Clinical and epidemiological profiles of severe malaria in children from Delhi, India

Plasmodium vivax is traditionally known to cause benign tertian malaria, although recent reports suggest that P. vivax can also cause severe life-threatening disease analogous to severe infection due to P. falciparum. There are limited published data on the clinical and epidemiological profiles of children suffering from 'severe malaria' in an urban setting of India. To assess the clinical and epidemiological profiles of children with severe malaria, a prospective study was carried out during June 2008-December 2008 in the Department of Pediatrics, Guru Teg Bahadur Hospital, a tertiary hospital located in East Delhi, India. Data on children aged < or = 12 years, diagnosed with severe malaria, were analyzed for their demographic, clinical and laboratory parameters. All patients were categorized and treated as per the guidelines of the World Health Organization. In total, 1,680 children were screened for malaria at the paediatric outpatient and casualty facilities of the hospital. Thirty-eight children tested positive for malaria on peripheral smear examination (2.26% slide positivity rate). Of these, 27 (71%) were admitted and categorized as severe malaria as per the definition of the WHO while another 11 (29%) received treatment on outpatient basis. Most (24/27; 88.8%) cases of severe malaria (n=27) were infected with P. vivax. Among the cases of severe malaria caused by Plasmodium vivax (n=24), 12 (50%) presented with altered sensorium (cerebral malaria), seven (29.1%) had severe anaemia (haemoglobin <5 g/dL), and 17 (70.8%) had thrombocytopaenia, of which two had spontaneous bleeding (epistaxis). Cases of severe vivax malaria are clinically indistinguishable from severe falciparum malaria. Our study demonstrated that majority (88.8%) of severe malaria cases in children from Delhi and adjoining districts of Uttar Pradesh were due to P. vivax-associated infection. P. vivax should, thus, be regarded as an important causative agent for severe malaria in children.     

一种简易疟原虫PCR模板制备方法

目的:寻找一种高效、简便、快速疟原虫PCR模板制备方法。方法:收集2010年-2011年自非洲、东南亚等疟疾流行区回国人员中疟疾现症患者(显微镜检查阳性),采滤纸血,用蒸馏水直接溶血法和Chel-ex-100法分别制备疟原虫PCR模板,采用巢式PCR技术检测疟原虫ss RNA基因,并对两法结果进行比较。结果:分别用蒸馏水直接溶血法和Chelex-100法制备疟原虫PCR模板32个,其中恶性疟原虫26个,间日疟原虫6个,直接溶血法巢式PCR检测全部出现特异性扩增带,而Chelex-100法均未出现扩增带。结论:蒸馏水直接溶血法是一种较为理想制备疟原虫PCR模板的方法。     

Effectiveness of primaquine terminal prophylaxis against late primary attacks of Plasmodium vivax malaria: a case-control study among troops of the Republic of Korea army

Plasmodium vivax malaria is an important cause of morbidity among troops operating in endemic areas near the Demilitarized Zone in the Republic of Korea (ROK). The ROK Army has been administering antimalarial chemoprophylaxis to those troops at greatest risk of malaria since 1997. The number of recipients increased from 15000 in 1997 to 90000 in 2001. We undertook a case-control study to estimate the effectiveness of primaquine prophylaxis against late primary attacks of P. vivax malaria in ROK Army troops. Microscopically confirmed cases of P. vivax malaria were identified through hospital-based surveillance. Controls were matched by unit. Between 1 November 2001 and 31 May 2002, 68 cases and 137 matched controls with confirmed chemoprophylaxis status were enrolled. The estimated effectiveness of primaquine prophylaxis was 32% (95% CI 23-63%). Our results suggest that the effectiveness of primaquine prophylaxis against late primary attacks of P. vivax malaria may be insufficient for soldiers of the ROK Army.     

Malaria relapse and recrudescence among travellers to the tropics

This study describes 14 cases of relapse and recrudescence of malaria, treated between 1991 and 2003. In that period, 146 patients were hospitalized in the Clinic of the Institute in Gdynia: 20 women and 126 men. In 103 cases the disease was caused by Plasmodium falciparum, in 31 cases by Plasmodium vivax, in 5 cases by Plasmodium malariae, and in 2 cases by Plasmodium ovale. Five patients were found to have mixed infections, with either P. falciparum and P. vivax or P. falciparum and P. ovale. Relapses in patients previously treated in the country or abroad accounted for 9.6% of all the treated cases of malaria. Recrudescences and relapses were diagnosed of both the tropical malaria (6 cases), and the tertian malaria caused by P. vivax (4 cases). Moreover, in 4 patients diagnosis was made of secondary malaria due to P. vivax infection, while the primary attack was caused by invasion of P. falciparum. Also discussed was the issue of drug-resistance of plasmodia and the resulting problems with the treatment.     

Demonstration by the polymerase chain reaction of mixed Plasmodium falciparum and P. vivax infections undetected by conventional microscopy.

Mixed malaria infections (Plasmodium falciparum and P. vivax) are suspected to occur at a greater frequency than is detected by conventional light microscopy. To determine this frequency we carried out a prospective 'blinded' comparison of diagnosis by conventional light microscopy and enzymatic amplification of the circumsporozoite gene extracted from dried spotted blood samples. Patients were previously healthy, active duty Thai soldiers assigned to a malaria risk area presenting with malaria. Microscopy (oil immersion objective at 1000 x magnification) involved examination of Giemsa-stained thick and thin blood films by an experienced microscopist. Whole blood samples (25 microliters) dried on filter paper were used for species-specific parasite deoxyribonucleic acid (DNA) amplification by the polymerase chain reaction (PCR) and hybridization with radiolabelled P. falciparum and P. vivax probes. Of 137 consecutive cases of malaria studied, 9% (3/32) of microscopically diagnosed P. falciparum infections and 5% (5/104) of microscopically diagnosed P. vivax infections were found to be mixed by the PCR/DNA probe systems, while 1 case was diagnosed as mixed by both microscopy and PCR. The possibility that malaria patients may have undetected mixed infections should be kept in mind because of the specific therapy required both for P. falciparum and for radical cure of P. vivax.     

Haematinic treatment of anaemia increases the risk of Plasmodium vivax malaria in pregnancy

Nutritional deficiency and malaria are 2 major causes of anaemia during pregnancy in tropical areas. The relationship between anaemia, its treatment with iron and folate, and malaria was studied in a prospective cohort of 2112 pregnant Karen women on the north-western border of Thailand between 1993 and 1997. The development of Plasmodium vivax malaria was associated with a past mean haematocrit > 30% (hazard ratio = 1.5, 95% CI 1.2-2, P = 0.001) and recent (< or = 30 d) iron and folate supplementation (hazard ratio = 1.7, 95% CI 1.1-2.6, P = 0.01). There were no associations with P. falciparum infections. Plasmodium vivax has a predilection for young erythrocytes, and these results suggest that pregnant women with larger numbers of circulating young red cells are at greater risk of developing P. vivax malaria. In P. vivax-endemic areas, systematic iron and folate supplementation confers both benefit and risk in pregnancy.     

Retinal haemorrhage in vivax malaria

Retinal haemorrhage is often observed in patients with Plasmodium falciparum, especially when combined with cerebral malaria. However, few cases of retinopathy have been reported in P. vivax malaria. Benign tertian malaria has re-emerged among soldiers in the South Korean demilitarized zone since 1993. We report an indigenous case of retinal haemorrhage caused by P. vivax and review the relevant literature.     

Endemic malaria in four villages of the Pakistani province of Punjab

The prevalence of malaria in 4 villages 60 km south of Lahore, Punjab, where Anopheles culicifacies is the vector was estimated from blood films made during three mass malaria surveys (MS) and at 204 clinics (CS) held over 18 consecutive months. The highest parasite rate occurred during October 1984 when 43% of the CS population had parasitaemias. Plasmodium vivax predominated early in the major transmission season (23% and 15% vivax parasitaemias in August 1983 and 1984 respectively) whereas P. falciparum was the most common species later in the transmission season (an average falciparum prevalence of 37% in October and November 1984) and following the transmission season through March. Despite the proximity and habitat similarity of the 4 villages, both total and species malaria prevalence rates showed inter-village differences. Asexual stage and gametocyte parasite rates in children were 2 to 5 times higher than in adults. No increased mortality due to malaria was detected among the 4000 study population. Malaria was stable and endemic, albeit seasonally transmitted, in these 4 villages during 1983 and 1984.     

The detection of Plasmodium falciparum and P. vivax in DNA-extracted blood samples using polymerase chain reaction

Seventeen pairs of published primer sets were compared for their relative sensitivity to detect malaria DNA extracted from blood samples, which were obtained from Pakistani patients suffering from malaria. The primer sets investigated consisted of: (i) 9 pairs of direct primers and 3 sets of nested primers for detecting Plasmodium falciparum, (ii) 2 pairs of direct primers and 2 sets of nested primers for detecting P. vivax, and (iii) 1 set of multiplex primers for detecting both P. falciparum and P. vivax, simultaneously. After a miniscreen of 9 DNA-extracted blood samples using the 17 primer sets stated above, 5 primer sets were short-listed (based on their superior sensitivity) and used for a maxi-screen of DNA extracted from 126 microscopy-positive blood samples from Pakistan, with the following results. (i) For the detection of P. falciparum, the direct primer pair 'PF1 + PF2' gave a sensitivity of 95% and the nested primer set 'RIT405 + RIT406/RIT371 + RIT372' gave a sensitivity of 97%. (ii) For the detection of P. vivax, the direct primer pair 'Forward + Reverse' and the nested primer set 'PLF + UNR/PLF + VIR' both gave a sensitivity of 94%. (iii) The nested multiplex primer set 'rPLU5 + rPLU6/rFAL1 + rFAL2 + rVIV1 + rVIV2' gave a sensitivity of 97% and 96% for P. falciparum and P. vivax, respectively. It was concluded that the nested multiplex primer set was the most optimal primer set to use for the detection of malaria DNA extracted from blood samples. Furthermore, the nested multiplex primer set has the advantage of simultaneously detecting and differentiating between P. vivax and P. falciparum.     

Plasmodium vivax circumsporozoite variants and Duffy blood group genotypes in the Brazilian Amazon region

The circumsporozoite protein (CSP) of the Plasmodium vivax infective sporozoite is considered to be a major target for the development of recombinant malaria vaccines. The Duffy blood group molecule acts as the red blood cell receptor for P. vivax. We review the frequency of P. vivax CSP variants and report their association with the Duffy blood group genotypes from Brazilian Amazon patients carrying P. vivax malaria. Peripheral blood samples were collected from 155 P. vivax-infected individuals from five Brazilian malaria-endemic areas. The P. vivax CSP variants and the Duffy blood group genotypes were assessed using PCR/RFLP. In single infections, the VK210 variant was the commonest followed by the P. vivax-like variant. The typing of P. vivax indicated that the frequency of variants among the study areas was significantly different from one to another. This is the first detection of the VK247 and P. vivax-like variant in single infections in endemic areas of Brazil. Association of the CSP P. vivax variants with the heterozygous Duffy blood group system genotype was significant for VK210 single infection. These observations provide additional data on the Plasmodium-host interactions concerning the Duffy blood group and P. vivax capability of causing human malaria.     

Malaria and splenomegaly in the Punjab

Splenomegaly and parasitaemia were correlated in 2891 children examined in outpatient clinics during 13 consecutive months in 4 Punjabi villages. The average monthly splenomegaly rate varied from 0.05-0.13 before the monsoon malaria transmission season to 0.18-0.27 during and after this season. Moderate splenic enlargement reached a peak during the malaria transmission season, while the highest proportion of very enlarged spleens occurred towards the end of, and after, the transmission season. Children with splenomegaly were 3 times more likely to have Plasmodium falciparum and 1.5 times more likely to have P. vivax parasitaemias than were children without palpable spleens. The larger the spleen the more likely a P. falciparum infection, whereas P. vivax was more commonly associated with minimal spleen enlargement. Although the probability of a child with splenomegaly having a malaria parasitaemia was highest (0.58-0.72) during and immediately following periods of malaria transmission, the odds ratio of malaria infections among those with splenomegaly to those without was at its lowest (1.41-2.11) during those months. Mean P. falciparum parasitaemias were significantly higher in infected children with moderately enlarged spleens than in infected children with nonpalpable spleens or in those with minimal or extensive splenomegaly. These results are compatible with splenomegaly being a result of both the malaria infection and the immune response. Early in infection many children had parasitaemia without splenomegaly; after the parasitaemia had cleared splenomegaly often persisted.     

Risk of Plasmodium vivax malaria reintroduction in Uzbekistan: genetic characterization of parasites and status of potential malaria vectors in the Surkhandarya region

Plasmodium vivax malaria was eradicated from Uzbekistan in 1961. Due to resurgence of the disease in neighbouring states and massive population migration, there has been an increase of P. vivax malaria, imported from Tajikistan, resulting in a number of indigenous cases being identified in areas bordering that country. A molecular study using the merozoite surface protein 1 (msp-1) gene as a marker was performed on 24 P. vivax genomic isolates from 12 indigenous and 10 imported malaria cases that occurred in the Surkhandarya region during the summer of 2002. Results have shown a significant difference in the frequency of msp-1 types between indigenous and imported isolates, the latter showing greater genetic heterogeneity. An entomological investigation in the area suggested that three Anopheles species, namely A. superpictus, A. pulcherrimus and A. hyrcanus may have a potential role in the endemic transmission of P. vivax.     

Therapeutic efficacy of chloroquine in Plasmodium vivax from areas with different epidemiological patterns in India and their Pvdhfr gene mutation pattern

Among the four human malaria parasites, drug resistance occurs mainly in Plasmodium falciparum. However, there are some reports of chloroquine (CQ) resistance in P. vivax from different geographical regions. In India, approximately 50% of a total of 2 million cases of malaria reported annually are due to P. vivax. CQ is the drug of choice for treatment. Since few cases of treatment failure have been reported from India, this study was undertaken to generate data systematically on the efficacy of CQ in 287 patients from different epidemiological regions. Cure rates for 28 days were 100% and there was a rapid parasite clearance rate in all age groups from all study sites. Although P. vivax has been reported to be inherently resistant to sulfonamide and pyrimethamine, Indian isolates exhibited only double mutations in dhfr in vitro.     

Sulfadoxine-pyrimethamine plus artesunate compared with chloroquine for the treatment of vivax malaria in areas co-endemic for Plasmodium falciparum and P. vivax: a randomised non-inferiority trial in eastern Afghanistan

Chloroquine (CQ) is an effective treatment of choice for vivax malaria in most settings, but with the spread of CQ-resistant Plasmodium falciparum, many countries now use artemisinin-based combination therapy for treatment of falciparum malaria. In areas co-endemic for falciparum and vivax malaria incorrect differential diagnosis is always a risk. In Afghanistan the adoption of sulfadoxine-pyrimethamine plus artesunate (SP+AS) as first-line falciparum treatment raises the prospect of a significant proportion of vivax malaria being misdiagnosed and treated with the combination. SP is considered to have limited efficacy against vivax malaria, and the efficacy of SP+AS against Plasmodium vivax has not been established in areas that are using SP+AS. A randomised, non-inferiority trial comparing SP+AS with CQ monotherapy was undertaken on 190 vivax malaria patients in eastern Afghanistan. Standard WHO procedures for in vivo evaluation of antimalarial drugs were followed. A total of 180 individuals completed the trial to day 42. Using a per protocol analysis, both regimens resulted in > or =96% treatment success at 28 d, but significantly more cases failed in the CQ arm (46%) than in the SP+AS arm (24%) by day 42. In areas where vivax infections might be misdiagnosed as falciparum infections and treated with SP+AS, patient management would be as good, or better than, with the standard CQ treatment.     

Therapeutic response of multidrug-resistant Plasmodium falciparum and P. vivax to chloroquine and sulfadoxine-pyrimethamine in southern Papua, Indonesia

To determine the level of antimalarial drug resistance in southern Papua, Indonesia, we assessed the therapeutic efficacy of chloroquine plus sulfadoxine-pyrimethamine (CQ+SP) for Plasmodium falciparum infections as well as CQ monotherapy for P. vivax infections. Patients with P. falciparum failing therapy were re-treated with unsupervised quinine+/-doxycycline therapy and those with P. vivax with either unsupervised quinine+/-doxycycline or amodiaquine. In total, 143 patients were enrolled in the study (103 treated with CQ+SP and 40 with CQ). Early treatment failures occurred in four patients (4%) with P. falciparum and six patients (15%) with P. vivax. The failure rate by Day 28 for P. vivax was 65% (95% CI 49-81). After PCR correction for re-infections, the Day 42 recrudescence rate for P. falciparum infections was 48% (95% CI 31-65). Re-treatment with unsupervised quinine+/-doxycycline resulted in further recurrence of malaria in 48% (95% CI 31-65) of P. falciparum infections and 70% (95% CI 37-100) of P. vivax infections. Eleven patients with recurrent P. vivax were re-treated with amodiaquine; there were no early or late treatment failures. In southern Papua, a high prevalence of drug resistance of P. falciparum and P. vivax exists both to first- and second-line therapies. Preliminary data indicate that amodiaquine retains superior efficacy compared with CQ for CQ-resistant P. vivax.     

Chloroquine blood concentrations and molecular markers of chloroquine-resistant Plasmodium falciparum in febrile children in northern Ghana

Plasmodium falciparum malaria is a predominant reason for health care utilization among children in sub-Saharan Africa. Despite the spread of resistance, chloroquine (CQ) is the most commonly used antimalarial. Little is known about the pattern of CQ use and resistance to the drug prior to attendance at a health care facility, and its impact on clinical presentation in children attending health care facilities in endemic regions. In a cross-sectional study among 840 febrile children presenting at a primary health care facility in northern Ghana from September to December 2000, CQ blood levels were measured by enzyme-linked immunosorbent assay and parasite isolates were genotyped for the CQ resistance markers pfcrt T76 and pfmdr1 Y86. Plasmodium falciparum was present in 95% by polymerase chain reaction and CQ was detected in 64% of the children. Concentrations of CQ in blood ranged from 31 to 3897 nmol/L (median 198 nmol/L). The pfcrt T76 and pfmdr1 Y86 resistance markers were detected in 84% and 57% of the isolates, respectively, and were selected by CQ. A significant trend for higher frequencies of the resistance markers with increasing CQ concentrations was observed. In this typical primary health care setting in sub-Saharan Africa, CQ use prior to attendance at a health care facility and CQ-resistant P. falciparum are frequent. As CQ selects resistant P. falciparum genotypes, CQ should be omitted as a first-line drug even in primary health care facilities when self-treatment with CQ is common.     

Malaria in southern Mozambique: malariometric indicators and malaria case definition in Manhiça district

A total of 2057 children aged <10 years were selected at random from a demographic surveillance system and enrolled in 4 malariometric cross-sectional surveys in different seasons in Manhi?a district in southern Mozambique. Plasmodium falciparum accounted for 90% of all malaria infections and the prevalence of asexual P. falciparum ranged from 13.7-21.7% at the end of the dry season to 30.5-34.0% at the end of rainy season. In order to determine the malaria attributable fraction (MAF) of fever, 1021 children from a nearby hospital acted as fever cases and from this separate case-control study the crude MAF was 36%, showing a marked age dependency. Plasmodium falciparum is the most common malaria species in Manhi?a. This malaria-mesoendemic area has year-round transmission. The importance of other non-malarial fever-causing conditions among infants was highlighted. Malaria appeared to be a major contributory factor to anaemia in the area.     

The malaria and typhoid fever burden in the slums of Kolkata, India: data from a prospective community-based study

Recent research has indicated that the malaria burden in Asia may have been vastly underestimated. We conducted a prospective community-based study in an impoverished urban site in Kolkata, India, to estimate the burden of malaria and typhoid fever and to identify risk factors for these diseases. In a population of 60452 people, 3605 fever episodes were detected over a 12-month period. The blood films of 93 febrile patients contained Plasmodium (90 P. vivax, 2 P. falciparum and 1 P. malariae). Blood cultures from 95 patients grew Salmonella enterica serotype Typhi. Malaria patients were found to be significantly older (mean age 29 years) compared with patients with typhoid fever (15 years; P<0.001) but had similar clinical features on presentation. Having a household member with malaria, illiteracy, low household income and living in a structure not built of bricks were associated with an increased risk for malaria. Having a household member with typhoid fever and poor hygiene were associated with typhoid fever. A geographic analysis of the spatial distribution of malaria and typhoid fever cases detected high-risk neighbourhoods for each disease. Focal interventions to minimise human-vector contact and improved personal hygiene and targeted vaccination campaigns could help to prevent malaria and typhoid fever in this site.     

Distribution of Plasmodium vivax variants (VK210, VK247 and P. vivax-like) in three endemic areas of the Amazon region of Brazil and their correlation with chloroquine treatment

The present study evaluated the glass fibre membrane (GFM)-polymerase chain reaction (PCR)-enzyme-linked immunosorbent assay (ELISA) technique for genotyping the Plasmodium vivax variants, to verify the distribution of P. vivax variants (VK210, VK247 and P. vivax-like) in parts of Brazil and their correlation with levels of parasitaemia, previous malaria experience and clearance of parasitaemia linked to different treatment schedules. The samples were taken from individuals living in Macapá, Porto Velho and Belém, all of which are endemic areas of vivax malaria in the Amazon region of Brazil. Blood samples were collected on GFMs. The gene that codes for the circumsporozoite proteins of P. vivax variants was amplified by PCR and the amplified fragments were hybridized to variant-specific, digoxigenin-labelled oligonucleotide probes by ELISA. The GFM-PCR-ELISA technique was shown to be accurate for epidemiological surveys of the vivax complex. All variants were detected in all 3 areas, but only P. vivax VK210 was found as a single agent of infection, while the other 2 occurred as mixed infections. The P. vivax-like variant was found to be associated with low parasitaemia and VK210 with the highest parasitaemia levels; none of the P. vivax variants was linked with a previous malaria experience. In all cases parasitaemia clearance was identical regarding the type of treatment and consequently it is not possible to confirm the previously reported correlation between P. vivax genotype and response to chloroquine.     

Health centre versus home presumptive diagnosis of malaria in southern Ghana: implications for home-based care policy

A study was conducted in 1997 to compare the accuracy of presumptive diagnosis of malaria in children aged 1-9 years performed by caretakers of the children to that of health centre staff in 2 ecological zones in southern Ghana. Similar symptoms were reported in the children at home and at the health centre. In the home setting, symptoms were reported the same day that they occurred, 77.6% of the children with a report of fever were febrile (axillary temperature > or = 37.5 degrees C) and 64.7% of the reports of malaria were parasitologically confirmed. In the health centre, the median duration of symptoms before a child was seen was 3 days (range 1-14 days), 58.5% of the children with a report of fever were febrile and 62.6% of the clinically diagnosed cases were parasitologically confirmed. In the 2 settings almost all the infections were due to Plasmodium falciparum. Parasite density was 3 times higher in the health centre cases compared to the home-diagnosed cases. Early and appropriate treatment of malaria detected in children by caretakers may prevent complications that arise as a result of persistence of symptoms and attainment of high parasitaemic levels.