Browsing by Author "Felger, Ingrid"

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    Comparison of detection methods to estimate asexual Plasmodium falciparum parasite prevalence and gametocyte carriage in a community survey in Tanzania
    (Springer Nature, 2014-11-18) Mwingira, Felista; Genton, Blaise; Kabanywanyi, Abdu-Noor M; Felger, Ingrid
    Background The use of molecular techniques to detect malaria parasites has been advocated to improve the accuracy of parasite prevalence estimates, especially in moderate to low endemic settings. Molecular work is time-consuming and costly, thus the effective gains of this technique need to be carefully evaluated. Light microscopy (LM) and rapid diagnostic tests (RDT) are commonly used to detect malaria infection in resource constrained areas, but their limited sensitivity results in underestimation of the proportion of people infected with Plasmodium falciparum. This study aimed to evaluate the extent of missed infections via a community survey in Tanzania, using polymerase chain reaction (PCR) to detect P. falciparum parasites and gametocytes. Methods Three hundred and thirty individuals of all ages from the Kilombero and Ulanga districts (Tanzania) were enrolled in a cross-sectional survey. Finger prick blood samples were collected for parasite detection by RDT, LM and molecular diagnosis using quantitative 18S rRNA PCR and msp2 nPCR. Gametocytes were detected by LM and by amplifying transcripts of the gametocyte-specific marker pfs25. Results Results from all three diagnostic methods were available for a subset of 226 individuals. Prevalence of P. falciparum was 38% (86/226; 95% CI 31.9–44.4%) by qPCR, 15.9% (36/226; 95% CI 11.1–20.7%) by RDT and 5.8% (13/226; 95% CI 2.69- 8.81%) by LM. qPCR was positive for 72% (26/36) of the RDT-positive samples. Gametocyte prevalence was 10.6% (24/226) by pfs25-qRT-PCR and 1.2% by LM.
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    Plasmodium falciparum msp1 , msp2 and glurp allele frequency and diversity in sub-Saharan Africa
    (Malaria Journal, 2011) Mwingira, Felista; Nkwengulila, Gamba; Schoepflin, Sonja; Sumari, Deborah; Beck, Hans-Peter; Snounou, Georges; Felger, Ingrid; Olliaro, Piero; Mugittu, Kefas
    Background: The efficacy of anti-malarial drugs is assessed over a period of 28-63 days (depending on the drugs’ residence time) following initiation of treatment in order to capture late failures. However, prolonged follow-up increases the likelihood of new infections depending on transmission intensity. Therefore, molecular genotyping of highly polymorphic regions of Plasmodium falciparum msp1, msp2 and glurp loci is usually carried out to distinguish recrudescence (true failures) from new infections. This tool has now been adopted as an integral part of anti-malarial efficacy studies and clinical trials. However, there are concerns over its utility and reliability because conclusions drawn from molecular typing depend on the genetic profile of the respective parasite populations, but this profile is not systematically documented in most endemic areas. This study presents the genetic diversity of P. falciparum msp1, msp2 and glurp markers in selected sub-Saharan Africa countries with varying levels of endemicity namely Malawi, Tanzania, Uganda, Burkina Faso and São Tomé. Methods: A total 780 baseline (Day 0) blood samples from children less than seven years, recruited in a randomized controlled clinical trials done between 1996 and 2000 were genotyped. DNA was extracted; allelic frequency and diversity were investigated by PCR followed by capillary electrophoresis for msp2 and fragment sizing by a digitalized gel imager for msp1 and glurp. Results and Conclusion: Plasmodium falciparum msp1, msp2 and glurp markers were highly polymorphic with low allele frequencies. A total of 17 msp1 genotypes [eight MAD20-, one RO33- and eight K1-types]; 116 msp2 genotypes [83 3D7 and 33 FC27- types] and 14 glurp genotypes were recorded. All five sites recorded very high expected heterozygosity (HE) values (0.68 - 0.99). HE was highest in msp2 locus (HE = 0.99), and lowest for msp1 (HE = 0.68) (P < 0.0001). The genetic diversity and allelic frequency recorded were independent of transmission intensity (P = 0.84, P = 0.25 respectively. A few genotypes had particularly high frequencies; however the most abundant showed only a 4% probability that a new infection would share the same genotype as the baseline infection. This is unlikely to confound the distinction of recrudescence from new infection, particularly if more than one marker is used for genotyping. Hence, this study supports the use of msp1, msp2 and glurp in malaria clinical trials in sub-Saharan Africa to discriminate new from recrudescent infections.
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    Plasmodium Falciparum Msp1, Msp2 And Glurp Allele Frequency and Diversity in Sub-Saharan Africa
    (BioMed Central, 2011) Mwingira, Felista; Nkwengulila, Gamba; Schoepflin, Sonja; Sumari, Deborah; Beck, Hans-Peter; Snounou, Georges; Felger, Ingrid; Olliaro, Piero; Mugittu, Kefas
    The efficacy of anti-malarial drugs is assessed over a period of 28-63 days (depending on the drugs’ residence time) following initiation of treatment in order to capture late failures. However, prolonged follow-up increases the likelihood of new infections depending on transmission intensity. Therefore, molecular genotyping of highly polymorphic regions of Plasmodium falciparum msp1, msp2 and glurp loci is usually carried out to distinguish recrudescence (true failures) from new infections. This tool has now been adopted as an integral part of anti-malarial efficacy studies and clinical trials. However, there are concerns over its utility and reliability because conclusions drawn from molecular typing depend on the genetic profile of the respective parasite populations, but this profile is not systematically documented in most endemic areas. This study presents the genetic diversity of P. falciparum msp1, msp2 and glurp markers in selected sub-Saharan Africa countries with varying levels of endemicity namely Malawi, Tanzania, Uganda, Burkina Faso and São Tomé. A total 780 baseline (Day 0) blood samples from children less than seven years, recruited in a randomized controlled clinical trials done between 1996 and 2000 were genotyped. DNA was extracted; allelic frequency and diversity were investigated by PCR followed by capillary electrophoresis for msp2 and fragment sizing by a digitalized gel imager for msp1 and glurp. Plasmodium falciparum msp1, msp2 and glurp markers were highly polymorphic with low allele frequencies. A total of 17 msp1 genotypes [eight MAD20-, one RO33- and eight K1-types]; 116 msp2 genotypes [83 3D7 and 33 FC27- types] and 14 glurp genotypes were recorded. All five sites recorded very high expected heterozygosity (HE) values (0.68 - 0.99). HE was highest in msp2 locus (HE = 0.99), and lowest for msp1 (HE = 0.68) (P < 0.0001). The genetic diversity and allelic frequency recorded were independent of transmission intensity (P = 0.84, P = 0.25 respectively. A few genotypes had particularly high frequencies; however the most abundant showed only a 4% probability that a new infection would share the same genotype as the baseline infection. This is unlikely to confound the distinction of recrudescence from new infection, particularly if more than one marker is used for genotyping. Hence, this study supports the use of msp1, msp2 and glurp in malaria clinical trials in sub-Saharan Africa to discriminate new from recrudescent infections.
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    The temporal dynamics and infectiousness of subpatent Plasmodium falciparum infections in relation to parasite density
    (Nature Communications, 2019-03-29) Slater, Hannah; Ross, Amanda; Felger, Ingrid; Hofmann, Natalie; Robison, Leanne; cook, Jackie; Gonçalves, Bronner; Bjòrkman, Andes; Ouedraogo, Andre; Morris, Ulrika; Msellem, Mwinyi; Koepfli, Christian; Muller, Ivo; Tadesse, Fitsum; Gadisa, Endalamaw; Das, Simita; Domingo, Gonzalo; Kapulu, Melissa; Midega, Janet; Owusu-Agyei, Seth; Nabet, Cecile; Piarroux, Renaud; Duombo, Ogobara; Niare, Safiatou; koram, Kwadwo; Lucci, Nami; Udhayakumar, Venkatachalam; Mosha, Jackline; Tiono, Alfred; Chandramohan, Daniel; Gosling, Roly; Mwingira, Felista; Sauerwein, Robert; Paul, Richard; Riley, Eleanor; White, Nicholas; Nosten, Francois; Imwong, Mallika; Bousema, Teun; Drakeley, Chris; Okell, Lucy
    Malaria infections occurring below the limit of detection of standard diagnostics are common in all endemic settings. However, key questions remain surrounding their contribution to sustaining transmission and whether they need to be detected and targeted to achieve malaria elimination. In this study we analyse a range of malaria datasets to quantify the density, detectability, course of infection and infectiousness of subpatent infections. Asymptomatically infected individuals have lower parasite densities on average in low transmission settings compared to individuals in higher transmission settings. In cohort studies, subpatent infections are found to be predictive of future periods of patent infection and in membrane feeding studies, individuals infected with subpatent asexual parasite densities are found to be approximately a third as infectious to mosquitoes as individuals with patent (asexual parasite) infection. These results indicate that subpatent infections contribute to the infectious reservoir, may be long lasting, and require more sensitive diagnostics to detect them in lower transmission settings.
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    Ultra-Sensitive Detection of Plasmodium falciparum by Amplification of Multi-Copy Subtelomeric Targets
    (PLOS, 2015-03-03) Hofmann, Natalie; Mwingira, Felista; Shekalaghe, Seif; Robison, Leanne; Muller, Ivo; Felger, Ingrid
    Background Planning and evaluating malaria control strategies relies on accurate definition of parasite prevalence in the population. A large proportion of asymptomatic parasite infections can only be identified by surveillance with molecular methods, yet these infections also contribute to onward transmission to mosquitoes. The sensitivity of molecular detection by PCR is limited by the abundance of the target sequence in a DNA sample; thus, detection becomes imperfect at low densities. We aimed to increase PCR diagnostic sensitivity by targeting multi-copy genomic sequences for reliable detection of low-density infections, and investigated the impact of these PCR assays on community prevalence data. Methods and Findings Two quantitative PCR (qPCR) assays were developed for ultra-sensitive detection of Plasmodium falciparum, targeting the high-copy telomere-associated repetitive element 2 (TARE-2, *250 copies/genome) and the var gene acidic terminal sequence (varATS, 59 copies/genome). Our assays reached a limit of detection of 0.03 to 0.15 parasites/μl blood and were 10× more sensitive than standard 18S rRNA qPCR. In a population cross-sectional study in Tanzania, 295/498 samples tested positive using ultra-sensitive assays. Light microscopy missed 169 infections (57%). 18S rRNA qPCR failed to identify 48 infections (16%), of which 40% carried gametocytes detected by pfs25 quantitative reverse-transcription PCR. To judge the suitability of the TARE-2 and varATS assays for high-throughput screens, their performance was tested on sample pools. Both ultra-sensitive assays correctly detected all pools containing one low-density P. falciparum–positive sample, which went undetected by 18S rRNA qPCR, among nine negatives. TARE-2 and varATS qPCRs improve estimates of prevalence rates, yet other infections might still remain undetected when absent in the limited blood volume sampled. Conclusions Measured malaria prevalence in communities is largely determined by the sensitivity of the diagnostic tool used. Even when applying standard molecular diagnostics, prevalence in our study population was underestimated by 8% compared to the new assays. Our findings highlight the need for highly sensitive tools such as TARE-2 and varATS qPCR in community surveillance and for monitoring interventions to better describe malaria epidemiology and inform malaria elimination efforts.