Geographic Structure of Plasmodium vivax: Microsatellite Analysis of Parasite Populations from Sri Lanka, Myanmar, and Ethiopia

Genetic diversity and population structure of Plasmodium viva-V parasites call predict the origin and Spread of novel Variants Within a population enabling Population specific malaria control measures. We analyzed the genetic diversity and population Structure of 425 P. vivax isolates from Sri Lanka, Myanmar, and Ethiopia using 12 trinucleotide and tetranucleotide microsatellite markers. All three parasite populations were highly polymorphic with 3-44 alleles per locus. Approximately 65% were multiple-clone infections. Mean genetic diversity (H(E)) was 0.7517 in Ethiopia, 0.8450 in Myanmar, and 0.8610 in Sri Lanka. Significant linkage disequilibrium Was maintained. Population structure showed two clusters (Asian and African) according to geography and ancestry Strong clustering of outbreak isolates from Sri Lanka and Ethiopia was observed. Predictive power of ancestry using two-thirds of the isolates as a model identified 78.2% of isolates accurately as being African or Asian. Microsatellite analysis is a useful tool for mapping short-term outbreaks of malaria and for predicting ancestry.

Comparative genomics of the neglected human malaria parasite Plasmodium vivax

The human malaria parasite Plasmodium vivax is responsible for 25 - 40% of the similar to 515 million annual cases of malaria worldwide. Although seldom fatal, the parasite elicits severe and incapacitating clinical symptoms and often causes relapses months after a primary infection has cleared. Despite its importance as a major human pathogen, P. vivax is little studied because it cannot be propagated continuously in the laboratory except in non- human primates. We sequenced the genome of P. vivax to shed light on its distinctive biological features, and as a means to drive development of new drugs and vaccines. Here we describe the synteny and isochore structure of P. vivax chromosomes, and show that the parasite resembles other malaria parasites in gene content and metabolic potential, but possesses novel gene families and potential alternative invasion pathways not recognized previously. Completion of the P. vivax genome provides the scientific community with a valuable resource that can be used to advance investigation into this neglected species.

Experimental infection of Leishmania chagasi in immunosuppressed balb/c mice: cellular immune response and parasite burden

The immune response to leishmaniasis can result in a polarization of a subpopulation of T lymphocytes, which leads to a different cell phenotype and results in immune protection or exacerbation of the disease. Leishmanias persist in the body both in asymptomatic infections and after treatment, representing risks in terms of immunosuppression. The objective of this study was to evaluate the effects of infection and immunosuppression by dexamethasone associated with pentoxifylline on animal weight, spleen weight, the parasitic load in the spleen and liver, as well as the production of IFN-gamma and IL-10 in spleen cell culture of Balb/c mice infected with Leishmania chagasi. The infection did not alter animal weight gain, but spleen weight and size increased. The immunosuppression, induced by dexamethasone associated with pentoxifylline, affected animal weight gain and weight and size of the spleen (in infected and not infected animals). The immunosuppression did not significantly alter the course of the parasite burden in the spleen and liver. Dexamethasone and pentoxifylline affected the studied cytokine production, but not influenced on Th1/Th2 response in infected animals.

Evaluation of TNF-alpha, IL-4, and IL-10 and parasite density in spleen and liver of L. (L.) chagasi naturally infected dogs

Dogs are the main domestic reservoirs of L. (L.) chagasi. Once in the vertebrate host, the parasite can cause visceral leishmaniasis, which can also be transmitted to humans. Cytokines are key elements of the host immune response against Leishmania spp. To investigate whether tumor necrosis factor (TNF)-alpha, interleukin (IL)-4 and IL-10 are associated with pattern infection in dogs, these cytokines were quantified in the spleen and liver of dogs naturally infected with L. (L.) chagasi, with or without clinical manifestations, and their levels were correlated with the parasite load verified in these organs. A total of 40 adult dogs naturally infected with L. (L.) chagasi were assessed, together with 12 uninfected control dogs. Samples from spleen and liver were used to determine the cytokine levels by capture ELISA and for quantifying parasite load by real-time PCR. Statistical analysis was performed using the minimum Chi square method and group means were compared using the Tukey test. TNF-alpha, IL-4 and IL-10 levels in infected dogs were higher than in control groups; the liver was the main cytokine-producing organ during infection. The level of splenic TNF-alpha showed correlation with parasite load and may represent an important marker for infection process evolution, with the participation of IL-10. These results may contribute to a clearer understanding of the immune response in dogs infected with L. (L.) chagasi, which may lead to the development of prophylactic or preventive measures for these animals.

Relationship of abomasal histology and parasite-specific immunoglobulin A with the resistance to Haemonchus contortus infection in three breeds of sheep

This study was carried out to evaluate the relationship of abomasal inflammatory cells and parasite-specific immunoglobulin A (IgA) in mucus, with the resistance to Haemonchus contortus infection in three breeds of sheep naturally infected with gastrointestinal nematodes. The breeds were the native Santa Ines sheep, and the European Suffolk and Ile de France breeds. Mast cells, eosinophils and globule leucocytes were enumerated in abomasal mucosa. Eosinophils within the sub-mucosa also were counted separately. Histamine concentration was estimated in abomasal tissue samples. Enzyme-linked immunosorbent assay was carried out in mucus samples to determine the level of IgA anti-H. contortus third and fifth instar. There were no significant differences among group means of these variables (P > 0.05). The correlation coefficients between fecal egg counts (FEC) x mast cells (r = -0.490; P < 0.05) and FEC x eosinophils in sub-mucosa (r = -0.714; P < 0.01) was significant in the Santa Ines sheep. In the Ile de France group, the correlation coefficients between globule leucocytes x FEC (r = -0.879; P < 0.001) and histamine x worm burden (r = -0.833; P < 0.01) were also significant. In the Santa Ines and Ile de France sheep, correlation coefficients between IgA anti-L3 x worm burden and IgA anti-L3 x FEC were negative. In general, inflammatory cells and IgA-parasite-specific in abomasum were inversely associated with H. contortus worm burden and FEC indicating that they may impair parasite development or fecundity in the three breeds of sheep. However, similar mean values of inflammatory cells and IgA were found in the resistant (Santa Ines) and in the susceptible (Suffolk and Ile de France) breeds of sheep. The enumeration of cells by histological assessment does not provide information on their functional activity, which may be different among breeds. Thus, the effect of breed on the functional activity of these and other inflammatory cells is an important area for further study. (c) 2004 Elsevier B.V. All rights reserved.

In vivo uptake of a haem analogue Zn protoporphyrin IX by the human malaria parasite P. falciparum-infected red blood cells

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Automated Nuclear Analysis of Leishmania major Telomeric Clusters Reveals Changes in Their Organization during the Parasite's Life Cycle

Parasite virulence genes are usually associated with telomeres. The clustering of the telomeres, together with their particular spatial distribution in the nucleus of human parasites such as Plasmodium falciparum and Trypanosoma brucei, has been suggested to play a role in facilitating ectopic recombination and in the emergence of new antigenic variants. Leishmania parasites, as well as other trypanosomes, have unusual gene expression characteristics, such as polycistronic and constitutive transcription of protein-coding genes. Leishmania subtelomeric regions are even more unique because unlike these regions in other trypanosomes they are devoid of virulence genes. Given these peculiarities of Leishmania, we sought to investigate how telomeres are organized in the nucleus of Leishmania major parasites at both the human and insect stages of their life cycle. We developed a new automated and precise method for identifying telomere position in the three-dimensional space of the nucleus, and we found that the telomeres are organized in clusters present in similar numbers in both the human and insect stages. While the number of clusters remained the same, their distribution differed between the two stages. The telomeric clusters were found more concentrated near the center of the nucleus in the human stage than in the insect stage suggesting reorganization during the parasite's differentiation process between the two hosts. These data provide the first 3D analysis of Leishmania telomere organization. The possible biological implications of these findings are discussed.

Comparison of naturally acquired parasite burdens among Florida Native, Rambouillet and crossbreed ewes

The response to natural infection by gastrointestinal nematodes was evaluated in three groups of lambing mature ewes: 16 Florida Native; 21 Rambouillet; and 18 1/2 Florida Native x 1/2 Rambouillet (F1 generation). Fecal egg counts (FEC),packed cell volume (PCV) and plasma protein levels were determined periodically through a one-year period. The highest FEC was recorded in Rambouillet ewes, with most of the means higher than 1000 eggs per gram (EPG). Florida Native and Fl ewes showed similar trends in FEC, with most of the means below 500 EPG. The highest mean PCV values were recorded in Florida Native ewes, followed in decreasing order by Fl and Rambouillet ewes. Fl ewes showed the lowest plasma protein levels and Florida Natives the highest. Fecal cultures revealed the highest percentages of Haemonchus larvae in Rambouillet ewes (>64%) and the lowest percentages in Florida Native ewes. Florida Native and Fl ewes were more resistant to gastrointestinal nematode infections than were Rambouillet ewes. High negative correlation coefficients between FEC x PCV and FEC x plasma protein levels coincided with peaks in mean FEC of Rambouillet (May and July), Florida Native and Fl ewes (October). This study showed that crossbreeding Rambouillet and Florida Native sheep resulted in the production of Fl ewes resistant against gastrointestinal nematode infections. (C) 1999 Elsevier B.V. B.V. All rights reserved.

Electrochemical genosensors for the detection of Bonamia parasite. Selection of single strand-DNA (ssDNA) probes by simulation of the secondary structure folding

A post-PCR nucleic acid work by comparing experimental data, from electrochemical genosensors, and bioinformatics data, derived from the simulation of the secondary structure folding and prediction of hybridisation reaction, was carried out in order to rationalize the selection of ssDNA probes for the detection of two Bonamia species, B. exitiosa and B. ostreae, parasites of Ostrea edulis.Six ssDNA probes (from 11 to 25 bases in length, 2 thiolated and 4 biotinylated) were selected within different regions of B. ostreae and B. exitiosa PCR amplicons (300 and 304 bases, respectively) with the aim to discriminate between these parasite species. ssDNA amplicons and probes were analyzed separately using the "Mfold Web Server" simulating the secondary structure folding behaviour. The hybridisation of amplicon-probe was predicted by means of "Dinamelt Web Server". The results were evaluated considering the number of hydrogen bonds broken and formed in the simulated folding and hybridisation process, variance in gaps for each sequence and number of available bases. In the experimental part, thermally denatured PCR products were captured at the sensor interface via sandwich hybridisation with surface-tethered probes (thiolated probes) and biotinylated signalling probes. A convergence between analytical signals and simulated results was observed, indicating the possibility to use bioinformatic data for ssDNA probes selection to be incorporated in genosensors. (C) 2011 Elsevier B.V. All rights reserved.

Description of Leishmania (Leishmania) forattinii sp. n., a new parasite infecting opossums and rodents in Brazil

A new parasite species of Leishmania is described, L. (Leishmania) forattinii sp. n., which was isolated from a pooled triturate of liver and spleen of a opossum (Didelphis marsupialis aurita) and from skin samples from a rodent (Proechmys iheringi denigratus), captured in primary forest on the Atlantic Cost of Brazil. Our results on the basis of biological and molecular criteria indicate that this taxonomically distinct parasite ias a new species of the L. mexicana complex, but closely related to L. (L.) aristidesi Laison & shaw, 1979, as revelated by phenetic and phylogenetic numerical analyses of the enzyme data. L. forattinii was clearly distinguishable from other Leishmania species of the genus usisng enzyme electrophoresis, monoclonal antibodies, molecular karyotypes, analysis of restriction enzyme digestion patterns of kinetoplast DNA (kDNA), as well as the use of kDNA hybridization procedures.

Characterization of an unidentified Sarcocystis falcatula-like parasite from the South American opossum, Didelphis albiventris from Brazil

An unidentified isolate of a Sarcocystis falcatula-like parasite was obtained from the lungs of budgerigars (Melopsittacus undulatus) fed sporocysts from a naturally-infected South American opossum, Didelphis albiventris from Brazil. Four captive budgerigars fed sporocysts from the opossum intestine died of acute sarcocystosis 8, 10, and 12 days after oral inoculation (DAI); one budgerigar was killed 12 DAI when it was lethargic. Schizonts and merozoites found in the lungs of the budgerigars reacted mildly with polyclonal S. falcatula antibody. The parasite was isolated in equine kidney cell cultures inoculated with lung tissue from a budgerigar that was killed 12 DAI. Two budgerigars inoculated subcutaneously with 100,000 culture-derived S. falcatula merozoites developed acute sarcocystosis and S. falcatula-like schizonts were found in their lungs 15 and 16 DAI. Four budgerigars kept as unfed controls in the same environment remained free of Sarcocystis infection. The parasite underwent schizogony in African green monkey kidney cells and bovine turbinate cells. Merozoites divided by endopolygeny, often leaving a residual body. Polymerase chain reaction studies using primers JNB33/JNB54 and Hinf I and Dra I digestion indicated that the isolate was not S. falcatula. Results of this study indicated that the South American opossum, D. albiventris, is a definitive host for yet another S. falcatula-like parasite.

Prevalence of Sarcocystis neurona and Neospora spp. infection in horses from Brazil based on presence of serum antibodies to parasite surface antigen

Sera from 961 horses from Brazil were tested for antibodies against the major surface antigens SnSAG4 and NhSAG1 to determine the seroprevalence of Sarcocystis neurona and Neospora hughesi, respectively. Antibodies against SnSAG4 were detected in 669 (69.6%) of the horses, while antibodies against NhSAG1 were detected in only 24 (2.5%) of the horses. These serologic results suggest that there is a high concentration of S. neurona in the environment of Brazil, which results in marked exposure of horses to this parasite. Additionally, the data further confirm that infection with Neospora spp. is relatively uncommon in horses. (c) 2005 Elsevier B.V. All tights reserved.

Purine nucleoside phosphorylase: A potential target for the development of drugs to treat T-cell- and apicomplexan parasite-mediated diseases

Purine nucleoside phosphorylase (PNP) catalyzes the reversible phosphorolysis of nucleosides and deoxynucleosides, generating ribose 1-phosphate and the purine base, which is an important step of purine catabolism pathway. The lack of such an activity in humans, owing to a genetic disorder, causes T-cell impairment, and thus drugs that inhibit human PNP activity have the potential of being utilized as modulators of the immunological system to treat leukemia, autoimmune diseases, and rejection in organ transplantation. Besides, the purine salvage pathway is the only possible way for apicomplexan parasites to obtain the building blocks for RNA and DNA synthesis, which makes PNP from these parasites an attractive target for drug development against diseases such as malaria. Hence, a number of research groups have made efforts to elucidate the mechanism of action of PNP based on structural and kinetic studies. It is conceivable that the mechanism may be different for PNPs from diverse sources, and influenced by the oligomeric state of the enzyme in solution. Furthermore, distinct transition state structures can make possible the rational design of specific inhibitors for human and apicomplexan enzymes. Here, we review the current status of these research efforts to elucidate the mechanism of PNP-catalyzed chemical reaction, focusing on the mammalian and Plamodium falciparum enzymes, targets for drug development against, respectively, T-Cell and Apicomplexan parasites-mediated diseases.

Biological control of sheep parasite nematodes by nematode-trapping fungi: In vitro activity and after passage through the gastrointestinal tract

The main method used for the control of gastrointestinal nematodes in sheep production is the application of chemotherapeutic agents, which often lead to the selection of parasites resistant to given active principles. Biological control can be considered a promising alternative, contributing to an increase in the efficacy of verminous control. We determined the in vitro activity and in situ survival of the predatory fungi Arthrobotrys musiformis and Arthrobotrys conoides during passage through the gastrointestinal tract of sheep after oral administration of conidia in microencapsulated form and as a liquid in natura. Initial in vitro tests showed that both fungi were efficient in the predation of trichostrongylid L3 larvae present in the faeces of sheep naturally infected with gastrointestinal nematodes. The fungi presented high nematophagous activity, which was 99.3% for A. conoides and 73.7% for A. musiformis. A. conoides did not survive passage through the gastrointestinal tract under the conditions of the present experiment. On the other hand, A. musiformis was reisolated after administration in either microencapsulated or liquid form, suggesting that this species is a promising alternative for the control of nematodes in sheep since it survives without any protection (in natura). © Springer 2005.

Ant-tended hemiptera in Amazonian myrmecophytes: Patterns of abundance and implications for mutualism function (Hymenoptera: Formicidae)

We assessed how the abundance of ant-tended Hemiptera associated with two Amazonian myrmecophytes, Tococa bullifera and Maieta guianensis, varied as a function of resident ant species. We collected five species or morpho-species of adult hemiptera in the domatia of M. guianensis, with four of these species also found in Tococa bullifera. Maieta guianensis plants inhabited by Crematogaster laevis had over four-fold more hemiptera in them than plants inhabited by Pheidole minutula. In contrast, the density of hemiptera in Tococa bullifera domatia was independent of the species of ant resident. For each of the two ant species inhabiting Maieta guianensis, there was a positive and significant relationship between the abundance of hemiptera and workers inhabiting a plant. This relationship was also significant and positive for the Tococa bullifera plants inhabited by C. laevis. However, there was no relationship between Azteca worker and hemipteran density, although there was a trend towards a positive relationship. Our results indicate that hemipteran abundance can vary significantly between different myrmecophyte species, but that the nature of this relationship is mediated by the identity of the ant associate. Because hemipterans are herbivores, the costs and benefits of different ant partners to the host plant may vary in ways that are often overlooked.