Comparative sensitivity to methyl eugenol of four putative Bactrocera dorsalis complex sibling species – further evidence that they belong to one and the same species B. dorsalis

Males of certain species belonging to the Bactrocera dorsalis complex are strongly attracted to, and readily feed on methyl eugenol (ME), a plant secondary compound that is found in over 480 plant species worldwide. Amongst those species is one of the world’s most severe fruit pests the Oriental fruit fly, Bactrocera dorsalis s.s., and the former taxonomic species Bactrocera invadens, Bactrocera papayae and Bactrocera philippinensis. The latter species have been recently synonymised with Bactrocera dorsalis based on their very similar morphology, mating compatibility, molecular genetics and identical sex pheromones following consumption of ME. Previous studies have shown that male fruit fly responsiveness to lures is a unique phenomenon that is dose species-specific, besides showing a close correlation to sexual maturity attainment. This led us to use ME sensitivity as a behavioural parameter to test if Bactrocera dorsalis and the three former taxonomic species had similar sensitivity towards odours of ME. Using Probit analysis, we estimated the median dose of ME required to elicit species’ positive response in 50% of each population tested (ED₅₀). ED₅₀ values were compared between Bactrocera dorsalis and the former species. Our results showed no significant differences between Bactrocera dorsalis s.s., and the former Bactrocera invadens, Bactrocera papayae and Bactrocera philippinensis in their response to ME. We consider that the Bactrocera males’ sensitivity to ME may be a useful behavioural parameter for species delimitation and, in addition to other integrative taxonomic tools used, provides further supportive evidence that the four taxa belong to one and the same biological species, Bactrocera dorsalis.     

Mechanism of hydrogen peroxide-induced keratinocyte migration in a scratch-wound model

Recent studies have shown that low concentrations of H₂O₂ are produced endogenously by nonphagocytes after wounding. We observed that H₂O₂ at such concentrations can stimulate proliferation as well as migration of keratinocytes in a scratch-wound assay. Both wounding and H₂O₂ can induce phosphorylation of ERK1/2 via EGFR, but the activation of ERK1/2 by H₂O₂ is more sustained and can last more than 8 h. Sustained ERK1/2 activation is required for the increased proliferation and migration induced by H₂O₂. The p38 MAPK was also found to be phosphorylated upon treatment with H₂O₂ but it was not required for H₂O₂-induced migration or proliferation. Furthermore, it was observed that there is a cross talk between the ERK1/2 and the p38 pathways whereby inhibition of either pathway can lead to activation of the other. As a result, the motogenic effects of H₂O₂ were further enhanced when p38 was inhibited. Our data are consistent with the view that H₂O₂ may play an important signaling role in wound healing.     

Multivariate selection under adverse genetic correlations: impacts of population sizes and selection strategies on gains and coancestry in forest tree breeding

We examined gene models for two traits with and without antagonistic pleiotropy using a locus-based simulation model to investigate the effects of different population sizes, heritabilities and economic weights, using index selection, and index selection with optimum selection (OS), over 10 generations. Gene models included additive and dominance gene action, with equal and varying initial allele frequencies with and without pleiotropy for a fixed level of resources (i.e. founder sizes each generation of 40, 80 and 160 with progeny arrays that totaled 800 per generation). Pleiotropy (with an initial r _g of −0.5), reduced gain by ~8–10% when heritabilities for both traits were the same (0.2), relative to non-pleiotropic cases. When traits had different heritabilities (i.e. 0.2 and 0.4), gains in the lower heritability trait were substantially lower, especially with pleiotropy present. In general, OS with slightly larger population sizes could offset losses in gain, but rarely overrode the large effects of different heritabilities or economic weights. Pleiotropy increased response variance among traits, which was magnified when heritabilities were different. Identifying an appropriate weight on relatedness in the OS process is important to manage coancestry expectations around the loss of alleles (or fixation of recessive alleles) and to minimise response variance. The dynamics of selection intensity, drift, rate of coancestry build-up, response variance, etc. are complex for multi-trait selection; however, a few economically viable strategies could reduce the adverse effects of selecting against genetic correlations without drastically impairing gain.     

Down-regulated NOD2 by immunosuppressants in peripheral blood cells in patients with SLE reduces the muramyl dipeptide-induced IL-10 production

Background

Pattern recognition receptors (PRRs) such as Toll-like receptors are aberrantly expressed of peripheral blood mononuclear cells (PBMCs) in systemic lupus erythematosus (SLE) patients, for playing immunopathological roles.

Methodology/Principal Findings

We investigated the expression and function of the PRR nucleotide-binding oligomerization domain (NOD2) in SLE. NOD2 expression in T, B lymphocytes, monocytes, myeloid dendritic cells (mDCs) and plasmacytoid dendritic cells (pDCs) was assessed in SLE patients and healthy controls (HCs) using flow cytometric analysis. Ex vivo production of cytokines from PBMCs upon NOD2 agonist muramyl dipeptide (MDP) stimulation was assessed using Cytometric Bead Array. Over-expression of NOD2 in monocytes was observed in immunosuppressant naïve SLE patients, and was positively associated with longer disease duration. Immunosuppressive therapy was an independent explanatory variable for downregulating NOD2 expression in CD8+ T, monocytes, mDCs and pDCs. Ex vivo basal productions of cytokines (IL-6, IL-8 and IL-10) were significantly increased in immunosuppressant naïve patients and patients with active disease despite immunosuppressants compared with HCs. Upon MDP stimulaiton, relative induction (%) of cytokines (IL-1β) from PBMC was significantly increased in immunosuppressant naïve patients with inactive disease, and patients with active disease despite immunosuppressant treatment compared with HCs. Immunosuppressant usage was associated with a decreased basal production and MDP induced relative induction (%) of IL-10 in patients with inactive disease compared with immunosuppressant naïve patients and HCs.

Conclusions/Significance

Bacterial exposure may increase the NOD2 expression in monocytes in immunosuppressant naïve SLE patients which can subsequently lead to aberrant activation of PBMCs to produce proinflammatory cytokines, implicating the innate immune response for extracellular pathogens in the immunopathological mechanisms in SLE. Immunosuppressant therapy may downregulate NOD2 expression in CD8+ T lymphocytes, monocytes, and DCs in SLE patients which subsequently IL-10 reduction, contributing towards the regulation of immunopathological mechanisms of SLE, at the expense of increasing risk of bacterial infection.     

Assessment of human pancreatic islet architecture and composition by laser scanning confocal microscopy.

The recent success of pancreatic islet transplantation has generated considerable enthusiasm. To better understand the quality and characteristics of human islets used for transplantation, we performed detailed analysis of islet architecture and composition using confocal laser scanning microscopy. Human islets from six separate isolations provided by three different islet isolation centers were compared with isolated mouse and non-human primate islets. As expected from histological sections of murine pancreas, in isolated murine islets alpha and delta cells resided at the periphery of the beta-cell core. However, human islets were markedly different in that alpha, beta, and delta cells were dispersed throughout the islet. This pattern of cell distribution was present in all human islet preparations and islets of various sizes and was also seen in histological sections of human pancreas. The architecture of isolated non-human primate islets was very similar to that of human islets. Using an image analysis program, we calculated the volume of alpha, beta, and delta cells. In contrast to murine islets, we found that populations of islet cell types varied considerably in human islets. The results indicate that human islets not only are quite heterogeneous in terms of cell composition but also have a substantially different architecture from widely studied murine islets.     

Discovery of an MIT-like atracotoxin family: spider venom peptides that share sequence homology but not pharmacological properties with AVIT family proteins

This project identified a novel family of six 66–68 residue peptides from the venom of two Australian funnel-web spiders, Hadronyche sp. 20 and H. infensa: Orchid Beach (Hexathelidae: Atracinae), that appear to undergo N- and/or C-terminal post-translational modifications and conform to an ancestral protein fold. These peptides all show significant amino acid sequence homology to atracotoxin-Hvf17 (ACTX–Hvf17), a non-toxic peptide isolated from the venom of H. versuta, and a variety of AVIT family proteins including mamba intestinal toxin 1 (MIT1) and its mammalian and piscine orthologs prokineticin 1 (PK1) and prokineticin 2 (PK2). These AVIT family proteins target prokineticin receptors involved in the sensitization of nociceptors and gastrointestinal smooth muscle activation. Given their sequence homology to MIT1, we have named these spider venom peptides the MIT-like atracotoxin (ACTX) family. Using isolated rat stomach fundus or guinea-pig ileum organ bath preparations we have shown that the prototypical ACTX–Hvf17, at concentrations up to 1 μM, did not stimulate smooth muscle contractility, nor did it inhibit contractions induced by human PK1 (hPK1). The peptide also lacked activity on other isolated smooth muscle preparations including rat aorta. Furthermore, a FLIPR Ca²⁺ flux assay using HEK293 cells expressing prokineticin receptors showed that ACTX–Hvf17 fails to activate or block hPK1 or hPK2 receptors. Therefore, while the MIT-like ACTX family appears to adopt the ancestral disulfide-directed β-hairpin protein fold of MIT1, a motif believed to be shared by other AVIT family peptides, variations in the amino acid sequence and surface charge result in a loss of activity on prokineticin receptors.     

The Use of Phase Inversion Temperature (PIT) Microemulsion Technology to Enhance Oil Utilisation during <Emphasis Type="Italic">Streptomyces rimosus</Emphasis> Fed-batch Fermentations to Produce Oxytetracycline

The use of a rapeseed oil emulsion feed, produced by a phase inversion temperature (PIT) process, produced more biomass, gave a 3-fold increase in oil utilisation and a higher oxytetracycline titre but a higher residual oil concentration when compared to a conventional fed-batch Streptomyces rimosus process fed with crude rapeseed oil. Importantly, microbial utilisation of the surfactant was confirmed for the first time.     

A new in vivo termination function for DNA polymerase I of Escherichia coli K12

Escherichia coli deleted for the tus gene are viable. Thus there must be at least one other mechanism for terminating chromosome synthesis. The tus deletion strain yielded a small fraction of cells that overproduce DNA, as determined by flow cytometry after run-out chromosome replication in the presence of rifampicin and cephalexin. A plasmid, paraBAD tus+, prevented the excess DNA replication only when arabinose was added to the medium to induce the synthesis of the Tus protein. Transduction studies were done to test whether or not additional chromosomal deletions could enhance the excess chromosome replication in the tus deletion strain. A strain containing a second deletion in metE udp overproduced DNA at a high level during run-out replication. Further studies demonstrated that a spontaneous unknown mutation had occurred during the transduction. This mutation was mapped and sequenced. It is polA(G544D). Transduction of polA(G544D) alone into the tus deletion strain produced the high DNA overproduction phenotype. The polA(G544D) and six other polA alleles were then tested in wild-type and in tus deletion backgrounds. The two alleles with low levels of 5'-->3' exonuclease (exo) overproduced DNA while those with either high or normal exo overproduce much less DNA in run-out assays in the wild-type background. In contrast, all seven mutant polA alleles caused the high DNA overproduction phenotype in a tus deletion background. To explain these results we propose a new in vivo function for wild-type DNA polymerase I in chromosome termination at site(s) not yet identified.