Colony variation in Sinorhizobium meliloti inoculant strain U 45.

A culture of Sinorhizobium meliloti strain U 45, maintained on yeast extract-mannitol (YM) agar, produced a mixture of Congo red-absorbing (R1) and non-absorbing (W1) colonies when grown on YM medium containing Congo red. The original freeze-dried (FD) culture formed gummy (G), white (W2) and small red (R2) colony types on the above medium. All colonies were stable except G, which segregated into G and W2-like types. Immune diffusion patterns of all colony types were identical. The W1 colony type dominated R1 when a 1:1 combination was sub-cultured on YM agar. The parent cultures and their variants exhibited a range of N2-fixing effectiveness and competitiveness when inoculated onto two cultivars of Medicago sativa. Variant R2 from the FD culture was ineffective on both cultivars. Genomic DNA fingerprinting with insertion elements ISRm3 and ISRm2011-2 suggested that transposition of these elements was not a cause of variation, but a DNA band was absent in the profiles of two out of three W2-like colonies. Protein profile comparisons showed high similarity (r = 0.98) between the colony types when grown in YM broth. When grown on Tryptone-Yeast extract medium, variants from the FD and agar-maintained cultures formed separate clusters with r = 0.79. Polymerase chain reaction fingerprinting using repetitive, site-directed and arbitrary primers failed to differentiate the variants. The results emphasize the need to monitor culture variability to maintain the quality of legume inoculants.

Characterization of pigmented methylotrophic bacteria which nodulate Lotononis bainesii.

Root nodule isolates from a shrubby legume, Lotononis bainesii, were characterized by 16S rRNA gene sequencing and morphologically by substrate utilization patterns. The symbiotic genome of these isolates was analysed by partial sequencing of the nifH gene. Based on the results of numerical taxonomy, the isolates formed a closely related cluster, showing no correspondence to any of the known rhizobial clusters. Analysis of nearly full-length 16S rDNA sequences demonstrated that these isolates were related to Methylobacterium nodulans (SY et al., 2001). In the absence of nifH sequence data for the genus Methylobacterium, the nifH phylogeny showed these isolates to be related to Azospirillum brasilense. The facultative methylotrophic nature of these isolates was also demonstrated by their ability to grow in the presence of methanol as a sole carbon source.

Caries prediction factors in children with primary dentition.

Previous studies have indicated an association between the dmft and the lactobacilli counts in small children. This study evaluated and compared a number of salivary factors that could have an effect on caries progression in two groups of children with primary dentition (group 1 = 3-6 years; group II = 9 years). The average dmft score was higher for group II. The dmft score of group I consisted mainly of a large dt component, while in group II a large ft component was found. Lactobacilli were present in 44.83% of group I and in 77.27% of group II. Significant positive correlations were found for group I between the dt component of the dmft and lactobacilli count (P < 0.05, r = 0.48) as well as the total dmft and lactobacilli count (P < 0.05, r = 0.45). Significant positive correlations were found for group II between the dmft and lactobacilli count (P < 0.05, r = 0.39) and the plaque index and lactobacilli count (P < 0.05, r = 0.31). Significant correlations between the dmft and the prevalence of lactobacilli in the oral cavity were also indicated (group I: P < 0.05, r = 0.45; group II: P < 0.05, r = 0.36). Significant correlations confirmed the association of lactobacilli with the caries process and indicated the reliability of lactobacilli counts to determine caries activity. Correlations between the dmft and the prevalence of lactobacilli in the oral cavity indicated the possibility of an excellent but simple test for the prediction of caries susceptibility in children.

Classification of heparinolytic bacteria into a new genus, Pedobacter, comprising four species: Pedobacter heparinus comb. nov., Pedobacter piscium comb. nov., Pedobacter africanus sp. nov. and Pedobacter saltans sp. nov. proposal of the family Sphingobacteriaceae fam. nov.

Sixteen heparinase-producing isolates, related to Sphingobacterium heparinum, were grouped into three major clusters by SDS-PAGE and DNA-rRNA hybridizations. Based on a polyphasic approach, it was shown that isolates of two of these clusters and S. heparinum species belong to a new genus for which the name Pedobacter is proposed. The genus consists of Pedobacter heparinus comb. nov. (formerly Sphingobacterium heparinum), which is the type species, Pedobacter piscium comb. nov. (formerly Sphingobacterium piscium), Pedobacter africanus sp. nov. and Pedobacter saltans sp. nov. and four as-yet-unnamed DNA hybridization groups. All the previously named taxa can be discriminated by phenotypic features, but have strong overall similarities with representatives of the genus Sphingobacterium and the misclassified species [Flexibacter] canadensis. All these organisms constitute a separate rRNA branch in rRNA superfamily V for which the family Sphingobacteriaceae fam. nov. is proposed.

In vivo effects of urease-producing bacteria involved with the pathogenesis of infection-induced urolithiasis on renal urokinase and sialidase activity.

Many hypotheses have been proposed for renal stone formation. It has been argued that with infection-induced renal stones the hydrolysis of urea by bacterial urease increases urinary pH, with consequent stone formation. Unfortunately, this theory is not applicable to the micro-organisms that do not produce urease (e.g. Escherichia coli). It has been recently reported that E. coli reduces the urinary urokinase activity of male rats, but does not influence the urinary sialidase activity. This study has now been expanded to the urease-producing bacteria Proteus mirabilis, Staphylococcus aureus, S. epidermidis, Pseudomonas aeruginosa and Micrococcus luteus. Subcutaneous injections with these bacteria were found to significantly (P < 0.003) reduce the UK activity of extrarenally obstructed kidneys. The urease-producing mammalian skin bacterium, M. luteus, was, however, the exception (P = 0.1079). In contrast to S. epidermidis, P. aeruginosa and M. luteus (P < 0.0213), P. mirabilis and S. aureus had no effect on renal sialidase activity (P < 0.4047). These results may explain why Proteus species are predominant in infection-induced renal stones. According to the urokinase-sialidase hypothesis, a decrease in urinary urokinase activity should increase the uromucoid levels, whilst no effect on the urinary sialidase activity should favour conversion of urinary uromucoid to mineralizable matrix. These conditions may lead to renal stone formation. An increase in urinary pH resulting from urease-producing micro-organisms will increase salt precipitation on the uromucoid. It is thus concluded that urease-producing bacteria may play a double role in renal stone formation.

Anaerobic bacteria in orofacial abscesses.

Obligate anaerobic bacteria were cultured from 15 orofacial abscesses. Bacteroides species constituted 43 per cent, Clostridium 21 per cent, Fusobacterium 14 per cent, Peptostreptococci 11 per cent, peptococci 7 per cent and Veillonella 4 per cent of the isolates. This study confirms the polymicrobial nature of orofacial infections. Sensitivity to antibiotics was unpredictable. Clostridium difficile, Clostridium tetani, Peptostreptococcus productus and Veillonella parvula showed resistance to some of the most frequently used antibiotics. Ampicillin and tetracycline were the most effective antibiotics and the highest resistance was shown against erythromycin.

Effects of bacteria involved with the pathogenesis of infection-induced urolithiasis on the urokinase and sialidase (neuraminidase) activity.

It has been hypothesized that urinary urokinase and sialidase may play a role in urolithiasis. If these theories have substance it is to be expected that microorganisms may also affect these enzymes, since the association between urinary tract infection and renal stone formation is well known. It is generally assumed that Proteus mirabilis and Staphylococcus albus, which produce the urea-splitting enzyme urease, are responsible for stone formation. However, the importance of non-urease-producing microorganisms (Escherichia coli and Enterococcus) in urolithiasis is unclear. Spectrophotometric studies were therefore devised to clarify this problem. Microorganisms associated with infection-induced stones (Proteus mirabilis and Escherichia coli) respectively inhibited the urokinase and stimulated the sialidase activity. In contrast, microorganisms which were not associated with infection stones (Bacillus subtilis) had significantly less effect on urokinase and sialidase activity. This study may explain infection-induced stone formation and could open a completely new line of research.

Reinterpretation of the taxonomic position of Xanthomonas maltophilia and taxonomic criteria in this genus. Request for an opinion.

The inclusion of "Pseudomonas maltophilia" Hugh 1981 in the genus Xanthomonas as Xanthomonas maltophilia (Hugh 1981) Swings et al. 1983 is questioned in view of the significant differences between these two taxa. This reclassification is not acceptable if practical means of differentiation in this genus are considered. The proposed alteration of the description of the genus Xanthomonas is also questionable because of the implications for everyday phytobacteriology. In view of the natural similarities, as well as the profound differences, between X. maltophilia and the genus Xanthomonas, we propose that a new genus should be created for X. maltophilia, which could be placed together with the genus Xanthomonas in a separate natural group.

Purification and characterization of a novel heparinase.

A unique heparinase was isolated from a recently discovered Gram-negative soil bacterium. The enzyme (heparinase III) was purified by hydroxylapatite chromatography, chromatofocusing, and gel permeation chromatography. The enrichment was 48x, and the specific activity of catalytically pure heparinase was 127 IU/mg of protein. Similar to the heparinase I from Flavobacterium heparinum, heparinase III also degrades heparin to mainly disaccharide fragments. It is specific for heparin and also breaks down heparan sulfate, but not hyaluronic acid and chondroitin sulfate. Heparinase III, however, differs markedly from heparinase I in several other aspects: it has a higher molecular mass (94 versus 43 kDa), pI (9.2 versus 8.5), its Km and kcat are different, and it has a higher energy of activation (15.6 versus 6.3 kcal/mol). Optimal activity was also found at higher pH (7.6 versus 6.5) and temperature (45 versus 37 degrees C). Furthermore, the amino acid composition of heparinase III is quite different from that of heparinase I.

Fluoroacetate-metabolizing pseudomonad isolated from Dichapetalum cymosum.

A pseudomonad was isolated from the fluoroacetate-producing plant Dichapetalum cymosum (Hook) Engl. and identified as Pseudomonas cepacia. We established that this isolate was capable of growing in fluoroacetate-enriched solutions without any reduction in growth rate. Our isolate of P. cepacia was capable of defluorinating 2.69 mg of fluoroacetate per 10(9) cells per h. Fluoroacetate was degraded to CO2 at a rate of 23.53 ng/10(9) cells per h.

Estradiol and catecholestradiols as possible genotoxic carcinogens.

Estradiol and other estrogens are not classified as genotoxic carcinogens, but rather as tumor promoters in the multistage process of carcinogenesis. This study is a reexamination of the carcinogenic status of estradiol and the catecholestradiols (2-OHE2 and 4-OHE2) with the recently developed bacterial assays for genotoxic carcinogens, the Chromotest. The bacterial kits revealed estradiol and catecholestradiols as biphasic and potential genotoxic carcinogens with the following SOS inducing potency values: E2 43,265 (SD 8,300); 2-OHE2 30,153 (SD 2,500), and 4-OHE2 68,939 (SD 4,500). The differences between these values are statistically highly significant (p less than 0.0005). These results were confirmed by studies on Escherichia coli, which showed an increase in cell number and a stimulation of DNA content in the presence of the estrogens. It is therefore concluded that estradiol and the catecholestradiols are possible genotoxic carcinogens which probably act as tumor inducers rather than tumor promoters.

Evaluation of Three Carcass Surface Microbial Sampling Techniques.

Three carcass surface microbial sampling techniques were evaluated: a double swab, an excision and an agar sausage technique. In each instance, a sampling area of 6,42 cm2 was used. For the double swab technique, two sterile dry swabs were used. A sterile meat borer was used to cut out the area of 6,42 cm2 for the excision technique. For the agar sausage technique, 50-cm3 medical syringes were used to take impression plate samples. All the samples obtained with the different techniques were subjected to serial dilutions, whereafter they were spread-plated in duplicate on prepoured plates. Results of the study indicated that there was a significant difference (P<0,05) between the three techniques. The excision technique was the most reliable while the agar sausage technique had a higher coefficient of determination (r2 value) with the excision technique than did the swab technique.

Ureolytic bacteria in sheep rumen.

Estimates were made of the numbers of viable bacteria in the rumens of sheep receiving different rations. Representative colonies were isolated and tested for urease production. Some urease-positive isolates were characterized and identified. The ureolytic activities of the urease-producing isolates were determined and compared with the activity of rumen fluid. The rations fed to the sheep did not exert a significant influence on the relative numbers of the urease-producting organisms in the rumen. No obligately anaerobic ureolytic bacteria were found. All urease-positive isolates were facultatively anaerobic, Gram-positive, catalase-positive cocci. Out of ten isolates, nine were identified as Staphylococcus saprophyticus and one as Micrococcus varians. The total urease activity of the different isolates based on the lowest numbers in which they were present in the rumen, compared favourably with the urease activity of rumen fluid. The facultatively anaerobic Gram-positive cocci were probably responsible for a large proportion of the urease activity of the rumen fluid. Conditions prevailing in the rumen were found to be conducive to urease production by the isolates tested.