Plasmodium falciparum msp-1 and msp-2 genetic diversity and multiplicity of infection in isolates from Congolese patients in the Republic of Congo.

With limited up to date data from the Republic of Congo, the aim of this study was to investigate allelic polymorphism of merozoite surface protein-1 (msp-1) and merozoite surface protein-2 (msp-2). This will help assess the genetic diversity and multiplicity of Plasmodium falciparum infection (MOI), from uncomplicated malaria individuals living in Brazzaville. Between March and October 2021, a cross-sectional study was carried out at a health center in Madibou District located in the south of Brazzaville. Plasmodium infection was diagnosed in human blood by microscopy and the block 2 of P. falciparum msp-1 and block 3 of msp-2 genes were genotyped by nested PCR. Overall, 57 genotypes with fragment sizes ranging from 110 to 410 bp were recorded for msp-1, among which 25, 21, and 11 genotypes identified for K1, MAD20, and RO33 allelic families respectively. RO33 (34.3%) and MAD20 (34.3%) allelic families were more frequent compared to K1 (31.4%) although the difference was not statistically significant. Also, 47 msp-2 genotypes were identified, including 26 FC27 genotypes type, and 21 genotypes belonging to the 3D7 allelic family. FC27 was more frequent (52.3%) compared to 3D7 (47.7%). The prevalence of the polyclonal infection was 90.0% while the MOI was 2.90 ± 1.0. The MOI and polyclonal infection were not significantly associated with the parasitaemia and anaemia. This study reveals a high genetic diversity and the trend of increasing MOI of P. falciparum isolates from the south of Brazzaville, compared to the reports from the same setting before the COVID-19 pandemic.

Human African trypanosomiasis (HAT) in the Republic of Congo: why the Congolese population is reluctant to screening?

Introduction: human African trypanosomiasis (HAT) is a neglected tropical infection, and surveillance of the disease relies on community participation in screening. This study aimed to identify the main factors associated with low community uptake of the HAT screening in endemic districts in the Republic of Congo. Methods: a cross-sectional survey was carried out during a sensitisation campaign about HAT in the districts of Mpouya, Ngabé and Loudima, which are endemic for the disease. After signing the informed consent form, participants were organized into groups of 10 for focus group discussions (FGDs). A list of questions was used for guiding the discussion, addressing understanding of the disease and reasons for refusing screening. Results: out of 220 recruited individuals (corresponding to 22 FGDs), 58.6% were men. The majority of the respondents described HAT as a rural disease (48.2%) or as a witchcraft (22.3%). Among the clinical signs cited by the participants, sleep disorder (40%) was the most common answer, followed by prolonged fever (19.5%) and madness (14.1%). The main reasons for non-adherence to HAT screening was the fear of lumbar puncture (45.9%) and stigmatisation (22.3%). Conclusion: the findings of this study suggest that more effort should be put into raising awareness of HAT and the benefits of screening amongst the Congolese population, in order to strengthen the national disease control program.

Seroprevalence of anti-SARS-CoV-2 antibodies in a population living in Bomassa village, Republic of Congo.

Objectives: With limited data available from Central Africa, the aim of our study was to evaluate the anti-SARS-CoV-2 Ab prevalence in indigenous residents of Bomassa, a village located in the Sangha region in the Republic of Congo. Methods: Plasma and oropharyngeal swab samples were collected from 304 healthy adult individuals, randomly recruited in May 2021 before vaccine introduction in the area. In addition, 82 plasma samples from the same area in 2019 were included as controls for the investigation of cross-reactivity against other coronaviruses. The SARS-CoV-2 virus was detected by qRT-PCR and sequenced using next-generation sequencing. ELISA was used for detecting IgG, IgM, and neutralizing Ab against SARS-CoV-2 antigens. Results: Around 4.9% (15/304) of the participants were SARS-CoV-2 positive, with B.1.631 being the only variant identified. Of 109 individuals harboring anti-SARS-CoV-2 IgG and/or IgM Ab, 45.9% (50/109) had anti-SARS-CoV-2 neutralizing Ab. Of the control samples collected before the pandemic, 3.7% (3/82) were positive for IgG, but negative for neutralizing Ab. Conclusions: Seroprevalence against SARS-CoV-2 occurred in 25% of the indigenous population sample, with almost 50% of these seropositive participants possessing neutralizing antibodies. These findings suggest that the spread of SARS-CoV-2 has been underestimated in the Republic of Congo.

Prevalence of non-Plasmodium falciparum species in southern districts of Brazzaville in The Republic of the Congo.

BACKGROUND: Although Plasmodium falciparum infection is largely documented and this parasite is the main target for malaria eradication, other Plasmodium species persist, and these require more attention in Africa. Information on the epidemiological situation of non-P. falciparum species infections is scarce in many countries, including in the Democratic Republic of the Congo (hereafter Republic of the Congo) where malaria is highly endemic. The aim of this study was to determine the prevalence and distribution of non-P. falciparum species infections in the region south of Brazzaville. METHODS: A cross-sectional survey was conducted in volunteers living in rural and urban settings during the dry and rainy seasons in 2021. Socio-demographic and clinical parameters were recorded. Plasmodium infection in blood samples was detected by microscopic analysis and nested PCR (sub-microscopic analysis). RESULTS: Of the 773 participants enrolled in the study, 93.7% were from the rural area, of whom 97% were afebrile. The prevalence of microscopic and sub-microscopic Plasmodium spp. infection was 31.2% and 63.7%, respectively. Microscopic Plasmodium malariae infection was found in 1.3% of participants, while sub-microscopic studies detected a prevalence of 14.9% for P. malariae and 5.3% for Plasmodium ovale. The rate of co-infection of P. malariae or P. ovale with P. falciparum was 8.3% and 2.6%, respectively. Higher rates of sub-microscopic infection were reported for the urban area without seasonal fluctuation. In contrast, non-P. falciparum species infection was more pronounced in the rural area, with the associated risk of the prevalence of sub-microscopic P. malariae infection increasing during the dry season. CONCLUSION: There is a need to include non-P. falciparum species in malaria control programs, surveillance measures and eradication strategies in the Republic of the Congo.

Plasmodium falciparum multiplicity of infection and pregnancy outcomes in Congolese women from southern Brazzaville, Republic of Congo.

BACKGROUND: Investigating whether the multiplicity of Plasmodium falciparum infection (MOI) is related to pregnancy outcomes, is of interest in sub-Saharan area where malaria is highly endemic. The present study aimed to characterize the genetic diversity of P. falciparum in women at delivery from Southern Brazzaville, and investigate whether the MOI is associated with maternal anaemia, preterm delivery, or low birth weight. METHODS: This was a cross sectional study carried out with samples collected between March 2014 and April 2015 from 371 women recruited at delivery at a Health Centre in southern Brazzaville, Republic of Congo. Matched peripheral, placental, and cord blood collected from each of the women at delivery were used for the detection of P. falciparum microscopic and submicroscopic parasitaemia, and parasite DNA genotyping by nested PCR. RESULTS: From 371 recruited women, 27 were positive to microscopic malaria parasitaemia while 223 women harboured submicroscopic parasitaemia. All msp-1 block 2 family allelic types (K1, MAD20 and RO33) were observed in all the three compartments of blood, with K1 being most abundant. K1 (with 12, 10, and 08 alleles in the peripheral, placental, and cord blood respectively) and MAD20 (with 10, 09, and 06 alleles in the respective blood compartments) were more diverse compared to RO33 (with 06, 06, and 05 alleles in the respective blood compartments). From the 250 women with microscopic and/or submicroscopic parasitaemia, 38.5%, 30.5%, and 18.4% of peripheral, placental and cord blood sample, respectively, harboured more than one parasite clone, and polyclonal infection was more prevalent in the peripheral blood of women with microscopic parasitaemia (54.5%) compared to those with submicroscopic parasitaemia (36.7%) (p = 0.02). The mean multiplicity of genotypes per microscopic and submicroscopic infection in peripheral blood was higher in anemic women (2.00 ± 0.23 and 1.66 ± 0.11, respectively) than in non-anaemic women (1.36 ± 0.15 and 1.45 ± 0.06, respectively) (p = 0.03 and 0.06). In logistic regression, women infected with four or more clones of the parasite were 9.4 times more likely to be anaemic than women harbouring one clone. This association, however, was only observed with the peripheral blood infection. No significant association was found between the MOI and low birth weight or preterm delivery. CONCLUSIONS: These results indicate that the genetic diversity of P. falciparum is high in pregnant women from southern Brazzaville in the Republic of Congo, and the multiplicity of the infection might represent a risk for maternal anaemia.

Occurrence of human astrovirus associated with gastroenteritis among Congolese children in Brazzaville, Republic of Congo.

INTRODUCTION: Acute viral gastroenteritis is a major public health concern, especially among children younger than 5 years of age. The aim of this study was to determine the occurrence of human astrovirus infection in children with acute gastroenteritis. METHODS: Stool specimens were collected from 506 children under 5 years of age hospitalized with acute diarrhoea (289 male and 208 female), and human astrovirus was investigated by RT-PCR. Associations of socio-demographic, clinical, and behavioural conditions with infection were analysed. RESULTS: The overall prevalence of human astrovirus was found to be 10.3%. The mean age of positive cases was 12.41 ± 6.21 months and this was associated with infection (p = 0.013). Children >18 months of age were at three times the risk of infection when compared to those aged 0-6 months (odds ratio (OR) 3.19, 95% confidence interval (CI) 1.15-8.88; p = 0.026). Children living in houses with more than one room (OR 0.60, 95% CI 0.28-0.96; p = 0.036) and mothers using treated water (OR 0.47, 95% CI 0.25-0.86; p = 0.015) were associated with reduced infection. CONCLUSIONS: In this study, infection with astrovirus was common in acute gastroenteritis cases among children younger than 5 years of age. Drinking treated water and living in non-crowded environments protected the children from infection.

High prevalence of norovirus and rotavirus co-infection in children with acute gastroenteritis hospitalised in Brazzaville, Republic of Congo.

OBJECTIVE: To evaluate the clinical severity of diarrhoea associated to viral co-infection in children with acute gastroenteritis. METHODS: About 461 children under five years hospitalised with acute diarrhoea (266 males and 187 females) were enrolled in the study. Using stool samples, rotavirus and adenovirus infections were investigated by ELISA, and norovirus infections by nested duplex RT-PCR. We assessed social, demographic, clinical and behavioural conditions that might influence the occurrence of rotavirus, adenovirus and norovirus infections. RESULTS: Mono-viral infection was detected in 49% and mixed viral infection in 12% of patients. The prevalence of mixed infection was neither dependent on age nor sex. Three samples were infected with all three viruses. A significant association was found between fever (axillary temperature> 37.5 °C) and rotavirus-norovirus dual infection (aOR (CI 95%) = 2.1 (1.14-3.84), P = 0.016; aOR (CI 95%) = 0.37 (0.19-0.73), P = 0.004). Mixed infection was the most common during the dry season from June to October (71.4% versus 54.7%, P = 0.023). CONCLUSION: Co-infection with both rotavirus and norovirus is common in under-five hospitalised children but does not contribute to the severity of the disease.

OBJECTIF: Evaluer la sévérité clinique de la diarrhée associée à la coinfection virale chez les enfants atteints de gastroentérite aiguë. MÉTHODES: 461 enfants de moins de cinq ans hospitalisés pour une diarrhée aiguë (266 garçons et 187 filles) ont été inclus dans l'étude. Sur des échantillons de selles, les infections à rotavirus et à adénovirus ont été investiguées par ELISA et les infections à norovirus par RT-PCR duplex imbriqué. Nous avons évalué les conditions sociales, démographiques, cliniques et comportementales susceptibles d'influencer la survenue d'infections à rotavirus, adénovirus et norovirus. RÉSULTATS: Une infection mono virale a été détectée chez 49% des patients et une infection virale mixte chez 12% des patients. La prévalence des infections mixtes ne dépendait ni de l'âge ni du sexe. Trois échantillons étaient infectés par tous les trois virus. Une association significative a été observée entre la fièvre (température axillaire > 37,5 °C) et la double infection rotavirus-norovirus (aOR (IC95%) = 2,1 (1,14-3,84), P = 0,016; aOR (IC95%) = 0,37 (0,19-0,73), P = 0,004). Les infections mixtes étaient les plus courantes pendant la saison sèche de juin à octobre (71,4% contre 54,7%, P = 0,023). CONCLUSION: La coinfection à la fois par le rotavirus et par le norovirus est fréquente chez les enfants de moins de cinq ans hospitalisés, mais ne contribue pas à la sévérité de la maladie.

Molecular characterization of norovirus infection responsible for acute diarrhea in Congolese hospitalized children under five years old in Brazzaville, Republic of Congo.

BACKGROUND: Acute diarrhea is a leading cause of morbidity and mortality among children under five worldwide. As no published data is available on the occurrence of this infection in the Republic of Congo, this study aimed at (1) determining the prevalence and (2) characterizing genotypes of norovirus strains in Brazzaville. METHODS: From June 2012 to June 2013, stool samples were collected from hospitalized young children with acute gastroenteritis. A total of 545 samples were tested for GI and GII norovirus infections using nested duplex reverse-transcription-polymerase chain reaction and sequencing. RESULTS: The GI and GII norovirus infection were detected in 148 samples. Males (28%) were not significantly more infected than females (25%). Norovirus infection was found exclusively in children aged under 24 months with a higher prevalence (P=0,048) in the age group of 7-12 months, and throughout the year with a peak in August and September. Genetic diversity of norovirus strains revealed that GII was the most prevalent (87%). No risk factor was significantly associated with norovirus infection. CONCLUSION: This study showed that noroviruses are important agents responsible for acute diarrhea in Congolese children and highlights the importance of continued surveillance.

Distribution of the cytochrome P450 CYP2C8*2 allele in Brazzaville, Republic of Congo.

BACKGROUND: Cytochrome P450 (CYP) enzymes are essential in the metabolism of most drugs used today. Single nucleotide polymorphism(s) occurring in CYP genes can adversely affect drug pharmacokinetics, efficacy, and safety. Individuals carrying the CYP2C8*2 c.805A > T (CYP2C8*2; rs11572103) allele have impaired amodiaquine metabolism, increased risk of amodiaquine-related adverse events, and may promote the selection of drug-resistant parasite strains. This study investigated the distribution of the CYP2C8*2 allele in Brazzaville, Republic of Congo, where artesunate + amodiaquine is used as the second-line treatment for uncomplicated Plasmodium falciparum malaria. METHODS: A total of 285 febrile children visiting the Marien Ngouabi paediatric hospital were genotyped for CYP2C8*2 using PCR-restriction fragment length polymorphism (PCR-RFLP). The allele frequencies and genotype distribution were determined. RESULTS: The CYP2C8*2 allele was successfully genotyped in 75% (213/285) of the study participants. The CYP2C8*2A allele had a frequency of 63%, whereas the CYP2C8*2T allele had a frequency of 37%. Genotypes CYP2C8*2AA (rapid metabolizer), CYP2C8*2AT (intermediate metabolizer), and CYP2C8*2TT (poor metabolizer) were observed in 44%, 38%, and 18% of the investigated participants, respectively. CONCLUSIONS: This study gives the first description of CYP2C8*2 allele distribution in the Republic of Congo and highlights the potential risk of amodiaquine-related adverse events. Information from this study will be beneficial during pharmacovigilance investigations.

Sub-microscopic Plasmodium falciparum infections in matched peripheral, placental and umbilical cord blood samples from asymptomatic Congolese women at delivery.

In malaria-endemic areas, most pregnant women are susceptible to asymptomatic Plasmodium falciparum infections. We present here the results of a cross-sectional study conducted in Madibou, a southern district of Brazzaville in the Republic of Congo, between March 2014 and April 2015. The main aim was to characterize P. falciparum infections. Blood samples corresponding to peripheral, placental and cord from 370 asymptomatic malaria women at delivery were diagnosed for plasmodium infection by thick blood smears (microscopic infection). Sub-microscopic infection was detected by PCR, using the MSP-2 gene as marker. Microscopic infections were detected in peripheral, placental and cord blood samples with a prevalence of respectively 7.3% (27/370), 2.7% (10/370) and 0%. The negative samples were submitted to sub-microscopic detection, with respective prevalence of 25.4% (87/343), 16.7% (60/360) and 9.4% (35/370) (P < 0.001). We further investigated the genetic diversity of the parasite by characterizing MSP2 allelic families 3D7 (24 distinct alleles) and FC27 (20 distinct alleles). The total number of alleles for these two families were 31, 25 and 19 in peripheral, placental and cord samples respectively. The 3D7 MSP-2 was the predominant allelic family. The multiplicity of infections (MOI) in peripheral (mean 1.4 ± 0.01; range 1-4), placental (mean 1.2 ± 0.01; range 1-3) and cord samples (1.4 ± 0.01; range 1-3) were similar (P = 0.9) and are unaffected by age, gravidity or sulfadoxine-pyrimethamine. These results shown a high prevalence of sub-microscopic infection and a high genetic diversity of Plasmodium falciparum strains in Congo. Age, gravidity and doses of preventive treatment based on sulfadoxine-pyrimethamine do not interfere with the multiplicity of infections.

Cytochrome P450 CYP2B6*6 distribution among Congolese individuals with HIV, Tuberculosis and Malaria infection.

BACKGROUND: The cytochrome P450 CYP2B6*6 (CYP2B6 c.516G>T; rs3745274) is one of the genetic factors that alters the drug metabolism in antimalarial, antiretroviral and TB first-line drugs. In Central African populations, the distribution of the CYP2B6*6 variant is poorly documented. This study investigated the distribution of CYP2B6 c.516G>T variant among Congolese individuals. METHODS: A total of 418 patients with HIV-1 mono-infection, HIV-1 and Tuberculosis coinfection and symptomatic P. falciparum malaria were genotyped for the CYP2B6 c.516G>T SNP using Restriction Fragment Length Polymorphism (RFLP). The allele frequencies and genotype distributions were determined. RESULTS: The CYP2B6 c.516G>T was successfully analysed in 69% (288/418) of the study participants. Among the investigated individuals, the distribution of the major allele CYP2B6*G was 45% and the minor CYP2B6*T allele was 55%. Significant differences in genotype distribution were also observed among the studied individuals. The CYP2B6*GG (rapid metabolizer) genotype was observed in 17% (49/288) followed by CYP2B6*GT (intermediate metabolizer) 55% (159/288) and CYP2B6*TT (poor metabolizers) 28% (80/288). CONCLUSION: This study contributes to increasing understanding on population pharmacogenetics and may help policy makers regulate treatment guidelines in the Congolese population with a high burden of HIV, Malaria and TB.

An update on glucose-6-phosphate dehydrogenase deficiency in children from Brazzaville, Republic of Congo.

BACKGROUND: Malaria transmission-blocking anti-malarial drugs, such as primaquine, offers an effective strategy for reducing the incidence of falciparum malaria. However, this drug induces haemolytic anaemia among glucose-6-phosphate dehydrogenase (G6PD) deficient individuals. The distribution of G6PD deficiency in Brazzaville, Republic of Congo and the association of G6PD deficiency with haemoglobin levels and blood cell counts were investigated. METHODS: A total of 212 febrile children were recruited for this study. Plasmodium falciparum diagnosis was conducted by microscopy and nested PCR. Sanger sequencing was used to assess G6PD deficiency by detecting 202G>A (rs1050828) and 376A>G (rs1050829) single nucleotide polymorphisms. RESULTS: Two hundred and twelve children were successfully genotyped for G6PD variants. Overall, 13% (27/212) of the children were G6PD deficient and 25% (25/100) females were heterozygous (11 BA- and 14 A+A-). The remaining 160 children had a normal G6PD genotype. The mean red blood and mean platelet counts were significantly lower in hemizygous male (G6PD A-) participants than in normal male (G6PD A+ or B) participants (p < 0.05). CONCLUSION: This study gives an update on G6PD deficiency among Congolese children. Understanding the distribution of G6PD deficiency in other geographical regions is recommended before primaquine is adopted in the malaria control programme.

Plasmodium falciparum merozoite protein-1 genetic diversity and multiplicity of infection in isolates from Congolese children consulting in a pediatric hospital in Brazzaville.

As in many sub-Saharan African countries, the burden of malaria has been reduced in the Republic of Congo as a result of massive deployment of insecticide treated nets and availability of artemisinin-combinations therapies (ACTs). High to moderate genetic diversity of msp-1 gene of Plasmodium falciparum (P. falciparum) has been reported from different parts of the world but limited data are available from Central Africa including the Republic of Congo. For this reason, the aim of study was to investigate the P. falciparum genetic diversity and to determine the multiplicity of infection in P. falciparum isolates from Congolese children in order to dispose of an additional parameter to measure the impact malaria control intervention. A total of 229 blood samples were collected from September 2014 to February 2015 in children aged from one to ten years presenting a paediatric hospital Marien NGOUABI located in Northern part of Brazzaville. Inclusion criterion was fever (axillary temperature ≥ 37.5 °C) or history of fever in the preceding 48 h before inclusion in this study. Then thick and thin blood smears were done to detect malaria parasites, to determine parasite density and to identify plasmodial species. Sub-microscopic infection was detected by PCR using the P. falciparum msp-1 gene as molecular marker. The prevalence of microscopic and sub-microscopic infection in this cohort was 10% and 27.5%, respectively. The K1 allelic family was predominant (45% of isolates) whereas the RO33 and MAD20 represented 35% and 20%, respectively of isolates. In this study 48% (38/79) of isolates harbored more than one parasite clone. Overall the multiplicity of infection (MOI) was 1.7. According to type of infection, the MOI was significantly higher in children with microscopic infection (2.5 vs 1.4 for submicroscopic infection, P = .001). When considering age, hemoglobin genotype (AA or AS) and level and parasite density, no association was observed with the MOI. This study reveals that the P. falciparum genetic diversity in isolates from Congolese children is high but with low multiplicity of infection.

Plasmodium falciparum infection in febrile Congolese children: prevalence of clinical malaria 10 years after introduction of artemisinin-combination therapies.

OBJECTIVES: To investigate the proportion of malaria infection in febrile children consulting a paediatric hospital in Brazzaville, to determine the prevalence of submicroscopic malaria infection, to characterise Plasmodium falciparum infection and compare the prevalence of uncomplicated P. falciparum malaria according to haemoglobin profiles. METHODS: Blood samples were collected from children aged <10 years with an axillary temperature ≥37.5 °C consulting the paediatric ward of Marien Ngouabi Hospital in Brazzaville. Parasite density was determined and all samples were screened for P. falciparum by nested polymerase chain reaction (PCR) using the P. falciparum msp-2 marker to detect submicroscopic infections and characterise P. falciparum infection. Sickle cell trait was screened by PCR. RESULTS: A total of 229 children with fever were recruited, of whom 10% were diagnosed with uncomplicated malaria and 21% with submicroscopic infection. The mean parasite density in children with uncomplicated malaria was 42 824 parasites/µl of blood. The multiplicity of infection (MOI) was 1.59 in children with uncomplicated malaria and 1.69 in children with submicroscopic infection. The mean haemoglobin level was 10.1 ± 1.7 for children with uncomplicated malaria and 12.0 ± 8.6 for children with submicroscopic infection. About 13% of the children harboured the sickle cell trait (HbAS); the rest had normal haemoglobin (HbAA). No difference in prevalence of uncomplicated malaria and submicroscopic infection, parasite density, haemoglobin level, MOI and P. falciparum genetic diversity was observed according to haemoglobin type. CONCLUSION: The low prevalence of uncomplicated malaria in febrile Congolese children indicates the necessity to investigate carefully other causes of fever.

Diagnosis of smear-negative pulmonary tuberculosis based on clinical signs in the Republic of Congo.

BACKGROUND: The diagnosis of pulmonary tuberculosis (PTB) and smear-negative pulmonary tuberculosis (SNPT) in resource-limited countries is often solely based on clinical signs, chest X-ray radiography and sputum smear microscopy. We investigated currently used methods for the routine diagnosis of SNPT in the Republic of Congo (RoC) among TB suspected patients. The specific case of HIV positive patients was also studied. METHODS: A cross-sectional study was conducted at the anti-tuberculosis center (CAT) of Brazzaville, RoC. Tuberculosis suspects were examined for physical signs of TB. Clinical signs, results from sputum smear microscopy, tuberculin skin test (TST) and chest X-ray were recorded. RESULTS: Of the 772 enrolled participants, 372 were diagnosed PTB. Cough was a common symptom for PTB and no PTB patients. Pale skin, positive TST, weight loss and chest X-ray with abnormalities compatible with PTB (PTB-CXR) were significant indicators of PTB. Thirty-six percent of PTB patients were diagnosed SNPT. This category of patients presented less persistent cough and less PTB-CXR. Anorexia and asthenia were significant indicators of SNPT. In the case of HIV+ patients, 57% were SNPT with anorexia, asthenia and shorter cough being strong indicators of SNPT. CONCLUSION: Chest X-ray abnormalities, weight loss, pale skin and positive TST were significant indicators of PTB. Anorexia and asthenia showed good diagnostic performance for SNPT, which deserve to be recommended as index indicators of SNPT diagnosis. Duration of cough is also a relevant indicator, especially for HIV+ patients.

High prevalence of sulphadoxine-pyrimethamine resistance-associated mutations in Plasmodium falciparum field isolates from pregnant women in Brazzaville, Republic of Congo.

Intermittent preventive treatment during pregnancy with sulfadoxine-pyrimethamine (IPTp-SP) has not been evaluated in the Republic of Congo since its implementation in 2006 and there is no published data on molecular markers of SP resistance among Plasmodium falciparum isolates from pregnant women. This first study in this country aimed to describe the prevalence of dihydrofolate reductase (dhfr) and dihydropteroate synthase (dhps) point mutations and haplotypes in P. falciparum isolates collected from pregnant women with asymptomatic infection. From March 2012 to December 2013, pregnant women attending Madibou health centre (in Southern Brazzaville) for antenatal visits were enrolled in this study after obtaining their written informed consent. Blood samples were collected and P. falciparum infections were characterized using PCR. A total of 363 pregnant women were enrolled. P. falciparum infection was detected in 67 (18.4%) samples as their PCR amplification of dhfr and dhps genes yielded bands and all the PCR products were successfully digested. Out of these 67 isolates, 59 (88%), 57 (85%) and 53 (79.1%) carried 51I, 59R and 108N dhfr mutant alleles, respectively. The prevalence of dhps 436A, 437G and 540E mutations were 67.1% (45/67), 98.5% (66/67) and 55.2% (37/67), respectively. More than one-half of the isolates carried quintuple mutations, with highly resistant haplotype dhfr51I/59R/108N + dhps437G/540E detected in 33% (22/67) whereas 25% (17/67) were found to carry sextuple mutations. We observed significantly higher frequencies of triple dhps mutations 436A/437G/540E and quintuple mutations dhfr51I/59R/108N+dhps437G/540E in isolates from women who received IPTp-SP than those who did not. Overall, this study shows high prevalence rates of SP-associated resistance mutations in P. falciparum isolates collected from pregnant women. The presence of the dhps mutant allele 540E and the high prevalence of isolates carrying quintuple dhfr/dhps mutations are here reported for the first time in the Republic of Congo. The increasing prevalence of multiple mutant alleles observed in this study is alarming and may present a challenge for the future interventions including IPTp-SP in the country.

Genetic polymorphism of merozoite surface protein 2 and prevalence of K76T pfcrt mutation in Plasmodium falciparum field isolates from Congolese children with asymptomatic infections.

BACKGROUND: In order to prepare the field site for future interventions, the prevalence of asymptomatic Plasmodium falciparum infection was evaluated in a cohort of children living in Brazzaville. Plasmodium falciparum merozoite surface protein 2 gene (msp2) was used to characterize the genetic diversity and the multiplicity of infection. The prevalence of mutant P. falciparum chloroquine resistance transporter (pfcrt) allele in isolates was also determined. METHODS: Between April and June 2010, 313 children below 10 years of age enrolled in the cohort for malaria surveillance were screened for P. falciparum infection using microscopy and polymerase chain reaction (PCR). The children were selected on the basis of being asymptomatic. Plasmodium falciparum msp2 gene was genotyped by allele-specific nested PCR and the pfcrt K76T mutation was detected using nested PCR followed by restriction endonuclease digestion. RESULTS: The prevalence of asymptomatic P. falciparum infections was 8.6% and 16% by microscopy and by PCR respectively. Allele typing of the msp2 gene detected 55% and 45% of 3D7 and FC27 allelic families respectively. The overall multiplicity of infections (MOI) was 1.3. A positive correlation between parasite density and multiplicity of infection was found. The prevalence of the mutant pfcrt allele (T76) in the isolates was 92%. CONCLUSION: This is the first molecular characterization of P. falciparum field isolates in Congolese children, four years after changing the malaria treatment policy from chloroquine (CQ) to artemisinin-based combination therapy (ACT). The low prevalence of asymptomatic infections and MOI is discussed in the light of similar studies conducted in Central Africa.