The detection of contamination in Trichophyton rubrum and trichophyton mentagrophytes

This report concerns the usefulness of two media, brain heart infusion agar (BHIA) and BCP milk dextrose agar in the detection of contamination inT. rubrum andT. mentagrophytes and provides cultural information in the identification of these species.Department of Microbiology, Public Health Laboratory, Ontario Department of Health, Box 9000, Postal Terminal A, Toronto 1, Ontario, Canada.Head, Medical Mycology Section.Mycologist, Medical Mycology Section.     

Morphological changes of trichophyton mentagrophytes under the influence of increased carbon dioxide tension

Summary Growth pigment production as well as macroconidia and arthrospore formation ofTrichophyton mentagrophytes are stimulated under the influence of increased carbon dioxide tension. Macroconidia with spirals and pistol forms of macroconidia are observed.The granular form ofT. mentagrophytes displays an asteroid growth (Dextrose agar of Sabouraud) with an ingrowth and elongation of the ray branches in the agar and a decrease of the air-mycelium. The culture completely resembles to those ofT. mentagrophytes varasteroides. The macroconidia appear elongated and they acquire an asteroid arrangement. The less granular strains ofT. mentagrophytes take the appearance ofT. interdigitale type.An analogy is made between these morphogenetic changes under the effect of increased CO₂ tension in vitro and the influence of the animal organism in the evolution of the growth forms of dermatophytes in vivo in the process of their parasitic adaptation.     

Trichophyton fischeri sp. nov.: a saprophyte resembling Trichophyton rubrum

A new species Trichophyton fischeri was isolated as a contaminant on blood agar plates. This fungus is believed to be a saprophyte. It may be confused with T. rubrum. On peptone dextrose agar plate, the growth is white and velvety to cottony. It occasionally forms furrows. The underside of the mature colony is brownish red. Clavate microaleuriospores are common. Trichophyton-type macroaleuriospores are produced occasionally on blood agar and potato dextrose agar. Erythritol does not stimulate T. fischeri to produce a red color on casamino erythritol albumen agar. Closterospore-like projections may be produced on the main filaments on peptone dextrose and potato dextrose agar.     

Ultrastructure of Nocardia Cell Growth and Development on Defined and Complex Agar Media

The cell growth of Nocardia strain 721-A on Brain Heart Infusion Agar (BHIA), nutrient agar, and chemically defined agar media was studied by light and electron microscopy. Light microscopy revealed a change in cell morphology induced by growth on BHIA. Electron microscopy demonstrated a concurrent change in intracellular complexity. On BHIA, the cells became bulbous and developed irregularly branched filaments which fragmented by multiple and random septation. These fragments appeared to undergo a secondary stage of development similar to that described for Arthrobacter. Cells grown on defined or nutrient agar did not become bulbous and lacked the unusual complexity found in cells grown on BHIA. Intracytoplasmic membranes were altered by the nutritional state of the cell and changed during cell development.     

Quantitative measurement of cell-bubble attachment in bubble columns using foam flotation techniques

A simple and convenient system for quantitatively measuring the number of adsorbed animal cells per unit of bubble surface area (, unit: cells/cm2) was developed. The system was successfully applied to recombinant Chinese hamster ovary (r-CHO) suspension cultures to investigate the dynamic cell-bubble attachment in a bubble column. In serum-free medium, values increased with bubble rising height (H) and cell concentration (C) and then became constant (about 1750 cells/cm2) when H and C were sufficiently high. In medium containing protective additives, the trends of values with H were similar to that in serum-free medium. Compared with serum-free medium, polyvinyl-pyrrolidone (PVP) increased the values to 1941 cell/cm2 whereas other tested additives decreased the values of in some different degree.     

The effect of sodium chloride on the growth and morphology of dermatophytes and some other keratolytic fungi.

A study of the influence of various concentrations of NaC1 on 21 species of dermatophytes and other keratolytic fungi was made. Based on the sensitivity of the species to various concentrations of NaC1, it was possible to divide them into five groups. Microsporum ferrugineum and Trichophyton concentricum and T. tonsurans were the most sensitive to NaC1 and were inhibited by 5%. The greatest number of species were inhibited by 12% NaC1. The fungi most tolerant to NaC1 were M. cookei and M. nanum, and T. mentagrophytes, T. schoenleinii, and T. terrestre. These species were inhibited by 15%. NaC1 prevented any variant change in Epidermophyton floccosum, T. mentagrophytes, and M. gypseum, but promoted a change in phenotype in M. audouinii and M. cookei, and T. gallinae. It is suggested that the word "pleomorphism" be replaced by the term sterile albinism. This term refers to that kind of cultural change when there is no evidence of sporulation and the white fluffy mycelium consists of fine sterile hyphae. Sterile albino strains of E. floccosum were induced to form a macroconidia on Sabouraud cycloheximide chloramphenicol gentamicin agar (SCCGA) containing 3-5% NaC1. Also, M. audouinii formed microconidia and macroconidia in velvety growth cultured on SCCGA containing NaC1.     

Correlation ofLegionella pneumophila virulence with the presence of a plasmid

Legionella pneumophila is the causative agent of Legionellosis in man and considered an opportunistic intracellular Gram-negative bacterium that preferentially infects macrophages. The presence of a plasmid in these organisms was determined in cultures of the bacteria grown in vitro. A correlation was observed between the growth of virulent strains of theLegionella in murine macrophages and growth on standard buffered charcoal yeast extract agar supplemented with 0.1% -ketoglutarate (BCYE) agar medium rich in cysteine and widely used for growth of the bacteria in vitro. In contrast, the avirulent isolates of these strains grew well on supplemented Mueller-Hinton (SMH) agar utilized for differentiating virulent from avirulentLegionella. However, one virulent strain ofLegionella (the Iowa strain) was found to grow moderately well on the SMH agar. In addition, test strains ofLegionella that infect in vitro human monocytes were found to grow moderately well on the BCYE- agar, but did not grow on the SMH agar. Examination of these strains for plasmid DNA expression showed that extra chromosomal DNA-containing molecules were present in theL. pneumophila strains characterized as virulent for in vitro growth in macrophages. However, the avirulent strains that replicated in the human monocytes readily but only poorly in the permissive murine macrophages did not show evidence of similar plasmid DNA expression.     

On the morphology of trichophyton quinckeanum

Summary The cultivation of four old non-sporulating strains ofTrichophyton quinckeanum on solid nutrient medium with 15% CO₂ (Sabouraud dextrose agar) gave an increased production of macroconidia. The macroconidia are multilocular with smooth walls. It is believed that the polymorphism of the macroconidia — some with pointed tip and other with round terminal cell — is characteristic of the species and has a taxonomic significance. Macroconidia with a pointed end cell are not of theMicrosporon type, since they have smooth walls and are not pointed on both ends Filiform appendages from the terminal cell occur regularly. Small and large macroconidia were seen. The microconidia are pyriform or elongated. No rounded microconidia withBotrytis type of arrangement were observed. The so-calledT. niveum is probably identical withT. quinckeannum.Together with the characteristics of the species as a parasite it is to be concluded thatT. quinckeanum is a peculiar dermatophyte species, corresponding toQuincke's -Pilz, but cannot be assimilated by the group ofT. mentagrophytes (Ctenomyces s. microides).     

Natural fermentation of soybean (glycine max) with added salt for ‘soy-daddawa’ production

Microbial growth, pH and titratable acidity decreased with increasing NaCl concentration during the fermentation of soybeans. NaCl at 1% (w/w) in the fermenting mash improved the organoleptic quality of soy-daddawa.The author is with the Department of Microbiology, Obafemi Awolowo University, lle-lfe, Nigeria.     

Restriction site and length polymorphism of the rDNA unit in the cultivated basidiomycetePleurotus cornucopiae

In the ribosomal DNA unit ofPleurotus cornucopiae, the rDNA coding regions are in the order 5, 5S-18S-5.8S-25S, 3, with the 5 location of the 5S gene differing from its 3 location found in other basidiomycetes. The most discriminating probe used to study the rDNA polymorphism consisted of a fragment that included the 5S, 18S and part of the 5.8S and 25S genes flanking three intergenic sequences. A high degree of rDNA polymorphism was observed in the sevenP. cornucopiae dikaryons studied. For the first time within a basidiomycete species, the restrictions maps distinguished two types of rDNA units (I and II). In each rDNA type, length variations in the external intergenic sequence IGS 1 located between the 25S and 5S genes allowed characterization of two different rDNA units in type I and four rDNA units in type II. This suggested that theP. cornucopiae rDNA units were derived from two kinds of ancestors (type I and II) by insertion or deletion events (100–700 bp) in the IGS 1. In four dikaryotic strains, two rDNA units of the same type (I or II) differing only by the IGS 1 length, were found in a similar number of copies, and presented a meiotic segregation in homokaryotic progeny. In one progeny, some homokaryotic strains possessed two different rDNA units: one with a high copy number and another with a lower one, showing that two different rDNA units could coexist in a single nucleus.     

Chydorus ‘mutilus’ Kreis, 1921 — a postephippial form of Chydorus sphaericus (O. F. Müller, 1785)

Since 1991 faunal collections have been made from the mountain lakes of NW Slovenia. These lakes are at altitudes of between 1250 and 2150 m a. s.l., and have rich biotas, both in terms of species-richness, and faunal abundance. Amongst the animals collected were hump-backed specimens of genus Chydorus. These are identical with Chydorus mutilus, a species described from Swiss mountain lakes by Kreis (1921). The abundance of specimen in the collections, coupled with the availability of data from four successive years of sampling, allowed the detailed analysis of these populations. This also includes an examination of chitinous remains preserved in subrecent sediments.The results show that C. mutilus Kreis, 1921 actually represents a postephippial form of C. sphaericus (O. F. Müller, 1785). From the data available, it appears that the hump-backed form only occurs under certain environmental conditions. Here I discuss environmental factors having the potential to trigger the formation of hump-backed Chydorus. These findings may prove significant in palaelimnological studies, and in the reconstruction of paleotemperatures.In addition, hump-backed animals, apparently identical to European C. mutilus, have also been found in a sample taken from Lake Titicaca (Peru) in 1954. This supports the hypothesis that the hump-backed morph is an environmentally-cued ecophenotype, and not an independent taxon.     

Presence of plastid and absence of mitochondrial DNA in male reproductive cells as evidence for cytoplasmic inheritance in Turnera ulmifolia and Zantedeschia aethiopica

Summary. Epifluorescence microscopy of mature pollen grains of Turnera ulmifolia and Zantedeschia aethiopica stained with 4,6-diamidino-2-phenylindole demonstrated the presence of fluorescent cytoplasmic DNA aggregates in the male reproductive cells of both species. Double staining of the cells with 4,6-diamidino-2-phenylindole and 3,3-dihexyloxacarbocyanine iodide in Technovit resin sections showed that the mitochondria of these cells did not correspond to the fluorescent cytoplasmic DNA aggregates. Electron microscopy studies revealed both plastids and mitochondria in the cells of these species. In addition, immunoelectron microscopy using an anti-DNA monoclonal antibody showed clear labeling of plastids but not mitochondria. These data provide cytological evidence for biparental plastid inheritance and maternal mitochondrial inheritance in these species.Correspondence and reprints: College of Life Sciences, Peking University, Beijing 100871, Peoples Republic of China.     

Persistent effects of changes in phytochrome status on internode growth in light-grown mustard: Occurrence,kinetics and locus of perception

Extension growth of the first internode in fully de-etiolated mustard (Sinapis alba L.) seedlings (11–12.5 d old) is under the control of both the current phytochrome photoequilibrium (Pfr/P, ratio of the far-red-absorbing form of phytochrome to total phytochrome) and that established by short (<12 h) pretreatments. Plants were pretreated with either light pulses providing different calculated Pfr/P followed by dark incubations of different durations (a), or with a 12-h period of white light establishing different Pfr/P (b). After the pretreatments, the plants received either light pulses providing different Pfr/P, followed by dark incubations (c), or continuous white light with or without addtional far-red light (d). Thus, four experimental approaches were followed: (a)(c); (a)(d); (b)(c) and (b)(d). Extension growth during the second period (c or d) was not only affected by the current phytochrome status, but also by that established during the pretreatment period (a or b). The results show the existence of a long-term promotion of stem growth which persists after the end of the low Pfr/P pretreatment. This effect is different from the previously reported rapid effect of far-red light added to background white light as follows: (i) the duration of low Pfr/P required to effect a full response is longer (2.5 h); (ii) the duration of the promotion after returning to high Pfr/P is longer (approx. 24 h) and (iii) the locus of perception is mainly in the leaves, rather than the growing internode.Abbreviations FR far-red light - PAR photosynthetically active radiation - Pfr/P ratio between the FR-absorbing form and total phytochrome - R red light - WL white light     

Antibiotic tetaine — a selective inhibitor of chitin and mannoprotein biosynthesis in Candida albicans

The antibiotic tetaine inhibits in Candida albicans the biosynthesis of two important cell wall constituents, chitin and mannoprotein. This effect is a consequence of inactivation of the enzyme glucosamine-6-phosphate synthetase. Due to the lack of glucosamine-6-phosphate the effective secretion of mannoprotein enzymes, acid phosphatase and invertase, by Candida albicans spheroplasts is inhibited. In the presence of tetaine, probably a modified mannoprotein, lacking a branched polymannan, is synthesized. The antibiotic action decreases the viability of Candida albicans cells, especially that of mycelial forms of this fungus.Abbreviations GlcNAc N-acetyl-d-glucosamine - GlcN-6-P d-glucosamine-6-phosphate - ManNAc N-acetyl-d-mannosamine - -MM -methylmannoside - EGTA 1,2 di/2-aminoethoxy/ethane - N,N,N,N tetra-acetic acid     

Changes in DNA content and chromosomal size during cell culture and plant regeneration of Scilla siberica: selective chromatin diminution in response to environmental conditions

The chromosomal complement and DNA content of cells of the monocotyledonous plant Scilla siberica were studied at various stages of growth, such as callus outgrowths from bulb tissue (bulb callus), callus grown from single protoplasts prepared from bulb callus (protoplast callus), regenerated plants kept for 2–4 years on agar medium and under constant climatic conditions, and regenerated plants grown first for 2 years on agar and then for 1 year in the garden. During callus culture, several different forms of chromatin loss were observed: (1) chromosome elimination early during cell culture, resulting in cells which were mostly diploid but still had large chromosomes similar to those of the original triploid plants (type 1 cells); from these cells no plants could be regenerated. (2) Dramatic reduction in heterochromatin containing the satellite DNA and, apparently subsequently, also of many other chromatin moieties, resulting in the formation of small chromosomes; frequent polyploidization in these cells, resulting in a variable number of chromosomes per cell (preferentially 30–40, in ca. 70% of the cells; type 2 cells). (3) Appearance of a large number of very small (< 1 m) Feulgenpositive chromatin particles (minute chromosomes; often arranged in metaphase-like arrays, suggesting that they were effectively distributed during mitosis). Such cells could survive and divide under cell culture conditions but did not regenerate plants (type 3 cells). In bulb calli, all three cell types were found whereas in protoplast calli only cells of type 2 and 3 were seen. Type 2 and 3 cells had lost, despite their frequent polyploidization, about 80% of their initial nuclear DNA content and an even higher proportion (>95%) of the satellite sequences. Southern DNA blot analysis revealed that sequences hybridizing with certain protein-coding genes such as that for chalcone synthase were also drastically reduced in copy number whereas the proportion of rDNA was even somewhat increased. In plants regenerated from type 2 cells of protoplast calli, which were aneuploid at near-pentaploidy to hexaploidy, further conspicuous changes in chromosomal and DNA content were not observed, as long as they were kept on agar medium and under constant climatic conditions. However, when such plants were grown in the garden for at least 1 year, the satellite DNA as well as the sequences hybridizing with the chalcone synthase gene were disproportionately increased to 30%–40% of their normal proportion, whereas the total DNA had increased by only approximately 15%. The chromosome numbers remained near-pentaploid to hexaploid as in the cell cultures from which these plants had been regenerated. These phenomena of selective loss and regain of chromatin in response to environmental conditions (cell culture, regenerated plants on agar, regenerated plants grown outdoors) are discussed in relation to other forms of chromatin loss, including developmentally controlled chromatin diminution in certain animals.     

Two approaches to obtaining low,extracellular deoxyribonuclease activity in cultures of heterocyst-forming cyanobacteria

Six out of 158 axenic strains of heterocyst-forming cyanobacteria consistently failed to produce circles of clearing in agar medium containing DNA-methyl green. When tested with [³H]DNA and coliphage DNA, supernatant fluids from cultures of two of these strains [University of Texas Culture Collection (UTEX) strain 2014 and 19-6C-C] showed no detectable deoxyribonuclease activity, and such fluids from another two of the six, and four others, showed low but detectable deoxyribonuclease activity. Covalently closed circular (plasmid) DNA was not detectably degraded by supernatant fluids from UTEX 2014 and 19-6C-C and from four of the other strains. When DNA was incubated with whole cells of certain strains, a sereis of fragments of discrete size was produced, perhaps by cell-bound, periplasmic, restriction endonucleases. Inclusion of one-tenth strength saline sodium citrate (SSC) in an eight-fold dilution of the medium of Allen and Arnon had little effect on growth of Anabaena variabilis American Type Culture Collection (ATCC) strain 29413 yet prevented all but slight degradation of plasmid pBR322 or of DNA.     

Refining the genetic map for the region flanking the X-linked hypophosphataemic rickets locus (Xp22.1–22.2)

An analysis of five of the most common cystic fibrosis (CF) mutations worldwide (F-508, R-553X, G-551D, N-1303K and G-542X) was performed in 36 Chilean patients. Polymerase chain reaction (PCR) amplification of the DNA followed by allele specific restriction enzyme analysis was used for detection. The overall frequencies of the mutations in the chromosomes analyzed were 29.2% for F-508 and 4.2% for R-553X (n=72). The G-542X, G-551D and N-1303 K mutations were absent in the Chilean sample. Our data suggest however that F-508 is not the most common CF mutation in Chilean patients. F-508 and R-553X account for only 33.4% of the alleles; 66.6% of them do not respond to the probes used and still remain uncharacterized.     

Production of chlamydospores by Candida albicans cultivated on dilute oxgall agar

Summary A simple easily prepared clear medium consisting of 2% oxgall in 1.8% agar is described. This has proved to be excellent for the rapid identification ofCandida albicans in swabs and cultures, provided that the inoculated area is covered by a coverslip and the incubation temperature period is maintained at 28°C for 24 to 48 hours. No special technical skill is required to prepare this medium. The results presented show thatC. albicans will always produce chlamydospores on this medium if the recommended procedure is followed.Head, Medical Mycology Section.Mycologist, Medical Mycology Section.     

Synthesis and release of proteases byBacteroides fragilis

Protease production byBacteroides fragilis ATCC 25285 was determined in batch and continuous cultures. During exponential growth in batch culture, the majority of proteolysis was cell associated. However, as the bacteria reached stationary phase, most of the intracellular proteases were released into the culture medium. Measurements of alkaline phosphatase and -galactosidase, which are respectively periplasmic and cytoplasmic marker enzymes inB. fragilis, showed that secretion of proteases in the stationary phase was a discrete event and was not associated with a general release of cytoplasmic contents. When the bacterium was grown in continuous culture, cell-associated protease activity increased concomitantly with dilution rate (D=0.03–0.23/h). The ratio of intracellular to whole cell protease activity also increased with growth rate (11 at D=0.03/h; 11.7 at D=0.23/h). Extracellular protease activity was detected only in trace amounts in continuous cultures at the lowest dilution rate. Determinations of the distribution of extracellular protease activity in batch culture after 48 h incubation showed that the majority of proteolysis (ca. 90%) was soluble. Nevertheless, a proportion was associated with particulate fractions, which had high specific activities.     

Differential characteristics of catalase-positive campylobacters correlated with DNA homology groups

Eighty-four strains of catalase-positive campylobacters could be placed into seven distinct DNA homology groups (species), corresponding to Campylobacter fetus, "C. hyointestinalis," C. jejuni, C. coli, "C. laridis," "C. fecalis," and aerotolerant campylobacters. The biochemical and physiological characteristics of the strains were examined for their correlation with the homology groups. The characterization tests that provided the most reliable differentiation at the species and subspecies level were growth at 25 and 42 degrees C, sensitivity to cephalothin and nalidixic acid, growth in semisolid media containing 1% glycine and 3.5% NaCl, growth on plates containing 1.5% NaCl, growth in a semisolid minimal medium, anaerobic growth in the presence of 0.1% trimethylamine-N-oxide, hydrogen sulfide production in SIM medium and triple-sugar iron agar, hippurate hydrolysis, nitrite reduction, and growth on plates under an air atmosphere.