No description available.

No description available.

No description available.

No description available.

The abundance, spatial distribution and diversity of class Flavobacteria, a bacterial group with a major role in marine secondary production, was investigated in the Southern Ocean euphotic zone of the ice pack off Eastern Antarctica and along a transect at 140-144 degrees East between latitudes 44.7 degrees South to 63.5 degrees South. Samples were comparatively analysed using 16S rRNA gene-based denaturing gradient gel electrophoresis (DGGE), fluorescent in situ hybridisation, real-time PCR and sequence analysis. The results were subsequently compared with direct cultivation approaches. Surface seawater samples were filter-fractionated into particulate and planktonic fractions and the abundance of particle-associated Flavobacteria, ascertained with real-time PCR and DGGE band analysis using Flavobacteriaspecific primers. Flavobacteria abundance was found to be significantly higher in Polar Front Zone (PFZ) and Antarctic Zone (AZ) water samples compared to warmer, nutrient limited Temperate Zone (TZ) and Sub-Antarctic Zone (SAZ) waters. Abundance of particle-associated Flavobacteria positively correlated with seawater chlorophyll a and nutrient concentrations. The abundance of planktonic Flavobacteria populations in the same samples remained relatively static, suggesting increased Flavobacteria abundance may relate to enhanced primary production in the PFZ and AZ. This was supported by comparisons of DGGE profiles that demonstrated significant differences occur in the total Flavobacteria community structure and 16S rRNA gene diversity between samples from the PFZ and AZ with samples from the TZ and SAZ. This suggests a shift to a different, more psychrophilic Flavobacteria community occurs across the Polar Front in the Southern Ocean. DGGE band sequences revealed a high diversity of class Flavobacteria within the Southern Ocean, with 24 genus-level lineages detected. Several of the phylotype clades detected were cosmopolitan in distribution, present in both polar and temperate oceans. Many of the phylotypes clustered in a large, so far uncultivated clade (previously termed DE cluster 2) widely distributed in seawater but apparently absent from sea-ice. Cosmopolitan phylotype clades occurred throughout the Southern Ocean, while several additional phylotype groups were found only in the colder waters of the PFZ and AZ. Examination of the cultivable diversity of Flavobacteria in Southern Ocean water samples, using a range of growth media, revealed a number of unique phylotypes including three novel genera, some grouping in clades for which only clones are currently available. Several other strains represented novel species belonging to the family Flavobacteriaceae, grouping in the genera Psychroserpens, Polaribacter and Tenacibaculum. A number of seawater microcosms were utilized to examine the colonization of bacteria-free diatom detritus by planktonic bacterial communities over a period of 30 days at 2 degrees Celsius. Flavobacteria phylotypes colonizing diatom detritus, determined by DGGE and real-time PCR analysis, were homologous with the dominant phylotypes in the particle-associated fraction of the samples from which they were taken. Colonisation occurred rapidly (less than 5 days) and comprised a succession of phylotypes, representing a diversity of Flavobacteria lineages. The increasing colonization corresponded to an increase in the dissolution of silicic acid compared with the corresponding control microcosms

Since the isolation of bacteria in pure culture, investigations of microbial physiology have focussed on model microorganisms grown in pure culture. However, in order to understand complex environmental processes, there is a need to investigate mixed microbial communities. This is true for enhanced biological phosphorus removal (EBPR), a wastewater treatment process that results in the enrichment of polyphosphateaccumulating organisms (PAOs) and glycogen non-polyphosphate accumulating organisms (GAOs). PAOs and GAOs are key groups of physiologically distinct microorganisms that can proliferate in EBPR processes. PAOs drive the EBPR process, removing Pi from the wastewater, whereas GAOs negatively impact the process. In situ studies of Defluviicoccus in full-scale plants. Putative GAOs from the Alphaproteobacteria, Defluviicoccus (including Defluviicoccus vanus) were studied in full-scale EBPR plants to determine their distribution, abundance and ecological physiology (ecophysiology). Quantitative fluorescence in situ hybridization (FISH) demonstrated that Defluviicoccus were generally in low abundance, however in one plant surveyed, Cluster 2 Defluviicoccus comprised 9% of Bacteria. FISH combined with microautoradiography (MAR) revealed that both Cluster 1 and Cluster 2 Defluviicoccus were capable of taking up a narrow range of substrates including acetate, propionate, and pyruvate under anaerobic and aerobic conditions. Formate, butyrate, ethanol and several other organic carbon substrates were not taken up. Cluster 2 Defluviicoccus demonstrated a phenotype consistent with the current metabolic model for GAOs - anaerobic assimilation of acetate as polyhydroxyalkanoates (PHAs) with concomitant glycogen catabolism and aerobic consumption of PHA. Evidence was provided that these GAOs are likely to be unable to denitrify. The PAO, Accumulibacter and other GAOs (Competibacter) co-existed in two full-scale plants with Cluster 2 Defluviicoccus, but in both plants, the latter organisms were more abundant. Thus Cluster 2 Defluviicoccus can be relatively abundant and could be carbon (C) competitors with PAOs and other GAOs in EBPR plants. Bioenergetic models for Accumulibacter and Defluviicoccus. Investigations of acetate and inorganic phosphate (Pi) uptake in enrichments of Accumulibacter and acetate uptake in enrichments of Defluviicoccus were carried on lab-scale enrichment cultures and bioenergetic models were proposed. For both enrichments anaerobic acetate uptake assays in the presence of the protonophore, carbonyl cyanide m-chlorophenylhydrazone or the electrical potential () uncoupler valinomycin, indicated that acetate is likely to be taken up by a permease driven by the component of the proton motive force (pmf). Further investigation with the sodium ionophore monensin, suggested that anaerobic acetate uptake by Defluviicoccus may in part be sodium-dependent. Results of this study suggest that Accumulibacter generate a pmf for anaerobic acetate uptake by efflux of protons in symport with Pi through an inorganic phosphate transport system. In contrast, this study suggests that the anaerobic pmf in Defluviicoccus is generated by an efflux of protons across the cell membrane by the fumarate respiratory system, or by extrusion of sodium ions via decarboxylation of methylmalonyl-CoA. Aerobic Pi uptake by the Accumulibacter enrichment was strongly inhibited in the presence of an ATPase inhibitor (N, N’-dicyclohexylcarbodiimide), suggesting that the phosphate specific transport (Pst) system is important even under relatively high concentrations of Pi. Acetate permease activity in Accumulibacter and Defluviicoccus may play an important role in the competition for acetate in the often acetate limited EBPR process. Activity of a highvelocity Pst system in Accumulibacter may further explain its ability to compete strongly in EBPR. Anaerobic central metabolism in Accumulibacter. This study integrated in situ structure-function techniques, community biochemical measurements, metagenomic and transcriptomic analysis to study the physiology of Accumulibacter enriched in EBPR sludge communities. Anaerobic acetate uptake and assimilation as PHA in Accumulibacter was confirmed using FISH-MAR and post-FISH chemical staining. The effect of inhibitors on acetate uptake and storage polymer metabolism in the Accumulibacter enrichment was consistent with C flux through the glycolytic pathway and the glyoxylate cycle. Bioinformatic analysis of sequenced strains of Accumulibacter suggested that this PAO may interconvert intermediates of glycolysis and the glyoxylate cycle via malate-pyruvate and oxaloacetate-phosphenolpyruvate cycling. Investigation of gene expression in Accumulibacter demonstrated anaerobic activity of aconitase, isocitrate lyase, succinate dehydrogenase and cytochrome b/b6. A fusion protein including a novel cytochrome b/b6 complex likely facilitates energetically unfavourable anaerobic C flux through succinate dehydrogenase in Accumulibacter by pushing electrons uphill to more electronegative electron carriers. Physiological data from this study is interpreted in light of previous metagenomic information from enriched EBPR sludge communities and integrated with previous metabolic models for PAOs to develop a model for anaerobic central metabolism in Accumulibacter. Anaerobic central metabolism in Defluviicoccus. A lab-scale GAO enrichment culture dominated by Defluviicoccus was investigated to determine central metabolic pathways involved in anaerobic formation of PHA, a key carbon storage polymer essential for survival and proliferation of microorganisms in EBPR systems. Glycogen levels decreased under anaerobic conditions in the enrichment culture. However, no decrease in glycogen was observed in the presence of the glyceraldehyde-3-phosphate dehydrogenase inhibitor iodoacetate, inferring that glycogen is catabolized anaerobically in Defluviicoccus. FISH-MAR and post-FISH chemical staining supported the idea that acetate is converted to PHA in Defluviicoccus under anaerobic conditions. Anaerobic acetate uptake rates and PHA formation in the presence of metabolic inhibitors of central metabolic pathways in the Defluviicoccus enrichment culture was determined. Inhibition of isocitrate lyase by 3-nitropropionate and itaconate, indicated that C is likely to be channelled through the glyoxylate cycle in Defluviicoccus. Inhibitors of aconitase and succinate dehydrogenase suggested that aconitase but not succinate dehydrogenase was active, providing further support for the role of the glyoxylate cycle in these GAOs. The observed fumarate reductase inhibitor effect on PHA production indicated reduction of fumarate to succinate and the operation of the reductive branch of the TCA cycle. Intracellular polyhydroxyvalerate was measured in the Defluviicoccus enrichment and was likely produced by degradation of succinate generated by the glyoxylate cycle or fumarate reduction to propionyl-CoA followed by condensation with acetyl-CoA via a methylmalonyl-CoA intermediate.

No description available.

1. Sixteen New Zealand and 10 overseas isolates of Phytophthora cinnamomi Rands have been compared for oospore production in paired and single cultures. Considerable variability existed among the 26 isolates in respect of mating ability and responses to physical and chemical factors. Addition of steroids and did not influence oospore production. However, hypoxanthine increased oospore production in two isolates. 2. Discovery of the role of steroids in relation to growth and asexual sporulation of Phytophthora ssp. makes it possible to replace natural media by a synthetic medium. For this purpose an investigation was made into the carbon, nitrogen, vitamin and calcium requirements of P. cinnamomi. Twenty carbon compounds were tested as major sources of carbon with Ca(NO3)2 and glutamic acid as nitrogen sources. Dextrin, starch, and sucrose were the best sources of carbon for growth and asexual reproduction. Xylose also supported abundant sporulation, although growth was slow. Organic acids and alcohols were poor sources of carbon. The best sources of organic nitrogen tested for growth were glutamine and glutamic acid. Tryptophan, phenyl-alanine, and cystine appeared to be inhibitory. Of the vitamins tested, only thiamine was required by the fungus. Five calcium compounds added singly to the basal medium gave no increase in the growth of the fungus. 3. & 4. The effect of soil extracts on sporulation of P. cinnamomi has been investigated. Non-sterile soil extracts from different soil types and from soils under different plan species all supported sporulation, stimulatory activity being markedly reduced only after 1000-fold dilution. Autumn soil was most active in stimulating sporulation, wheras summer soil was inactive. When peptone was added to summer soil, the stimulatory activity of the extract was restored. A wide variety of materials added to distilled water failed to stimulate sporulation. Soil extract lost its stimulatory properties after sterilisation by autoclaving, steaming, Seitz or sintered glass filtration, dialysis, ultrasonic disintegration, ultracentrifugation or treatment with propylene oxide or alcohol. Root exudates and microorganisms other than bacteria had no effect on sporulation. Fifty-nine bacterial cultures classified in 15 genera stimulated sporulation on one or more occasions when transferred from nutrient broth cultures but not from nutrient agar slopes. Of 16 morphologically distinct bacteria isolated from wet autumn soil, two Pseudomonas species stimulated abundant sporulation when transferred from agar plates to sterile soil extract. The activity of these isolates was lost after six weeks in culture, but was partially restored by growing them in nutrient broth. Extracts from sterilised soil inoculated with attenuated cultures of these Pseudomonas species were stimulatory but subcultures from these extracts were only stimulatory if grown in broth. The difference in ability to stimulate sporulation between cultures from broth and from agar was not due to bacterial numbers, flagellation, or fimbriation, but was associated with the additional nutrient transferred with the bacterial inoculum from a broth culture. Streptomycin was the only one of several antibiotics tested which inhibited sporulation. Irrigation of P. cinnamomi mycelium with continuously-produced sterile filtrate from soil extracts produced sporangia in 48 hours.

1. Spirodela oligorrhiza on sterile glucose-mineral medium ceased to grow three days after transfer into darkness. 2. Cytokinins, supplied in the medium, allowed continuous growth of Spirodela after transfer into darkness. Other plant growth substances, or adenine analogues, were ineffective. 3. Kinetin stimulated production of new fronds after a 24 hour lag period when added to dormant cultures although it was rapidly taken up. Kinetin reached a constant concentration in the plantlets within 30 to 60 minutes of addition to the medium. 4. In the absence of cytokinin, dormancy continued for three or four weeks after which growth spontaneously resumed in darkness. The growth rate then reached almost half that achieved with optimum kinetin concentrations in darkness. Growth continued in darkness for at least eight weeks without cytokinin. 5. Pretreatment in the light with either metabolic inhibitors, kinetin, abscisic acid, or high or low temperature, essentially eliminated the period of dormancy of Spirodela transferred to darkness in the absence of cytokinin. Growth was reduced 50% to 90% during the pretreatments. 6. The pretreatments designed to affect plastid RNA, protein or ATP production were the most effective. The fastest growth rate achieved in darkness without cytokinin after pre-treatment was 10% less than that promoted by optimum kinetin. 7. Dormant Spirodela in darkness continued to incorporate precursors into RNA, DNA and protein at a rate 50% that in growing (plus kinetin) cultures. Net rates of macromolecule accumulation were extremely slow, indicating extensive degradation. 8. Addition of kinetine to non-growing Spirodela in darkness stimulated the synthesis of RNA, DNA and protein simultaneously after a lag of approximately one hour. The rates of precursor incorporation increased to equal those in continuously growing cultures. 9. Non-growing Spriodela in darkness rapidly accumulated starch. Kinetin had little or no effect on accumulation and mobilisation of starch. 10. 14C-glucose was taken up by growing (plus kinetin) and non-growing (minus kinetin) Spirodela in darkness, and was metabolized equally in each. Three times as much 14C-glucose entered starch in the non-growing cultures. 11. A model scheme for control of dormancy in Spirodela is proposed based on an inhibitory mediator. The mediator may be similar to the hypothetical mediator of the pleiotypic response shown by animal cells.

1. Sixteen New Zealand and 10 overseas isolates of Phytophthora cinnamomi Rands have been compared for oospore production in paired and single cultures. Considerable variability existed among the 26 isolates in respect of mating ability and responses to physical and chemical factors. Addition of steroids and did not influence oospore production. However, hypoxanthine increased oospore production in two isolates. 2. Discovery of the role of steroids in relation to growth and asexual sporulation of Phytophthora ssp. makes it possible to replace natural media by a synthetic medium. For this purpose an investigation was made into the carbon, nitrogen, vitamin and calcium requirements of P. cinnamomi. Twenty carbon compounds were tested as major sources of carbon with Ca(NO3)2 and glutamic acid as nitrogen sources. Dextrin, starch, and sucrose were the best sources of carbon for growth and asexual reproduction. Xylose also supported abundant sporulation, although growth was slow. Organic acids and alcohols were poor sources of carbon. The best sources of organic nitrogen tested for growth were glutamine and glutamic acid. Tryptophan, phenyl-alanine, and cystine appeared to be inhibitory. Of the vitamins tested, only thiamine was required by the fungus. Five calcium compounds added singly to the basal medium gave no increase in the growth of the fungus. 3. & 4. The effect of soil extracts on sporulation of P. cinnamomi has been investigated. Non-sterile soil extracts from different soil types and from soils under different plan species all supported sporulation, stimulatory activity being markedly reduced only after 1000-fold dilution. Autumn soil was most active in stimulating sporulation, wheras summer soil was inactive. When peptone was added to summer soil, the stimulatory activity of the extract was restored. A wide variety of materials added to distilled water failed to stimulate sporulation. Soil extract lost its stimulatory properties after sterilisation by autoclaving, steaming, Seitz or sintered glass filtration, dialysis, ultrasonic disintegration, ultracentrifugation or treatment with propylene oxide or alcohol. Root exudates and microorganisms other than bacteria had no effect on sporulation. Fifty-nine bacterial cultures classified in 15 genera stimulated sporulation on one or more occasions when transferred from nutrient broth cultures but not from nutrient agar slopes. Of 16 morphologically distinct bacteria isolated from wet autumn soil, two Pseudomonas species stimulated abundant sporulation when transferred from agar plates to sterile soil extract. The activity of these isolates was lost after six weeks in culture, but was partially restored by growing them in nutrient broth. Extracts from sterilised soil inoculated with attenuated cultures of these Pseudomonas species were stimulatory but subcultures from these extracts were only stimulatory if grown in broth. The difference in ability to stimulate sporulation between cultures from broth and from agar was not due to bacterial numbers, flagellation, or fimbriation, but was associated with the additional nutrient transferred with the bacterial inoculum from a broth culture. Streptomycin was the only one of several antibiotics tested which inhibited sporulation. Irrigation of P. cinnamomi mycelium with continuously-produced sterile filtrate from soil extracts produced sporangia in 48 hours.