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The SD Bioline® IgG4 rapid diagnostic test (RDT) detects IgG4 antibodies induced by the Onchocerca volvulus-specific antigen Ov16. We evaluated the stability of the RDT results over 1 month, at different time points after completion of each assay, using eluted dried blood spots collected in central Cameroon. Agreement coefficients regarding positivity between 30 minutes and 24 hours, 1, 2, 3, and 4 weeks were, 96.4%, 93.4%, 93.3%, 93.2%, and 93.2%, respectively. Between 30 minutes and 24 hours, 3.6% of the 15,444 tests showed inconsistent results with 81.2% of these tests changing from negative to positive, increasing O. volvulus antibody prevalence from 23.9% to 26.2% (P < 0.0001). This change from negative to positive outcome was confirmed at the subsequent timepoints. Depending on the desired accuracy of prevalence estimates, reading time may have to be redefined more strictly.

Monocyte dysfunction in helminth infection is one of the mechanisms proposed to explain the diminished parasite antigen-specific T cell responses seen with patent filarial infection. In fact, monocytes from filariae-infected individuals demonstrate internalized filarial antigens and, as a consequence, express inhibitory surface molecules and have diminished cytokine production. To investigate the mechanisms underlying monocyte dysfunction in filarial infections, purified human monocytes were exposed to live microfilariae (mf) of Brugia malayi, and the mRNA and protein expression of important inhibitory and/or autophagy-related molecules were assessed. Our data indicate that mf-induced autophagy in human monocytes shown by the formation of autophagic vesicles, by the upregulation in the mRNA expression of autophagy-related genes BCN1, LC3B, ATG5, ATG7 (P < 0.05), and by increase in the levels of LC3B protein. Furthermore, this mf-induced autophagy increased the levels of monocyte CD206 expression. In addition, mf significantly induced the frequency of interferon (IFN)-γ+ human monocytes and at the same time induced the mRNA expression of indoleamine 2,3-dioxygenase (IDO) through an IFN-γ-dependent mechanism; significantly enhanced tryptophan degradation (an indicator of IDO activity; P < 0.005). Interestingly, this autophagy induction by mf in monocytes was IFN-γ-dependent but IDO-independent as was reversed by anti-IFN-γ but not by an IDO inhibitor. Our data collectively suggest that mf of Brugia malayi regulate the function of monocytes by induction of IDO and IFN-γ, induce autophagy through an IFN-γ-dependent mechanism, and increase M2 phenotype through induction of autophagy; all acting in concert to drive monocyte dysfunction.

Lymphatic filariasis (LF) is a debilitating disease that afflicts over 70 million people worldwide. It is caused by the parasitic nematodes Wuchereria bancrofti, Brugia malayi, and Brugia timori. While efforts to eliminate LF have seen substantial success, complete eradication will likely require more time and resources than predicted. Identifying new drug and vaccine targets in adult filariae could help elimination efforts.This study’s aim was to evaluate intestinal proteins in adult Brugia malayi worms as possible therapeutic targets. Using siRNA, we successfully inhibited transcripts of four candidate genes: Bma-Serpin, Bma-ShTK, Bma-Reprolysin, and Bma-LAD-2. Of those, Bma-LAD-2, an immunoglobulin superfamily cell adhesion molecule (IgSF CAM), was determined to be essential for adult worm survival. We observed a 70.42% knockdown in Bma-LAD-2 transcript levels 1 day post-siRNA incubation and an 87.02% reduction in protein expression 2 days post-siRNA incubation. This inhibition of Bma-LAD-2 expression resulted in an 80% decrease in worm motility over 6 days, a 93.43% reduction in microfilaria release (Mf) by day 6 post-siRNA incubation, and a significant decrease in MTT reduction. Transmission electron microscopy revealed the loss of microvilli and unraveling of mitochondrial cristae in the intestinal epithelium of Bma-LAD-2 siRNA-treated worms. Strikingly, Bma-LAD-2 siRNA-treated worms exhibited an almost complete loss of pseudocoelomic fluid, suggesting that loss of these tight junctions led to the leakage and subsequent loss of the worm’s structural integrity. Luciferase immunoprecipitation system assay demonstrated that serum from 30 patients with LF did not have detectable IgE antibodies against Bma-LAD-2, indicating that LF exposure does not result in IgE sensitization to this antigen.These results indicate that Bma-LAD-2 is an essential protein for adult Brugia malayi and may be an effective drug or vaccine target. In addition, these findings further validate the strategy of targeting the worm intestine to prevent and treat helminthic infections.Author SummaryBrugia malayi is a parasitic nematode that can cause lymphatic filariasis, a debilitating disease prevalent in tropical and subtropical countries. Significant progress has been made towards eliminating the disease. However, complete eradication may require new therapeutics such as drugs or a vaccine that kill adult filariae. In this study, we identified an immunoglobulin superfamily cell adhesion molecule (Bma-LAD-2) as a potential drug and vaccine candidate. When we knocked down Bma-LAD-2 expression, we observed a decrease in worm motility, fecundity, and metabolism. We also visualized the loss of microvilli, destruction of the mitochondria in the intestinal epithelium, and loss of pseudocoelomic fluid contents after Bma-LAD-2 siRNA treatment. Finally, we demonstrated that serum from filaria-infected patients does not contain preexisting IgE to Bma-LAD-2, which indicates that this antigen would likely be safe to administer as a vaccine in endemic populations.

Angiostrongylus cantonensis is one of the leading causes of eosinophilic meningitis worldwide. A field-deployable molecular detection method could enhance both environmental surveillance and clinical diagnosis of this emerging pathogen. Accordingly, RPAcan3990, a recombinase polymerase assay (RPA), was developed to target a region predicted to be highly repeated in the A. cantonensis genome. The assay was then adapted to produce a visually interpretable fluorescent readout using an orange camera lens filter and a blue light. Using A. cantonensis genomic DNA, the limit of detection was found to be 1 fg/μl by both fluorometer measurement and visual reading. All clinical samples known to be positive for A. cantonensis from various areas of the globe were positive by RPAcan3990. Cerebrospinal fluid samples from other etiologies of eosinophilic meningitis (i.e., Toxocara sp. and Gnathostoma sp.) were negative in the RPAcan3990 assay. The optimal incubation temperature range for the reaction was between 35°C and 40°C. The assay successfully detected 1 fg/μl of A. cantonensis genomic DNA after incubation at human body temperature (in a shirt pocket). In conclusion, these data suggest RPAcan3990 is potentially a point-of-contact molecular assay capable of sensitively detecting A. cantonensis by producing visually interpretable results with minimal instrumentation.

The most common causes of eosinophilia globally are helminth parasites. Refugees from high endemic areas are at increased risk of infection compared with the general U.S. population. It is widely accepted that eosinophilia is a good marker for helminth infection in this population, yet its absence has little predictive value for excluding infection. During an enhanced premigration health program, the CDC offered voluntary testing and management of intestinal parasites, among other conditions, to U.S.-bound refugees in Thailand. Stool specimens were tested for Ascaris lumbricoides, Strongyloides stercoralis, Trichuris trichiura, hookworms, Giardia lamblia, Cryptosporidium spp., and Entamoeba histolytica using quantitative polymerase chain reaction. Complete blood counts were performed to identify eosinophilia. Predictive values of eosinophilia for parasitic infections were calculated within nematode groups. Between July 9, 2012 and November 29, 2013, 2,004 participants were enrolled. About 73% were infected with at least one parasite. The overall median eosinophil count was 483 cells/μL (interquartile range [IQR] = 235–876 cells/μL). Compared with participants who did not test positive for any infection, higher eosinophil counts were observed in those infected with A. lumbricoides (RR = 1.3, 95% CI = 1.1–1.4), S. stercoralis (RR = 1.8, 95% CI = 1.4–2.4), Necator americanus (RR = 1.2, 95% CI = 1.1–1.4), and Ancylostoma ceylanicum (RR = 1.8, 95% CI = 1.5–2.2). Eosinophil counts were higher in younger participants (2–4 years versus 65+ years: RR = 4.2, 95% CI = 2.5–6.9), and lower in female participants (RR = 0.9, 95% CI = 0.8–0.9). Sensitivities ranged from 51% to 73%, specificities from 48% to 65%, and predictive values from 4% to 98%. The predictive value of eosinophilia is poor for the most common parasitic infections, and it should not be used alone for screening refugees.

Angiostrongylus cantonensis is one of the leading causes of eosinophilic meningitis worldwide. A field-deployable molecular detection method could enhance both environmental surveillance and clinical diagnosis of this emerging pathogen. Accordingly, RPAcan3990, a recombinase polymerase assay (RPA), was developed to target a region predicted to be highly repeated in the A. cantonensis genome. The assay was then adapted to produce a visually interpretable fluorescent readout using an orange camera lens filter and a blue light. Using A. cantonensis genomic DNA, the limit of detection was found to be 1 fg/μl by both fluorometer measurement and visual reading. All clinical samples known to be positive for A. cantonensis from various areas of the globe were positive by RPAcan3990. Cerebrospinal fluid samples from other etiologies of eosinophilic meningitis (i.e., Toxocara sp. and Gnathostoma sp.) were negative in the RPAcan3990 assay. The optimal incubation temperature range for the reaction was between 35°C and 40°C. The assay successfully detected 1 fg/μl of A. cantonensis genomic DNA after incubation at human body temperature (in a shirt pocket). In conclusion, these data suggest RPAcan3990 is potentially a point-of-contact molecular assay capable of sensitively detecting A. cantonensis by producing visually interpretable results with minimal instrumentation.

A molecule termed BmIL5Rbp (aka Bm8757) was identified from Brugia malayi filarial worms and found to competitively inhibit human IL-5 binding to its human receptor. After the expression and purification of a recombinant BmIL5Rbp and generation of BmIL5Rbp-specific rabbit antibody, we localized the molecule on B. malayi worms through immunohistochemistry and immunoelectron microscopy. RNA interference was used to inhibit BmIL5Rbp mRNA and protein production. BmIL5Rbp was shown to localize to the cuticle of Brugia malayi and to be released in their excretory/secretory products. RNAi inhibited BmIL5Rbp mRNA production by 33% and reduced the surface protein expression by ~50% and suppressed the release of BmIL5Rbp in the excretory/secretory products. RNAi has been used successfully to knock down the mRNA and protein expression of BmIL5Rbp in the early larval stages of B. malayi and provided a proof-of-principle for the local inhibition of the human IL5 receptor. These findings provide evidence that a parasite encoded IL5R antagonist could be utilized therapeutically.

The establishment of infection with the lymphatic dwelling filarial parasites is dependent on the infectivity and subsequent development of the infective larvae (L3) within the human host to later stages (L4, adults) that require several developmental molts. The molecular mechanisms underlying the developmental processes in parasitic nematodes are not clearly defined. We report the proteomic profiles throughout the entire L3 to L4 molt using an established in vitro molting process for the human pathogen B. malayi. A total of 3466 proteins of B. malayi and 54 from Wolbachia were detected at one or more time points. Based on the proteomic profiling, the L3 to L4 molting proteome can be broadly divided into an early, middle and late phase. Enrichment of proteins, protein families and functional categories between each time point or between phases primarily relate to energy metabolism, immune evasion through secreted proteins, protein modification, and extracellular matrix-related processes involved in the development of new cuticle. Comparative analyses with somatic proteomes and transcriptomes highlighted the differential usage of cysteine proteinases (CPLs), BmCPL-1, -4 and -5 in the L3-L4 molt compared to the adults and microfilariae. Inhibition of the CPLs effectively blocked the in-vitro L3 to L4 molt. Overall, only 4 Wolbachia proteins (Wbm0495, Wbm0793, Wbm0635, and Wbm0786) were detected across all time points and suggest that they play an inconsequential role in the early developmental process.ImportanceThe neglected tropical diseases of lymphatic filariasis, onchocerciasis (or river blindness), and loiasis are the three major filarial infections of humans that cause long-term disability, impaired childhood growth, reduced reproductive capacity. Global efforts to control and/or eliminate these infections as a public health concern are based on strategies and tools to strengthen the diagnostics, therapeutic and prophylactic measures. A deeper understanding of the genes, proteins and pathways critical for the development of the parasite is needed to help further investigate the mechanisms of parasite establishment and disease progression, because not all the transmitted infective larvae get to develop successfully and establish infections. The significance of this study is in identifying the proteins and the pathways that are needed by the parasite for successful developmental molts, that in turn will allow for investigating targets of therapeutic and prophylactic potential.

Using multiparameter flow cytometry human innate lymphoid cell (ILC) subsets can be detected in the circulation, in relatively low frequencies. Despite the low frequency of ILCs in circulation, ex vivo experiments have demonstrated that these ILCs release extremely large per cell quantities of signature ILC cytokines following activation. To determine how activated ILC cytokine production is regulated, ILC subsets were activated in the presence or absence of the immunoregulatory cytokines IL-10 and TGF-β. An examination of circulating ILC subsets revealed surface expression of IL-10Rα and mRNA expression of both IL-10Rα and TGF-βR1 for all ILC subsets. Stimulated ILC1 production of IFN-γ was decreased by TGF-β and not IL-10. Interestingly, ILC2s stimulated in the presence of IL-10 had a marked reduction in cytokine production of IL-5 and IL-13 while TGF-β had no effect on ILC2 cytokine production. Ex vivo activated ILC1 and ILC2 subsets were also found to be a source of the immunoregulatory cytokine IL-10, raising the potential for ILC-mediated regulation of immune cells. These findings demonstrate the differential effects of immunoregulatory cytokines IL-10 and TGF-β on activated ILC1 and ILC2 populations ex vivo.

This study investigates the relationship between helminth infection and allergic sensitization by assessing the influence of preexisting allergy on the outcome of helminth infections, rather than the more traditional approach in which the helminth infection precedes the onset of allergy. Here we used a murine model of house dust mite–induced (HDM-induced) allergic inflammation followed by Ascaris infection to demonstrate that allergic sensitization drives an eosinophil-rich pulmonary type 2 immune response (Th2 cells, M2 macrophages, type 2 innate lymphoid cells, IL-33, IL-4, IL-13, and mucus) that directly hinders larval development and reduces markedly the parasite burden in the lungs. This effect is dependent on the presence of eosinophils, as eosinophil-deficient mice were unable to limit parasite development or numbers. In vivo administration of neutralizing antibodies against CD4 prior to HDM sensitization significantly reduced eosinophils in the lungs, resulting in the reversal of the HDM-induced Ascaris larval killing. Our data suggest that HDM allergic sensitization drives a response that mimics a primary Ascaris infection, such that CD4(+) Th2-mediated eosinophil-dependent helminth larval killing in the lung tissue occurs. This study provides insight into the mechanisms underlying tissue-specific responses that drive a protective response against the early stages of the helminths prior to their establishing long-lasting infections in the host.

Most of the 30 to 100 million people infected with Strongyloides stercoralis have subclinical (or asymptomatic) infections. These infections are commonly chronic and longstanding. A change in immune status can increase parasite numbers, leading to hyperinfection syndrome, dissemination, and death if unrecognized. The use of corticosteroids and HTLV-1 infection are most commonly associated with the hyperinfection syndrome. Strongyloides adult parasites reside in the small intestine and induce immune responses that are like other nematodes. Definitive diagnosis of S stercoralis infection is based on stool examinations for larvae. S stercoralis remains largely neglected.

Coexistent helminth infections are known to modulate T cell and cytokine responses in latent infection with Mycobacterium tuberculosis. However, their role in modulating chemokine responses in latent tuberculosis (LTB) has not been explored. Because chemokines play a vital role in the protective immune responses in LTB, we postulated that coexistent helminth infection could modulate chemokine production in helminth-LTB coinfection. To test this, we measured the levels of a panel of CC and CXC chemokines at baseline and following mycobacterial Ag or mitogen stimulation in individuals with LTB with (Strongyloides stercoralis+LTB+) or without S. stercoralis (S. stercoralis−LTB+) infection and in individuals without both infections, healthy controls (HC). At baseline (in the absence of a stimulus), S. stercoralis+LTB+ individuals exhibited significantly diminished production of CCL1, CCL2, CCL4, CCL11, CXCL9, CXCL10, and CXCL11 in comparison with S. stercoralis−LTB+ and/or HC individuals. Upon mycobacterial Ag stimulation, S. stercoralis+LTB+ individuals exhibited significantly diminished production of CCL1, CCL2, CCL4, CCL11, CXCL2, CXCL9, and CXCL10 in comparison with S. stercoralis−LTB+ and/or HC individuals. No differences were observed upon mitogen stimulation. Finally, after anthelmintic treatment, the baseline levels of CCL1, CCL2, CCL4, CCL11, and CXCL11 and mycobacterial Ag–stimulated levels of CCL1, CCL2, CCL11, CXCL2, and CXCL10 were significantly increased in S. stercoralis+LTB+ individuals. Thus, our data demonstrate that S. stercoralis+LTB+ individuals are associated with a compromised ability to express both CC and CXC chemokines and that this defect is at least partially reversible upon treatment. Hence, coexistent helminth infection induces downmodulation of chemokine responses in LTB individuals with likely potential effects on tuberculosis pathogenesis.

Highlights • Pfs230 is a candidate malaria transmission blocking vaccine against P. falciparum. • Pfs230 vaccine is being tested in areas where malaria and helminth infections are co-endemic. • Chronic helminth infection induces a marked increase in systemic Th2 and regulatory cytokine levels in mice. • Chronic H. polygyrus bakeri infection does not alter Pfs230 vaccine specific-antibody levels. • Functional activity of Pfs230 vaccine was not impaired by chronic helminth infection in mice., Introduction Malaria transmission blocking vaccines (TBV) are innovative approaches that aim to induce immunity in humans against Plasmodium during mosquito stage, neutralizing the capacity of the infected vectors to transmit malaria. Pfs230D1-EPA/Alhydrogel®, a promising protein-protein conjugate malaria TBV, is currently being tested in human clinical trials in areas where P. falciparum malaria is coendemic with helminth parasites. Helminths are complex metazoans that share the master capacity to downregulate the host immune response towards themselves and also to bystander antigens, including vaccines. However, it is not known whether the activity of a protein-based malaria TBV may be affected by a chronic helminth infection. Methods Using an experimental murine model for a chronic helminth infection (Heligmosomoides polygyrus bakeri - Hpb), we evaluated whether prior infection alters the activity of Pfs230D1-EPA/Alhydrogel® TBV in mice. Results After establishment of a chronic infection, characterized by a marked increase of parasite antigen-specific IgG1, IgA and IgE antibody responses, concomitant with an increase of systemic IL-10, IL-5 and IL-6 levels, the Hpb-infected mice were immunized with Pfs230D1-EPA/Alhydrogel® and the vaccine-specific immune response was compared with that in non-infected immunized mice. TBV immunizations induced an elevated vaccine specific-antibody response, however Pfs230D1 specific-IgG levels were similar between infected and uninfected mice at days 15, 25 and 35 post-vaccination. Absolute numbers of Pfs230D1-activated B cells generated in response to the vaccine were also similar among the vaccinated groups. Finally, vaccine activity assessed by reduction of oocyst number in P. falciparum infected mosquitoes was similar between Hpb-infected and immunized mice with non-infected immunized mice. Conclusion Pfs230D1-EPA/Alhydrogel® efficacy is not impaired by a chronic helminth infection in mice.

Background Human and animal studies have demonstrated that helminth infections are associated with a decreased prevalence of type 2 diabetes mellitus (T2DM). However, very little is known about their biochemical and immunological interactions. Methods To assess the relationship between a soil-transmitted helminth, Strongyloides stercoralis (Ss), and T2DM, we examined analytes associated with glycemic control, metabolic processes, and T-cell–driven inflammation at the time of Ss diagnosis and 6 months after definitive anthelmintic treatment. We measured plasma levels of hemoglobin A1c, glucose, insulin, glucagon, adipocytokines, and T-helper (TH) 1-, 2-, and 17- associated cytokines in patients with T2DM with (INF group) or without (UN group) Ss infection. In INF individuals, we again assessed the levels of these analytes 6 months following anthelmintic treatment. Results Compared to UN individuals, INF individuals exhibited significantly diminished levels of insulin and glucagon that increased significantly following therapy. Similarly, INF individuals exhibited significantly diminished levels of adiponectin and adipsin that reversed following therapy. INF individuals also exhibited significantly decreased levels of the TH1- and TH17- associated cytokines in comparison to UN individuals; again, anthelmintic therapy augmented these levels. As expected, INF individuals had elevated levels of TH2-associated and regulatory cytokines that normalized following definitive therapy. Multivariate analysis revealed that these changes were independent of age, sex, body mass index, and liver and renal function. Conclusions Strongyloides stercoralis infection is associated with a significant modulation of glycemic, hormonal, and cytokine parameters in T2DM and its reversal following anthelmintic therapy. Hence, Ss infection has a protective effect on diabetes-related parameters.

Background Various epidemiological and experimental studies propose that helminths could play a preventive role against the progression of type 2 diabetes mellitus (T2DM). T2DM induces microvascular and large vessel complications mediated by elevated levels of angiogenic factors and soluble receptor for advanced glycation end product (RAGE) ligands. However, the interactions between helminths and host angiogenic factors and RAGE ligands are unexplored. Methods To assess the relationship between a soil-transmitted helminth, Strongyloides stercoralis (Ss), and T2DM, we measured plasma levels of vascular endothelial growth factor (VEGF)–A, -C, and -D; angiopoietins 1 and 2 (Ang-1 and Ang-2); and their receptors VEGF-R1, -R2, and -R3 as well as soluble RAGE (sRAGE) and their ligands advanced glycation end products (AGEs), S100A12, and high mobility group box 1 (HMGB-1) in individuals with T2DM with or those without Ss infection. In Ss-infected individuals, we also measured the levels of aforementioned factors 6 months following anthelmintic therapy. Results Ss-infected individuals exhibited significantly decreased levels of VEGF-A, VEGF-C, VEGF-D, Ang-1, and Ang-2 and their soluble receptors VEGF-R1, -R2, and -R3, that increased following anthelmintic therapy. Likewise, Ss-infected individuals exhibited significantly decreased levels of AGEs and their ligands sRAGE, S100A12, and HMGB-1, which reversed following anthelmintic therapy. Conclusions Our data suggest that Ss infection could play a beneficial role by limiting or delaying T2DM-related vascular complications.

We have previously shown that the microfilarial (mf) stage of Brugia malayi can inhibit the mammalian target of rapamycin (mTOR; a conserved serine/threonine kinase critical for immune regulation and cellular growth) in human dendritic cells (DC) and we have proposed that this mTOR inhibition is associated with the DC dysfunction seen in filarial infections. Extracellular vesicles (EVs) contain many proteins and nucleic acids including microRNAs (miRNAs) that might affect a variety of intracellular pathways. Thus, EVs secreted from mf may elucidate the mechanism by which the parasite is able to modulate the host immune response during infection. EVs, purified from mf of Brugia malayi and confirmed by size through nanoparticle tracking analysis, were assessed by miRNA microarrays (accession number GSE157226) and shown to be enriched (>2-fold, p-value, Author summary Lymphatic filariasis, a neglected tropical disease caused by parasitic worms which affects millions of individuals, represents an important public health concern due to its high morbidity that significantly diminishes quality of life. The parasite is able to establish a chronic infection by manipulating its host’s immune responses. The larval mf stage of the parasite is of particular interest as the parasites are present in the peripheral blood and in constant contact with the host’s immune cells. We decided to investigate the role of mf-derived EVs in the modulation of human antigen presenting cells during infection. We showed that mf release EVs that are similar to exosomes, and these parasite vesicles are readily internalized by human DC. The mf-derived EVs possess a unique miRNA profile and are enriched in miRNAs that can target the mTOR pathway. We have also demonstrated that purified mf-derived EVs are capable of inhibiting mTOR signaling in human monocytes. Collectively, our results suggest that mf release exosome-like vesicles that modulate the immune metabolism of host antigen presenting cells.

Background Ivermectin-based onchocerciasis elimination, reported in 2009–2012, for Bakoye and Falémé, Mali, supported policy-shifting from morbidity control to elimination of transmission (EOT). These foci are coendemic with lymphatic filariasis (LF). In 2007–2016 mass ivermectin plus albendazole administration was implemented. We report Ov16 (onchocerciasis) and Wb123 (LF) seroprevalence after 24–25 years of treatment to determine if onchocerciasis EOT and LF elimination as a public health problem (EPHP) have been achieved. Methods The SD Bioline Onchocerciasis/LF Ig[immunoglobulin]G4 biplex rapid diagnostic test (RDT) was used in 2186 children aged 3–10 years in 13 villages (plus 2 hamlets) in Bakoye and in 2270 children in 15 villages (plus 1 hamlet) in Falémé. In Bakoye, all-age serosurveys were conducted in 3 historically hyperendemic villages (1867 individuals aged 3 -78 years). Results In Bakoye, IgG4 seropositivity was 0.27% (95% confidence interval [CI] = .13%–.60%) for both Ov16 and Wb123 antigens. In Falémé, Ov16 and Wb123 seroprevalence was 0.04% (95% CI = .01%–.25%) and 0.09% (95% CI = .02%–.32%), respectively. Ov16-seropositive children were from historically meso/hyperendemic villages. Ov16 positivity was, Ov16–Wb123 (onchocerciasis– lymphatic filariasis [LF]) seroprevalence was measured in children aged ≤10 years to determine if onchocerciasis and elimination was achieved in 2 coendemic foci (Bakoye and Falémé) in Mali under long-term ivermectin treatment, which stopped in 2016. These goals seem to have been achieved.

Background Angiostrongylus cantonensis (Ac), or the rat lungworm, is a major cause of eosinophilic meningitis. Humans are infected by ingesting the 3rd stage larvae from primary hosts, snails, and slugs, or paratenic hosts. The currently used molecular test is a qPCR assay targeting the ITS1 rDNA region (ITS1) of Ac. Methods In silico design of a more sensitive qPCR assay was performed based on tandem repeats predicted to be the most abundant by the RepeatExplorer algorithm. Genomic DNA (gDNA) of Ac were used to determine the analytical sensitivity and specificity of the best primer/probe combination. This assay was then applied to clinical and environmental samples. Results The limit of detection of the best performing assay, AcanR3990, was 1 fg (the DNA equivalent of 1/100 000 dilution of a single 3rd stage larvae). Out of 127 CDC archived CSF samples from varied geographic locations, the AcanR3990 qPCR detected the presence of Ac in 49/49 ITS1 confirmed angiostrongyliasis patients, along with 15/73 samples previously negative by ITS1 qPCR despite strong clinical suspicion for angiostrongyliasis. Intermediate hosts (gastropods) and an accidental host, a symptomatic horse, were also tested with similar improvement in detection observed. AcanR3990 qPCR did not cross-react in 5 CSF from patients with proven neurocysticercosis, toxocariasis, gnathostomiasis, and baylisascariasis. AcanR3990 qPCR failed to amplify genomic DNA from the other related Angiostrongylus species tested except for Angiostrongylus mackerrasae (Am), a neurotropic species limited to Australia that would be expected to present with a clinical syndrome indistinguishable from Ac. Conclusion These results suggest AcanR3990 qPCR assay is highly sensitive and specific with potential wide applicability as a One Health detection method for Ac and Am.

Human ascariasis is a major neglected tropical disease caused by the nematodeAscaris lumbricoides. We report a 296 megabase (Mb) reference-quality genome comprised of 17,902 protein-coding genes derived from a single, representativeAscarisworm. An additional 68 worms were collected from 60 human hosts in Kenyan villages where pig husbandry is rare. Notably, the majority of these worms (63/68) possessed mitochondrial genomes that clustered closer to the pig parasiteAscaris suumthan toA. lumbricoides. Comparative phylogenomic analyses identified over 11 million nuclear-encoded SNPs but just two distinct genetic types that had recombined across the genomes analyzed. The nuclear genomes had extensive heterozygosity, and all samples existed as genetic mosaics with eitherA. suum-like orA. lumbricoides-like inheritance patterns supporting a highly interbredAscarisspecies genetic complex. As no barriers appear to exist for anthroponotic transmission of these ‘hybrid’ worms, a one-health approach to control the spread of human ascariasis will be necessary.

Background Cutaneous leishmaniasis (CL) is a neglected tropical disease causing an estimated 1 million new cases annually. While antimonial compounds are the standard of care worldwide, they are associated with significant adverse effects. Miltefosine, an oral medication, is United States (US) Food and Drug Administration approved to treat CL caused by Leishmania braziliensis, Leishmania guyanensis, and Leishmania panamensis. Evidence of efficacy in other species and side-effect profiles in CL has been limited. Methods Twenty-six patients with CL were treated with miltefosine at the US National Institutes of Health. Species included L. braziliensis (n = 7), L. panamensis (n = 5), Leishmania mexicana (n = 1), Leishmania infantum (n = 3), Leishmania aethiopica (n = 4), Leishmania tropica (n = 2), Leishmania major (n = 1), and unspeciated (n = 3). Demographic and clinic characteristics of the participants, response to treatment, and associated adverse events were analyzed. Results Treatment with miltefosine resulted in cure in 77 % (20/26) of cases, with cures among all species. Common adverse events included nausea/vomiting (97%) and lack of appetite (54%). Clinical management or dose reduction was required in a third of cases. Gout occurred in 3 individuals with a prior history of gout. Most laboratory abnormalities, including elevated creatinine and aminotransferases, were mild and normalized after treatment. Conclusions Our data suggest that miltefosine has good but imperfect efficacy to a wide variety of Leishmania species. While side effects were common and mostly mild to moderate, some resulted in discontinuation of therapy. Due to oral administration, broad efficacy, and manageable toxicities, miltefosine is a viable alternative treatment option for CL, though cost and lack of local availability may limit its widespread use.

Type 2 diabetes mellitus (T2DM) is characterized by heightened systemic inflammation and microbial translocation. Whether concomitant helminth infections can modulate this systemic response is unclear. We examined the presence of markers of systemic inflammation (levels of acute phase proteins) and of microbial translocation [levels of lipopolysaccharide (LPS) and its associated products] in T2DM individuals with (Ss +) or without (Ss -) Strongyloides stercoralis (Ss) infection. We also analyzed these parameters at 6 months following anthelmintic treatment in Ss + individuals. Ss + individuals exhibited significantly diminished levels of alpha-2 macroglobulin, C-reactive protein, haptoglobin and serum amyloid protein A1 compared to Ss - individuals and these levels increased significantly following therapy. Similarly, Ss + individuals exhibited significantly diminished levels of LPS, sCD14, intestinal fatty acid binding protein, LPS binding protein and endotoxin IgG antibody and most of these levels increased significantly following therapy. Thus, helminth infection is associated with attenuation of systemic inflammation and microbial translocation in T2DM and its reversal following anthelmintic therapy.

Several epidemiological and immunological studies indicate a reciprocal association between obesity/metabolic syndrome and helminth infections. Numerous studies demonstrated that obesity is concomitant with chronic low-grade inflammation, which is marked by vital changes in cellular composition and function of adipose tissue. However, the effect of helminth infection on the homeostatic milieu in obesity is not well-understood. To determine the relationship between Strongyloides stercoralis (Ss) infection and obesity, we examined an array of parameters linked with obesity both before and at 6 months following anthelmintic treatment. To this end, we measured serum levels of pancreatic hormones, incretins, adipokines and Type-1, Type-2, Type-17, and other proinflammatory cytokines in those with non-diabetic obesity with (INF) or without Ss infection (UN). In INF individuals, we evaluated the levels of these parameters at 6 months following anthelmintic treatment. INF individuals revealed significantly lower levels of insulin, glucagon, C-peptide, and GLP-1 and significantly elevated levels of GIP compared to UN individuals. INF individuals also showed significantly lower levels of Type-1, Type-17 and other pro-inflammatory cytokines and significantly increased levels of Type-2 and regulatory cytokines in comparison to UN individuals. Most of these changes were significantly reversed following anthelmintic treatment. Ss infection is associated with a significant alteration of pancreatic hormones, incretins, adipokines, and cytokines in obese individuals and its partial reversal following anthelmintic treatment. Our data offer a possible biological mechanism for the protective effect of Ss infection on obesity.

Background As new lymphatic filariasis infections are eliminated through mass chemotherapy, previously affected individuals are left with the sequellae, especially chronic progressive lymphoedema. Currently this is managed by careful attention to limb hygiene to prevent infection. Studies over the past 15 years have suggested that the incorporation of doxycycline treatment may arrest or even reverse progression of lymphoedema. Most of this work has been observational or based on small studies, and if this intervention is effective, studies need to be conducted on a larger scale and under diverse geographical and social conditions before it can be incorporated into treatment policy. Methods/Design The double-blind, placebo-controlled study was designed to investigate the impact of six weeks treatment with doxycycline added to standard limb hygiene on early stage filarial lymphoedema in five sites in Africa and the Indian subcontinent. One site in Cameroon is selected for studying lymphoedema in podoconiosis. Each site was individually powered with the potential to undertake a meta-analysis on completion. Evaluation methods followed those used in Ghana in 2012 with additions resulting from advances in technology. The details of the core protocol and how it was varied to take account of differing situations at each of the sites are provided. The study will enrol up to 1800 patients and will complete in mid-2021. Conclusions This paper provides details of what challenges were faced during its development and discusses the issues and how they were resolved. In particular, the reasons for inclusion of new technology and the problems encountered with the supply of drugs for the studies are described in detail. By making these details available, it is hoped that the study protocol will help others interested in improving treatment for filarial lymphoedema in the design of future studies. Trial registration India: Clintrials.gov. NCT02929121 registered 10 Oct 2016: https://clinicaltrials.gov/ct2/show/NCT02929121 Mali: Clintrials.gov. NCT02927496 registered 7 Oct 2016: https://clinicaltrials.gov/ct2/show/NCT0292749 Sri Lanka: Clintrials.gov. NCT02929134 registered 10 Oct 2016: https://clinicaltrials.gov/ct2/show/NCT02929134 Ghana: ISRCTN. 14042737 registered 10 July 2017: 10.1186/ISRCTN14042737 Tanzania: ISRCTN. 65756724 registered 21 July 2017: 10.1186/ISRCTN65756724 Cameroon: ISRCTN. 1181662 registered 25 July 2017: 10.1186/ISRCTN11881662

Background We have previously demonstrated that eosinophil-associated processes underlie some of the differences in clinical presentation among patients with Loa loa infection prior to therapy and that some posttreatment adverse events appear to be dependent on eosinophil activation. Methods We first conducted a retrospective review of 204 patients (70 microfilaria [MF] positive/134 negative) with Loa loa both before and following definitive therapy. We then measured filarial-specific antibodies, eosinophil- and Th2-associated cytokines, and eosinophil granule proteins in their banked serum prior to and at 1 year following definitive treatment. We also evaluated the influence of pretreatment corticosteroids and/or apheresis in altering the efficacy of treatment. Results Patients without circulating microfilariae (MF negative) not only had a higher likelihood of peripheral eosinophilia and increased antifilarial antibody levels but also had significantly increased concentrations of granulocyte-macrophage colony–stimulating factor, interleukin (IL) 5, and IL-4 compared with MF-positive patients. However, these differences had all resolved by 1 year after treatment, when all parameters approached the levels seen in uninfected individuals. Neither pretreatment with corticosteroids nor apheresis reduced the efficacy of the diethylcarbamazine used to treat these subjects. Conclusions Our results highlight that, by 1 year following treatment, infection-associated immunologic abnormalities had resolved in nearly all patients treated for loiasis, and pretreatment corticosteroids had no influence on the resolution of the immunologic perturbations nor on the efficacy of diethylcarbamazine as a curative agent in loiasis. Clinical Trials Registration NCT00001230.

Current assays to detect allergen-specific IgE have constraints related to obtaining pure, conformationally active allergen, variability in allergen extracts, sample volume required, and turnaround time. The luciferase immunoprecipitation systems (LIPS) immunoassay is a fast, sensitive assay created using recombinant antigens that requires a low specimen volume. These assays can also be easily modified to detect multiple antigens and antibody isotypes. Here, we demonstrate the use of LIPS assays as an innovative method to quantitatively measure allergen component-specific IgE in small sample volumes. Sera from healthy volunteers, helminth-infected adults, and peanut-allergic children were screened for IgE to cat using ImmunoCAP. These samples were also measured for IgE against Fel d 1 using LIPS. LIPS signal correlated to cat IgE levels with rS = 0.6204, p < 0.001. The LIPS signal: noise ratio differed significantly between cat IgE-samples and cat IgE+ samples with values > 0.5 kU/L, with the ability to differentiate cat IgE – individuals from cat IgE+ individuals with 85% sensitivity and 76% specificity. Given their rapidity, efficiency, sensitivity, and quantitation over a broad dynamic range, LIPS immunoassays can be a robust and flexible tool with potential uses in allergy research, diagnostics, and treatment.

Nodding syndrome is a rare, enigmatic form of pediatric epilepsy that has occurred in an epidemic fashion beginning in the early 2000s in geographically distinct regions of Africa. Despite extensive investigation, the etiology of nodding syndrome remains unclear, although much progress has been made in understanding the pathogenesis of the disease, as well as in treatment and prevention. Nodding syndrome is recognized as a defined disease entity, but it is likely one manifestation along a continuum of Onchocerca volvulus–associated neurological complications. This review examines the epidemiology of nodding syndrome and its association with environmental factors. It provides a critical analysis of the data that support or contradict the leading hypotheses of the etiologies underlying the pathogenesis of the syndrome. It also highlights the important progress made in treating and preventing this devastating neurological disease and prioritizes important areas for future research.

Summary Background A test-and-not-treat (TaNT) strategy has been developed to prevent people with high concentrations of circulating Loa loa microfilariae (>20 000 microfilariae per mL) developing serious adverse events after ivermectin treatment during mass drug administration to eliminate onchocerciasis. An important question related to cost and programmatic issues is whether annual retesting is required for everyone. We therefore aimed to investigate changes in L loa microfilarial densities during TaNT campaigns run 18 months apart. Methods In this observational cohort study, we assessed the participants of two TaNT campaigns for onchocerciasis. These campaigns, which were run by a research team, together with personnel from the Ministry of Health and community health workers, were done in six health areas (in 89 communities) in Okola health district (Cameroon); the first campaign was run between Aug 10, and Oct 29, 2015, and the second was run between March 7, and May 26, 2017. All individuals aged 5 years and older were invited to be screened for Loa loa microfilaraemia before being offered ivermectin (unless contraindicated). L loa microfilarial density was measured at the point of care using the LoaScope. All those with a L loa microfilarial density of 20 000 microfilariae per mL or less were offered treatment; in the first 2 weeks of the 2015 campaign, a higher exclusion threshold of 26 000 microfilariae per mL or less was used. At both rounds of the intervention, participants were registered with a paper form, in which personal information were collected. In 2017, we also recorded whether each individual reported participation in the 2015 campaign. The primary outcome, assessed in all participants, was whether L loa microfilarial density was above or below the exclusion threshold (ie, the criteria that guided the decision to treat). Findings In the 2015 TaNT campaign, 26 415 people were censused versus 29 587 people in the 2017 TaNT campaign. All individuals aged 5 years and older without other contraindications to treatment (22 842 people in 2015 and 25 421 people in 2017) were invited to be screened for L loa microfilaraemia before being offered ivermectin. In 2015, 16 182 individuals were examined with the LoaScope, versus 18 697 individuals in the same communities in 2017. 344 (2·1%) individuals were excluded from ivermectin treatment because of a high L loa microfilarial density in 2015, versus 283 (1·5%) individuals in 2017 (p99·9%) of 6983 individuals treated with ivermectin in 2015 had L loa microfilariae density below the level associated with neurological serious adverse events. Interpretation Individuals treated with ivermectin do not need to be retested for L loa microfilaraemia before the next treatment, provided that they can be re-identified. This adjusted approach will enable substantial cost savings and facilitate reaching programmatic goals for elimination of onchocerciasis in areas that are co-endemic for loiasis. Funding Bill & Melinda Gates Foundation, Division of Intramural Research (National Institute of Allergy and Infectious Diseases, US National Institutes of Health).

Yemen is a country that has been treating severe cases of oncho-dermatitis since 1992 and is now moving to a program aimed at the elimination of the transmission of Onchocerca volvulus. It is important to ensure that the currently acceptable tools used in epidemiological assessment of onchocerciasis in Africa and Latin America also apply to Yemen. Five hundred and ten blood samples from three known O. volvulus–endemic areas, locations that have never been under a mass treatment program, were tested for the presence of antibodies against a panel of O. volvulus–specific antigens using enzyme-linked immunosorbent assay (Ov16) and luciferase immunoprecipitation system (Ov-FAR-1 and Ov-MSA-1) assays. Overall, 31.4% of the samples tested were positive, with positivity increasing with age. Positivity was seen in 76.5% of those presenting with clinical onchocerciasis but importantly also in more than 28.5% of those defined as free of oncho-dermatitis; these latter individuals are likely to be serving as a source for persistent reinfection. This study supports the use of the current O. volvulus–specific serologic methodology in Yemen.

Chronic helminth infections are known to be associated with the modulation of antigen-specific T-cell responses. Strongyloides stercoralis infection is characterized by the downmodulation of antigen-specific Th1 and Th17 responses and the upregulation of Th2 and Th9 responses. Immune homeostasis is partially maintained by negative regulators of T-cell activation, cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) and programmed death 1 (PD-1), which dampen effector responses during chronic infections. However, their roles in S. stercoralis infection are yet to be defined. Therefore, we sought to determine the role of CTLA-4 and PD-1 in regulating CD4+ and CD8+ T-cell responses and examined the frequencies of monofunctional and dual functional Th1/T cytotoxic type 1 (Tc1), Th17/Tc17, Th2/Tc2, and Th9/Tc9 cells in S. stercoralis infection in 15 infected individuals stimulated with parasite antigen following CTLA-4 or PD-1 blockade. Our data reveal that CTLA-4 or PD-1 blockade results in significantly enhanced frequencies of monofunctional and dual functional Th1/Tc1 and Th17/Tc17 cells and, in contrast, diminishes the frequencies of monofunctional and dual functional Th2/Tc2 and Th9/Tc9 cells with parasite antigen stimulation in whole-blood cultures. Thus, we demonstrate that CTLA-4 and PD-1 limit the induction of particular T-cell subsets in S. stercoralis infection, which suggests the importance of CTLA-4 and PD-1 in immune modulation in a chronic helminth infection.

This hookworm, uncommonly found in humans, has a higher cure rate than that for Necator americanus hookworm., During 2012–2015, US-bound refugees living in Myanmar–Thailand border camps (n = 1,839) were surveyed for hookworm infection and treatment response by using quantitative PCR. Samples were collected at 3 time points: after each of 2 treatments with albendazole and after resettlement in the United States. Baseline prevalence of Necator americanus hookworm was 25.4%, Ancylostoma duodenale 0%, and Ancylostoma ceylanicum (a zoonosis) 5.4%. Compared with N. americanus prevalence, A. ceylanicum hookworm prevalence peaked in younger age groups, and blood eosinophil concentrations during A. ceylanicum infection were higher than those for N. americanus infection. Female sex was associated with a lower risk for either hookworm infection. Cure rates after 1 dose of albendazole were greater for A. ceylanicum (93.3%) than N. americanus (65.9%) hookworm (p

Ivermectin treatment can cause central nervous system adverse events (CNS-AEs) in persons with very high-density Loa loa microfilaremia (≥ 30,000 mf/mL blood). Hypoendemic onchocerciasis areas where L. loa is endemic have been excluded from ivermectin mass drug administration programs (MDA) because of the concern for CNS AEs. The rapid assessment procedure for L. loa (RAPLOA) is a questionnaire survey to assess history of eye worm. If ≥ 40% of respondents report eye worm, this correlates with ≥ 2% prevalence of very high-density loiasis microfilaremia, posing an unacceptable risk of CNS-AEs after MDA. In 2016, we conducted a L. loa study in 110 ivermectin-naive, suspected onchocerciasis hypoendemic villages in southern Nigeria. In previous RAPLOA surveys these villages had prevalences between 10% and 67%. We examined 10,605 residents using the LoaScope, a cell phone-based imaging device for rapidly determining the microfilaria (mf) density of L. loa infections. The mean L. loa village mf prevalence was 6.3% (range 0-29%) and the mean individual mf count among positives was 326 mf/mL. The maximum individual mf count was only 11,429 mf/mL, and among 2,748 persons sampled from the 28 villages with ≥ 40% RAPLOA, the ≥ 2% threshold of very high Loa mf density could be excluded with high statistical confidence (P < 0.01). These findings indicate that ivermectin MDA can be delivered in this area with extremely low risk of L. loa-related CNS-AEs. We also concluded that in Nigeria the RAPLOA survey methodology is not predictive of ≥ 2% prevalence of very high-density L. loa microfilaremia.

Although Wuchereria bancrofti (Wb), the causative agent of lymphatic filariasis, is endemic throughout Mali, the prevalence of Wb microfilaremia (Mf) can vary widely between villages despite similar prevalence of infection as assessed by circulating antigen. To examine this variation, cross-sectional data obtained during screening prior to an interventional study in two neighboring villages in Mali were analyzed. The overall prevalence of Wb, as assessed by Wb CAg(circulating antigen), was 50.3% among 373 participants, aged 14-65. Wuchereria bancrofti Mf-positive and negative individuals appeared randomly distributed across the two villages (Moran's I spatial statistic = -0.01, Z score = 0.1, P>0.05). Among the 187 subjects positive for Wb CAg, 117 (62.5%) had detectable Mansonella perstans microfilaremia (Mp Mf) and 64 (34.2%) had detectable Wb microfilaremia. The prevalence of Mp microfilaremia was 73.4% in the Wb Mf-positive group (as compared to 56.9% in the Wb Mf-negative group; p=0.01), and median Wb Mf load was increased in co-infected subjects (267Mf/ml vs 100 Mf/ml; p

Filarial infections of humans cause some of the most important neglected tropical diseases. The global efforts for eliminating filarial infections by mass drug administration programs may require additional tools (safe macrofilaricidal drugs, vaccines, and diagnostic biomarkers). The accurate and sensitive detection of viable parasites is essential for diagnosis and for surveillance programs. Current community-wide treatment modalities do not kill the adult filarial worms effectively; hence, there is a need to identify and develop safe macrofilaricidal drugs. High-throughput sequencing, mass spectroscopy methods and advances in computational biology have greatly accelerated the discovery process. Here, we describe post-genomic developments toward the identification of diagnostic biomarkers and drug targets for the filarial infection of humans.

A patient in Pennsylvania, USA, with common variable immunodeficiency sought care for fever, cough, and abdominal pain. Imaging revealed lesions involving multiple organs. Liver resection demonstrated necrotizing granulomas, recognizable tegument, and calcareous corpuscles indicative of an invasive cestode infection. Sequencing revealed 98% identity to a Versteria species of cestode found in mink.

A patient in Pennsylvania, USA, with common variable immunodeficiency sought care for fever, cough, and abdominal pain. Imaging revealed lesions involving multiple organs. Liver resection demonstrated necrotizing granulomas, recognizable tegument, and calcareous corpuscles indicative of an invasive cestode infection. Sequencing revealed 98% identity to a Versteria species of cestode found in mink.

Antigen-based immunoassays are currently needed for point-of-care quantification of Loa loa microfilariae (mf). Coupling transcriptomic approaches with bioinformatic analysis, we have identified 11 specific putative proteins (coding mRNAs) with potential utility as biomarkers of patent (mf + ) L. loa infection. We successfully developed antigen capture immunoassays to quantify 2 (LOAG_14221 and LOAG_15846) of these proteins in individual plasma/serum samples. Of the 2 quantifiable circulating biomarkers, LOAG_14221 showed the highest degree of specificity, particularly with a monoclonal antibody-based immunoassay. Moreover, the levels of LOAG_14221 in L. loa mf + patients were positively correlated to the mf densities in the corresponding blood samples ( r = 0.53 and P = 0.008 for polyclonal assay; r = 0.54 and P = 0.004 for monoclonal assay). Thus, LOAG_14221 is a very promising biomarker that will be exploited in a quantitative point-of-care immunoassay for determination of L. loa mf densities.

Summary Background Plasmodium falciparum sporozite (PfSPZ) Vaccine is a metabolically active, non-replicating, whole malaria sporozoite vaccine that has been reported to be safe and protective against P falciparum controlled human malaria infection in malaria-naive individuals. We aimed to assess the safety and protective efficacy of PfSPZ Vaccine against naturally acquired P falciparum in malaria-experienced adults in Mali. Methods After an open-label dose-escalation study in a pilot safety cohort, we did a double-blind, randomised, placebo-controlled trial based in Doneguebougou and surrounding villages in Mali. We recruited 18–35-year-old healthy adults who were randomly assigned (1:1) in a double-blind manner, with stratification by village and block randomisation, to receive either five doses of 2·7 × 10 5 PfSPZ or normal saline at days 0, 28, 56, 84, and 140 during the dry season (January to July inclusive). Participants and investigators were masked to group assignments, which were unmasked at the final study visit, 6 months after receipt of the last vaccination. Participants received combined artemether and lumefantrine (four tablets, each containing 20 mg artemether and 120 mg lumefantrine, given twice per day over 3 days for a total of six doses) to eliminate P falciparum before the first and last vaccinations. We collected blood smears every 2 weeks and during any illness for 24 weeks after the fifth vaccination. The primary outcome was the safety and tolerability of the vaccine, assessed as local and systemic reactogenicity and adverse events. The sample size was calculated for the exploratory efficacy endpoint of time to first P falciparum infection beginning 28 days after the fifth vaccination. The safety analysis included all participants who received at least one dose of investigational product, whereas the efficacy analyses included only participants who received all five vaccinations. This trial is registered at ClinicalTrials.gov, number NCT01988636. Findings Between Jan 18 and Feb 24, 2014, we enrolled 93 participants into the main study cohort with 46 participants assigned PfSPZ Vaccine and 47 assigned placebo, all of whom were evaluable for safety. We detected no significant differences in local or systemic adverse events or laboratory abnormalities between the PfSPZ Vaccine and placebo groups, and only grade 1 (mild) local or systemic adverse events occurred in both groups. The most common solicited systemic adverse event in the vaccine and placebo groups was headache (three [7%] people in the vaccine group vs four [9%] in the placebo group) followed by fatigue (one [2%] person in the placebo group), fever (one [2%] person in the placebo group), and myalgia (one [2%] person in each group). The exploratory efficacy analysis included 41 participants from the vaccine group and 40 from the placebo group. Of these participants, 37 (93%) from the placebo group and 27 (66%) from the vaccine group developed P falciparum infection. The hazard ratio for vaccine efficacy was 0·517 (95% CI 0·313–0·856) by time-to-infection analysis (log-rank p=0·01), and 0·712 (0·528–0·918) by proportional analysis (p=0·006). Interpretation PfSPZ Vaccine was well tolerated and safe. PfSPZ Vaccine showed significant protection in African adults against P falciparum infection throughout an entire malaria season. Funding US National Institutes of Health Intramural Research Program, Sanaria.

Background Severe adverse reactions have been observed in individuals with Loa loa infection treated with either diethylcarbamazine (DEC), the drug of choice for loiasis, or ivermectin (IVM), which is used in mass drug administration programs for control of onchocerciasis and lymphatic filariasis in Africa. In this study, posttreatment clinical and immunologic reactions were compared following single-dose therapy with DEC or IVM to assess whether these reactions have the same underlying pathophysiology. Methods Twelve patients with loiasis and microfilarial counts

Since the mid-2000s, the immunochromatographic card test (ICT), a point-of-care test for detecting Wuchereria bancrofti circulating filarial antigens (CFAs), has been the backbone for mapping and monitoring lymphatic filariasis (LF) worldwide. Recently, there have been instances in which CFA positivity has been associated with Loa loa microfilaremia. Here, we examined the association, at both the community and individual levels, between L. loa and CFA using additional diagnostic tools (quantitative polymerase chain reaction [qPCR], Og4C3 enzyme-linked immunosorbent assay, and IgG4 antibodies to Wb123 assays) to demonstrate the relationship between L. loa microfilaremia and ICT positivity. In May 2013, peripheral blood was collected during the day from 1,812 individuals living in southern Cameroon. ICT tests were done on the spot, and positive individuals were resampled at night. Results of qPCR and Wb123 assays concurred proving the absence of W. bancrofti infection. Og4C3 assays indicate a quantitative relationship between the level of L. loa microfilaremia and that of CFA. This was confirmed by epidemiological analyses, which reveal a strong association between L. loa microfilaremia and ICT positivity, with 50% of ICT reacting to L. loa when its microfilarial density exceeds 30,000 microfilariae/mL. At the community level, the proportion of positive ICT would exceed 2% when the prevalence of L. loa microfilaremia in the total population is above 20%. This has significant implications in terms of mapping and control of LF caused by W. bancrofti in Loa-endemic areas. Cross-reactivity of ICT with L. loa has to be considered in the context of both individual and community diagnostics.

Helminth infections are known to influence T- and B- cell responses in latent Mycobacterium tuberculosis infection (LTBI). Whether helminth infections also modulate monocyte responses in helminth-LTBI co-infection has not been fully explored. To this end, we examined the activation, polarization and function of human monocytes isolated from individuals with LTBI with (n=25) or without (n=25) co-incident Strongyloides stercoralis infection (Ss+ and Ss− respectively). Our data reveal that the presence of Ss infection is associated with lower frequencies of monocytes expressing CD54, CD80, CD86 at baseline (absence of stimulation) and in response to mycobacterial-antigen stimulation than monocytes from Ss−individuals. In contrast, Ss infection was associated with higher frequencies of M2-like monocytes, as determined by expression of CD206 and CD163. Monocytes from Ss+ individuals had a reduced capacity to phagocytose or exhibit respiratory burst activity following mycobacterial-antigen or LPS stimulation and were less capable of expression of IL-1β, TNFα, IL-6 and IL-12 at baseline and/or following antigen stimulation compared to those without Ss infection. In addition, definitive treatment of Ss infection resulted in a significant reversal of the altered monocyte function 6 months after anthelmintic therapy. Finally, T cells from Ss+ individuals exhibited significantly lower activation at baseline or following mycobacterial-antigen-stimulation. Therefore, our data highlights the induction of dampened monocyte activation, enhanced M2 polarization and impaired monocyte function in helminth-LTBI coinfection. Our data also reveal a different mechanism by which helminth infection modulates immune function in LTBI.

Background Lymphedema is a public health problem in countries with lymphatic filariasis (LF) including Mali. We studied the epidemiology and clinical presentation of lymphedema in three previously LF-endemic health districts of Mali after at least five consecutive rounds of mass drug administration (MDA) with albendazole and ivermectin. Methods From 2016 to 2018, we used passive and active case finding methods to identify lymphedema cases in three health districts with high pre-MDA LF prevalence: Kolondieba (66%), Bougouni (44%) and Kolokani (34%). Results Three hundred and thirty nine cases of lymphedema were identified, 235 (69.32%) through active case finding. Their median age was 56 years (range 2–90) and 286 (84.36%) were women. Lymphedema was reported in 226 (78.5%) people aged 41 years and older compared to 73 (21.5%) people below the age of 41 years (Chi2 = 17.28, df = 5, p = 0.004). One hundred and seventy five cases of lymphedema were found in Kolondieba (66 per 100,000 people), 116 in Bougouni (19 per 100,000) and 48 in Kolokani (16 per 100,000). Stage III lymphedema was observed in 131 (38.64%), stage II in 108 (31.86%), stage IV in 46 (13.57%), stage I in 23 (6.78%), stage V in 21 (6.19%) and stage VI in ten (2.95%). In the three study districts, lymphedema affected the legs in 281 (82.89%), the arms in 42 (12.39%) and both in 16 (4.72%) (Chi2 = 13.63, p = 0.008). Conclusion Health districts in Mali with the highest pre-MDA LF prevalences had the highest prevalence of lymphedema. Efforts to actively identify lymphedema cases should be scaled up in previous LF-endemic areas, and should be supplemented by a morbidity management and disability prevention plan at the peripheral health system level.

Background With the expansion of soil transmitted helminth (STH) intervention efforts and the corresponding decline in infection prevalence, there is an increased need for sensitive and specific STH diagnostic assays. Previously, through next generation sequencing (NGS)-based identification and targeting of non-coding, high copy-number repetitive DNA sequences, we described the development of a panel of improved quantitative real-time PCR (qPCR)-based assays for the detection of Necator americanus, Ancylostoma duodenale, Ancylostoma ceylanicum, Trichuris trichiura, and Strongyloides stercoralis. However, due to the phenomenon of chromosome diminution, a similar assay based on high copy-number repetitive DNA was not developed for the detection of Ascaris lumbricoides. Recently, the publication of a reference-level germline genome sequence for A. lumbricoides has facilitated our development of an improved assay for this human pathogen of vast global importance. Methodology/Principal findings Repurposing raw DNA sequence reads from a previously published Illumina-generated, NGS-based A. lumbricoides germline genome sequencing project, we performed a cluster-based repeat analysis utilizing RepeatExplorer2 software. This analysis identified the most prevalent repetitive DNA element of the A. lumbricoides germline genome (AGR, Ascaris germline repeat), which was then used to develop an improved qPCR assay. During experimental validation, this assay demonstrated a fold increase in sensitivity of ~3,100, as determined by relative Cq values, when compared with an assay utilizing a previously published, frequently employed, ribosomal internal transcribed spacer (ITS) DNA target. A comparative analysis of 2,784 field-collected samples was then performed, successfully verifying this improved sensitivity. Conclusions/Significance Through analysis of the germline genome sequence of A. lumbricoides, a vastly improved qPCR assay has been developed. This assay, utilizing a high copy-number repeat target found in eggs and embryos (the AGR repeat), will improve prevalence estimates that are fundamental to the programmatic decision-making process, while simultaneously strengthening mathematical models used to examine STH infection rates. Furthermore, through the identification of an optimal target for PCR, future assay development efforts will also benefit, as the identity of the optimized repeat DNA target is likely to remain unchanged despite continued improvement in PCR-based diagnostic technologies., Author summary With an at-risk population in the billions, Ascaris lumbricoides is a pathogen of great global importance. In recent years, efforts to control the spread of this parasitic helminth have expanded, resulting in declining infection rates and worm burdens in some regions. While immeasurably important for global health, these declines have also served to expose the shortcomings of traditional diagnostic methods, as low-levels of pathogen generate a need for more sensitive tools, and microscopy-based techniques are proving ill-suited to the task at hand. Thankfully, improved sensitivity can be achieved through the careful selection of optimal repetitive DNA targets for PCR. However, previous attempts to identify such targets in A. lumbricoides were unsuccessful, largely due to chromosome diminution, an unusual phenomenon occurring in the Ascaridida, whereby large portions of the germline genome are reproducibly eliminated during early development, resulting in their absence in larvae or adult worms. As the stool-based molecular diagnosis of A. lumbricoides infection is primarily dependent upon the identification of egg-derived DNA, utilizing genomic DNA from adult worms for molecular target selection eliminates germline candidates and results in suboptimal target sequence choices. Recently, the publication of a pre-diminution germline genome of A. lumbricoides has provided us with an opportunity to re-evaluate target selection, facilitating the development of a novel quantitative real-time PCR assay with greatly improved sensitivity (~3100-fold as determined by relative Cq value) over previously developed assays that were based on ribosomal repeat DNA sequences with lower copy numbers.

Background Severe adverse events after treatment with ivermectin in individuals with high levels of Loa loa microfilariae in the blood preclude onchocerciasis elimination through community-directed treatment with ivermectin (CDTI) in Central Africa. We measured the cost of a community-based pilot using a test-and-not-treat (TaNT) strategy in the Soa health district in Cameroon. Methods Based on actual expenditures, we empirically estimated the economic cost of the Soa TaNT campaign, including financial costs and opportunity costs that will likely be borne by control programs and stakeholders in the future. In addition to the empirical analyses, we estimated base-case, less intensive, and more intensive resource use scenarios to explore how costs might differ if TaNT were implemented programmatically. Results The total costs of US$283 938 divided by total population, people tested, and people treated with 42% coverage were US$4.0, US$9.2, and US$9.5, respectively. In programmatic implementation, these costs (base-case estimates with less and more intensive scenarios) could be US$2.2 ($1.9–$3.6), US$5.2 ($4.5–$8.3), and US$5.4 ($4.6–$8.6), respectively. Conclusions TaNT clearly provides a safe strategy for large-scale ivermectin treatment and overcomes a major obstacle to the elimination of onchocerciasis in areas coendemic for Loa loa. Although it is more expensive than standard CDTI, costs vary depending on the setting, the implementation choices made by the institutions involved, and the community participation rate. Research on the required duration of TaNT is needed to improve the affordability assessment, and more experience is needed to understand how to implement TaNT optimally., We measured the costs of a community-based pilot using a safe test-and-not-treat strategy to treat onchocerciasis with ivermectin in areas coendemic for Loa loa. Costs per person were US$4.0 (population), US$9.2 (tested), and US$9.5 (treated).

The human disease lymphatic filariasis causes the debilitating effects of elephantiasis and hydrocele. Lymphatic filariasis currently affects the lives of 90 million people in 52 countries. There are three nematodes that cause lymphatic filariasis,Brugia malayi, B. timori, andWuchereria bancrofti, but 90% of all cases of lymphatic filariasis are caused solely byW. bancrofti. Here we use population genomics to identify the geographic origin ofW. bancroftiand reconstruct its spread. Previous genomic sequencing efforts have suffered from difficulties in obtaining Wb DNA. We used selective whole genome amplification to enrichW. bancroftiDNA from infected blood samples and were able to analyze 47 whole genomes ofW. bancroftifrom endemic locations in Haiti, Mali, Kenya, and Papua New Guinea. Our results are consistent with a Southeast Asia or East Asia origin forW. bancroftispread around the globe by infecting migrating populations of humans. Austronesians probably introducedW. ban-croftito Madagascar where later migrations moved it to continental Africa. From Africa,W. bancroftispread to the New World during the transatlantic slave trade. The greater genetic diversity ofW. bancroftipopulations from Haiti are also consistent with genetic admixture from multiple source populations. Genome scans for locally adapted haplotypes identified genes associated with human immune suppression and insecticide sensitivity. Locally adapted haplotypes may provide a foundation to understand the distribution ofW. bancrofticompared to that of other filarial nematodes and how populations may differ in response to eradication efforts.