Detection and genetic diversity of porcine rotavirus A,B and C in eastern Australian piggeries

Rotaviruses (RV) have a high prevalence in piggeries worldwide and are one of the major pathogens causing severe diarrhoea in young pigs. RV species A, B, and C have been linked to piglet diarrhoea in Australian pig herds, but their genetic diversity has not been studied in detail. Based on sequencing of the structural viral protein 7 (VP7) RVA G genotypes G3, G4 and G5, and RVC types G1, G3, G5, and G6 have been identified in Australian piggeries in previous studies. Although occurrence of RVB was reported in Australia in 1988, no further genetic analysis has been conducted. To improve health management decisions in Australian pig herds, more information on RV prevalence and genetic diversity is needed. Here, 243 enteric samples collected from 20 pig farms within Eastern Australia were analysed for the presence of RV in different age groups using a novel PCR-based multiplex assay (Pork MultiPath™ enteric panel). RVA, RVB, and RVC were detected in 10, 14, and 14 farms, respectively. Further sequencing of VP7 in selected RV-positive samples revealed G genotypes G2, G5, G9 (RVA), G6, G8, G14, G16, G20 (RVB), and G1, G3, G5, G6 (RVC) present. RVA was only detected in young (<10 weeks old) pigs whereas RVB and RVC were also detected in older animals (>11 weeks old). Interestingly, RVB and RVC G-type occurrence differed between age groups. In conclusion, this study provides new insights on the prevalence and diversity of different RV species in pig herds of Eastern Australia whilst demonstrating the ability of the Pork MultiPath™ technology to accurately differentiate between these RV species.     

Contribution of the natural biota associated with substrates to the nutritional requirements of the post-larval shrimp, Penaeus esculentus (Haswell), inhigh-density rearing systems

The contribution of epiphytes associated with physical substrates to the nutritional requirements of post‐larval shrimp, Penaeus esculentus Haswell, was determined in high‐density rearing systems (3000, 6000 and 11 000 m^?3). Stable isotope signatures of epiphytes on polyethylene mesh substrate, AquaMats? and tank walls were compared with shrimp signatures. Two methods were used: the determination of carbon and nitrogen natural abundance ratios; and ¹⁵N‐nitrogen enrichment ratios after the addition of ¹⁵N‐ammonium to tanks. Using the natural abundance technique and a simple mixing model, epiphytes were found to contribute substantially to the carbon requirements of post‐larval shrimp (39–53%). This was despite the addition of formulated feed at satiation levels. There was no indication of a reduced contribution of carbon from epiphytes to shrimp nutrition at higher shrimp densities. The lack of a difference in the ¹⁵N/¹⁴N ratios of the two food sources meant that mixing models could not be used to calculate the contribution of nitrogen from epiphytes vs. artificial feed to shrimp nutrition. Using the ¹⁵N‐nitrogenenrichment method, the amount of nitrogen contributed by epiphytes to shrimp nutrition over 24 h could be determined. This method showed that nitrogen from epiphytes was assimilated by shrimp. ¹⁵N‐enrichment methods provided a more accurate alternative to natural abundance techniques, particularly when the stable isotope signals ofthe food sources are similar. This experiment hasshown the benefits in providing substrates for P.esculentus in high‐density rearing systems to provide an additional food source for shrimp.     

Development of new real-time PCR methods for detection of Decapod iridescent virus 1 in shrimp

Novel Decapod Iridescent virus (DIV1) infections emerged in mainland China around 2014 and have devastated shrimp aquaculture operations in Chinese coastal provinces. In 2020, DIV1 has spread to Taiwan with devastating results to shrimp and crayfish farms, in addition to being found in wild caught Penaeus monodon from the Indian Ocean. This trend is a major cause for concern and an urgent reminder to expand the tools needed to monitor the spread of DIV1 globally. Here, we describe a set of four different real-time polymerase chain reaction (PCR) assays positioned across the genome of DIV1 to detect the virus in shrimp tissues. All four assays show a wide dynamic range and high analytical sensitivity and specificity. In addition, the newly developed assays show excellent diagnostic sensitivity and specificity in clinical Litopenaeus vannamei samples of North Asian origin. The new molecular toolset will enhance global capabilities to monitor the spread of DIV1 and ultimately be used as an early warning system for farmers and authorities to engage in appropriate risk mitigation strategies.     

Feed Containing Novacq Improves Resilience of Black Tiger Shrimp,Penaeus Monodon,to Gill‐associated Virus‐induced Mortality

The ability of Novacq to improve resilience of black tiger shrimp, Penaeus monodon, to infection and mortality induced by gill‐associated virus (GAV) was investigated. Over a 26‐d period, shrimp were fed pellets with or without 10% Novacq. Following this, four replicate tanks, each containing 10 shrimp that had been fed either diet, were maintained as‐is, injected with saline or injected with GAV inoculum (i.e., 40 shrimp for each of the six groups). For shrimp (n = 20) in two of each group of four tanks, survival was monitored daily over 14 d and a pleopod was sampled from each shrimp on Days 0 and 14. For the other two tanks, a pleopod was sampled from each shrimp on Days 0, 3, 7, 10, and 14 to track changes in GAV loads over time. Survival was significantly higher (P < 0.05) from Day 7 onward among the group fed Novacq. GAV infection loads appeared to vary more between individuals in the Novacq diet cohort, but overall were not reduced significantly at any time points post‐challenge compared to shrimp tested from the Control diet cohort.     

Intensive production of juvenile tiger shrimp Penaeus monodon: An evaluation of stocking density and artificial substrates

Tiger shrimp Penaeus monodon were intensively grown from PL₁₅ for 56 d in tank systems at stocking densities of 1000 and 2000 shrimp m^− 3, with and without the addition of artificial substrates (AquaMat® (buoyant and non-buoyant) and polyethylene mesh) at each density. Shrimp growth was significantly greater at the lower density and when substrates were added. Mean shrimp weight at harvest ranged from 0.64 ± 0.06 g (2000 shrimp m^− 3, no added substrate) to 1.17 ± 0.01 g (1000 shrimp m^− 3, added substrate). Survival was high and averaged 79.5 ± 2.7% across all treatments. The addition of substrates significantly increased survival at both stocking densities; however, survival was not significantly affected by stocking density. A maximum harvest density of 1645 shrimp m^− 3 and biomass of 1.27 kg m^− 3 were produced at a stocking density of 2000 m^− 3 with added substrates. Both harvest density and biomass significantly increased with stocking density and addition of substrates. The feed conversion ratio (FCR) of formulated feed was significantly lower when substrates were added. The results show that growth of P. monodon juveniles was inversely related to stocking density during intensive production. However, production output was significantly increased by addition of artificial substrates, which enhanced both growth and survival.     

Spawning of female Black tiger shrimp (Penaeus monodon) is not impacted by muscle injection of dsRNA targeted to gill‐associated virus

The ability of domesticated Penaeus monodon, Black Tiger shrimp, to spawn following tail‐muscle injection of dsRNA was examined. Ablated domesticated female broodstock infected subclinically with gill‐associated virus (GAV) were injected with saline or a cocktail of five‐dsRNAs targeting different regions in the GAV ORF1a/1b gene. To track changes in GAV infection loads, TaqMan real‐time PCR was used to quantify mean viral RNA amounts in each of three pleopod clips collected at the time of injection (Day 0) and either immediately after a female spawned or on Day 11 when the trial was terminated. Over the trial, 4 of 19 (21%) saline‐injected shrimp spawned and 12 of 25 (48%) dsRNA‐injected shrimp spawned, with one spawning twice. Egg numbers varied from 25 600 to 459 800 for the saline‐injected shrimp and from 4900 to 213 900 for the dsRNA‐injected shrimp. Of these, one of the four egg batches hatched from saline‐injected shrimp and 9 of the 13 egg batches hatched from dsRNA‐injected shrimp. While variable, egg numbers and hatch rates recorded were typical of those obtained from domesticated broodstock at the commercial hatchery and particularly among females previously spawned. Mean GAV RNA amounts detected in pleopod samples increased in five of the eight saline‐injected shrimp tested by 1.6–227.4‐fold and decreased in 12 of the 15 ds‐RNA‐injected shrimp tested by ?1.1 to ?45.1‐fold. The study demonstrated that tail‐muscle injection of GAV‐specific dsRNA does not adversely impact the ability of P. monodon to spawn.     

Productivity benefits of selectively breeding Black Tiger shrimp (Penaeus monodon) in Australia

Advances in the domestication and selective breeding of Australian Black Tiger shrimp, Penaeus monodon, opens the opportunity for world producers to reconsider the benefits of farming this species. Just over a decade ago this species was the world's most farmed shrimp species, however, difficulty in its domestication, in part, led to the widespread establishment of Penaeus vannamei (Pacific White shrimp) as the most farmed shrimp species in the world. This study empirically evaluates the productivity benefits of commercially domesticating P. monodon against production from wild broodstock of the same species. The evaluation compared the relative production from commercial ponds stocked with the progeny of wild P. monodon broodstock and ponds stocked with the progeny of domesticated stocks. The production data were from 164 ponds of domesticated stocks and 30 ponds of wild stocks, collected over 4 years (2009–2013) from two separate farm sites of the same Australian shrimp farming company. The wild stocks were sourced from the east coast of Australia. The results suggested that the productivity of the selectively bred stocks was 39% greater compared with production from wild stocks given equivalent amounts of feed and other inputs. Furthermore, productivity was additionally enhanced depending on the choice of feeds and whether stocking took place in September rather than later in the year (i.e. in early spring rather than late spring/early summer in the Southern hemisphere). This suggests that there is significant potential to further enhance the productivity of P. monodon farms via integrating advances in domestication, feeds and management practices.     

Tetraploid inductions of Penaeus monodon using cold shock

Genetic protection of improved lines of Giant Tiger shrimp, Penaeus monodon, may be achieved through the reliable production of all triploid families, as triploid shrimp are reproductively sterile. The problem with current triploid induction methods is in producing 100?% triploid progeny reliably and doing so on a commercial scale. All triploid offspring may be achievable by mating tetraploids with diploids, overcoming current limitations. However, tetraploidy in shrimp to date has been unsuccessful, with all tetraploids dying very early in development. As the induction method of heat was found to be lethal, this study investigated an alternative method using cold water to assess induction rates and lethality. Cold-water treatments between ?2 and 14?°C were applied at 18, 20, and 22?min post-spawning detection for various durations with the aim of inhibiting first mitosis. Tetraploidy was induced in treatments at or less than 1?°C; however, no nauplii hatched from these treatments. As no diploid or tetraploid nauplii hatched from any of the treatments below 3?°C, it was determined that this cold-water treatment method is not suitable for tetraploid production in P. monodon.