Engineered detoxification confers resistance against a pathogenic bacterium

We generated transgenic sugarcane plants that express an albicidin detoxifying gene (albD), which was cloned from a bacterium that provides biocontrol against leaf scald disease. Plants with albicidin detoxification capacity equivalent to 1-10 ng of AlbD enzyme per mg of leaf protein did not develop chlorotic disease symptoms in inoculated leaves, whereas all untransformed control plants developed severe symptoms. Transgenic lines with high AlbD activity in young stems were also protected against systemic multiplication of the pathogen, which is the precursor to economic disease. We have shown that genetic modification to express a toxin-resistance gene can confer resistance to both disease symptoms and multiplication of a toxigenic pathogen in its host.

Structure of the tegument and ectocommensal microorganisms of Gyliauchen nahaensis (Digenea : Gyliauchenidae), an inhabitant of herbivorous fish of the Great Barrier Reef, Australia

The ultrastructure of the tegument and tegument-associated microorganisms of the gyliauchenid digenean Gyliauchen nahaensis is described by transmission and scanning electron microscopy. The tegument is devoid of surface spines and is characterized by a moderately folded apical membrane, abundant vesicles, basal mitochondria, a folded basal plasma membrane, and a thick basal matrix. Microorganisms form a dense biofilm on the tegument of the posterodorsal surface and the excretory papilla. At least 7 microbial morphotypes were identified, including eubacteria, spirochaetes, and nanobacteria.

First published record of the pathogenic monogenean parasite Neobenenenia melleni (Capsalidae) from Australia

The monogenean Neobenedenia melleni (Mac- Callum, 1927) Yamaguti 1963 is a well-known and virulent pathogen in culture conditions recorded from the skin of many teleost fish species worldwide. Until now, N. melleni has not been reported from wild or cultured fish in Australian waters. This study documents a recent outbreak of N. melleni that occurred on Lates calcarifer (barramundi) cultivated in sea cages in Hinchinbrook Channel between Hinchinbrook Island and mainland Queensland, Australia, which resulted in the loss of 200 000 fish (50 tonnes). The origin of this outbreak is unclear because N. melleni has not been recorded from any wild host species in Australia and strict quarantine regulations exclude the possibility of its introduction on imported fish. We propose that N. melleni occurs naturally on wild populations of some teleost species in Australian waters and that the few surveys of wild fish conducted along the eastcoast have failed to report this species. The possibility that uncharacteristically low water temperatures led to the outbreak is discussed.

Saprophytic microorganisms with potential for biological control of Botrytis cinerea on Geraldton waxflower flowers

Saprophytic bacteria, yeasts and filamentous fungi were isolated from Geraldton waxflower flowers and screened to identify potential antagonism towards Botrytis cinerea. Isolates from other sources (e.g. avocado) were also tested. Isolates were initially screened in vitro for inhibition of B. cinerea conidial germination, germ tube elongation and mycelial growth. The most antagonistic bacteria, yeasts and fungi were selected for further testing on detached waxflower flowers. Conidia of the pathogen were mixed with conidia or cells of the selected antagonists, co-inoculated onto waxflower flowers, and the flowers were sealed in glass jars and incubated at 20 degreesC. The number of days required for the pathogen to cause flower abscission was determined. The most antagonistic bacterial isolate, Pseudomonas sp. 677, significantly reduced conidial germination and retarded germ tube elongation of B. cinerea. None of the yeast or fungal isolates tested was found to significantly reduce conidial germination or retard germ tube elongation, but several significantly inhibited growth of B. cinerea. Fusarium sp., Epicoccum sp. and Trichoderma spp. were the most antagonistic of these isolates. Of the isolates tested on waxflower, Pseudomonas sp. 677 was highly antagonistic towards B. cinerea and delayed waxflower abscission by about 3 days. Trichoderma harzianum also significantly delayed flower abscission. However, as with most of the fungal antagonists used, inoculation of waxflower flowers with this isolate resulted in unsightly mycelial growth.

Potential of Cactoblastis cactorum as a vector for fungi pathogenic to pricklypear, Opuntia inermis

In Australia, fungi associated with larvae of the biological control agent Cactoblastis cactorum may contribute to the control of the exotic weed pricklypear (Opuntia inermis), C, cactorum larvae were assessed for their ability to vector pathogenic fungi into O, inermis by the infestation of larvae with fungal suspensions. Six fungal isolates caused disease after being carried into the host on external surfaces of larvae, and propagules of one isolate (UQ5109) initiated disease after being transferred from the cladode epidermis into the host by larvae feeding on the plant. Scanning electron microscopy revealed extensive hyphal growth on the external surfaces of larvae infested with several of the isolates. Fungi isolated from field-grown O, inermis cladodes were tested for pathogenicity to this plant in an in vivo plant assay. In total, 152 isolates were screened, 22 of which infected the host in pathogenicity tests. Only 1 (UQ5115) infected undamaged host tissue, whereas the remainder required the host to be wounded before infection could proceed. The majority of isolates were only weakly pathogenic, even when inoculated via wounds, suggesting that most were either saprophytes or weak parasites. This study demonstrates that it is possible for larvae of C, cactorum to transmit fungal pathogens into O, inermis tissue and it has provided a sound basis for future field work to determine the contribution that fungi make to the control of O. inermis, (C) 2001 Academic Press.

Pathogenic specialisation within Colletotrichum trifolii in Australia, and lucerne cultivar reactions to all known Australian pathotypes

Anthracnose and crown rot, caused by Colletotrichum trifolii, are serious diseases of lucerne (Medicago saliva L.) in humid regions of the world. A race survey was conducted by inoculating individual lucerne clones (genotypes) with C. trifolii isolates collected from a range of Medicago hosts, locations, and years in south-eastern Queensland. This survey revealed for the first time in Australia the presence of race 2 (virulence on anthracnose resistance gene An I) and the first world report of race 4 (virulence on An(2)). A collection of North American race I and race 2 C. trifolii isolates, when inoculated onto the Australian differential clones, gave responses that were in agreement with their North American reactions. A RAPD analysis was conducted on 9 Australian C. trifolii isolates including races 1, 2, and 4; two C. destructivum and one C. gloeosporioides isolate were included as known outliers. For the C. trifolii isolates, 94.6% similarity was found regardless of host origin or race, compared with 2.2% similarity between this group and the C. gloeosporioides and C. destructivum isolates, confirming that the new races belong to C. trifolii. Currently, it is hypothesised that only plants carrying genes An, and An2 are resistant to the 3 races. Of 22 cultivars screened against the 3 races, only UQL-1, Hallmark, and Pioneer 54Q53 had >30% of plants resistant to the 3 races in separate screenings. The research highlights the need to find new sources of resistance to C. trifolii in lucerne.

Combining nonthermal technologies to control foodborne microorganisms

Novel nonthermal processes, such as high hydrostatic pressure (HHP), pulsed electric fields (PEFs), ionizing radiation and ultrasonication, are able to inactivate microorganisms at ambient or sublethal temperatures. Many of these processes require very high treatment intensities, however, to achieve adequate microbial destruction in low-acid foods. Combining nonthermal processes with conventional preservation methods enhances their antimicrobial effect so that lower process intensities can be used. Combining two or more nonthermal processes can also enhance microbial inactivation and allow the use of lower individual treatment intensities. For conventional preservation treatments, optimal microbial control is achieved through the hurdle concept, with synergistic effects resulting from different components of the microbial cell being targeted simultaneously. The mechanisms of inactivation by nonthermal processes are still unclear; thus, the bases of synergistic combinations remain speculative. This paper reviews literature on the antimicrobial efficiencies of nonthermal processes combined with conventional and novel nonthermal technologies. Where possible, the proposed mechanisms of synergy is mentioned. (C) 2003 Elsevier Science B.V. All rights reserved.

A Sco homologue plays a role in defence against oxidative stress in pathogenic Neisseria

Sco proteins are found in mitochondria and in a variety of oxidase positive bacteria. Although Sco is required for the formation of the Cu-A centre in a cytochrome oxidase of the aa(3) type, it was observed that oxidases with a Cu-A centre are not present in many bacteria that contain a Sco homologue. Two bacteria of this type are the pathogens Neisseria meningitidis and Neisseria gonorrhoeae. The sco genes of N. gonorrhoeae strain 1291 and N. meningitidis strain MC58 were cloned, inactivated by inserting a kanamycin resistance cassette and used to make knockout mutants by allelic exchange. Both N. gonorrhoeae and N. meningitidis sco mutants were highly sensitive to oxidative killing by paraquat, indicating that Sco is involved in protection against oxidative stress in these bacteria. (C) 2003 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.

Isolation and characterization of integron-containing bacteria without antibiotic selection

The emergence of antibiotic resistance among pathogenic and commensal bacteria has become a serious problem worldwide. The use and overuse of antibiotics in a number of settings are contributing to the development of antibiotic-resistant microorganisms. The class 1 and 2 integrase genes (intI1 and intI2, respectively) were identified in mixed bacterial cultures enriched from bovine feces by growth in buffered peptone water (BPW) followed by integrase-specific PCR. Integrase-positive bacterial colonies from the enrichment cultures were then isolated by using hydrophobic grid membrane filters and integrase-specific gene probes. Bacterial clones isolated by this technique were then confirmed to carry integrons by further testing by PCR and DNA sequencing. Integron-associated antibiotic resistance genes were detected in bacteria such as Escherichia coli, Aeromonas spp., Proteus spp., Morganella morganii, Shewanella spp., and urea-positive Providencia stuartii isolates from bovine fecal samples without the use of selective enrichment media containing antibiotics. Streptomycin and trimethoprim resistance were commonly associated with integrons. The advantages conferred by this methodology are that a wide variety of integron-containing bacteria may be simultaneously cultured in BPW enrichments and culture biases due to antibiotic selection can be avoided. Rapid and efficient identification, isolation, and characterization of antibiotic resistance-associated integrons are possible by this protocol. These methods will facilitate greater understanding of the factors that contribute to the presence and transfer of integron-associated antibiotic resistance genes in bacterial isolates from red meat production animals.

Azurin of pathogenic Neisseria spp. is involved in defense against hydrogen peroxide and survival within cervical epithelial cells

Laz, a lipid-modified azurin of the human pathogens Neisseria gonorrhoeae and Neisseria meningitidis, is involved in defense against oxidative stress and copper toxicity; laz mutant strains are hypersensitive to hydrogen peroxide and copper. The N. gonorrhoeae laz mutant also has decreased survival in an ex vivo primary human ectocervical epithelial assay.

Identification and characterization of non-pathogenic Fusarium oxysporum capable of increasing and decreasing Fusarium wilt severity

Fusarium wilt of banana is a potentially devastating disease throughout the world. Options for control of the causal organism, Fusarium oxysporum f.sp. cubense (Foc) are limited. Suppressive soil sites have previously been identified where, despite the presence of Foc, Fusarium wilt does not develop. In order to understand some aspects of this disease suppression, endophytic Fusarium oxysporum isolates were obtained from banana roots. These isolates were genetically characterized and compared with an isolate of Fusarium oxysporum previously identified as being capable of suppressing Fusarium wilt of banana in glasshouse trials. Three additional isolates were selected for glasshouse trials to assess suppression of Fusarium wilt in two different cultivars of banana, Cavendish and Lady Finger. One isolate (BRIP 29089) was identified as a potential biocontrol organism, reducing the disease severity of Fusarium wilt in Lady Finger and Cavendish cultivars. Interestingly, one isolate (BRIP 45952) increased Fusarium wilt disease severity on Cavendish. The implications of an isolate of Fusarium oxysporum, non-pathogenic on banana, increasing disease severity and the potential role of non-pathogenic isolates of Fusarium oxysporum in disease complexes are discussed. (c) 2006 Published by Elsevier Ltd on behalf of The British Mycological Society.