Virulence of the Nine Serovar Reference Strains of Haemophilus paragallinarum

The virulence of the reference strains of the nine currently recognized Kume serovars of Haemophilus paragallinarum was investigated. The capacity of the H. paragallinarum strains to cause the typical clinical signs of upper respiratory tract disease associated with infectious coryza in unvaccinated, nasal-challenged chickens was assessed. Differences in virulence were assessed by means of a standardized scoring system for clinical signs. All nine strains were pathogenic to chickens, producing typical clinical signs of infectious coryza. The highest clinical signs score was obtained for serovar C-1 (1.72), while the lowest clinical signs score was obtained for serovar C-4 (0.32). Our results indicate that virulence differences exist among the serovars of H. paragallinarum.

Selection and characterisation of two attenuated vaccine lines of Eimeria tenella in Australia

Objective To attenuate two strains of Eimeria tenella by selecting for precocious development and evaluate the strains in characterisation trials and by field evaluation, to choose one precocious line for incorporation into an Australian live coccidiosis vaccine for poultry. Design Two strains from non-commercial flocks were passaged through chickens while selecting for precocious development. Each strain was characterised for drug sensitivity, pathogenicity, protection against homologous and heterologous challenge, and oocyst output in replicated experiments in which the experimental unit was a cage of three birds. Oocyst output and/or body weight gain data collected over a 10 to 12 day period following final inoculation were measured. Feed conversion ratios were also calculated where possible. Results Fifteen passages resulted in prepatent periods reduced by 24 h for the Redlands strain (from 144 h to 120 h)and 23 h for the Darryl strain (from 139 h to 116 h). Characterisation trials demonstrated that each precocious line was significantly less pathogenic than its parent strain and each effectively induced immunity that protected chickens against challenge with both the parent strain and other virulent field strains. Both lines had oocyst outputs that, although significantly reduced relative to the parent strains, remained sufficiently high for commercial vaccine production, and both showed susceptibility to coccidiostats. Conclusion Two attenuated lines have been produced that exhibit the appropriate characteristics for use in an Australian live coccidiosis vaccine.

Comparative growth and pathogenicity of geographical isolates of Sclerotinia sclerotiorum on lentil genotypes

Isolates of Sclerotinia sclerotiorum were collected from infected lentil plants from 2 agro-ecological zones of Syria and used to study their comparative growth on culture media and pathogenicity on different lentil genotypes. The growth studies were carried out on Potato Dextrose Agar (PDA) growth media under laboratory conditions. Mycelial radial growth and sclerotial production were the parameters used to compare the isolates. Pathogenicity studies were carried out with selected isolates on 10 lentil genotypes, infected as detached shoots and as whole potted-plants in the plastic house. The isolates showed considerable variation in cultural characteristics through mycelial growth, mycelial pigmentation and sclerotial production in the media plates. There were significant differences in the growth and sclerotial production of most of the isolates, but no apparent correlation between mycelial growth and sclerotial production among the isolates. Genotype by isolate interactions was significant for the isolates tested for pathogenicity. These interactions, however, appeared to be caused by differences in virulence of the isolates and did not suggest the occurrence of distinct pathogenic races of the pathogen isolates.

Phylogenetic considerations for predicting the host range of Ustilago sporoboli-indici, a potential biological control agent for Sporobolus species in Australia

The ribosomal DNA internal transcribed spacer region was amplified and sequenced from a selection of specimens of the Sporobolus smut Ustilago sporoboli-indici. Phylogenetic comparison with other Ustilago and Sporisorium species revealed strong support for an evolutionary radiation of Ustilago species infecting the Chloridoideae and Pooideae, of which U. sporoboli-indici forms a major lineage. Comparisons are made with other groups of plant pathogenic fungi, and it is concluded that phylogenetic analyses of potential biocontrol agents are useful for identifying pathogens that are derived from evolutionary lineages that parasitize a wide range of unrelated plants. Such pathogens are less desirable as biocontrol agents as they may have a greater likelihood of infecting plants outside their normal host ranges.

Phytogeny of the Quambalariaceae fam. nov., including important Eucalyptus pathogens in South Africa and Australia

The genus Quambalaria consists of plant-pathogenic fungi causing disease on leaves and shoots of species of Eucalyptus and its close relative, Corymbia. The phylogenetic relationship of Quambalaria spp., previously classified in genera such as Sporothrix and Ramularia, has never been addressed. It has, however, been suggested that they belong to the basidiomycete orders Exobasidiales or Ustilaginales. The aim of this study was thus to consider the ordinal relationships of Q. eucalypti and Q. pitereka using ribosomal LSU sequences. Sequence data from the ITS nrDNA were used to determine the phylogenetic relationship of the two Quambalaria species together with Fugomyces (= Cerinosterus) cyanescens. In addition to sequence data, the ultrastructure of the septal pores of the species in question was compared. From the LSU sequence data it was concluded that Quambalaria spp. and F. cyanescens form a monophyletic clade in the Microstromatales, an order of the Ustilaginomycetes. Sequences from the ITS region confirmed that Q. pitereka and Q. eucalypti are distinct species. The ex-type isolate of F. cyanescens, together with another isolate from Eucalyptus in Australia, constitute a third species of Quambalaria, Q. cyanescens (de Hoog & G.A. de Vries) Z.W. de Beer, Begerow & R. Bauer comb. nov. Transmission electron-microscopic studies of the septal pores confirm that all three Quambalaria spp. have dolipores with swollen lips, which differ from other members of the Microstromatales (i.e. the Microstromataceae and Volvocisporiaceae) that have simple pores with more or less rounded pore lips. Based on their unique ultrastructural features and the monophyly of the three Quambalaria spp. in the Microstromatales, a new family, Quambalariaceae Z.W. de Beer, Begerow & R. Bauer fam. nov., is described.

Antimicrobial susceptibility testing of Brachyspira intermedia and Brachyspira pilosicoli isolates from Australian chickens

Susceptibilities of predominantly Australian isolates of the pathogenic intestinal spirochaetes Brachyspira intermedia (n=25) and Brachyspira pilosicoli (n=17) from chickens were tested in agar dilution against four concentrations each of the antimicrobials tiamulin, lincomycin, tylosin, metronidazole, tetracycline and ampicillin. Based on available minimum inhibitory concentration (MIC) breakpoint values for Brachyspira hyodysenteriae or other Gram-negative enteric veterinary pathogens, isolates of both species generally were susceptible to tiamulin, lincomycin, metronidazole and tetracycline. Although not classed as resistant, four isolates of B. intermedia had an elevated MIC range for tiamulin (1 to 4 mg/l), 11 isolates of B. intermedia and five of B. pilosicoli had an elevated MIC range for lincomycin (10 to 50 mg/l), one isolate of B. pilosicoli had an elevated MIC range for tetracycline (10 to 20 mg/l), and one isolate of B. intermedia and five of B. pilosicoli had an elevated MIC range for ampicillin (10 to 50 mg/l). A clear lack of susceptibility to tylosin (MIC >4 mg/l) was seen in 11 isolates each of B. intermedia and B. pilosicoli, and to ampicillin (MIC >32 mg/l) in two isolates of B. pilosicoli. These data suggest that some resistance to common antimicrobials exists among intestinal spirochetes obtained from laying hens and supports the need of MIC data for clinical isolates before any treatment is considered.

Ceratocystis atrox sp. nov. associated with Phoracantha acanthocera infestations on Eucalyptus grandis in Australia

Ceratocystis spp. include important pathogens of trees as well as apparently saprophytic species. Four species have been recorded on Eucalyptus grandis in Australia, of which only one, C. pirilliformis Barnes and M.J. Wingf., is known to be pathogenic. A recent survey of pests and diseases of Eucalyptus trees in northern Queensland revealed a species of Ceratocystis associated with the tunnels made by the aggressive wood-boring insect Phoracantha acanthocera (Macleay) (Cerambicydae: Coleoptera). The aim of the present study was to identify the fungus based on morphological characteristics and comparisons of DNA sequence data for three gene regions. The fungus peripherally resembles C. fimbriata Ell. and Halst. but differs from this species most obviously by having much darker mycelium, longer ascomatal necks, segmented hyphae and an absence of aleuroconidia. Comparisons of combined sequence data confirmed that the Ceratocystis sp. from P. acanthocera represents an undescribed taxon, which is provided with the name Ceratocystis atrox sp. nov. C. atrox appears to have a close relationship with P. acanthocera, although its role in the biology of the insect is unknown and its pathogenicity has not been considered.

Insecticide resistance and implications for future aphid management in Australian grains and pastures: A review

Aphids can cause substantial damage to cereals, oilseeds and legumes through direct feeding and through the transmission of plant pathogenic viruses. Aphid-resistant varieties are only available for a limited number of crops. In Australia, growers often use prophylactic sprays to control aphids, but this strategy can lead to non-target effects and the development of insecticide resistance. Insecticide resistance is a problem in one aphid pest of Australian grains in Australia, the green peach aphid (Myzus persicae). Molecular analyses of field-collected samples demonstrate that amplified E4 esterase resistance to organophosphate insecticides is widespread in Australian grains across Australia. Knockdown resistance to pyrethroids is less abundant, but has an increased frequency in areas with known frequent use of these insecticides. Modified acetylcholinesterase resistance to dimethyl carbamates, such as pirimicarb, has not been found in Australia, nor has resistance to imidacloprid. Australian grain growers should consider control options that are less likely to promote insecticide resistance, and have reduced impacts on natural enemies. Research is ongoing in Australia and overseas to provide new strategies for aphid management in the future.

First fungal pathogen to be utilized for weed biocontrol in Fiji and Papua New Guinea

Biological control of weeds has been carried out in Fiji since 1911, when the seed-fly Ophiomyia lantanae was introduced in an attempt to control Lantana camara. In 1988, the thrips Liothrips mikaniae was introduced from Trinidad into the Solomon Islands in an attempt to undertake biocontrol of Mikania micrantha (mikania) in the Pacific. A small colony of the thrips was subsequently taken from the Solomon Islands to the Kerevat Lowlands Agricultural Experimental Station in New Britain, Papua New Guinea (PNG). Now two decades later and for the first time, a pathogenic rust fungus has been imported for use against mikania, one of Fiji’s and the Pacific’s worst invasive weeds.

Studies on the rust Prospodium tuberculatum, a new classical biological control agent released against the invasive alien weed Lantana camara in Australia. 2. Host range

A strain of the rust Prospodium tuberculatum from Brazil was screened as a potential biocontrol agent against 40 Australian Lantana camara forms and 52 closely related, non-target plant species. Results under glasshouse conditions showed that the Brazilian rust strain is pathogenic to only two flower colour forms: pink and pink-edged red. Macro- and microsymptoms were recorded using 11 assessment categories and four susceptibility ratings. No macrosymptoms were observed on any of the non-target plants.

The infectivity and host range of Orgyia anartoides nucleopolyhedrovirus.

The painted apple moth (PAM), Teia anartoides (Walker) (Lepidoptera: Lymantriidae) made a recent incursion into New Zealand. A nucleopolyhedrovirus (NPV), Orgyia anartoides NPV (OranNPV), originally isolated from PAM in Australia, was tested for its pathogenicity to PAM and a range of non-target insect species found in New Zealand, to evaluate its suitability as a microbial control for this insect invader. Dosage-mortality tests showed that OranNPV was highly pathogenic to PAM larvae; mean LT50 values for third instars ranged from 17.9 to 8.1 days for doses from 102 to 105 polyhedral inclusion bodies/larva, respectively. The cause of death in infected insects was confirmed as OranNPV. Molecular analysis established that OranNPV can be identified by PCR and restriction digestion, and this process complemented microscopic examination of infected larvae. No lymantriid species occur in New Zealand; however, the virus had no significant effects on species from five other lepidopteran families (Noctuidae, Tortricidae, Geometridae, Nymphalidae and Plutellidae) or on adult honeybees. Thus, all indications from this initial investigation are that OranNPV would be an important tool in the control of PAM in a future incursion of this species into New Zealand.

Prospects for the biological control of Lantana camara (Verbenaceae) in New Zealand

Lantana camara is an environmental weed in the northern North Island of New Zealand. It is an increasingly observed problem in forest margins, coastal scrublands, dunes, plantations and island habitats, and its rapid, uncontrolled growth can create dense impenetrable thickets, suppressing vegetation and bush regeneration. Biological control options are being considered for its management. A strain of the Brazilian rust Prospodium tuberculatum was released against lantana in Australia in 2001. This rust was screened against invasive forms of the weed that occur in New Zealand and was found to be pathogenic under glasshouse conditions. A survey found no evidence that the rust occurs in New Zealand. It is concluded that P. tuberculatum is potentially a suitable agent for the biocontrol of lantana in New Zealand and further research should be carried out prior to importation of the organism.

Controlled exposure as a management tool for Glasser's disease.

In this study, nasal swabs taken from multiparous sows at weaning time or from sick pigs displaying symptoms of Glasser's disease from farms in Australia [date not given] were cultured and analysed by polymerase chain reaction (PCR). Within each genotype detected on a farm, representative isolates were serotyped by gel diffusion (GD) testing or indirect haemagglutination (IHA) test. Isolates which did not react in any of the tests were regarded as non-typable and were termed serovar NT. Serovars 1, 5, 12, 13 and 14 were classified as highly pathogenic; serovars 2, 4 and 15 being moderately pathogenic; serovar 8 being slightly pathogenic and serovars 3, 6, 7, 9 and 11 being non-pathogenic. Sows were inoculated with the strain of Haemophilus parasuis (serovars 4, 6 and 9 from Farms 1, 2 and 4, respectively) used for controlled challenge 3 and 5 weeks before farrowing. Before farrowing the sows were divided into control and treatment groups. Five to seven days after birth, the piglets of the treatment group were challenged with a strain from the farm which had were used to vaccinate the sows. The effectiveness of the controlled exposure was evaluated by number of piglets displaying clinical signs possibly related to infection, number of antibiotic treatments and pig mortality. Nasal swabs of sick pigs were taken twice a week to find a correlation to infection. A subsample of pigs was weighed after leaving the weaning sheds. The specificity of a realtime PCR amplifying the infB gene was evaluated with 68 H. parasuis isolates and 36 strains of closely related species. 239 samples of DNA from tissues and fluids of 16 experimentally challenged animals were also tested with the realtime PCR, and the results compared with culture and a conventional PCR. The farm experiments showed that none of the controlled challenge pigs showed any signs of illness due to Glasser's disease, although the treatment groups required more antibiotics than the controls. A total of 556 H. parasuis isolates were genotyped, while 150 isolates were serotyped. H. parasuis was detected on 19 of 20 farms, including 2 farms with an extensive history of freedom from Glasser's disease. Isolates belonging to serovars regarded as potentially pathogenic were obtained from healthy pigs at weaning on 8 of the 10 farms with a history of Glasser's disease outbreaks. Sampling 213 sick pigs yielded 115 isolates, 99 of which belonged to serovars that were either potentially pathogenic or of unknown pathogenicity. Only 16 isolates from these sick pigs were of a serovar known to be non-pathogenic. Healthy pigs also had H. parasuis, even on farms free of Glasser's disease. The realtime PCR gave positive results for all 68 H. parasuis isolates and negative results for all 36 non-target bacteria. When used on the clinical material from experimental infections, the realtime PCR produced significantly more positive results than the conventional PCR (165 compared to 86).

Interactions between rotation breaks, tillage and N management on sugarcane grown at Bundaberg and Ingham.

The impact of cropping histories (sugarcane, maize and soybean), tillage practices (conventional tillage and direct drill) and fertiliser N in the plant and 1st ratoon (1R) crops of sugarcane were examined in field trials at Bundaberg and Ingham. Average yields at Ingham (Q200) and Bundaberg (Q151) were quite similar in both the plant crop (83 t/ha and 80 t/ha, respectively) and the 1R (89 t/ha v 94 t/ha, respectively), with only minor treatment effects on CCS at each site. Cane yield responses to tillage, break history and N fertiliser varied significantly between sites. There was a 27% yield increase in the plant crop from the soybean fallow at Ingham, with soybeans producing a yield advantage over continuous cane, but there were no clear break effects at Bundaberg - possibly due to a complex of pathogenic nematodes that responded differently to soybeans and maize breaks. There was no carryover benefit of the soybean break into the 1R crop at Ingham, while at Bundaberg the maize break produced a 15% yield advantage over soybeans and continuous cane. The Ingham site recorded positive responses to N fertiliser addition in both the plant (20% yield increase) and 1R (34% yield increase) crops, but there was negligible carryover benefit from plant crop N in the 1R crop, or of a reduced N response after a soybean rotation. By contrast, the Bundaberg site showed no N response in any history in the plant crop, and only a small (5%) yield increase with N applied in the 1R crop. There was again no evidence of a reduced N response in the 1R crop after a soybean fallow. There were no significant effects of tillage on cane yields at either site, although there were some minor interactions between tillage, breaks and N management in the 1R crop at both sites. Crop N contents at Bundaberg were more than 3 times those recorded at Ingham in both the plant and 1R crops, with N concentrations in millable stalk at Ingham suggesting N deficiencies in all treatments. There was negligible additional N recovered in crop biomass from N fertiliser application or soybean residues at the Ingham site. There was additional N recovered in crop biomass in response to N fertiliser and soybean breaks at Bundaberg, but effects were small and fertiliser use efficiencies poor. Loss pathways could not be quantified, but denitrification or losses in runoff were the likely causes at Ingham while leaching predominated at Bundaberg. Results highlight the complexity involved in developing sustainable farming systems for contrasting soil types and climatic conditions. A better understanding of key sugarcane pathogens and their host range, as well as improved capacity to predict in-crop N mineralisation, will be key factors in future improvements to sugarcane farming systems.

Outbreak of angular leaf spot, caused by Xanthomonas fragariae, in a Queensland strawberry germplasm collection.

Strawberry (Fragaria (x) ananassa) plants exhibiting leaf lesions consistent with angular leaf spot (ALS, caused by Xanthomonas fragariae Kennedy and King 1962) were identified in the Queensland strawberry germplasm at Bundeberg in May 2010. Water suspensions of bacterial ooze tested positive using a previously described primer set. However, the slow growth rate of X. fragariae and the presence of a fast-growing, non-pathogenic, undescribed Xanthomonas species presented problems that were overcome by dilution plating and DNA sequence analysis. Sequencing of the gyrB locus of putative colonies of X. fragariae indicated 100% sequence similarity to other X. fragariae isolates. A new set of diagnostic primers for X. fragariae based on the gyrB locus is presented.