RESUMEN
BACKGROUND: Recent studies from different malaria-endemic regions including western Africa have now shown that Plasmodium vivax can infect red blood cells (RBCs) and cause clinical disease in Duffy-negative people, though the Duffy-negative phenotype was thought to confer complete refractoriness against blood invasion with P. vivax. The actual prevalence of P. vivax in local populations in Ghana is unknown and little information is available about the distribution of Duffy genotypes. The aim of this study was to assess the prevalence of P. vivax in both asymptomatic and symptomatic outpatients and the distribution of Duffy genotypes in Ghana. METHODS: DNA was extracted from dried blood spots (DBS) collected from 952 subjects (845 malaria patients and 107 asymptomatic persons) from nine locations in Ghana. Plasmodium species identification was carried out by nested polymerase chain reaction (PCR) amplification of the small-subunit (SSU) rRNA genes. For P. vivax detection, a second PCR of the central region of the Pvcsp gene was carried out. Duffy blood group genotyping was performed by allele-specific PCR to detect the presence of the FYES allele. RESULTS: No cases of P. vivax were detected in any of the samples by both PCR methods used. Majority of infections (542, 94.8%) in the malaria patient samples were due to P. falciparum with only 1 infection (0.0017%) due to Plasmodium malariae, and 2 infections (0.0034%) due to Plasmodium ovale. No case of mixed infection was identified. Of the samples tested for the FYES allele from all the sites, 90.5% (862/952) had the FYES allele. All positive samples were genotyped as FY*B-33/FY*B-33 (Duffy-negative homozygous) and therefore classified as Fy(a-b-). CONCLUSIONS: No cases of P. vivax were detected by both PCRs and majority of the subjects tested carried the FYES allele. The lack of P. vivax infections observed can be attributed to the high frequency of the FYES allele that silences erythroid expression of the Duffy. These results provide insights on the host susceptibility for P. vivax infections that had not been investigated in Ghana before.
Asunto(s)
Sistema del Grupo Sanguíneo Duffy/genética , Frecuencia de los Genes , Genotipo , Malaria Vivax/epidemiología , Ghana/epidemiología , Malaria Vivax/parasitología , Plasmodium vivax/fisiología , PrevalenciaRESUMEN
Infections with the malaria parasite Plasmodium falciparum typically comprise multiple strains, especially in high-transmission areas where infectious mosquito bites occur frequently. However, little is known about the dynamics of mixed-strain infections, particularly whether strains sharing a host compete or grow independently. Competition between drug-sensitive and drug-resistant strains, if it occurs, could be a crucial determinant of the spread of resistance. We analysed 1341 P. falciparum infections in children from Angola, Ghana and Tanzania and found compelling evidence for competition in mixed-strain infections: overall parasite density did not increase with additional strains, and densities of individual chloroquine-sensitive (CQS) and chloroquine-resistant (CQR) strains were reduced in the presence of competitors. We also found that CQR strains exhibited low densities compared with CQS strains (in the absence of chloroquine), which may underlie observed declines of chloroquine resistance in many countries following retirement of chloroquine as a first-line therapy. Our observations support a key role for within-host competition in the evolution of drug-resistant malaria. Malaria control and resistance-management efforts in high-transmission regions may be significantly aided or hindered by the effects of competition in mixed-strain infections. Consideration of within-host dynamics may spur development of novel strategies to minimize resistance while maximizing the benefits of control measures.
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Antimaláricos/farmacología , Cloroquina/farmacología , Resistencia a Medicamentos , Malaria Falciparum/parasitología , Plasmodium falciparum/efectos de los fármacos , Plasmodium falciparum/fisiología , Angola , Niño , Preescolar , Ghana , Humanos , Lactante , Recién Nacido , Malaria Falciparum/tratamiento farmacológico , Plasmodium falciparum/genética , TanzaníaRESUMEN
BACKGROUND: Case management based on prompt diagnosis and adequate treatment using artemisinin-based combination therapy (ACT) remains the main focus of malaria control in Ghana. As part of routine surveillance on the therapeutic efficacy of ACT in Ghana, the efficacy of amodiaquine-artesunate (AS-AQ) and artemether-lumefantrine (AL) were studied in six sentinel sites representing the forest and savannah zones of the country. METHODS: Three sites representing the two ecological zones studied AS-AQ whilst the other three sites studied AL. In each site, the study was a one-arm prospective evaluation of the clinical, parasitological, and haematological responses to directly observed therapy for uncomplicated malaria with either AS-AQ or AL among children aged 6 months and 9 years. The WHO 2009 protocol for monitoring anti-malarial drug efficacy was used for the study between July 2013 and March 2014. RESULTS: Per-protocol analyses on day 28 showed an overall PCR-corrected cure rate of 100% for AS-AQ and 97.6% (95% CI 93.1, 99.5) for AL: 97.2% (95% CI 92.0, 99.4) in the forest zone and 100% in the savannah zone. Kaplan-Meier survival analysis showed similar outcomes. Prevalence of fever decreased by about 75% after the first day of treatment with each ACT in the two ecological zones. No child studied was parasitaemic on day 3, and gametocytaemia was generally maintained at low levels (<5%). Post-treatment mean haemoglobin concentrations significantly increased in the two ecological zones. CONCLUSIONS: Therapeutic efficacy of AS-AQ and AL remains over 90% in the forest and savannah zones of Ghana. Additionally, post-treatment parasitaemia on day 3 is rare suggesting that artemisinin is still efficacious in Ghana.
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Amodiaquina/uso terapéutico , Antimaláricos/uso terapéutico , Artemisininas/uso terapéutico , Malaria/tratamiento farmacológico , Artesunato , Niño , Preescolar , Combinación de Medicamentos , Ecología , Femenino , Ghana , Humanos , Lactante , Masculino , Plasmodium falciparum/efectos de los fármacos , Plasmodium falciparum/patogenicidad , Estudios ProspectivosRESUMEN
BACKGROUND: The recently introduced SYBR Green1 (SG) assay for testing parasites susceptibility to anti-malarial drugs needs further improvement. This has been necessitated by various setbacks, the major one being the low fluorescence intensity associated with it use. This shortcoming diminishes the anticipated hope that this novel method was going to replace the more traditional ones, such as the isotopic and microscopy. In order to restore confidence in its use, series of experiments to determine conditions that give the best fluorescence intensity were conducted. METHODS: Conditions that yield the maximum fluorescent signal were ascertained by measuring the fluorescence after incubation of Plasmodium falciparum culture at different parasites concentration with lysis buffer containing SYBR Green (LBS) at different time period. In order to ascertain the effect of freeze-thaw on fluorescence intensity, P. falciparum culture was frozen for 1 h, thawed, incubated with LBS and the fluorescence measured. The optimized conditions determined in this study were then used to assess the susceptibility of clinical isolates of P. falciparum to artesunate, chloroquine and mefloquine. The concentration of anti-malarial drug inhibiting parasite growth by 50 % (IC50) for each drug was estimated using the online ICEstimator. The IC50 generated using the optimized SG method determined in this study was compared with that obtained using microscopic method and the previously reported standard SG method. RESULTS: Over all, the SG method was found to be easy to perform and sensitive. Freeze-thaw of parasite culture followed by incubation with lysis buffer containing the dye for 3 h was consistently observed to give the highest fluorescence signal. The IC50 values for chloroquine, mefloquine and artesunate determined were consistent and comparable with that determined with the previously reported standard SG method and the microscopic method. CONCLUSION: The authors conclude that freezing and thawing of parasite culture, followed by incubation with LBS in the dark for 3 h provided a significant improvement in fluorescence signal. The IC50 generated using the improved SG method is comparable with that from microscopy and the standard method.
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Antimaláricos/farmacología , Resistencia a Medicamentos , Fluorometría/métodos , Compuestos Orgánicos/metabolismo , Pruebas de Sensibilidad Parasitaria/métodos , Plasmodium falciparum/efectos de los fármacos , Coloración y Etiquetado/métodos , Artemisininas/farmacología , Artesunato , Benzotiazoles , Niño , Preescolar , Cloroquina/farmacología , Diaminas , Humanos , Concentración 50 Inhibidora , Mefloquina/farmacología , Plasmodium falciparum/genética , QuinolinasRESUMEN
BACKGROUND: Congenital malaria is defined as malaria parasitaemia in the first week of life. The reported prevalence of congenital malaria in sub-Saharan Africa is variable (0 - 46%). Even though the clinical significance of congenital malaria parasitaemia is uncertain, anti-malarial drugs are empirically prescribed for sick newborns by frontline health care workers. Data on prevalence of congenital malaria in high-risk newborns will inform appropriate drug use and timely referral of sick newborns. METHODS: Blood samples of untreated newborns less than 1 week of age at the time of referral to Korle Bu Teaching hospital in Accra, Ghana during the peak malaria seasons (April to July) of 2008 and 2010 were examined for malaria parasites by, i) Giemsa-stained thick and thin blood smears for parasite count and species identification, ii) histidine-rich protein- and lactic dehydrogenase-based rapid diagnosis tests, or iii) polymerase chain reaction amplification of the merozoite surface protein 2 gene, for identification of sub-microscopic parasitaemia. Other investigations were also done as clinically indicated. RESULTS: In 2008, nine cases of Plasmodium falciparum parasitaemia were diagnosed by microscopy in 405 (2.2%) newborns. All the nine newborns had low parasite densities (≤ 50 per microlitre). In 2010, there was no case of parasitaemia by either microscopy or rapid diagnosis tests in 522 newborns; however, 56/467 (12%) cases of P. falciparum were detected by polymerase chain reaction. CONCLUSION: Congenital malaria is an uncommon cause of clinical illness in high-risk untreated newborns referred to a tertiary hospital in the first week of life. Empirical anti-malarial drug treatment for sick newborns without laboratory confirmation of parasitaemia is imprudent. Early referral of sick newborns to hospitals with resources and skills for appropriate care is recommended.
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Malaria Falciparum/congénito , Malaria Falciparum/epidemiología , Antígenos de Protozoos/sangre , Sangre/parasitología , Estudios Transversales , ADN Protozoario/sangre , Femenino , Ghana/epidemiología , Humanos , Inmunoensayo , Recién Nacido , Masculino , Microscopía , Parasitemia/congénito , Parasitemia/epidemiología , Reacción en Cadena de la Polimerasa , PrevalenciaRESUMEN
BACKGROUND: Based on report of declining efficacy of chloroquine, Ghana shifted to the use of artemisinin-based combination therapy (ACT) in 2005 as the first-line anti-malarial drug. Since then, there has not been any major evaluation of the efficacy of anti-malarial drugs in Ghana in vitro. The sensitivity of Ghanaian Plasmodium falciparum isolates to anti-malarial drugs was, therefore, assessed and the data compared with that obtained prior to the change in the malaria treatment policy. METHODS: A SYBR Green 1 fluorescent-based in vitro drug sensitivity assay was used to assess the susceptibility of clinical isolates of P. falciparum to a panel of 12 anti-malarial drugs in three distinct eco-epidemiological zones in Ghana. The isolates were obtained from children visiting health facilities in sentinel sites located in Hohoe, Navrongo and Cape Coast municipalities. The concentration of anti-malarial drug inhibiting parasite growth by 50% (IC50) for each drug was estimated using the online program, ICEstimator. RESULTS: Pooled results from all the sentinel sites indicated geometric mean IC50 values of 1.60, 3.80, 4.00, 4.56, 5.20, 6.11, 10.12, 28.32, 31.56, 93.60, 107.20, and 8952.50 nM for atovaquone, artesunate, dihydroartemisin, artemether, lumefantrine, amodiaquine, mefloquine, piperaquine, chloroquine, tafenoquine, quinine, and doxycycline, respectively. With reference to the literature threshold value indicative of resistance, the parasites showed resistance to all the test drugs except the artemisinin derivatives, atovaquone and to a lesser extent, lumefantrine. There was nearly a two-fold decrease in the IC50 value determined for chloroquine in this study compared to that determined in 2004 (57.56 nM). This observation is important, since it suggests a significant improvement in the efficacy of chloroquine, probably as a direct consequence of reduced drug pressure after cessation of its use. Compared to that measured prior to the change in treatment policy, significant elevation of artesunate IC50 value was observed. The results also suggest the existence of possible cross-resistance among some of the test drugs. CONCLUSION: Ghanaian P. falciparum isolates, to some extent, have become susceptible to chloroquine in vitro, however the increasing trend in artesunate IC50 value observed should be of concern. Continuous monitoring of ACT in Ghana is recommended.
Asunto(s)
Antimaláricos/farmacología , Malaria Falciparum/parasitología , Compuestos Orgánicos/química , Plasmodium falciparum/efectos de los fármacos , Benzotiazoles , Niño , Preescolar , Cloroquina/farmacología , Diaminas , Resistencia a Medicamentos , Ghana , Humanos , Lactante , Concentración 50 Inhibidora , QuinolinasRESUMEN
BACKGROUND: With the introduction of artemisinin-based combination therapy (ACT) in 2005, monitoring of anti-malarial drug efficacy, which includes the use of molecular tools to detect known genetic markers of parasite resistance, is important for first-hand information on the changes in parasite susceptibility to drugs in Ghana. This study investigated the Plasmodium falciparum multidrug resistance gene (pfmdr1) copy number, mutations and the chloroquine resistance transporter gene (pfcrt) mutations in Ghanaian isolates collected in seven years to detect the trends in prevalence of mutations. METHODS: Archived filter paper blood blots collected from children aged below five years with uncomplicated malaria in 2003-2010 at sentinel sites were used. Using quantitative real-time polymerase chain reaction (qRT-PCR), 756 samples were assessed for pfmdr1 gene copy number. PCR and restriction fragment length polymorphism (RFLP) were used to detect alleles of pfmdr1 86 in 1,102 samples, pfmdr1 184, 1034, 1042 and 1246 in 832 samples and pfcrt 76 in 1,063 samples. Merozoite surface protein 2 (msp2) genotyping was done to select monoclonal infections for copy number analysis. RESULTS: The percentage of isolates with increased pfmdr1 copy number were 4, 27, 9, and 18% for 2003-04, 2005-06, 2007-08 and 2010, respectively. Significant increasing trends for prevalence of pfmdr1 N86 (×(2) = 96.31, p <0.001) and pfcrt K76 (×(2) = 64.50, p <0.001) and decreasing trends in pfmdr1 Y86 (x(2) = 38.52, p <0.001) and pfcrt T76 (x(2) = 43.49, p <0.001) were observed from 2003-2010. The pfmdr1 F184 and Y184 prevalence showed an increasing and decreasing trends respectively but were not significant (×(2) = 7.39,p=0.060; ×(2) = 7.49, p = 0.057 respectively). The pfmdr1 N86-F184-D1246 haplotype, which is alleged to be selected by artemether-lumefantrine showed a significant increasing trend (×(2) = 20.75, p < 0.001). CONCLUSION: Increased pfmdr1 gene copy number was observed in the isolates analysed and this finding has implications for the use of ACT in the country although no resistance has been reported. The decreasing trend in the prevalence of chloroquine resistance markers after change of treatment policy presents the possibility for future introduction of chloroquine as prophylaxis for malaria risk groups such as children and pregnant women in Ghana.
Asunto(s)
Antimaláricos/farmacología , Resistencia a Medicamentos , Malaria Falciparum/parasitología , Proteínas de Transporte de Membrana/genética , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Plasmodium falciparum/genética , Proteínas Protozoarias/genética , Antimaláricos/uso terapéutico , Preescolar , ADN Protozoario/genética , Femenino , Dosificación de Gen , Frecuencia de los Genes , Ghana/epidemiología , Política de Salud , Humanos , Lactante , Malaria Falciparum/epidemiología , Masculino , Mutación Missense , Plasmodium falciparum/clasificación , Plasmodium falciparum/efectos de los fármacos , Plasmodium falciparum/aislamiento & purificación , Polimorfismo de Longitud del Fragmento de Restricción , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
BACKGROUND: In 2008, artemether - lumefantrine (AL) and dihydroartemisinin - piperaquine (DHAP) were added to artesunate - amodiaquine (AS-AQ) as first-line drugs for uncomplicated malaria in Ghana. The introduction of new drugs calls for continuous monitoring of these drugs to provide timely information on trends of their efficacy and safety to enhance timely evidence-based decision making by the National Malaria Control Programme. In this regard, the therapeutic efficacy of AL was monitored from September 2010 to April 2011 in four sentinel sites representing the three main ecological zones of the country. METHODS: The study was a one-arm prospective evaluation of clinical and parasitological responses to directly observed treatment for uncomplicated malaria among children aged 6 months to 59 months using the 2009 WHO protocol for surveillance of anti-malarial drug efficacy. Children recruited into the study received weight-based 20/120 mg AL at 0, 8, 24, 36, 48, and 60 hrs. Parasitaemia levels were assessed on days 2, 3, 7, 14, 21, 28, and at any time a study child was brought to the clinic with fever. RESULTS: A total of 175 children were enrolled into the study: 56 in the savanna zone, 78 in the forest zone and 41 in the coastal zone. Per-protocol analysis showed that the overall PCR-corrected cure rates on day 14 and day 28 were 96.5% (95% CI: 92.1, 98.6) and 95.4% (95% CI: 90.3, 98.0), respectively, with statistically significant differences between the ecological zones. The 90.4% day-28 cure rate observed in the savannah zone (95% CI: 78.2, 96.4) was significantly the lowest compared with 100% (95% CI: 93.2, 99.9) in the forest zone and 93.8% (95% CI: 77.8, 98.9) in the coastal zone (P = 0.017). Fever and parasite clearance were slower among children enrolled in the savannah zone. Gametocytaemia after day-3 post-treatment was rare in all the zones. CONCLUSIONS: The study has shown that AL remains efficacious in Ghana with significant ecologic zonal differences. The savannah zone may be a potential zone for any emergence of resistant alleles as a result of the slower parasite clearance observed in the zone.
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Antimaláricos/administración & dosificación , Artemisininas/administración & dosificación , Etanolaminas/administración & dosificación , Fluorenos/administración & dosificación , Malaria/tratamiento farmacológico , Combinación Arteméter y Lumefantrina , Preescolar , Combinación de Medicamentos , Femenino , Ghana , Humanos , Lactante , Masculino , Estudios Prospectivos , Resultado del TratamientoRESUMEN
BACKGROUND: In 2005, Ghana adopted artemisinin-based combination therapy (ACT) for primary treatment of falciparum malaria. A comprehensive study of the drug-resistance-associated mutations and their genetic lineages will lead to a better understanding of the evolution of antimalarial drug resistance in this region. METHODS: The pfcrt, pfmdr1, dhps, and dhfr mutations associated with chloroquine (CQ) and sulfadoxine-pyrimethamine (SP) resistance and the microsatellite loci flanking these genes were genotyped in Plasmodium falciparum isolates from Ghana. RESULTS: The prevalence of mutations associated with both CQ and SP resistance was high in Ghana. However, we observed a decrease in prevalence of the pfcrt K76T mutation in northern Ghana after the change in drug policy from CQ to ACT. Analysis of genetic diversity and differentiation at microsatellite loci flanking all 4 genes indicated that they have been under strong selection, because of CQ and SP use. The triple-mutant pfcrt and dhfr alleles in Ghana were derived from Southeast Asia, whereas the double-mutant dhfr, dhps, and pfmdr1 alleles were of African lineage. CONCLUSION: Because of the possible role of pfmdr1 in amodiaquine and mefloquine resistance, demonstrating selection on pfmdr1 and defining lineages of resistant alleles in an African population holds great importance.
Asunto(s)
Alelos , Antimaláricos/farmacología , Resistencia a Medicamentos , Malaria Falciparum/parasitología , Plasmodium falciparum/efectos de los fármacos , Plasmodium falciparum/genética , Sustitución de Aminoácidos , Evolución Biológica , Preescolar , Cloroquina/farmacología , ADN Protozoario/genética , Dihidropteroato Sintasa/genética , Combinación de Medicamentos , Evolución Molecular , Genotipo , Ghana , Humanos , Lactante , Recién Nacido , Proteínas de Transporte de Membrana/genética , Repeticiones de Microsatélite , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Mutación Missense , Plasmodium falciparum/clasificación , Plasmodium falciparum/aislamiento & purificación , Proteínas Protozoarias/genética , Pirimetamina/farmacología , Sulfadoxina/farmacología , Tetrahidrofolato Deshidrogenasa/genéticaRESUMEN
The recent World Malaria report shows that progress in malaria elimination has stalled. Current data acquisition by NMCPs depend on passive case detection and clinical reports focused mainly on Plasmodium falciparum (Pf). In recent times, several countries in sub-Saharan Africa have reported cases of Plasmodium vivax (Pv) with a considerable number being Duffy negative. The burden of Pv and Plasmodium ovale (Po) appear to be more than acknowledged. Similarly, the contribution of asymptomatic malaria in transmission is hardly considered by NMCPs in Africa. Inclusion of these as targets in malaria elimination agenda is necessary to achieve elimination goal, as these harbor hypnozoites. The Pan African Vivax and Ovale Network (PAVON) is a new consortium of African Scientists working in Africa on the transmission profile of Pv and Po. The group collaborates with African NMCPs to train in Plasmodium molecular diagnostics, microscopy, and interpretation of molecular data from active surveys to translate into policy. Details of the mission, rational and modus operandi of the group are outlined.
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Malaria , Plasmodium ovale , Plasmodium vivax , África , Infecciones Asintomáticas/epidemiología , Malaria/epidemiología , Malaria/parasitología , Malaria/prevención & control , Malaria/transmisión , Malaria Vivax/epidemiología , Malaria Vivax/parasitología , Malaria Vivax/prevención & control , Malaria Vivax/transmisiónRESUMEN
BACKGROUND: Malaria is a major cause of morbidity and mortality in Africa. International effort and funding for control has been stepped up, with substantial increases from 2003 in the delivery of malaria interventions to pregnant women and children younger than 5 years in The Gambia. We investigated the changes in malaria indices in this country, and the causes and public-health significance of these changes. METHODS: We undertook a retrospective analysis of original records to establish numbers and proportions of malaria inpatients, deaths, and blood-slide examinations at one hospital over 9 years (January, 1999-December, 2007), and at four health facilities in three different administrative regions over 7 years (January, 2001-December, 2007). We obtained additional data from single sites for haemoglobin concentrations in paediatric admissions and for age distribution of malaria admissions. FINDINGS: From 2003 to 2007, at four sites with complete slide examination records, the proportions of malaria-positive slides decreased by 82% (3397/10861 in 2003 to 337/6142 in 2007), 85% (137/1259 to 6/368), 73% (3664/16932 to 666/11333), and 50% (1206/3304 to 336/1853). At three sites with complete admission records, the proportions of malaria admissions fell by 74% (435/2530 to 69/1531), 69% (797/2824 to 89/1032), and 27% (2204/4056 to 496/1251). Proportions of deaths attributed to malaria in two hospitals decreased by 100% (seven of 115 in 2003 to none of 117 in 2007) and 90% (22/122 in 2003 to one of 58 in 2007). Since 2004, mean haemoglobin concentrations for all-cause admissions increased by 12 g/L (85 g/L in 2000-04 to 97 g/L in 2005-07), and mean age of paediatric malaria admissions increased from 3.9 years (95% CI 3.7-4.0) to 5.6 years (5.0-6.2). INTERPRETATION: A large proportion of the malaria burden has been alleviated in The Gambia. Our results encourage consideration of a policy to eliminate malaria as a public-health problem, while emphasising the importance of accurate and continuous surveillance.
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Hospitalización/estadística & datos numéricos , Malaria Falciparum/epidemiología , Adolescente , Distribución por Edad , Animales , Anticuerpos Antiprotozoarios/sangre , Niño , Preescolar , Femenino , Gambia/epidemiología , Registros de Hospitales/estadística & datos numéricos , Hospitalización/tendencias , Humanos , Lactante , Recién Nacido , Malaria Falciparum/mortalidad , Malaria Falciparum/prevención & control , Estudios Multicéntricos como Asunto , Plasmodium falciparum/inmunología , Embarazo , Estudios Retrospectivos , Estaciones del AñoRESUMEN
Naturally acquired antibody responses provide partial protection from clinical malaria, and blood-stage parasite vaccines under development aim to prime such responses. To investigate the determinants of antibody response longevity, immunoglobulin G (IgG) antibodies to several blood-stage vaccine candidate antigens in the sera of two cohorts of children of up to 6 years of age during the dry seasons of 2003 and 2004 in The Gambia were examined. The first cohort showed that most antibodies were lost within less than 4 months of the first sampling if a persistent infection was not present, so the study of the second-year cohort involved collecting samples from individuals every 2 weeks over a 3-month period. Antibody responses in the second cohort were also influenced by persistent malaria infection, so analysis focused particularly on children in whom parasites were not detected after the first time point. Antibodies to most antigens declined more slowly in children in the oldest age group (>5 years old) and more rapidly in children in the youngest group (<3 years old). However, antibodies to merozoite surface protein 2 were shorter lived than antibodies to other antigens and were not more persistent in older children. The age-specific and antigen-specific differences were not explained by different IgG subclass response profiles, indicating the probable importance of differential longevities of plasma cell populations rather than antibody molecules. It is likely that young children mostly have short-lived plasma cells and thus experience rapid declines in antibody levels but that older children have longer-lasting antibody responses that depend on long-lived plasma cells.
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Envejecimiento/inmunología , Anticuerpos Antiprotozoarios/inmunología , Antígenos de Protozoos/inmunología , Malaria Falciparum/inmunología , Plasmodium falciparum/inmunología , Animales , Niño , Preescolar , Estudios de Cohortes , Gambia/epidemiología , Humanos , Lactante , Malaria Falciparum/epidemiología , Estaciones del Año , Factores de TiempoRESUMEN
Sulfadoxine-pyrimethamine (SP) is used as malaria chemoprophylaxis for pregnant women and children in Ghana. Plasmodium falciparum resistance to SP is linked to mutations in the dihydropteroate synthase gene (pfdhps), dihydrofolate reductase gene (pfdhfr) and amplification of GTP cyclohydrolase 1 (pfgch1) gene. The pfgch1 duplication is associated with pfdhfr L164, a crucial mutant for high level pyrimethamine resistance which is rare in Ghana. The presence of amplified pfgch1 in Ghanaian isolates could be an indicator of the evolution of the L164 mutant. This study therefore determined the pfgch1 copy number variations and SP resistance mutations in clinical isolates from Ghana. One hundred and ninety-two (192) blood samples collected from children aged ≤14 years with uncomplicated malaria in 2013-14 and 2015-16 were used. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the pfgch1 copy number and nested PCR-Sanger sequencing used to detect mutations in pfdhps and pfdhfr genes. Twelve parasites (6.3%) harbored double copies of the pfgch1 gene out of the 192 samples. Of the 12, 75% had the pfdhfr I51-R59-N108, 92% had the pfdhps G437 mutant, 8% had the pfdhps E540 and 67% had the SP resistance haplotype IRNG. No L164 was detected in samples with amplified pfgch1. The rare T108 mutant associated with cycloguanil resistance showed predominance (60%) over N108 in the 2015-16 isolates. The observation of parasites with increased copy number of pfgch1 gene is indicative of the future evolution of the rare quadruple pfdhfr mutant, I51-R59-N108-L164, in Ghanaian parasites. Mutant pfdhps isolates also had increased gch1 copy number suggestive that it may also facilitate sulphadoxine resistance. The selection of parasites with pfgch1 gene amplification will enhance the sustenance and persistence of parasites with SP resistance in the country. Policy makers need to begin the search for a replacement chemoprophylaxis drug for malaria vulnerable groups in Ghana.
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Dihidropteroato Sintasa/genética , Resistencia a Medicamentos/genética , GTP Ciclohidrolasa/genética , Mutación , Plasmodium falciparum/genética , Proteínas Protozoarias/genética , Tetrahidrofolato Deshidrogenasa/genética , Combinación de Medicamentos , Femenino , Ghana , Humanos , Masculino , Plasmodium falciparum/enzimología , Reacción en Cadena de la Polimerasa , Pirimetamina , SulfadoxinaRESUMEN
BACKGROUND: Plasmodium falciparum delayed clearance with the use of artemisinin-based combination therapy (ACTs) has been reported in some African countries. Single nucleotide polymorphisms (SNPs) in two genes, P. falciparum adaptor protein complex 2 mu subunit (pfap2mu) and ubiquitin specific protease 1 (pfubp1), have been linked to delayed clearance with ACT use in Kenya and recurrent imported malaria in Britain. With over 12 years of ACT use in Ghana, this study investigated the prevalence of SNPs in the pfap2mu and pfubp1 in Ghanaian clinical P. falciparum isolates to provide baseline data for antimalarial drug resistance surveillance in the country. METHODS: Filter paper blood blots collected in 2015-2016 from children aged below 9 years presenting with uncomplicated malaria at hospitals in three sentinel sites Begoro, Cape Coast and Navrongo were used. Parasite DNA was extracted from 120 samples followed by nested polymerase chain reaction (nPCR). Sanger sequencing was performed to detect and identify SNPs in pfap2mu and pfubp1 genes. RESULTS: In all, 11.1% (9/81) of the isolates carried the wildtype genotypes for both genes. A total of 164 pfap2mu mutations were detected in 67 isolates whilst 271 pfubp1 mutations were observed in 72 isolates. The majority of the mutations were non-synonymous (NS): 78% (128/164) for pfap2mu and 92.3% (250/271) for pfubp1. Five unique samples had a total of 215 pfap2mu SNPs, ranging between 15 and 63 SNPs per sample. Genotypes reportedly associated with ART resistance detected in this study included pfap2mu S160N (7.4%, 6/81) and pfubp1 E1528D (7.4%, 6/81) as well as D1525E (4.9%, 4/81). There was no significant difference in the prevalence of the SNPs between the three ecologically distinct study sites (pfap2mu: χ2 = 6.905, df = 2, P = 0.546; pfubp1: χ2 = 4.883, df = 2, P = 0.769). CONCLUSIONS: The detection of pfap2mu and pfubp1 genotypes associated with ACT delayed parasite clearance is evidence of gradual nascent emergence of resistance in Ghana. The results will serve as baseline data for surveillance and the selection of the genotypes with drug pressure over time. The pfap2mu S160N, pfubp1 E1528D and D1525E must be monitored in Ghanaian isolates in ACT susceptibility studies, especially when cure rates of ACTs, particularly AL, is less than 100%.
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Complejo 2 de Proteína Adaptadora/genética , Resistencia a Medicamentos/genética , Plasmodium falciparum/genética , Polimorfismo de Nucleótido Simple/genética , Proteínas de Unión al ARN/genética , Proteasas Ubiquitina-Específicas/genética , Antimaláricos/farmacología , Antimaláricos/uso terapéutico , Artemisininas/efectos adversos , Artemisininas/uso terapéutico , Niño , Preescolar , Femenino , Genotipo , Ghana/epidemiología , Humanos , Lactante , Malaria Falciparum/tratamiento farmacológico , Malaria Falciparum/epidemiología , Malaria Falciparum/parasitología , Masculino , Mutación , Plasmodium falciparum/aislamiento & purificación , PrevalenciaRESUMEN
Unlike other countries, the chloroquine resistant marker Pfcrt T76 mutant has remained fairly stable in Ghana several years after official disuse of chloroquine. Certain mutations in Pfmdr1 may potentiate Pfcrt T76, offering a possible explanation for this observation. To understand the phenomenon, the co-existence of mutations in Pfmdr1 with Pfcrt T76 in Ghanaian Plasmodium falciparum isolates was studied. The reported presence of parasites with reduced sensitivity to amodiaquine and quinine in the country was also studied. Blood samples collected from confirmed malaria patients presenting at health facilities in two distinct ecological zones were analyzed. The prevalence of Pfcrt K76T and the five point mutations in Pfmdr1 were determined using nested PCR followed by RFLP analysis. The association between genes was determined by chi square analysis, and synergism between the two genes was ascertained using the Jonckheere-Terptra (J-T) test followed by Monte Carlo simulation (MCS). Nearly fifty-four percent (53.7%) of the P. falciparum isolates examined had the Pfcrt T76 gene, out of which 18.3% had both K76 and T76 alleles. Mutations at codon 86, 184, 1034, 1042 and 1246 of the Pfmdr1 gene were detected in 36.0%, 87.9%, 71.0%, 91.6% and 8.4% of the isolates, respectively. The haplotypes of Pfmdr1 present were NFCDD (43.46%), YFCDD (27.57%), NFSDD (7.48%), NYSNY (5.14%) and YFSDD (4.67%). Pfcrt T76 was significantly associated with a double mutation at codon 86 and 184 of Pfmdr1 (YF; χ2=18.045, p=0.006). Associations were observed between Pfcrt K76T and Pfmdr1 triple mutation at codons 86, 184 and 1034 (NFC; χ2=13.770, p=0.032 and YFC; χ2=16.489, p=0.011). The J-T test showed significant synergism between Pfcrt 76 and Pfmdr1 polymorphisms (p<0.0001), which was confirmed by MCS at 99% CI. Synergism between Pfcrt and Pfmdr1 mutant genes could account for the slow recovery of chloroquine sensitive P. falciparum in Ghana. The same phenomenon could explain resistance to amodiaquine and quinine. The outcomes of this study also indicated a possible emergence of artemether-lumefantrine resistance in Ghana.
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Antimaláricos/farmacología , Cloroquina/farmacología , Resistencia a Medicamentos , Inestabilidad Genómica , Proteínas de Transporte de Membrana/genética , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Plasmodium falciparum/efectos de los fármacos , Proteínas Protozoarias/genética , Estudios Transversales , Frecuencia de los Genes , Ghana , Humanos , Proteínas de Transporte de Membrana/metabolismo , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Mutación Missense , Mutación Puntual , Proteínas Protozoarias/metabolismoRESUMEN
BACKGROUND: We investigated the prevalence of sub-microscopic Plasmodium falciparum infections and gametocyte carriage in asymptomatic individuals in Navrongo in northern Ghana, an area of seasonal malaria transmission. DESIGN: A cross sectional study of 209 randomly selected participants of all age-groups was conducted in February and March, 2015. METHODS: Capillary blood samples collected from these individuals were used for the detection of both asexual and gametocyte stage parasites by microscopy, reverse transcriptase polymerase chain reaction (RT-PCR) and conventional nested PCR methods. The prevalence data as determined by microscopy and molecular methods were compared using chi-square tests. RESULTS: Parasitaemia from these asymptomatic infections ranged from 40 to 3,520 parasites/µl of blood (geometric mean parasitaemia = 732 parasites/µl). The prevalence of asymptomatic P. falciparum carriage was 4.8% (10/209) and 13.9% (29/209) using microscopy and RT-PCR respectively. The overall prevalence of sub-microscopic infections in the total number of samples analysed was 9.1% (19/209) and 66% (19/29) of the asymptomatic infections. P. falciparum gametocytemia detected by microscopy was 1% (2/209) and 3.8% (8/209) by PCR. CONCLUSION: This is the first report of sub-microscopic asexual and gametocytes infections in the dry season in a seasonal malaria transmission area in Ghana. It has established that persistent latent malaria infections occur and that these could supply the source of parasites for the next transmission season. The findings highlight the presence of sub-microscopic infections and therefore the need for active case detection surveillance to eliminate "asymptomatic reservoir" parasites and consequently break the transmission of the disease in Ghana. FUNDING: Bill and Melinda Gates Foundation grant awarded to Noguchi Memorial Institute for Medical Research Postdoctoral and Postgraduate Training in Infectious Diseases Research (Global Health Grant # OPP52155); National Institutes of Health grant (NIH-NIAID RO1 # 1RO1AI099623) to MDW; European Developing Countries Clinical Trials Partnership (EDCTP)-West African Network of Excellence for Clinical Trials in TB, AIDS and Malaria (WANETAM) (Project code CB.07.41700.007).
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Enfermedades Asintomáticas , Portador Sano/epidemiología , Malaria Falciparum/epidemiología , Adolescente , Adulto , Animales , Niño , Preescolar , Estudios Transversales , Femenino , Ghana/epidemiología , Humanos , Masculino , Microscopía , Persona de Mediana Edad , Plasmodium falciparum/genética , Reacción en Cadena de la Polimerasa , Prevalencia , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Estaciones del Año , Adulto JovenRESUMEN
BACKGROUND: Genotyping malaria parasites to assess their diversity in different geographic settings have become necessary for the selection of antigenic epitopes for vaccine development and for antimalarial drug efficacy or resistance investigations. This study describes the genetic diversity of Plasmodium falciparum isolates from uncomplicated malaria cases over a ten year period (2003-2013) in Ghana using the polymorphic antigenic marker, merozoite surface protein 2 (msp2). METHODS: Archived filter paper blood blots from children aged nine years and below with uncomplicated malaria collected from nine sites in Ghana were typed for the presence of the markers. A total of 880 samples were genotyped for msp2 for the two major allelic families, FC27 and 3D7, using nested polymerase chain reaction (PCR). The allele frequencies and the multiplicity of infection were determined for the nine sites for five time points over a period of ten years, 2003-2004, 2005-2006, 2007-2008, 2010 and 2012-2013 malaria transmission seasons. RESULTS: The number of different alleles detected for the msp2 gene by resolving PCR products on agarose gels was 14. Both of the major allelic families, 3D7 and FC27 were common in all population samples. The highest multiplicity of infection (MOI) was observed in isolates from Begoro (forest zone, rural site): 3.31 for the time point 2007-2008. A significant variation was observed among the sites in the MOIs detected per infection (Fisher's exact test, P < 0.001) for the 2007 isolates and also at each of the three sites with data for three different years, Hohoe, P = 0.03; Navrongo, P < 0.001; Cape Coast, P < 0.001. Overall, there was no significant difference between the MOIs of the three ecological zones over the years (P = 0.37) and between the time points when data from all sites were pooled (P = 0.40). CONCLUSIONS: The diversity and variation between isolates detected using the msp2 gene in Ghanaian isolates were observed to be profound; however, there was homogeneity throughout the three ecological zones studied. This is indicative of gene flow between the parasite populations across the country probably due to human population movements (HPM).
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Antígenos de Protozoos/genética , Variación Genética , Malaria Falciparum/parasitología , Plasmodium falciparum/clasificación , Plasmodium falciparum/genética , Proteínas Protozoarias/genética , Niño , Preescolar , Femenino , Frecuencia de los Genes , Técnicas de Genotipaje , Ghana , Humanos , Lactante , Recién Nacido , Masculino , Plasmodium falciparum/aislamiento & purificación , Reacción en Cadena de la PolimerasaRESUMEN
The emergence and spread of Plasmodium falciparum resistance to commonly used antimalarials such as chloroquine and sulphadoxine/pyrimethamine poses major challenges to malaria control in sub-Saharan Africa. We undertook a study on the efficacy of some antimalarial drugs in 2003 with the view of supporting the National Malaria Control Programme in the review of the antimalarial drug treatment policy in Ghana. Children aged 6-59 months with signs/symptoms of uncomplicated malaria including axillary temperature > or =37.5 degrees C; mono infection with P. falciparum; and parent's willingness to give consent, were randomized into four treatment groups and followed up for a maximum of 28 days. The treatment groups were chloroquine (CHQ), sulphadoxine/pyrimethamine (SP), amodiaquine+artesunate (ADQ+ART) combination, and artemether+lumefantrine (Coartem) combination. Clinical evaluation of 168 children studied showed that cumulative pcr-corrected cure rates on day 28 were 100% for ADQ+ART; 97.5% for coartem, 60% for SP and 25% for CHQ. The artemisinin-based combinations effected rapid fever and parasite clearance. Prevalence of gametocytaemia was highest in the SP group whilst the CHQ group did not show any significant changes in haemoglobin levels during the follow-up period. The findings are in agreement with current recommendations for using artemisinin-based combinations for treating uncomplicated malaria in areas of high CHQ failure such as Ghana.
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Amodiaquina/uso terapéutico , Antimaláricos/uso terapéutico , Artemisininas/uso terapéutico , Cloroquina/uso terapéutico , Malaria Falciparum/tratamiento farmacológico , Pirimetamina/uso terapéutico , Sesquiterpenos/uso terapéutico , Sulfadoxina/uso terapéutico , Amodiaquina/administración & dosificación , Antimaláricos/administración & dosificación , Artemisininas/administración & dosificación , Artesunato , Preescolar , Cloroquina/administración & dosificación , Combinación de Medicamentos , Quimioterapia Combinada , Femenino , Ghana , Humanos , Lactante , Masculino , Pirimetamina/administración & dosificación , Sesquiterpenos/administración & dosificación , Sulfadoxina/administración & dosificación , Resultado del TratamientoRESUMEN
In 2005, sulphadoxine-pyrimethamine (SP) became the drug of choice for intermittent preventive treatment of Plasmodium falciparum malaria in pregnancy (IPTp) in Ghana. Reports suggest the use of SP by others to treat uncomplicated malaria. Because of the increased use of SP, the prevalence of mutations in the genes, dihydrofolate reductase (dhfr), and dihydropteroate synthetase (dhps), linked to SP resistance in P. falciparum were determined. Blood samples from 945 children with uncomplicated malaria collected at nine sites from 2003 to 2010 were analyzed using polymerase chain reaction and restriction fragment length polymorphism. Prevalence of the dhfr triple and dhfr plus dhps quadruple mutations showed significant increase in trend from 2003 to 2010 (χ(2) = 18.78, P < 0.001, χ(2) = 15.11, P < 0.001, respectively). For dhps double mutant G437 + E540 the prevalence was low (1.12%) caused by the very low prevalence of E540. Our findings show the wide use of SP in Ghana and therefore its use for IPTp needs to be closely monitored.
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Antimaláricos/farmacología , Resistencia a Medicamentos , Malaria Falciparum/parasitología , Plasmodium falciparum/efectos de los fármacos , Pirimetamina/farmacología , Sulfadoxina/farmacología , Oxidorreductasas de Alcohol , Alelos , Antimaláricos/uso terapéutico , Biomarcadores , Dihidropteroato Sintasa/genética , Dihidropteroato Sintasa/metabolismo , Combinación de Medicamentos , Regulación Enzimológica de la Expresión Génica , Ghana/epidemiología , Política de Salud , Humanos , Lactante , Malaria Falciparum/tratamiento farmacológico , Malaria Falciparum/epidemiología , Plasmodium falciparum/enzimología , Reacción en Cadena de la Polimerasa , Polimorfismo de Longitud del Fragmento de Restricción , Vigilancia de la Población , Pirimetamina/uso terapéutico , Sulfadoxina/uso terapéutico , Tetrahidrofolato Deshidrogenasa/genética , Tetrahidrofolato Deshidrogenasa/metabolismo , Factores de TiempoRESUMEN
BACKGROUND: Epstein-Barr virus (EBV) and cytomegalovirus (CMV) are persistent herpesviruses that have various immunomodulatory effects on their hosts. Both viruses are usually acquired in infancy in Sub-Saharan Africa, a region where childhood vaccines are less effective than in high income settings. To establish whether there is an association between these two observations, we tested the hypothesis that infection with one or both viruses modulate antibody responses to the T-cell independent meningococcal polysaccharide vaccine and the T-cell dependent measles vaccines. METHODOLOGY/PRINCIPAL FINDINGS: Infection with EBV and CMV was diagnosed by the presence of virus-specific IgM in the peripheral blood or by the presence of IgG at higher levels than that found in umbilical cord blood. Anti-meningococcus IgG and IgM were quantified by ELISA. Anti-measles antibody responses were quantified by haemagglutinin antibody inhibition assay. Infants infected with EBV had reduced IgG and IgM antibody responses to meningococcal polysaccharides and to measles vaccine. Infection with CMV alone predicted no changes in the response to meningococcal polysaccharide. While CMV alone had no discernable effect on the antibody response to measles, the response of infants infected with both CMV and EBV was similar to that of infants infected with neither, suggesting that the effects of CMV infection countered the effects of EBV on measles antibody responses. CONCLUSIONS: The results of this exploratory study indicate that infection with EBV is associated with reduced antibody responses to polysaccharides and to measles vaccine, but suggest that the response to T-cell dependent antigens such as measles haemagglutinin may be restored by infection with CMV.