Plant hosts of the phytoplasmas and rickettsia-like-organisms associated with strawberry lethal yellows and green petal diseases

Candidatus Phytoplasma australiense (Ca. P. australiense) is associated with the plant diseases strawberry lethal yellows (SLY), strawberry green petal (SGP), papaya dieback (PDB), Australian grapevine yellows (AGY) and Phormium yellow leaf (PYL; New Zealand). Strawberry lethal yellows disease is also associated with a rickettsia-like-organism (RLO) or infrequently with the tomato big bud (TBB) phytoplasma, the latter being associated with a wide range of plant diseases throughout Australia. In contrast, the RLO has been identified only in association with SLY disease, and Ca. P. australiense has been detected only in a limited number of plant host species. The aim of this study was to identify plant hosts that are possible reservoirs of Ca. P. australiense and the SLY RLO. Thirty-one plant species from south-east Queensland were observed with disease between 2001 and 2003 and, of these, 18 species tested positive using phytoplasma-specific primers. The RLO was detected in diseased Jacksonia scoparia and Modiola caroliniana samples collected at Stanthorpe. The TBB phytoplasma was detected in 16 different plant species and Ca. P. australiense Australian grapevine yellows strain was detected in six species. The TBB phytoplasma was detected in plants collected at Nambour, Stanthorpe, Warwick and Brisbane. Ca. P. australiense was detected in plants collected at Nambour, Stanthorpe, Gatton and Allora. All four phytoplasmas were detected in diseased Gomphocarpus physocarpus plants collected at Toowoomba, Allora, Nambour and Gatton. These results indicated that the vector(s) of Ca. P. australiense are distributed throughout south-east Queensland and the diversity of phytoplasmas detected in G. physocarpus suggests it is a feeding source for phytoplasma insect vectors or it has a broad susceptibility to a range of phytoplasmas.

Rickettsia-like-organisms and phytoplasmas associated with diseases in Australian strawberries

Strawberry lethal yellows (SLY) disease in Australia is associated with the phytoplasmas Candidatus Phytoplasma australiense and tomato big bud, and a rickettsia-like-organism (RLO). Ca. P. australiense is also associated with strawberry green petal (SGP) disease. This study investigated the strength of the association of the different agents with SLY disease. We also documented the location of SLY or SGP plants, and measured whether they were RLO or phytoplasma positive. Symptomatic strawberry plants collected from south-east Queensland (Australia) between January 2000 and October 2002 were screened by PCR for both phytoplasmas and the RLO. Two previously unreported disease symptoms termed severe fruit distortion (SFD) and strawberry leaves from fruit (SLF) were observed during this study but there was no clear association between these symptoms and phytoplasmas or the RLO. Only two SGP diseased plants were observed and collected, compared with 363 plants with SLY disease symptoms. Of the 363 SLY samples, 117 tested positive for the RLO, 67 tested positive for Ca. P. australiense AGY strain and 11 plants tested positive for Ca. P. australiense PYL variant strain. On runner production farms at Stanthorpe, Queensland the RLO was detected in SLY diseased plants more frequently than for the phytoplasmas. On fruit production farms on the Sunshine Coast, Queensland, Ca. P. australiense was detected in SLY disease plants more frequently than the RLO.

Development of a New Zealand database of plant virus and virus-like organisms.

The recent 8th Australasian plant virology workshop in Rotorua, New Zealand, discussed the development of a New Zealand database of plant virus and virus-like organisms. Key points of discussion included: (i) the purpose of such a database; (ii) who would benefit from the information in a database; (iii) the scope of a database and its associated collections; (iv) database information and format; and (v) potential funding of such a database. From the workshop and further research, we conclude that the preservation and verification of specimens within the collections and the development of a New Zealand database of plant virus and virus-like organisms is essential. Such a collection will help to fulfil statutory requirements in New Zealand and assist in fulfilling international obligations under the International Plant Protection Convention. Sustaining such a database will assist New Zealand virologists and statutory bodies to undertake scientifically sound research. Establishing reliable records and an interactive database will help to ensure accurate and timely diagnoses of diseases caused by plant viruses and virus-like organisms. Detection of new incursions and their diagnosis will be further enhanced by the use of such reference collections and their associated database. Connecting and associating this information to similar overseas databases would assist international collaborations and allow access to the latest taxonomic and diagnostic resources. Associated scientists working in the areas of plant breeding, export phytosanitary assurance and in the area of the conservation estate would also benefit from access to verified specimens of plant viruses and virus-like organisms. We conclude that funding of a New Zealand database of virus and virus-like organisms and its associated collections should be based partly on Crown funds, as it is a nationally significant biological resource.

Biodegradation of 1080: Testing soils in south-eastern Australia for sodium fluoroacetate-degrading micro-organisms

Sodium fluoroacetate (1080) is a vertebrate poison commonly used for the control of vertebrate pests in Australia. Long-term environmental persistence of 1080 from baiting operations has likely nontarget species and environmental impacts and is a matter of public concern. Defluorinating micro-organisms have been detected in soils of Western and central Australia, and Queensland, but not in south-eastern Australia. The presence or absence of defluorinating micro-organisms in soils from south-eastern Australia will assist in determining whether long-term environmental persistence of 1080 is or is not occurring. Soils from the Central West Slopes and Plains and Central Tablelands of New South Wales were sampled to investigate the presence and capability of 1080 defluorinating soil micro-organisms. Thirty-one species of micro-organisms were isolated from soils from each site after 10 days incubation in a 20 mM 1080 solution. Of these, 13 isolates showed measurable defluorinating ability when grown in a 1080 and sterile soil suspension. Two species, the bacteria Micromonospora, and the actinomycete Streptosporangium, have not been previously reported for their defluorinating ability. These results indicate that defluorinating micro-organisms are present in soils in south-eastern Australia, which adds weight to other studies that found that 1080 is subject to microbiological degradative processes following removal from the bait substrate. Soil micro-organism defluorination, in combination with physical breakdown and uptake by plants, indicates that fluoroacetate in soils and natural water ways is unlikely to persist. This has implications for the better informed use of 1080 in pest animal management programmes in south-eastern Australia.

Volatile components associated with bacterial spoilage of tropical prawns

Analysis of headspace volatiles by gas chromatography/mass spectrometry from king (Penaeus plebejus), banana (P. merguiensis), tiger (P. esculentus/semisulcatus) and greasy (Metapenaeus bennettae) prawns stored in ice or ice slurry, which is effectively an environment of low oxygen tension, indicated the presence of amines at the early stages of storage (less than 8 days) irrespective of the nature of the storage media. Esters were more prevalent in prawns stored on ice (normal oxygen conditions) at the latter stages of storage (more than 8 days) and were only produced by Pseudomonas fragi, whereas sulphides and amines occurred whether the predominant spoilage organism was Ps.fragi or Shewanella putrefaciens. The free amino acid profiles of banana and king prawns were high in arginine (12–14%) and low in cysteine (0.1–0.17%) and methionine (0.1–0.2%). Filter sterilised raw banana prawn broth inoculated with a total of 15 cultures of Ps. fragi and S. putrefaciens and incubated for two weeks at 5°C, showed the presence of 17 major compounds in the headspace volatiles analysed using gas chromatography/mass spectrometry (GC/MS). These were mainly amines, sulphides, ketones and esters. Principal Component Analysis of the results for the comparative levels of the volatiles produced by pure cultures, inoculated into sterile prawn broth, indicated three subgroupings of the organisms; I, Ps. fragi from a particular geographic location; II, S. putrefaciens from another geographic location; and III, a mixture of Ps. fragi and S. putrefaciens from different geographic locations. The sensory impression created by the cultures was strongly related to the chemical profile as determined by GC/MS. Organisms, even within the same subgrouping classified as identical by the usual tests, produced a different range of volatiles in the same uniform substrate.

Coliforms in processed mango: Significance and control

The aims of this investigation were to enumerate coliforms in fresh mangoes, puree, cheeks, and cheeks-in-puree in order to determine the source of these organisms in the processed products, to determine methods for their control, and to identify coliforms isolated from cheeks-in-puree to determine whether they have any public health significance. Product from four processors was tested on two occasions. The retail packs of cheeks-in-puree having the highest coliform counts were those in which raw puree was added to the cheeks. Coliform counts in these samples ranged between 1.4 × 103 and 5.4 × 104 cfu/g. Pasteurisation reduced the coliform count of raw puree to < 5 cfu/g. Forty-seven percent of the 73 colonies, isolated as coliforms on the basis of their colony morphology on violet red bile agar, were identified as Klebsiella pneumoniae using the ATB 32E Identification System. Klebsiella strains were tested for growth at 10 °C, faecal coliform response, and fermentation of -melizitose, to differentiate the three phenotypically similar strains, K. pneumoniae, K. terrigena and K planticola. Results indicated that 41% of K. pneumoniae isolates gave reactions typical of K. pneumoniae. A further 44% of strains gave an atypical reaction pattern for these tests and were designated ‘psychrotrophic’ K. pneumoniae. Klebsiella pneumoniae counts of between 2.1 × 103 and 4.9 × 104 cfu/g were predicted to occur in the retail packs of mango cheeks-in-puree produced by the processors who constituted this product with raw puree. In view of the opportunistic pathogenic nature of K. pneumoniae, its presence in these products is considered undesirable and steps, such as pasteurisation of puree, should be taken in order to inactivate it

Microbiological status of piggery effluent from 13 piggeries in the south east Queensland region of Australia

Aims: To assist in the development of safe piggery effluent re-use guidelines by determining the level of selected pathogens and indicator organisms in the effluent ponds of 13 south-east Queensland piggeries. Methods and Results: The numbers of thermotolerant coliforms, Campylobacter jejuni/coli, Erysipelothrix rhusiopathiae, Escherichia coli, Salmonella and rotavirus were determined in 29 samples derived from the 13 piggeries. The study demonstrated that the 13 final effluent ponds contained an average of 1Æ2 · 105 colony-forming units (CFU) 100 ml)1 of thermotolerant coliforms and 1Æ03 · 105 CFU 100 ml)1 of E. coli. The Campylobacter level varied from none detectable (two of 13 piggeries) to a maximum of 930 most probable number (MPN) 100 ml)1 (two of 13 piggeries). Salmonella was detected in the final ponds of only four of the 13 piggeries and then only at a low level (highest level being 51 MPN 100 ml)1). No rotavirus and no Erysip. rhusiopathiae were detected. The average log10 reductions across the ponding systems to the final irrigation pond were 1Æ77 for thermotolerant coliforms, 1Æ71 for E. coli and 1Æ04 for Campylobacter. Conclusions: This study has provided a baseline knowledge on the levels of indicator organisms and selected pathogens in piggery effluent. Significance and Impact of the Study: The knowledge gained in this study will assist in the development of guidelines to ensure the safe and sustainable re-use of piggery effluent.

Investigation and application of methods for enumerating heterotrophs and Escherichia coli in the air within piggery sheds

Aims: To investigate methods for the recovery of airborne bacteria within pig sheds and to then use the appropriate methods to determine the levels of heterotrophs and Escherichia coli in the air within sheds. Methods and Results: AGI-30 impingers and a six-stage Andersen multi-stage sampler (AMS) were used for the collection of aerosols. Betaine and catalase were added to impinger collection fluid and the agar plates used in the AMS. Suitable media for enumerating E. coli with the Andersen sampler were also evaluated. The addition of betaine and catalase gave no marked increase in the recovery of heterotrophs or E. coli. No marked differences were found in the media used for enumeration of E. coli. The levels of heterotrophs and E. coli in three piggeries, during normal pig activities, were 2Æ2 · 105 and 21 CFU m)3 respectively. Conclusions: The failure of the additives to improve the recovery of either heterotrophs or E. coli suggests that these organisms are not stressed in the piggery environment. The levels of heterotrophs in the air inside the three Queensland piggeries investigated are consistent with those previously reported in other studies. Flushing with ponded effluent had no marked or consistent effect on the heterotroph or E. coli levels. Significance and Impact of the Study: Our work suggests that levels of airborne heterotrophs and E. coli inside pig sheds have no strong link with effluent flushing. It would seem unlikely that any single management activity within a pig shed has a dominant influence on levels of airborne heterotrophs and E. coli

Reclassification of [Pasteurella] trehalosi as Bibersteinia trehalosi gen. nov., comb. nov.

[Pasteurella] trehalosi is an important pathogen of sheep, being primarily associated with serious systemic infections in lambs but also having an association with pneumonia. The aim of the present investigation was to characterize a broad collection of strains tentatively identified as [P.] trehalosi in order to reclassify and rename this taxon to support improvements in our understanding of the pathogenesis and epidemiology of this important organism. The type strain for [P.] trehalosi, strain NCTC 10370T, was included along with 42 field isolates from sheep (21), cattle (14), goats (1), roe deer (3) and unknown sources (3). An extended phenotypic characterization was performed on all 43 strains. Amplified fragment length polymorphism (AFLP) was also performed on the isolates. Two of the field isolates were subjected to 16S rRNA gene sequencing. These sequences, along with five existing sequences for [P.] trehalosi strains and 12 sequences for other taxa in the family Pasteurellaceae, were subjected to a phylogenetic analysis. All the isolates and the reference strains were identified as [P.] trehalosi. A total of 17 out of 22 ovine isolates produced acid from all glycosides, while only four out of 14 bovine isolates produced acid from all glycosides. All 22 ovine isolates were haemolytic and CAMP-positive, while no other isolate was haemolytic and only two bovine isolates were CAMP-positive. Nineteen AFLP types were found within the [P.] trehalosi isolates. All [P.] trehalosi isolates shared at least 70% similarity in AFLP patterns. The largest AFLP type included the type strain and 7 ovine field isolates. Phylogenetic analysis indicated that the seven strains studied (two field isolates and the five serovar reference strains) are closely related, with 98.6% or higher 16S rRNA gene sequence similarity. As both genotypic and phenotypic testing support the separate and distinct nature of these organisms, we propose the transfer of [P.] trehalosi to a new genus, Bibersteinia, as Bibersteinia trehalosi comb. nov. The type strain is NCTC 10370T (=ATCC 29703T). Bibersteinia trehalosi can be distinguished from the existing genera of the family by the observation of only nine characteristics; catalase, porphyrin, urease, indole, phosphatase, acid from dulcitol, (+)-D-galactose, (+)-D mannose and (+)-D-trehalose.

Developments in controlled green-water larval culture technologies for estuarine fishes in Queensland, Australia and elsewhere

Since 1989, researchers with the Department of Primary Industries and Fisheries (DPI&F) in Queensland, Australia, have successfully used controlled low-water exchange green-water cultures to rear the larvae of estuarine fishes and crustaceans through to metamorphosis. High survivals and excellent fry condition have been achieved for several commercially important endemic species produced for various projects. They include barramundi or sea bass, Lates calcarifer, Australian bass, Macquaria novemaculeata, dusky flathead, Platycephalus fuscus, sand whiting, Sillago ciliata, red sea bream or snapper, Pagrus auratus, banana prawn, Fenneropenaeus merguiensis, and others. The consistent success of our standardised and relatively simple approach at different localities has led to it being incorporated into general fingerling production practices at several establishments in Australia. Although post-metamorphosis rearing methods have differed for each species investigated, due to various biological and behavioural traits and project requirements, these larval rearing methods have been successful with few species-specific modifications. Initially modelled on the Taiwanese approach to rearing Penaeids in aerated low-water exchange cultures, the approach similarly appears to rely on a beneficial assemblage of micro-organisms. Conceptually, these micro-organisms may include a mixture of the air-borne primary invaders of pure phytoplankton cultures when exposed to outdoor conditions. Whilst this would vary with different sites, our experiences with these methods have consistently been favourable. Mass microalgal cultures with eco-physiological youth are used to regularly augment larval fish cultures so that rearing conditions simulate an exponential growth-phase microalgal bloom. Moderate to heavy aeration prevents settlement of particulate matter and encourages aerobic bacterial decomposition of wastes. The green-water larval rearing approach described herein has demonstrated high practical utility in research and commercial applications, and has greatly simplified marine finfish hatchery operations whilst generally lifting production capacities for metamorphosed fry in Australia. Its potential uses in areas of aquaculture other than larviculture are also discussed.

Gene interactions constrain the course of evolution of phosphine resistance in the lesser grain borer, Rhyzopertha dominica

Phosphine, a widely used fumigant for the protection of stored grain from insect pests, kills organisms indirectly by inducing oxidative stress. High levels of heritable resistance to phosphine in the insect pest of stored grain, Rhyzopertha dominica have been detected in Asia, Australia and South America. In order to understand the evolution of phosphine resistance and to isolate the responsible genes, we have undertaken genetic linkage analysis of fully sensitive (QRD14), moderately resistant (QRD369) and highly resistant (QRD569) strains of R. dominica collected in Australia. We previously determined that two loci, rph1 and rph2, confer high-level resistance on strain QRD569, which was collected in 1997. We have now confirmed that rph1 is responsible for the moderate resistance of strain QRD369, which was collected in 1990, and is shared with a highly resistant strain from the same geographical region, QRD569. In contrast, rph2 by itself confers only very weak resistance, either as a heterozygote or as a homozygote and was not discovered in the field until weak resistance (probably due to rph1) had become ubiquitous. Thus, high-level resistance against phosphine has evolved via stepwise acquisition of resistance alleles, first at rph1 and thereafter at rph2. The semi-dominance of rph2 together with the synergistic interaction between rph1 and rph2 would have led to rapid selection for homozygosity. A lack of visible fitness cost associated with alleles at either locus suggests that the resistance phenotype will persist in the field.

Productivity and health effects of anaplasmosis and babesiosis on Bos indicus cattle and their crosses, and the effects of differing intensity of tick control in Australia

Tick fever is an important disease of cattle where Rhipicephalus (Boophilus) microplus acts as a vector for the three causal organisms Babesia bovis, Babesia bigemina and Anaplasma marginale. Bos indicus cattle and their crosses are more resistant to the clinical effects of infection with B. bovis and B. bigemina than are Bos taurus cattle. Resistance is not complete, however, and herds of B. indicus-cross cattle are still at risk of babesiosis in environments where exposure to B. bovis is light in most years but occasionally high. The susceptibility of B. indicus cattle and their crosses to infection with A. marginale is similar to that of B. taurus cattle. In herds of B. indicus cattle and their crosses the infection rate of Babesia spp. and A. marginale is lowered because fewer ticks are likely to attach per day due to reduced numbers of ticks in the field (long-term effect on population, arising from high host resistance) and because a smaller proportion of ticks that do develop to feed on infected cattle will in turn be infected (due to lower parasitaemia). As a consequence, herds of B. indicus cattle are less likely than herds of B. taurus cattle to have high levels of population immunity to babesiosis or anaplasmosis. The effects of acaricide application on the probability of clinical disease due to anaplasmosis and babesiosis are unpredictable and dependent on the prevalence of infection in ticks and in cattle at the time of application. Attempting to manipulate population immunity through the toleration of specific threshold numbers of ticks with the aim of controlling tick fever is not reliable and the justification for acaricide application should be for the control of ticks rather than for tick fever. Vaccination of B. indicus cattle and their crosses is advisable in all areas where ticks exist, although vaccination against B. bigemina is probably not essential in pure B. indicus animals.

Detection of Helicobacter pullorum in meat chickens in Australia

The bacterial genus Helicobacter is a member of the Campylobacteriales in the Epsilonproteobacteria subphylum, and is comprised of organisms that are morphologically similar to Campylobacter. The term ‘campylobacteria’ is used to encompass the genera Campylobacter, Arcobacter, Helicobacter and Anaerobiospirillum. Helicobacter was separated from the genus Campylobacter in 1989. Helicobacter spp have been isolated from gastric tissue or intestinal contents of humans and a wide range of animal species, with some associated with disease...

Impacts of pesticides and fertilisers on soil

The effects of inorganic amendments (fertilisers and pesticides) on soil biota that are reported in the scientific literature are, to say the least, variable. Though there is clear evidence that certain products can have significant impacts, the effects can be positive or negative. This is not surprising when you consider the number of organisms and amount of different functional groups, the number of products and various rates at which they could be applied, the methods of application and the environmental differences that occur in soil at a micro scale (within centimetres) in a paddock, let alone between paddocks, farms, catchments, regions etc. It therefore becomes extremely difficult to draw definitive conclusions from the reported results in order to summarise the impacts of these inputs. Several research trials and review papers have been published on this subject and most similarly conclude that the implications of many of the effects are still uncertain.