Search

La Trobe University Faculty of Science, Technology and Engineering Murray Darling Freshwater Research CentreMDFRC item.Routine microscopic examination of biomass samples from many activated sludge plants around the world reveals the presence of large numbers of both Gram positive and Gram negative coccoid cells often arranged in distinctive tetrads or sheets of tetrads. Three Gram negative tetrad cocci were successfully isolated into pure culture using micromanipulation from samples of biomass from plants in Tamworth, N.S.W., Australia, Verona, Italy and Macau. After extensive phenotypic characterisation and 16S rDNA sequence analysis, all isolates showed high overall similarity to each other, but were phylogenetically quite different to any previously sequenced bacterium. They have therefore been classified as three species of a new genus, the genus Amaricoccus in the a Proteobacteria in the domain Bacteria. None could accumulate polyphosphate granules, and production of intracellular poly-beta-hydroxybutyrate (PHB) granules depended on the carbon source used in the medium. A similar characterisation of the original isolate of the G-bacteria described by Cech and Hartman showed this organism also belonged to the same genus as the other three isolates, but as a distinct species. The practical implications of these observations are discussed.

Urinary tract infection (UTI) is a disease of extremely high incidence in both community and nosocomial settings. UTIs cause significant morbidity and mortality, with approximately 150 million cases globally per year. Uropathogenic Escherichia coli (UPEC) is the primary cause of UTI and is generally treated empirically. However, the rapidly increasing incidence of UTIs caused by multidrug-resistant UPEC strains has led to limited available treatment options and highlights the urgent need to develop alternative treatment and prevention strategies. In this study, we performed a comprehensive analysis to define the regulation, structure, function, and immunogenicity of recently identified UPEC vaccine candidate C1275 (here referred to as IrmA). We showed that the irmA gene is highly prevalent in UPEC, is cotranscribed with the biofilm-associated antigen 43 gene, and is regulated by the global oxidative stress response OxyR protein. Localization studies identified IrmA in the UPEC culture supernatant. We determined the structure of IrmA and showed that it adopts a unique domain-swapped dimer architecture. The dimeric structure of IrmA displays similarity to those of human cytokine receptors, including the interleukin-2 receptor (IL-2R), interleukin-4 receptor (IL-4R), and interleukin-10 receptor (IL-10R) binding domains, and we showed that purified IrmA can bind to their cognate cytokines. Finally, we showed that plasma from convalescent urosepsis patients contains high IrmA antibody titers, demonstrating the strong immunogenicity of IrmA. Taken together, our results indicate that IrmA may play an important role during UPEC infection., IMPORTANCE Uropathogenic E. coli (UPEC) is the primary cause of urinary tract infection (UTI), a disease of major significance to human health. Globally, the incidence of UPEC-mediated UTI is strongly associated with increasing antibiotic resistance, making this extremely common infection a major public health concern. In this report, we describe the regulatory, structural, functional, and immunogenic properties of a candidate UPEC vaccine antigen, IrmA. We demonstrate that IrmA is a small UPEC protein that forms a unique domain-swapped dimer with structural mimicry to several human cytokine receptors. We also show that IrmA binds to IL-2, IL-4, and IL-10, is strongly immunogenic in urosepsis patients, and is coexpressed with factors associated with biofilm formation. Overall, this work suggests a potential novel contribution for IrmA in UPEC infection.

© 2017 Future Medicine Ltd. Chlamydia trachomatis is the most prevalent sexually transmitted bacterial infection worldwide and the leading cause of preventable blindness. Reports have emerged of treatment failure, suggesting a need to develop new antibiotics to battle Chlamydia infection. One possible candidate for a new treatment is the protease inhibitor JO146, which is an effective anti-Chlamydia agent that targets the CtHtrA protein. CtHtrA is a lynchpin on the chlamydial cell surface due to its essential and multifunctional roles in the bacteria's stress response, replicative phase of development, virulence and outer-membrane protein assembly. This review summarizes the current understanding of CtHtrA function and presents a mechanistic model that highlights CtHtrA as an effective target for anti-Chlamydia drug development.

Neisseria meningitidis serogroups B and C have been responsible for the majority of invasive meningococcal disease in Australia, with serogroup B strains causing an increasing proportion of cases in recent years. Serogroup Y has typically caused sporadic disease in Australia. In 2002, a cluster of 4 cases was reported from a rural region in Queensland. Three of these cases were serogroup C, with 1 case diagnosed by molecular detection only, and the fourth case was identified as a serogroup Y infection. Genomic analysis, including antigen finetyping, multilocus sequence typing (MLST), and core genome MLST, demonstrated that the serogroup Y case, though spatially and temporally linked to a serogroup C disease cluster, was not the product of a capsule switch and that one of the serogroup C isolates had a deletion of the entire porA sequence.

Nano-textured surfaces (NTS) are critical to organisms as self-adaptation and survival tools. These NTS have been actively mimicked in the process of developing bactericidal surfaces for diverse biomedical and hygiene applications. To design and fabricate bactericidal topographies effectively for various applications, understanding the bactericidal mechanism of NTS in nature is essential. The current mechanistic explanations on natural bactericidal activity of nanopillars have not utilized recent advances in microscopy to study the natural interaction. This research reveals the natural bactericidal interaction between E.coli and a dragonfly wing's NTS using advanced microscopy techniques and propose a model. Contrary to the existing mechanistic models, this experimental approach demonstrated that the NTS of dragonfly wings has two prominent nanopillar populations and the resolved interface shows membrane damage occurred without direct contact of the bacterial cell membrane with the nanopillars. We propose that the bacterial membrane damage is initiated by a combination of strong adhesion between nanopillars and bacterium EPS layer as well as shear force when immobilised bacterium attempt to move on the NTS. These findings could help guide the design of novel bio-mimetic nanomaterials by maximising the synergies between both biochemical and mechanical bactericidal effects.Nano-textured surfaces (NTS) are critical to organisms as self-adaptation and survival tools. These NTS have been actively mimicked in the process of developing bactericidal surfaces for diverse biomedical and hygiene applications. To design and fabricate bactericidal topographies effectively for various applications, understanding the bactericidal mechanism of NTS in nature is essential. The current mechanistic explanations on natural bactericidal activity of nanopillars have not utilized recent advances in microscopy to study the natural interaction. This research reveals the natural bactericidal interaction between E.coli and a dragonfly wing's NTS using advanced microscopy techniques and propose a model. Contrary to the existing mechanistic models, this experimental approach demonstrated that the NTS of dragonfly wings has two prominent nanopillar populations and the resolved interface shows membrane damage occurred without direct contact of the bacterial cell membrane with the nanopillars. We propose that the bacterial membrane damage is initiated by a combination of strong adhesion between nanopillars and bacterium EPS layer as well as shear force when immobilised bacterium attempt to move on the NTS. These findings could help guide the design of novel bio-mimetic nanomaterials by maximising the synergies between both biochemical and mechanical bactericidal effects.

Melioidosis, caused by the bacterium Burkholderia pseudomallei , is an often severe infection that regularly involves respiratory disease following inhalation exposure. Intranasal (i.n.) inoculation of mice represents an experimental approach used to study the contributions of bacterial capsular polysaccharide I (CPS I) to virulence during acute disease. We used aerosol delivery of B. pseudomallei to establish respiratory infection in mice and studied CPS I in the context of innate immune responses. CPS I improved B. pseudomallei survival in vivo and triggered multiple cytokine responses, neutrophil infiltration, and acute inflammatory histopathology in the spleen, liver, nasal-associated lymphoid tissue, and olfactory mucosa (OM). To further explore the role of the OM response to B. pseudomallei infection, we infected human olfactory ensheathing cells (OECs) in vitro and measured bacterial invasion and the cytokine responses induced following infection. Human OECs killed >90% of the B. pseudomallei in a CPS I-independent manner and exhibited an antibacterial cytokine response comprising granulocyte colony-stimulating factor, tumor necrosis factor alpha, and several regulatory cytokines. In-depth genome-wide transcriptomic profiling of the OEC response by RNA-Seq revealed a network of signaling pathways activated in OECs following infection involving a novel group of 378 genes that encode biological pathways controlling cellular movement, inflammation, immunological disease, and molecular transport. This represents the first antimicrobial program to be described in human OECs and establishes the extensive transcriptional defense network accessible in these cells. Collectively, these findings show a role for CPS I in B. pseudomallei survival in vivo following inhalation infection and the antibacterial signaling network that exists in human OM and OECs.

Neutral capsular polysaccharides (CPSs) were isolated from Acinetobacter baumannii NIPH190, NIPH201, and NIPH615. The CPSs were found to contain common monosaccharides only and to be branched with a side-chain 1→3-linked β-d-glucopyranose residue. Structures of the oligosaccharide repeat units (K units) of the CPSs were elucidated by 1D and 2D (1)H and (13)C NMR spectroscopy. Novel CPS biosynthesis gene clusters, designated KL30, KL45, and KL48, were found at the K locus in the genome sequences of NIPH190, NIPH201, and NIPH615, respectively. The genetic content of each gene cluster correlated with the structure of the CPS unit established, and therefore, the capsular types of the strains studied were designated as K30, K45, and K48, respectively. The initiating sugar of each K unit was predicted, and glycosyltransferases encoded by each gene cluster were assigned to the formation of the linkages between sugars in the corresponding K unit.

Free to read on publisher website Type K82 c apsular polysaccharide (CPS) w as isolat ed from Acinetobacter baumannii LUH5534 . The s t ructure of a linear tetra saccharide repeating un it of the CPS w as established by sugar analysi s along with one - and two - dimensional 1 H and 13 C NMR spectroscopy . Proteins encoded by the KL82 capsule gene cluster in the genome of LUH5534 were assigned to roles in the synthesis of the K82 CPS . In particular, functions were assigned to two new glycosylt ransferases (Gtr152 and Gtr153) and a novel pyru vyltransferase, Ptr5, responsible for the synthesis of D - galactose 4,6 - ( R ) - pyruvic acid acetal ( D - Gal 4,6 R Pyr) .

The structures of capsular polysaccharides (CPSs) produced by different Acinetobacter baumannii strains have proven to be invaluable in confirming the role of specific genes in the synthesis of rare sugars through the correlation of genetic content at the CPS biosynthesis locus with sugars found in corresponding CPS structures. A module of four genes (rmlA, rmlB, vioA and vioB) was identified in the KL57 capsule biosynthesis gene cluster of A. baumannii isolate BAL_212 from Vietnam. These genes were predicted to direct the synthesis of 4-acetamido-4,6-dideoxy-d-glucose (N-acetylviosamine, d-Qui4NAc) and the K57 CPS was found to contain this monosaccharide. The K57 structure was determined and, in addition to d-Qui4NAc, included three N-acetylgalactosamine residues in the main chain, with a single glucose side branch. The KL57 gene cluster has not been found in any other A. baumannii genomes, but the rmlA-rmlB-vioA-vioB module is present in the KL119 gene cluster that would likely produce a d-Qui4NAc-containing CPS.

Hypocrealean fungi of the genus Metarhizium are known rhizospheric-competent symbionts that support increased plant growth and development. However, the establishment of inocula in the rhizosphere is typically low. We investigated the role of plant: fungal signals and specifically of the plant hormone strigolactone in colonisation of pea plant roots by M. anisopliae. Germination of conidiospores in root exudate and colonisation of the root by M. anisopliae were estimated on wild variety pea (Pisum sativum L. cv Torsdag) and the mutants, rms 5-3: strigolactone-deficient and rms 4-1: strigolactone-overproducing mutants. Condiospore germination was significantly lower in root exudates from the strigolactone-deficient pea (rms 5-3) than in exudates from wild and strigolactone -overproducing (rms 4-1) mutant plants. Colonisation of root segments was higher in wild and strigolactone -overproducing (rms 4-1) mutant plants than the strigolactone-deficient (rms 5-3) plants. This is the first report indicating that plant strigolactone increases both conidiospore germination and colonisation of the root by species of Metarhizium.

Australian eusocial stingless bees of the tribe Meliponini raise young on honey and pollen stored in pots made of resin-beeswax. These provisions harbour a diverse microbiome, including bacteria and fungi, that can contribute to improved nutritional value and preservation and thus to the overall health of the bee colony. Chemical analyses of Australian pot-pollen have suggested that some bioactive volatiles might originate from microbial fermentation. The gut microbiota of Australian Meliponini has been studied, but the microbiota associated with their nests remains unexplored. In this study, fungi and yeasts were isolated using selective high-osmolarity media and antibacterial agents from pot-pollen, pot-honey, brood cell walls (‘cerumen’) and food provisions from nine colonies of three Australian Meliponini species (Tetragonula carbonaria, T. hockingsi and Austroplebeia australis) located at the same site in South East Queensland. Bees from the 9 colonies were assumed to be foraging within the same radius. Fungi and yeasts were characterised by microscopy, sequencing of ITS and LSU ‘barcodes’ and subsequent BLAST search against GenBank, phylogenetic tree construction and amplicon concatenation. The relative abundance of bacteria in pot-pollen was estimated from 16S ‘barcodes’ by PCR amplification and Next Generation Sequencing using the MiSeq platform. Organisms identified include Ascomycota, Basidiomycota, Firmicutes and Proteobacteria previously unreported in nest materials of Australian Meliponini. The distribution of the microorganisms across the three bee species and their nest materials are described. Further analysis of yeast-derived volatiles is on-going as part of a larger project on the relationships between Australian Meliponini bees and yeast symbionts.

The composition of the bee microbiome has been shown to have remained stable across millennia. However, over a shorter timeframe, variations within and between the microbiota of colonies may illuminate key questions on the environmental versus inherited origins of the microbiome and, potentially, of interactions between symbionts. We used next generation sequencing to compare the microbiome of Australian stingless bees (Tetragonula carbonaria Smith) from pairs of related hives that had been separated across locations, preventing overlap in foraging. DNA was extracted from 6 whole bees from each colony and the V3-V4 region of the 16S rRNA gene ‘barcode’ was amplified and sequenced using the Illumina MiSeq. Reads were analysed using new bioinformatics tools to identify Exact Sequence Variants and to determine diversity within our sequencing data at the highest resolution possible. Database-independent analysis was used to extend the exploration beyond taxonomic assignment, and statistical measures were used to identify highly similar sequences that have dissimilar patterns of occurrence. Patterns of co-occurrence and absence of variants were used to describe diversity and explore potential cooperation and competition between symbionts. Microsatellite analysis of the individual bees will be used to determine the effects of kinship and environment on bee microbiota.

SUMMARYCampylobacter jejuniis responsible for most foodborne bacterial infections worldwide including Australia. The aim of this study was to investigate a combination of typing methods in the characterization ofC. jejuniisolated from clinical diarrhoeal samples (n= 20) and chicken meat (n= 26) in order to identify the source of infection and rank isolates based on their relative risk to humans. Sequencing of theflaAshort variable region demonstrated that 86% of clinical isolates had genotypes that were also found in chicken meat. A polymerase chain reaction binary typing system identified 27 different codes based on the presence or absence of genes that have been reported to be associated with various aspects ofC. jejunipathogenicity, indicating that not all isolates may be of equal risk to human health. The lipooligosaccharide (LOS) of theC. jejuniisolates was classified into six classes (A, B, C, E, F, H) with 10·4% remaining unclassified. The majority (72·7%) of clinical isolates possessed sialylated LOS classes. Sialylated LOS classes were also detected in chicken isolates (80·7%). Antimicrobial tests indicated a low level of resistance, with no phenotypic resistance found to most antibiotics tested. A combination of typing approaches was useful to assign isolates to a source of infection and assess their risk to humans.

Microbacterium sp. strain TNHR37B was isolated from a geothermal bore well sample (50°C) collected from a region of coal seam gas extraction activities. The 3.5-Mb genome with a G+C content of 69.9% contained unique genes, and a low similarity value for average nucleotide identity using BLAST was observed with the available 73 Microbacterium sp. genomes.

O-antigen polysaccharide is a major immunogenic feature of the lipopolysaccharide of Gram-negative bacteria, and most species produce a large variety of forms that differ substantially from one another. There are 18 known O-antigen forms in the Yersinia pseudotuberculosis complex, which are typical in being composed of multiple copies of a short oligosaccharide called an O unit. The O-antigen gene clusters are located between the hemH and gsk genes, and are atypical as 15 of them are closely related, each having one of five downstream gene modules for alternative main-chain synthesis, and one of seven upstream modules for alternative side-branch sugar synthesis. As a result, many of the genes are in more than one gene cluster. The gene order in each module is such that, in general, the earlier a gene product functions in O-unit synthesis, the closer the gene is to the 5΄ end for side-branch modules or the 3΄ end for main-chain modules. We propose a model whereby natural selection could generate the observed pattern in gene order, a pattern that has also been observed in other species.

Cellulosilyticum sp. strain I15G10I2 was isolated from a coal seam gas water treatment pond at the Spring Gully water treatment facility, Roma, Queensland, Australia. Analysis of the genome of 4,489,861 bp and G+C content of 35.23% revealed that strain I15G10I2 shared limited similarity to members of the genus Cellulosilyticum , family Lachnospiraceae .

We have developed a system called the Operon Assembly Protocol (OAP), which takes advantage of the homologous recombination DNA repair pathway in Saccharomyces cerevisiae to assemble full-length operons from a series of overlapping PCR products into a specially engineered yeast-Escherichia coli shuttle vector. This flexible, streamlined system can be used to assemble several operon clones simultaneously, and each clone can be expressed in the same E. coli tester strain to facilitate direct functional comparisons. We demonstrated the utility of the OAP by assembling and expressing a series of E. coli O1A O-antigen gene cluster clones containing various gene deletions or replacements. We then used these constructs to assess the substrate preferences of several Wzx flippases, which are responsible for translocation of oligosaccharide repeat units (O units) across the inner membrane during O-antigen biosynthesis. We were able to identify several O unit structural features that appear to be important determinants of Wzx substrate preference. The OAP system should be broadly applicable for the genetic manipulation of any bacterial operon and can be modified for use in other host species. It could also have potential uses in fields such as glycoengineering.

The Community for Open Antimicrobial Drug Discovery (CO-ADD) is a not-for-profit, collaborative approach to discovering new antibiotics. We access novel chemical diversity from academic synthetic chemists, who collectively possess millions of untested compounds with chemical diversity that lie outside commercial collections. We perform high-throughput antimicrobial screening of pure compounds derived from both synthetic and natural sources free of charge. The resulting data can be used by participants for publication, patenting and development purposes, and is fed back into the research community through an open-access database after a 2-year period during which information is kept confidential to the provider. CO-ADD is fundamentally asking two questions: can the community work together to address the global threat of antimicrobial resistance; and are there as yet undiscovered, novel antimicrobial compounds already present within our diverse global chemistry community?

Project DAQ540 has shown that pathogenic fungi have significant potential to be used as biopesticides against mirids, green vegetable bug (GVB) and Helicoverpa without impact on natural enemies, particularly to reduce early season pest pressure. Cloned, characterised fungal isolates of at least one fungal biopesticide were supplied to commercial interests and data to support registration and commercial production was analysed and reported. A decision model for mirids was developed and application extended to GVB. Focus groups with up to 12 growers or consultants were conducted near Toowoomba and at least one on-farm trial was carried out in Darling Downs, the Burnett or Central Queensland (QLD).

Genomes of 82 Acinetobacter baumannii global clones 1 (GC1) and 2 (GC2) isolates were sequenced and different forms of the locus predicted to direct synthesis of the outer core (OC) of the lipooligosaccharide were identified. OCL1 was in all GC2 genomes, whereas GC1 isolates carried OCL1, OCL3 or a new locus, OCL5. Three mutants in which an insertion sequence (ISAba1 or ISAba23) interrupted OCL1 were identified. Isolates with OCL1 intact produced only lipooligosaccharide, while the mutants produced lipooligosaccharide of reduced molecular weight. Thus, the assignment of the OC locus as that responsible for the synthesis of the OC is correct.

Free to read on publisher website Lipopolysaccharide is a major immunogenic structure for the pathogen Yersinia pseudotuberculosis, which contains the O-specific polysaccharide (OPS) that is presented on the cell surface. The OPS contains many repeats of the oligosaccharide O-unit and exhibits a preferred modal chain length that has been shown to be crucial for cell protection in Yersinia. It is well established that the Wzz protein determines the preferred chain length of the OPS, and in its absence, the polymerization of O units by the Wzy polymerase is uncontrolled. However, for Y. pseudotuberculosis, a wzz mutation has never been described. In this study, we examine the effect of Wzz loss in Y. pseudotuberculosis serotype O:2a and compare the lipopolysaccharide chain-length profile to that of Escherichia coli serotype O111. In the absence of Wzz, the lipopolysaccharides of the two species showed significant differences in Wzy polymerization. Yersinia pseudotuberculosis O:2a exhibited only OPS with very short chain lengths, which is atypical of wzz-mutant phenotypes that have been observed for other species. We hypothesise that the Wzy polymerase of Y. pseudotuberculosis O:2a has a unique default activity in the absence of the Wzz, revealing the requirement of Wzz to drive O-unit polymerization to greater lengths.

Histological analysis of gill samples taken from individuals of Latris lineata reared in aquaculture in Tasmania, Australia, and those sampled from the wild revealed the presence of epitheliocystis-like basophilic inclusions. Subsequent morphological, in situ hybridization, and molecular analyses were performed to confirm the presence of this disease and discovered a Chlamydia -like organism associated with this condition, and the criteria set by Fredericks and Relman's postulates were used to establish disease causation. Three distinct 16S rRNA genotypes were sequenced from 16 fish, and phylogenetic analyses of the nearly full-length 16S rRNA sequences generated for this bacterial agent indicated that they were nearly identical novel members of the order Chlamydiales . This new taxon formed a well-supported clade with “ Candidatus Parilichlamydia carangidicola” from the yellowtail kingfish ( Seriola lalandi ). On the basis of sequence divergence over the 16S rRNA region relative to all other members of the order Chlamydiales , a new genus and species are proposed here for the Chlamydia -like bacterium from L. lineata , i.e., “ Candidatus Similichlamydia latridicola” gen. nov., sp. nov.

Complementary sequences at the 5′ and 3′ ends of the dengue virus RNA genome are essential for viral replication, and are believed to cyclise the genome through long-range base pairing in cis. Although consistent with evidence in the literature, this view neglects possible biologically active multimeric forms that are equally consistent with the data. Here, we propose alternative multimeric structures, and suggest that multigenome noncovalent concatemers are more likely to exist under cellular conditions than single cyclised monomers. Concatemers provide a plausible mechanism for the dengue virus to overcome the single-stranded (+)-sense RNA virus dilemma, and can potentially assist genome transport from the virus-induced vesicles into the cytosol.

Deleterious responses to pathogens during infancy may contribute to infection and associated asthma. Chlamydia respiratory infections in early life are common causes of pneumonia and lead to reduced lung function and asthma. We investigated the role of interleukin-13 (IL-13) in promoting early-life Chlamydia respiratory infection, infection-induced airway hyperresponsiveness (AHR), and severe allergic airway disease (AAD). Infected infant Il13(-/-) mice had reduced infection, inflammation, and mucus-secreting cell hyperplasia. Surprisingly, infection of wild-type (WT) mice did not increase IL-13 production but reduced IL-13Rα2 decoy receptor levels compared with sham-inoculated controls. Infection of WT but not Il13(-/-) mice induced persistent AHR. Infection and associated pathology were restored in infected Il13(-/-) mice by reconstitution with IL-13. Stat6(-/-) mice were also largely protected. Neutralization of IL-13 during infection prevented subsequent infection-induced severe AAD. Thus, early-life Chlamydia respiratory infection reduces IL-13Rα2 production, which may enhance the effects of constitutive IL-13 and promote more severe infection, persistent AHR, and AAD.

Infection with Burkholderia pseudomallei causes melioidosis, a disease with a high mortality rate (20% in Australia and 40% in Southeast Asia). Neurological melioidosis is particularly prevalent in northern Australian patients and involves brain stem infection, which can progress to the spinal cord; however, the route by which the bacteria invade the central nervous system (CNS) is unknown. We have previously demonstrated that B. pseudomallei can infect the olfactory and trigeminal nerves within the nasal cavity following intranasal inoculation. As the trigeminal nerve projects into the brain stem, we investigated whether the bacteria could continue along this nerve to penetrate the CNS. After intranasal inoculation of mice, B. pseudomallei caused low-level localized infection within the nasal cavity epithelium, prior to invasion of the trigeminal nerve in small numbers. B. pseudomallei rapidly invaded the trigeminal nerve and crossed the astrocytic barrier to enter the brain stem within 24 h and then rapidly progressed over 2,000 μm into the spinal cord. To rule out that the bacteria used a hematogenous route, we used a capsule-deficient mutant of B. pseudomallei that does not survive in the blood and found that it also entered the CNS via the trigeminal nerve. This suggests that the primary route of entry is via the nerves that innervate the nasal cavity. We found that actin-mediated motility could facilitate initial infection of the olfactory epithelium. Thus, we have demonstrated that B. pseudomallei can rapidly infect the brain and spinal cord via the trigeminal nerve branches that innervate the nasal cavity.

Urinary tract infections (UTIs) caused by uropathogenic Escherichia coli (UPEC) are common infections in humans. Despite the substantial healthcare cost represented by these infections, the human immune response associated with the infection immediately following the onset of symptoms in patients remains largely undefined. We performed a prospective study aimed at defining the milieu of urinary cytokines in adult inpatients in the 24-48 h period immediately following hospital admission for acute cystitis due to UPEC. Urine samples, analyzed using 27-target multiplex protein assays, were used to generate immune profiles for patients and compared to age- and gender-matched healthy controls. The levels of multiple pro-inflammatory cytokines were significantly elevated in urine as a result of infection, an observation consistent with prior findings in murine models and clinical literature. We also identified significant responses for several novel factors not previously associated with the human response to UTI, including Interleukin (IL)-4, IL-7, IL-9, IL-17A, eotaxin, Granulocyte-macrophage colony-stimulating factor (GM-CSF) and several growth factors. These data establish crucial parallels between the human immune response to UPEC and murine model UTI studies, and emphasize the complex but poorly defined nature of the human immune response to UPEC, particularly in the immediate period following the onset of symptoms for acute cystitis.

The genome sequence of Caloramator mitchellensis strain VF08, a rod-shaped, heterotrophic, strictly anaerobic bacterium isolated from the free-flowing waters of a Great Artesian Basin (GAB) bore well located in Mitchell, an outback Queensland town in Australia, is reported here. The analysis of the 2.42-Mb genome sequence indicates that the attributes of the genome are consistent with its physiological and phenotypic traits.

One of the earliest plant responses to pathogens is the induced accumulation of reactive oxygen species (ROS). The superoxide ion is an important intermediate in the generation of ROS having a key regulatory function during plant–microbe interactions and is an important component in fungal development. The superoxide dismutase (SOD) family contributes to frontline defense via detoxification of reactive superoxide radical anions. Sclerotinia sclerotiorum (Lib.) de Bary is a necrotrophic fungal pathogen with a broad host range. S. sclerotiorum produces the non-specific phytotoxin and key pathogenicity factor, oxalic acid (OA). We have identified an S. sclerotiorum SOD (Sssod1) with high similarity to CuZnSODs. Sssod1 contains an open reading frame of 908 bp in length and is predicted to encode a protein of 155 amino acids that harbors the entire hallmark motifs associated with SOD function. Treatment with the CuZnSOD inhibitor diethyldithiocarbamate (DETC) resulted in delayed hyphal growth and sclerotial development in a dose-dependent manner. Mutants generated carrying an Sssod1 deletion (ΔSssod1) exhibited morphological defects similar to those observed with the inhibitor treatment. Moreover, ΔSssod1 was more sensitive than wild-type to menadione, a redox cycling agent. Expression of Sssod1 was induced following treatment with oxidizing agents and during interaction with plant host tissue the ΔSssod1 mutant was significantly reduced in virulence on both tomato and tobacco plants compared to wild-type. Interestingly, pathogenicity of the superoxide dismutase mutant was mostly restored following supplementation with oxalate. We also observed that ΔSssod1 was reduced in oxalate production by half. In accordance with reduced virulence, ΔSssod1 induced a host oxidative burst in adjacent uninfected cells, a phenotype indicative of active pathogen recognition by the host. Intriguingly, during wild-type infection, host ROS production was significantly reduced. These results suggest that wild-type Sclerotinia suppresses host defense responses during infection.

© 2015 Roulis et al. Background: The obligate intracellular bacterium Chlamydia pneumoniae is a common respiratory pathogen, which has been found in a range of hosts including humans, marsupials and amphibians. Whole genome comparisons of human C. pneumoniae have previously highlighted a highly conserved nucleotide sequence, with minor but key polymorphisms and additional coding capacity when human and animal strains are compared. Results: In this study, we sequenced three Australian human C. pneumoniae strains, two of which were isolated from patients in remote indigenous communities, and compared them to all available C. pneumoniae genomes. Our study demonstrated a phylogenetically distinct human C. pneumoniae clade containing the two indigenous Australian strains, with estimates that the most recent common ancestor of these strains predates the arrival of European settlers to Australia. We describe several polymorphisms characteristic to these strains, some of which are similar in sequence to animal C. pneumoniae strains, as well as evidence to suggest that several recombination events have shaped these distinct strains. Conclusions: Our study reveals a greater sequence diversity amongst both human and animal C. pneumoniae strains, and suggests that a wider range of strains may be circulating in the human population than current sampling indicates.

A strictly thermophilic anaerobe, designated strain VF08T, was isolated from a water sample collected from a Great Artesian Basin bore (registered bore number 22981) situated at Mitchell, QLD, Australia. Cells of isolate VF08Twere slightly curved, non-sporulating rods (1.5–3.5×0.4–0.8 μm), which stained Gram-negative but possessed a Gram-positive cell-wall ultrastructure. The strain grew optimally in tryptone-yeast extract-glucose (TYEG) medium at 55 °C (temperature growth range between 37 and 60 °C) and a pH of 7 (pH growth range, 6.0–9.0). Yeast extract or tryptone was required for growth on glucose, fructose, xylose, maltose, sucrose, raffinose, cellobiose, ribose, pyruvate, tryptone, peptone, Casamino acids, amyl media and serine, but could also support growth as the sole carbon source. End products from glucose fermentation were acetate, ethanol, CO2and H2. The strain reduced vanadium(V), but not iron(III), manganese(IV), elemental sulfur, sulfate, thiosulfate, sulfite, nitrate or nitrite in the presence of 0.2 % yeast extract, peptone, tryptone, glucose, sucrose and Casamino acids, but an increase in the growth rate or cell yield was not observed. Growth was inhibited by chloramphenicol, streptomycin, tetracycline, penicillin, ampicillin and ≥2 % NaCl (w/v). The G+C content of the DNA was 38.4±0.8 mol% as determined by the thermal denaturation (Tm) method. 16S rRNA gene sequence analysis revealed that isolate VF08Twas a member of the genusCaloramatorwithCaloramator australicusandCaloramator fervidus(formerlyClostridium fervidus) being the closest relatives with similarity values of 85.0 and 86.1 %, respectively, when helix 6 nucleotides were included in the analysis, and 95.2 % and 94 %, respectively, when these nucleotides were masked from the analysis. Further analysis revealed that strain VF08Tformed an individual cluster (cluster II) within the genusCaloramatorand could be distinguished from other species within the genusCaloramator(clusters I, III and IV) on the basis of signature nucleotides and differences in phenotypic traits. These data suggest that strain VF08Tis a novel species of the genusCaloramator, for which the nameCaloramator mitchellensissp. nov. is proposed. The type strain is VF08T(=JCM 15828T=KCTC 5735T). An emended description of the genusCaloramatoris also provided.

Encapsulation of probiotic bacteria in cross-linked alginate beads is of major interest for improving the survivability in harsh acid and bile environment and also in food matrices. Alginate micro beads (10-40 μm) containing the probiotics Lactobacillus rhamnosus GG and Lactobacillus acidophilus NCFM were produced by a novel technique based on dual aerosols of alginate solution and CaCl(2) cross linking solution. Extruded macro beads (approximately 2mm diameter) produced by the conventional method and micro beads produced by novel aerosols technique offered comparable protection to L. rhamnosus in high acid and bile environment. Chitosan coating of micro beads resulted in a significant increase in survival time of L. rhamnosus from 40 to 120 min in acid condition and the reduction in cell numbers was confined to 0.94 log over this time. Alginate macro beads are more effective than micro beads in protecting L. acidophilus against high acid and bile. Chitosan coating of micro beads resulted in similar protection to L. acidophilus in macro beads in acid and extended the survival time from 90 to at least 120 min. Viability of this organism in micro beads was 3.5 log after 120 min. The continuous processing capability and scale-up potential of the dual aerosol technique offers potential for an efficient encapsulation of probiotics in very small alginate micro beads below sensorial detection limits while still being able to confer effective protection in acid and bile environment.

A Gram-negative, aerobic, mesophilic, non-spore-forming, chemotrophic, chlorophyll-lacking, nitrogen-fixing bacterium, designated strain USBA 355T, was isolated from the saline spring ‘Salado de Consotá’ situated in the Colombian Andes. The non-flagellated cells of strain USBA 355T were straight to slightly curved rods (0.6–0.7 × 3.0–3.5 μm). Growth occurred optimally at 30 °C (growth temperature range between 20 and 40 °C), at pH 6.5–6.7 (pH growth range between 5.0 and 8.0) and at 0.5 % NaCl (w/v) (range between 0 and 4 %). The major quinone present was Q-10 and the predominant fatty acids identified were C19 : 0 cyclo ω8c, C18 : 1 ω7c and C18 : 0. The G+C content of the chromosomal DNA was 71±1 mol%. 16S rRNA gene sequence analysis indicated that strain USBA 355T formed a distant phylogenetic line of descent with members of the genus Thalassobaculum, family Rhodospirillaceae, class Alphaproteobacteria (90 % gene sequence similarity). Comparison of the phylogenetic, chemotaxonomic and physiological features of strain USBA 355T with all other members of the family Rhodospirillaceae suggested that it represents a novel genus and species for which the name Tistlia consotensis gen. nov., sp. nov. is proposed. The type strain of the type species is USBA 355T (=JCM 15529T=KCTC 22406T).

A mesophilic, strictly anaerobic, slightly halophilic bacterium, designated strain USBA 82T, was isolated from a terrestrial saline spring in the Colombian Andes. The non-spore-forming curved rods (5–7×1.3 μm) with pointed or rounded ends, stained Gram-negative and were motile by means of laterally inserted flagella. The strain grew optimally at 30 °C (growth range 20–40 °C), pH 7.3 (growth range pH 5.5–8.5) and 2 % (w/v) NaCl (growth range 0.1–7 % NaCl). The strain fermented peptides, amino acids and a few organic acids, but growth was not observed on carbohydrates, alcohols or fatty acids. The strain reduced thiosulfate and sulfur to sulfide. Sulfate, sulfite, nitrate and nitrite were not used as electron acceptors. On peptone alone, acetate, succinate, propionate and traces of ethanol were formed, but in the presence of thiosulfate, acetate and succinate were formed. The G+C content of the chromosomal DNA was 52 mol% (T m).16S rRNA gene sequence analysis indicated that strain USBA 82T was affiliated to Dethiosulfovibrio peptidovorans within the phylum Synergistetes with a similarity value of approximately 93 %. Based on the differences between the new strain and the type species of the genus Dethiosulfovibrio, we suggest that strain USBA 82T represents a novel species of the genus for which the name Dethiosulfovibrio salsuginis sp. nov. is proposed. The type strain is USBA 82T (=DSM 21565T=KCTC 5659T).

A strictly anaerobic, sluggishly motile, spore-forming, thermophilic bacterium, designated strain AeGT, was isolated from microbial mats colonizing a runoff channel formed by free-flowing thermal waters of a bore well (New Lorne Bore; registered number 17263) in the Great Artesian Basin, Australia. Cells of strain AeGT were curved rods (2.0–10.0×0.8–1.0 μm) and stained Gram-negative. The strain grew optimally in tryptone-yeast extract-citrate medium at 55 °C (range for growth between 45 and 60 °C) and pH 7.0 (range for growth between pH 6.5 and 8.0). Citrate and malate, but no other organic acids, carbohydrates or amino acids could be used in the presence of up to 0.1 % yeast extract. Although yeast extract and/or tryptone were required for growth on citrate, they did not support growth as sole carbon sources. Strain AeGT reduced thiosulfate and sulfite in the presence of 0.2 % yeast extract, but not Fe(III), Mn(IV), sulfate, elemental sulfur, nitrate or nitrite. Growth was inhibited by chloramphenicol, streptomycin, tetracycline, penicillin and ampicillin and in the presence of NaCl concentrations >1 %. The DNA G+C content was 55.4±1.0 mol% as determined by the thermal denaturation method. 16S rRNA gene sequence analysis indicated that strain AeGT was a member of the family Veillonellaceae, class ‘Clostridia’, phylum ‘Firmicutes’ and was most closely related to members of the genus Propionispora (mean 16S rRNA gene sequence similarity value to type strains was 90.8 %). Based on these results, strain AeGT is considered to represent a novel species in a new genus, for which the name Sporolituus thermophilus gen. nov., sp. nov. is proposed. The type strain of the type species is AeGT (=JCM 15556T=KCTC 5668T).

Soil-transmitted helminth (STH) infections are endemic in northern Viet Nam where the climate and agricultural practices, such as the use of human excreta as fertiliser and the use of wastewater for irrigation, favour transmission. An intervention was conducted in Yen Bai Province, north-west Viet Nam, to measure the effectiveness of single dose albendazole (400mg) administered every 4 months for reducing the prevalence of STH infections in women of reproductive age. Stool samples were collected from women before the intervention and 3 and 12 months post-intervention. Information on a range of demographic and socio-economic variables was also collected to measure the major risk factors for high STH burden in this area. The prevalence of hookworm, Ascaris lumbricoides and Trichuris trichiura infection in the baseline sample of 366 women were 76.2%, 19.2% and 29.1%, respectively. In the women who were surveyed at baseline and again at 3 and 12 months after the intervention (n=118) cure rates were 71.3% for hookworm, 87.0% for A. lumbricoides and 81.4% for T. trichiura by the end of the 12 month study period (i.e. after three doses of albendazole). The main risk factor for hookworm infection was if women worked outside (odds ratio (OR)=3.2 (95% Confidence Interval (CI) 1.6-6.2), P=0.001) and the major risk factor for A. lumbricoides and T. trichiura infection was a lack of education. Low educational attainment was also the strongest risk factor for co-infection with all three species of STH (OR=7.5 (95% CI 3.4-16.4), P

Bacterial consortium-AIE2 with a capability of contemporaneous Cr(VI) reduction and azo dye RV5 decolourization was developed from industrial wastewaters by enrichment culture technique. The 16S rRNA gene based molecular analyses revealed that the consortium bacterial community structure consisted of four bacterial strains namely, Alcaligenes sp. DMA, Bacillus sp. DMB, Stenotrophomonas sp. DMS and Enterococcus sp. DME. Cumulative mechanism of Cr(VI) reduction by the consortium was determined using in vitro Cr(VI) reduction assays. Similarly, the complete degradation of Reactive Violet 5 (RV5) dye was confirmed by FTIR spectroscopic analysis. Consortium-AIE2 exhibited simultaneous bioremediation efficiencies of (97.8 ± 1.4) % and (74.1 ± 1.2) % in treatment of both 50 mg l−1 Cr(VI) and RV5 dye concentrations within 48 h of incubation at pH 7 and 37°C in batch systems. Continuous bioreactor systems achieved simultaneous bioremediation efficiencies of (98.4 ± 1.5) % and (97.5 ± 1.4) % after the onset of steady-state at 50 mg l−1 input Cr(VI) and 25 mg l−1 input RV5 concentrations, respectively, at medium dilution rate (D) of 0.014 h−1. The 16S rRNA gene copy numbers in the continuous bioreactor as determined by real-time PCR assay indicated that Alcaligenes sp. DMA and Bacillus sp. DMB dominated consortium bacterial community during the active continuous bioremediation process.

A strictly anaerobic Fe(III)-reducing bacterium, designated strain Red1T, was isolated from the production water of the Redwash oilfield, USA. The cells were motile rods (1–5×0.5–0.6 μm) that stained Gram-negative and possessed polar flagella. Strain Red1T obtained energy from the reduction of Fe(III), Mn(IV), nitrate, elemental sulfur and trimethylamine N-oxide in the presence of a wide range of electron donors, including a variety of organic acids, alcohols, biological extracts and hydrogen. Strain Red1T was incapable of fermentative growth. The novel isolate grew optimally at 40 °C (temperature range for growth, 30–50 °C) and at pH 7 (pH range, 6–9) with 2 % (w/v) NaCl (NaCl range, 0.1–10 %, w/v). The DNA G+C content was 52.5 mol%. Phylogenetic analysis of the 16S rRNA gene sequence indicated that strain Red1T was a member of the order Desulfuromonadales within the class Deltaproteobacteria and most closely related to Geoalkalibacter ferrihydriticus Z-0531T (95.8 %), Desulfuromonas palmitatis SDBY1T (92.5 %) and ‘Desulfuromonas michiganensis’ BB1 (92.4 %). On the basis of phenotypic and phylogenetic differences, the novel strain is proposed to represent a novel species, Geoalkalibacter subterraneus sp. nov. (type strain Red1T=JCM 15104T=KCTC 5626T).

Paenibacillus sabina strain JD2, a chitinolytic marine bacterium, was isolated from sea dumps collected at Sultanpur near Bhavnagar, India and its nutritional requirement for chitinase production was defined using statistical optimisation method. Effect of 8 different medium components on chitinase production by Paenibacillus sabina strain JD2 was screened by Plackett-Burman design. The screened medium components were further used in central composite design where chitinase production, pH and biomass responses were used in different models to evaluate fit ones. After performing power transformation, quadratic model was found to be fit for chitinase response and 2F1 model was found to be fit for biomass response. Whilst for pH response, quadratic model was found to be fit without any requirement of power transformation. In multiresponse analysis, medium formulation consisting of (g/l): chitin 18, yeast extract 0.50, and CaCl2 0.08, was found to predict 82.93 U/ml of chitinase with overall highest desirability of 0.842 as compared to other formulations. The selection of model was done on basis of high adjusted R2 value and lowered p-value for each model in individual analysis of each response. Through desirability analysis, it was found that biomass and pH played an important role in increasing the chitinase production by Paenibacillus sabina strain JD2. Through statistical optimisation method, 2.74-fold increase in chitinase production was achieved as compared to unoptimised medium.

Herpes simplex virus type 1 variants selected by virus propagation in cultured cells in the presence of the sulfated oligosaccharide PI-88 were analyzed. Many of these variants were substantially resistant to the presence of PI-88 during their initial infection of cells and/or their cell-to-cell spread. Nucleotide sequence analysis revealed that the deletion of amino acids 33–116 of gC but not lack of gC expression provided the virus with selective advantage to infect cells in the presence of PI-88. Purified gC (Δ33–116) was more resistant to PI-88 than unaltered protein in its binding to cells. Alterations that partly contributed to the virus resistance to PI-88 in its cell-to-cell spread activity were amino acid substitutions Q27R in gD and R770W in gB. These results suggest that PI-88 targets several distinct viral glycoproteins during the course of initial virus infection and cell-to-cell spread.

Rosiglitazone is commonly used in the treatment of Type 2 diabetes. However, the cardiovascular outcomes with rosiglitazone have only been evaluated recently. Firstly, they were evaluated by meta-analysis. A total of 42 short-term trials, not designed to determine cardiovascular outcomes, were included in the meta-analysis. There were 72 myocardial infarctions in the control group of 12283 subjects (0.59%) and 86 in the rosiglitazone group of 15560 subjects (0.55%). This gave an odds ratio of 1.43 and a p-value of 0.03. As a consequence of this meta-analysis, the safety of this widely used medicine in subjects with Type 2 diabetes is being questioned. Interim analysis of the long-term RECORD (Rosiglitazone Evaluated for Cardiac Outcomes and Regulation of glycaemia in Diabetes) trial, which was designed to determine cardiovascular outcomes, showed that these occurred in 9.1% of control subjects and 9.8% of the rosiglitazone subjects, and these values were not significantly different. Both the meta-analysis and the interim analysis of RECORD have many limitations. In conclusion, it seems that it is possible that rosiglitazone may have a small harmful effect on cardiovascular outcomes. As the effect (if any) is small, this should not be cause for alarm, but rather for further review of how rosiglitazone should be used and for a larger trial to determine the unequivocal effect of rosiglitazone on cardiovascular outcomes.

RATIONALE: Chlamydial lung infection has been associated with asthma in children and adults. However, how chlamydial infection influences the development of immune responses that promote asthma remains unknown. OBJECTIVES: To determine the effect of chlamydial infection at various ages on the development of allergic airway disease (AAD). METHODS: Mouse models of chlamydial lung infection and ovalbumin-induced AAD were established in neonatal and adult BALB/c mice. Neonatal or adult mice were given a chlamydial infection and 6 weeks later were sensitized and subsequently challenged with ovalbumin. Features of AAD and inflammation were compared between uninfected or unsensitized controls. MEASUREMENTS AND MAIN RESULTS: Mild Chlamydia-induced lung disease was observed 10-15 days after infection, as evidenced by increased bacterial numbers and histopathology in the lung and a reduction in weight gain. After 6 weeks, infection and histopathology had resolved and the rate of weight gain had recovered. Neonatal but not adult infection resulted in significant decreases in interleukin-5 production from helper T cells and by the numbers of eosinophils recruited to the lung in response to ovalbumin exposure. Remarkably, the effects of early-life infection were associated with the generation of both type 1 and 2 ovalbumin-specific helper T-cell cytokine and antibody responses. Furthermore, although neonatal infection significantly attenuated eosinophilia, the generation of the mixed T-cell response exacerbated other hallmark features of asthma: mucus hypersecretion and airway hyperresponsiveness. Moreover, infection prolonged the expression of AAD and these effects were restricted to early-life infection. CONCLUSIONS: Early-life chlamydial infection induces a mixed type 1 and 2 T-cell response to antigen, which differentially affects the development of key features of AAD in the adult.

Strain ILE-2T was isolated from an upflow anaerobic sludge bed reactor treating brewery wastewater. The motile, non-sporulating, slightly curved cells (2–4×0.1 μm) stained Gram-negative and grew optimally at 42 °C and pH 7.1 with 0.5 % NaCl. The strain required yeast extract for growth and fermented Casamino acids, peptone, isoleucine, arginine, lysine, alanine, valine, glutamate, histidine, glutamine, methionine, malate, fumarate, glycerol and pyruvate to acetate, propionate and minor amounts of branched-chain fatty acids. Carbohydrates, formate, acetate, propionate, butyrate, isovalerate, methanol, ethanol, 1-propanol, butanol, lactate, succinate, starch, casein, gelatin, xylan and a number of other amino acids were not utilized. The DNA G+C content of strain ILE-2T was 52.7 mol%. 16S rRNA gene sequence analysis revealed that ILE-2T was distantly related to members of the genera Aminobacterium (83 % similarity) and Aminomonas (85 % similarity) in the family Syntrophomonadaceae, order Clostridiales, phylum Firmicutes. On the basis of the results of our polyphasic analysis, strain ILE-2T represents a novel species and genus within the family Syntrophomonadaceae, for which the name Aminiphilus circumscriptus gen. nov., sp. nov. is proposed. The type strain of Aminiphilus circumscriptus is ILE-2T (=DSM 16581T =JCM 14039T).

An aerobic bacterium, designated strain E1HC-02T, was isolated from the decaying leaf litter of a slash pine forest located in southeast Queensland, Australia. Cells of strain E1HC-02T were short irregular rods (0.5–1.0×0.2–0.4 μm) which stained Gram-positive and possessed a cell-wall ultrastructure which appeared to be made of protein subunits. The novel strain grew optimally in 1 % trypticase soy broth (TSB) at 25 °C and at a pH of 9.1. Strain E1HC-02T metabolized a range of carbohydrates, organic acids and amino acids. The G+C content of the DNA was 71±1 mol% as determined by the thermal denaturation method. 16S rRNA gene sequence analysis of strain E1HC-02T showed that it was a member of the family Microbacteriaceae, phylum Actinobacteria. The cell wall contained a type B2β peptidoglycan, the dominant cellular fatty acid was 18 : 1ω7c and the major hydroxy fatty acid was 2-OH 14 : 0. The major menaquinones were MK-8 (76 %) and MK-7 (24 %) and the glycolipids present were disphosphatidylglycerol, phosphatidylglycerol and three unidentified phospholipids. The chemotaxonomic properties of strain E1HC-02T were distinctly different to all of the 17 genera of the family Microbacteriaceae and hence strain E1HC-02T is designated as representing a novel species of a new genus, Frondicola australicus gen. nov., sp. nov. The type strain of the type species is E1HC-02T (=JCM 13598T=DSM 17894T).

Nucleopolyhedrovirus (NPV) has become an integral part of integrated pest management (IPM) in many Australian agricultural and horticultural crops. This is the culmination of years of work conducted by researchers at the Queensland Department of Primary Industries and Fisheries (QDPI&F) and Ag Biotech Australia Pty Ltd. In the early 1970’s researchers at QDPI&F identified and isolated a virus in Helicoverpa armigera populations in the field. This NPV was extensively studied and shown to be highly specific to Helicoverpa and Heliothis species. Further work showed that when used appropriately the virus could be used effectively to manage these insects in crops such as sorghum, cotton, chickpea and sweet corn. A similar virus was first commercially produced in the USA in the 1970’s. This product, Elcar®, was introduced into Australia in the late 1970’s by Shell Chemicals with limited success. A major factor contributing to the poor adoption of Elcar was the concurrent enormous success of the synthetic pyrethroids. The importance of integrated pest management was probably also not widely accepted at that time. Gradual development of insect resistance to synthetic pyrethroids and other synthetic insecticides in Australia and the increased awareness of the importance of IPM meant that researchers once again turned their attentions to environmentally friendly pest management tools such NPV and beneficial insects. In the 1990’s a company called Rhone-Poulenc registered an NPV for use in Australian sorghum, chickpea and cotton. This product, Gemstar®, was imported from the USA. In 2000 Ag Biotech Australia established an in-vivo production facility in Australia to produce commercial volumes of a product similar to the imported product. This product was branded, ViVUS®, and was first registered and sold commercially in Australia in 2003. The initial production of ViVUS used a virus identical to the American product but replicating it in an Australian Helicoverpa species, H. armigera. Subsequent research collaboration between QDPI&F and Ag Biotech reinvigorated interest in the local virus strain. This was purified and the production system adapted to produce it on a commercial scale. This new version of ViVUS, which was branded ViVUS Gold®, was first registered and sold commercially in 2004. Widespread insect resistance to insecticides and a greater understanding of integrated pest management is leading to increased adoption of technologies such NPV in Australian agriculture.

This protocol is used to induce transgenic roots on soybean to study the function of genes required in biological processes of the root. Young seedlings with unfolded cotyledons are infected at the cotyledonary node and/or hypocotyl with Agrobacterium rhizogenes carrying the gene construct to be tested and the infection sites are kept in an environment of high humidity. When the emerged hairy roots can support the plants, the main roots are removed and the transgenic roots can be tested. Using this method, almost 100% of the infected plants form hairy roots within 1 month from the start of the experiments.

Capsular polysaccharide (CPS) was isolated from Acinetobacter baumannii NIPH146, and the following structure of branched pentasaccharide repeating unit was established by sugar analyses along with 1D and 2D NMR spectroscopy: In comparison to most other known capsular polysaccharides of A. baumannii , the CPS studied is neutral and lacks any specific monosaccharide component. The synthesis, assembly and export of this structure could be attributed to genes in a novel capsule biosynthesis gene cluster, designated KL37, which was found in the NIPH146 genome. The CPS of A. baumannii NIPH146 shares the α- d -Gal p -(1→6)-β- d -Glc p -(1→3)- d -Gal p NAc-(1→ trisaccharide fragment with the CPS units of several A. baumannii strains, including ATCC 17978 and LUH 5537 that carry the KL3 and KL22 gene clusters, respectively. KL37 contains two genes for glycosyltransferases that are related to two glycosyltransferase genes present in both KL3 and KL22, and the encoded proteins could be tentatively assigned to linkages between sugars in the CPS repeat.

An Acinetobacter baumannii global clone 1 (GC1) isolate was found to carry a novel capsule biosynthesis gene cluster, designated KL12. KL12 contains genes predicted to be involved in the synthesis of simple sugars, as well as ones for N-acetyl-L-fucosamine (L-FucpNAc) and N-acetyl-D-fucosamine (D-FucpNAc). It also contains a module of 10 genes, 6 of which are required for 5,7-di-N-acetyl-legionaminic acid synthesis. Analysis of the composition of the capsule revealed the presence of N-acetyl-D-galactosamine, L-FucpNAc and D-FucpNAc, confirming the role of fnlABC and fnr/gdr genes in the synthesis of L-FucpNAc and D-FucpNAc, respectively. A non-2-ulosonic acid, shown to be 5,7-diacetamido-3,5,7,9-tetradeoxy-L-glycero-L-altro-non-2-ulosonic acid, was also detected. This sugar has not previously been recovered from biological source, and was designated 5,7-di-N-acetyl-acinetaminic acid (Aci5Ac7Ac). Proteins encoded by novel genes, named aciABCD, were predicted to be involved in the conversion of 5,7-di-N-acetyl-legionaminic acid to Aci5Ac7Ac. A pathway for 5,7-di-N-acetyl-8-epilegionaminic acid biosynthesis was also proposed. In available A. baumannii genomes, genes for the synthesis of 5,7-di-N-acetyl-acinetaminic acid were only detected in two closely related capsule gene clusters, KL12 and KL13, which differ only in the wzy gene. KL12 and KL13 are carried by isolates belonging to clinically important clonal groups, GC1, GC2 and ST25. Genes for the synthesis of N-acyl derivatives of legionaminic acid were also found in 10 further A. baumannii capsule gene clusters, and three carried additional genes for production of 5,7-di-N-acetyl- 8-epilegionaminic acid.

Acinetobacter baumannii isolate A1 was recovered in the United Kingdom in 1982 and belongs to global clone 1 (GC1). Here, we present its complete 3.91-Mbp genome sequence, generated via a combination of short-read sequencing (Illumina), long-read sequencing (PacBio), and manual finishing.

Streptococcus pneumoniae is a potentially deadly human pathogen associated with high morbidity, mortality and global economic burden. The universally used bacterial genotyping methods are multilocus sequence typing and pulsed field gel electrophoresis. However, another highly discriminatory, rapid and less expensive genotyping technique,multilocus variable number of tandem repeat analysis (MLVA), has been developed. Unfortunately, no universal MLVA protocol exists, and some MLVA protocols do not amplify certain loci for all pneumococcal serotypes, leaving genotyping profiles incomplete. A number of other genotyping or characterization methods have been developed and will be discussed. This review examines the various protocols for genotyping S. pneumoniae and highlights the current direction technology and research is heading to understand this bacterium.

The crystal structure of the H. orenii glycosidase was determined by molecular replacement and refined at 1.10 Å resolution., A gene from the heterotrophic, halothermophilic marine bacterium Halothermothrix orenii has been cloned and overexpressed in Escherichia coli. This gene encodes the only glycoside hydrolase of family 43 (GH43) produced by H. orenii. The crystal structure of the H. orenii glycosidase was determined by molecular replacement and refined at 1.10 Å resolution. As for other GH43 members, the enzyme folds as a five-bladed β-propeller. The structure features a metal-binding site on the propeller axis, near the active site. Based on thermal denaturation data, the H. orenii glycosidase depends on divalent cations in combination with high salt for optimal thermal stability against unfolding. A maximum melting temperature of 76°C was observed in the presence of 4 M NaCl and Mn2+ at pH 6.5. The gene encoding the H. orenii GH43 enzyme has previously been annotated as a putative α-l-arabinofuranosidase. Activity was detected with p-nitrophenyl-α-l-arabinofuranoside as a substrate, and therefore the name HoAraf43 was suggested for the enzyme. In agreement with the conditions for optimal thermal stability against unfolding, the highest arabinofuranosidase activity was obtained in the presence of 4 M NaCl and Mn2+ at pH 6.5, giving a specific activity of 20–36 µmol min−1 mg−1. The active site is structurally distinct from those of other GH43 members, including arabinanases, arabinofuranosidases and xylanases. This probably reflects the special requirements for degrading the unique biomass available in highly saline aqueous ecosystems, such as halophilic algae and halophytes. The amino-acid distribution of HoAraf43 has similarities to those of mesophiles, thermophiles and halophiles, but also has unique features, for example more hydrophobic amino acids on the surface and fewer buried charged residues.