Plant hormones and oxidative stress in Hevea brasiliensis

Plant hormones are naturally occurring organic substances that are produced within the plant at low concentrations which regulate the growth and metabolism. It was observed that over-harvesting latex through high intensity tapping had a direct effect on the endogenous hormone levels in rubber plants. This could induce the development of oxidative stress leading to several complex physiological disorders including tapping panel dryness (TPD). During oxidative stress, the levels of stress hormones increased and the growth hormones decreased in the bark tissue. Both ethylene (ET) and abscisic acid (ABA) concentrations were high in trees that are exposed to oxidative stress. The levels of hydrogen peroxide (H2O2) and its scavenging enzyme, peroxidase (Px), present in healthy trees were appeared to be capable of scavenging the H2O2 molecule produced in the tissue. Hence, the minimum stress response was noticed in the bark tissues of normal trees. The regular wounding of the bark tissues for harvesting latex cannot be avoided in rubber trees. But, the amount of Px produced in the bark tissue was inadequate to detoxify the H2O2 produced under certain physiological state of the tree (TPD) and thus leading to oxidative stress. Accumulation of malondealdehyde (MDA) was evidenced as the peroxidative damage occurred in the bark tissues of stressed trees. The tissue cyanide (CN) level was very high in stressed trees due to the low levels of CN scavenging enzyme, β-cyanolalanine synthase (β-CAS). Trees under oxidative stress had increased levels of stress hormone in the bark tissue and hence, the low levels of growth hormones and high levels of stress hormones in the soft bark tissue would have caused disorders in the cellular differentiation and metabolism in the laticiferous tissues of Hevea trees limiting the production leading to significant crop loss

Cloning and expression of hmgr1 gene from Hevea brasiliensis

Biosynthesis of natural rubber (cis-1,4-polyisoprene) takes place through mevalonate pathway in Hevea. The enzyme 3-hydroxy- 3-methyl glutaryl-CoA reductase (HMGR), which catalyses the synthesis of mevalonate from HMG-CoA is a key regulatory enzyme in this pathway. This study aimed to clone and express hmgr1 gene, in order to obtain the HMGR protein in vitro and to further use this protein as a marker for yield potential in Hevea. For this purpose, mRNA was isolated from the latex of Hevea (clone RRII 105). cDNA was synthesized and PCR amplification of coding region of hmgr1 was performed using hmgr1 specific primers. The PCR amplified product (~1.8 kb) was cloned into an expression vector (pRSET-A) and transformed into E. coli (BL21DE3) cells. Protein expression in transformed cells when monitored by SDS-PAGE analysis indicated the presence of HMGR protein (61.6 kDa). The protein would be used for developing specific antibody that could be further utilized for the quantification of HMGR in different Hevea clones for screening the yield potential. The details of cloning and expression of hmgr1 are presented and discussed

A novel vaccine platform using glucan particles for induction of protective responses against Francisella tularensis and other pathogens

Vaccines are considered the bedrock of preventive medicine. However, for many pathogens, it has been challenging to develop vaccines that stimulate protective, long-lasting immunity. We have developed a novel approach using beta-1,3-D-glucans (BGs), natural polysaccharides abundantly present in fungal cell walls, as a biomaterial platform for vaccine delivery. BGs simultaneously provide for receptor-targeted antigen delivery to specialized antigen-presenting cells together with adjuvant properties to stimulate antigen-specific and trained non-specific immune responses. This review focuses on various approaches of using BG particles (GPs) to develop bacterial and fungal vaccine candidates. A special case history for the development of an effective GP tularaemia vaccine candidate is highlighted

Expression analysis of rubber biosynthetic pathway genes in Hevea brasiliensis

Hevea brasiliensisis, the primary commercial source of natural rubber (cis-1, 4-polyisoprene), is a fundamental raw material used for manufacturing a wide range of industrial and domestic rubber products in automobile, medical and defense industries. In Hevea, biosynthesis of rubber takes place through mevalonate pathway. Clonal variations in the productivity of rubber may be the result of variations in the activities of the enzymes involved in rubber biosynthesis in different Hevea clones. In this study, expression of 14 genes corresponding to enzymes/regulatory proteins involved in rubber biosynthesis was analyzed in high and low latex yielding clones of Hevea brasiliensis. The level of expression of HbSUT3, a sucrose transporter and enzymes related to the synthesis of rubber such as hydroxymethyl glutaryl-CoA synthase (hmgs), HMG-CoA reductase (hmgr) and mevalonate diphosphate decarboxylase (MVD) were found to be significantly higher in high rubber yielding clones compared to the low rubber yielding clones. The higher expression of these genes might result in an increased supply of IPP, the isoprenoid monomer, required for rubber biosynthesis. Expression of genes in the downstream rubber biosynthetic pathway such as FPPS, RuT and REF2 were also found to be significantly higher in high rubber yielding clones than low yielders. The results suggest that high rubber yield is associated with high expression of these genes and these genes can be used as markers for high yield potential in Hevea

Krox-20 inhibits Jun-NH2-terminal kinase/c-Jun to control Schwann cell proliferation and death

The transcription factor Krox-20 controls Schwann cell myelination. Schwann cells in Krox-20 null mice fail to myelinate, and unlike myelinating Schwann cells, continue to proliferate and are susceptible to death. We find that enforced Krox-20 expression in Schwann cells cell-autonomously inactivates the proliferative response of Schwann cells to the major axonal mitogen β–neuregulin-1 and the death response to TGFβ or serum deprivation. Even in 3T3 fibroblasts, Krox-20 not only blocks proliferation and death but also activates the myelin genes periaxin and protein zero, showing properties in common with master regulatory genes in other cell types. Significantly, a major function of Krox-20 is to suppress the c-Jun NH2-terminal protein kinase (JNK)–c-Jun pathway, activation of which is required for both proliferation and death. Thus, Krox-20 can coordinately control suppression of mitogenic and death responses. Krox-20 also up-regulates the scaffold protein JNK-interacting protein 1 (JIP-1). We propose this as a possible component of the mechanism by which Krox-20 regulates JNK activity during Schwann cell development

Sequence analysis of VP1, VP2 and VP3 genes of Infectious Bursal Disease Virus from a field outbreak in Kerala, India

Infectious bursal disease (IBD), caused by infectious bursal disease virus (IBDV), is one among the top five infectious diseases of poultry that discernibly affects commercial poultry industry. The mutating viral genome of IBDV accounts for disease outbreaks in fields even after following stringent biosecurity measures and vaccination protocols. The present study is focussed on the characterisation of an IBDV field virus, IBD/CVAS/6, from a vaccinated flock in Kerala, based on the sequence analysis of VP1, VP2 and VP3 genes. Bursa of Fabricius samples collected from a 26 days-old chicken flock from a suspected outbreak in the Thiruvananthapuram district of Kerala formed the subject of the study. Reverse transcriptase-polymerase chain reaction (RT-PCR) targeting VP2 gene confirmed the presence of virus in the sample. The sequence analysis revealed that the deduced amino acid sequence of VP1 gene of IBD/CVAS/6 was 100 per cent homologous with an attenuated very virulent vaccine strain of Israel, mb and the VP2 gene was 100 per cent homologous with mb and Ventri IBDV plus vaccine strain of India. The analysis of VP3 gene also revealed the similarity with vaccine strains except for a single variation S745N in its deduced amino acid sequence. The phylogenetic analysis of IBD/CVAS/6 revealed it’s close relation with mb, Ventri IBDV plus and a very virulent strain of Israel, ks. The characteristic virulent marker amino acid motifs ‘SWSASGS’ and ‘TDN’ were present in the VP2 and VP1 genes, respectively. Hence, the study revealed that the obtained virus has emerged from an attenuated very virulent vaccine strain and hence the present study is a report of involvement of the intermediate plus vaccine strain in field outbreaks in Kerala. The role of S745N in virulence cannot be accounted from the present study, however the involvement of IBD/CVAS/6 in the outbreak might be related to S745N variation or variations in other genes of the virus or due to the inefficiency of the vaccine or vaccination protocol followed, which can be defined only after further studies. The present report is the first characterisation study in Kerala focussing on the analysis of VP1, VP2 and VP3 genes of IBDV

Collaborating to Cure the Most Common Parasites on the Planet

Soil-transmitted helminths (STHs), most notably, hookworms, whipworms, and Ascaris, are nematodes that infect more than 1.5 billion of the poorest people and are leading causes of morbidity worldwide. Only one class of de-worming drugs (anthelmintic) is commonly used in mass drug administrations. New anthelmintics are urgently needed to overcome emerging resistance and to produce higher cure rates. Crystal (Cry) proteins, in particular Cry5B, made by Bacillus thuringiensis (Bt) are promising new candidates. Cry5B has excellent anthelmintic properties against many free-living and parasitic nematodes, including in vivo efficacy against multiple STH infections in rodents (Heligomasmidoes polygyrus and Ancylostoma ceylanicum) and in pigs (Ascaris suum). An enormous challenge for STHs, very different from most diseases worked on in the developing world, is the requirement that therapies be very cheap (the people infected are very poor and current drugs costs pennies a dose), massively scalable (over 4 billion people are at risk from infection), and have a long shelf life in harsh environments, that have high temperature and humidity and no cold chain. Working together, we have made excellent progress in our development efforts to produce a deployable version of Cry5B that is cheap, safe, scalable, and stable. These efforts are focused on microbiology, bacterial engineering, expression, and formulation. In the process of this work, we have discovered a novel bacterial expression system that meets these key requirements. In addition, we will provide latest information about the broad spectrum of activity of Cry5B against key parasites that make this therapeutic a very attractive alternative from current treatments

Occurrence of thermophilic Campylobacter spp. in pigs and the assessment of biosecurity measures employed at unorganized pig farms in Thrissur, Kerala

Campylobacter spp. is considered as one of the major causes of foodborne illnesses worldwide. A total of 130 samples including faecal samples (n=40), rectal swabs (n=40) and sewage samples (n=50) were collected from the two unorganized pig farms to study the occurrence of Campylobacter spp. The biosecurity measures on the farms were also assessed. An overall occurrence of 26.15 per cent with a higher rate of isolation from rectal swabs (57.5 per cent) than faecal and sewage samples (25 per cent and 2 per cent) were observed. The occurrence of C. coli was found to be 55 per cent, while that of C. jejuni and C. coli was 5 per cent in rectal swabs collected from Farm A. Campylobacter coli could be isolated only from the sewage sample from farm B. Direct multiplex PCR screening detected C. coli in 32 per cent and 44 per cent of sewage samples from farms A and B, respectively. This indicates that the Campylobacter organisms in sewage samples might have attained viable but not culturable form. In both farms, no effective biosecurity measures were followed. The lack of biosecurity measures in farms contributes to the transmission of Campylobacter spp. from the environment to the animals. Farm workers of both the farms were unaware of hygienic practices and biosecurity measures. Furthermore, little attention was paid to personal protective measures, which could pose a significant occupational risk of contracting campylobacteriosis, resulting in complex sequelae