Influence of season, age and breed on prevalence of haemoprotozoan diseases in cattle of Tamil Nadu, India

Aim: To assess the prevalence of haemoprotozoan diseases in cross-bred and indigenous cattle in relation to season, age and breed in Western part of Tamil Nadu, India. Materials and Methods: A total of 2637 blood smears were screened for haemoprotozoan diseases and samples were received from the college hospital and veterinary dispensaries in Western part of Tamil Nadu, India. Blood smears were stained using Giemsa's technique and examined under oil immersion. Results: Microscopic examination of blood smears revealed an overall prevalence of 16.64 %; of which theileriosis was 13 %, followed by anaplasmosis 2.64 % and then babesiosis 1.0%. Among the haemoprotozoan diseases, the prevalence of theileriosis was significantly (p<0.05) high during summer (14.4%), followed by moderate in monsoon (13.8%) and less in fair (11.5%) seasons. However, there was no significant seasonal influence on the prevalence of babesiosis and anaplasmosis. The data on influence of breed revealed that there was a significantly (p<0.05) high prevalence of haemoprotozoan diseases in Holstein Friesian (HF) and Jersey cross breeds than indigenous breed and the occurrence of these haemoprotozoan diseases was found to be high among the age groups of 2-7 years in cross-bred animals and below 2 years in indigenous animals. Conclusion: The present study suggests that Western part of Tamil Nadu is highly endemic for theileriosis and occurrence of the disease was high during summer. Cross-bred animals aged 2-7 years are highly susceptible to these haemoprotozoan diseases than indigenous animals

Comparative proteomics of related symbiotic mussel species reveals high variability of host-symbiont interactions.

Deep-sea Bathymodiolus mussels and their chemoautotrophic symbionts are well-studied representatives of mutualistic host-microbe associations. However, how host-symbiont interactions vary on the molecular level between related host and symbiont species remains unclear. Therefore, we compared the host and symbiont metaproteomes of Pacific B. thermophilus, hosting a thiotrophic symbiont, and Atlantic B. azoricus, containing two symbionts, a thiotroph and a methanotroph. We identified common strategies of metabolic support between hosts and symbionts, such as the oxidation of sulfide by the host, which provides a thiosulfate reservoir for the thiotrophic symbionts, and a cycling mechanism that could supply the host with symbiont-derived amino acids. However, expression levels of these processes differed substantially between both symbioses. Backed up by genomic comparisons, our results furthermore revealed an exceptionally large repertoire of attachment-related proteins in the B. thermophilus symbiont. These findings imply that host-microbe interactions can be quite variable, even between closely related systems

Removal of Heterologous Sequences from Plasmodium falciparum Mutants Using FLPe-Recombinase

Genetically-modified mutants are now indispensable Plasmodium gene-function reagents, which are also being pursued as genetically attenuated parasite vaccines. Currently, the generation of transgenic malaria-parasites requires the use of drug-resistance markers. Here we present the development of an FRT/FLP-recombinase system that enables the generation of transgenic parasites free of resistance genes. We demonstrate in the human malaria parasite, P. falciparum, the complete and efficient removal of the introduced resistance gene. We targeted two neighbouring genes, p52 and p36, using a construct that has a selectable marker cassette flanked by FRT-sequences. This permitted the subsequent removal of the selectable marker cassette by transient transfection of a plasmid that expressed a 37°C thermostable and enhanced FLP-recombinase. This method of removing heterologous DNA sequences from the genome opens up new possibilities in Plasmodium research to sequentially target multiple genes and for using genetically-modified parasites as live, attenuated malaria vaccines

Gene Disruption of Plasmodium falciparum p52 Results in Attenuation of Malaria Liver Stage Development in Cultured Primary Human Hepatocytes

Difficulties with inducing sterile and long lasting protective immunity against malaria with subunit vaccines has renewed interest in vaccinations with attenuated Plasmodium parasites. Immunizations with sporozoites that are attenuated by radiation (RAS) can induce strong protective immunity both in humans and rodent models of malaria. Recently, in rodent parasites it has been shown that through the deletion of a single gene, sporozoites can also become attenuated in liver stage development and, importantly, immunization with these sporozoites results in immune responses identical to RAS. The promise of vaccination using these genetically attenuated sporozoites (GAS) depends on translating the results in rodent malaria models to human malaria. In this study, we perform the first essential step in this transition by disrupting, p52, in P. falciparum an ortholog of the rodent parasite gene, p36p, which we had previously shown can confer long lasting protective immunity in mice. These P. falciparum P52 deficient sporozoites demonstrate gliding motility, cell traversal and an invasion rate into primary human hepatocytes in vitro that is comparable to wild type sporozoites. However, inside the host hepatocyte development is arrested very soon after invasion. This study reveals, for the first time, that disrupting the equivalent gene in both P. falciparum and rodent malaria Plasmodium species generates parasites that become similarly arrested during liver stage development and these results pave the way for further development of GAS for human use

Improving statistical inference on pathogen densities estimated by quantitative molecular methods: malaria gametocytaemia as a case study

BACKGROUND: Quantitative molecular methods (QMMs) such as quantitative real-time polymerase chain reaction (q-PCR), reverse-transcriptase PCR (qRT-PCR) and quantitative nucleic acid sequence-based amplification (QT-NASBA) are increasingly used to estimate pathogen density in a variety of clinical and epidemiological contexts. These methods are often classified as semi-quantitative, yet estimates of reliability or sensitivity are seldom reported. Here, a statistical framework is developed for assessing the reliability (uncertainty) of pathogen densities estimated using QMMs and the associated diagnostic sensitivity. The method is illustrated with quantification of Plasmodium falciparum gametocytaemia by QT-NASBA. RESULTS: The reliability of pathogen (e.g. gametocyte) densities, and the accompanying diagnostic sensitivity, estimated by two contrasting statistical calibration techniques, are compared; a traditional method and a mixed model Bayesian approach. The latter accounts for statistical dependence of QMM assays run under identical laboratory protocols and permits structural modelling of experimental measurements, allowing precision to vary with pathogen density. Traditional calibration cannot account for inter-assay variability arising from imperfect QMMs and generates estimates of pathogen density that have poor reliability, are variable among assays and inaccurately reflect diagnostic sensitivity. The Bayesian mixed model approach assimilates information from replica QMM assays, improving reliability and inter-assay homogeneity, providing an accurate appraisal of quantitative and diagnostic performance. CONCLUSIONS: Bayesian mixed model statistical calibration supersedes traditional techniques in the context of QMM-derived estimates of pathogen density, offering the potential to improve substantially the depth and quality of clinical and epidemiological inference for a wide variety of pathogens

Evaluation and optimization of membrane feeding compared to direct feeding as an assay for infectivity

<p>Abstract</p> <p>Background</p> <p>Malaria parasite infectivity to mosquitoes has been measured in a variety of ways and setting, includind direct feeds of and/or membrane feeding blood collected from randomly selected or gametocytemic volunteers. <it>Anopheles gambiae s.l </it>is the main vector responsible of <it>Plasmodium falciparum </it>transmission in Bancoumana and represents about 90% of the laboratory findings, whereas <it>Plasmodium malariae </it>and <it>Plasmodium ovale </it>together represent only 10%.</p> <p>Materials and methods</p> <p>Between August 1996 and December 1998, direct and membrane feeding methods were compared for the infectivity of children and adolescent gametocyte carriers to anopheline mosquitoes in the village of Bancoumana in Mali. Gametocyte carriers were recruited twice a month through a screening of members of 30 families using Giemsa-stained thick blood smears. F1 generation mosquitoes issued from individual female wild mosquitoes from Bancoumana were reared in a controlled insectary conditions and fed 5% sugar solution in the laboratory in Bamako, until the feeding day when they are starved 12 hours before the feeding experiment. These F1 generation mosquitoes were divided in two groups, one group fed directly on gametocyte carriers and the other fed using membrane feeding method.</p> <p>Results</p> <p>Results from 372 <it>Plasmodium falciparum </it>gametocyte carriers showed that children aged 4–9 years were more infectious than adolescents (p = 0.039), especially during the rainy season. Data from 35 carriers showed that mosquitoes which were used for direct feeding were about 1.5 times more likely to feed (p < 0.001) and two times more likely to become infected, if they fed (p < 0.001), than were those which were used for membrane feeding. Overall, infectivity was about three-times higher for direct feeding than for membrane feeding (p < 0.001).</p> <p>Conclusion</p> <p>Although intensity of infectivity was lower for membrane feeding, it could be a surrogate to direct feeding for evaluating transmission-blocking activity of candidate malaria vaccines. An optimization of the method for future trials would involve using about three-times more mosquitoes than would be used for direct feeding.</p

The Transmembrane Isoform of Plasmodium falciparum MAEBL Is Essential for the Invasion of Anopheles Salivary Glands

Malaria transmission depends on infective stages in the mosquito salivary glands. Plasmodium sporozoites that mature in midgut oocysts must traverse the hemocoel and invade the mosquito salivary glands in a process thought to be mediated by parasite ligands. MAEBL, a homologue of the transmembrane EBP ligands essential in merozoite invasion, is expressed abundantly in midgut sporozoites. Alternative splicing generates different MAEBL isoforms and so it is unclear what form is functionally essential. To identify the MAEBL isoform required for P. falciparum (NF54) sporozoite invasion of salivary glands, we created knockout and allelic replacements each carrying CDS of a single MAEBL isoform. Only the transmembrane form of MAEBL is essential and is the first P. falciparum ligand validated as essential for invasion of Anopheles salivary glands. MAEBL is the first P. falciparum ligand experimentally determined to be essential for this important step in the life cycle where the vector becomes infectious for transmitting sporozoites to people. With an increasing emphasis on advancing vector-based transgenic methods for suppression of malaria, it is important that this type of study, using modern molecular genetic tools, is done with the agent of the human disease. Understanding what P. falciparum sporozoite ligands are critical for mosquito transmission will help validate targets for vector-based transmission-blocking strategies

The factors that influence the reading habit among Malaysian: a systematic literature review

Reading is an action that a person does to obtain a wider knowledge through his or her experience and this process may lead that person into a very eye-opening self-discovery. In Malaysia, reading habits rate is still at a lower level, Malaysian on average read only two books per year. About 80% of Malaysian university students are reluctant readers. They can read but choose not to read. Therefore, the objective of this study is to identify the factors affecting the reading habit among Malaysia. To answer the research question of this study we performed a systematic literature review (SLR). We defined a mapping study process adapted from existing guidelines to categorize and to structure the research evidence that has been published in the area of reading habits among Malaysian and world-wide. Twenty-Four papers were used in our synthesis of evidence, and five factors were identified. The results of this SLR showed the five factors that can affect the reading habits among Malaysian: (1) Internet (2) Environmental influence (3) Lack of motivation (4) Peer pressure (5) Entertainment. The findings of this SLR would be beneficial for understanding the needs of Malaysian towards the reading habit by looking at the factors, that might be a measurement of reading habit environment in other specific scopes

Comparative assessment of An. gambiae and An. stephensi mosquitoes to determine transmission-reducing activity of antibodies against P. falciparum sexual stage antigens.

BACKGROUND: With the increasing interest in vaccines to interrupt malaria transmission, there is a demand for harmonization of current methods to assess Plasmodium transmission in laboratory settings. Potential vaccine candidates are currently tested in the standard membrane feeding assay (SMFA) that commonly relies on Anopheles stephensi mosquitoes. Other mosquito species including Anopheles gambiae are the dominant malaria vectors for Plasmodium falciparum in sub-Saharan Africa. METHODS: Using human serum and monoclonal pre-fertilization (anti-Pfs48/45) and post-fertilization (anti-Pfs25) antibodies known to effectively inhibit sporogony, we directly compared SMFA based estimates of transmission-reducing activity (TRA) for An. stephensi and An. gambiae mosquitoes. RESULTS: In the absence of transmission-reducing antibodies, average numbers of oocysts were similar between An. gambiae and An. stephensi. Antibody-mediated TRA was strongly correlated between both mosquito species, and absolute TRA estimates for pre-fertilisation monoclonal antibodies (mAb) showed no significant difference between the two species. TRA estimates for IgG of naturally exposed individuals and partially effective concentrations of anti-Pfs25 mAb were higher for An. stephensi than for An. gambiae. CONCLUSION: Our findings support the use of An. stephensi in the SMFA for target prioritization. As a vaccine moves through product development, better estimates of TRA and transmission-blocking activity (TBA) may need to be obtained in epidemiologically relevant parasite-species combination