or extremely oxygen-sensitive organisms were recovered. This suggests that such organisms may not play a significant role in causing clinical infections.     

Selective Enrichment with a Resuscitation Step for Isolation of Freeze-Injured Escherichia coli O157:H7 from Foods

We studied injury of Escherichia coli O157:H7 cells in 11 food items during freeze storage and methods of isolating freeze-injured E. coli O157:H7 cells from foods. Food samples inoculated with E. coli O157:H7 were stored for 16 weeks at −20°C in a freezer. Noninjured and injured cells were counted by using tryptic soy agar and sorbitol MacConkey agar supplemented with cefixime and potassium tellurite. Large populations of E. coli O157:H7 cells were injured in salted cabbage, grated radish, seaweed, and tomato samples. In an experiment to detect E. coli O157:H7 in food samples artificially contaminated with freeze-injured E. coli O157:H7 cells, the organism was recovered most efficiently after the samples were incubated in modified E. coli broth without bile salts at 25°C for 2 h and then selectively enriched at 42°C for 18 h by adding bile salts and novobiocin. Our enrichment method was further evaluated by isolating E. coli O157:H7 from frozen foods inoculated with the organism prior to freezing. Two hours of resuscitation at 25°C in nonselective broth improved recovery of E. coli O157:H7 from frozen grated radishes and strawberries, demonstrating that the resuscitation step is very effective for isolating E. coli O157:H7 from frozen foods contaminated with injured E. coli O157:H7 cells.     

Selective Isolation of Leptospiras from Contaminated Material by Incorporation of Neomycin to Culture Media

Incorporation of neomycin to the culture medium was found to be effective in inhibiting Escherichia coli contaminants without interfering with the growth of serotype L. autumnalis. The growth of 12 other Leptospira serotypes was unaffected by the addition of 300 mug of neomycin per ml to Ellinghausen medium or 5 mug/ml to Fletcher medium. Neomycin-containing medium was found to be of value in the isolation of leptospiras from cultures of blood from infected laboratory animals. A higher percentage of isolates was obtained in swine kidneys from an abattoir in medium containing neomycin than resulted from the same medium without antibiotic or with 5-fluorouracil. Contaminated leptospiral cultures growing in media with 5-fluorouracil were purified by subculturing into neomycin-containing media.