The differentiation of asaccharolytic anaerobic gram-positive cocci by protein electrophoresis.

Conventional biochemical and antibiotic sensitivity tests were used to allocate 87 clinical isolates of anaerobic gram-positive cocci to currently recognised species, in comparison with type and other authentic reference strains. Whole-cell protein electrophoresis was then performed with extracts of each strain. Allowing for difficulties of standardisation, it was possible to allocate most of the organisms to species-related groups on the basis of protein patterns. Organisms identified conventionally as Peptostreptococcus anaerobius and P. micros formed homogeneous groups by protein electrophoresis. There was evidence for heterogeneity amongst strains identified as P. asaccharolyticus (two groups, including P. indolicus), P. prevotii and P. magnus. However, aberrant P. prevotii strains were allocated to the P. asaccharolyticus groups, leaving a homogeneous P. prevotii group, and if P. variabilis were re-instated as a species, the remaining P. magnus strains could be divided into two groups. Of the anaerobic gram-positive cocci in the National Collection of Type Cultures deposited by Hare, Group IV is P. magnus, Group IX is P. micros and Groups I, III and VIII appear to be related to the butyrate-producing species P. asaccharolyticus and P. prevotii, but are strongly saccharolytic.

Detection and quantification of microorganisms in a heterogeneous foodstuff by image analysis.

A relatively inexpensive image analysis system has been developed to semi-automate the detection and quantification of microbial growth in sections of food. A system based on an IBM PC compatible, with a frame store card, was programmed to scan Gram-stained sections using a motorized stage. Each field of view was thresholded after subtraction of a background image and the area between two thresholds measured. In the food studied it was found that, by using a size limit, it was possible to reduce the number of fields that needed to be examined by a microscopist to approximately 3% of those scanned. Visual examination was still required to distinguish bacterial cells from other stained objects which occasionally occur.

Characterization of Corynebacterium group JK by whole-cell protein patterns.

A total of 102 strains received as Corynebacterium 'group JK' were characterized by SDS-PAGE of their whole-cell proteins. Numerical taxonomy based on the protein pattern absorbance profiles indicated that 91 of the strains formed a cluster. Seventy strains isolated in the UK were identified as group JK, indicating the increasing detection of this group as opportunistic pathogens. Fine differences between strain patterns were visible but it was not possible to associate these with any particular clinical source.

Testosterone metabolism by isolated human axillary Corynebacterium spp.: a gas-chromatographic mass-spectrometric study.

Transformations of [4-14C]testosterone have been studied in Corynebacterium spp. isolated from the axillae of men. Metabolites have been separated by TLC and capillary gas chromatography and have been identified by gas chromatography-mass spectrometry (GC-MS). The introduction of a clean-up step using Florisil columns, prior to TLC, removed Tween-80 which co-extracted from the medium with the metabolites. This procedure greatly improved TLC resolution. Testosterone was converted enzymically to 5 alpha- and 5 beta-DHT, identification being assisted by the inclusion of [3,4-13C]testosterone in some incubations. Other metabolites formed enzymically were 4-androstene-3,17-dione, 5 beta-androstane-3,17-dione, 3 beta-hydroxy-5 beta-androstan-17-one and 5 beta-androstane-3 alpha .17 alpha-diol. Some spontaneous breakdown of [14C]testosterone occurred giving rise to 5 alpha (beta)-DHT, androstanediol and a monohydroxy-diketo-androstene, the latter being reduced enzymically to 2 monohydroxy-diketo-androstanes. Under the conditions used, no clear evidence has been obtained for the formation of 5 alpha-androst-16-en-3-one, an odorous steroid that occurs in the axillae of men; the possible reasons why we were unable to prove the biosynthesis of this compound are discussed.

Transformation of steroids by axillary coryneform bacteria.

Synopsis The contribution made by the apocrine glands to body odour has been discussed. In particular, the role of odorous steroids, such as 5alpha-androst-16-en-3-one (5alpha-A), and their formation by axillary bacteria, has been explored. Isotopically labelled testosterone was incubated (21 days at 37 degrees C) with strains of axillary bacteria, both separately and in mixtures, and the products characterized by capillary gas chromatography (GC) and GC-mass spectrometry. In many cases testosterone was converted into large yields of 5beta-dihydrotestosterone and 4-androstenedione, although 5alpha-dihydrotestosterone, 5beta-androstanedione and androstanediols were also produced. Mixing certain coryneform bacteria appeared to cause some additive and some inhibitory effects. A non-polar steroid, with polarity similar to that of 5alpha-A, was formed in some incubations, but identification could not be confirmed. Nevertheless, preliminary experiments have indicated that 5alpha-A is present in axillary hair of men, and the possible reasons why we have failed to prove the biosynthesis of this odorous steroid are discussed.

Fingerprinting Acinetobacter strains from clinical sources by numerical analysis of electrophoretic protein patterns.

A total of 57 strains of Acinetobacter calcoaceticus was fingerprinted by SDS-PAGE of cellular protein. All strains were also examined by conventional, API and N/F-Tek methods, and antibiotic sensitivity patterns were determined. In general, using the API 20E and N/F-Tek methods, it was possible to assign isolates of A. calcoaceticus to the two accepted "biotypes" or "varieties", A.c. anitratus and A.c. lwoffi, but these methods did not offer useful subdivision of the biotypes. Gel electrophoresis permitted subdivision of the strains into clusters in a manner which suggests that the technique may be valuable in typing strains isolated during outbreaks of infection in hospital.

The similarities between Pseudomonas paucimobilis and allied bacteria derived from analysis of deoxyribonucleic acids and electrophoretic protein patterns.

The chromosomal DNA was isolated and purified from 17 strains of Pseudomonas paucimobilis, and from the type or reference strains of Flavobacterium capsulatum, F. devorans, F. multivorum, 'Chromobacterium lividum', Xanthomonas campestris and seven species of Pseudomonas. The DNA base compositions (mol% G + C) of P. paucimobilis strains were between 62.2 and 68.6%, and typical strains had a mean value of 65.3 +/- 0.4 mol%, determined from thermal denaturation temperature. DNA-DNA molecular hybridization with 3H-labelled probe DNA from NCTC 11030 P. paucimobilis (the type strain) indicated that the species comprised a core of 13 closely related strains (74 to 96%), which included F. devorans NCIB 8195 (= ATCC 10829). Four P. paucimobilis strains displayed lower levels of hybridization (less than or equal to 38%). The hybridization results showed that P. paucimobilis was not closely related to allied yellow-pigmented bacteria or to other reference pseudomonads. The electrophoretic protein patterns of representative strains were analysed by computer-assisted techniques and similarity coefficients were calculated. A high degree of congruence was obtained with the DNA hybridization data, and the protein analyses indicated that the four atypical P. paucimobilis strains were heterogeneous and not a single group within the species. The generic affinities of P. paucimobilis, F. capsulatum, 'P. azotocolligans' and P. echinoides are uncertain, but available chemotaxonomic data indicate these species could provide the basis for a new genus.

Classification of Corynebacterium species from axillary skin by numerical analysis of electrophoretic protein patterns.

The genus Corynebacterium, defined by the presence of mesodiaminopimelic acid, arabinose and corynomycolic acids in the cell wall, contains a very diverse group of organisms according to the results of numerical analysis of protein patterns; 13 groups were distinguished in this study. Strains isolated from axillary skin and hair included representatives of several groups but many strains were placed in groups 1 and 6A. Neither of these groups contained any reference strains and may constitute hitherto undescribed species. The reference strains of C. diphtheriae formed a coherent group not closely related to any non-pathogenic Corynebacterium species.