Showing papers on "Agar plate published in 1984"

Correlation of elastase production by some strains of Aspergillus fumigatus with ability to cause pulmonary invasive aspergillosis in mice.

Abstract: Seventy-five strains of Aspergillus fumigatus were screened for production of elastase in liquid and agar media containing elastin in yeast carbon base buffered with 0.05 M borate, pH 7.6. Of 71 strains which cleared elastin in agar plates, 33 produced elastase in liquid medium, as measured spectrophotometrically with elastin-Congo red. Six strains producing elastase and four nonproducers were tested for ability to cause invasive aspergillosis in immunocompromised mice (six mice per strain). All 36 mice exposed to elastase-producing strains died within 48 to 96 h. Lung tissue from dead mice showed hyphae and necrosis of the alveoli. Lungs of mice exposed to spores of strains not producing elastase showed few germinated spores and no destruction of alveoli. These results indicate that elastase may be significant in the invasion process.

New, simple medium for selective recovery of Klebsiella pneumoniae and Klebsiella oxytoca from human feces.

Abstract: A culture medium was developed which selectively favored the growth of Klebsiella pneumoniae and Klebsiella oxytoca in Escherichia coli-rich fecal cultures, without the use of antibiotics. The discriminative capacity of this medium was based on the presence of only two carbon sources, citrate and inositol, which can be utilized by nearly all K. pneumoniae and K. oxytoca strains but not by E. coli. The medium consisted of Simmons citrate agar (SCA) with 1% inositol (SCAI). Klebsiella strains from fecal samples subcultured on SCAI grew unhampered as yellow, dome-shaped, often mucoid colonies, whereas E. coli appeared as tiny, watery colonies. Apart from some Enterobacter strains, no other types of bacteria were found to mimic the typical appearance of klebsiellae. Recovery experiments from stool samples revealed a limiting ratio of Klebsiella to E. coli of 1:10(6) or more when samples were plated on SCAI versus ratios of 1:10(2) to 1:10(3) on blood agar or Macconkey agar. Compared with an existing Klebsiella culture method, the combination of SCA and MacConkey-inositol-carbenicillin (MIC) agar, Klebsiella yields with SCAI were not lower than those with the combination of MIC and SCA. Furthermore, the efficiency of the SCAI method was twice that of the latter combination. The SCAI plate could be a valuable tool in studies on the epidemiology of K. pneumoniae and K. oxytoca, for example in nosocomial infections, especially those concerning immunocompromised patients.

The Use of Mudlac Transposons as Tools for Vital Staining to Visualize Clonal and Non-clonal Patterns of Organization in Bacterial Growth on Agar Surfaces

Abstract: When a histochemical stain for β-galactosidase activity is applied to growth of Gram-negative bacteria on agar medium, the pigmentation is non-uniform and capable of revealing internal colony organization into different cell types. Use of an Escherichia coli strain with a thermo-sensitive lac repressor indicates that colonies expand by addition of new cells at the periphery and that older cells which have synthesized β-galactosidase early in development remain in the centre. Mixed inocula of different strains show clonal exclusiveness as they proliferate outwards. Mudlac transposons can create genetic fusions that place β-galactosidase expression under a variety of regulatory systems. Stained surface cultures of E. coli and Pseudomonas putida strains carrying Mudlac insertions in plasmids reveal a variety of flower-like staining patterns. These patterns display both clonal (i.e. sectorial) and non-clonal (circular and radial) features which are heritable within a given strain. The non-clonal aspects of the patterns reflect phenotypic differentiation without genetic change. These observations indicate that bacterial growth on agar surfaces is a highly regulated process similar, in many respects, to the development of specific multicellular tissues and organisms.

Glucose-sucrose-potassium tellurite-bacitracin agar, an alternative to mitis salivarius-bacitracin agar for enumeration of Streptococcus mutans.

Abstract: An agar medium for selective recovery and enumeration of Streptococcus mutans was developed as an alternative to mitis salivarius-bacitracin (MSB) agar. Combinations of dyes, antibiotics, and tellurite were added to a nonselective medium which, because of its sucrose content, allowed easy recognition of S. mutans colonies. Candle jar incubation for 2 days, by comparison with anaerobic incubation, reduced background flora but did not diminish S. mutans recoveries from clinical samples. Quantitative comparisons were made of the simultaneous recoveries of a number of authentic S. mutans serotype representatives and fresh clinical isolates, using various glucose-sucrose-potassium tellurite-bacitracin (GSTB) formulations and mitis salivarius, MSB, and blood agars. Mitis salivarius counts were not detectably different from blood counts, but counts on MSB were distinctly lower. A formulation of the new medium containing 5% glucose 5% sucrose, 0.001% potassium tellurite, 0.3 U of bacitracin per ml (hence GSTB), and 2% agar gave recoveries nearly equal to those on mitis salivarius agar and much greater than those on MSB. The medium yielded readily recognized S. mutans colonies and facilitated detection of intracellular polysaccharide formers upon flooding with I2 reagent. Freshly isolated serotype c, E, and f colonies could often be distinguished from serotype d and g colonies, a distinction made reliable by testing for intracellular polysaccharide. A study of 300 salivary samples revealed GSTB to give significantly higher recoveries than MSB. About 72% of all samples were substantially underestimated for S. mutans with MSB, and 6.7% of samples were falsely negative for S. mutans with MSB. Recovery of background flora on GSTB was as low or lower than on MSB, and both types of agar could be stored for at least 9 weeks without notable change of selectivity. Thus, GSTB agar appears to be simple and reliable to use and requires no anaerobic incubation. Caution is voiced about interpretation of data previously reported which evaluated S. mutans on MSB agar.

Influence of culture conditions on expression of the mucoid mode of growth of Pseudomonas aeruginosa.

Abstract: Five strains of Pseudomonas aeruginosa that are routinely distinguished by diagnostic laboratories on the basis of their colony morphology on agar media were grown in different media to assess the effects of culture conditions on mucoid growth, which we define as the copious production of exopolysaccharide. On brain heart infusion agar, only two of these strains (mucoid and gelatinous) grew as slimy mucoid colonies. None of the five strains produced a mucoid pattern of growth in Mueller-Hinton broth, in which all grew as turbid, nonmucoid, homogeneous suspensions of bacterial cells. When Mueller-Hinton broth was supplemented with Mg2+, all of the strains produced some mucoid aggregated growth, but growth in a modified version of the chemically defined medium of Vogel and Bonner, with elevated levels of Mg2+ and gluconate, produced patently mucoid growth in all strains. This mucoid growth in a liquid medium takes the form of large, coherent, slimy aggregates within the medium and of a "collar" of adherent microcolonies at the air-medium interface. Direct observations by light and electron microscopy showed the submerged aggregates and the adherent microcolonies to be composed of bacterial cells enmeshed in a copious exopolysaccharide matrix. When agar was added to the supplemented medium of Vogel and Bonner, only the mucoid and gelatinous strains produced slimy mucoid colonies on its surface. We conclude that both chemical and physical factors affect exopolysaccharide production by these clinical strains of P. aeruginosa and that the colony morphology on a single agar medium is an insufficient criterion for the designation of a given isolate as being mucoid or nonmucoid. Images

Colistin-oxolinic acid-blood agar: a new selective medium for streptococci.

Abstract: The development and evaluation of a new selective medium (colistin-oxolinic acid-blood agar) for streptococci is described. Streptococci of medical and veterinary importance grew well on the medium. Gram-negative organisms, staphylococci. Bacillus spp., and coryneforms are all inhibited. It was concluded that the medium is valuable for the isolation of streptococci in pure culture from mixed flora and has advantages over other media previously described. Increased isolation rates were obtained together with earlier identification of the isolated strains.

Selective enrichment broth culture for detection of Clostridium difficile and associated cytotoxin.

Abstract: A procedure was devised for routine examination of feces for Clostridium difficile with selective enrichment broth culture containing increased levels of carbohydrates and antibiotics to detect cytotoxin and volatile acids in broths inoculated with fecal samples. C. difficile was detected and identified with a rapidity comparable to that of conventional culture on selective cycloserine-cefoxitin fructose agar. Detection rates for C. difficile in inoculated broths (111/401 or 27%) were significantly higher than for culture on cycloserine-cefoxitin fructose agar (47/401 or 11%, P greater than 0.001). All fecal samples containing C. difficile and cytotoxin were correctly identified by the procedure. Isocaproic acid peak heights greater than 2 mm in selective enrichment broths inoculated with fecal samples indicated that C. difficile was present in the fecal sample examined. Of the positive specimens examined, 58% (64/111) produced peak heights greater than 10 mm. Peak heights less than 2 mm were not associated with C. difficile in the fecal sample. The investigated procedure provided a reliable alternative to the routine processing of feces for detecting C. difficile and associated cytotoxin in feces. Inoculated broths with isocaproic acid peak heights greater than 2 mm, after 24 to 48 h of incubation, and in which cytotoxin was detected, were subcultured to blood agar to obtain isolates of the organism as required. Broths which showed isocaproic acid peak heights less than 2 mm, and in which cytotoxin was not detected, were discarded as negative for C. difficile. The procedure was deemed potentially useful for epidemiological surveys of C. difficile.

A selective enrichment broth for the isolation of Clostridium difficile.

Abstract: The cycloserine, cefoxitin, fructose agar medium (CCF agar) devised by George et al' and the modification of it where egg yolk is replaced by horse blood (Oxoid CCF agar), have been widely and successfully used for the isolation of Clostridium diffiwile from faecal specimens.'-4 For epidemiological and environmental work where only small numbers of organisms may be present, it is likely that a more sensitive method of culture is required. Hafiz et al used a broth containing p-cresol in studying vaginal carriage of C diffcile5 whilst Wilson et a16 have reported that the incorporation of sodium taurocholate in CCF agar (in place of egg yolk) enhances the recovery of spore forms of C difficile from solid media. We describe the use of cycloserine cefoxitin fructose broth containing 0 1% sodium taurocholate (CCFT broth) for the isolation of C diffjile from vaginal and faecal specimens. women attending the Department of Genital Medicine (DGM). Groups IVa and IVb were mothers attending the matemity unit. Predelivery high vaginal swabs were taken by the midwife at onset of labour. Post delivery vaginal swabs were taken just prior to discharge. Group V were the babies delivered to mothers in group IV.

Sampling of high concentrations of airborne fungi.

Abstract: The paper discusses a method developed for determining airborne fungi particles in environments highly contaminated with mold fungi. The collection of airborne fungi was performed with two slit samplers. These sampling devices were found to give the highest values out of the three different types tested simultaneously. After spores had been collected with the slit samplers, the collection medium, an agar gel, was removed from the petri dishes and homogenized in a sterile 0.9% sodium chloride solution. The homogenate was diluted stepwise and spread on agar plates prior to the cultivation and the determination of viable counts. When the homogenization procedure was tested on samples collected from three different work environments, no increase or decrease in the number of colony-forming units could be detected. Storage of the homogenate at 2 degrees C over 9 d did not increase the number of viable fungal colonies.

Modified agar medium for detecting environmental salmonellae by the most-probable-number method.

Abstract: Salmonellae in the environment remain a potential source of disease. Low numbers of salmonellae have been detected and enumerated from environmental samples by most-probable-number methods which require careful colony selection from a plated agar medium. A modified xylose lysine brilliant green medium was prepared to control the loss of selectivity caused by heating the brilliant green component. Added agar reduced colony spreading. The medium contained 47 g of xylose lysine agar base per liter; the agar content was adjusted to 2%, autoclaved, cooled to 50 degrees C, and then amended just before pouring to include H2S indicator and 7 ppm (7 ml of 1:1,000 brilliant green per liter) of unheated brilliant green dye. H2S-positive salmonellae were easily detected from sewage sludge compost to the exclusion of most other gram-negative bacteria. As a result, fewer non-salmonellae were picked for further most-probable-number analysis, greatly reducing the work load associated with the most-probable-number method. Direct plating was possible for enumerating salmonellae in laboratory composts containing ca. 10(3) or more salmonellae.

Efficacy of a topical antibiotic irrigant in decreasing or eliminating bacterial contamination in surgical wounds.

Abstract: Using a simple in vitro system, the authors showed that colony counts of Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, and Pseudomonas species can be reduced by 12%-56% with saline irrigation; the reduction of colony numbers, however, especially for S. aureus, was not always statistically significant. However, even if it were statistically significant, the amount of reduction would not be clinically significant. A topical antibiotic irrigant containing bacitracin/neomycin was effective against S. aureus, S. epidermidis-, and E. coli-treated agar plates. Except for a single plate containing Pseudomonas organisms, the growth of Pseudomonas colonies was also prevented by antibiotic irrigation. Tissue samples of muscle, fat, and bone obtained during operation showed antibiotic levels comparable with, and in most cases greater than, those found in the blood agar plates. These data may be clinically significant and suggest that bacitracin/neomycin irrigation can be safely used to reduce the incidence of postoperative infection.

The DNA repair host-mediated assay as a rapid and sensitive in vivo procedure for the determination of genotoxic factors present in various organs of mice. Some preliminary results with mitomycin C.

Abstract: The DNA repair host-mediated assay, in which repairable DNA damage is determined in E. coli cells present in various organs of mice exposed to genotoxic agents, was further developed to broaden the range of organs under study and to simplify the procedure of assessing differential bacterial cell survival. A pair of derivatives of E. coli K-12 strain 343/113 was constructed which differed vastly in DNA repair capacity (uvr+/rec+ vs uvrB/recA), as a means of assessing DNA damaging effects; furthermore, the strains differed in their ability to ferment lactose (Δ Lac vs Lac+), so that the individual survival of both strains could be determined on a single agar medium (containing neutral red as pH indicator), on which the strains had different colony colour morphology (red, Lac+ vs white, Lac− colonies). Finally, the strains were made streptomycin-dependent, to prevent uncontrolled growth of the bacterial cells within the various organs and also to inhibit contamination of the survival agar medium by representatives of the normal intestinal microflora.

Preparing inoculum for susceptibility testing of anaerobes.

Abstract: We studied six methods for preparation of inoculum to use in susceptibility tests of anaerobic bacteria to determine viable counts of 15 different species of anaerobes We counted viable bacteria for each method-organism combination Methods studied included those used for the more routine tests (broth microdilution and broth-disk elution) and for the National Committee for Clinical Laboratory Standards standard reference agar method Inocula prepared from broth suspensions of organisms from 24- and 48-h anaerobe blood agar plates and adjusted to the turbidity of a 05 McFarland standard gave the most consistent counts for all organisms and also the highest numbers Counts from these suspensions were higher than those from overnight growth in thioglycolate or Schaedler broth when all were adjusted against the same turbidity standard Preparing inoculum directly from agar plates may also speed up the sometimes lengthy process of susceptibility testing of anaerobes and thus make results more clinically useful

Conversion of Blastomyces dermatitidis to the yeast form at 37 degrees C and 26 degrees C.

Abstract: A partially defined agar medium, KT, has been developed and compared with brain heart infusion agar for the conversion of Blastomyces dermatitidis to the yeast form. On the KT medium, the mold form converted to a yeast form within 72 h of incubation at 37 degrees C or after 3 weeks at 26 degrees C. A nutritionally dependent dimorphism in B. dermatitidis was observed.

Media for isolation of aeromonas spp. from faeces

Abstract: Summary Five solid media were evaluated for isolation of Aeromonas spp. from faeces: desoxycholate citrate agar (DCA), MacConkey's agar (MAC), xylose-desoxycholate-citrate agar (XDCA), Rogol's medium (ROG), which contained ampicillin 20 mg/L and p-nitrophenyl-glycer-ine 25 mg/L as inhibitors, and blood agar (BA) with ampicillin 10 mg/L. False negative oxidase tests limited the usefulness of DCA and MAC and, although the use of XDCA avoided the problem of fermentation of lactose, some Aeromonas spp. failed to grow on XDCA or produced minute colonies unsuitable for oxidase tests. BA yielded the highest rate of isolation for Aeromonas spp. from 323 faecal samples—15·2% for all Aeromonas spp. and 9·3% for enterotoxigenic (ENT+) strains. This compares with 10·8% for all strains and 6·5% for ENT+ strains isolated on DCA, 7·1% for all strains and 4% for ENT+ strains on MAC and 4% for all strains and 1·5% for ENT+ strains on ROG. Blood agar with ampicillin is recommended for isolation of Aeromonas spp. from faeces.

Value of blood agar for primary plating and clinical implication of simultaneous isolation of Aeromonas hydrophila and Aeromonas caviae from a patient with gastroenteritis.

Abstract: The simultaneous recovery of Aeromonas hydrophila and Aeromonas caviae from the stool of a 49-year-old woman with watery diarrhea was facilitated through the use of a blood agar medium which detected the hemolytic capability of A. hydrophila. In vitro phenotypic tests support the conclusion that only the A. hydrophila isolate was clinically significant.

Clonal Growth of Human Acute Myeloid Leukemia Cells (ML-1 and HL-60) in Serum-free Agar Medium

Abstract: Human acute myeloid leukemia (ML-1 and HL-60) cells grew continuously in the serum-free liquid medium supplemented with human transferrin and bovine insulin. Both ML-1 and HL-60 cells formed clusters and colonies in the serum-free agar medium supplemented with bovine serum albumin, human transferrin, cholesterol, and L-alpha-phosphatidylcholine. Medium conditioned by phytohemagglutinin-stimulated leukocytes prepared in the absence of serum had three types of colony-stimulating factors on normal human bone marrow cells. When fetal calf serum was present, medium conditioned by phytohemagglutinin-stimulated leukocytes stimulated the clonal growth of HL-60 cells at the lower concentration. However, it inhibited that of ML-1 cells. In contrast, under serum-free conditions, medium conditioned by phytohemagglutinin-stimulated leukocytes promoted the clonal growth of both ML-1 and HL-60 cells at the lower concentrations. The study using a Sephadex G-200 column revealed that, in the serum-supplemented cultures, HL-60 cells responded to one of the three colony-stimulating factors and an activity with molecular weight of around 12,000, while ML-1 cells responded only to an activity with molecular weight of around 12,000. In the serum-free cultures, both ML-1 and HL-60 cells were stimulated by activities with molecular weights of 62,000 and 54,000, respectively. These studies demonstrate that the determination of growth factors for cell lines is dependent on culture conditions, particularly on serum component; that there is a heterogeneity of ML-1 and HL-60 cells in response to the growth factors; and that there is potential importance of demonstration of heterogeneity among different cell lines in establishing requirements for different stages of differentiation.

Hyphal interactions between Rhizoctonia solani and some Verticillium species

Abstract: The biological control of soil-borne plant pathogens with mycoparasitic fungi (e.g., Trichoderma and Gliocladium spp.) has received considerable attention (Ayers and Adams, 1981; Lumsden, 1981; Papavizas and Lumsden, 1980). Species of Verticillium have been often reported in the literature as mycoparasites but only a few taxa have been cited as parasites of soil-borne plant pathogens. For most common Verticillium species host range studies have not been conducted and their ability to attack soil-borne plant pathogens has not been determined. Hyphal interactions between selected species of Verticillium and the plant pathogen R. solani were examined in this study. Species and isolates of Verticil? lium were chosen to represent the common soil-borne members of the genus. Although most taxa are known as mycoparasites, isolates were not obtained di? rectly from fungal substrates (Table I). A measure ofthe variability within a single species was obtained by including six isolates of V. lecanii from different sources. Identification of the isolates of V. psalliotae and V. lamellicola was verified by determining growth rates on 2% malt extract agar at 20 to 30 C (Gams and Van Zaayen, 1982). All cultures were maintained on Difco potato dextrose agar slants (PDA) and stored at 4 C. Two isolates of R. solani (R-19 and R-33) were tested in combination with each isolate of Verticillium. Both were highly virulent members of anastomosis group AG-4 and isolated from diseased bedding plants (Stephens et al, 1982). Hyphal interactions between Verticillium and R. solani isolates were observed by growing paired cultures from agar discs (5 mm diam) placed at opposite sides of agar plates (9 cm). All results were based on minimum of two plates prepared on two different occasions for each Verticillium/Rhizoctonia combination on at least two media: 2% water agar and Difco potato dextrose agar. In preliminary tests with some isolates, hyphal interactions were also examined on Difco cornmeal agar and 2% agar in an aqueous extract of composted hardwood bark container medium. The extract was prepared by soaking one liter of container media in one liter of distilled water overnight followed by filtering the mixture through cheesecloth. This medium had a final pH of 7.1 and was used to approximate the nutritional conditions of the environment from which two species were isolated. All plates were incubated at 24 C under continuous light and were examined after 5-14 da by direct examination with a compound microscope using either brightfield or Nomarski interference illumination. A coverslip was placed over the zone of contact for examination at high magnification and for photography. In some cases, a small drop of cotton blue in lactophenol was used to stain the hyphae. Replicate plates were prepared with a layer of cellulose dialysis membrane on the

Some characteristics of aerial and submerged spores of kitasatosporia setalba

Abstract: Kitasatosporia setalba KM-6054 forms two kinds of spores: aerial spores on an agar medium and submerged spores in a liquid medium. In this study, both kinds of spores were resistant to sonication, but sensitive to lysozyme digestion and moist heat. LL-2, 6-Diaminopimelic acid (A2pm), alanine, glutamic acid and glycine were detected as the main constituents in the cell walls of the aerial and submerged spores. The respective molar ratios were similar; 1.0:1.6:0.5:1.0 and 1.0:1.5:0.7:0.8. On the other hand, the vegetative mycelium on agar media and the filamentous mycelium in submerged cultures were sensitive to sonication and differed from the two kinds of spores in both amino acid composition and A2pm type.

Colonial Morphology of Treponemes Observed by Electron Microscopy

Abstract: The colonial morphology of three strains of cultivable, nonpathogenic treponemes including a human oral treponeme was examined by light and electron microscopy. Treponema phagedenis strains Kazan and Reiter produced large white colonies on the surface of solid media composed of sterility test broth, 0.9 to 3.1% agar, rifampin, and 12.5% rabbit or horse serum. A human oral treponeme, strain G7201 , grew as diffused white zones on 0.9 to 3.1% agar plates. Under the cultural conditions employed agar concentrations slightly affected the time of appearance of colonies of the three strains of treponemes. When the colonies of these three strains were viewed by scanning electron microscopy, differences in their colonial morphology were observed. The 11-day-old colonies of human oral strain G7201 were very small, 5 to 15 micron in diameter, and had a slight irregular border. Kazan treponemes developed circular, entire and low convex colonies. Scanning and transmission electron microscopy revealed that the colonies of Reiter treponemes contained spherical forms almost up to 5 micron in diameter, each consisting of an outer membrane and a treponemal main body. They were very similar to the spherical bodies produced by strain G7201 in sucrose-containing broth.

Uterine culture in mares.

Abstract: A guarded, sterile swab is used to obtain samples for uterine culture. With the mare in stocks, the tail bandage and the perineum washed, the culture rod is introduced into the vagina with a gloved hand. After the rod is guided through the cervix, the guard cap is dislodged and the swab is rubbed along the endometrium, after which the rod is extracted. Samples for uterine culture should only be obtained during full estrus. Swabs should be directly plated onto agar within 2 hours of collection. Blood agar is appropriate for initial screening, but use of specialized types of agar expedites identification of microbes. Plates are incubated at 37 C and inspected for growth every 12 hours. The type and number of bacterial colonies should be coupled with the history and clinical signs in deciding on the necessity and type of treatment. Pure, heavy bacterial growth is usually accompanied by clinical signs of infection. Interpretation of the significance of moderate bacterial growth may be aided by cytologic examination of endometrial smears, made by rolling the swab onto a glass slide and staining with Diff - Quik . Large numbers of neutrophils indicate the need for antibiotic therapy. Mixed bacterial growth and variable numbers of neutrophils usually indicate faulty sampling technic. Microaerophilic or anaerobic cultures may aid diagnosis in cases of equivocal aerobic culture results.

Efficient isolation of thermophilic and thermotolerant mucoralean fungi

Abstract: Rhizomucor miehei (Cooney & Emerson) Schipper, R. pusillus (Lindt) Schipper, Rhizopus rhizopodiformis (Cohn) Zopf, and Absidia corymbifera (Cohn) Sacc. & Trotter were isolated from numerous materials immersed in YpSs agar or 2% malt extract agar containing 50 μg/ml Benlate, 100 U/ml penicillin G, and 40 μg/ml streptomycin sulfate; agar plates were incubated at 45 °C. Alternative media and procedures useful for special circumstances are described. These four pathogenic thermophilic or thermotolerant species were readily isolated from spices, herbal teas, sunflower seeds, and other materials.

Cell culture models as ancillary tools in the isolation and characterization of mycoplasmas.

Abstract: Since the late 1960s, there have been an increasing number of reports describing the isolation and identification of fastidious strains of Mycoplasma hyorhinis in cell cultures but not in conventional mycoplasma media, i.e., agar and broth. The application of those techniques normally used in studying viruses, i.e., infection of cell cultures coupled with the subsequent use of immunological and biological test procedures, has provided a reliable alternative procedure for M. hyorhinis detection. Major isolation surveys have revealed that as many as 60 to 80% of M. hyorhinis isolates from contaminated tissue cultures failed to grow in agar medium. Efforts to elucidate the mechanisms involved in failure of the cell culture-derived M. hyorhinis strain to grow in standard cell-free mycoplasma media are ongoing. Initial data indicated that extracts prepared from tissue cultures (BHK 21) and incorporated in Macpherson's broth and agar medium would permit growth of fastidious strains. Moreover, it appears that these strains are particularly sensitive to inhibition by yeast products often found in mycoplasma media. While M. hyorhinis appears to be unique with respect to its sensitivity to medium components, these fastidious strains are isolated with such frequency that the routine use of an indicator cell system is strongly recommended.

Enzyme immunoassay for the detection of group A streptococcal antigen.

Abstract: A competitive inhibition enzyme immunoassay for the detection of Streptococcus pyogenes directly from throat specimens or from solid bacteriological medium is described Group A-specific polysaccharide adsorbed onto treated polystyrene beads, in conjunction with rabbit antibody to S pyogenes, was used to determine the presence of the polysaccharide antigen Inhibition values in excess of 65% were observed with 10(4) or more CFU of S pyogenes per test An inhibition of 25% was demonstrated with as few as 10(3) CFU per test Heterologous microorganisms tested at 10(6) CFU per test reacted at levels of inhibition less than 25% Two types of bacterial transport medium and swabs of different fiber compositions did not alter the assay performance Accurate identification of S pyogenes was achieved by testing single colonies picked directly from blood agar plates which had been incubated for 18 to 24 h In addition, the assay was performed on throat specimens from children and adults having pharyngitis A single-swab, blind study was conducted in which enzyme immunoassay reactivity was compared with results of blood agar culture and bacitracin sensitivity When there were discordant results, serological identification was used as the confirmatory test At an optimal cutoff value of 40% inhibition, sensitivity and specificity by enzyme immunoassay were 970% and 979%, respectively, as compared with confirmed culture results The assay has an incubation time of 3 h and is a sensitive and specific method for the detection of S pyogenes antigen

Typing of Enterobacter spp. by bacteriocin susceptibility and its use in epidemiological analysis.

Abstract: Most clinical isolates of Enterobacter cloacae are bacteriocinogenic and susceptible to bacteriocins. Both rapidly diffusing, nonsedimentable, protease-susceptible and slowly diffusing, sedimentable, protease-resistant bacteriocins are produced. A practicable system was devised for epidemiological typing of E. cloacae isolates by their patterns of susceptibility to bacteriocins. A set of eight bacteriocin-producing strains was grown on tryptic soy agar plates for 16 h. After removal of the producer lawn, the isolates to be typed were inoculated on the agar media by a multipoint inoculator. After a second 16-h period of incubation, the strains were classified into bacteriocin types according to the patterns of growth inhibition. Typability of 134 clinical isolates was 96.3%. Only 11 (8.2%) of the isolates fell into the largest group. Repeat testing of isolates from the same patients within 2 months gave identical bacteriocin types. Other species of Enterobacter (E. agglomerans and E. aerogenes) are also typable by this method.

Changes in Fatty Acids During Morphogenesis in Sclerotium rolfsii

Abstract: Changes in the lipids of Sclerotium rolfsii were followed during its growth in agar and liquid synthetic media. The fungus was grown either aerobically on a cellophane membrane placed on a synthetic agar medium, or under submerged conditions in the same liquid medium, using a rotary shaker, with sclerotial formation induced by pouring the culture into Petri dishes. In the aerobically-grown fungus, lipid content decreased from 119-7 μg (mg dry wt)-1 in the mycelium to a low content of 7.4 μg (mg dry wt)-1 in mature sclerotia. The non-polar lipids were mainly utilized at the sclerotial initiation stage, while the polar lipids were utilized during sclerotial maturation. Total lipid content in submerged mycelium was 10-fold less than in aerial mycelium; however initial and mature sclerotia produced from either the submerged or aerial mycelium were similar in their lipid content and in the composition and distribution of fatty acids. Increasing glucose concentrations in the growth medium increased the fungal biomass but did not change lipid content or composition in the two growth systems. A supplement of 10-2 M-L-threonine to the growth media significantly increased the number of sclerotia produced in submerged culture. L-Threonine also prolonged the lag phase of a submerged culture and caused a delay in lipid accumulation. Moreover, L-threonine increased the lipid level but did not change the lipid or fatty acid distribution. This may help in elucidating the role of lipids in the mycelium during sclerotial formation.