Prevalence of asymptomatic P. falciparum gametocyte carriage among school children in Mbita, Western Kenya and assessment of the association between gametocyte density, multiplicity of infection and mosquito infection prevalence.

Background: Asymptomatic Plasmodium falciparum gametocyte carriers are reservoirs for sustaining transmission in malaria endemic regions. Gametocyte presence in the host peripheral blood is a predictor of capacity to transmit malaria. However, it does not always directly translate to mosquito infectivity. Factors that affect mosquito infectivity include, gametocyte sex-ratio and density, multiplicity of infection (MOI), and host and vector anti-parasite immunity. We assess the prevalence of gametocyte carriage and some of its associated risk factors among asymptomatic schoolchildren in Western Kenya and to further analyse the association between gametocyte density, multiplicity of infection (MOI) and mosquito infection prevalence. Methods: P. falciparum parasite infections were detected by RDT (Rapid Diagnostic Test) and microscopy among schoolchildren (5-15 years old). Blood from 37 microscopy positive gametocyte carriers offered to laboratory reared An. gambiae s.l. mosquitoes. A total of 3395 fully fed mosquitoes were screened for Plasmodium sporozoites by ELISA. P. falciparum was genotyped using 10 polymorphic microsatellite markers. The association between MOI and gametocyte density and mosquito infection prevalence was investigated. Results: A significantly higher prevalence of P. falciparum infection was found in males 31.54% (764/2422) (p-value < 0.001) compared to females 26.72% (657/2459). The microscopic gametocyte prevalence among the study population was 2% (84/4881). Children aged 5-9 years have a higher prevalence of gametocyte carriage (odds ratios = 2.1 [95% CI = 1.3–3.4], P = 0.002) as compared to children aged 10-15 years. After offering gametocyte positive blood to An. gambiae s.l. by membrane feeding assay, our results indicated that 68.1% of the variation in mosquito infection prevalence was accounted for by gametocyte density and MOI (R-SQR. = 0.681, p < 0.001). Conclusions: We observed a higher risk of gametocyte carriage among the younger children (5-9 years). Gametocyte density and MOI significantly predicted mosquito infection prevalence.

Gametocyte carriage after seasonal malaria chemoprevention in Plasmodium falciparum infected asymptomatic children

Background Treatment of clinical Plasmodium falciparum malaria with sulfadoxine-pyrimethamine (SP) and amodiaquine (AQ) is associated with increased post-treatment gametocyte carriage. The effect of seasonal malaria chemoprevention (SMC) with SP and AQ on gametocyte carriage was assessed in asymptomatic P. falciparum infected children. Methods The study was carried out in eastern Gambia. Asymptomatic P. falciparum malaria infected children aged 24–59 months old who were eligible to receive SMC (SMC group) and children 5–8 years that were not eligible to receive SMC (comparison group) were recruited. Gametocytaemia was determined by molecular methods before and after SMC administration. Gametocyte carriage between the groups was compared using the chi-squared test and within-person using conditional logistic regression. Results During the 2017 and 2018 malaria transmission seasons, 65 and 75 children were recruited in the SMC and comparison groups, respectively. Before SMC administration, gametocyte prevalence was 10.7% (7/65) in the SMC group and 13.3% (10/75) in the comparison group (p = 0.64). At day 13 (IQR 12, 13) after SMC administration, this was 9.4% (5/53) in children who received at least the first dose of SMC treatment and 12.7% (9/71) for those in the comparison group (p = 0.57). Similarly, there was no difference in prevalence of gametocytes between children that adhered to all 3-day doses of SMC treatment 15.6% (5/32) and those in the comparison group (p = 0.68). In the SMC group, within-group gametocyte carriage was similar before and after SMC administration in children that received at least the first dose of SMC treatment (OR 0.6, 95% CI 0.14–2.51; p = 0.48) and in those that adhered to all 3-day doses of SMC treatment (OR 1.0, 95% CI 0.20–4.95; p = 1.0). Conclusion In this study with relative low gametocyte prevalence prior to SMC treatment, no evidence was observed that SMC treatment increased gametocyte carriage in asymptomatic P. falciparum malaria infected children.

Asymptomatic Plasmodium infection and associated factors among pregnant women in the Merti district, Oromia, Ethiopia

Background Asymptomatic Plasmodium infection (API) that occurs during pregnancy increases the risk of stillbirths, abortion, premature delivery, and low birth weight. API also hinders the control and prevention of malaria as infected hosts serve as silent reservoirs for transmission of Plasmodium species in the community. Objective The aim of this study was to determine the prevalence of API and associated factors among pregnant women. This community-based cross-sectional study was conducted at Merti district, Oromia, Ethiopia among 364 pregnant women from March to September 2018. Methods Sociodemographic and obstetrics features were collected using a structured questionnaire. About 2ml of blood was collected from participants to detect Plasmodium species, gametocyte carriage rate, parasite density, and anemia. Results The prevalence of API among pregnant women was 3.6%. The proportion of Plasmodium falciparum and Plasmodium vivax was 6(46.2%) and 7(53.8%) respectively. Out of 13 Plasmodium species identified, Gametocyte carriage rate was 4(30.7%). The geometric mean density of the asexual stage of the parasites was 994.7(interquartile [IQR], 320 to 2200) parasites/ul. The geometric mean gametocyte density was 303.3 (interquartile range [IQR], 160 to 600). The proportion of anemia among Plasmodium-infected participants was 12(92.3%). Previous infection by Plasmodium species (AOR = 5.42; 95% CI: 1.19–29.03, p = 0.047), lack of insecticide-treated bed net use (AOR = 6.52; 95% CI: 1.17–36.44, p = 0.032), and living close to stagnant water (AOR = 4.18; 95% CI (1.12–17.36, p = 0.049) were significantly associated with API. Anemia was significantly higher among Plasmodium-infected than non-infected pregnant women (x2 = 27.62, p <0.001). Conclusion In the current study, a relatively high prevalence of API was detected among pregnant women. Identifying API in the community is important to prevent the unwanted outcomes of Plasmodium infection and its transmission.

HIV-1 Infection Is Associated With Increased Prevalence and Abundance of Plasmodium falciparum Gametocyte-Specific Transcripts in Asymptomatic Adults in Western Kenya

As morbidity and mortality due to malaria continue to decline, the identification of individuals with a high likelihood of transmitting malaria is needed to further reduce the prevalence of malaria. In areas of holoendemic malaria transmission, asymptomatically infected adults may be infected with transmissible gametocytes. The impact of HIV-1 on gametocyte carriage is unknown, but co-infection may lead to an increase in gametocytemia. In this study, a panel of qPCR assays was used to quantify gametocyte stage-specific transcripts present in dried blood spots obtained from asymptomatic adults seeking voluntary HIV testing in Kombewa, Kenya. A total of 1,116 Plasmodium-specific 18S-positive samples were tested and 20.5% of these individuals had detectable gametocyte-specific transcripts. Individuals also infected with HIV-1 were 1.82 times more likely to be gametocyte positive (P&lt;0.0001) and had significantly higher gametocyte copy numbers when compared to HIV-negative individuals. Additionally, HIV-1 positivity was associated with higher gametocyte prevalence in men and increased gametocyte carriage with age. Overall, these data suggest that HIV-positive individuals may have an increased risk of transmitting malaria parasites in regions with endemic malaria transmission and therefore should be at a higher priority for treatment with gametocidal antimalarial drugs.

Antibody Responses to Crude Gametocyte Extract Predict Plasmodium falciparum Gametocyte Carriage in Kenya

BackgroundMalaria caused by Plasmodium falciparum remains a serious global public health challenge especially in Africa. Interventions that aim to reduce malaria transmission by targeting the gametocyte reservoir are key to malaria elimination and/or eradication. However, factors that are associated with gametocyte carriage have not been fully explored. Consequently, identifying predictors of the infectious reservoir is fundamental in the elimination campaign.MethodsWe cultured P. falciparum NF54 gametocytes (to stage V) and prepared crude gametocyte extract. Samples from a total of 687 participants (aged 6 months to 67 years) representing two cross-sectional study cohorts in Kilifi, Kenya were used to assess IgG antibody responses by ELISA. We also analyzed IgG antibody responses to the blood-stage antigen AMA1 as a marker of asexual parasite exposure. Gametocytemia and asexual parasitemia data quantified by microscopy and molecular detection (QT-NASBA) were used to determine the relationship with antibody responses, season, age, and transmission setting. Multivariable logistic regression models were used to study the association between antibody responses and gametocyte carriage. The predictive power of the models was tested using the receiver operating characteristic (ROC) curve.ResultsMultivariable logistic regression analysis showed that IgG antibody response to crude gametocyte extract predicted both microscopic (OR=1.81 95% CI: 1.06–3.07, p=0.028) and molecular (OR=1.91, 95% CI: 1.11–3.29, p=0.019) P. falciparum gametocyte carriage. Antibody responses to AMA1 were also associated with both microscopic (OR=1.61 95% CI: 1.08–2.42, p=0.020) and molecular (OR=3.73 95% CI: 2.03–6.74, p&lt;0.001) gametocytemia. ROC analysis showed that molecular (AUC=0.897, 95% CI: 0.868–0.926) and microscopic (AUC=0.812, 95% CI: 0.758–0.865) multivariable models adjusted for gametocyte extract showed very high predictive power. Molecular (AUC=0.917, 95% CI: 0.891–0.943) and microscopic (AUC=0.806, 95% CI: 0.755–0.858) multivariable models adjusted for AMA1 were equally highly predictive.ConclusionIn our study, it appears that IgG responses to crude gametocyte extract are not an independent predictor of gametocyte carriage after adjusting for AMA1 responses but may predict gametocyte carriage as a proxy marker of exposure to parasites. Serological responses to AMA1 or to gametocyte extract may facilitate identification of individuals within populations who contribute to malaria transmission and support implementation of transmission-blocking interventions.

Plasmodium falciparum gametocyte carriage in symptomatic patients shows significant association with genetically diverse infections, anaemia, and asexual stage density

Background Multi-genotype malaria infections are frequent in endemic area, and people commonly harbour several genetically distinct Plasmodium falciparum variants. The influence of genetic multiplicity and whether some specific genetic variants are more or less likely to invest into gametocyte production is not clearly understood. This study explored host and parasite-related risk factors for gametocyte carriage, and the extent to which some specific P. falciparum genetic variants are associated with gametocyte carriage. Methods Gametocytes and asexual forms were detected by light microscopy on thick smears collected between 2010 and 2012 in Nanoro, Burkina Faso. Merozoite surface protein 1 and 2 were genotyped by nested PCR on clinical samples. Associations between gametocyte carriage and factors, including multiplicity of infection, parasite density, patient age, gender, haemoglobin (Hb) level, and body temperature were assessed. The relationship between the presence of a particular msp1 and msp2 genetic variants and gametocyte carriage was also explored. Results Of the 724 samples positive to P. falciparum and successfully genotyped, gametocytes were found in 48 samples (6.63%). There was no effect of patient gender, age and body temperature on gametocyte carriage. However, the probability of gametocyte carriage significantly increased with increasing values of multiplicity of infection (MOI). Furthermore, there was a negative association between parasite density and gametocyte carriage. MOI decreased with parasite density in gametocyte-negative patients, but increased in gametocyte carriers. The probability of gametocyte carriage decreased with Hb level. Finally, the genetic composition of the infection influenced gametocyte carriage. In particular, the presence of RO33 increased the odds of developing gametocytes by 2 while the other allelic families K1, MAD20, FC27, and 3D7 had no significant impact on the occurrence of gametocytes in infected patients. Conclusion This study provides insight into potential factors influencing gametocyte production in symptomatic patients. The findings contribute to enhance understanding of risk factors associated with gametocyte carriage in humans. Trial registration NCT01232530.

Gametocyte Carriages of Plasmodium falciparum (pfs25) and Plasmodium vivax (pvs25) during Mass Screening and Treatment in West Timor, Indonesia a Longitudinal Prospective Study

Background: Three rounds of mass screening and treatment (MST) demonstrated no effect on Plasmodium falciparum and P. vivax incidence in West Timor, Indonesia. This study nested within that trial evaluated the effect of MST on gametocyte carriage.Methods: Microscopy and PCR diagnostics were applied to study subjects through 3 months of MST involving dihydroartemisinin-piperaquine (DHP) and primaquine based on infecting Plasmodium species. RT-qPCR targeting the pfs25 and pvs25 sequences was conducted to detect and quantify gametocytes in blood samples of P. falciparum and P. vivax- infected subjects. Data from the baseline and endpoint were compared (p<=0.05 as the significance threshold).Results:Between baseline and endpoint, prevalence remained unchanged for P. falciparum (6%=52/811 versus 7%=50/740, p=0.838) and decreased slightly for P. vivax (24%=192/811 versus 19%=142/740, p=0.035). No significant difference was observed in gametocyte carriage rates for either P. falciparum (43%=19/44 versus 59%=23/39, OR & 95%CI=1.89, 0.72-4.97) or P. vivax (39%=49/125 versus 37%=39/106, OR & 95%CI=0.90, 0.51-1.59). Despite the insignificant differences between the two time points, the majority of positive subjects at the endpoint were newly parasitemic individuals (new infections) (Pf:92%=46/50, Pv: 65%=93/142). This result was similarly demonstrated for the transmissible stage - where new gametocyte carriers dominated endpoint positive subjects (Pf: 96%=22/23, Pv: 95%=37/39). Conclusion:Although three rounds of MST decreased individual parasitemia and gametocytemia based on diagnosis and treatment, this failed to impact malaria prevalence at a population level due to the development of new infected individuals that can sustain transmission. These observations in part explains why MST failed to impact transmission of malaria in this transmission context.

Gametocyte Carriage After Seasonal Malaria Chemoprevention in Plasmodium Falciparum Infected Asymptomatic Children

BackgroundTreatment of clinical Plasmodium falciparum malaria with sulfadoxine-pyrimethamine (SP) and amodiaquine (AQ) is associated with increased post-treatment gametocyte carriage. The effect of seasonal malaria chemoprevention (SMC) with SP and AQ on gametocyte carriage was assessed in asymptomatic Plasmodium falciparum infected children. MethodsThe study was carried out in eastern Gambia. Asymptomatic malaria infected children aged 24-59 months old eligible for SMC (SMC group) and children 5-8 years that did not receive SMC treatment (comparison group) were recruited. Baseline gametocyte prevalence was determined by molecular methods before SMC administration and 13 days after. Gametocyte carriage between the groups was compared using the Chi-squared test and within person using conditional logistic regression.ResultsDuring the 2017 and 2018 malaria transmission seasons, 65 and 75 children were recruited in the SMC and comparison groups respectively. Before SMC administration, gametocyte prevalence was 10.7% (7/65) in the SMC group and 13.3% (10/75) in the comparison group (p=0.64). At day 13 after SMC administration, this was 9.4%, (5/53) for the SMC group and 12.7%, (9/71) for the comparison group (p=0.57). Within the SMC group, gametocyte carriage before and after SMC administration was similar (OR 0.6, 95% CI 0.14 – 2.51) (p=0.48).ConclusionIn this study with relative low gametocyte prevalence prior to SMC treatment, no evidence was observed that SMC increased gametocyte carriage in asymptomatic malaria infected children.

Prevalence of asymptomatic P. falciparum gametocyte carriage in schoolchildren and assessment of the association between gametocyte density, multiplicity of infection and mosquito infection prevalence

Background: Malaria is a major public health threat in sub-Saharan Africa. Asymptomatic Plasmodium falciparum gametocyte carriers are potential infectious reservoirs for sustaining transmission in many malaria endemic regions. The aim of the study was to assess the prevalence of gametocyte carriage and some of its associated risk factors among asymptomatic schoolchildren in Western Kenya and further analyse the association between gametocyte density, multiplicity of infection (MOI) and mosquito infection prevalence. Methods: Rapid diagnostic tests were used to screen for P. falciparum parasite infection among schoolchildren (5-15 years old) and the results were verified using microscopy. Microscopy positive gametocyte carriers were selected to feed laboratory reared An. gambiae s.l. mosquitoes using membrane feeding method. Genomic DNA was extracted from dry blood spot samples and P. falciparum populations were genotyped using 10 polymorphic microsatellite markers. Assessment of the association between MOI and gametocyte density and mosquito infection prevalence was conducted. Results: A significantly higher prevalence of P. falciparum infection was found in males 31.54% (764/2422) (p-value < 0.001) compared to females 26.72% (657/2459). The microscopy gametocyte prevalence among the study population was 2% (84/4881). Children aged 5-9 years have a higher prevalence of gametocyte carriage (odds ratios = 2.1 [95% CI = 1.3–3.4], P = 0.002) as compared to children aged 10-15 years. After challenging An. gambiae s.l. by membrane feeding assay on gametocyte positive patient blood, our results indicate that 68.1% of the variation in mosquito infection prevalence is accounted for by gametocyte density and MOI (R-SQR. = 0.681, p < 0.001). Conclusions: Age was a significant risk factor for gametocyte carriage, as indicated by the higher risk of gametocyte carriage among the younger children (5-9 years). Gametocyte density and MOI statistically significantly predicted mosquito infection prevalence. Both of the variables added significantly to the prediction (p < 0.05).

Assessment of the Relationship Between Gametocyte Density, Multiplicity of Infection and Mosquito Infectivity in Plasmodium Falciparum Infections

Background: Malaria is a major public health threat in sub-Saharan Africa. Asymptomatic P. falciparum gametocyte carriers are potential infectious reservoirs for sustaining transmission in many malaria endemic regions. The aim of the study was to assess the prevalence of gametocyte carriage and some of its associated risk factors among asymptomatic schoolchildren (age 5-15 years) in Mbita, western Kenya and further analyse the association between gametocyte density, multiplicity of infection (MOI) and mosquito infection prevalence.Methods: Rapid diagnostic test (RDT) was used to screen for P. falciparum parasite infection among asymptomatic schoolchildren (5-15 years old) residing in Mbita, Western Kenya and the results were further verified using microscopy. Microscopy positive gametocyte carriers were selected to feed laboratory reared An. gambiae s.s. mosquitoes using membrane feeding method. Genomic DNA was extracted from dry blood spots (DBS) samples and P. falciparum populations were genotyped using 10 polymorphic microsatellite markers. Assessment of the association between MOI and gametocyte density and mosquito infection rates was conducted. Results: The prevalence of Plasmodium falciparum infection among the study population was 29.11% (1421/4881). A significantly higher prevalence of P. falciparum infection was found among the male gender 31.54% (764/2422) (p-value < 0.001) compared to the females 26.72% (657/2459). The microscopy gametocyte prevalence among the study population was 2% (84/4881). Children (5-9 years) have a higher risk of gametocyte carriage (Odd Ratios = 2.1 [95% CI = 1.3–3.4], P = 0.002) as compared to those between the ages of 10-15 years. Our results indicate that, about 68.1% of the variation in mosquito infection prevalence is accounted for by the gametocytes density and MOI (R-SQR. = 0.681, p < 0.001).Conclusion: The study reports a P. falciparum infection prevalence of 29.11% with a gametocyte prevalence of 2% among the study population as determined by microscopy. Age was a significant risk factor for gametocyte carriage as indicated by the higher risk of gametocyte carriage among the younger children (5-9 years). The gametocyte density and multiplicity of infection statistically significantly predicted mosquito infection prevalence. Both of the variables added significantly to the prediction, (p < 0.05).