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Biomedical subjects

L L Blackall

Publications and source records attributed to L L Blackall.

13 recordsLinked to original sources

Phylogenetic positions of phytoplasmas associated with dieback, yellow crinkle and mosaic diseases of papaya, and their proposed inclusion in 'Candidatus Phytoplasma australiense' and a new taxon, 'Candidatus Phytoplasma australasia'.

DNA extracted from three papaya (Carica papaya L.) plants, individually affected by dieback, yellow crinkle or mosaic diseases, was subjected to PCR using phytoplasma-specific primers to amplify the 16S rRNA gene plus 16S-23S rRNA intergenic spacer region. Near-complete DNA sequences obtained for the three PCR amplimers were subjected to phylogenetic analyses and direct sequence comparison with other phytoplasma 16S rDNA and 16S-23S spacer region DNA sequences. The papaya yellow crinkle (PpYC) and papaya mosaic (PpM) sequences were identical to each other, but distinctly different from the papaya dieback (PpDB) sequence, showing 90.3% identity in the he 16S rDNA and 87.8% identity in the 16S-23S spacer region DNA sequences. A phylogenetic tree based on 16S rDNA sequences was calculated, in which PpYC and PpM are most closely related to the tomato big bud phytoplasma (TBB; 99.7% 16S rDNA sequence identity) from Australia, within subclade iii. This subclade consists of strains only reported occurring in the Southern Asian region and Australia, which indicates an Asian/Australasian origin. PpDB is most closely related to the Phormium yellow leaf phytoplasma from new Zealand (PYL; 99.9% identity) and the Australian grapevine yellows phytoplasma (AGY; 99.7% identity). These three phytoplasma strains form a distinct clade within subclade xii, which also includes the European strains STOL and VK as another distinct clade. The origin of the closely related but geographically separated AGY-like strains and STOL-like strains of subclade xii is unclear. It is proposed that phytoplasma strains PpDB, PYL and AGY be included in the previously described taxon 'Candidatus Phytoplasma australiense', and that PbYC, PpM and TBB be assigned to a new taxon, "Candidatus Phytoplasma australasia'.

Base Sequence

Microbiology of a nitrite-oxidizing bioreactor.

The microbiology of the biomass from a nitrite-oxidizing sequencing batch reactor (NOSBR) fed with an inorganic salts solution and nitrite as the sole energy source that had been operating for 6 months was investigated by microscopy, by culture-dependent methods, and by molecular biological methods, and the seed sludge that was used to inoculate the NOSBR was investigated by molecular biological methods. The NOSBR sludge comprised a complex and diverse microbial community containing gram-negative and gram-positive rods, cocci, and filaments. By culture-dependent methods (i.e., micromanipulation and sample dilution and spread plate inoculation), 16 heterotrophs (6 gram positive and 10 gram negative) were identified in the NOSBR sludge (RC), but no autotrophs were isolated. 16S ribosomal DNA clone libraries of the two microbial communities revealed that the seed sludge (GC) comprised a complex microbial community dominated by Proteobacteria (29% beta subclass; 18% gamma subclass) and high G + C gram-positive bacteria (10%). Three clones (4%) were closely related to the autotrophic nitrite-oxidizer Nitrospira moscoviensis. The NOSBR sludge was overwhelmingly dominated by bacteria closely related to N. moscoviensis (89%). Two clone sequences were similar to those of the genus Nitrobacter. Near-complete insert sequences of eight RC and one GC N. moscoviensis clone were determined and phylogenetically analyzed. This is the first report of the presence of bacteria from the Nitrospira phylum in wastewater treatment systems, and it is hypothesized that these bacteria are the unknown nitrite oxidizers in these processes.

Bacteria

The filamentous morphotype Eikelboom type 1863 is not a single genetic entity.

Five isolates of a filamentous bacterial morphotype with the distinctive diagnostic microscopic features of Eikelboom Type 1863 were obtained from activated sludge sewage treatment plants in Victoria, Australia. On the basis of phenotypic evidence and 16S rDNA sequence data, these isolates proved to be polyphyletic. Two (Ben 06 and Ben 06C) are from the Chryseobacterium subgroup which is in the Cytophaga group, subdivision I of the Flexibacter-Cytophaga-Bacteroides phylum. Two (Ben 56 and Ben 59) belong to the genus Acinetobacter, and one (Ben 58) is a Moraxella sp., closest to Mor. osloensis. The significance of these findings to the reliance on microscopic features for identification of these filamentous bacteria in activated sludge is discussed.

DNA, Bacterial

Phenotypic and phylogenetic description of an Italian isolate of "Microthrix parvicella".

"Microthrix parvicella" strain RN1 was isolated from an activated sludge treatment plant in Italy using micromanipulation techniques. The strain grows as thin unbranchedfilaments which are Gram-positive with Neisser-positive granules. The isolate was characterized by analysis of the 16S rDNA which was amplified directly from cell biomass by the polymerase chain reaction and sequenced. "Microthrix parvicella" strain RN1 presents a very high similarity (100%) with another "M. parvicella" strain recently isolated in Australia, suggesting that this micro-organism, a novel, deep branching member of the actinomycetes subphylum, is the same causing the common events of bulking and foaming phenomena in activated sludge treatment plants throughout the world.

DNA, Bacterial

The characterization and description of representatives of 'G' bacteria from activated sludge plants.

The name Tetracoccus cechii is proposed for two strains of the tetrad arranged cocci, previously known as 'G' bacteria, which were isolated from laboratory scale activated sludge plants in the Czech Republic and in Italy. They were morphologically, phenotypically and phylogenetically characterized and found to comprise a novel lineage in the alpha-3 group of the proteobacterial phylum in the domain Bacteria. The strains are Gram-negative and produce intracellular inclusions of poly-beta-hydroxybutyrate. Although commonly seen in activated sludge mixed liquor as cocci 1-2 microns in diameter, arranged in tetrads, in pure culture they can also grow in amorphous aggregations and the cells are generally more variable in their size and shape with coccobacilli as well as cocci being present. They are not able to grow phototrophically, nor can they reduce nitrate beyond nitrite nor grow anaerobically. The closest phylogenetic neighbours of T. cechii are Rhodobacter sphaeroides and R. capsulatus which are 93% similar by 16S rDNA comparison. Tetracoccus cechii is oxidase- and catalase-positive, non-motile and has an optimal growth temperature between 25 degrees and 35 degrees C. The 16S rRNA of T. cechii has a 21 nucleotide deletion in the V9 region (Escherichia coli positions 1258-1278) and this feature is a unique molecular synapomorphy in the alpha-3 group.

Base Sequence

PCR detection of Clostridium perfringens producing different toxins in faeces of goats.

A polymerase chain reaction (PCR) was used to identify the genes encoding the major toxins of Clostridium perfringens in faeces of goats. When pure cultures of Cl. perfringens types A, B, C, D and E were used as templates in the PCR, amplicons were observed on the agarose gel as bands at approximately the 247 (alpha primers), 1025 (beta primers), 403 (epsilon primers) and 298 (iota primers) bp level of the DNA marker. When used to identify different types of Cl. perfringens in samples artificially spiked with these micro-organisms, the PCR detected as few as 1-1.5 x 10(2) cfu g-1 of the five types of Cl. perfringens tested. The PCR technique allowed the identification and typing of Cl. perfringens strains in faeces of goats, without recourse to other techniques such as the mouse neutralization test.

Animals

A proposal to reclassify Nocardia pinensis Blackall et al. as Skermania piniformis gen. nov., comb. nov.

The type strain of Nocardia pinensis was the subject of chemotaxonomic and 16S ribosomal DNA sequencing studies. The resultant nucleotide sequence was aligned with the sequences of representatives of the genera Corynebacterium, Dietzia, Gordona, Mycobacterium, Nocardia, Rhodococcus, and Tsukamurella, and phylogenetic trees were generated by using the Fitch-Margoliash, maximum-parsimony, maximum-likelihood, and neighbor-joining methods. It was evident from the phylogenetic analyses that N. pinensis represents a distinct phyletic line that is most closely associated with the Gordona clade. This genealogical evidence, together with chemotaxonomic and phenotypic data derived from this and previous studies, indicates that N. pinensis merits generic status within the family Nocardiaceae. Therefore, we propose that N. pinensis Blackall et al. 1989 be reclassified as Skermania piniformis gen. nov., comb. nov. The type strain of Skermania piniformis cleaved an array of conjugated substrates based on the fluorophores 7-amino-4-methylcoumarin and 4-methylumbelliferone.

Bacteriological Techniques

rRNA sequences and evolutionary relationships among toxic and nontoxic cyanobacteria of the genus Microcystis.

A primary-structure analysis of the 16S rRNA gene was performed with 10 strains representing five described and one unidentified species of the genus Microcystis. The phylogenies determined illustrate the evolutionary affiliations among Microcystis strains, other cyanobacteria, and related plastids and bacteria. A cluster of 10 strains that included hepatotoxic isolates identified as Microcystis aeruginosa formed a monophyletic group. However, the genus Microcystis appeared to be polyphyletic and contained two strains that clustered with unicellular cyanobacteria belonging to the genus Synechococcus. The clustering of related Microcystis strains, including strains involved in the production of the cyclic peptide toxin microcystin, was consistent with cell morphology, gas vacuolation, and the low G + C contents of the genomes. The Microcystis lineage was also distinct from the lineage containing the unicellular genus Synechocystis and the filamentous, heterocyst-forming genus Nostoc. The secondary structure of a Microcystis 16S rRNA molecule was determined, and genus-specific sequence signatures were used to design primers that permitted identification of the potentially toxic cyanobacteria belonging to the genus Microcystis via DNA amplification.

Bacterial Proteins

"Candidatus Microthrix parvicella", a filamentous bacterium from activated sludge sewage treatment plants.

"Candidatus Microthrix parvicella" is a filamentous bacterium that grows with great difficulty in cultures from the mixed liquor of activated sludge sewage treatment plants. It is gram positive, and the ultrastructure of its cell walls has been determined to be of the gram-positive type by electron microscopical examination. Phylogenetically, it is a deep-branching member of the subphylum actinomycetes within the gram-positive phylum of the domain Bacteria. As for phenotypic features, it is known that the organism contains a polyphosphate inclusions and that it is catalase positive. In mixed cultures in activated sludge plants and in pure culture in the laboratory, it has a characteristic and distinctive winding filamentous morphology, with filaments hundreds of micrometers long.

Gram-Positive Bacteria

Detection by polymerase chain reaction of Clostridium perfringens producing epsilon toxin in faeces and in gastrointestinal contents of goats.

A polymerase chain reaction (PCR) was used to identify the gene-encoding epsilon toxin production in Clostridium perfringens types B and D in faeces and in gastrointestinal contents of goats. The samples were cultured in thioglycollate broth and centrifuged. The upper layer of the pellet was used as a template for PCR, obviating the need for DNA extraction. This technique specifically differentiated Cl. perfringens types B and D from Cl. perfringens types A and C and from Escherichia coli. When used to identify Cl. perfringens type D in samples artificially spiked with the micro-organism, the PCR detected as few as 1.4 x 10(2) cfu g-1 of sample. Gastrointestinal contents and faeces were collected from 20 goats at slaughter and processed by PCR. Several positive results were obtained from the first five goats that were slaughtered and sampled a few days after their arrival at the abattoir, but only a few samples gave positive results during the following weeks, after the goats had been fed a concentrated ration containing monensin. A possible role of this drug in control of enterotoxaemia is suggested.

Animals

The opportunistic pathogen Nocardia farcinica is a foam-producing bacterium in activated sludge plants.

A Gram-positive unicellular coccal-diphtheroid rod causing foam in an activated sludge plant was successfully isolated by micromanipulation. Phenotypic characterization and 16S rDNA sequencing identified it as Nocardia farcinica. This is the first report that this opportunistic pathogen is a foam-causing bacterium in activated sludge, and the clinical implications of these observations are discussed.

DNA, Bacterial

Bacterial community structures of phosphate-removing and non-phosphate-removing activated sludges from sequencing batch reactors.

The bacterial community structures of phosphate- and non-phosphate-removing activated sludges were compared. Sludge samples were obtained from two sequencing batch reactors (SBRs), and 16S rDNA clone libraries of the bacterial sludge populations were established. Community structures were determined by phylogenetic analyses of 97 and 92 partial clone sequences from SBR1 (phosphate-removing sludge) and SBR2 (non-phosphate-removing sludge), respectively. For both sludges, the predominant bacterial group with which clones were affiliated was the beta subclass of the proteobacteria. Other major groups represented were the alpha proteobacterial subclass, planctomycete group, and Flexibacter-Cytophaga-Bacteroides group. In addition, several clone groups unaffiliated with known bacterial assemblages were identified in the clone libraries. Acinetobacter spp., thought to be important in phosphate removal in activated sludge, were poorly represented by clone sequences in both libraries. Differences in community structure were observed between the phosphate- and non-phosphate-removing sludges; in particular, the Rhodocyclus group within the beta subclass was represented to a greater extent in the phosphate-removing community. Such differences may account for the differing phosphate-removing capabilities of the two activated sludge communities.

Acinetobacter