PCR-mediated screening and cloning of a malate synthase genefrom Streptomyces griseus NCIMB 9001

Malate synthase is an essential metabolic enzyme of the glyoxylate bypass that makes possible the replenishment of carbon intermediates to cells grown on acetate. A polymerase chain reaction (PCR)-based molecular screening investigation of full-length malate synthase genes from Streptomyces spp. was initiated by our group. To this end, consensus primers were designed based on known streptomycete malate synthase sequences and successful amplification was obtained for Streptomyces griseus, S. fimbriatus and S. lipmanii. The putative full-length malate synthase gene from S. griseus was subsequently cloned, sequenced and expressed. Sequence analysis of this gene showed very high identity with other streptomycete malate synthase genes. Furthermore, high malate synthase activity was detected after heterologous expression in Escherichia coli, thus demonstrating successfully the rapid cloning and functional verification of a streptomycete malate synthase gene. Growth studies of S. griseus revealed that malate synthase activity was induced by the presence of acetate, which is a two-carbon source. Interestingly, the activity peaked during late growth phase when the biomass was declining, suggesting that the enzyme may have a late role in metabolism.     

Sexual maturity and intraspecific mating success of two sibling species of the Bactrocera dorsalis complex

Bactrocera carambolae Drew & Hancock and B. papayae Drew & Hancock (Diptera: Tephritidae), two closely related sibling species in the B. dorsalis (Hendel) complex were shown to have different rates of sexual maturity. The response of B. carambolae males to methyl eugenol was observed to begin 10 days after emergence and increased with age. The attractancy peaked at 28 days after emergence and above. The males were also found to require a higher dosage of methyl eugenol (1 mg) for optimal response. These results are in contrast with those of B. papayae reported previously. The response to methyl eugenol was also observed to correspond with the age when first mating was performed.After eclosion, both sexes of B. carambolae mated approximately 2 weeks later and showed significantly lower matings/individual when compared with B. papayae in a 60-day observation period. Both sexes of B. papayae and males of B. carambolae were able to remate on the next day. However, B. carambolae females required a minimum refractory period of 8 days in order to be sexually receptive again.During the 60-day period, the mating propensity in B. papayae (100% success) was higher than B. carambolae (71–87% success) (P < 0.05). This observation was substantiated by a higher intraspecific mating success of B. papayae as compared to B. carambolae in a field cage (P < 0.001).     

Household environmental microbiota influences early-life eczema development

Exposure to a diverse microbial environment during pregnancy and early postnatal period is important in determining predisposition towards allergy. However, the effect of environmental microbiota exposure during preconception, pregnancy and postnatal life on development of allergy in the child has not been investigated so far. In the S-PRESTO (Singapore PREconception Study of long Term maternal and child Outcomes) cohort, we collected house dust during all three critical window periods and analysed microbial composition using 16S rRNA gene sequencing. At 6 and 18 months, the child was assessed for eczema by clinicians. In the eczema group, household environmental microbiota was characterized by presence of human-associated bacteria Actinomyces, Anaerococcus, Finegoldia, Micrococcus, Prevotella and Propionibacterium at all time points, suggesting their possible contributions to regulating host immunity and increasing the susceptibility to eczema. In the home environment of the control group, putative protective effect of an environmental microbe Planomicrobium (Planococcaceae family) was observed to be significantly higher than that in the eczema group. Network correlation analysis demonstrated inverse relationships between beneficial Planomicrobium and human-associated bacteria (Actinomyces, Anaerococcus, Finegoldia, Micrococcus, Prevotella and Propionibacterium). Exposure to natural environmental microbiota may be beneficial to modulate shed human-associated microbiota in an indoor environment.     

Factors affecting the characteristics of melamine resin microcapsules containing fragrant oils

Microcapsules containing fragrant oils as a core material were prepared byin situ polymerization, using melamine-formaldehyde prepolymer as the wall material. The several parameters, such as stirring times, stirring rates, emulsifier types, emulsifier concentrations, and the viscosity of the core materials, affect the characteristics of the microcapsules. These parameters were investigated by the analyses of microcapsule size, particle size distribution, and morphology. The average microcapsule size decreased with an increase in stirring time, stirring rate, emulsifier concentration, and viscosity of the core material. It was also found that poly(vinyl alcohol) as a protective colloid could enhance the stability of the melamine-formaldehyde microcapsules.     

The GAR/RGG motif defines a family of nuclear alarmins

The nucleus is the target of autoantibodies in many diseases, which suggests intrinsic nuclear adjuvants that confer its high autoimmunogenicity. Nucleolin (NCL) is one abundant nucleolar autoantigen in systemic lupus erythematosus (SLE) patients and, in lupus-prone mice, it elicits autoantibodies early. With purified NCL, we observed that it was a potent alarmin that activated monocytes, macrophages and dendritic cells and it was a ligand for TLR2 and TLR4. NCL released by necrotic cells also exhibited alarmin activity. The NCL alarmin activity resides in its glycine/arginine-rich (GAR/RGG) motif and can be displayed by synthetic GAR/RGG peptides. Two more GAR/RGG-containing nucleolar proteins, fibrillarin (FBRL) and GAR1, were also confirmed to be novel alarmins. Therefore, the GAR/RGG alarmin motif predicts a family of nucleolar alarmins. The apparent prevalence of nucleolar alarmins suggests their positive contribution to tissue homeostasis by inducing self-limiting tissue inflammation with autoimmunity only occurring when surveillance is broken down.Subject terms: Cell death and immune response, Systemic lupus erythematosus     

Interferon-β Is a Potent Promoter of Nerve Growth Factor Production by Astrocytes

Abstract: Recent clinical evidence has suggested that interferon-β is efficacious in the treatment of the demyelinating disease, multiple sclerosis. The mechanism of its efficacy remains unclear, and suggested modes of action have focused on immune modulation. Nonimmune effects of interferon-β may also contribute to its efficacy. Given that astrocytes produce a range of neurotrophic factors, we examined the possibility that interferon-β could increase the astrocytic production of nerve growth factor (NGF), which has been reported to cause oligodendrocytes to proliferate and to extend their processes; these phenotypes can impact favorably on remyelination. When the recombinant form of mouse interferon-β was added to mouse astrocyte cultures, a dose-dependent increase in NGF mRNA was obtained. The 40-fold increase in NGF mRNA elicited by 1,000 U/ml interferon-β was far more potent than that produced by other NGF-elevating agents in this study. In concordance, the protein for NGF was elevated by interferon-β. The production of NGF by interferon-β may be relevant to its clinical efficacy in multiple sclerosis. Furthermore, we suggest the potential utility of interferon-β in Alzheimer's disease.     

Courtship behavior of the mosquitoSabethes cyaneus (Diptera: Culicidae)

Unlike any other mosquito reported, Sabethes cyaneus(Fabricius) displays an elaborate courtship before and during copulation. A male approaches a female suspended from a horizontal stick, suspends himself in front of her as he grasps her folded wings, and proceeds with a series of discrete stereotyped behaviors that involve proboscis vibration and movement of iridescent blue paddles on his midlegs. The sequence of these behaviors is as follows: freeleg waving, swinging, copulation attempt, superficial coupling, waving, genital shift, waggling, and release. Insemination occurs after genital shift. The only overt reciprocation by the female is abdomen lowering during the male's swinging. Courtship is often unsuccessful, and males are usually rejected during freeleg waving. The relation between male performance and mating success remains obscure.     

Role of tumor-derived exosomes in cancer metastasis

The highlights of cancer research include the discovery of exosomes, which are small (30-100?nm) sized vesicular nanoparticles released virtually by all cells. Tumor-derived exosomes (TDEs) are notoriously known for orchestrating the invasion-metastasis cascade via systemic pathways that we have previously proposed (1), resulting in a paradigm shift of our understanding about the pathobiology of metastases. In principle, exosomes serve as transport medium for proteins, mRNAs and miRNAs to transmit targeted cues from the primary cell to distant sites via horizontal transfer or cell-receptor interaction. In this chapter, we seek to explore in-depth the mechanisms engendering TDE in the metastatic cascade, along with experimental models to augment our understanding. The aforementioned has also paved way for parallel advancements in the therapeutic armamentarium, as evident from pronounced efforts to exploit the metastatic process for therapeutic targeting. In this light, we aim to examine potential anti-metastatic therapeutic opportunities derived from exosomal research. Lastly, exosomes may play a crucial role in the contemporary era of “liquid biopsies”, given the array of molecular information with diagnostic and predictive indications. We thus intend to end this chapter off by exploring future applications of exosomes that could illuminate shortcomings and propel advancements in biomarker research.     

Presence of gamma-aminobutyric acid in embryonic palates of AJ and SWV mouse strains

The presence of gamma-aminobutyric acid (GABA) in the embryonic palate was sought as a criterion for its role in regulating palate development. GABA was measured by a gas chromatographic-mass spectrometric (GC-MS) method using the heptafluorobutyryl (HFB)-cyclohexyl-GABA derivative, which gave the necessary sensitivity and specificity to measure low levels of GABA in the presence of contaminating substances. GABA was measured in dissected embryonic palates at various times of development in the AJ mouse strain. GABA levels were lower in day 14 AJ palates (0.19 +/- 0.01 nmol/mg protein) than at days 13 (0.28 +/- 0.03) and 15 (0.30 +/- 0.04). Comparable levels were observed in fore- and hindlimbs at day 14, whereas levels were lower in embryonic tongue and higher, as was expected, in embryonic brain. To confirm the presence of GABA in the palate, it was analyzed in growing palate mesenchymal cells in primary and secondary cultures as well as in serum-free medium. In addition, GABA levels were compared in the SWV mouse strain; this strain exhibits a more efficient active uptake mechanism and diazepam produces a higher frequency of cleft palate in this strain than in AJ. SWV contained one and one-half to three times higher concentrations of GABA in excised palates and cultured palate cells than the AJ strain. Furthermore, when GABA levels in skin fibroblasts of the two strains were measured, SWV cells contained 2.7-fold greater GABA than AJ cells. The present results provide additional evidence for the role of GABA in palate development.