Jennifer Elliman

Impact of herbicide Ametryn on microbial communities in mixed liquor of a membrane bioreactor (MBR).

Ametryn, which is a second generation herbicide, was introduced to a lab-scale MBR at a concentration of 1mg/L and a 20-40% removal was observed at HRT ranging from 7.8 to 15.6h for an average influent Ametryn concentration of 0.8 mg/L. Components of EPS (protein and carbohydrates) increased in the bioreactor and the observed biomass production reduced after the addition of Ametryn. In a batch study, GAC was added to MBR mixed liquor and removal of Ametryn via biodegradation and adsorption were measured. Five common bacterial colony types (Gram negative and positive bacilli and Gram negative cocci) were identified and three of these were resistant to Ametryn up to 5mg/L. GAC was found to be a very effective Ametryn adsorption medium and in some occasions Ametryn may have acted as a nutrient source for bacteria.

Growth, osmoregulatory responses and changes to the lipid and fatty acid composition of organs from the mud crab, Scylla serrata, over a broad salinity range

Abstract Aquatic animals can often undergo substantial physiological responses to salinity; however, associated lipid/fatty acid alterations to their various tissues have received little attention. To investigate this, we measured the growth of mud crab, Scylla serrata, juveniles over two moults (duration of 23–60 days) at salinities of 4, 12, 20, 28, 36 and 44‰ (30 replicates/treatment). After the second moult, 6-day post moult crabs were sampled for hepatosomatic index (HSI), haemolymph osmolality, Na+, K+ and Ca2+ levels, anterior/posterior gill Na+/K+-ATPase activity as well as the lipid/fatty acid composition of the anterior and posterior gills, hepatopancreas and muscle. High salinities of 36 and 44‰ significantly lowered crab growth and HSI (p<0.01). S. serrata strongly hyper-regulated haemolymph osmolality and ions likely due to significantly enhanced posterior gill Na+/K+-ATPase activity. At decreasing salinities, eicosapentaenoic acid, arachidonic acid, n-3 and long chain polyunsaturated fatty acid (PUFA) significantly increased (p<0.05), likely to maintain Na+/K+-ATPase activity via increased membrane fluidity. Muscle and hepatopancreas n-3/n-6 PUFA ratios significantly decreased (p<0.05) at increasing salinities indicating possible fatty acid metabolic modifications. Results indicate that S. serrata juveniles are well adapted to low salinities, with higher salinities likely reducing their metabolism, reflected by lower growth, HSI and higher posterior gill and hepatopancreatic triacylglycerol.

Bacteriophage adenine methyltransferase: a life cycle regulator? Modelled using Vibrio harveyi myovirus like.

The adenine methyltransferase (DAM) gene methylates GATC sequences that have been demonstrated in various bacteria to be a powerful gene regulator functioning as an epigenetic switch, particularly with virulence gene regulation. However, overproduction of DAM can lead to mutations, giving rise to variability that may be important for adaptation to environmental change. While most bacterial hosts carry a DAM gene, not all bacteriophage carry this gene. Currently, there is no literature regarding the role DAM plays in life cycle regulation of bacteriophage. Vibrio campbellii strain 642 carries the bacteriophage Vibrio harveyi myovirus like (VHML) that has been proven to increase virulence. The complete genome sequence of VHML bacteriophage revealed a putative adenine methyltransferase gene. Using VHML, a new model of phage life cycle regulation, where DAM plays a central role between the lysogenic and lytic states, will be hypothesized. In short, DAM methylates the rha antirepressor gene and once methylation is removed, homologous CI repressor protein becomes repressed and non‐functional leading to the switching to the lytic cycle. Greater understanding of life cycle regulation at the genetic level can, in the future, lead to the genesis of chimeric bacteriophage with greater control over their life cycle for their safe use as probiotics within the aquaculture industry.

First complete genome of an Ambidensovirus; Cherax quadricarinatus densovirus, from freshwater crayfish Cherax quadricarinatus.

In 1999, the causative agent of an epizootic in Cherax quadricarinatus was described, and given the provisional name Cherax quadricarinatus parvovirus-like. Sequencing of the 6334 nt genome identified three open-reading frames on the top strand coding NS3 (35.55 kDa), NS1 (67.36 kDa) and NS2 (35.18 kDa) and on the bottom strand a single open reading frame which most likely encodes 4 structural proteins. Motifs characteristic of the Densovirinae were found in the ORFs. Phylogenetic analysis of the amino acids in NS1 places the genome in the genus Ambidensovirus, most closely related to the marine sea star densovirus (75%, E=0.0) and distantly related to Acheta domestica densovirus (44.1%). The virus name is proposed as species Decapod ambidensovirus, variant Cherax quadricarinatus densovirus. This is the first Ambidensovirus to be found in decapod crustaceans and the first of the subfamily Densovirinae to be sequenced from a freshwater crayfish. Cherax quadricarinatus densovirus and sea star densovirus are the first highly related Densovirinae to infect phylogenetically disparate hosts and are thus far, unique among the Densovirinae.

Effects of Colonization of the Roots of Domestic Rice (Oryza sativa L. cv. Amaroo) by Burkholderia pseudomallei

ABSTRACT Burkholderia pseudomallei is a saprophytic bacterium that causes melioidosis and is often isolated from rice fields in Southeast Asia, where the infection incidence is high among rice field workers. The aim of this study was to investigate the relationship between this bacterium and rice through growth experiments where the effect of colonization of domestic rice (Oryza sativa L. cv Amaroo) roots by B. pseudomallei could be observed. When B. pseudomallei was exposed to surface-sterilized seeds, the growth of both the root and the aerosphere was retarded compared to that in controls. The organism was found to localize in the root hairs and endodermis of the plant. A biofilm formed around the root and root structures that were colonized. Growth experiments with a wild rice species (Oryza meridionalis) produced similar retardation of growth, while another domestic cultivar (O. sativa L. cv Koshihikari) did not show retarded growth. Here we report B. pseudomallei infection and inhibition of O. sativa L. cv Amaroo, which might provide insights into plant interactions with this important human pathogen.

Assessment of a cricket, Acheta domesticus, bioassay for Chequa Iflavirus and bunya-like virus from redclaw crayfish Cherax quadricarinatus

Chequa iflavirus and a bunya-like virus infect redclaw crayfish (Cherax quadricarinatus) and they may cause mortality reaching 20-40% after about three weeks after a stress event. To complete Rivers postulates for viruses, virus-free animals are needed. Due to a lack of chequa iflavirus and bunya-like virus-free crayfish (testing shows>85% infection rate) coupled with the facts that iflavirus and bunyaviruses are found in insects and that crickets had been successful alternate hosts for crustacean viruses before, Acheta domesticus was trialled asa bioassay animal. There was no significant difference (P>0.05) in mortality rates between uninfected control crickets and infected crickets. Reverse transcriptase polymerase chain reaction for both viruses failed to find any trace of the RNA viruses in fed or inoculated crickets after 30days. The search for an alternative bioassay host will have to be widened.

A novel virus (order Bunyavirales) from stressed redclaw crayfish (Cherax quadricarinatus) from farms in northern Australia

Athtabvirus, a bunya-like virus and chequa iflavirus infect redclaw crayfish (Cherax quadricarinatus) and they may cause mortality reaching 20-40% after about three weeks following transportation stress. Lesions were seen in the muscles of broodstock and juveniles and nerve cords of craylings. Using NextGen sequencing, the whole transcriptomes of a farmed case crayfish and a tank-reared, unaffected crayfish were assembled producing over 500,000 contigs. The average depth of reads was 18 replicates with a range from 15 to 44. The near complete sequences of the large and middle genome segments of a bunya-like virus were detected along with chequa iflavirus. The internal bunya-like motifs; RNA-dependent RNA polymerase on the L segment, and glycoprotein n (Gn) on the M segment were easily identified. In the opposite, positive-sense direction on the M segment, another presumed glycoprotein (glycoprotein c) with a low-density lipoprotein receptor (cysteine-rich) motif was identified by position specific iterated (psi)-BLASTp. The athtabvirus was related to Whenzhou Shrimp Virus 2 (E = 0.0, 43% amino acid identity), an unassigned, -ve sense ssRNA virus, and to peribunyaviruses (E = 10-50-20). In descending order of the number of RNA copies/0.2 mg of tissue, the organs most heavily infected were muscle (9.4 × 106), nerve cord (5.24 × 106), heart (4.07 × 106), gills (3.96 × 106), hepatopancreas (1.58 × 106) and antennal gland (6.6 × 105). Given the tissue tropism (muscle and nerves) of athtabvirus and the original lesions, this virus is implicated in being involved in the mortalities in crayfish after transportation.

VERO CELL LINES EXPRESSING NUCLEAR LOCATION SIGNALS OF Penaeus merguiensis HEPANDENSOVIRUS: AN EARLY STUDY

Milkfish culture in marine cages contributes significantly to the total fisheries production in the Philippines and has the potential of replacing traditional brackishwater production systems in mangrove areas. For years however, it has been plagued with problems related to pollution [1,2] and erratic harvests primarily attributed to improper and unregulated culture management practices. The present study introduced bottom feeding in high density culture of milkfish in marine cages as a strategy to improve productivity without compromising environmental integrity. Aside from the work of the authors, there have been no records of bottom feeding in milkfish farming in the Philippines. The use of high density stocking (100 fish m) on the other hand, is not a recommended practice although demonstrated to be feasible [3]. Lack of sufficient scientific evidence on the relationship of increased density with growth and the absence of strategies to improve growth and survival under high density stocking in milkfish cage culture prevents its adoption. The use of underwater hydro-kinetic bottom feeder designed to improve feeding access even in highly crowded milkfish cage culture conditions addresses this issue. With improved feeding access at high density stocking, yield per unit volume is also expected to increase thus diminish the area required in generating equivalent production at lower stock densities. Poor growth with increasing density have been observed in farmed fish [3,4] although the underlying mechanisms are still poorly understood [4]. In farmed rainbow trout, the commonly reported effects with increasing density are reductions in food conversion efficiency, nutritional conditions, growth and an increase in fin erosion [5]. These contrast with the results on other species which showed positive relationships. For instance, higher growth and elevated scramble feeding were observed in hybrid striped bass held in large memberships [6]. Similarly, lower feed intake, higher growth and higher feed efficiency with increasing group size were reported in juvenile perch [7]. Growth variance in fish is often caused by aggression arising from food distribution and ration size [8,9].The hemoglobins produced by Tegillarca granosa have antibacterial activity toward some Gram-positive and Gram-negative bacteria. In this study, the genes encoding the recombinant proteins Tg-HbIIA and Tg-HbIIB were cloned from T. granosa hemocytes by RT-PCR, and the proteins were expressed in Escherichia coli Transetta (DE3). The proteins were purified using a HisTrap FF affinity chromatography column under denaturing conditions and refolded at 4 °C by urea gradient dialysis, and the antibacterial activity of the recombinant proteins was determined. The Tg-HbIIA protein had antibacterial activity toward Vibrio harveyi and Pseudomonas putida, with the minimum inhibition concentration (MIC) values of 65.8 ug/ml and 4.11 ug/ml, respectively. The Tg-HbIIB protein had antibacterial activity toward V. harveyi, P. putida and Acinetobacter baumanii, with MIC values of 158 ug/ml, 39.5 ug/ml and 79 ug/ml, respectively. They had no antibacterial activity against Staphylococcus aureus, E. coli, B. firmus, B. subtilis, S. epidermidis, or V. parahaemolyticus. This study provides a basis for further research on the antibacterial function and mechanism of hemoglobin.Aims: Penaeus merguiensis hepandensovirus (PmeHDV) (GenBank No. DQ458781) is a shrimp hepatopancreatic parvovirus (HPV), belonging to subfamily Densovirinae. Transportation of Densovirinae into and out of nucleus is allowed by the binding of nuclear location signals (NLSs) to importins (Imp). PmeHDV has putative NLSs that need to be experimentally tested. The aims of this study is to determine if the three putative NLSs of PmeHDV are functioning by transfecting NLS-inserted-plasmid DNAs into Vero cell lines using a transfection reagent.Place and Duration of the Study: Data for this study was collected from the Veterinary and Biomedical Sciences Laboratories at James Cook University (JCU) during the duration from May 2015 to December 2016. Methodology: Each plasmid has been synthetically inserted with each sequence of the putative NLSs and afluorescent protein. The presence of the NLS in the cell nucleus and cytoplasm was screened. The overlay of visualization of transfected plasmids is presented.Results: It appears the NLSs are not functioning well as that the proteins are blocked at the nuclear membrane, probably linked to importin beta-1 and not frequently entering the nucleus. Our study demonstrated small noticeable differences in the outer nuclei within transfected-Vero cells with the experimental NLSs genes.Conclusion: In conclusion, our fluorescent study was not sufficiently sensitive to be confident of the detection in NLS-transfected cells under different filters. The study of crustacean virus-host interactions using proxy cell cultures as models remains a major challenge.