High-resolution melt-curve analysis of random amplified polymorphic DNA (RAPD-HRM) for the characterisation of pathogenic leptospires: intra-serovar divergence, interserovar convergence, and evidence of attenuation in Leptospira reference collections.

High-resolution melt-curve analysis of random amplified polymorphic DNA (RAPD-HRM) is a novel technology that has emerged as a possible method to characterise leptospires to serovar level. RAPD-HRM has recently been used to measure intra-serovar convergence between strains of the same serovar as well as inter-serovar divergence between strains of different serovars. The results indicate that intra-serovar heterogeneity and inter-serovar homogeneity may limit the application of RAPD-HRM in routine diagnostics. They also indicate that genetic attenuation of aged, high-passage-number isolates could undermine the use of RAPD-HRM or any other molecular technology. Such genetic attenuation may account for a general decrease seen in titres of rabbit hyperimmune antibodies over time. Before RAPD-HRM can be further advanced as a routine diagnostic tool, strains more representative of the wild-type serovars of a given region need to be identified. Further, RAPD-HRM analysis of reference strains indicates that the routine renewal of reference collections, with new isolates, may be needed to maintain the genetic integrity of the collections.

Microbial indicators related to yield and disease and changes in soil microbial community structure with ginger farm management practices

TRFLP (terminal restriction fragment length polymorphism) was used to assess whether management practices that improved disease suppression and/or yield in a 4-year ginger field trial were related to changes in soil microbial community structure. Bacterial and fungal community profiles were defined by presence and abundance of terminal restriction fragments (TRFs), where each TRF represents one or more species. Results indicated inclusion of an organic amendment and minimum tillage increased the relative diversity of dominant fungal populations in a system dependant way. Inclusion of an organic amendment increased bacterial species richness in the pasture treatment. Redundancy analysis showed shifts in microbial community structure associated with different management practices and treatments grouped according to TRF abundance in relation to yield and disease incidence. ANOVA also indicated the abundance of certain TRFs was significantly affected by farming system management practices, and a number of these TRFs were also correlated with yield or disease suppression. Further analyses are required to determine whether identified TRFs can be used as general or soil-type specific bio-indicators of productivity (increased and decreased) and Pythium myriotylum suppressiveness.

Rhipicephalus (Boophilus) microplus tick in vitro feeding methods for functional (dsRNA) and vaccine candidate (antibody) screening

Rhipicephalus (Boophilus) microplus (Acari: Ixodidae) ticks cause economic losses for cattle industries throughout tropical and subtropical regions of the world estimated at $US2.5 billion annually. Lack of access to efficacious long-lasting vaccination regimes and increases in tick acaricide resistance have led to the investigation of targets for the development of novel tick vaccines and treatments. In vitro tick feeding has been used for many tick species to study the effect of new acaricides on the transmission of tick-borne pathogens. Few studies have reported the use of in vitro feeding for functional genomic studies using RNA interference and/or the effect of specific anti-tick antibodies. In particular, in vitro feeding reports for the cattle tick are limited due to its relatively short hypostome. Previously published methods were further modified to broaden optimal tick sizes/weights, feeding sources including bovine and ovine serum, optimisation of commercially available blood anti-coagulant tubes, and IgG concentrations for effective antibody delivery. Ticks are fed overnight and monitored for ∼5–6 weeks to determine egg output and success of larval emergence using a humidified incubator. Lithium-heparin blood tubes provided the most reliable anti-coagulant for bovine blood feeding compared with commercial citrated (CPDA) and EDTA tubes. Although >30 mg semi-engorged ticks fed more reliably, ticks as small as 15 mg also fed to repletion to lay viable eggs. Ticks which gained less than ∼10 mg during in vitro feeding typically did not lay eggs. One mg/ml IgG from Bm86-vaccinated cattle produced a potent anti-tick effect in vitro (83% efficacy) similar to that observed in vivo. Alternatively, feeding of dsRNA targeting Bm86 did not demonstrate anti-tick effects (11% efficacy) compared with the potent effects of ubiquitin dsRNA. This study optimises R. microplus tick in vitro feeding methods which support the development of cattle tick vaccines and treatments.

Postharvest manual answers many questions

Many authors have noted that consumer confidence in buying fresh flowers is strongly related to their perceived value in that quality and vase life must be high and consistent over time for consumers to repeat buy. Growers, wholesalers, exporters and retailers seek practical information about recommended handling and treatments at the harvest and postharvest stages, including that relating to flowers native to Australia and South Africa ("wildflowers"). This information is essential for products to be of high quality with an acceptable vase life for the end consumer, especially if exported. Published postharvest manuals generally focus on traditional flower crops and so rarely include many, or any, wildflowers. A manual entitled Postharvest Handling of Australian flowers from Native Plants and Related Species was published in 2002 and addressed this gap, but required updating. This situation presented an opportunity to provide in-depth information to compliment the Australian wildflower quality specifications (see accompanying paper in the same volume), and to assemble the latest knowledge on wildflower quality and postharvest issues. The resultant manual contains extensive information about harvesting, quality issues and recommended postharvest care focussed on wildflowers. Much of the information is documented for the first time, being based on the most up to date research and development (R&D) as well as practical experience of the floral supply chain, researchers and other technical experts. The manual provides practical and detailed information on postharvest treatment of fresh wildflowers for growers, florists, wholesalers and exporters to use on a daily basis. It discusses the many unique features of wildflowers that must be understood and managed in order to maximise their quality and vase life after marketing and export. The manual also includes postharvest advice for 16 flower- and foliage lines for which quality specifications were not produced. This advice is presented according to the same template as the specifications.