Black Sigatoka disease: new technologies to strengthen eradication strategies in Australia

In 2001, an incursion of Mycosphaerella fijiensis, the causal agent of black Sigatoka, was detected in Australia's largest commercial banana growing region, the Tully Banana Production Area in North Queensland. An intensive surveillance and eradication campaign was undertaken which resulted in the reinstatement of the disease-free status for black Sigatoka in 2005. This was the first time black Sigatoka had ever been eradicated from commercial plantations. The success of the eradication campaign was testament to good working relationships between scientists, growers, crop monitors, quarantine regulatory bodies and industry. A key contributing factor to the success was the deployment of a PCR-based molecular diagnostic assay, developed by the Cooperative Research Centre for Tropical Plant Protection (CRCTPP). This assay complemented morphological identification and allowed high throughput diagnosis of samples facilitating rapid decision-making during the eradication campaign. This paper describes the development and successful deployment of molecular diagnostics for black Sigatoka. Shortcomings in the gel-based assay are discussed and the advantages of highly specific real-time PCR assays, capable of differentiating between Mycosphaerella fijiensis, Mycosphaerella musicola and Mycosphaerella eumusae are outlined. Real-time assays may provide a powerful diagnostic tool for applications in surveillance, disease forecasting and resistance testing for Sigatoka leaf spot diseases.

Heritability of reproductive traits and genetic correlations with growth in the black tiger prawn Penaeus monodon reared in tanks

Captive-reared broodstock of the black tiger prawn (Penaeus monodon) have exhibited poor reproductive performance limiting attempts to domesticate this species. The potential for improved reproductive performance was assessed by determining heritabilities of four measures of reproductive performance, their genetic correlations with each other and with growth rate and weight at age. Heritability estimates (h2 ± S.E.) obtained from √ (days to spawn), √ (egg number), √ (nauplii number) and arcsin √ (proportion hatched) were 0.47 ± 0.15, 0.41 ± 0.18, 0.27 ± 0.16, and 0.18 ± 0.16, respectively. Estimates of genetic correlations between reproductive traits and weight at age, or growth rate were less than 0.5 except for √ (egg number) and weight at 54 weeks (0.93 ± 0.19) and √ (egg number) and 16-54-week growth (0.63 ± 0.29).

Simple sequence repeat markers associated with three quantitative trait loci for black point resistance can be used to enrich selection populations in bread wheat

Black point in wheat has the potential to cost the Australian industry $A30.4 million a year. It is difficult and expensive to screen for resistance, so the aim of this study was to validate 3 previously identified quantitative trait loci (QTLs) for black point resistance on chromosomes 2B, 4A, and 3D of the wheat variety Sunco. Black point resistance data and simple sequence repeat (SSR) markers, linked to the resistance QTLs and suited to high-throughput assay, were analysed in the doubled haploid population, Batavia (susceptible) × Pelsart (resistant). Sunco and Pelsart both have Cook in their pedigree and both have the Triticum timopheevii translocation on 2B. SSR markers identified for the 3 genetic regions were gwm319 (2B, T. timopheevii translocation), wmc048 (4AS), and gwm341 (3DS). Gwm319 and wmc048 were associated with black point resistance in the validation population. Gwm341 may have an epistatic influence on the trait because when resistance alleles were present at both gwm319 and wmc048, the Batavia-derived allele at gwm341 was associated with a higher proportion of resistant lines. Data are presented showing the level of enrichment achieved for black point resistance, using 1, 2, or 3 of these molecular markers, and the number of associated discarded resistant lines. The level of population enrichment was found to be 1.83-fold with 6 of 17 resistant lines discarded when gwm319 and wmc048 were both used for selection. Interactions among the 3 QTLs appear complex and other genetic and epigenetic factors influence susceptibility to black point. Polymorphism was assessed for these markers within potential breeding material. This indicated that alternative markers to wmc048 may be required for some parental combinations. Based on these results, marker-assisted selection for the major black point resistance QTLs can increase the rate of genetic gain by improving the selection efficiency and may facilitate stacking of black point resistances from different sources.

Atrazine degradation and transport in runoff on a Black Vertosol

In Australia communities are concerned about atrazine being detected in drinking water supplies. It is important to understand mechanisms by which atrazine is transported from paddocks to waterways if we are to reduce movement of agricultural chemicals from the site of application. Two paddocks cropped with grain sorghum on a Black Vertosol were monitored for atrazine, potassium chloride (KCl) extractable atrazine, desethylatrazine (DEA), and desisopropylatrazine (DIA) at 4 soil depths (0-0.05, 0.05-0.10, 0.10-0.20, and 0.20-0.30 m) and in runoff water and runoff sediment. Atrazine + DEA + DIA (total atrazine) had a half-life in soil of 16-20 days, more rapid dissipation than in many earlier reports. Atrazine extracted in dilute potassium chloride, considered available for weed control, was initially 34% of the total and had a half-life of 15-20 days until day 30, after which it dissipated rapidly with a half life of 6 days. We conclude that, in this region, atrazine may not pose a risk for groundwater contamination, as only 0.5% of applied atrazine moved deeper than 0.20 m into the soil, where it dissipated rapidly. In runoff (including suspended sediment) atrazine concentrations were greatest during the first runoff event (57 days after application) (85 μg/L) and declined with time. After 160 days, the total atrazine lost in runoff was 0.4% of the initial application. The total atrazine concentration in runoff was strongly related to the total concentration in soil, as expected. Even after 98% of the KCl-extractable atrazine had dissipated (and no longer provided weed control), runoff concentrations still exceeded the human health guideline value of 40 μg/L. For total atrazine in soil (0-0.05 m), the range for coefficient of soil sorption (Kd) was 1.9-28.4 mL/g and for soil organic carbon sorption (KOC) was 100-2184 mL/g, increasing with time of contact with the soil and rapid dissipation of the more soluble, available phase. Partition coefficients in runoff for total atrazine were initially 3, increasing to 32 and 51 with time, values for DEA being half these. To minimise atrazine losses, cultural practices that maximise rain infiltration, and thereby minimise runoff, and minimise concentrations in the soil surface should be adopted.

Transmission of Japanese Encephalitis Virus from the Black Flying Fox, Pteropus alecto, to Culex annulirostris Mosquitoes, Despite the Absence of Detectable Viremia.

To determine the potential role of flying foxes in transmission cycles of Japanese encephalitis virus (JEV) in Australia, we exposed Pteropus alecto (Megachiroptera: Pteropididae) to JEV via infected Culex annulirostris mosquitoes or inoculation. No flying foxes developed symptoms consistent with JEV infection. Anti-JEV IgG antibodies developed in 6/10 flying foxes exposed to infected Cx. annulirostris and in 5/5 inoculated flying foxes. Low-level viremia was detected by real-time reverse transcriptase polymerase chain reaction in 1/5 inoculated flying foxes and this animal was able to infect recipient mosquitoes. Although viremia was not detected in any of the 10 flying foxes that were exposed to JEV by mosquito bite, two animals infected recipient mosquitoes. Likewise, an inoculated flying fox without detectable viremia infected recipient mosquitoes. Although infection rates in recipient mosquitoes were low, the high population densities in roosting camps, coupled with migratory behavior indicate that flying foxes could play a role in the dispersal of JEV.

Identification of genetic regions associated with black point in barley.

Black point (BP) can cause severe losses to the barley industry through downgrading and discounting of malting barley. The genetic improvement in BP resistance of barley is complex, requiring reliable screening tools, an understanding of genotype by environment interactions and an understanding of the biochemical mechanisms of melanisation involved in BP development. Thus the application of molecular markers for resistance to BP may be a useful tool for plant breeders. We have investigated the genetic regions associated with BP resistance in the barley F2 population, Valier/Binalong. Quantitative trait loci (QTLs) contributed by the resistant parent Valier, were detected on chromosomes 2HS, 2HC, 3HL, 4HL and a QTL contributed by the susceptible parent, Binalong was detected on 5HL. Three of the four QTLs were detected in two distinctly different environments. The differences observed in BP resistance between these two environments and the implications for accelerated screening are discussed. Identified SSR markers in these regions may be useful for selecting black point resistance in related breeding materials.

Impacts of grazing management options on pasture and animal productivity in a Heteropogon contortus (black speargrass) pasture in central Queensland. 4. Animal production.

Steer liveweight gains were measured in an extensive grazing study conducted in a Heteropogon contortus (black speargrass) pasture in central Queensland between 1988 and 2001. Treatments included a range of stocking rates in native pastures, legume-oversown native pasture and animal diet supplement/spring-burning pastures. Seasonal rainfall throughout this study was below the long-term mean. Mean annual pasture utilisation ranged from 13 to 61%. Annual liveweight gains per head in native pasture were highly variable among years and ranged from a low of 43 kg/steer at 2 ha/steer to a high of 182 kg/steer at 8 ha/steer. Annual liveweight gains were consistently highest at light stocking and decreased with increasing stocking rate. Annual liveweight gain per hectare increased linearly with stocking rate. These stocking rate trends were also evident in legume-oversown pastures although both the intercept and slope of the regressions for legume-oversown pastures were higher than that for native pasture. The highest annual liveweight gain for legume-oversown pasture was 221 kg/steer at 4 ha/steer. After 13 years, annual liveweight gain per unit area occurred at the heaviest stocking rate despite deleterious changes in the pasture. Across all years, the annual liveweight advantage for legume-oversown pastures was 37 kg/steer. Compared with native pasture, changes in annual liveweight gain with burning were variable. It was concluded that cattle productivity is sustainable when stocking rates are maintained at 4 ha/steer or lighter (equivalent to a utilisation rate around 30%). Although steer liveweight gain occurred at all stocking rates and economic returns were highest at heaviest stocking rates, stocking rates heavier than 4 ha/steer are unsustainable because of their long-term impact on pasture productivity.

Impacts of grazing management options on pasture and animal productivity in a Heteropogon contortus (black speargrass) pasture in central Queensland. 3. Diet composition in autumn

The diet selected in autumn by steers fistulated at the oesophageous was studied in a subset of treatments in an extensive grazing study conducted in a Heteropogon contortus pasture in central Queensland between 1988 and 2001. These treatments were a factorial array of three stocking rates (4, 3 and 2 ha/steer) and three pasture types (native pasture, legume-oversown native pasture and animal diet supplement/spring-burning native pasture). Seasonal rainfall throughout this study was below the long-term mean and mean annual pasture utilisation ranged from 30 to 61%. Steers consistently selected H. contortus with levels decreasing from 47 to 18% of the diet as stocking rate increased from 4 ha/steer to 2 ha/steer. Stylosanthes scabra cv. Seca was always selected in legume-oversown pastures with diet composition varying from 35 to 66% despite its plant density increasing from 7 to 65 plants/m(2) and pasture composition from 20 to 50%. Steers also selected a diet containing Chrysopogon fallax, forbs and sedges in higher proportions than they were present in the pasture. Greater availability of the intermediate grasses Chloris divaricata and Eragrostis spp. was associated with increased stocking rates. Bothriochloa bladhii was seldom selected in the diet, especially when other palatable species were present in the pasture, despite B. bladhii often being the major contributor to total pasture yield. It was concluded that a stocking rate of 4 ha/steer will maintain the availability of H. contortus in the pasture.

Impacts of grazing management options on pasture and animal productivity in a Heteropogon contortus (black speargrass) pasture in central Queensland. 2. Population dynamics of Heteropogon contortus and Stylosanthes scabra cv. Seca.

The dynamics of Heteropogon contortus and Stylosanthes scabra cv. Seca populations were studied in a subset of treatments in an extensive grazing study conducted in central Queensland between 1988 and 2001. These treatments were 4 stocking rates in native pasture and 2 of these stocking rates in legume oversown and supplement/spring burning treatments. For the 1999-2000 summer, population data for H. contortus in 5 of these native pasture and supplement/burning treatments were compared with those for an additional burnt treatment. Seasonal rainfall throughout this study was below the long-term mean and mean annual pasture utilisation ranged from 24 to 61%. Increasing stocking rate from 5 to 2 ha/steer in native pasture reduced H. contortus plant density. Increasing stocking rate reduced seedling recruitment as a result of its effect on soil seedbanks. Seedling recruitment was the major determinant of change in plant density, although some individual H. contortus plants did survive throughout the study. Burning in spring 1999, particularly at light stocking rate, promoted seedling recruitment above that in both unburnt native and legume oversown pasture and resulted in increased H. contortus plant density. In the legume oversown treatments, S. scabra cv. Seca density increased rapidly from 15 plants/m2 in 1988 to 140 plants/m2 in 2001 following a lag phase between 1988 and 1993. This increased S. scabra density was associated with an eventual decline in H. contortus plant density through reduced seedling recruitment. It was concluded that H. contortus population density is sustainable at stocking rates of 4 and 5 ha/steer (30% pasture utilisation) and that spring burning at light stocking rate can promote H. contortus populations. Increasing densities of S. scabra need to be managed to prevent its dominance.

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Organic diet development for black tiger prawns

The survival and growth of black tiger prawn (Penaeus monodon) juveniles (~3.3 g) were compared after feeding in tanks over one month with several prepared diets based on organically certified ingredients. The extrusion process in the manufacture of pelletised experimental diets was similar to processes used in commercial plants and was closely documented. The daily feeding rate (6% of starting mean body weight) was split equally into two feeds, one in the morning and one in the afternoon. All diets tested produced high survival (97-100%). A widely-used commercial Australian prawn feed was used as a control diet. It contained 41.2% protein with 29.5 g kg-1 lysine, and produced the highest (P<0.05) growth (117% weight gain). Three of the experimental organic diets tested (namely, 1. wheat + soy, 2. pig weaner diet + soy, and 3. pig weaner diet + dried fish waste) produced moderate growth (73–77% weight gain). These contained 33%, 36% or 31% protein, respectively, and produced better (P<0.05) growth than diets utilising a range of other prospective ingredients (eg: wheat + dried scallop gut, wheat + fish waste, wheat + chickpea, or wheat + macadamia meal, containing 23%, 25%, 29% or 24% protein, respectively). An unfed control-treatment produced the lowest (P<0.05) growth (4% weight gain). The water stability of the experimental diets that produced the best growth was poorer than the commercial diet, suggesting that improvements in this aspect of these organic feed’s manufacture could result in additional performance benefits and possibly reduced feed wastage. Analyses revealed a linear relationship between diet performance (in terms of weight gains) and the protein and lysine contents of diets. About 70% of diet performance was explained by these factors. The superior performance of the commercial diet could be attributed primarily to its formulation using mainly marine proteins, as well as a range of other unknown factors (commercial in confidence). These other factors range from use of feed attractants, better knowledge of ingredient nutrient availability, different extrusion conditions and the use of other unspecified micro-nutrients not present in the experimental diets. The organic diets studied still require a degree of fine-tuning before structured commercial uptake. This would sensibly include further detailed investigations of the composition and nutrient availabilities of these and other organic dietary ingredients, and refinement of the extrusion process for formulated diets.

Burning, then resting, reduces wiregrass (Aristida spp.) in black speargrass pastures

Wiregrasses (Aristida spp.) are becoming more prevalent, causing reduced productivity, in the black speargrass (Heterpogon contortus) pastures of south-east Queensland. Burning a native pasture of poor botanical composition (with a high proportion of wiregrass) in spring, and resting a heavily-grazed pasture also of poor composition, both improved the condition of these pastures. However, burning in late summer did not. Three months after burning in spring, relative density of wiregrass had decreased and that of a desirable species, black speargrass, had increased. However, this effect did not persist under continuous and heavy grazing. Resting increased threefold both the yield of pasture and the proportion of black speargrass, and decreased the proportion of wiregrass by two-thirds. Burning in late summer had no effect on the relative density of black speargrass but relative density of wiregrass increased. Selective grazing after burning in late summer kept this pasture in very poor condition for 14 months. These results suggest that the best way to increase the proportion of black speargrass and reduce the proportion of wiregrass in a pasture would be to burn in spring and then rest the pasture

Ionophore-mediated transmembrane movement of divalent cations in small unilamellar liposomes: An evaluation of the chlortetracycline fluorescence technique and correlations with black lipid membrane studies

Conceptual advances in the field of membrane transport have, in the main, utilized artificial membranes, both planar and vesicular. Systems of biological interest,viz., cells and organelles, resemble vesicles in size and geometry. Methods are, therefore, required to extend the results obtained with planar membranes to liposome systems. In this report we present an analysis of a fluorescence technique, using the divalent cation probe chlortetracycline, in small, unilamellar vesicles, for the study of divalent cation fluxes. An ion carrier (X537 A) and a pore former (alamethicin) have been studied. The rate of rise of fluorescence signal and the transmembrane ion gradient have been related to transmembrane current and potential, respectively. A second power dependence of ion conduction-including the electrically silent portion thereof — on X537 A concentration, has been observed. An exponential dependence of ldquocurrentrdquo on ldquotransmembrane potentialrdquo in the case of alamethicin is also confirmed. Possible errors in the technique are discussed.

Modified Atmosphere Packaging (MAP) for Control of Black Spot Formation in Chilled Prawns

The marketing of organically labeled prawns is predominately in a cooked or raw frozen form to avoid the development of melanosis (black spot). Certification for organic status prohibits the use of any added chemicals. The application of 60% CO2/40%N2 modified atmosphere to chilled (raw) prawns using two species of prawn was investigated for the ability to control black spot formation. Sensory assessment and microbiological counts were used to determine the end of product shelf life. Modified atmosphere packaged (MAP) prawns exhibited no melanosis for up to 16 days. The high quality life was retained for 12 days; shelf life of 16 days, according to standard microbiological criteria, was achieved, which is more than twice previously reported for non-MAP prawns. Results suggest MAP may be an effective method for the marketing of organically grown prawns as well as those produced by conventional prawn aquaculture without application of the normal chemicals used to prevent black spot. Copyright © 2014 Crown Copyright.