Characterization of Within-Host Plasmodium falciparum Diversity Using Next-Generation Sequence Data

Our understanding of the composition of multi-clonal malarial infections and the epidemiological factors which shape their diversity remain poorly understood. Traditionally within-host diversity has been defined in terms of the multiplicity of infection (MOI) derived by PCR-based genotyping. Massively parallel, single molecule sequencing technologies now enable individual read counts to be derived on genome-wide datasets facilitating the development of new statistical approaches to describe within-host diversity. In this class of measures the FWS metric characterizes within-host diversity and its relationship to population level diversity. Utilizing P. falciparum field isolates from patients in West Africa we here explore the relationship between the traditional MOI and FWS approaches. FWS statistics were derived from read count data at 86,158 SNPs in 64 samples sequenced on the Illumina GA platform. MOI estimates were derived by PCR at the msp-1 and -2 loci. Significant correlations were observed between the two measures, particularly with the msp-1 locus (P = 5.92×10−5). The FWS metric should be more robust than the PCR-based approach owing to reduced sensitivity to potential locus-specific artifacts. Furthermore the FWS metric captures information on a range of parameters which influence out-crossing risk including the number of clones (MOI), their relative proportions and genetic divergence. This approach should provide novel insights into the factors which correlate with, and shape within-host diversity

Experimental hut evaluation of DawaPlus 3.0 LN and DawaPlus 4.0 LN treated with deltamethrin and PBO against free-flying populations of Anopheles gambiae s.l. in Vallée du Kou, Burkina Faso.

BackgroundIn view of widespread pyrethroid resistance in malaria vectors in Africa, two long-lasting insecticidal nets (LLINs) incorporated with a synergist, piperonyl butoxide (PBO), DawaPlus 3.0 (deltamethrin + PBO in the roof panel; deltamethrin alone in the side panels) and DawaPlus 4.0 (deltamethrin + PBO in all panels), were evaluated in an experimental hut trial in a rice growing irrigated area in Burkina Faso. Efficacy of nets was tested against free-flying malaria vector, Anopheles gambiae s.l., with high pyrethroid resistance involving L1014F kdr and CYP6P3P450 resistance mechanisms.MethodologyThe efficacy of unwashed and 20-times washed DawaPlus 3.0 (polyethylene roof panel with 120 mg/m2 deltamethrin and 440 mg/m2 PBO; polyester side panels with deltamethrin 100 mg/m2) and DawaPlus 4.0 (same composition as roof of DawaPlus 3.0) was evaluated against DawaPlus 2.0 (80 mg/m2 deltamethrin; positive control). Volunteer sleepers and treatments were rotated in huts using a Latin square design on 63 consecutive nights during August-October 2016. Mortality, human blood-feeding inhibition, deterrence and exit rates of An. gambiae s.l. were monitored.Principal findingsSignificantly higher rates of mortality and blood-feeding inhibition were observed with unwashed DawaPlus 4.0 (36%; 47.5%) than unwashed DawaPlus 3.0 (11.8%; 33.3%), DawaPlus 2.0 (4.3%; 6.4%) or untreated net (P ConclusionsThe PBO-containing DawaPlus 4.0 significantly protected against An. gambiae s.l. in the study area. Unwashed DawaPlus 3.0 gave low to moderate protection against the positive control. PBO inhibits oxidase action; hence in areas with active malaria transmission having oxidase mechanisms, PBO nets could confer additional personal protection

Programmatic Implementation of Contact Investigation in Eight African Countries

The objective was to implement CI under national tuberculosis programmatic conditions and to advocate for its scaling up. Contact investigation was implemented in 150 Basic Management Units identified across eight countries. The target populations (children <5 years and persons living with HIV (PLHIV)) were evaluated during home and clinic visits using standardized tools, clinical examinations and, according to each country, additional tests. Contacts with active TB received TB treatment and those eligible received TB preventive therapy (TPT). Data were collected each quarter using standardized forms. Meetings were organized with partners to share preliminary results and advocate for scaling up. From October 2020 to December 2021, 9049 home visits were performed. The proportions of children <5 years and PLHIV who were screened and diagnosed with active TB were, respectively, 2.6% and 10.1%. Ninety-three percent of children <5 years and 98% of PLHIV living at home received TPT or TB treatment, respectively. The scale-up for contact investigation partially or at national level in 2022 was effective in six of the eight countries included in the project. These results indicate that CI is feasible under programmatic conditions within the National TB Programs of African countries

Does mosquito mass-rearing produce an inferior mosquito?

Abstract Background The success of the sterile insect technique depends, among other things, on continuous releases of sexually competitive sterile males within the target area. Several factors (including high rearing density and physical manipulation, such as larvae and pupae separation) can influence the quality of males produced in mass-rearing facilities. The different steps in mass production in the laboratory may modify the behaviour of mosquitoes, directly or through loss of natural characters as a result of adaptation to lab rearing, and lead to the competitiveness of sterile male being reduced. In the present study, the objective was to evaluate the effect of mass-rearing conditions on sterile male sexual competitiveness in semi-field cages compared to routine small scale laboratory rearing methods. Methods Anopheles arabiensis immature stages were reared both on a large scale using a rack and tray system developed by the FAO/IAEA (MRS), and on a small scale using standard laboratory rearing trays (SRS). Mosquito life history traits such as pupation rate, emergence rate, adult size as well as the effect of irradiation on adult longevity were evaluated. Moreover, 5–6 day old mosquitoes were released into field cages and left for two nights to mate and the mating competitiveness between sterile mass-reared males and fertile males reared on a small scale when competing for small scale reared virgin females was investigated. Resulting fertility in a treatment ratio of 1:1:1 (100 irradiated males: 100 non-irradiated males: 100 virgin females) was compared to control cages with 0:100:100 (non-irradiated control) and 100:0:100 (irradiated control). Results No significant differences in life history parameters were observed between rearing methods. The competitiveness index of mass reared males (0.58) was similar to males reared on a small scale (0.59). A residual fertility rate of 20% was observed in the irradiated control (100:0:100), measured as the percentage of eggs collected from the cages which developed to adulthood. No significant difference was observed (t = 0.2896, df = 4, P = 0.7865) between the rearing treatments (MRS and SRS) in the fertility rate, a measure of mating competitiveness. Conclusions The results showed that the FAO/IAEA mass-rearing process did not affect mosquito life history parameters or the mating competitiveness of males

Co-production of surfactin and a novel bacteriocin by Bacillus subtilis subsp. subtilis H4 isolated from bikalga, an African alkaline Hibiscus sabdariffa seed fermented condiment.

Bikalga is a Hibiscus sabdariffa seed fermented condiment widely consumed in Burkina Faso and neighboring countries. The fermentation is dominated by Bacillus subtilis group species. Ten B. subtilis subsp. subtilis (six isolates) and Bacillus licheniformis (four isolates) isolated from traditional Bikalga were examined for their antimicrobial activity against a panel of 36 indicator organisms including Gram-positive and Gram-negative bacteria and yeasts. The Bacillus spp. isolates showed variable inhibitory abilities depending on the method used. Both Gram-positive and Gram-negative bacteria were inhibited in the agar spot assay while only Gram-positive pathogens were inhibited in the agar well diffusion assay. Cell free supernatants (CFS) of pure cultures of 3 B. subtilis subsp. subtilis (G2, H4 and F1) strains inhibited growth of Listeria monocytogenes, Micrococcus luteus, Staphylococcus aureus and Bacillus cereus, while CFS of 2 B. licheniformis (E3 and F9) strains only inhibited M. luteus. The antimicrobial substance(s) produced by B. subtilis subsp. subtilis H4 was further characterized. The antimicrobial substance(s) produced by H4 was detected from mid-exponential growth phase. The activity was sensitive to protease and trypsin, but resistant to the proteolytic action of proteinase K and papain. Treatment with α-amylase and lipase II resulted in a complete loss of antimicrobial effect, indicating that a sugar moiety and lipid moiety are necessary for the activity. Treatment with mercapto-ethanol resulted in a significant loss, indicative of the presence of disulfide bridges. The antimicrobial activity of H4 was heat resistant and active at pH3–10. PCR detection of yiwB, sboA, spoX, albA and spaS, etnS genes and genes coding for surfactins and plipastatins (fengycins) indicated a potential for subtilosin, subtilin and lipopeptide production, respectively. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was carried out and a single band of approximately 4kDa had antimicrobial activity. Ultra high performance liquid chromatography-time of flight mass spectrometry (UHPLC-TOFMS) analysis of the 4kDa band allowed identification of surfactin and a protein with a monoisotopic mass of 3346.59Da, which is dissimilar in size to subtilosin and subtilin. Surfactin is a cyclic lipoheptapeptide, which contains a β-hydroxy fatty acid, but no di-sulfide bridges or sugar residues. The complete loss of activity upon amylase treatment indicates that surfactin was not responsible for the observed antimicrobial effect. However, it cannot completely be ruled out that surfactin acts synergistically with the detected protein, though further investigations are needed to confirm this