Reduction In Number Of Oocysts Research Articles

Gliding motility protein LIMP promotes optimal mosquito midgut traversal and infection by Plasmodium berghei

Substrate-dependent gliding motility is key to malaria transmission. It mediates host cell traversal, invasion and infection by Plasmodium and related apicomplexan parasites. The 110 amino acid-long cell surface protein LIMP is essential for P. berghei sporozoites where it is required for the invasion of the mosquito’s salivary glands and the liver cells of the rodent host. Here we define an additional role for LIMP during mosquito invasion by the ookinete. limp mRNA is provided as a translationally repressed mRNP (messenger ribonucleoprotein) by the female gametocyte and the protein translated in the ookinete. Parasites depleted of limp (Δlimp) develop ookinetes with apparent normal morphology and no defect during in vitro gliding motility, and yet display a pronounced reduction in oocyst numbers; compared to wildtype 82 % more Δlimp ookinetes remain within the mosquito blood meal explaining the decrease in oocysts. As in the sporozoite, LIMP exerts a profound role on ookinete infection of the mosquito.

An inter-laboratory comparison of standard membrane-feeding assays for evaluation of malaria transmission-blocking vaccines.

BackgroundAn effective malaria transmission-blocking vaccine may play an important role in malaria elimination efforts, and a robust biological assay is essential for its development. The standard membrane-feeding assay (SMFA) for Plasmodium falciparum infection of mosquitoes is considered a “gold standard” assay to measure transmission-blocking activity of test antibodies, and has been utilized widely in both non-clinical and clinical studies. While several studies have discussed the inherent variability of SMFA within a study group, there has been no assessment of inter-laboratory variation. Therefore, there is currently no assurance that SMFA results are comparable between different studies.MethodsMouse anti-Pfs25 monoclonal antibody (mAb, 4B7 mAb), rat anti-Pfs48/45 mAb (85RF45.1 mAb) and a human polyclonal antibody (pAb) collected from a malaria-exposed adult were tested at the same concentrations (6–94 μg/mL for 4B7, 1.2–31.3 μg/mL for 85RF45.1 and 23–630 μg/mL for human pAb) in two laboratories following their own standardized SMFA protocols. The mAbs and pAb, previously shown to have strong inhibition activities in the SMFA, were tested at three or four concentrations in two or three independent assays in each laboratory, and percent inhibition in mean oocyst intensity relative to a control in the same feed was determined in each feeding experiment.ResultsBoth monoclonal and polyclonal antibodies dose-dependently reduced oocyst intensity in all experiments performed at the two test sites. In both laboratories, the inter-assay variability in percent inhibition in oocyst intensity decreased at higher levels of inhibition, regardless of which antibody was tested. At antibody concentrations that led to a >80 % reduction in oocyst numbers, the inter-laboratory variations were in the same range compared with the inter-assay variation observed within a single laboratory, and the differences in best estimates from multiple feeds between the two laboratories were <5 percentage points.ConclusionsThis study confirms previous reports that the precision of the SMFA increases with increasing percent inhibition. Moreover, the variation between the two laboratories is not greater than the variation observed within a laboratory. The findings of this study provide guidance for comparison of SMFA data from different laboratories.Electronic supplementary materialThe online version of this article (doi:10.1186/s12936-016-1515-z) contains supplementary material, which is available to authorized users.

Synergistic approach for treatment of chicken coccidiosis using berberine – A plant natural product

Despite the advent of anticoccidial drugs and vaccines, coccidiosis continues to result in substantial economic losses to the poultry industry. Berberine, a natural alkaloid is well known in studies involving synergistic approaches, thereby reducing the dosage of principal drugs. Therefore, a study was designed to see whether a synergistic anticoccidial effect could be obtained between amprolium and berberine, in vivo using broiler chicken. Anticoccidial activity was measured in comparison to the reference drug amprolium on the basis of oocyst output reduction, mean weight gain and feed conversion ratio. Oocyst output was measured using Mc-Masters counting technique. Different combinations of berberine and amprolium were tested and out of which 1:1 ratio was the most effective for controlling these parasites. Oral gavaging of 100(50 + 50) mg/kg body weight of 1:1 ratio of amprolium and berberine caused the equivalent reduction in number of oocysts (38.85 ± 9.61) one day prior to that of standard drug amprolium (49.95 ± 16.65) as well as pure berberine (44.4 ± 9.61) used in the study. Weight gain of birds was also highest in the synergistic group (1547.43 ± 12.86) among all the infected groups. Besides feed conversion ratio in the synergistic group was also better (1.387 ± 0.026). The results of this study proved the effectiveness of both amprolium and berberine and revealed synergism between amprolium and berberine against coccidian oocysts, confirmed by significant reduction in the number of coccidian oocysts shed in the feces, leading to better weight gain and improved feed conversion ratio. The study deep-rooted the synergistic potential of berberine, a natural bioactive compound for controlling a protozoan parasite and the results of this study corroborate with its use for treatment of severe diarrhoea, amoebiasis and intestinal infections.

Perforin-like protein PPLP4 is crucial for mosquito midgut infection by Plasmodium falciparum

The genomes of Plasmodium parasites encode for five perforin-like proteins, PPLP1-5, and four of them have previously been demonstrated to be involved in disruption of host cell barriers. We now show that the fifth perforin, PPLP4, is crucial for infection of the mosquito vector by Plasmodium falciparum parasites. PPLP4 is expressed in the blood and mosquito midgut stages in granular structures. In gametocytes, PPLP4 expression is specific to the female gender, while ookinetes show a PPLP4 localization at the apical pole. Gene disruption of pplp4 results in no phenotypical change during blood stage replication, gametocyte development or gametogenesis, while mosquitoes fed with PPLP4-deficient gametocytes display a severe reduction in oocyst numbers, and an accumulation of ookinetes in the mosquito midguts was observed. In conclusion, we propose an essential role for PPLP4 in infection of the mosquito midgut, presumably by mediating ookinete traversal through the midgut epithelium.

Plasmodium falciparum Produce Lower Infection Intensities in Local versus Foreign Anopheles gambiae Populations

Both Plasmodium falciparum and Anopheles gambiae show great diversity in Africa, in their own genetic makeup and population dynamics. The genetics of the individual mosquito and parasite are known to play a role in determining the outcome of infection in the vector, but whether differences in infection phenotype vary between populations remains to be investigated. Here we established two A. gambiae s.s. M molecular form colonies from Cameroon and Burkina Faso, representing a local and a foreign population for each of the geographical sites. Experimental infections of both colonies were conducted in Cameroon and Burkina Faso using local wild P. falciparum, giving a sympatric and allopatric vector-parasite combination in each site. Infection phenotype was determined in terms of oocyst prevalence and intensity for at least nine infections for each vector-parasite combination. Sympatric infections were found to produce 25% fewer oocysts per midgut than allopatric infections, while prevalence was not affected by local/foreign interactions. The reduction in oocyst numbers in sympatric couples may be the result of evolutionary processes where the mosquito populations have locally adapted to their parasite populations. Future research on vector-parasite interactions must take into account the geographic scale of adaptation revealed here by conducting experiments in natural sympatric populations to give epidemiologically meaningful results.

Transmission-blocking activity induced by malaria vaccine candidates Pfs25/Pvs25 is a direct and predictable function of antibody titer

BackgroundMosquito stage malaria vaccines are designed to induce an immune response in the human host that will block the parasite's growth in the mosquito and consequently block transmission of the parasite. A mosquito membrane-feeding assay (MFA) is used to test transmission-blocking activity (TBA), but in this technique cannot accommodate many samples. A clear understanding of the relationship between antibody levels and TBA may allow ELISA determinations to be used to predict TBA and assist in planning vaccine development.MethodsRabbit anti-Pfs25 sera and monkey anti-Pvs25 sera were generated and the antibody titers were determined by a standardized ELISA. The biological activity of the same sera was tested by MFA using Plasmodium gametocytes (cultured Plasmodium falciparum or Plasmodium vivax from malaria patients) and Anopheles mosquitoes.ResultsAnti-Pfs25 and anti-Pvs25 sera showed that ELISA antibody units correlate with the percent reduction in the oocyst density per mosquito (Spearman Rank correlations: 0.934 and 0.616, respectively), and fit a hyperbolic curve when percent reduction in oocyst density is plotted against antibody units of the tested sample. Antibody levels also correlated with the number of mosquitoes that failed to become infected, and this proportion can be calculated from the reduction in oocyst numbers and the distribution of oocysts per infected mosquito in control group.ConclusionELISA data may be used as a surrogate for the MFA to evaluate transmission-blocking vaccine efficacy. This will facilitate the evaluation of transmission-blocking vaccines and implementation of this malaria control strategy.

Induction of Plasmodium falciparum transmission-blocking antibodies in nonhuman primates by a combination of DNA and protein immunizations.

Malaria transmission-blocking vaccination can effectively reduce and/or eliminate transmission of parasites from the human host to the mosquito vector. The immunity achieved by inducing an antibody response to surface antigens of male and female gametes and parasite stages in the mosquito. Our laboratory has developed DNA vaccine constructs, based on Pfs25 (a Plasmodium falciparum surface protein of 25 kDa), that induce a transmission-blocking immune response in mice (C. A. Lobo, R. Dhar, and N. Kumar, Infect. Immun. 67:1688-1693, 1999). To evaluate the safety, immunogenicity, and efficacy of the Pfs25 DNA vaccine in nonhuman primates, we immunized rhesus macaques (Macaca mulatta) with a DNA vaccine plasmid encoding Pfs25 or a Pfg27-Pfs25 hybrid or with the plasmid (empty plasmid) alone. Immunization with four doses of these DNA vaccine constructs elicited antibody titers that were high but nonetheless unable to reduce the parasite's infectivity in membrane feeding assays. Further boosting of the antibody response with recombinant Pfs25 formulated in Montanide ISA-720 increased antibody titers (30-fold) and significantly blocked transmission of P. falciparum gametocytes to Anopheles mosquitoes (approximately 90% reduction in oocyst numbers in the midgut). Our data show that a DNA prime-protein boost regimen holds promise for achieving transmission-blocking immunity in areas where malaria is endemic and could be effective in eradicating malaria in isolated areas where the level of malaria endemicity is low.

The effects of Nosema algerae on the development of Plasmodium yoelii nigeriensis in Anopheles stephensi.

Experimental simultaneous infections of Anopheles stephensi (Diptera: Culicidae) with Nosema algerae (Microsporida: Nosematidae) and Plasmodium yoelii nigeriensis under standardized laboratory conditions showed partial suppression of the malaria parasite. At 9 days after an infective bloodmeal, the oocysts in the midgut were counted; 12.1%-66.6% of the double-infected mosquitoes exhibited no oocysts, whereas only 4.5%-12% of the control group showed no oocysts. The mean reduction in oocyst numbers under the influence of Nosema was 84.68%. At 14 days after infection with Plasmodium, the amount of sporozoites was examined; their mean reduction in eight experiments was 70%.