Sporulating Strain Research Articles

In vitro and in vivo antifungal efficacy of individual and consortium Bacillus strains in controlling potato black scurf and possible development of spore-based fungicide

Potato is a major crop whose yield and quality are impacted by the pathogenic fungus Rhizoctonia solani. Here, a control approach based on the use of endophytic bacillus spores and bacterial consortia was assessed as an innovative solution to improve tuber productivity and health. Endophytic potato bacteria extracted from several organs (leaves, stems and roots) evaluated for their antagonistic activity against R. solani by the double culture technique and their ability to rot potato slices. Additionally, the ability of Bacillus strains to generate hydrolytic enzymes and produce lipopeptides (using ESI-MS/MS mass spectrometry) was assessed. Then, the effectiveness of selected bacterial strains, either individually or in consortium (after carrying out compatibility tests) in the suppression of R. solani was evaluated in the greenhouse. Six endophytes not affecting potato tubers and having an antagonistic effect against R. solani were identified: B. halotolerans SpS5, B. velezensis KnL15, B. aryabhattai FaR1, B. amyloliquefaciens LiR9, B. haynesii ReR10 which produce several types and combinations of lipopeptides belonging to different families (iturins, fengycins and surfactins) as well as hydrolytic enzymes (chitinase, protease and cellulase). The vegetative and sporulating strain SpS5, as well as the 5 other Bacillus and their Sps5-based consortia, were found to be very effective in increasing the yield of healthy, high-quality tubers and reducing disease severity. Based on the results mentioned above, a microbial fungicide from the spores of (i) B. halotolerans SpS5 for which we sequenced the whole genome or (ii) bacterial consortia could be developed to combat black scurf potato and other fungal infections.

Sporulating behavior of Bacillus licheniformis strains influences their population dynamics during raw milk holding

To understand the role of strain variability, population dynamics of 2 strains of Bacillus licheniformis, ATCC 6634 and ATCC 14580, were modeled as a function of temperature (4.0-12.0°C) and duration (0-72 h) using regression analysis. Based on the initial spiking of vegetative cells (approximately 4.0 log cfu/mL) and spores (approximately 2.0 log cfu/mL), regression equations, elucidating B. licheniformis growth behavior during raw milk holding at low temperature, were obtained. Contour plots were developed to determine the time-temperature combinations, keeping the population changes to less than 1.0 log. In vegetative cell spiking study of B. licheniformis ATCC 6634 (S1), cell population changes remained below 1.0 log up to 72 h at 8°C. For B. licheniformis ATCC 14580 (S2), 1.0 log shift was not observed only after 80 h at 8°C, indicating higher multiplication potential of S1 as compared with S2. As S2 was a readily sporulating strain, the vegetative spiking study showed spore formation at different storage temperatures. Evidence of some parallel germination was observed for this strain at 8°C or higher, when raw milk samples were spiked with spores. The experimental values obtained for sporeformers and spore counts were validated with contour plot-generated values. Overall, for raw milk samples predominated by the low sporulating strain, the contour plots suggested holding at 8°C or below for up to 72 h. In the case of the readily sporulating strain (S2), raw milk could be held at 8°C for 80 h, where little or no sporulation is observed. Sporulation behavior, germination and multiplication ability, strain variability, and temperature and duration of holding raw milk influenced the population dynamics of Bacillus species. However, in the presence of equivalent numbers of both types of sporulating strains in raw milk, despite strain variability, holding milk at 8°C for not more than 72 h would keep any cell population changes below 1.0 log. In addition, under these storage conditions, the population would remain as vegetative cells that are likely to be inactivated by pasteurization. The contour plots, so generated, would help predict the population shifts and define optimum holding conditions for raw milk before further processing.

Draft Genome Sequence of a Sporulating and Motile Strain of Lachnotalea glycerini Isolated from Water in Québec City, Canada.

ABSTRACTLachnotalea glycerini CCRI-19302 belongs to the genus Lachnotalea. The strain was isolated from a water sample harvested in Québec City, Canada. The genome assembly comprised 4,694,231 bp, with 34.6% GC content. This is the first documentation to report the genome sequence of a sporulating and motile strain of L. glycerini.

Fundamental principles governing sporulation efficiency: A network theory approach

Using network theory on an integrated time-resolved genome-wide gene expression data, we investigated the intricate dynamic regulatory relationships of transcription factors and target genes to unravel signatures that contribute to extreme phenotypic differences in yeast, Saccharomyces cerevisiae. We performed a comparative analysis of the gene expression profiles of two yeast strains SK1 and S288c which are known for high and low sporulation efficiency, respectively. The results based on various structural attributes of the networks, such as clustering coefficient, degree-degree correlations, and betweenness centrality suggested that a delay in crosstalk between functional modules can be construed as one of the prime reasons behind low sporulation efficiency of S288c strain. A more hierarchical structure in the late phase of sporulation in S288c indicated an attempt of this low sporulating strain to obtain modularity, which is a feature of early sporulation phase. Further, the weak ties analysis revealed that mostly meiosis-associated genes were the end nodes of the weak ties for the high sporulating SK1 strain, while for the low sporulating S288c strain these nodes were mitotic genes. This again was a clear indication of the delay in regulatory activities in the S288c strain, which are essential to initiate sporulation. Our results demonstrate the potential of this framework in identifying candidate nodes contributing to phenotypic diversity in natural populations with application prospects in drug target discovery and personalized health.

Ogataea thermophilasp. nov., the teleomorph ofCandida thermophila

Ascospore formation was observed in the type strain of Candida thermophila Shin K-S, Shin YK, Yoon and Park on some yeast sporulation media. In addition, a further sporulating strain was found that proved to be conspecific with C. thermophila on the basis of sequences of the D1/D2 domain of the large-subunit (26S) rRNA gene and the internal transcribed spacer (ITS)1-5.8S rRNA gene - ITS2 region. Therefore, Ogataea thermophila Péter, Tornai-Lehoczki, Shin K-S & Dlauchy sp. nov. is proposed as the teleomorph of C. thermophila. The type strain is Y94(T)=JCM 10994(T)=KCCM 50661(T)=KCTC 17233(T).

Four Linked Genes Participate in Controlling Sporulation Efficiency in Budding Yeast

Quantitative traits are conditioned by several genetic determinants. Since such genes influence many important complex traits in various organisms, the identification of quantitative trait loci (QTLs) is of major interest, but still encounters serious difficulties. We detected four linked genes within one QTL, which participate in controlling sporulation efficiency in Saccharomyces cerevisiae. Following the identification of single nucleotide polymorphisms by comparing the sequences of 145 genes between the parental strains SK1 and S288c, we analyzed the segregating progeny of the cross between them. Through reciprocal hemizygosity analysis, four genes, RAS2, PMS1, SWS2, and FKH2, located in a region of 60 kilobases on Chromosome 14, were found to be associated with sporulation efficiency. Three of the four “high” sporulation alleles are derived from the “low” sporulating strain. Two of these sporulation-related genes were verified through allele replacements. For RAS2, the causative variation was suggested to be a single nucleotide difference in the upstream region of the gene. This quantitative trait nucleotide accounts for sporulation variability among a set of ten closely related winery yeast strains. Our results provide a detailed view of genetic complexity in one “QTL region” that controls a quantitative trait and reports a single nucleotide polymorphism-trait association in wild strains. Moreover, these findings have implications on QTL identification in higher eukaryotes.

Isolation, characterisation and expression patterns of a RAD51 ortholog from Pleuotus ostreatus

Using degenerated primers for conserved regions of Rec A homologs we have isolated a gene from Pleurotus ostreatus that shows characteristic features of RAD51 homologs. The encoded amino acid sequence of P. ostreatus RAD51 ( PoRAD51 ) shows greatest sequence similarities with RAD51 from Coprinus cinereus (89% identity). Furthermore the genomic organisation of PoRAD51 is almost identical to that of RAD51 from C. cinereus. Northern analysis shows that the expression of PoRAD51 is found in vegetative mycelium, and fruit body tissue, and that it is expressed at elevated levels in lamellae/basidia and following DNA damage. A sporulation deficient mutant strain of P. ostreatus (ATTC 58937) showed expression patterns of the RAD51 gene that are similar those of the normal sporulating strain.

Metabolic Pathway Rates and Fluorescence Measurements During Bioconversions by Non‐Growing Immobilized Clostridium Acetobutylicum

AbstractMetabolic and physiological aspects of non‐growing calcium alginate‐immobilized Clostridium acetobutylicum were investigated in a continuous‐flow reactor system. Changes in metabolic reaction rates and product selectivity were analyzed via metabolic pathway rate analysis, and variations in intracellular NAD(P)H‐dependent fluorescence were monitored using an on‐line fluorometer. In addition, the effects of sporulation were studied by comparing experiments utilizing either a sporulating strain (ATCC 824) or a sporulation mutant (ATCC 39236). In both cases, biocatalytic activity was lost through an initial period of cell lysis; afterwards, the sporulation mutant retained overall activity for a longer time because of the much lower degree of sporulation in this strain. The analysis of metabolic pathway rates in both experiments showed that early formation of solvent products was replaced by acid product formation, that the specific rates of all metabolic reactions were likely enhanced by the release of cytoplasmic compounds from the lysed cells (to a greater extent for the ATCC 824 cells), and that the patterns of specific activity loss were different between the two strains.

Netropsin increases formation of mRNA coding for a neutral metalloproteinase in Bacillus megaterium.

The anticancer drug netropsin increases the synthesis of an exocellular metalloproteinase during exponential growth as well as in the stationary phase of a sporulating strain of Bacillus megaterium. Its effect is due to a stimulation of the synthesis of the mRNA coding for the proteinase, determined as a residual synthesis of the enzyme in the presence of actinomycin D. The half-life of the proteinase mRNA (5-6 min at 35 degrees C) is not affected by netropsin. Netropsin relieves partially the repression of the proteinase mRNA caused by amino acids, whereas the repression brought about by an increased temperature is almost unaffected by the drug.

The Influence of Nutrition on the Growth and Sporulation of Two Strains ofEpicoccum Nigrum

SUMMARY Two strains of Epicoccum nigrum were cultured on potato-dextrose agar and on 27 test media. Morphological differences, colony size, and total number of spores were determined. On PDA, one strain, the sporulating strain, averaged 11 times more spores and 25% greater colony diam than the other, the mycelial strain. On the test media, nutritional differences also appeared. Starch was the preferred carbon source for the mycelial strain and glucose the preferred carbon source for the sporulating strain. Epicoccum is a cosmopolitan genus of the Moniliales. It occurs as a saprophyte or weak parasite on many plant species (13) and has been isolated from soil, air, insects, human skin, and textiles (15). In 1959 Schol-Schwarz studied over 70 isolates of Epicoccum and reduced most of them to synonymy with the type E. nigrum Link. Her studies

South American leaf blight of Hevea brasiliensis: culture of Microcyclus ulei

Microcyclus ulei (P. Henn.) Arx was isolated by depositing fresh conidia on plain agar and then transferring it to potato sucrose agar. Dried rubber leaf extract stimulated growth, but some brands of agar inhibited growth and sporulation. Two morphological strains occurred amongst field isolations. The free sporulating strain appeared to be associated with resistant clones. One isolate produced ascospores. Three distinct colony types were formed from cultures of laboratory-produced conidia and variants with a high degree of sporulation were obtained by successive subcultures. Single conidium cultures differed in mycelial growth, conidial and pycnidiospore production but single ascospore cultures were uniform in appearance. The fungus grew best at 23 °C, produced most conidia after three weeks, but optimum germination occurred with younger spores. Stromata were produced saprophytically on leaf disks bearing conidial lesions or pycnidia.

Motility of Bacillus subtilis during growth and sporulation.

The change of motility and the presence of flagella were followed throughout growth and sporulation in a standard sporulating strain and in 19 cacogenic sporulation mutants of Bacillus subtilis. For the standard strain, the fraction of motile cells decreased during the developmental period to less than 10% at T4. Motility was lost well before the cells lose their flagella. Conditions reducing the decrease of motility also reduced sporulation: motile cells never contained spores. The decrease of motility was not coupled with a decrease in the cellular concentration of adenosine 5'-triphosphate or a decline in oxygen consumption, but an uncoupling agent immediately destroyed motility at any time. Apparently, motility decreased during development because it became increasingly uncoupled from the energy generating systems of the cell. The motility of sporulation mutants decreased after the end of growth at the same time as or earlier than the motility of the standard strain; the early decrease of motility in an aconitase mutant, but not that in an alpha-ketoglurate dehydrogenase mutant, could be avoided by addition of L-glutamate. Sporulation or related events such as extracellular antibiotic or protease production were not needed for the motility decline.

Characteristics of phi T, the temperate bacteriophage carried by Bacillus megaterium 899a.

Phage T was the only phage observed in lysates of Bacillus megaterium 899a induced with mitomycin C, 0.35 mug/ml. The phage adsorbed slowly to its host in nutrient agar, giving rise to plaques of varying sizes and turbidity. Only clear plaques were observed when the phage and host cells were preincubated in an adsorption buffer and plated under optimum conditions. Plaque turbidity was caused by either the addition of 0.5 x 10(-2) to 1.0 x 10(-2) M CaCl(2) to the phage assay medium, or by raising the incubation temperature to 34 C. Phage T purified on a CsCl gradient had a density of 1.48 g/ml in CsCl and the extracted phage DNA had a buoyant density in CsCl of 1.6975 g/ml, equivalent to 38.2% guanine plus cytosine. The phage was rapidly inactivated at 75 C and was unstable in the presence of chloroform at 4 C, but it was stable in buffer stored in ice. When stage I sporulating cells were induced with mitomycin C, phage were carried into spores which when germinated lyse with the release of phi T. The burst size on induction of early-log vegetative cells was 52, whereas the burst size of induced T(0) sporulating cells, diluted in fresh medium, was 47 for a sporulating strain and 140 for an asporogenous mutant. A typical phage T had a long, noncontracting tail 240 nm long, 9 to 11 nm wide, with a repeating disk unit along the tail, 4 nm in size center to center. The tail ended in a small disk (15 nm wide) which is presumably for attachment to the host. The hexagonal head measures 68 by 57 nm and is composed of donut-shaped units 9 nm in diameter.

Effect of drying onParacoccidioides brasiliensis

The susceptibility to drying of Paracoccidioides brasiliensis growing on sterilized soils, was investigated using 2 strains with different sporulating capacities. Decreased fungal viability was noted during the 31 day experimental periods. The sporulating strain surpassed its initial viability at the fifth day when 75 days old culture was used, but not with 135 days old culture. Higher water losses, as observed at the 12th and 31st days, were not tolerated by any of the strains. Some viable particles resisted desiccation for at least 31 days and maintained their pathogenicity for hamsters. Colonies developed on small pieces of wood in the inoculum.A progressive drop of viability of the fungus was observed when exposed to a 2°C. environmental temperature with 100% relative humidity. The sporulating strain seemed to resist the deteriorative effect more than did the non-sporulating strain.

Zellteilungsverhalten von Bacillus megaterium beim Übergang zu Kohlenstoffhunger

AbstractIn cultures of a sporulating strain of B. megaterium grown at different growth rates the rate of division and the distribution of cell length were measured electronmicroscopically. Samples were taken from two doubling times before carbon limitation through 2.5 hours after the shift down.Contrary to the optical density, which increased exponentially until the carbon source was exhausted, the frequency of division changed 1–2 generations before. In fast growing glucose‐cultures the cells were longer than those in the slow growing arabinose‐cultures. These first mentioned cells became shorter already one doubling before the shift down by a transitory increase in division rate. No difference in length of growing and starvating cells was found with the arabinose‐culture. In all cultures the final decrease of the frequency of division also took place before the stationary phase was reached. This indicates that the initiation of division is more sensitive to carbon starvation than the net growth.The shapes of the curves reflecting the distribution of cell size were nearly identical with growing and starving populations. Thus, cell elongation is inhibited by carbon exhaustion in all cells at the same time.During the starvation period not all division septa were completed. 3–10% of the cells with in the populations remained uncompletely divided.

SYNTHESIS AND DEGRADATION OF POLY-β-HYDROXYBUTYRIC ACID IN CONNECTION WITH SPORULATION OF BACILLUS MEGATERIUM

Slepecky, Ralph A. (Northwestern University, Evanston, Ill.), and John H. Law. Synthesis and degradation of poly-beta-hydroxybutyric acid in connection with sporulation of Bacillus megaterium. J. Bacteriol. 82:37-42. 1961.-The production of poly-beta-hydroxybutyrate has been followed in Bacillus megaterium, a sporulating strain, and B. megaterium strain KM, a nonsporulating strain, by an improved assay procedure and by the use of C(14)-acetate. The production of polymer in the KM strain follows the growth curve very slowly and reaches a peak at the time the cells are entering the stationary phase of growth. Slow utilization of polymer follows. When the sporulating strain is grown under conditions favorable for polymer production, no spores are formed; polymer production and utilization follow kinetics similar to those observed with asporogenous strains. When the sporulating strain is grown under conditions unfavorable for polymer production but favorable for sporulation, less polymer is produced and peak production occurs during the log phase of growth. Rapid utilization of the polymer precedes sporulation. If the medium is made favorable for polymer production by the addition of glucose and acetate and vigorous aeration conditions are used, sporulation can be obtained after good polymer production and subsequent utilization.