Two cases of long-lasting, sub-microscopic Plasmodium malariae infections in adults from coastal Tanzania.

BACKGROUND: Malaria is endemic in Tanzania with majority of clinical cases caused by Plasmodium falciparum. Additionally, Plasmodium malariae and Plasmodium ovale spp. are also present and clinical manifestations caused by these infections are not well described. Clinical episodes caused by P. malariae infections are often characterized by a relatively mild illness with a low number of parasites, which can persist for long periods. In this report, two cases of P. malariae infections that were identified during a clinical trial evaluating the P. falciparum malaria vaccine candidate, PfSPZ Vaccine are described. The two participants were followed up and monitored for clinical and laboratory parameters to assess vaccine safety providing the opportunity to study clinical manifestations of P. malariae over 4 months. CASE PRESENTATION: Two young, healthy Tanzanian men infected with low density asexual blood stage P. malariae diagnosed by quantitative polymerase chain reaction (qPCR) are described. Retrospective analysis of collected and stored blood samples revealed that the two volunteers had constant asexual blood stage parasitaemia for more than 4 months. During the 132 days of infection, the volunteers' vital signs, body temperature and serum biochemistry all remained within normal ranges. Haematological abnormalities, which were transiently outside normal ranges, were regarded as not clinically significant. During this time period, four consecutive evaluations of blood samples by thick blood smear microscopy conducted by an experienced microscopist were all negative, indicating the presence of low-density sub-microscopic infections. CONCLUSIONS: The two cases of P. malariae infections presented here confirm the ability of this Plasmodium species to persist at low density in the human host for extended time periods without causing clinical symptoms. The presented data also demonstrate that clinical study sites in malaria endemic regions need to have a strong malaria diagnostic infrastructure, including the ability of capturing sub-microscopic parasitaemia and differentiation of Plasmodium species. Trial registration ClinicalTrials.gov: NCT02613520, https://clinicaltrials.gov/ct2/show/NCT02613520 , Registered: November 24th 2015, Enrolment of the first participant to the trial: December 15th 2015, Trial was registered before the first participant was enrolled.

Pilot deployment of a community health care worker in distributing and offering the COVID-19 AgRDT in Tanzania.

A pilot implementation of the rapid diagnostic test program was performed to collect evidence of the feasibility, acceptability, and uptake of the COVID-19 AgRDT in Tanzania. We conducted a prospective cross-sectional study in the community to provide quantitative details of the pilot implementation of the antigen rapid diagnostic test (AgRDT) in Tanzania. This study was undertaken between March 2022 and September 2022. The pilot was implemented by distributing and offering test kits to people suspected of having COVID-19 in Dar es Salaam through community health workers. A total of 1039 participants consented to participate in the survey. All the participants reported having heard about the disease. The radio was the main source (93.2%) of information on COVID-19. With regard to prevention measures, approximately 930 (89.5%) of the respondents thought that COVID-19 could be prevented. Approximately 1035 (99.6%) participants reported that they were willing to have a COVID-19 AgRDT test and wait for 20 min for the results. With regard to the participants' opinions on the AgRDT device, the majority 907 (87.3%) felt comfortable with the test, and 1,029 (99.0%) were very likely to recommend the AgRDT test to their friends. The majority of participants 848 (83.1%) mentioned that they would be willing to pay for the test if it was not available for free. The results suggest overall good acceptance of the COVID-19 AgRDT test. It is evident that the use of trained community healthcare workers allows easy screening of all possible suspects and helps them receive early treatment.

Blood-stage malaria vaccine candidate RH5.1/Matrix-M in healthy Tanzanian adults and children; an open-label, non-randomised, first-in-human, single-centre, phase 1b trial.

BACKGROUND: A blood-stage Plasmodium falciparum malaria vaccine would provide a second line of defence to complement partially effective or waning immunity conferred by the approved pre-erythrocytic vaccines. RH5.1 is a soluble protein vaccine candidate for blood-stage P falciparum, formulated with Matrix-M adjuvant to assess safety and immunogenicity in a malaria-endemic adult and paediatric population for the first time. METHODS: We did a non-randomised, phase 1b, single-centre, dose-escalation, age de-escalation, first-in-human trial of RH5.1/Matrix-M in Bagamoyo, Tanzania. We recruited healthy adults (aged 18-45 years) and children (aged 5-17 months) to receive the RH5.1/Matrix-M vaccine candidate in the following three-dose regimens: 10 µg RH5.1 at 0, 1, and 2 months (Adults 10M), and the higher dose of 50 µg RH5.1 at 0 and 1 month and 10 µg RH5.1 at 6 months (delayed-fractional third dose regimen; Adults DFx). Children received either 10 µg RH5.1 at 0, 1, and 2 months (Children 10M) or 10 µg RH5.1 at 0, 1, and 6 months (delayed third dose regimen; Children 10D), and were recruited in parallel, followed by children who received the dose-escalation regimen (Children DFx) and children with higher malaria pre-exposure who also received the dose-escalation regimen (High Children DFx). All RH5.1 doses were formulated with 50 µg Matrix-M adjuvant. Primary outcomes for vaccine safety were solicited and unsolicited adverse events after each vaccination, along with any serious adverse events during the study period. The secondary outcome measures for immunogenicity were the concentration and avidity of anti-RH5.1 serum IgG antibodies and their percentage growth inhibition activity (GIA) in vitro, as well as cellular immunogenicity to RH5.1. All participants receiving at least one dose of vaccine were included in the primary analyses. This trial is registered at ClinicalTrials.gov, NCT04318002, and is now complete. FINDINGS: Between Jan 25, 2021, and April 15, 2021, we recruited 12 adults (six [50%] in the Adults 10M group and six [50%] in the Adults DFx group) and 48 children (12 each in the Children 10M, Children 10D, Children DFx, and High Children DFx groups). 57 (95%) of 60 participants completed the vaccination series and 55 (92%) completed 22 months of follow-up following the third vaccination. Vaccinations were well-tolerated across both age groups. There were five serious adverse events involving four child participants during the trial, none of which were deemed related to vaccination. RH5-specific T cell and serum IgG antibody responses were induced by vaccination and purified total IgG showed in vitro GIA against P falciparum. We found similar functional quality (ie, GIA per µg RH5-specific IgG) across all age groups and dosing regimens at 14 days after the final vaccination; the concentration of RH5.1-specific polyclonal IgG required to give 50% GIA was 14·3 µg/mL (95% CI 13·4-15·2). 11 children were vaccinated with the delayed third dose regimen and showed the highest median anti-RH5 serum IgG concentration 14 days following the third vaccination (723 µg/mL [IQR 511-1000]), resulting in all 11 who received the full series showing greater than 60% GIA following dilution of total IgG to 2·5 mg/mL (median 88% [IQR 81-94]). INTERPRETATION: The RH5.1/Matrix-M vaccine candidate shows an acceptable safety and reactogenicity profile in both adults and 5-17-month-old children residing in a malaria-endemic area, with all children in the delayed third dose regimen reaching a level of GIA previously associated with protective outcome against blood-stage P falciparum challenge in non-human primates. These data support onward efficacy assessment of this vaccine candidate against clinical malaria in young African children. FUNDING: The European and Developing Countries Clinical Trials Partnership; the UK Medical Research Council; the UK Department for International Development; the National Institute for Health and Care Research Oxford Biomedical Research Centre; the Division of Intramural Research, National Institute of Allergy and Infectious Diseases; the US Agency for International Development; and the Wellcome Trust.

Field evaluation of the novel One Step Malaria Pf and Pf/Pv rapid diagnostic tests and the proportion of HRP-2 gene deletion identified on samples collected in the Pwani region, Tanzania.

Background: Malaria rapid diagnostic tests (mRDTs) have played an important role in the early detection of clinical malaria in an endemic area. While several mRDTs are currently on the market, the availability of mRDTs with high sensitivity and specificity will merit the fight against malaria. We evaluated the field performance of a novel One Step Malaria (P.f/P.v) Tri-line and One Step Malaria (P.f) rapid test kits in Pwani, Tanzania. Methods: In a cross-sectional study conducted in Bagamoyo and Kibiti districts in Tanzania, symptomatic patients were tested using the SD BIOLINE, One Step Malaria (P.f/P.v) Tri-line and One Step Malaria (P.f) rapid test kits, microscope, and quantitative Polymerase Chain Reaction (qPCR). An additional qPCR assay was carried out to detect Histidine-Rich Protein 2 (HRP-2) gene deletion on mRDT negative but microscope and qPCR positive samples. Microscope results confirmed by qPCR were used for analysis, where qPCR was used as a reference method. Results: The sensitivity and specificity of One Step P.f/P.v Tri-line mRDTs were 96.0% (CI 93.5-97.7%) and 98.3% (CI 96.8-99.2%), respectively. One Step P.f mRDT had sensitivity and specificity of 95.2% (CI 92.5-97.1%) and 97.9% (CI 96.3-99.0%) respectively. Positive predictive value (PPV) was 97.6% (CI 95.4-98.7%) and negative predictive value (NPV) was 96.2% (CI 95.5-98.3%) for the One Step P.f/P.v Tri-line mRDTs respectively, while One Step P.f mRDT had positive predictive value (PPV) and negative predictive value (NPV) of 97.0% (CI 94.8-98.3%) and 96.7 (CI 94.9-97.9%) respectively. 9.8% (CI 7.84-11.76) of all samples tested and reported to be malaria-negative by mRDT had HRP-2 gene deletion. Conclusion: One Step Malaria P.f/P.v Tri-line and One Step Malaria P.f rapid test kits have similar sensitivity and specificity as the standard mRDT that is currently in the market, demonstrating the potential to contribute in the fight against malaria in endemic settings. However, the identified malaria parasites population with HRP-2 gene deletion pose a threat to the current mRDT usability in the field and warrants further investigations.

Superior antibody immunogenicity of a viral-vectored RH5 blood-stage malaria vaccine in Tanzanian infants as compared to adults.

BACKGROUND: RH5 is a leading blood-stage candidate antigen for a Plasmodium falciparum vaccine; however, its safety and immunogenicity in malaria-endemic populations are unknown. METHODS: A phase 1b, single-center, dose-escalation, age-de-escalation, double-blind, randomized, controlled trial was conducted in Bagamoyo, Tanzania (NCT03435874). Between 12th April and 25th October 2018, 63 healthy adults (18-35 years), young children (1-6 years), and infants (6-11 months) received a priming dose of viral-vectored ChAd63 RH5 or rabies control vaccine. Sixty participants were boosted with modified vaccinia virus Ankara (MVA) RH5 or rabies control vaccine 8 weeks later and completed 6 months of follow-up post priming. Primary outcomes were the number of solicited and unsolicited adverse events post vaccination and the number of serious adverse events over the study period. Secondary outcomes included measures of the anti-RH5 immune response. FINDINGS: Vaccinations were well tolerated, with profiles comparable across groups. No serious adverse events were reported. Vaccination induced RH5-specific cellular and humoral responses. Higher anti-RH5 serum immunoglobulin G (IgG) responses were observed post boost in young children and infants compared to adults. Vaccine-induced antibodies showed growth inhibition activity (GIA) in vitro against P. falciparum blood-stage parasites; their highest levels were observed in infants. CONCLUSIONS: The ChAd63-MVA RH5 vaccine shows acceptable safety and reactogenicity and encouraging immunogenicity in children and infants residing in a malaria-endemic area. The levels of functional GIA observed in RH5-vaccinated infants are the highest reported to date following human vaccination. These data support onward clinical development of RH5-based blood-stage vaccines to protect against clinical malaria in young African infants. FUNDING: Medical Research Council, London, UK.

Understanding the role of serological and clinical data on assessing the dynamic of malaria transmission: a case study of Bagamoyo district, Tanzania.

Introduction: naturally acquired blood-stage malaria antibodies and malaria clinical data have been reported to be useful in monitoring malaria change over time and as a marker of malaria exposure. This study assessed the total immunoglobulin G (IgG) levels to Plasmodium falciparum schizont among infants (5-17 months), estimated malaria incidence using routine health facility-based surveillance data and predicted trend relation between anti-schizont antibodies and malaria incidence in Bagamoyo. Methods: 252 serum samples were used for assessment of total IgG by enzyme-linked immunosorbent assay and results were expressed in arbitrary units (AU). 147/252 samples were collected in 2021 during a blood-stage malaria vaccine trial [ClinicalTrials.gov NCT04318002], and 105/252 were archived samples of malaria vaccine trial conducted in 2012 [ClinicalTrials.gov NCT00866619]. Malaria incidence was calculated from outpatient clinic data of malaria rapid test or blood smear positive results retrieved from District-Health-Information-Software-2 (DHIS2) between 2013 and 2020. Cross-sectional data from both studies were analysed using STATA version 14. Results: this study demonstrated a decline in total anti-schizont IgG levels from 490.21AU in 2012 to 97.07AU in 2021 which was related to a fall in incidence from 58.25 cases/1000 person-year in 2013 to 14.28 cases/1000 person-year in 2020. We also observed a significant difference in incidence when comparing high and low malaria transmission areas and by gender. However, we did not observe differences when comparing total anti-schizont antibodies by gender and study year. Conclusion: total anti-schizont antibody levels appear to be an important serological marker of exposure for assessing the dynamic of malaria transmission in infants living in malaria-endemic regions.

Using haematophagous fly blood meals to study the diversity of blood-borne pathogens infecting wild mammals.

Many emerging infectious diseases originate from wild animals, so there is a profound need for surveillance and monitoring of their pathogens. However, the practical difficulty of sample acquisition from wild animals tends to limit the feasibility and effectiveness of such surveys. Xenosurveillance, using blood-feeding invertebrates to obtain tissue samples from wild animals and then detect their pathogens, is a promising method to do so. Here, we describe the use of tsetse fly blood meals to determine (directly through molecular diagnostic and indirectly through serology), the diversity of circulating blood-borne pathogens (including bacteria, viruses and protozoa) in a natural mammalian community of Tanzania. Molecular analyses of captured tsetse flies (182 pools of flies totalizing 1728 flies) revealed that the blood meals obtained came from 18 different vertebrate species including 16 non-human mammals, representing approximately 25% of the large mammal species present in the study area. Molecular diagnostic demonstrated the presence of different protozoa parasites and bacteria of medical and/or veterinary interest. None of the six virus species searched for by molecular methods were detected but an ELISA test detected antibodies against African swine fever virus among warthogs, indicating that the virus had been circulating in the area. Sampling of blood-feeding insects represents an efficient and practical approach to tracking a diversity of pathogens from multiple mammalian species, directly through molecular diagnostic or indirectly through serology, which could readily expand and enhance our understanding of the ecology and evolution of infectious agents and their interactions with their hosts in wild animal communities.

Efficacy and safety of intravenous ferric carboxymaltose compared with oral iron for the treatment of iron deficiency anaemia in women after childbirth in Tanzania: a parallel-group, open-label, randomised controlled phase 3 trial.

BACKGROUND: Iron deficiency anaemia is of major concern in low-income settings, especially for women of childbearing age. Oral iron substitution efficacy is limited by poor compliance and iron depletion severity. We aimed to assess the efficacy and safety of intravenous ferric carboxymaltose versus oral iron substitution following childbirth in women with iron deficiency anaemia in Tanzania. METHODS: This parallel-group, open-label, randomised controlled phase 3 trial was done at Bagamoyo District Hospital and Mwananyamala Hospital, Tanzania. Eligible participants were close to delivery and had iron deficiency anaemia defined as a haemoglobin concentration of less than 110 g/L and a ferritin concentration of less than 50 µg/L measured within 14 days before childbirth. Participants were randomly assigned 1:1 to receive intravenous ferric carboxymaltose or oral iron, stratified by haemoglobin concentration and site. Intravenous ferric carboxymaltose was administered at a dose determined by the haemoglobin concentration and bodyweight (bodyweight 35 kg to <70 kg and haemoglobin ≥100 g/L: 1000 mg in one dose; bodyweight 35 kg to <70 kg and haemoglobin <100 g/L, or bodyweight ≥70 kg and haemoglobin ≥100 g/L: 1500 mg in two doses at least 7 days apart; bodyweight ≥70 kg and haemoglobin <100 g/L: 2000 mg in two doses at least 7 days apart). Oral iron treatment consisted of three dried ferrous sulphate tablets of 200 mg containing 60 mg of elementary iron and 5 mg of folic acid every morning. Oral treatment was to be taken for 3 months after haemoglobin normalisation. The primary outcome was haemoglobin normalisation (>115 g/L) at 6 weeks. Follow-up visits were at 6 weeks, and 3, 6, and 12 months. Analyses were done in the modified intention-to-treat population of participants who had a 6-week haemoglobin concentration result, using logistic and linear regression models for binary and continuous outcomes, adjusted for baseline haemoglobin concentration and site. This trial is registered with ClinicalTrials.gov, NCT02541708. FINDINGS: Between Oct 8, 2015, and March 14, 2017, 533 individuals were screened and 230 were enrolled and randomly assigned to a study group (114 to intravenous iron, 116 to oral iron). At 6 weeks, 94 (82%) participants in the intravenous iron group and 92 (79%) in the oral iron group were assessed for the primary outcome. 75 (80%) participants in the intravenous iron group and 47 (51%) in the oral iron group had normalised haemoglobin (odds ratio 4·65, 95% CI 2·33-9·27). There were two mild to moderate infusion-related adverse events; and five serious adverse events (three in the intravenous iron group, two in the oral iron group), unrelated to the study medication. INTERPRETATION: Intravenous iron substitution with ferric carboxymaltose was safe and yielded a better haemoglobin response than oral iron. To our knowledge, this is the first study to provide evidence of the benefits and safety of intravenous iron substitution in a low-income setting. FUNDING: Vifor Pharma, R Geigy-Stiftung, Freiwillige Akademische Gesellschaft, and Swiss Tropical and Public Health Institute.

The Clean pilot study: evaluation of an environmental hygiene intervention bundle in three Tanzanian hospitals.

BACKGROUND: Healthcare associated infections (HAI) are estimated to affect up to 15% of hospital inpatients in low-income countries (LICs). A critical but often neglected aspect of HAI prevention is basic environmental hygiene, particularly surface cleaning and linen management. TEACH CLEAN is an educational intervention aimed at improving environmental hygiene. We evaluated the effectiveness of this intervention in a pilot study in three high-volume maternity and newborn units in Dar es Salaam, Tanzania. METHODS: This study design prospectively evaluated the intervention as a whole, and offered a before-and-after comparison of the impact of the main training. We measured changes in microbiological cleanliness [Aerobic Colony Counts (ACC) and presence of Staphylococcus aureus] using dipslides, and physical cleaning action using gel dots. These were analysed with descriptive statistics and logistic regression models. We used qualitative (focus group discussions, in-depth interviews, and semi-structured observation) and quantitative (observation checklist) tools to measure why and how the intervention worked. We describe these findings across the themes of adaptation, fidelity, dose, reach and context. RESULTS: Microbiological cleanliness improved during the study period (ACC pre-training: 19%; post-training: 41%). The odds of cleanliness increased on average by 1.33 weekly during the pre-training period (CI = 1.11-1.60), and by 1.08 (CI = 1.03-1.13) during the post-training period. Cleaning action improved only in the pre-training period. Detection of S. aureus on hospital surfaces did not change substantially. The intervention was well received and considered feasible in this context. The major pitfalls in the implementation were the limited number of training sessions at the hospital level and the lack of supportive supervision. A systems barrier to implementation was lack of regular cleaning supplies. CONCLUSIONS: The evaluation suggests that improvements in microbiological cleanliness are possible using this intervention and can be sustained. Improved microbiological cleanliness is a key step on the pathway to infection prevention in hospitals. Future research should assess whether this bundle is cost-effective in reducing bacterial and viral transmission and infection using a rigorous study design.

ELIMU-MDx: a web-based, open-source platform for storage, management and analysis of diagnostic qPCR data.

The Electronic Laboratory Information and Management Utensil for Molecular Diagnostics (ELIMU-MDx) is a user-friendly platform designed and built to accelerate the turnaround time of diagnostic qPCR assays. ELIMU-MDx is compliant with Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines and has extensive data-import capabilities for all major qPCR instruments by using the RDML data standard. This platform was designed as an open-source software tool and can be accessed through the web browser on all major operating systems.