Geospatial modeling of pre-intervention prevalence of Onchocerca volvulus infection in Ethiopia as an aid to onchocerciasis elimination

Background Onchocerciasis is a neglected tropical and filarial disease transmitted by the bites of blackflies, causing blindness and severe skin lesions. The change in focus for onchocerciasis management from control to elimination requires thorough mapping of pre-control endemicity to identify areas requiring interventions and to monitor progress. Onchocerca volvulus infection prevalence in sub-Saharan Africa is spatially continuous and heterogeneous, and highly endemic areas may contribute to transmission in areas of low endemicity or vice-versa. Ethiopia is one such onchocerciasis-endemic country with heterogeneous O. volvulus infection prevalence, and many districts are still unmapped despite their potential for O. volvulus infection transmission. Methodology/Principle findings A Bayesian geostatistical model was fitted for retrospective pre-intervention nodule prevalence data collected from 916 unique sites and 35,077 people across Ethiopia. We used multiple environmental, socio-demographic, and climate variables to estimate the pre-intervention prevalence of O. volvulus infection across Ethiopia and to explore their relationship with prevalence. Prevalence was high in southern and northwestern Ethiopia and low in Ethiopia's central and eastern parts. Distance to the nearest river (-0.015, 95% BCI: -0.025 - -0.005), precipitation seasonality (-0.017, 95% BCI: -0.032 - -0.001), and flow accumulation (-0.042, 95% BCI: -0.07 - -0.019) were negatively associated with O. volvulus infection prevalence, while soil moisture (0.0216, 95% BCI: 0.014 - 0.03) was positively associated. Conclusions/Significance Infection distribution was correlated with habitat suitability for vector breeding and associated biting behavior. The modeled pre-intervention prevalence can be used as a guide for determining priority for intervention in regions of Ethiopia that are currently unmapped, most of which have comparatively low infection prevalence.

A qPCR to quantify Wolbachia from few Onchocerca volvulus microfilariae as a surrogate for adult worm histology in clinical trials of antiwolbachial drugs

The filarial nematode Onchocerca volvulus causes onchocerciasis (river blindness), a neglected tropical disease affecting 21 million people, mostly in Sub-Saharan Africa. Targeting the endosymbiont Wolbachia with antibiotics leads to permanent sterilization and killing of adult worms. The gold standard to assess Wolbachia depletion is the histological examination of adult worms in nodules beginning at 6 months post-treatment. However, nodules can only be used once, limiting the time points to monitor Wolbachia depletion. A diagnostic to longitudinally monitor Wolbachia depletion from microfilariae (MF) at more frequent intervals < 6 months post-treatment would accelerate clinical trials of antiwolbachials. We developed a TaqMan qPCR amplifying the single-copy gene wOvftsZ to quantify Wolbachia from as few as one MF that had migrated from skin biopsies and compared quantification using circular and linearized plasmids or synthetic dsDNA (gBlock®). qPCR for MF from the rodent nematode Litomosoides sigmodontis was used to support the reproducibility and validate the principle. The qPCR using as few as 2 MF from O. volvulus and L. sigmodontis reproducibly quantified Wolbachia. Use of a linearized plasmid standard or synthesized dsDNA resulted in numbers of Wolbachia/MF congruent with biologically plausible estimates in O. volvulus and L. sigmodontis MF. The qPCR assay yielded a median of 48.8 (range 1.5–280.5) Wolbachia/O. volvulus MF. The qPCR is a sensitive tool for quantifying Wolbachia in a few MF from skin biopsies and allows for establishing the qPCR as a surrogate parameter for monitoring Wolbachia depletion in adult worms of new antiwolbachial candidates.

Bacteriome Diversity of Blackflies’ Gut and Association with Onchocerca volvulus, the Causative Agent of Onchocerciasis in Mbam Valley (Center Region, Cameroon)

Vector control using larvicides is the main alternative strategy to address limits of preventive chemotherapy using ivermectin for the control of onchocerciasis. However, it remains substantially limited by implementation difficulties, ecological concerns and the resistance of vector populations. Therefore, efficient and environmentally safe alternative control strategies are still needed. This study explores the composition of the blackfly bacteriome and its variability in the presence of Onchocerca volvulus infection, in order to determine their potential as a novel vector control-based approach to fight onchocerciasis. An entomological survey of a collection of samples was performed in the Bafia health district, a historical endemic focus for onchocerciasis in Cameroon. A total of 1270 blackflies were dissected and the infection rate was 10.1%, indicative of ongoing transmission of onchocerciasis in the surveyed communities. Sequencing process of blackflies’ gut DNA for bacteria screening revealed 14 phyla and 123 genera, highlighting the diversity of gut blackflies bacterial communities. Eight bacteria formed the core of blackfly bacteriome and Wolbachia was the predominant genus with 73.4% of relative abundance of blackflies’ gut bacterial communities. Acidomonas and Roseanomas genera were significantly abundant among infected blackflies (p = 0.01), whereas other genera such as Brevibacterium and Fructobacillus were associated with the absence of infection (p = 0.0009). Differences in gut bacterial distribution of blackflies according to their infection status by the parasite suggest a causal relationship between the bacteriome composition and the onset of blackflies’ infection by O. volvulus or vice versa. Blackfly native bacteria are then potentially involved in infection by O. volvulus, either by facilitating or preventing the parasite infestation of the vector. These bacteria represent an interesting potential as a biological tool/target for a novel approach of vector control to fight onchocerciasis.

The genome sequence of the Australian filarial nematode, Cercopithifilaria johnstoni

We present a genome assembly and annotation of an individual female Cercopithifilaria johnstoni, a parasitic filarial nematode that is transmitted by hard ticks (Ixodidae) to infect a broad range of native Australian murid and marsupial hosts. The genome sequence is 76.9 Mbp in length, and although in draft form (N50 = 99 kbp, N50[n] = 232), is largely complete based on universally conserved orthologs (BUSCOs; genome = 94.9%, protein = 96.5%) and relative to other related filarial species. These data represent the first genomic resources for the genus Cercopithifilaria, a group of parasites with a broad host range, and form the basis for comparative analysis with the human-infective parasite, Onchocerca volvulus, both of which are responsible for similar eye and skin pathologies in their respective hosts.

A Reevaluation of the Tolerability and Effects of Single-Dose Ivermectin Treatment on Onchocerca volvulus Microfilariae in the Skin and Eyes in Eastern Ghana

Mass administration of ivermectin (IVM) has significantly reduced onchocerciasis prevalence, intensity, and morbidity in most endemic areas. Most IVM clinical trials were performed long ago in persons with high-intensity infections that are uncommon in West Africa today. This cohort treatment study recruited participants from a hypoendemic area in eastern Ghana to reevaluate the efficacy and tolerability of IVM with a special focus on the kinetics of microfilaria (Mf) clearance. Mf in the skin and anterior chambers (ACs) were assessed by skin snip and slit lamp examinations at baseline and at 3 and 6 months after treatment with IVM 150 μg/kg. Out of 231 people, 79 enrolled were treatment-naïve. The baseline geometric mean skin Mf count was 12.67/mg (range 3–86). Although persons with MfAC at baseline (64/231, 27%) had significantly higher skin Mf counts than people without MfAC, 7 of 39 (15%) of persons with skin Mf counts in the range of 3–5 Mf/mg had MfAC. Skin Mf were detected in 14% (31/218) and 45% (96/216) of participants 3 and 6 months after IVM treatment, respectively. MfAC were detected in 12 of 212 (5.7%) study participants at 6 months 81% (187 of 231) of participants experienced 439 adverse events within 7 days after treatment; all adverse events were mild (96.1%) or moderate. This study has provided new data on the kinetics of Mf in the skin and eyes after IVM treatment of persons with light to moderate intensity Onchocerca volvulus infections that are common in Africa at this time.

Multimodal biomarker discovery for active Onchocerca volvulus infection

The neglected tropical disease onchocerciasis, or river blindness, is caused by infection with the filarial nematode Onchocerca volvulus. Current estimates indicate that 17 million people are infected worldwide, the majority of them living in Africa. Today there are no non-invasive tests available that can detect ongoing infection, and that can be used for effective monitoring of elimination programs. In addition, to enable pharmacodynamic studies with novel macrofilaricide drug candidates, surrogate endpoints and efficacy biomarkers are needed but are non-existent. We describe the use of a multimodal untargeted mass spectrometry-based approach (metabolomics and lipidomics) to identify onchocerciasis-associated metabolites in urine and plasma, and of specific lipid features in plasma of infected individuals (O. volvulus infected cases: 68 individuals with palpable nodules; lymphatic filariasis cases: 8 individuals; non-endemic controls: 20 individuals). This work resulted in the identification of elevated concentrations of the plasma metabolites inosine and hypoxanthine as biomarkers for filarial infection, and of the urine metabolite cis-cinnamoylglycine (CCG) as biomarker for O. volvulus. During the targeted validation study, metabolite-specific cutoffs were determined (inosine: 34.2 ng/ml; hypoxanthine: 1380 ng/ml; CCG: 29.7 ng/ml) and sensitivity and specificity profiles were established. Subsequent evaluation of these biomarkers in a non-endemic population from a different geographical region invalidated the urine metabolite CCG as biomarker for O. volvulus. The plasma metabolites inosine and hypoxanthine were confirmed as biomarkers for filarial infection. With the availability of targeted LC-MS procedures, the full potential of these 2 biomarkers in macrofilaricide clinical trials, MDA efficacy surveys, and epidemiological transmission studies can be investigated.

Tandem Use of OvMANE1 and Ov-16 ELISA Tests Increases the Sensitivity for the Diagnosis of Human Onchocerciasis

The current serological test for human onchocerciasis relies on IgG4 reactivity against the parasite Ov-16 antigen, with reported sensitivities of only 60–80%. As control programs move from control to elimination, it is imperative to identify novel molecules that could improve the serodiagnosis reliability of this disease. In this study we compared the sensitivity of total IgG against OvMANE1—a chimeric antigen previously identified as a potential biomarker of human onchocerciasis—with that of an Ov-16 antibody test to detect an Onchocerca volvulus infection in persons presenting with microfilaria in skin snips. One hundred and ninety serum samples were obtained from persons with epilepsy in an onchocerciasis-endemic area at Ituri in the Democratic Republic of Congo where ivermectin has never been distributed. Fifty-nine (31.1%) samples were from individuals with a positive skin snip test; 41 (69.5%) of these 59 samples were positive with the OvMANE1 test and 41 (69.5%) with the Ov-16 test; 30 (50.8%) samples were positive for both tests and in 52 (88.1%) at least one of the tests was positive. Testing the 131 sera from persons with a negative skin snip result revealed that 63 (48.1%) were positive exclusively with the OvMANE1 test, 13 (9.9%) exclusively with the Ov-16 test and 25 (19.1%) with both tests. Nine European samples from individuals without past travel history in onchocerciasis endemic zones and 15 samples from Rwanda, a hypoendemic country for onchocerciasis were all negative for the OvMANE1 and Ov-16 tests. However, the specificity of both tests was difficult to determine due to the lack of a gold standard for antibody tests. In conclusion, the tandem use of OvMANE1 and Ov-16 tests improves the sensitivity of detecting Onchocerca volvulus seropositive individuals but, the OvMANE1 test needs to be further evaluated on samples from a population infected with other helminths to cautiously address its specificity.

Onchocerca volvulus transmission in the Mbam valley of Cameroon following 16 years of annual community-directed treatment with ivermectin, and the description of a new cytotype of Simulium squamosum

Background The onchocerciasis focus surrounding the lower Mbam and Sanaga rivers, where Onchocerca volvulus is transmitted by Simulium damnosum s.l. (Diptera: Simuliidae), was historically the largest in the southern regions of Cameroon. Annual community-directed treatment with ivermectin (CDTI) has been taking place since 2000, but recent studies have shown that new infections are occurring in children. We aimed to investigate blackfly biting and O. volvulus transmission rates along the lower Mbam river 16 years after the formal onset of annual CDTI. Methods Black flies were collected for three consecutive days each month between July 2016 and June 2017 at two riverside villages and two inland sites situated 4.9 km and 7.9 km from the riverside. Specimens collected at each site were dissected on one of the three collection days each month to estimate parity rates and O. volvulus infection rates, while the remaining samples were preserved for pool screening. Results In total, 93,573 S. damnosum s.l. black flies were recorded biting humans and 9281 were dissected. Annual biting rates of up to 606,370 were estimated at the riverside, decreasing to 20,540 at 7.9 km, while, based on dissections, annual transmission potentials of up to 4488 were estimated at the riverside, decreasing to 102 and 0 at 4.9 km and 7.9 km, respectively. However, pool screening showed evidence of infection in black flies at the furthest distance from the river. Results of both methods demonstrated the percentage of infective flies to be relatively low (0.10–0.36%), but above the WHO threshold for interruption of transmission. In addition, a small number of larvae collected during the dry season revealed the presence of Simulium squamosum E. This is the first time S. squamosum E has been found east of Lake Volta in Ghana, but our material was chromosomally distinctive, and we call it S. squamosum E2. Conclusions Relatively low O. volvulus infection rates appear to be offset by extremely high densities of biting black flies which are sustaining transmission along the banks of the lower Mbam river. Graphical Abstract