Major Australian tropical fruits biodiversity: Bioactive compounds and their bioactivities

The plant kingdom harbours many diverse bioactive molecules of pharmacological relevance. Temperate fruits and vegetables have been highly studied in this regard, but there have been fewer studies of fruits and vegetables from the tropics. As global consumers demand and are prepared to pay for new appealing and exotic foods, tropical fruits are now being more intensively investigated. Polyphenols and major classes of compounds like flavonoids or carotenoids are ubiquitously present in these fruits, as they are in the temperate ones, but particular classes of compounds are unique to tropical fruits and other plant parts. Bioactivity studies of compounds specific to tropical fruit plants may lead to new drug discoveries, while the synergistic action of the wide range of diverse compounds contained in plant extracts underlies nutritional and health properties of tropical fruits and vegetables. The evidence for in vitro and animal bioactivities is a strong indicator of the pharmacological promise shown in tropical fruit plant biodiversity. In this review, we will discuss both the occurrence of potential bioactive compounds isolated and identified from a selection of tropical fruit plants of importance in Australia, as well as recent studies of bioactivity associated with such fruits and other fruit plant parts.

An innovative microplate assay to facilitate the detection of antimicrobial activity in plant extracts.

A microplate assay was modified for the detection of antimicrobial activity in plant extracts. The aim was to develop an in vitro assay that could rapidly screen plant extracts to provide quantitative data on inhibition of microbial growth. A spectrophotometric assay using a microplate with serial dilutions of the plant extract and the bacteria was developed. Two bacteria, Staphylococcus aureus and Escherichia coli, were used for this study. Essential oils, oregano (Origanum vulgare) and lemon myrtle (Backhousia citriodora), and three active components carvacrol, thymol and citral were evaluated. The reproducibility of the assay was high, with correlation coefficients (r aureus and E. coli between 0.9321 and 0.9816. Similarly, r and 0.9814. This assay could also be used to measure antimicrobial activity in plant extracts which vary in pH and color.

The genetic diversity of ampeloviruses in Australian pineapples and their association with mealybug wilt disease

Pineapple mealybug wilt-associated virus 1 (PMWaV-1), 2 (PMWaV-2) and -3 (PMWaV-3) have been detected in Australian commercial pineapple crops, along with a previously undescribed ampelovirus, for which the name Pineapple mealybug wilt-associated virus 5 (PMWaV-5) is proposed. Partial sequences extending from open reading frame 1b through to the heat shock protein homologue were obtained for PMWaV-1, -3 and -5. Phylogenetic analyses of selected regions of these sequences indicated that PMWaV-5 is a distinct species and most closely related to PMWaV-1. The amino acid sequence variation observed in the RNA-dependent RNA polymerase region of PMWaV-1 isolates was 95.8–98.4% and of PMWaV-3 isolates was 92.2–99.5%. In surveys of mealybug wilt disease (MWD) affected crops, none of the four viruses was clearly associated with the disease at all survey sites. A statistically significant association (P < 0.001) between the presence of PMWaV-2 and symptoms was observed at one survey site (site 3), but the virus was at a low incidence at the remaining three survey sites. By contrast, although PMWaV-1 and -3 were equally distributed between symptomless and MWD-affected plants at site 3, there was a statistically significant (P < 0.001) association between each of these two viruses and MWD at sites 1 and 4. At site 2, there was a statistically significant (P < 0.001) association only between PMWaV-3 and MWD. PMWaV-1 was the most commonly found of the four viruses and conversely PMWaV-5 was only occasionally found. Australian isolates of PMWaV-1, -2 and -3 were transmitted by the mealybug species Dysmicoccus brevipes.

Isolation and characterization of novel microsatellite markers and their application for diversity assessment in cultivated groundnut (Arachis hypogaea)

Background: Cultivated peanut or groundnut (Arachis hypogaea L.) is the fourth most important oilseed crop in the world, grown mainly in tropical, subtropical and warm temperate climates. Due to its origin through a single and recent polyploidization event, followed by successive selection during breeding efforts, cultivated groundnut has a limited genetic background. In such species, microsatellite or simple sequence repeat (SSR) markers are very informative and useful for breeding applications. The low level of polymorphism in cultivated germplasm, however, warrants a need of larger number of polymorphic microsatellite markers for cultivated groundnut. Results: A microsatellite- enriched library was constructed from the genotype TMV2. Sequencing of 720 putative SSR-positive clones from a total of 3,072 provided 490 SSRs. 71.2% of these SSRs were perfect type, 13.1% were imperfect and 15.7% were compound. Among these SSRs, the GT/CA repeat motifs were the most common (37.6%) followed by GA/CT repeat motifs (25.9%). The primer pairs could be designed for a total of 170 SSRs and were optimized initially on two genotypes. 104 (61.2%) primer pairs yielded scorable amplicon and 46 (44.2%) primers showed polymorphism among 32 cultivated groundnut genotypes. The polymorphic SSR markers detected 2 to 5 alleles with an average of 2.44 per locus. The polymorphic information content (PIC) value for these markers varied from 0.12 to 0.75 with an average of 0.46. Based on 112 alleles obtained by 46 markers, a phenogram was constructed to understand the relationships among the 32 genotypes. Majority of the genotypes representing subspecies hypogaea were grouped together in one cluster, while the genotypes belonging to subspecies fastigiata were grouped mainly under two clusters. Conclusion. Newly developed set of 104 markers extends the repertoire of SSR markers for cultivated groundnut. These markers showed a good level of PIC value in cultivated germplasm and therefore would be very useful for germplasm analysis, linkage mapping, diversity studies and phylogenetic relationships in cultivated groundnut as well as related Arachis species.

Spatial patterns of sub-tidal seagrasses and their tissue nutrients in the Torres Strait, northern Australia: Implications for management

The distribution and nutritional profiles of sub-tidal seagrasses from the Torres Strait were surveyed and mapped across an area of 31,000 km2. Benthic sediment composition, water depth, seagrass species type and nutrients were sampled at 168 points selected in a stratified representative pattern. Eleven species of seagrass were present at 56 (33.3%) of the sample points. Halophila spinulosa, Halophila ovalis, Cymodocea serrulata and Syringodium isoetifolium were the most common species and these were nutrient profiled. Sub-tidal seagrass distribution (and associated seagrass nutrient concentrations) was generally confined to northern-central and south-western regions of the survey area (and where sediments were comprised primarily muddy sand to gravelly sand. Seagrass nitrogen and starch content, the most important nutrients for marine herbivores, were significantly correlated with species and with the plant component (above or below ground). For all seagrass species, the above-ground component (shoots and leaves) possessed greater nitrogen concentrations than the below-ground component (roots and rhizomes), which possessed greater starch concentrations. S. isoetifolium had the highest total nitrogen concentrations (1.40±0.05% DW). However, it also had higher fibre concentrations (38.2±0.68% DW) relative to the other four species. H. ovalis possessed the highest starch concentrations (2.76±0.12% DW) and highest digestibility (83.24±0.66% DW) as well as the lowest fibre (27.2±0.66% DW). The high relative abundance (found at 55% of the sites that had seagrass) and nutrient quality characteristics of H. ovalis make it an important source of energy to marine herbivores that forage sub-tidally in the Torres Strait. There were two regions in Torres Strait (north-central and south-western) where sub-tidal seagrass meadows were prevalent and of relatively higher nutritional value. This spatial and nutritional information can be used by local agencies to manage and to protect the ecological, economic and cultural values of the sub-tidal seagrass ecosystems and associated fisheries of the Torres Strait.

Host-plant resistance and biopesticides: Ingredients for successful integrated pest management (IPM) in Australian sorghum production

There are two major pests of sorghum in Australia, the sorghum midge, Stenodiplosis sorghicola (Coquillett), and the corn earworm, Helicoverpa armigera (Hübner). During the past 10 years the management of these pests has undergone a revolution, due principally to the development of sorghum hybrids with resistance to sorghum midge. Also contributing has been the adoption of a nucleopolyhedrovirus for the management of corn earworm. The practical application of these developments has led to a massive reduction in the use of synthetic insecticides for the management of major pests of sorghum in Australia. These changes have produced immediate economic, environmental and social benefits. Other flow-on benefits include providing flexibility in planting times, the maintenance of beneficial arthropods and utilisation of sorghum as a beneficial arthropod nursery, a reduction in midge populations and a reduction in insecticide resistance development in corn earworm. Future developments in sorghum pest management are discussed.

Cyanide poisoning in cattle from Dysphania glomulifera (red crumbweed): Using the internet for rapid plant identification and diagnostic advice: Case Report

A 300-strong Angus-Brahman cattle herd near Springsure, central Queensland, was being fed Acacia shirleyi (lancewood) browse during drought and crossed a 5-hectare, previously burnt area with an almost pure growth of Dysphania glomulifera subspecies glomulifera (red crumbweed) on their way to drinking water. Forty cows died of cyanide poisoning over 2 days before further access to the plant was prevented. A digital image of a plant specimen made on a flat-bed scanner and transmitted by email was used to identify D glomulifera. Specific advice on the plant's poisonous properties and management of the case was then provided by email within 2 hours of an initial telephone call by the field veterinarian to the laboratory some 600 km away. The conventional method using physical transport of a pressed dried plant specimen to confirm the identification took 5 days. D glomulifera was identified in the rumen of one of two cows necropsied. The cyanogenic potential of D glomulifera measured 4 days after collection from the site of cattle deaths was 18,600 mg HCN/kg in dry matter. The lethal dose of D glomulifera for a 420 kg cow was estimated as 150 to 190 g wet weight. The plant also contained 4.8% KNO3 equivalent in dry matter, but nitrate-nitrite poisoning was not involved in the deaths.

Resistance to root-lesion nematodes (Pratylenchus thornei and P-neglectus) in synthetic hexaploid wheats and their durum and Aegilops tauschii parents.

Root-lesion nematodes (Pratylenchus thornei Sher and Allen and P. neglectus (Rensch) Filipijev and Schuurmans Stekhoven) cause substantial yield loss to wheat crops in the northern grain region of Australia. Resistance to P. thornei for use in wheat breeding programs was sought among synthetic hexaploid wheats (2n= 6x = 42, AABBDD) produced through hybridisations of Triticum turgidum L. subsp. durum (Desf.) Husn (2n= 4x = 28, AABB) with Aegilops tauschii Coss. (2n= 2x = 14, DD). Resistance was determined for the synthetic hexaploid wheats and their durum and Ae. tauschii parents from the numbers of nematodes in the roots of plants grown for 16 weeks in pots of pasteurised soil inoculated with P. thornei. Fifty-nine (32%) of 186 accessions of synthetic hexaploid wheats had lower numbers of nematodes than Gatcher Selection 50a (GS50a), a partially resistant bread wheat. Greater frequencies of partial resistance were present in the durum parents (72% of 39 lines having lower nematode numbers than GS50a) and in the Ae. tauschii parents (55% of 53 lines). The 59 synthetic hexaploids were re-tested in a second experiment along with their parents. In a third experiment, 11 resistant synthetic hexaploid wheats and their F-1 hybrids with Janz, a susceptible bread wheat, were tested and the F(1)s were found to give nematode counts intermediate between the respective two parents. Synthetic hexaploid wheats with higher levels of resistance resulted from hybridisations where both the durum and Ae. tauschii parents were partially resistant, rather than where only one parent was partially resistant. These results suggest that resistance to P. thornei in synthetic hexaploid wheats is polygenic, with resistances located both in the D genome from Ae. tauschii and in the A and/or B genomes from durum. Five synthetic hexaploid wheats were selected for further study on the basis of (1) a high level of resistance to P. thornei of the synthetic hexaploid wheats and of both their durum and Ae. tauschii parents, (2) being representative of both Australian and CIMMYT (International Maize and Wheat Improvement Centre) durums, and (3) being representative of the morphological subspecies and varieties of Ae. tauschii. These 5 synthetic hexaploid wheats were also shown to be resistant to P. neglectus, whereas GS50a and 2 P. thornei-resistant derivatives were quite susceptible. Results of P. thornei resistance of F(1)s and F(2)s from a half diallel of these 5 synthetic hexaploid wheats, GS50a, and Janz from another study indicate polygenic additive resistance and better general combining ability for the synthetic hexaploid wheats than for GS50a. Published molecular marker studies on a doubled haploid population between the synthetic hexaploid wheat with best general combining ability (CPI133872) and Janz have shown quantitative trait loci for resistance located in all 3 genomes. Synthetic hexaploid wheats offer a convenient way of introgressing new resistances to P. thornei and P. neglectus from both durum and Ae. tauschii into commercial bread wheats.

Seasonal changes in pasture quality and diet selection and their relationship with liveweight gain of steers grazing tropical grass and grass-legume pastures in northern Australia.

The variation in liveweight gain in grazing beef cattle as influenced by pasture type, season and year effects has important economic implications for mixed crop-livestock systems and the ability to better predict such variation would benefit beef producers by providing a guide for decision making. To identify key determinants of liveweight change of Brahman-cross steers grazing subtropical pastures, measurements of pasture quality and quantity, and diet quality in parallel with liveweight were made over two consecutive grazing seasons (48 and 46 weeks, respectively), on mixed Clitoria ternatea/grass, Stylosanthes seabrana/grass and grass swards (grass being a mixture of Bothriochloa insculpta cv. Bisset, Dichanthium sericeum and Panicum maximum var. trichoglume cv. Petrie). Steers grazing the legume-based pastures had the highest growth rate and gained between 64 and 142 kg more than those grazing the grass pastures in under 12 months. Using an exponential model, green leaf mass, green leaf %, adjusted green leaf % (adjusted for inedible woody legume stems), faecal near infrared reflectance spectroscopy predictions of diet crude protein and diet dry matter digestibility, accounted for 77, 74, 80, 63 and 60%, respectively, of the variation in daily weight gain when data were pooled across pasture types and grazing seasons. The standard error of the regressions indicated that 95% prediction intervals were large (+/- 0.42-0.64 kg/head.day) suggesting that derived regression relationships have limited practical application for accurately estimating growth rate. In this study, animal factors, especially compensatory growth effects, appeared to have a major influence on growth rate in relation to pasture and diet attributes. It was concluded that predictions of growth rate based only on pasture or diet attributes are unlikely to be accurate or reliable. Nevertheless, key pasture attributes such as green leaf mass and green leaf% provide a robust indication of what proportion of the potential growth rate of the grazing animals can be achieved.

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.

Water and thermal regimes for field pea in Australia and their implications for breeding.

There is a large gap between the refined approaches to characterise genotypes and the common use of location and season as a coarse surrogate for environmental characterisation of breeding trials. As a framework for breeding, the aim of this paper is quantifying the spatial and temporal patterns of thermal and water stress for field pea in Australia. We compiled a dataset for yield of the cv. Kaspa measured in 185 environments, and investigated the associations between yield and seasonal patterns of actual temperature and modelled water stress. Correlations between yield and temperature indicated two distinct stages. In the first stage, during crop establishment and canopy expansion before flowering, yield was positively associated with minimum temperature. Mean minimum temperature below similar to 7 degrees C suggests that crops were under suboptimal temperature for both canopy expansion and radiation-use efficiency during a significant part of this early growth period. In the second stage, during critical reproductive phases, grain yield was negatively associated with maximum temperature over 25 degrees C. Correlations between yield and modelled water supply/demand ratio showed a consistent pattern with three phases: no correlation at early stages of the growth cycle, a progressive increase in the association that peaked as the crop approached the flowering window, and a progressive decline at later reproductive stages. Using long-term weather records (1957-2010) and modelled water stress for 104 locations, we identified three major patterns of water deficit nation wide. Environment type 1 (ET1) represents the most favourable condition, with no stress during most of the pre-flowering phase and gradual development of mild stress after flowering. Type 2 is characterised by increasing water deficit between 400 degree-days before flowering and 200 degree-days after flowering and rainfall that relieves stress late in the season. Type 3 represents the more stressful condition with increasing water deficit between 400 degree-days before flowering and maturity. Across Australia, the frequency of occurrence was 24% for ET1, 32% for ET2 and 43% for ET3, highlighting the dominance of the most stressful condition. Actual yield averaged 2.2 t/ha for ET1, 1.9 t/ha for ET2 and 1.4 t/ha for ET3, and the frequency of each pattern varied substantially among locations. Shifting from a nominal (i.e. location and season) to a quantitative (i.e. stress type) characterisation of environments could help improving breeding efficiency of field pea in Australia.

Resistance gene analogues in sugarcane and sorghum and their association with quantitative trait loci for rust resistance

Fifty-four different sugarcane resistance gene analogue (RGA) sequences were isolated, characterized, and used to identify molecular markers linked to major disease-resistance loci in sugarcane. Ten RGAs were identified from a sugarcane stem expressed sequence tag (EST) library; the remaining 44 were isolated from sugarcane stem, leaf, and root tissue using primers designed to conserved RGA motifs. The map location of 31 of the RGAs was determined in sugarcane and compared with the location of quantitative trait loci (QTL) for brown rust resistance. After 2 years of phenotyping, 3 RGAs were shown to generate markers that were significantly associated with resistance to this disease. To assist in the understanding of the complex genetic structure of sugarcane, 17 of the 31 RGAs were also mapped in sorghum. Comparative mapping between sugarcane and sorghum revealed syntenic localization of several RGA clusters. The 3 brown rust associated RGAs were shown to map to the same linkage group (LG) in sorghum with 2 mapping to one region and the third to a region previously shown to contain a major rust-resistance QTL in sorghum. These results illustrate the value of using RGAs for the identification of markers linked to disease resistance loci and the value of simultaneous mapping in sugarcane and sorghum.

Panel of real-time PCRs for the multiplexed detection of two tomato-infecting begomoviruses and their cognate whitefly vector species

A new approach for the simultaneous identification of the viruses and vectors responsible for tomato yellow leaf curl disease (TYLCD) epidemics is presented. A panel of quantitative multiplexed real-time PCR assays was developed for the sensitive and reliable detection of Tomato yellow leaf curl virus-Israel (TYLCV-IL), Tomato leaf curl virus (ToLCV), Bemisia tabaci Middle East Asia Minor 1 species (MEAM1, B biotype) and B.tabaci Mediterranean species (MED, Q biotype) from either plant or whitefly samples. For quality-assurance purposes, two internal control assays were included in the assay panel for the co-amplification of solanaceous plant DNA or B.tabaci DNA. All assays were shown to be specific and reproducible. The multiplexed assays were able to reliably detect as few as 10 plasmid copies of TYLCV-IL, 100 plasmid copies of ToLCV, 500fg B.tabaci MEAM1 and 300fg B.tabaci MED DNA. Evaluated methods for routine testing of field-collected whiteflies are presented, including protocols for processing B.tabaci captured on yellow sticky traps and for bulking of multiple B.tabaci individuals prior to DNA extraction. This work assembles all of the essential features of a validated and quality-assured diagnostic method for the identification and discrimination of tomato-infecting begomovirus and B.tabaci vector species in Australia. This flexible panel of assays will facilitate improved quarantine, biosecurity and disease-management programmes both in Australia and worldwide.

Pythium soft rot of ginger: Detection and identification of the causal pathogens, and their control

Ginger is considered by many people to be the outstanding member among 1400 other species in the family Zingiberaceae. Not only it is a valuable spice used by cooks throughout the world to impart unique flavour to their dishes but it also has a long track record in some Chinese and Indian cultures for treating common human ailments such as colds and headaches. Ginger has recently attracted considerable attention for its anti-inflammatory, antibacterial and antifungal properties. However, ginger as a crop is also susceptible to at least 24 different plant pathogens, including viruses, bacteria, fungi and nematodes. Of these, Pythium spp. (within the kingdom Stramenopila, phyllum Oomycota) are of most concern because various species can cause rotting and yield loss on ginger at any of the growth stages including during postharvest storage. Pythium gracile was the first species in the genus to be reported as a ginger pathogen, causing Pythium soft rot disease in India in 1907. Thereafter, numerous other Pythium spp. have been recorded from ginger growing regions throughout the world. Today, 15 Pythium species have been implicated as pathogens of the soft rot disease. Because accurate identification of a pathogen is the cornerstone of effective disease management programs, this review will focus on how to detect, identify and control Pythium spp. in general, with special emphasis on Pythium spp. associated with soft rot on ginger.

The host range of three biotypes of Dactylopius tomentosus (Lamarck) (Hemiptera: Dactylopiidae) and their potential as biological control agents of Cylindropuntia spp. (Cactaceae) in Australia

The host range of two newly imported biotypes of Dactylopius tomentosus and their potential as biological control agents of Cylindropuntia spp. were investigated. A third biotype (imbricata) of D. tomentosus previously released in Australia to control C. imbricata was also screened to determine if it will feed on other species of Cylindropuntia occurring in Australia. Efficacy trials were conducted to evaluate the ability of the biotypes to retard the growth or kill those plant species supporting development of four or more individuals in the host test trials. The host range of the three biotypes of D. tomentosus was restricted to the genus Cylindropuntia. However, the biotypes showed varying degrees of specificity within this genus. The imbricata biotype was the only biotype to develop on Australian C. rosea provenances, albeit with a range of developmental success on all C. rosea provenances tested. The Spanish provenance supported the highest development success followed by Grawin (NSW), Lorne Station (NSW) while the least preferred was the Mexican provenance. The rosea and cholla biotypes were unsuitable candidates to control C. rosea in Australia. However, the efficacy trials showed that the cholla biotype had a high impact on four of the eight naturalised Cylindropuntia species in Australia. This biotype established rapidly and the sustained feeding of one fecund female and her progeny killed potted plants of C. imbricata and C. fulgida at week 18. This biotype has the potential to be an effective agent against C. fulgida, C. imbricata, C. kleiniae and C. tunicata and, as a consequence, an application seeking its release in Australia has been lodged.