Affinities between Asian non-human Schistosoma species,the S. indicum group,and the African human schistosomes

Schistosoma species have traditionally been arranged in groups based on egg morphology, geographical origins, and the genus or family of snail intermediate host. One of these groups is the 'S. indicum group' comprising species from Asia that use pulmonate snails as intermediate hosts. DNA sequences were obtained from the four members of this group (S. indicum, S. spindale, S. nasale and S. incognitum) to provide information concerning their phylogenetic relationships with other Asian and African species and species groups. The sequences came from the second internal transcribed spacer (ITS2) of the ribosomal gene repeat, part of the 28S ribosomal RNA gene (28S), and part of the mitochondrial cytochrome c oxidase subunit 1 (CO1) gene. Tree analyses using both distance and parsimony methods showed the S. indicum group not to be monophyletic. Schistosoma indicum, S. spindale and S. nasale were clustered among African schistosomes, while S. incognitum was placed as sister to the African species (using ITS2 and 28S nucleotide sequences and CO1 amino acid sequences), or as sister to all other species of Schistosoma (CO1 nucleotide sequences). Based on the present molecular data, a scenario for the evolution of the S. indicum group is discussed.     

Trio under threat: can we secure the future of rhinos,elephants and tigers in Malaysia?

Three of Malaysia’s endangered large mammal species are experiencing contrasting futures. Populations of the Sumatran rhino (Dicerorhinus sumatrensis) have dwindled to critically low numbers in Peninsular Malaysia (current estimates need to be revised) and the state of Sabah (less than 40 individuals estimated). In the latter region, a bold intervention involving the translocation of isolated rhinos is being developed to concentrate them into a protected area to improve reproduction success rates. For the Asian elephant (Elephas maximus), recently established baselines for Peninsular Malaysia (0.09 elephants/km² estimated from one site) and Sabah (between 0.56 and 2.15 elephants/km² estimated from four sites) seem to indicate globally significant populations based on dung count surveys. Similar surveys are required to monitor elephant population trends at these sites and to determine baselines elsewhere. The population status of the Malayan tiger (Panthera tigris jacksoni) in Peninsular Malaysia, however, remains uncertain as only a couple of scientifically defensible camera-trapping surveys (1.66 and 2.59 tigers/100 km² estimated from two sites) have been conducted to date. As conservation resources are limited, it may be prudent to focus tiger monitoring and protection efforts in priority areas identified by the National Tiger Action Plan for Malaysia. Apart from reviewing the conservation status of rhinos, elephants and tigers and threats facing them, we highlight existing and novel conservation initiatives, policies and frameworks that can help secure the long-term future of these iconic species in Malaysia.     

Ape conservation physiology: fecal glucocorticoid responses in wild Pongo pygmaeus morio following human visitation

Nature-based tourism can generate important revenue to support conservation of biodiversity. However, constant exposure to tourists and subsequent chronic activation of stress responses can produce pathological effects, including impaired cognition, growth, reproduction, and immunity in the same animals we are interested in protecting. Utilizing fecal samples (N = 53) from 2 wild habituated orangutans (Pongo pygmaeus morio) (in addition to 26 fecal samples from 4 wild unhabituated orangutans) in the Lower Kinabatangan Wildlife Sanctuary of Sabah, Malaysian Borneo, we predicted that i) fecal glucocorticoid metabolite concentrations would be elevated on the day after tourist visitation (indicative of normal stress response to exposure to tourists on the previous day) compared to samples taken before or during tourist visitation in wild, habituated orangutans, and ii) that samples collected from habituated animals would have lower fecal glucocorticoid metabolites than unhabituated animals not used for tourism. Among the habituated animals used for tourism, fecal glucocorticoid metabolite levels were significantly elevated in samples collected the day after tourist visitation (indicative of elevated cortisol production on the previous day during tourist visitation). Fecal glucocorticoid metabolite levels were also lower in the habituated animals compared to their age-matched unhabituated counterparts. We conclude that the habituated animals used for this singular ecotourism project are not chronically stressed, unlike other species/populations with documented permanent alterations in stress responses. Animal temperament, species, the presence of coping/escape mechanisms, social confounders, and variation in amount of tourism may explain differences among previous experiments. Acute alterations in glucocorticoid measures in wildlife exposed to tourism must be interpreted conservatively. While permanently altered stress responses can be detrimental, preliminary results in these wild habituated orangutans suggest that low levels of predictable disturbance can likely result in low physiological impact on these animals.     

Brain copper, iron, magnesium, zinc, calcium, sulfur and phosphorus storage in Wilson's disease.

PROJECT: Wilson's disease (WD) is an inherited disorder of copper metabolism characterised by juvenile liver cirrhosis and by neurological symptoms. Copper levels in brain in WD have been reported to be 10 to 15 fold normal values, depending on the different brain regions. Being very few data on copper distribution in central nervous system in WD available, it seemed of interest to study the concentration of copper and of other trace elements (Zn, P, Mg, Ca, Fe and S) in the brain of a patient died for WD. PROCEDURE: a 56 year old woman affected by WD was admitted to our hospital with signs of hepatic failure and died few days later. At autopsy, a brain slice extending from the left to the right hemisphere was divided in 28 samples. On each sample Copper, Iron, Magnesium, Phosphorus, Sulphur, Zinc and Calcium were determined by Induced Coupled Plasma Atomic Emission Spectroscopy. RESULTS: the mean concentration of copper, ranging from 88 to 158 microg/g of dry tissue in all the brain specimens was higher than literature reference values, while that of the other tested elements was considerably lower. CONCLUSIONS: 1) In the brain of WD patient examined the status of trace elements was extensively altered. Further studies are necessary to correlate the concentration of trace elements with pathological lesions and with clinical pictures. 2) The elements considered in our study showed an uneven distribution in different brain areas.     

Epidemiological Characteristics of Strongyloidiasis in Inhabitants of Indigenous Communities in Borneo Island,Malaysia

Epidemiological study on strongyloidiasis in humans is currently lacking in Malaysia. Thus, a cross-sectional study was carried out to determine the prevalence of Strongyloides stercoralis infection among the inhabitants of longhouse indigenous communities in Sarawak. A single stool and blood sample were collected from each participant and subjected to microscopy, serological and molecular techniques. Five species of intestinal parasites were identified by stool microscopy. None of the stool samples were positive for S. stercoralis. However, 11% of 236 serum samples were seropositive for strongyloidiasis. Further confirmation using molecular technique on stool samples of the seropositive individuals successfully amplified 5 samples, suggesting current active infections. The prevalence was significantly higher in adult males and tended to increase with age. S. stercoralis should no longer be neglected in any intestinal parasitic survey. Combination of more than 1 diagnostic technique is necessary to increase the likelihood of estimating the ‘true’ prevalence of S. stercoralis.     

Generation of SNP datasets for orangutan population genomics using improved reduced-representation sequencing and direct comparisons of SNP calling algorithms

Background

High-throughput sequencing has opened up exciting possibilities in population and conservation genetics by enabling the assessment of genetic variation at genome-wide scales. One approach to reduce genome complexity, i.e. investigating only parts of the genome, is reduced-representation library (RRL) sequencing. Like similar approaches, RRL sequencing reduces ascertainment bias due to simultaneous discovery and genotyping of single-nucleotide polymorphisms (SNPs) and does not require reference genomes. Yet, generating such datasets remains challenging due to laboratory and bioinformatical issues. In the laboratory, current protocols require improvements with regards to sequencing homologous fragments to reduce the number of missing genotypes. From the bioinformatical perspective, the reliance of most studies on a single SNP caller disregards the possibility that different algorithms may produce disparate SNP datasets.

Results

We present an improved RRL (iRRL) protocol that maximizes the generation of homologous DNA sequences, thus achieving improved genotyping-by-sequencing efficiency. Our modifications facilitate generation of single-sample libraries, enabling individual genotype assignments instead of pooled-sample analysis. We sequenced ~1% of the orangutan genome with 41-fold median coverage in 31 wild-born individuals from two populations. SNPs and genotypes were called using three different algorithms. We obtained substantially different SNP datasets depending on the SNP caller. Genotype validations revealed that the Unified Genotyper of the Genome Analysis Toolkit and SAMtools performed significantly better than a caller from CLC Genomics Workbench (CLC). Of all conflicting genotype calls, CLC was only correct in 17% of the cases. Furthermore, conflicting genotypes between two algorithms showed a systematic bias in that one caller almost exclusively assigned heterozygotes, while the other one almost exclusively assigned homozygotes.

Conclusions

Our enhanced iRRL approach greatly facilitates genotyping-by-sequencing and thus direct estimates of allele frequencies. Our direct comparison of three commonly used SNP callers emphasizes the need to question the accuracy of SNP and genotype calling, as we obtained considerably different SNP datasets depending on caller algorithms, sequencing depths and filtering criteria. These differences affected scans for signatures of natural selection, but will also exert undue influences on demographic inferences. This study presents the first effort to generate a population genomic dataset for wild-born orangutans with known population provenance.     

Preparation of soybean seed samples for FT-IR microspectroscopy

Typical preparation of seed samples for infrared (IR) microspectroscopy involves imbibition of the seed for varying time periods followed by cryosectioning. Imbibition, however, may initiate germination even at 4° C with associated changes in the chemistry of the sample. We have found that it is possible to section seeds that are sufficiently hard, such as soybeans, on a standard laboratory microtome without imbibition. The use of dry sectioning of unimbibed seeds is reported here, as well as a comparison of different mounting media and modes of analysis. Glycerol, Tissue-Tek, and ethanol were used as mounting media, and the quality of the resulting spectra was assessed. Ethanol was the preferred mountant, because it dried quickly with no residue and thus did not interfere with the spectrum of interest. Analysis in transmission mode using barium fluoride windows to hold the samples was compared with transmission-reflection analysis with sections mounted on special infrared-reflecting slides. The two modes of analysis performed well in different regions of the spectrum. The mode of analysis (transmission vs. transmission-reflection) should be based on the components of greatest interest in the sample.     

Efficacy of ivermectin against Parastrongylus malaysiensis infection in rats.

The efficacy of ivermectin on experimental infections of P. malaysiensis in rats was determined. Ivermectin was 99.4% and 97.9% effective at a dosage of 400 meg and 800 meg respectively at seven days post-infection. The same two dosages of ivermectin when given at 14 days post infection had an efficacy of 100%. However, as an adulticide it had only 40.7% efficacy. Ivermectin may therefore be useful for the treatment of parastrongyliasis due to the larval stages of the worm which can cause significant pathology in man and animals.     

Immunohistochemical Markers of Neural Progenitor Cells in the Early Embryonic Human Cerebral Cortex

The development of the human central nervous system represents a delicate moment of embryogenesis. The purpose of this study was to analyze the expression of multiple immunohistochemical markers in the stem/progenitor cells in the human cerebral cortex during the early phases of development. To this end, samples from cerebral cortex were obtained from 4 human embryos of 11 weeks of gestation. Each sample was formalin-fixed, paraffin embedded and immunostained with several markers including GFAP, WT1, Nestin, Vimentin, CD117, S100B, Sox2, PAX2, PAX5, Tβ4, Neurofilament, CD44, CD133, Synaptophysin and Cyclin D1. Our study shows the ability of the different immunohistochemical markers to evidence different zones of the developing human cerebral cortex, allowing the identification of the multiple stages of differentiation of neuronal and glial precursors. Three important markers of radial glial cells are evidenced in this early gestational age: Vimentin, Nestin and WT1. Sox2 was expressed by the stem/progenitor cells of the ventricular zone, whereas the postmitotic neurons of the cortical plate were immunostained by PAX2 and NSE. Future studies are needed to test other important stem/progenitor cells markers and to better analyze differences in the immunohistochemical expression of these markers during gestation.Key words: Cerebral cortex, human embryo, human development, immunohistochemistry, fetal stem cells     

Anti-tumor therapy with macroencapsulated endostatin producer cells

Background  

Theracyte is a polytetrafluoroethylene membrane macroencapsulation system designed to induce neovascularization at the tissue interface, protecting the cells from host's immune rejection, thereby circumventing the problem of limited half-life and variation in circulating levels. Endostatin is a potent inhibitor of angiogenesis and tumor growth. Continuous delivery of endostatin improves the efficacy and potency of the antitumoral therapy. The purpose of this study was to determine whether recombinant fibroblasts expressing endostatin encapsulated in Theracyte immunoisolation devices can be used for delivery of this therapeutic protein for treatment of mice bearing B16F10 melanoma and Ehrlich tumors.