Knowledge, attitudes, and practices related to soil-transmitted helminth infections among residents of Bata district, Equatorial Guinea; a cross-sectional study.

BACKGROUND: Soil-transmitted helminth (STH) infection control remains a priority in endemic regions where local epidemiological data are needed for sustainable control strategies, particularly regarding population knowledge, attitudes, and practices (KAP). Our work assessed KAP toward STH infection and associated factors among residents of Bata district, Equatorial Guinea. METHODS: A community-based cross-sectional study was conducted among 14 randomly selected communities in the Bata district. Eligible participants were interviewed face-to-face using a standardized questionnaire. Participants aged under 18 years were interviewed in the presence of their parents or legal guardians. For participants aged less than ten, a simplified version of the main questionnaire was developed focusing on children's practices toward STH and was administered to their parents or legal guardians. RESULTS: A total of 399 participants were included in the present analysis. Among them, 58% responded to the main questionnaire. The mean (± SD) age of participants aged 10 and over was 37.5 (± 22.2) years, and 60% of them were females, while the mean (± SD) age of those aged less than ten was 5.0 (± 2.5) years. The respondents' overall knowledge, attitudes, and practices to STH were rated as bad (33%), very good (77%), and good (55%), respectively. Knowledge was significantly associated with education level (p = 0.04) with the knowledge level lower for participants with no formal education than for those with secondary/university education (ß = -0.56, 95% CI: -1.00 - -0.12, p = 0.01); Appropriate attitudes level was significantly associated with occupation (p = 0.02) and education levels (p = 0.049) with the appropriate attitude level lower for students than for farmers/fishers (ß = -1.24, 95% CI: -2.17--0.32, p = 0.01) and for primary-level participants than for those with secondary/university education (ß = -0.68, 95% CI: -1.23--0.13, p = 0.02); while appropriate practice level were significantly associated with age (p = 0.01), occupation (p = 0.01), and education (p = 0.02), with the appropriate practices level increasing with age (ß = 0.03, 95% CI: 0.005 - 0.05, p = 0.01) and lower in participants with no formal education than in those with secondary/university education (ß = -1.19, -2.05 - -0.32, p = 0.007). CONCLUSION: The present study revealed a lack of knowledge about STH in the study population, particularly regarding disease causes and transmission ways, highlighting the need for the implementation of integrated health education strategies, both at the community and school levels.

Analysis of nucleic acids extracted from rapid diagnostic tests reveals a significant proportion of false positive test results associated with recent malaria treatment.

BACKGROUND: Surveillance programmes often use malaria rapid diagnostic tests (RDTs) to determine the proportion of the population carrying parasites in their peripheral blood to assess the malaria transmission intensity. Despite an increasing number of reports on false-negative and false-positive RDT results, there is a lack of systematic quality control activities for RDTs deployed in malaria surveillance programmes. METHODS: The diagnostic performance of field-deployed RDTs used for malaria surveys was assessed by retrospective molecular analysis of the blood retained on the tests. RESULTS: Of the 2865 RDTs that were collected in 2018 on Bioko Island and analysed in this study, 4.7% had a false-negative result. These false-negative RDTs were associated with low parasite density infections. In 16.6% of analysed samples, masked pfhrp2 and pfhrp3 gene deletions were identified, in which at least one Plasmodium falciparum strain carried a gene deletion. Among all positive RDTs analysed, 28.4% were tested negative by qPCR and therefore considered to be false-positive. Analysing the questionnaire data collected from the participants, this high proportion of false-positive RDTs could be explained by P. falciparum histidine rich protein 2 (PfHRP2) antigen persistence after recent malaria treatment. CONCLUSION: Malaria surveillance depending solely on RDTs needs well-integrated quality control procedures to assess the extent and impact of reduced sensitivity and specificity of RDTs on malaria control programmes.

Rapid Identification of SARS-CoV-2 Variants of Concern Using a Portable peakPCR Platform.

The need for tools that facilitate rapid detection and continuous monitoring of SARS-CoV-2 variants of concern (VOCs) is greater than ever, as these variants are more transmissible and therefore increase the pressure of COVID-19 on healthcare systems. To address this demand, we aimed at developing and evaluating a robust and fast diagnostic approach for the identification of SARS-CoV-2 VOC-associated spike gene mutations. Our diagnostic assays detect the E484K and N501Y single-nucleotide polymorphisms (SNPs) as well as a spike gene deletion (HV69/70) and can be run on standard laboratory equipment or on the portable rapid diagnostic technology platform peakPCR. The assays achieved excellent diagnostic performance when tested with RNA extracted from culture-derived SARS-CoV-2 VOC lineages and clinical samples collected in Equatorial Guinea, Central-West Africa. Simplicity of usage and the relatively low cost are advantages that make our approach well suitable for decentralized and rapid testing, especially in resource-limited settings.

Incidence of Plasmodium falciparum malaria infection in 6-month to 45-year-olds on selected areas of Bioko Island, Equatorial Guinea.

BACKGROUND: Extensive malaria control measures have been implemented on Bioko Island, Equatorial Guinea over the past 16 years, reducing parasite prevalence and malaria-related morbidity and mortality, but without achieving elimination. Malaria vaccines offer hope for reducing the burden to zero. Three phase 1/2 studies have been conducted successfully on Bioko Island to evaluate the safety and efficacy of whole Plasmodium falciparum (Pf) sporozoite (SPZ) malaria vaccines. A large, pivotal trial of the safety and efficacy of the radiation-attenuated Sanaria® PfSPZ Vaccine against P. falciparum is planned for 2022. This study assessed the incidence of malaria at the phase 3 study site and characterized the influence of socio-demographic factors on the burden of malaria to guide trial design. METHODS: A cohort of 240 randomly selected individuals aged 6 months to 45 years from selected areas of North Bioko Province, Bioko Island, was followed for 24 weeks after clearance of parasitaemia. Assessment of clinical presentation consistent with malaria and thick blood smears were performed every 2 weeks. Incidence of first and multiple malaria infections per person-time of follow-up was estimated, compared between age groups, and examined for associated socio-demographic risk factors. RESULTS: There were 58 malaria infection episodes observed during the follow up period, including 47 first and 11 repeat infections. The incidence of malaria was 0.25 [95% CI (0.19, 0.32)] and of first malaria was 0.23 [95% CI (0.17, 0.30)] per person per 24 weeks (0.22 in 6-59-month-olds, 0.26 in 5-17-year-olds, 0.20 in 18-45-year-olds). Incidence of first malaria with symptoms was 0.13 [95% CI (0.09, 0.19)] per person per 24 weeks (0.16 in 6-59-month-olds, 0.10 in 5-17-year-olds, 0.11 in 18-45-year-olds). Multivariate assessment showed that study area, gender, malaria positivity at screening, and household socioeconomic status independently predicted the observed incidence of malaria. CONCLUSION: Despite intensive malaria control efforts on Bioko Island, local transmission remains and is spread evenly throughout age groups. These incidence rates indicate moderate malaria transmission which may be sufficient to support future larger trials of PfSPZ Vaccine. The long-term goal is to conduct mass vaccination programmes to halt transmission and eliminate P. falciparum malaria.

Quantifying malaria acquired during travel and its role in malaria elimination on Bioko Island.

BACKGROUND: Malaria elimination is the goal for Bioko Island, Equatorial Guinea. Intensive interventions implemented since 2004 have reduced prevalence, but progress has stalled in recent years. A challenge for elimination has been malaria infections in residents acquired during travel to mainland Equatorial Guinea. The present article quantifies how off-island contributes to remaining malaria prevalence on Bioko Island, and investigates the potential role of a pre-erythrocytic vaccine in making further progress towards elimination. METHODS: Malaria transmission on Bioko Island was simulated using a model calibrated based on data from the Malaria Indicator Surveys (MIS) from 2015 to 2018, including detailed travel histories and malaria positivity by rapid-diagnostic tests (RDTs), as well as geospatial estimates of malaria prevalence. Mosquito population density was adjusted to fit local transmission, conditional on importation rates under current levels of control and within-island mobility. The simulations were then used to evaluate the impact of two pre-erythrocytic vaccine distribution strategies: mass treat and vaccinate, and prophylactic vaccination for off-island travellers. Lastly, a sensitivity analysis was performed through an ensemble of simulations fit to the Bayesian joint posterior probability distribution of the geospatial prevalence estimates. RESULTS: The simulations suggest that in Malabo, an urban city containing 80% of the population, there are some pockets of residual transmission, but a large proportion of infections are acquired off-island by travellers to the mainland. Outside of Malabo, prevalence was mainly attributable to local transmission. The uncertainty in the local transmission vs. importation is lowest within Malabo and highest outside. Using a pre-erythrocytic vaccine to protect travellers would have larger benefits than using the vaccine to protect residents of Bioko Island from local transmission. In simulations, mass treatment and vaccination had short-lived benefits, as malaria prevalence returned to current levels as the vaccine's efficacy waned. Prophylactic vaccination of travellers resulted in longer-lasting reductions in prevalence. These projections were robust to underlying uncertainty in prevalence estimates. CONCLUSIONS: The modelled outcomes suggest that the volume of malaria cases imported from the mainland is a partial driver of continued endemic malaria on Bioko Island, and that continued elimination efforts on must account for human travel activity.

Genetic diversity and natural selection on the thrombospondin-related adhesive protein (TRAP) gene of Plasmodium falciparum on Bioko Island, Equatorial Guinea and global comparative analysis.

BACKGROUND: Thrombospondin-related adhesive protein (TRAP) is a transmembrane protein that plays a crucial role during the invasion of Plasmodium falciparum into liver cells. As a potential malaria vaccine candidate, the genetic diversity and natural selection of PfTRAP was assessed and the global PfTRAP polymorphism pattern was described. METHODS: 153 blood spot samples from Bioko malaria patients were collected during 2016-2018 and the target TRAP gene was amplified. Together with the sequences from database, nucleotide diversity and natural selection analysis, and the structural prediction were preformed using bioinformatical tools. RESULTS: A total of 119 Bioko PfTRAP sequences were amplified successfully. On Bioko Island, PfTRAP shows its high degree of genetic diversity and heterogeneity, with π value for 0.01046 and Hd for 0.99. The value of dN-dS (6.2231, p < 0.05) hinted at natural selection of PfTRAP on Bioko Island. Globally, the African PfTRAPs showed more diverse than the Asian ones, and significant genetic differentiation was discovered by the fixation index between African and Asian countries (Fst > 0.15, p < 0.05). 667 Asian isolates clustered in 136 haplotypes and 739 African isolates clustered in 528 haplotypes by network analysis. The mutations I116T, L221I, Y128F, G228V and P299S were predicted as probably damaging by PolyPhen online service, while mutations L49V, R285G, R285S, P299S and K421N would lead to a significant increase of free energy difference (ΔΔG > 1) indicated a destabilization of protein structure. CONCLUSIONS: Evidences in the present investigation supported that PfTRAP gene from Bioko Island and other malaria endemic countries is highly polymorphic (especially at T cell epitopes), which provided the genetic information background for developing an PfTRAP-based universal effective vaccine. Moreover, some mutations have been shown to be detrimental to the protein structure or function and deserve further study and continuous monitoring.

Clustering of subpatent infections in households with asymptomatic rapid diagnostic test-positive cases in Bioko Island, Equatorial Guinea independent of travel to regions of higher malaria endemicity: a cross-sectional study.

BACKGROUND: Prevalence of falciparum malaria on Bioko Island remains high despite sustained, intensive control. Progress may be hindered by high proportions of subpatent infections that are not detected by rapid diagnostic tests (RDT) but contribute to onward transmission, and by imported infections. Better understanding of the relationship between subpatent infections and RDT-detected infections, and whether this relationship is different from imported versus locally acquired infections, is imperative to better understand the sources of infection and mechanisms of transmission to tailor more effective interventions. METHODS: Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was performed on a sub-set of samples from the 2015 Malaria Indicator Survey to identify subpatent infections. Households with RDT(+) individuals were matched 1:4 with households with no RDT(+) individuals. The association between living in a household with an RDT(+) individual and having a subpatent infection was evaluated using multivariate hierarchical logistic regression models with inverse probability weights for selection. To evaluate possible modification of the association by potential importation of the RDT(+) case, the analysis was repeated among strata of matched sets based on the reported eight-week travel history of the RDT(+) individual(s). RESULTS: There were 142 subpatent infections detected in 1,400 individuals (10.0%). The prevalence of subpatent infections was higher in households with versus without an RDT(+) individual (15.0 vs 9.1%). The adjusted prevalence odds of subpatent infection were 2.59-fold greater (95% CI: 1.31, 5.09) for those in a household with an RDT(+) individual compared to individuals in a household without RDT(+) individuals. When stratifying by travel history of the RDT(+) individual, the association between subpatent infections and RDT(+) infections was stronger in the strata in which the RDT(+) individual(s) had not recently travelled (adjusted prevalence odds ratio (aPOR) 2.95; 95% CI:1.17, 7.41), and attenuated in the strata in which recent travel was reported (aPOR 1.76; 95% CI: 0.54, 5.67). CONCLUSIONS: There is clustering of subpatent infections around RDT(+) individual(s) when both imported and local infection are suspected. Future control strategies that aim to treat whole households in which an RDT(+) individual is found may target a substantial portion of infections that would otherwise not be detected.

Rapid selection of a pyrethroid metabolic enzyme CYP9K1 by operational malaria control activities.

Since 2004, indoor residual spraying (IRS) and long-lasting insecticide-impregnated bednets (LLINs) have reduced the malaria parasite prevalence in children on Bioko Island, Equatorial Guinea, from 45% to 12%. After target site-based (knockdown resistance; kdr) pyrethroid resistance was detected in 2004 in Anopheles coluzzii (formerly known as the M form of the Anopheles gambiae complex), the carbamate bendiocarb was introduced. Subsequent analysis showed that kdr alone was not operationally significant, so pyrethroid-based IRS was successfully reintroduced in 2012. In 2007 and 2014-2015, mass distribution of new pyrethroid LLINs was undertaken to increase the net coverage levels. The combined selection pressure of IRS and LLINs resulted in an increase in the frequency of pyrethroid resistance in 2015. In addition to a significant increase in kdr frequency, an additional metabolic pyrethroid resistance mechanism had been selected. Increased metabolism of the pyrethroid deltamethrin was linked with up-regulation of the cytochrome P450 CYP9K1. The increase in resistance prompted a reversion to bendiocarb IRS in 2016 to avoid a resurgence of malaria, in line with the national Malaria Control Program plan.

Nutritional, clinical and immunological status of children at HIV diagnosis in the continental region of Equatorial Guinea.

OBJECTIVE: To evaluate the nutritional, clinical and immunological status of children at HIV diagnosis in the continental region of Equatorial Guinea. METHODS: Children <18 years diagnosed with HIV between 2009 and 2017 were included. Clinical, immunological and nutritional data were collected. Weight-for-height, weight-for-age and height-for-age Z-scores were calculated using WHO Child Growth Standards. The population was assessed in two equal periods (2009-2013 and 2014-2017) from the time of diagnosis. RESULTS: A total of 213 children were diagnosed with HIV (49.3% males), median age 3.8 years (IQR: 1.5-8.2). A total of 121 cases (56.8%) were at WHO clinical stage III, and 53 (24.9%) were at WHO clinical stage IV. CDC immunological stage II was diagnosed in 51/130 (39.2%) and CDC stage III in 44/130 (33.8%). About 56.2% of children were underweight (weight-for-age <-2 SD); 20.1% moderately and 36.1% severely so. About 27.6% of children were wasted (weight-for-height <-2 SD); 11.9% moderately and 15.7% severely so. About 56.3% of children were stunted (height-for-age <-2 SD); 20.7% moderately and 35.6% severely so. The prevalence of wasting was higher in children ≤5 years than in children >5 years (36.4% vs. 19.1%, P = 0.026). In the second period, the prevalence of moderate-severe immunodeficiency decreased (87.2% to 67.0%, P = 0.018), without significant differences in the other nutritional or clinical data. Severe underweight was a risk factor for moderate-severe immunodeficiency (aOR: 4 [95% CI: 1.4-11.4], P = 0.010). CONCLUSIONS: We highlight a high proportion of malnutrition at the time of HIV diagnosis in Guinea´s paediatric population. Early diagnosis of HIV infection is a priority, achievable by training Guinea´s physicians to suspect HIV early, introducing HIV molecular diagnostic techniques and ensuring intensive nutritional treatment.

OBJECTIF: Evaluer le statut nutritionnel, clinique et immunologique des enfants au moment du diagnostic du VIH dans la région continentale de la Guinée équatoriale. MÉTHODES: Les enfants de moins de 18 ans diagnostiqués avec le VIH entre 2009 et 2017 ont été inclus. Les données cliniques, immunologiques et nutritionnelles ont été collectées. Les scores Z du poids pour la taille, du poids pour l'âge et de la taille pour l'âge ont été calculés en utilisant les normes de croissance de l'enfant de l'OMS. La population a été évaluée en deux périodes égales (2009-2013 et 2014-2017) à partir du moment du diagnostic. RÉSULTATS: 213 enfants ont été diagnostiqués avec le VIH (49,3% de sexe masculin), âge médian de 3,8 ans (IQR: 1,5-8,2). 121 cas (56,8%) étaient au stade clinique III de l'OMS et 53 (24,9%) étaient au stade clinique IV de l'OMS. Le stade immunologique II du CDC a été diagnostiqué chez 51/130 (39,2%) et le stade III du CDC chez 44/130 (33,8%). 56,2% des enfants avaient un poids insuffisant (poids pour âge <-2 écart type [ET]); 20,1% modérément et 36,1% sévèrement. 27,6% des enfants étaient émaciés (poids pour taille <-2 ET); 11,9% modérément et 15,7% sévèrement. 56,3% des enfants avaient un retard de croissance (taille pour l'âge <-2 ET); 20,7% modérément et 35,6% sévèrement. La prévalence de l'émaciation était plus élevée chez les enfants ≤5 ans que chez les enfants > 5 ans (36,4% contre 19,1%; P = 0,026). Dans la deuxième période, la prévalence de l'immunodéficience modérée à sévère a diminué (87,2% à 67,0% ; P = 0,018), sans différences significatives dans les autres données nutritionnelles ou cliniques. L'insuffisance pondérale sévère était un facteur de risque d'immunodéficience modérée à sévère (aOR: 4 [IC95%: 1,4-11,4] ; P = 0,010). CONCLUSIONS: Nous mettons en évidence ici une forte proportion de malnutrition au moment du diagnostic du VIH dans la population pédiatrique guinéenne. Le diagnostic précoce de l'infection par le VIH est une priorité, réalisable en formant les médecins guinéens à identifier le VIH tôt, en introduisant des techniques de diagnostic moléculaire du VIH et en assurant un traitement nutritionnel intensif.

First evidence of the deletion in the pfhrp2 and pfhrp3 genes in Plasmodium falciparum from Equatorial Guinea.

BACKGROUND: The World Health Organization (WHO) recommends rapid diagnostic tests (RDTs) as a good alternative malaria-diagnosis method in remote parts of sub-Saharan Africa. The majority of commercial RDTs currently available detect the Plasmodium falciparum protein histidine-rich protein 2 (PfHRP2). There have also been recent reports of pfhrp2 gene deletions being found in parasites collected from several African countries. The WHO has concluded that lacking the pfhrp2 gene must be monitored in Africa. The purpose of the study was to analyse why the samples that were positive by PCR were negative by RDTs and, therefore, to determine whether there have been deletions in the pfhrp2 and/or pfhrp3 genes. METHODS: Malaria NM-PCR was carried out on all the samples collected in the field. A group of 128 samples was positive by PCR but negative by RDT; these samples were classified as RDT false-negatives. PCR was carried out for exon2 of pfhrp2 and pfhrp3 genes to detect the presence or absence of these two genes. Frequencies with 95% confidence intervals (CIs) were used for prevalence estimates. Associations were assessed by the Chi square test or Fisher´s exact test. The level of significance was set at p ≤ 0.05. Statistical analyses were performed using the software package SPSSv.15.0. RESULTS: After PCR, 81 samples were identified (4.7%, 95% CI 3.8-5.8) which had deletion in both genes, pfhrp2 and pfhrp3. Overall, however, 11 samples (0.6%, 95% CI 0.36-1.14) had deletion only in pfhrp2 but not in pfhrp3, and 15 (0.9%, 95% CI 0.6-1.5) presented with deletion only in pfhrp3 but not in pfhrp2. Considering the pfhrp2 gene separately, within the total of 1724 samples, 92 (5.3%, 95% CI 4.37-6.5) had evidence of deletion. CONCLUSION: The present study provides the first evidence of deletion in the pfhrp2 and pfhrp3 genes in P. falciparum isolates from Equatorial Guinea. However, larger studies across different regions within the country and across different seasonal profiles are needed to determine the full extent of pfhrp2 and pfhrp3 deletion. It is strongly recommended to implement an active surveillance programme in order to detect any increases in pfhrp2 and pfhrp3 deletion frequencies.

Malaria outbreak in Riaba district, Bioko Island: lessons learned.

At the beginning of 2019, a sudden surge of malaria cases was observed in the district of Riaba, Bioko Island. Between January and April, confirmed malaria cases increased 3.8-fold compared to the same period in 2018. Concurrently, anopheline human biting rate (HBR) increased 2.1-fold. During the outbreak, 82.2% of the district population was tested for malaria with a rapid diagnostic test; 37.2% of those tested had a detectable infection and were treated according to national guidelines. Vector control interventions, including indoor residual spraying and larval source management were scaled-up. After the interventions, the number of confirmed cases decreased by 70% and the overall parasite prevalence in the communities by 43.8%. Observed prevalence in a follow up malaria indicator survey, however, was significantly higher than elsewhere on the island, and higher than in previous years. There was no significant reduction in HBR, which remained high for the rest of the year. The surge was attributed to various factors, chiefly increased rainfall and a large number of anthropogenic anopheline breeding sites created by construction works. This case study highlights the need for sustained vector control interventions and multi-sector participation, particularly in malaria control and elimination settings with persistently high local malaria receptivity.

HDV infection in immigrant populations.

AIMS: Little data have been published so far on the epidemiological aspects of hepatitis D virus (HDV) infection in immigrant populations and even poorer is the information on the virological, phylogenetic, and clinical aspects of this infection in these populations. This review article, aimed primarily at physicians caring for immigrants, summarizes the information available on HDV infection and analyzes data on this topic concerning the immigrant populations. METHODS AND RESULTS: The prevalence of HDV infection in HBsAg-positive immigrants varies according to the country of origin. For example, in immigrants from sub-Saharan Africa, this prevalence is higher in those born in Equatorial Guinea (24.4%) than those from other African countries (10.3%). The epidemiological impact of HDV infection linked to migratory flows is a function of the different endemicity between countries of origin and countries in which a new existence has been established. This impact is high when immigrants from areas endemic to HDV infection (eg, Equatorial Guinea) settle in areas of low endemicity (eg, Germany or England, with a prevalence of around 4%), while the impact is lesser or nonexistent if the migratory flows are directed toward countries with intermediate endemicity (eg, Italy and Greece, with a prevalence of around 10%). CONCLUSION: This impact of immigration on HDV epidemiology can be strong when HDV endemicity is high in the country of origin and low in the host country and slight when immigrants move to high or medium endemic countries.

Characterising malaria connectivity using malaria indicator survey data.

Malaria connectivity describes the flow of parasites among transmission sources and sinks within a given landscape. Because of the spatial and temporal scales at which parasites are transported by their hosts, malaria sub-populations are largely defined by mosquito movement and malaria connectivity among them is largely driven by human movement. Characterising malaria connectivity thus requires characterising human travel between areas with differing levels of exposure to malaria. Whilst understanding malaria connectivity is fundamental for optimising interventions, particularly in areas seeking or sustaining elimination, there is a dearth of human movement data required to achieve this goal. Malaria indicator surveys (MIS) are a generally under utilised but potentially rich source of travel data that provide a unique opportunity to study simple associations between malaria infection and human travel in large population samples. This paper shares the experience working with MIS data from Bioko Island that revealed programmatically useful information regarding malaria importation through human travel. Simple additions to MIS questionnaires greatly augmented the level of detail of the travel data, which can be used to characterise human travel patterns and malaria connectivity to assist targeting interventions. It is argued that MIS potentially represent very important and timely sources of travel data that need to be further exploited.

Molecular monitoring of the diversity of human pathogenic malaria species in blood donations on Bioko Island, Equatorial Guinea.

BACKGROUND: Malaria can be transmitted by blood transfusion from human to human and it is responsible for the majority of transfusion-transmitted infectious diseases worldwide. In sub-Saharan Africa, it had been estimated that almost a quarter of blood donations contain malaria parasites. Since rapid diagnostic tests and thick blood smear microscopy lack sensitivity for low density parasitaemia, particularly in asymptomatic adults, the most reliable method to assess the problem of transfusion-transmitted malaria are nucleic acid-based molecular approaches such as quantitative polymerase chain reaction. The study was undertaken to determine the prevalence of sub-microscopic malaria parasite infection among blood donors in Malabo, Equatorial Guinea. METHODS: Between July and August 2017, a total of 200 individual blood samples from blood donors at the Malabo Blood Bank were collected and screened by rapid diagnostic tests and thick blood smear microscopy. Retrospectively, the same samples were analysed for the presence of undetected, low-density malaria parasites using quantitative polymerase chain reaction. RESULTS: In comparison to 6.5% (13/200) by rapid diagnostic test and 2.0% (4/200) by microscopy, the proportion of Plasmodium falciparum positive blood donations analysed by quantitative polymerase chain reaction was significantly higher (26%, 52/200). Densities of P. falciparum positive blood donations were ranging from 0.06 to 3707.0 parasites/µL with 79.6% below 100 parasites/µL and therefore not detectable by non-molecular malaria diagnostic tests. qPCR based species identification revealed that P. falciparum was the dominating species responsible for 88.1% (52/59) of positive blood donations, followed by Plasmodium malariae (15.3%, 9/59) and Plasmodium ovale (3.4%, 2/59). CONCLUSIONS: This study confirms that in malaria endemic settings, sub-patent malaria infections among blood donors are prevalent. In blood collected from healthy donors living in Malabo, P. falciparum, P. malariae and P. ovale parasites were identified. Currently widely used malaria diagnostic tools have missed more than 75% of P. falciparum containing blood donations, demonstrating the value of quantitative polymerase chain reaction to reliably detect low density P. falciparum infections. Since the availability of molecular diagnostic methods in malaria endemic countries is still limited, the blood recipients living in malaria endemic countries should be treated following WHO recommendations.

Malaria determining risk factors at the household level in two rural villages of mainland Equatorial Guinea.

BACKGROUND: After the introduction of an artemisinin-based combination therapy, the reduction of prevalence of malaria infections has shown a remarkable progress during the last decade. However due to the lack of a consistent malaria control programme and socioeconomic inequalities, Plasmodium infection is still one of the major cause of disease in Equatorial Guinea, namely in the rural communities. This study explored the associated risk factors of malaria transmission at the microeconomic level (households) in two rural villages of mainland Equatorial Guinea. METHODS: This survey involved 232 individuals living in 69 households located in two rural villages, Ngonamanga and Miyobo, of coastal and interior of Equatorial Guinea, respectively. Malaria prevalence was measured by PCR and parasitaemia level by optical microscopy; household socioeconomic status (SES) was measured based on house characteristics using a 2-step cluster analysis. Logistic regression analysis was performed to investigate the relationship of a diverse set of independent variables on being diagnosed with malaria and on showing high levels of parasitaemia. RESULTS: The prevalence of Plasmodium spp. infection was 69%, with 80% of households having at least one parasitaemic member. The majority of houses have eaves (80%), walls of clay/wood (90%) and zinc roof (99%) and only 10% of them have basic sanitation facilities. The studied areas showed reduced rates of indoor residual spraying coverage (9%), and long-lasting insecticide-treated net ownership (35%), with none of these preventive tools showing any significant effects on malaria risk in these areas. Neither the risk of malaria infection (PCR positive result) or the development of high parasitaemia did show association with SES. CONCLUSIONS: This study has contributed to reinforce the importance of living conditions associated to a high risk of malaria infection and vulnerability to develop high parasitaemia. This study also contributes to future malaria control interventions to be implemented in mainland Equatorial Guinea or in other countries with similar environmental conditions.

Genetic diversity and allele frequencies of Plasmodium falciparum msp1 and msp2 in parasite isolates from Bioko Island, Equatorial Guinea.

BACKGROUND: Malaria is still a serious public health problem on Bioko Island (Equatorial Guinea), although the number of annual cases has been greatly reduced since 2004 through the Bioko Island Malaria Control Project (BIMCP). A better understanding of malaria parasite population diversity and transmission dynamics is critical for assessing the effectiveness of malaria control measures. The objective of this study is to investigate the genetic diversity of Plasmodium falciparum populations and multiplicity of infection (MOI) on Bioko Island 7 years after BIMCP. METHODS: A total of 181 patients with uncomplicated P. falciparum malaria diagnosed with microscopy were collected from Bioko Island from January 2011 to December 2014. Parasite DNA was extracted using chelex-100 and species were identified using a real-time PCR followed by high-resolution melting. Plasmodium falciparum msp1 and msp2 allelic families were determined using nested PCR. RESULTS: Three msp1 alleles (K1, MAD20, and RO33) and two msp2 alleles (FC27 and 3D7) were analysed in all samples. In msp1, the MAD20 allelic family was predominant with 96.69% (175/178) followed respectively by the K1 allelic family with 96.07% (171/178) and R033 allelic family with 70.78% (126/178). In msp2, the FC27 allelic family was the most frequently detected with 97.69% (169/173) compared to 3D7 with 72.25% (125/173). Twenty-six different alleles were observed in msp1 with 9 alleles for K1, 9 alleles for MAD20 and 8 alleles for R033. In msp2, 25 individual alleles were detected with 5 alleles for FC27 and 20 alleles for 3D7. The overall MOI was 5.51 with respectively 3.5 and 2.01 for msp1 and msp2. A significant increase in overall MOI was correlated with the age group of the patients (P = 0.026) or parasite densities (P = 0.04). CONCLUSIONS: The present data showed high genetic diversity and MOI values among the P. falciparum population in the study, reflecting both the high endemic level and malaria transmission on Bioko Island. These data provide valuable information for surveillance of P. falciparum infection and for assessing the appropriateness of the current malarial control strategies in the endemic area.

Trends in parasite prevalence following 13 years of malaria interventions on Bioko island, Equatorial Guinea: 2004-2016.

BACKGROUND: Whilst there have been substantial reductions in malaria transmission over the past decade, in many countries in West and Central Africa the malaria burden remains high. Monitoring and evaluation of malaria transmission trends and intervention strategies are key elements for malaria control programmes. This study uses a time series of annual malaria indicator surveys to track the progress of malaria control in Bioko Island, Equatorial Guinea, over a 13 year period of intensive interventions. Malaria infection and haemoglobin were measured annually in children (1 to 14 years) in cross-sectional household surveys from 2004 to 2016 in 18 sentinel sites across the island. Trends in transmission patterns were assessed and the impact of the vector control interventions (net use and spray coverage) was evaluated. RESULTS: Between 2004 and 2016 approximately 106,500 individual tests for parasitaemia were conducted using rapid diagnostic tests. Although spray coverage remained relatively high (> 70%) over the time period, reported net usage was generally below 40%. Parasite prevalence reduced from 43.3 to 10.5% between 2004 and 2016. The prevalence of moderate to severe anaemia in children aged 1-5 years reduced from 14.9 to 1.6%. Impact in individual sites ranged from 57 to 100% reductions in parasite prevalence between 2004 and 2016. Sleeping under a net and living in a house that had been sprayed in the past 12 months were independently protective against infection (OR = 0.69 [95%CI 0.61-0.80] and OR = 0.87 [95% CI 0.78-0.97], respectively), whilst recent travel to the mainland increased the odds of infection nearly fourfold (OR = 3.94 [95%CI 2.79-5.56]). CONCLUSION: Island-wide interventions have resulted in a substantial reduction in malaria transmission on Bioko Island. This unique time series of 13 consecutive annual malaria indicator surveys clearly demonstrates the long-term effectiveness of the sustained use of two vector control interventions, indoor residual spraying and LLINs, and the value of comprehensive and sustained surveillance. Despite considerable success in reducing the burden on the island, malaria is still endemic, with populations in some areas remaining at high risk of infection.

Spatial clustering and risk factors of malaria infections in Bata district, Equatorial Guinea.

BACKGROUND: The transmission of malaria is intense in the majority of the countries of sub-Saharan Africa, particularly in those that are located along the Equatorial strip. The present study aimed to describe the current distribution of malaria prevalence among children and its environment-related factors as well as to detect malaria spatial clusters in the district of Bata, in Equatorial Guinea. METHODS: From June to August 2013 a representative cross-sectional survey using a multistage, stratified, cluster-selected sample was carried out of children in urban and rural areas of Bata District. All children were tested for malaria using rapid diagnostic tests (RDTs). Results were linked to each household by global position system data. Two cluster analysis methods were used: hot spot analysis using the Getis-Ord Gi statistic, and the SaTScan™ spatial statistic estimates, based on the assumption of a Poisson distribution to detect spatial clusters. In addition, univariate associations and Poisson regression model were used to explore the association between malaria prevalence at household level with different environmental factors. RESULTS: A total of 1416 children aged 2 months to 15 years living in 417 households were included in this study. Malaria prevalence by RDTs was 47.53%, being highest in the age group 6-15 years (63.24%, p < 0.001). Those children living in rural areas were there malaria risk is greater (65.81%) (p < 0.001). Malaria prevalence was higher in those houses located <1 km from a river and <3 km to a forest (IRR: 1.31; 95% CI 1.13-1.51 and IRR: 1.44; 95% CI 1.25-1.66, respectively). Poisson regression analysis also showed a decrease in malaria prevalence with altitude (IRR: 0.73; 95% CI 0.62-0.86). A significant cluster inland of the district, in rural areas has been found. CONCLUSIONS: This study reveals a high prevalence of RDT-based malaria among children in Bata district. Those households situated in inland rural areas, near to a river, a green area and/or at low altitude were a risk factor for malaria. Spatial tools can help policy makers to promote new recommendations for malaria control.

Seroprevalence of antibodies against bacterial pathogens in sheep from Equatorial Guinea.

Equatorial Guinea (EG) is a country in Central Africa with typical tropical weather. Sheep are an important source of food in EG, but the absence of information regarding infectious diseases that affect the native flocks of small ruminants is a concern. The country is currently implementing several new practices in the sheep industry associated with the importation of sheep from other countries. This study aimed to evaluate the seroprevalence of bacterial infections that are important to the sheep industry in EG sheep flocks. The detection of specific antibodies for the four agents studied was performed using enzyme-linked immunosorbent assay in 1,002 serum samples from EG sheep. The results showed a true prevalence of 13.37% for antibodies against Corynebacterium pseudotuberculosis, 0.59% for Brucella ovis, 19.89% for Chlamydophila abortus and 0.79% for Mycoplasma agalactiae in animals from production flocks. Among a group of 35 samples from isolated native animals, 47.56% were seropositive for antibodies against C. pseudotuberculosis, 42.84% for B. ovis, 54.28% for C. abortus and 11.35% for M. agalactiae. These results comprise the first report of the prevalence of infectious diseases in sheep in EG. They highlight the importance of adopting adequate measures to prevent infection by bacteria endemic to EG native flocks during the development of the sheep industry in the country.

La Guinée équatoriale est un pays d'Afrique centrale au climat typiquement tropical. Les moutons constituent une source importante d'aliments pour le pays, ce qui rend particulièrement alarmante l'absence d'informations concernant les maladies infectieuses affectant le cheptel autochtone de petits ruminants. Le pays met actuellement en place de nouvelles pratiques d'élevage ovin, en lien avec l'importation de moutons d'autres pays. La présente étude avait pour objet d'évaluer la prévalence sérologique de quelques infections bactériennes majeures pour le secteur de l'élevage dans le cheptel ovin de la Guinée équatoriale. Pour les besoins de l'étude, 1 002 échantillons de sérum prélevés de moutons autochtones ont été soumis à une épreuve immuno-enzymatique pour la détection d'anticorps spécifiques dirigés contre quatre agents bactériens. La prévalence réelle de moutons issus de troupeaux d'élevage possédant des anticorps vis-à-vis de chacune des bactéries recherchées s'est élevée à 13,37 % pour Corynebacterium pseudotuberculosis, à 0,59 % pour Brucella ovis, à 19,89 % pour Chlamydophila abortus et à 0,79 % pour Mycoplasma agalactiae. Dans un groupe de 35 prélèvements issus d'animaux isolés autochtones, le taux d'échantillons positifs était de 47,56 % pour C. pseudotuberculosis, de 42,84 % pour B. ovis, de 54,28 % pour C. abortus et de 11,35 % pour M. agalactiae. Ces résultats constituent le premier rapport sur la prévalence des maladies infectieuses chez le mouton en Guinée équatoriale. Ils soulignent l'importance d'adopter des mesures appropriées afin de prévenir les infections dues à des bactéries endémiques dans le cheptel autochtone de la Guinée équatoriale, à un moment où ce pays cherche à développer l'élevage ovin.

Guinea Ecuatorial es un país centroafricano con un clima tropical típico, en el que las ovejas constituyen una importante fuente de alimento. Por ello es preocupante la ausencia de información sobre las enfermedades infecciosas que afectan a los rebaños autóctonos de pequeños rumiantes. El país está implantando ahora una serie de nuevos procedimientos en el sector ovino, vinculados a la importación de ovejas de otros países. Los autores describen un estudio encaminado a evaluar la seroprevalencia en los rebaños de ovejas del país de infecciones bacterianas de importancia para el sector. Para detectar anticuerpos específicos contra los cuatro agentes estudiados se aplicó la técnica del ensayo inmunoenzimático a 1 002 muestras de suero de ovejas autóctonas. Los resultados pusieron de manifiesto una prevalencia real de anticuerpos contra Corynebacterium pseudotuberculosis del 13,37%, del 0,59% contra Brucella ovis, del 19,89% contra Chlamydophila abortus y del 0,79% contra Mycoplasma agalactiae en ejemplares pertenecientes a rebaños productivos. De un grupo de 35 muestras procedentes de animales autóctonos aislados, un 47,56% resultaron seropositivas para los anticuerpos contra C. pseudotuberculosis, un 42,84% en el caso de B. ovis, un 54,28% en el de C. abortus y un 11,35% en el de M. agalactiae. Estos resultados, que constituyen el primer informe sobre la prevalencia de enfermedades infecciosas en las ovejas de Guinea Ecuatorial, demuestran la importancia de instituir medidas adecuadas para prevenir la infección por bacterias endémicas de los rebaños autóctonos de Guinea Ecuatorial en un momento en que la industria ovina del país está en pleno crecimiento.