Do Ecological Niche Model Predictions Reflect the Adaptive Landscape of Species?: A Test Using Myristica malabarica Lam., an Endemic Tree in the Western Ghats,India

Ecological niche models (ENM) have become a popular tool to define and predict the “ecological niche” of a species. An implicit assumption of the ENMs is that the predicted ecological niche of a species actually reflects the adaptive landscape of the species. Thus in sites predicted to be highly suitable, species would have maximum fitness compared to in sites predicted to be poorly suitable. As yet there are very few attempts to address this assumption. Here we evaluate this assumption. We used Bioclim (DIVA GIS version 7.3) and Maxent (version 3.3.2) to predict the habitat suitability of Myristica malabarica Lam., an economically important tree occurring in the Western Ghats, India. We located populations of the trees naturally occurring in different habitat suitability regimes (from highly suitable to poorly suitable) and evaluated them for their regeneration ability and genetic diversity. We also evaluated them for two plant functional traits, fluctuating asymmetry – an index of genetic homeostasis, and specific leaf weight – an index of primary productivity, often assumed to be good surrogates of fitness. We show a significant positive correlation between the predicted habitat quality and plant functional traits, regeneration index and genetic diversity of populations. Populations at sites predicted to be highly suitable had a higher regeneration and gene diversity compared to populations in sites predicted to be poor or unsuitable. Further, individuals in the highly suitable sites exhibited significantly less fluctuating asymmetry and significantly higher specific leaf weight compared to individuals in the poorly suitable habitats. These results for the first time provide an explicit test of the ENM with respect to the plant functional traits, regeneration ability and genetic diversity of populations along a habitat suitability gradient. We discuss the implication of these resultsfor designing viable species conservation and restoration programs.     

Section: bioprocessing and biological engineering gene fragment polymerization for increased yield of recombinant HIV fusion inhibitor enfuvirtide

Fuzeon (Enfuvirtide, T20) is the first fusion inhibitor approved by the FDA of the USA for the treatment of HIV/AIDS in combination with other anti-retroviral drugs. Enfuvirtide is a synthetic peptide that blocks the entry of HIV into healthy host CD₄ cells, which requires very high (90 mg twice daily) therapeutic doses. To increase the yield of Enfuvirtide, a gene polymerization strategy was introduced and recombinant T20 (rT20) was expressed in Escherichia coli as a five copy repeat polypeptide with a histidine-tag. The five copy rT20 was purified by Ni-affinity chromatography and cleaved to single rT20 units by cyanogen bromide. Finally, single rT20 units were purified by reversed phase chromatography giving a yield (400 mg/l) with a purity >95 %, which exhibited specific biological activity similar to Fuzeon.     

Glycocalyx of bodies versus tails of Schistosoma mansoni cercariae. Lectin-binding, size, charge, and electron microscopic characterization

Infection with Schistosoma mansoni is initiated by penetration of the intact skin of the mammalian host with the head but not the tail of the parasite cercariae. The surface of cercariae is covered by a 1-2-micron thick carbohydrate-rich glycocalyx (gx). Furthermore, the transformation of cercariae to schistosomula (the next parasitic stage in the mammalian host) is associated with loss of gx from the bodies. To understand the role of gx in the host-parasite relationship, we have characterized the gx of both bodies and tails of S. mansoni cercariae. A fluorescent fucose-specific lectin-stained bodies and not tails of the organism. Moreover, when an enriched preparation of gx obtained by extraction with 40% aqueous phenol and exclusion from Sepharose CL-6B was subjected to affinity chromatography on insolubilized Lotus lectin, which binds fucose-containing glycans, only body gx was retained. Body gx is smaller and less negatively charged than tail gx. Electron microscopy showed that gx from bodies and tails is composed of 25-40-nm particles and fibrillar material. Carbohydrate composition of gx of bodies and tails indicate that fucose and glucose are major components, respectively. beta-Elimination experiments indicate that the linkage sugar is N-acetylgalactosamine in both cases. Upon treatment with alkaline borohydride, nearly 90% of gx of both bodies and tails was recovered as two glycan chains: I and II (Mr approximately 10,500 and 5,600, respectively). Glycan I was in both cases more negatively charged than glycan II. Fucose is the predominant sugar in glycan I of the bodies while glycan I of tails is mainly composed of glucose. The gx was resistant to several proteases. This resistance and the abundance of carbohydrate in gx may be of biological importance for survival of cercariae. The substantial differences observed between the gx of bodies and tails may provide the basis for understanding the mechanism of selective release of body gx during transformation.     

Isolation and characterization of a protective antigen for adjuvant-free immunization against Schistosoma mansoni

A single, 68,000 m.w. glycoprotein antigen from adult Schistosoma mansoni was purified by immunoaffinity chromatography with the use of a newly developed, protective, anti-schistosome murine monoclonal antibody. Immunization with two doses of 0.5 microgram or 1 microgram of purified antigen, without adjuvants, afforded a mean 28% reduction in parasite recovery in CF1 mice, and 2-% reduction in parasite BALB/c mice. On immunoblotting, the 68,000 m.w. antigen was common to S. mansoni adults and schistosomula, whereas parasite eggs contained only cross-reacting low m.w. antigens of 19,100 and 16,000. Immunization resulted in the development of anti-antigen antibody and enhanced immediate cutaneous hypersensitivity to the 31-3B6 antigen. By contrast, delayed-type hypersensitivity and sensitization to circumoval granuloma formation were not observed in immunized mice. It was concluded that the 68,000 m.w. 31-3B6 antigen represents a candidate vaccine for adjuvant-free immunization against S. mansoni.     

Concurrent determination of ezetimibe and its phase-I and II metabolites by HPLC with UV detection: quantitative application to various in vitro metabolic stability studies and for qualitative estimation in bile

Simultaneous separation and quantification of ezetimibe (EZM) and its phase-I metabolite i.e., ezetimibe ketone (EZM-K) and phase-II metabolite i.e., ezetimibe glucuronide (EZM-G) in various matrices was accomplished by gradient HPLC with UV detection. The assay procedure involved deproteinization of 500 microL of either incubation or bile sample containing analytes and internal standard (IS, theophylline) with 75 microL acetonitrile containing 25% perchloric acid. An aliquot of 100 microL supernatant was injected onto a C18 column. The chromatographic separation was achieved by gradient elution consisting of 0.05 M formic acid:acetonitrile:methanol:water at a flow rate of 1.0 mL/min. The detection of analyte peaks were achieved by monitoring the eluate using an UV detector set at 250 nm. Nominal retention times of IS, EZM-G, ezetimibe ketone glucuronide (EZM-KG), EZM and EZM-K were 9.39, 24.23, 27.82, 29.04 and 30.56 min, respectively. Average extraction efficiencies of EZM, EZM-G and IS was >75-80% and for EZM-K was >50% from all the matrices tested. Limit of quantitation (LOQ) for EZM, EZM-K and EZM-G was 0.02 microg/mL. Due to the lack of availability of reference standard of EZM-KG, the recovery and LOQ aspects for this metabolite were not assessed. Overall, the method is suitable for simultaneous measurement of EZM, and its phase-I and phase-II metabolite (EZM-G) in in vitro and in vivo studies.     

Bovine viral diarrhea virus infection affects the expression of proteins related to professional antigen presentation in bovine monocytes

The complete annotation of the cattle genome allows reliable protein identification by tandem mass spectrometry (MS(2)) and greatly facilitates proteomics. Previously, we reported that differential detergent fractionation (DDF) analysis of bovine monocytes reveals proteins related to antigen pattern recognition, uptake and presentation to immunocompetent lymphocytes. Here we have identified 47 bovine proteins, involved in immune function of professional antigen-presenting cells (APC) that have been significantly altered after cytopathic (cp) Bovine Viral Diarrhea Virus (BVDV) infection. In particular, proteins related to immune responses such as cell adhesion, apoptosis, antigen uptake, processing and presentation, acute phase response proteins, MHC class I- and II-related proteins and other molecules involved in immune function of professional antigen presentation have been significantly altered after BVDV infection. Our data suggest that cp BVDV, while promoting monocyte activation and differentiation, is inhibiting their antigen presentation to immunocompetent T cells, thus resulting in the uncontrolled inflammation mediated by activated macrophages, enhanced viral spread, and impaired anti-viral defense mechanisms in the host.     

Development of polymorphic microsatellite loci in Nothapodytes nimmoniana, a medicinally important tree from the Western Ghats, India

Nothapodytes nimmoniana is a medicinally important tree species that occur in the Western Ghats, a megadiversity hotspot in southern India. Inner stem bark of the tree contains an important anti-cancer alkaloid, camptothecin for which the natural population of the tree is heavily harvested. In this paper, we report the isolation and characterization of eight polymorphic microsatellite loci using enrichment hybridization protocol. Analysis of 36 individuals representing two populations revealed three to 12 alleles per locus. Observed heterozygosity ranged from 0.21 to 0.94 for the two populations. None of the loci tested showed linkage disequilibrium. These markers are invaluable for evaluating the genetic structure and assessing the genetic impacts of harvesting of N. nimmoniana in the Western Ghats to formulate strategies for conservation of the species.