Early symptom-associated inflammatory responses shift to type 2 responses in controlled human schistosome infection.

Schistosomiasis is an infection caused by contact with Schistosoma-contaminated water and affects more than 230 million people worldwide with varying morbidity. The roles of T helper 2 (TH2) cells and regulatory immune responses in chronic infection are well documented, but less is known about human immune responses during acute infection. Here, we comprehensively map immune responses during controlled human Schistosoma mansoni infection using male or female cercariae. Immune responses to male or female parasite single-sex infection were comparable. An early TH1-biased inflammatory response was observed at week 4 after infection, which was particularly apparent in individuals experiencing symptoms of acute schistosomiasis. By week 8 after infection, inflammatory responses were followed by an expansion of TH2 and regulatory cell subsets. This study demonstrates the shift from TH1 to both TH2 and regulatory responses, typical of chronic schistosomiasis, in the absence of egg production and provides immunological insight into the clinical manifestations of acute schistosomiasis.

Ageing of Plasmodium falciparum malaria sporozoites alters their motility, infectivity and reduces immune activation in vitro.

BACKGROUND: Sporozoites (SPZ), the infective form of Plasmodium falciparum malaria, can be inoculated into the human host skin by Anopheline mosquitoes. These SPZ migrate at approximately 1 µm/s to find a blood vessel and travel to the liver where they infect hepatocytes and multiply. In the skin they are still low in number (50-100 SPZ) and vulnerable to immune attack by antibodies and skin macrophages. This is why whole SPZ and SPZ proteins are used as the basis for most malaria vaccines currently deployed and undergoing late clinical testing. Mosquitoes typically inoculate SPZ into a human host between 14 and 25 days after their previous infective blood meal. However, it is unknown whether residing time within the mosquito affects SPZ condition, infectivity or immunogenicity. This study aimed to unravel how the age of P. falciparum SPZ in salivary glands (14, 17, or 20 days post blood meal) affects their infectivity and the ensuing immune responses. METHODS: SPZ numbers, viability by live/dead staining, motility using dedicated sporozoite motility orienting and organizing tool software (SMOOT), and infectivity of HC-04.j7 liver cells at 14, 17 and 20 days after mosquito feeding have been investigated. In vitro co-culture assays with SPZ stimulated monocyte-derived macrophages (MoMɸ) and CD8+ T-cells, analysed by flow cytometry, were used to investigate immune responses. RESULTS: SPZ age did not result in different SPZ numbers or viability. However, a markedly different motility pattern, whereby motility decreased from 89% at day 14 to 80% at day 17 and 71% at day 20 was observed (p ≤ 0.0001). Similarly, infectivity of day 20 SPZ dropped to ~ 50% compared with day 14 SPZ (p = 0.004). MoMɸ were better able to take up day 14 SPZ than day 20 SPZ (from 7.6% to 4.1%, p = 0.03) and displayed an increased expression of pro-inflammatory CD80, IL-6 (p = 0.005), regulatory markers PDL1 (p = 0.02), IL-10 (p = 0.009) and cytokines upon phagocytosis of younger SPZ. Interestingly, co-culture of these cells with CD8+ T-cells revealed a decreased expression of activation marker CD137 and cytokine IFNγ compared to their day 20 counterparts. These findings suggest that older (day 17-20) P. falciparum SPZ are less infectious and have decreased immune regulatory potential. CONCLUSION: Overall, this data is a first step in enhancing the understanding of how mosquito residing time affects P. falciparum SPZ and could impact the understanding of the P. falciparum infectious reservoir and the potency of whole SPZ vaccines.

Protective efficacy of short-term infection with Necator americanus hookworm larvae in healthy volunteers in the Netherlands: a single-centre, placebo-controlled, randomised, controlled, phase 1 trial.

BACKGROUND: Vaccine development against hookworm is hampered by the absence of the development of protective immunity in populations repeatedly exposed to hookworm, limiting identification of mechanisms of protective immunity and new vaccine targets. Immunisation with attenuated larvae has proven effective in dogs and partial immunity has been achieved using an irradiated larvae model in healthy volunteers. We aimed to investigate the protective efficacy of immunisation with short-term larval infection against hookworm challenge. METHODS: We did a single-centre, placebo-controlled, randomised, controlled, phase 1 trial at Leiden University Medical Center (Leiden, Netherlands). Healthy volunteers (aged 18-45 years) were recruited using advertisements on social media and in publicly accessible areas. Volunteers were randomly assigned (2:1) to receive three short-term infections with 50 infectious Necator americanus third-stage filariform larvae (50L3) or placebo. Infection was abrogated with a 3-day course of albendazole 400 mg, 2 weeks after each exposure. Subsequently all volunteers were challenged with two doses of 50L3 at a 2-week interval. The primary endpoint was egg load (geometric mean per g faeces) measured weekly between weeks 12 and 16 after first challenge, assessed in the per-protocol population, which included all randomly assigned volunteers with available data on egg counts at week 12-16 after challenge. This study is registered with ClinicalTrials.gov, NCT03702530. FINDINGS: Between Nov 8 and Dec 14, 2018, 26 volunteers were screened, of whom 23 enrolled in the trial. The first immunisation was conducted on Dec 18, 2018. 23 volunteers were randomly assigned (15 to the intervention group and eight to the placebo group). Egg load after challenge was lower in the intervention group than the placebo group (geometric mean 571 eggs per g [range 372-992] vs 873 eggs per g [268-1484]); however, this difference was not statistically significant (p=0·10). Five volunteers in the intervention group developed a severe skin rash, which was associated with 40% reduction in egg counts after challenge (geometric mean 742 eggs per g [range 268-1484] vs 441 eggs per g [range 380-520] after challenge; p=0·0025) and associated with higher peak IgG1 titres. INTERPRETATION: To our knowledge, this is the first study to describe a protective effect of short-term exposure to hookworm larvae and show an association with skin response, eosinophilic response, and IgG1. These findings could inform future hookworm vaccine development. FUNDING: Dioraphte Foundation.

Safety and infectivity of female cercariae in Schistosoma-naïve, healthy participants: a controlled human Schistosoma mansoni infection study.

BACKGROUND: A controlled human infection model for schistosomiasis (CHI-S) can speed up vaccine development and provides insight into early immune responses following schistosome exposure. Recently, we established CHI-S model using single-sex male-only Schistosoma mansoni (Sm) cercariae in Schistosoma-naïve individuals. Given important differences in antigenic profile and human immune responses to schistosomes of different sex, we pioneered a single-sex female-only CHI-S model for future use in vaccine development. METHODS: We exposed 13 healthy, Schistosoma-naïve adult participants to 10 (n = 3) or 20 (n = 10) female cercariae and followed for 20 weeks, receiving treatment with praziquantel (PZQ) 60 mg/kg at week 8 and 12 after exposure. FINDINGS: The majority (11/13) participants reported rash and/or itch at the site of exposure, 5/13 had transient symptoms of acute schistosomiasis. Exposure to 20 cercariae led to detectable infection, defined as serum circulating anodic antigen levels >1.0 pg/mL, in 6/10 participants. Despite two rounds of PZQ treatment, 4/13 participants showed signs of persistent infection. Additional one- or three-day PZQ treatment (1 × 60 mg/kg and 3 × 60 mg/kg) or artemether did not result in cure, but over time three participants self-cured. Antibody, cellular, and cytokine responses peaked at week 4 post infection, with a mixed Th1, Th2, and regulatory profile. Cellular responses were (most) discriminative for symptoms. INTERPRETATION: Female-only infections exhibit similar clinical and immunological profiles as male-only infections but are more resistant to PZQ treatment. This limits future use of this model and may have important implications for disease control programs. FUNDING: European Union's Horizon 2020 (grant no. 81564).

Excretion patterns of Schistosoma mansoni antigens CCA and CAA by adult male and female worms, using a mouse model and ex vivo parasite cultures.

Assays which enable the detection of schistosome gut-associated circulating anodic (CAA) and cathodic (CCA) antigen in serum or urine are increasingly used as a diagnostic tool for schistosome infection. However, little is known about the production and clearance of these circulating antigens in relation to the sex and reproductive maturity of the parasite. Here we describe CAA and CCA excretion patterns by exploring a mouse model after exposure to 36 male-only, female-only and mixed (male/female) Schistosoma mansoni cercariae. We found that serum and urine CAA levels, analysed at 3 weeks intervals, peaked at 6 weeks post-infection. Worms recovered after perfusion at 14 weeks were cultured ex vivo. Male parasites excreted more circulating antigens than females, in the mouse model as well as ex vivo. In mixed infections (supporting egg production), serum CAA levels correlated to the number of recovered worms, whereas faecal egg counts or Schistosoma DNA in stool did not. No viable eggs and no inflammation were seen in the livers from mice infected with female worms only. Ex vivo, CAA levels were higher than CCA levels. Our study confirms that CAA levels reflect worm burden and allows detection of low-level single-sex infections.