Schistosoma Group Project Medical Parasitology PDF

Seminars in Immunopathology (2020) 42:355–371 https://doi.org/10.1007/s00281-020-00789-x REVIEW Schistosomiasis—from immunopathology to vaccines Donald P. McManus 1 & Robert Bergquist 2 & Pengfei Cai 1 & Shiwanthi Ranasinghe 1 & Biniam Mathewos Tebeje 1 & Hong You 1 Received: 4 S...

Seminars in Immunopathology (2020) 42:355–371 https://doi.org/10.1007/s00281-020-00789-x REVIEW Schistosomiasis—from immunopathology to vaccines Donald P. McManus 1 & Robert Bergquist 2 & Pengfei Cai 1 & Shiwanthi Ranasinghe 1 & Biniam Mathewos Tebeje 1 & Hong You 1 Received: 4 September 2019 / Accepted: 5 February 2020 / Published online: 19 February 2020 # Springer-Verlag GmbH Germany, part of Springer Nature 2020, corrected publication 2020 Abstract Schistosomiasis (bilharzia) is a neglected tropical disease caused by trematode worms of the genus Schistosoma. The transmis- sion cycle involves human (or other mammalian) water contact with surface water contaminated by faeces or urine, as well as specific freshwater snails acting as intermediate hosts. The main disease-causing species are S. haematobium, S. mansoni and S. japonicum. According to the World Health Organisation, over 250 million people are infected worldwide, leading to consid- erable morbidity and the estimated loss of 1.9 million disability-adjusted life years (DALYs), a likely underestimated figure. Schistosomiasis is characterised by focal epidemiology and an over-dispersed population distribution, with higher infection rates in children. Complex immune mechanisms lead to the slow acquisition of immune resistance, but innate factors also play a part. Acute schistosomiasis, a feverish syndrome, is most evident in travellers following a primary infection. Chronic schistosomiasis affects mainly individuals with long-standing infections residing in poor rural areas. Immunopathological reactions against schistosome eggs trapped in host tissues lead to inflammatory and obstructive disease in the urinary system (S. haematobium) or intestinal disease, hepatosplenic inflammation and liver fibrosis (S. mansoni and S. japonicum). An effective drug— praziquantel—is available for treatment but, despite intensive efforts, no schistosomiasis vaccines have yet been accepted for public use. In this review, we briefly introduce the schistosome parasites and the immunopathogenic manifestations resulting from schistosomiasis. We then explore aspects of the immunology and host-parasite interplay in schistosome infections paying special attention to the current status of schistosomiasis vaccine development highlighting the advancement of a new controlled human challenge infection model for testing schistosomiasis vaccines. Keywords Schistosoma. Schistosomiasis. Urogenital schistosomiasis. Gastrointestinal/hepatosplenic schistosomiasis. Schistosomiasis vaccine. Controlled human infection model Introduction subtropics, impacting one billion people, with 250 million infected, in 74 countries [1–4]. The infection is caused by Schistosomiasis, also called bilharzia, is a neglected tropical trematode blood flukes of the genus Schistosoma; these flat- disease of poverty that is widespread in the tropics and worms have a complex life cycle that involves, depending on species, aquatic or amphibious snails as intermediate hosts and mammalian definitive hosts (Fig. 1). The main clinically important species are as follows: S. japonicum, transmitted by This article is a contribution to the special issue on: Immunopathology of the amphibious snail Oncomelania and resulting in intestinal unresolved tropical diseases - Guest Editor: Marcel Tanner and hepatosplenic schistosomiasis in the People’s Republic of China, the Philippines and Indonesia; S. haematobium, the * Donald P. McManus [email protected] most common species, transmitted by Bulinus snails and caus- ing urogenital schistosomiasis in Africa and in some countries 1 of the Arabian peninsula (it has also recently emerged on the Immunology Department, QIMR Berghofer Medical Research Institute, Brisbane, Queensland, Australia French island of Corsica ); and S. mansoni which is trans- 2 mitted by Biomphalaria snails and causes intestinal and he- Ingerod, Brastad, Sweden, formerly UNICEF/UNDP/World Bank/WHO Special Programme for Research and Training in patic disease in Africa, the Arabian peninsula and Latin Tropical Diseases (TDR), World Health Organization, America. S. guineensis and S. intercalatum (both endemic in Geneva, Switzerland Central and West Africa) and S. mekongi (restricted to a short 356 Semin Immunopathol (2020) 42:355–371 Fig. 1 Life cycle of human schistosomes. Adult schistosome worms from the lumen of the blood vessels through the intestinal or bladder parasitise the blood vessels of humans and other vertebrates which act mucosa and are shed in the faeces (S. mansoni and S. japonicum) or as definitive hosts, but their life cycle necessitates a phase of asexual urine (S. haematobium), but as many as 50% of them wind up in multiplication and development within a freshwater snail. Infective adjacent tissues or are flushed into the liver (mainly S. mansoni and larvae (cercariae) are periodically released from the snail, and these S. japonicum) but can in rare cases find more remote organs. Eggs actively seek and penetrate the skin of the human definitive host; in failing to be excreted from the host are trapped in the tissues inducing zoonotic schistosomiasis (caused by S. japonicum), other mammals, inflammatory responses and resulting in pathological lesions that are especially bovines, can also be involved. Following penetration of the ultimately revealed as clinical disease. The life cycle is completed when skin, the cercarial tails drop off in the skin and the parasites transform into the eggs hatch, releasing free-swimming ciliated miracidia which, in turn, schistosomula which enter the venous blood vessels and are transported locate and infect specific freshwater snails (Biomphalaria spp. for to the lungs via the right heart, before reaching the left side of the heart to S. mansoni; Oncomelania spp. for S. japonicum, Bulinus spp. for eventually enter the arterial circulation. The schistosomula migrate to, and S. haematobium). Cercariae are released after two generations of live in, the mesenteric veins of the bowel (S. mansoni, S. japonicum) or primary sporocysts and then daughter sporocysts within the snail. From the pelvic venous plexus (S. haematobium). There they mature into Reference 1: Ross AG, Bartley PB, Sleigh AC, Olds GR, Li Y, Williams female or male adult worms with the females ending up in the male’s GM, McManus DP (2002) Schistosomiasis New Engl J Med 346: 1212– gynaecophoral canal and ultimately producing eggs over a period that in 1220; Massachusetts Medical Society. Reprinted with permission. extreme cases may be as long as 20 years. The eggs are aimed at passing stretch of the Mekong River in southern Lao People’s regional importance [1–3]. Globally, over 250 million individ- Democratic Republic and eastern Cambodia) are of local, uals are infected with Schistosoma spp. with some 780 million Semin Immunopathol (2020) 42:355–371 357 people at risk of infection [2–4]. The disease results in con- of eggs released daily [1–4]. Importantly, morbidity/mortality siderable human morbidity, even leading to mortality, notably associated with S. japonicum infections is exacerbated due to in sub-Saharan Africa where more than 90% of all infections the substantially higher number of eggs, often released in occur. The Global Burden of Disease (GBD) study of 2016 packages, by this species. The three phases of the disease estimated the global burden due to schistosomiasis at 1.9 mil- mentioned above are dictated by the length of infection. lion disability-adjusted life years (DALYs) , although re- These differ with respect to symptoms and clinical conse- ported DALY figures vary considerably with 70 million being quences as well as to the rates of schistosome egg excretion the highest. in urine or faeces. A recently published review covered the main features of schistosome biology and the epidemiology, clinical features, Immediate manifestations pathogenesis, diagnosis, management and control of schisto- somiasis. Here we provide a more in-depth consideration The first clinical manifestation following infection is a of the pathophysiology and progression of clinical disease in maculopapular pruritic eruption, termed cercarial dermatitis, schistosomiasis, the immunology and host-parasite interplay which may arise at the site of skin penetration by schistosome in schistosome infections and recent updates on progress in cercariae. Cercarial dermatitis results from an innate im- schistosomiasis vaccine development. mune response giving rise to hypersensitivity reactions to dead or dying parasites and can be observed with all human schistosome species. These skin reactions comprise discrete Immunopathogenesis and clinical disease erythematous raised lesions varying in size (1–3 cm) which may develop within a few hours post-infection, although a The main lesions in schistosome infection are due not to the rash may appear up to 7 days later in tourists or migrants adult parasites but to the eggs laid by the female worms [1–4]. moving into an area endemic for schistosomiasis for the first Whereas many eggs reach the external environment to contin- time. The resulting dermatitis is similar to, but less severe than ue schistosome transmission, a large proportion is trapped in swimmers’ itch, which is a response in persons when infected the liver, intestine or bladder of the definitive host. Here, they by schistosomes not adapted to humans (e.g. those adapted to secrete various components including proteolytic enzymes birds) [1, 7]. that elicit eosinophilic inflammatory immune responses, in- cluding the formation of granulomas, which are progressively Acute schistosomiasis replaced by fibrotic deposits eventually resulting in urogenital disease (S. haematobium) or, alternatively, intestinal or ad- Following cercarial penetration and schistosomula matu- vanced hepatosplenic schistosomiasis (S. mansoni and ration, the infection may proceed to the symptomatic S. japonicum) [1–4]. acute schistosomiasis stage although the first clinical There are three distinct stages of disease progression in manifestations in non-immune subjects may be delayed schistosomiasis: acute, established active and late chronic by several weeks or even months following exposure [1,. Acute disease occurs mainly in travellers to areas endemic 7]. The symptom complex, known as Katayama fever or for schistosomiasis following a primary schistosome infection Katayama syndrome, is common in areas with high rates with the common presenting symptoms in S. mansoni– and of schistosomiasis transmission and is an early clinical S. japonicum–infected individuals being abdominal and mus- indicator of a first infection or a heavy reinfection with cle (myalgia) pain, fever, malaise and fatigue; haematuria Schistosoma spp. cercariae. Acute schistosomiasis is the (blood in the urine) is characteristic of individuals infected disease manifestation most likely to be underdiagnosed or with S. haematobium [1–4]. Established active and late chron- misdiagnosed by physicians in non-endemic countries and ic schistosomiasis are found mainly in people with long- remains, compared with the other clinical stages of schis- standing schistosome infection who live in poor rural areas. tosomiasis, poorly understood. The syndrome is The clinical manifestations are inflammatory and obstructive named after the Katayama District of Hiroshima prefec- and result from immunopathological reactions against schis- ture in Japan where the first human case of S. japonicum tosome eggs trapped in host tissues [1–4]. In S. japonicum and was described in 1904. Patients commonly recall prior S. mansoni infections, we see intestinal disease by advanced contact with natural freshwater bodies, such as streams or hepatosplenic schistosomiasis, whereas in S. haematobium in- lakes between 2 weeks and 3 months earlier. Common fection, the disease is urogenital mainly involving lesions of symptoms include generalised myalgia, right-upper- not only the bladder wall but also the reproductive system quadrant pain, bloody diarrhoea, headache, fever, eosino- [1–4]. philia, non-productive cough and patchy pulmonary infil- Morbidity is especially pronounced in high-intensity infec- trates observed on chest radiography [3, 7]. These vary in tions (i.e. those with a high worm burden) with large numbers severity depending on the infecting species but are 358 Semin Immunopathol (2020) 42:355–371 thought to be due to systemic hypersensitivity reactions antigenic glycoproteins (that generate an inflammatory re- mediated by the immune system against antigens released sponse mediated by CD4+ T-helper-2 lymphocytes), the func- by migrating schistosomula or, in most cases, eggs fol- tion of which is to help facilitate their passage from the blood lowing their deposition in host tissues [3, 7]. Whereas vessels, where they are released from the female worm, to the the majority of patients recover spontaneously after 2– lumen of the urinary bladder or intestine, to eventually close 10 weeks, some develop a generalised rash and more se- the life cycle by reaching and infecting the intermediate snail rious and persistent disease with dyspnoea, abdominal host. However, these glycoproteins also induce the formation pain, weight loss, hepatomegaly and diarrhoea [3, 7]. of granulomas (an assembly of inflammatory cells such as While Katayama fever caused by S. japonicum infections is macrophages, lymphocytes, neutrophils and eosinophils) that commonly reported, it is rarely observed in relation to amass around the eggs. The granulomas generally develop at S. haematobium or S. mansoni in individuals from chronically the sites of maximal egg accumulation and, while killing the exposed communities and may be the result of in-utero de- eggs, the end result is fibrotic lesions in the host tissues [1–4]. sensitisation and a lowering of immune responsiveness to The organ-specific clinical symptoms in schistosomi- schistosome antigens in infants born to infected mothers [4, asis often correlate positively with the intensity of in- 7]. An alternative explanation might be repeated exposure to fection (i.e. the number of eggs excreted) and are me- schistosome cercarial antigens inducing CD4+ T cells that diated by egg-induced inflammation followed by the produce IL-10 in the skin leading to downregulation of the granulomatous reaction [1–4]. Established active infec- immune response ; however, it is also possible that cases tions, characteristic of children in schistosomiasis- from endemic areas simply remain undiagnosed. Katayama endemic areas, are partially reversible following killing fever caused by S. japonicum infections not only is not re- of the adult worms through repeated treatment with stricted to a primary infection but also occurs in people living PZQ or ART + PZQ killing both young and adult in areas with a history of previous infections, notably in the worms. As the disease becomes chronic, the elevat- People’s Republic of China where such infections have been ed lymphocyte proliferative responses generated by the recorded in endemic communities exposed to flooding [6, 7]. soluble egg antigens at this phase of infection progres- Both repeated doses of 60 mg per kg body weight of the sively decrease [2, 4]. acylated quinoline-pyrazine compound, praziquantel (PZQ), and a prolonged daily course at 20 mg per kg body weight have been used as treatment. The anti-malarial artemether Chronic and advanced disease (ART) (and other artemisinin derivatives, produced from leaves of the Chinese medicinal plant Artemisia annua) Mature, patent schistosome infections, associated with chron- should kill all invading schistosomula if it is given every ic local inflammatory responses to schistosome eggs trapped 2 weeks, thereby providing an avenue for preventing acute in host tissues, may in urogenital infection (caused by cases in high-risk groups such as tourists, fishermen and flood S. haematobium) lead to obstructive disease in the urinary relief workers in areas endemic for schistosomiasis [6, 9]. and reproductive system or, alternatively, intestinal disease, hepatosplenic inflammation and liver fibrosis in disease due Established active infection to S. mansoni and S. japonicum [1, 2, 10]. Granulomas, which develop at the sites of maximal accumulation of eggs, destroy In the majority of schistosomiasis cases, particularly in indi- the eggs but result in fibrosis in host tissues. Granuloma for- viduals residing in endemic areas, symptomatic, acute disease mation and the local inflammatory response mediated by does not occur; instead, the disease develops directly into an CD4+ T-helper-2 lymphocytes help in facilitating the passage established active infection characterised by the presence of of eggs into the lumen of the gut or urinary tract prior to their mature adult worms, the production of eggs and their excre- exit in faeces or urine [11–13]. However, the worm burden in tion in urine or faeces. Live adults present in blood vessels most individuals exposed continuously to schistosome infec- do not generate an immune response with local inflammation, tion gradually declines over time as partial acquired immunity which is believed to be due to the worms’ ability to masquer- develops against new infections; concomitantly, due to natural ade by acquiring host antigens or through molecular mimicry death of parasites, there is a reduction in established worm whereby the parasite antigens are hidden and possibly also numbers which results in smaller numbers of new eggs being resulting from the continuous regeneration of the parasite lodged in host tissue or excreted [4, 10]. In addition, owing to tegumental external surface effectuated by unique somatic immunological down-regulation, newly formed granulomas stem cells. Schistosome eggs, on the other hand, are unpro- are smaller, while as eggs within them are killed, any earlier tected, so all lesions in affected host organs and the resulting formed granulomas gradually resolve to be replaced by scar- symptoms are entirely due to the inflammatory responses gen- ring (fibrous tissue), thereby contributing to a reduced severity erated against them (Fig. 2). The eggs actively secrete soluble of symptoms. Semin Immunopathol (2020) 42:355–371 359 Fig. 2 Features of schistosome-induced granuloma formation. Adult neutrophils. Chemokine-binding proteins secreted by the eggs of schistosome worm pairs residing in mesenteric veins produce eggs S. mansoni eggs bind neutrophil chemoattractant C-X-C-motif some of which become entrapped in the host’s liver (or other organs) chemokine ligand 8 (CXCL8), thereby blocking the infiltration of tissue where they evoke a dominant CD4(+) TH2 immune response neutrophils to the granuloma. In contrast, these proteins do not bind to mediated by IL-4 and IL-13. This leads to the development of eosinophil chemoattractant CC-chemokine ligand 11 (CCL11) and, granulomas and fibrosis with hepatic stellate cells, macrophages, therefore, do not inhibit the recruitment of eosinophils. b A granuloma lymphocytes, neutrophils, and eosinophils, all identified as major in the liver of a S. mansoni-infected mouse with hepatic stellate cells cellular contributors to these events. a Major cellular populations (HSCs). Part a adapted with permission from Chuah, C., Jones, M. K., located within and adjacent to the hepatic granuloma induced in either Burke, M. L., McManus, D. P. & Gobert, G. N. Cellular and chemokine- S. japonicum or S. mansoni infection. Whereas a dense population of mediated regulation in schistosome-induced hepatic pathology. Trends eosinophils are present at the core of a S. mansoni-induced hepatic Parasitol. 30, 141–150 (2014), Elsevier. Part b courtesy of A. M. O. granuloma, the core in a S. japonicum infection is comprised chiefly of Kildemoes, University of Copenhagen, Denmark Urogenital schistosomiasis important clinical complication of infection; indeed, S. haematobium has been classified as a carcinogenic infection Adult S. haematobium worms settle and reside in the small by the International Agency for Research on Cancer [4, 19, 20]. venules around the ureter and bladder of the urinary tract. The process of carcinogenesis is closely associated with tissue Many eggs are trapped in the tissues, mainly the bladder wall inflammation and well differentiated squamous cell carcinomas where they eventually become calcified, while those that pass that metastasise locally. Inflammatory gene damage, β- into the bladder lumen leave the body with the urine. In an glucuronidase and nitrosamines are possible factors contribut- established active infection, live egg clusters in the urogenital ing to the process of carcinogenesis although S. haematobium tissues are found surrounded by pronounced tissue eosinophilia lesions may intensify the exposure of the bladder epithelium to and an intense inflammatory reaction [14, 15], while passage mutagenic substrates such as tobacco. through the bladder wall is commonly accompanied by In the genital system, parasite eggs are frequently deposited sloughing off of the epithelial surface followed by ulceration in the prostate and the seminal vesicles (in men) or in the and bleeding. Intense egg-induced tissue inflammation results cervix (in women), where characteristic cervical lesions are in thickening of the bladder wall and development of found with pronounced tissue inflammation with calcified pseudopolyps and hypertrophic masses [16, 17]. Haematuria, (in the chronic stage) as well as live eggs in relatively new appearing 10–12 weeks after infection, is the first sign of disease infections. Infected females may present with other genital due to an established active S. haematobium infection and many lesions (e.g. vulvar nodules), vaginal bleeding, fallopian tube children from endemic areas present with this symptom. damage and dyspareunia. Men with genital schistosomiasis Along with dysuria, haematuria also occurs in the late disease have increased levels of inflammatory cytokines and in- phase. Late disease manifestations additionally include ureter creased numbers of leucocytes in semen. In both men and urethra obstruction, bladder calcification, secondary bacte- and women, S. haematobium infection has been correlated rial infections in the urinary tract, proteinuria (often nephrotic with increased risk of infection by, and transmission of, the syndrome), hydronephrosis, renal colic and renal failure. human immunodeficiency virus (HIV) with attraction of in- Epidemiological studies have found an association of chron- flammatory cells and possibly presence of the virus in semen. ic, urogenital schistosomiasis with squamous cell carcinoma of In women, intercourse can cause bleeding lesions on the fria- the bladder in Egypt and other parts of Africa. This is an ble mucosa of the cervix and the vagina. 360 Semin Immunopathol (2020) 42:355–371 Gastrointestinal and hepatosplenic schistosomiasis S. mansoni and (rarely) S. haematobium, but not S. japonicum, can lead to pulmonary schistosomiasis. In Schistosome eggs retained in the gut wall induce inflamma- this manifestation, eggs have been transported to the lung tion, polyposis, ulceration, micro-abscess formation and hy- capillaries where they induce granuloma formation which perplasia [1, 3, 4]. Symptoms of gastrointestinal disease tend may lead to fibrosis in the perialveolar area resulting in pul- to be more pronounced in individuals with high infection in- monary hypertension and cor pulmonale (increased size of the tensities and include loss of appetite, diarrhoea (particularly in right heart ventricle due to elevated pressure in the lung children) that may alternate with constipation, blood in stool capillaries). (haematochezia) and colicky hypogastric pain or pain in the left iliac fossa [1, 2, 4]. Severe chronic intestinal disease may result in rectal or colonic stenosis. Colonic polyposis may Immunology and host-parasite interplay manifest as a protein-losing enteropathy, whereas an inflam- in schistosome infections matory mass in the colon may even mimic cancer. When eggs of S. japonicum and S. mansoni are flushed to Animal models the liver (which they commonly are), the granulomatous in- flammatory response induces pre-sinusoidal inflammation Experimental animal models provide invaluable information and periportal (clay-pipe) stem fibrosis [1, 2, 4]; sustained on the immunobiology of schistosomiasis, including the na- heavy infection is frequently associated with this clinical pic- ture of the host innate and adaptive responses to schistosomes ture, but it normally takes many years to develop [1, 2, 4]. and the strategies implemented to manipulate these responses Hepatomegaly occurs early in the evolution of chronic schis-. Indeed, much of our understanding of the human im- tosomiasis, while late-stage periportal collagen deposits lead mune responses to schistosomes has been facilitated by the to the progressive obstruction of the blood flow, marked portal availability of murine experimental S. japonicum and hypertension, which results in the development of alternative S. mansoni infection models [28–30] which have provided routes of the blood circulation (commonly causing varices invaluable help in the design of immunological studies in inside the oesophagus, often leading to bleeding and vomiting people with schistosomiasis. Briefly, these studies show that of blood). Other common clinical signs are abdominal ascites T cell-mediated immunity is pivotal in the development of and splenomegaly (enlargement and hardening of the spleen) acquired resistance to schistosomes in mice and that the pro- [1, 2, 4, 21]. Periportal fibrosis, characteristic of schistosomi- cesses of immune regulation and T cell regulation involved asis, can be seen on magnetic resonance imaging (MRI), com- are complex. Whereas the T-helper (TH) response typically puted tomography scan and ultrasonography [1, 21]. induced by these blood flukes is a pronounced TH2 one, it is the development of a balanced TH reaction that is critical in Other disease manifestations preventing disease progression since both TH1 and TH2 com- ponents, if one-sided and excessive, can lead to damaging The most severe clinical outcome associated with schistosome pathology. infection is probably neuroschistosomiasis, which results In recent years, the use of transgenic or gene knockout mice from the inflammatory response against eggs deposited in has considerably advanced our understanding of the immuno- the brain or spinal cord following the aberrant migration of pathological mechanisms and the process of granuloma for- adult worms to these locations [23–25]. Common signs and mation that are important in both schistosomiasis and other symptoms of neuroschistosomiasis are radiculopathy, an in- fibrotic diseases. However, whereas mouse models of crease in intracranial pressure, myelopathy and subsequent schistosomiasis have proven useful in identifying factors in- clinical sequelae [23–25]. Acute encephalitis of the cortex, volved in granuloma formation and in immunopathology, be- subcortical white matter, basal ganglia or internal capsule is ing naive hosts that do not live sufficiently long to develop typical of S. japonicum infection, whereas myelopathy (acute protective immune responses after primary infection, they transverse myelitis and subacute myeloradiculopathy) of the cannot be used for analysing immune correlates to reinfection lumbosacral region is the most common neurological mani- resistance; consequently, much of what is known about these festation of S. mansoni or S. haematobium infection. host–parasite interactions derives from human epidemiologi- Cerebral complications include encephalopathy with head- cal studies considered in detail below. ache, visual impairment, delirium, seizures, motor deficits and ataxia, whereas spinal symptoms include lumbar pain, Human studies lower limb radicular pain, muscle weakness, sensory loss and bladder dysfunction. The interaction between schistosomes and the host immune Schistosome-induced lesions in the heart and appendix system in the parasitised mammal is highly complex involving have been documented and chronic infection with a number of discrete phases (characterised by penetrating Semin Immunopathol (2020) 42:355–371 361 cercariae, migrating schistosomula, adult worms, eggs excret- spectrum in endemic settings can only be achieved by long- ed from the host or eggs entrapped in tissues). Immune re- term protection involving vaccination [2, 34]. Indeed, the in- sponses against egg antigens are responsible, as indicated ear- ability of PZQ to interfere prominently in transmission of the lier, for the pathology in schistosomiasis, whereas responses infection has led to a renewed interest in vaccine development. towards cercariae, and possibly some of the other stages, no- tably schistosomula, are likely important for the development of resistance to re-infection. Protective immune responses in Schistosomiasis vaccine development schistosomiasis do develop in people in Schistosoma-endemic areas but slowly (over a period of 10 to 15 years) [2, 4]. Why is a vaccine needed? Accordingly, with adults usually resistant to reinfection fol- lowing treatment, children less than 10 years old are highly Extensive efforts have been made in schistosomiasis control susceptible which provides an explanation for the age-preva- mainly as the result of population-based preventive chemo- lence/age-intensity curves typically recorded for individuals in therapy delivered through mass drug administration (MDA), areas endemic for schistosomiasis. i.e. the periodic oral PZQ administration, targeting mainly Immune correlate studies in different geographical loca- school-aged children at risk without prior diagnosis. tions and a variety of epidemiological settings suggest that Indeed, the global reduction in schistosomiasis morbidity acquired anti-schistosome protective immunity after curative resulting from this sustained PZQ treatment led to a strong drug therapy is mediated (although not exclusively) by TH2 emphasis (perhaps overly so) on chemotherapy as the exclu- cell–associated responses characterised by eosinophilia and sive method to achieve elimination of the disease. production of cytokines such as IL-4 and IL-5 as well as However, this progress notwithstanding, schistosomiasis schistosome-specific IgE. High levels of IgG4 are also pro- prevalence has remained largely unaffected as rapid reinfec- duced during infection, potentially blocking the protective ef- tion quickly restores the prevailing levels of infection after fects of other immunoglobulins at a degree associated with each scheduled MDA programme. Notable also is that susceptibility to infection. Indeed, the age-dependent con- PZQ is not 100% effective, and particularly, since it is given as comitant immunity against reinfection resulting from repeated a single dose in each treatment cycle, the untouched migrating natural adult worm death over time leads to establishment of schistosomula give rise to a new generation of adult worms 1– protective immunity over several years and that this is more 2 months after MDA. Furthermore, due to the widespread closely related to the IgE/IgG4 balance than to the absolute use of the drug, the potential emergence of PZQ drug resistance level of each isotype. It has been hypothesised that the is a continual concern, although there is, as yet, no direct killing of adult worms, either after PZQ treatment or naturally, definitive evidence that clinically relevant drug-resistant leads to release of antigens which may cross-react with larval schistosomes have evolved in the field [4, 35]. antigens and stimulate protective IgE responses, implying that In only a small number of schistosomiasis-endemic areas, the more dead worms an infected individual has experienced notably the People’s Republic of China where elimination is the more intense the generated protective responses. As now on the horizon, has a clear reduction in prevalence been schistosome worms generally live for 3–10 years, this hypoth- reached. This effect can, however, be credited not only to the esis could provide an explanation, at least in part, why chil- use of PZQ but also as much to effective snail control and dren, who have experienced few dying worms are susceptible other measures such as environmental modification and health to infection and accumulate more worms through continuous education. It must in this connection be mentioned that exposure, whereas adults who have experienced many dying the snail species acting as intermediate hosts outside Southeast worms develop protective immune responses; it may also ex- Asia are non-amphibious and, therefore, considerably more plain why anti-schistosome immunity in human populations difficult to control. There is therefore now an almost takes such a long time to develop [2, 4]. general acceptance that sustainable schistosomiasis preven- It is important to stress that the deployment of a vaccine for tion and control will require a multifaceted, integrated ap- schistosomiasis dependent on IgE would likely be problemat- proach, a scenario that underscores support for the develop- ic and hampered by regulatory and safety issues since this ment and deployment of a vaccine as a key element in the line class of antibodies is associated with allergic responses [3, 4, of attack making up for the short-term effect of chemotherapy 33, 34]. Therefore, looking to the immune responses of chron-. Indeed, Science ranks a schistosomiasis-vaccine as one ically infected individuals, and even those who become refrac- of the top 10 vaccines requiring urgent development based on tory by producing IgE after PZQ drug treatment, should be feasibility and need. Accordingly, the development and approached with care. Nevertheless, although vaccines positioning of an effective vaccine within the spectrum of do not currently figure prominently in the context of schisto- disease control is an option that would add a much needed somiasis control, it is imperative that research continues in this long-term attribute to be applied in combination with MDA important area as genuine and lasting change of the disease [34–36, 38]. Furthermore, the schistosomiasis vaccine does 362 Semin Immunopathol (2020) 42:355–371 not need to be 100% effective as mathematical modelling important defensive mechanisms might differ, whereas human provides support to the view that even a partially protective population studies in endemic areas cannot provide all the vaccine would contribute to reducing schistosome infections important information needed. It has thus been advocated and interrupting endemic transmission [34, 36, 39, 40]. that it is useful to also carry out testing of schistosome vaccine The sad reality that no commercial vaccine is available antigen candidates in non-human primates, even if only to currently against any of the human schistosomes emphasises strengthen the safety aspect of the vaccination strategies. the need for continued efforts towards achieving this elusive Furthermore, vaccines based on studies performed only in goal. Researchers in the field have met with an uphill battle for the mouse model could even have undesirable effects if taken many years, partly due to the scarce funding in face of the prematurely to human clinical trials [46, 47]. It has even been complicated immunology involved when the human host bal- speculated that schistosome vaccine/challenge trials undertak- ances between resisting new infections and avoiding serious en in mice might be flawed, resulting in erroneous protective pathology when reacting against entrapped tissue eggs from efficacy data. Nevertheless, immunisation of mice with a prior infections. In addition, over the course of evolution, single dose of irradiated cercariae , or schistosomula treat- schistosomes have evolved intricate survival strategies en- ed with artemether , induces high levels of protection abling successful relocation between various, completely dif- [49–51]. Although it is doubtful that such approaches could ferent environments to allow them access to the human host be used clinically as they would likely carry too high a risk of [41, 42]. In retrospect, one cannot avoid the sobering thought adverse events, these pioneering findings motivated the search that it was unfortunate that the initially adequate allocation of for the key antigens simulating these responses. For example, funding for schistosomiasis vaccine development was not an early study of mice vaccinated with S. mansoni irradiated sustained. As donors dropped out, the dwindling levels of cercariae identified a group of five such molecules, i.e. triose funding restricted the possibility of rapidly reaching the goal, phosphate isomerase (TPI), glutathione S-transferase (GST), making the development of an effective product even more heat shock protein 70 (HSP-70), paramyosin and a 23-kDa challenging. There is, however, some light at the end of integral membrane protein. The search for defined schis- the tunnel provided by the fact schistosomes do not replicate tosomiasis vaccine candidates of this kind intensified thereaf- in their definitive hosts provides some respite in contrast to ter, resulting in a large suite of putatively protective antigens viral and bacterial infections. Thus, even a partially protective. Recent data on S. mansoni and S. haematobium vaccine vaccine could contribute to reducing infection rates and candidates including their location, function, formulation and interrupting transmission, a view indicated earlier supported mode of delivery, as well as the adjuvants employed and the by mathematical modelling [34, 36, 39, 40]. In addition, re- protective efficacy generated, are presented in Table 1. Similar cent progress has shown some promise, not only due to the data are available for S. japonicum. availability of novel adjuvants that can selectively manipulate Generally, the work followed a step-by-step approach that the immune responses, but also because immunological re- included identification of protective antigens, selection of the search makes it possible to assess the specific responses each most promising ones, their molecular cloning, expression and vaccine antigen elicits (and what it needs to elicit) through cell purification followed by testing for vaccine efficacy in vacci- signalling studies. It has been argued that immune re- nated and challenged mice. In the next section, we con- sponses directed against the adult schistosome worm might sider the evaluation of the relatively few schistosomiasis vac- not be the major mechanism sought. If this assumption cines that have advanced to clinical trials or have the potential is correct, the short interval between cercarial skin penetration to enter into clinical testing. and the presence of schistosomula in the lungs about 72 h later would be the time when the parasite should be the most vul- nerable for immune attack. This susceptibility is likely to Human vaccine development at least partly spill over into the juvenile schistosome in the liver sinusoids after which time the parasite is already resistant Three lead human schistosomiasis vaccines (Sh28GST, Sm- to antibody-dependent cellular cytotoxicity (ADCC) con- 14 and Sm-TSP-2) are currently at differing phases of clinical trolled by complement. development with a fourth (Sm-P80) to follow shortly. Animal models and vaccine development Sh28GST Animal experiments, particularly using the mouse challenge Recombinant 28-kDa glutathione S-transferase of model, carried out over the past 50 years have provided a rich S. haematobium (rSh28GST), presented with Alhydrogel (al- appreciation of Schistosoma spp.–host interactions. uminium hydroxide) and designated as Bilhvax, was devel- However, although the protective immune responses to schis- oped in partnership by Professor Andre Capron and his team tosome infection might be similar in humans and mice, at Inserm-Université Lille and Institut Pasteur de Lille in Table 1 Recent data on Schistosoma mansoni vaccine antigen candidates (modified from ) Antigen Location in Identity/function Immunisation Adjuvant Host Worm burden Liver egg adult worm strategy reduction (%) burden reduction (%) Sm-p80 Associated with tegument inner Calpain-neutral Recombinant protein Resiquimod Mouse 50 16 membrane cysteine protease Primed with pcDNA3 and Resiquimod Mouse 49 30 boosted with recombinant protein Primed and boosted with Oligodeoxynucleotide Mouse 70 75 recombinant protein Recombinant protein Resiquimod Baboon 58 – DNA vaccine – Baboon 38–46 32–28 Semin Immunopathol (2020) 42:355–371 Fatty acid binding protein Whole body, cytosolic Absorbs, transports and Recombinant protein – Mouse 67 – (FABP) (Sm14) compartmentalises fatty acids from the host Tetraspanin protein 2 Tegument apical membrane Tetraspanin integral Recombinant protein Freunds Mouse 57 64 (Sm-TSP2) membrane protein Recombinant protein Alum/CpG Mouse 25 27 Glutathione S-transferase (Sh28GST) Whole body Enzyme involved in Recombinant protein Aluminium Baboon 0–80 – fatty acid metabolism and prostaglandin D2 synthesis Sm29 Tegument apical membrane Unknown, but has a C-terminal DNA vaccine with pUMVC3 – Mouse 17–22 – domain plasmid Recombinant protein Complete Freunds and Mouse 51 – incomplete Freunds Sm14 + Sm-29 – – Multivalent recombinant proteins Poly (I;C) Mouse 40 68 Sm29 + Sm-TSP-2 – – Multivalent DNA vaccine with – Mouse 24–32 – pUMVC3 plasmid Multivalent Recombinant proteins CpG-Alum Mouse 35 – Oesophageal gland secretion Oesophagus Digestive tract proteins Recombinant proteins Freunds Mouse 25–32 33–44 (Sm100.3) (oesophageal) Cathepsin B1(SmCB1) Gut (gastrodermis) Gut protease (cysteine Recombinant proteins Postulated to have Mouse 73 83 (Combined peptidase) inbuilt with SG3PDHa + PRX- adjuvant properties MAPb) S. mansoni cathepsin B Gut (gastrodermis) Gut protease (cysteine Recombinant proteins CpG Mouse 59 56 (Sm-CB) peptidase) oligodeoxynucleot- ides Montanide ISA 750 Mouse 60 62 VG Schistosome cysteine proteinase, Gut Gut protease (Asparaginyl DNA vaccine – Mouse No significant 37 asparaginyl endopeptidase peptidase) reduction (SmAE) (Sm32) Lysosome-associated membrane Gastrodermis Processing of ingested blood Recombinant protein alum-CpG Mouse 16–25 – glycoprotein (Sm-LAMP) Dynein light chain proteins Unknown Evolutionarily conserved among Recombinant protein Alhydrogel Mouse different organisms -DLC 12 43 -DLC 13 51 363 364 Semin Immunopathol (2020) 42:355–371 reduction (%) France together with scientists from Eurogentec, Liège, Liver egg Belgium. The vaccine was shown to be safe and burden immunogenic, generating a TH2-type response in young 73 – healthy Caucasian male adults in a phase-1 randomised, clin- ical study; this encouraging outcome provided the impetus to Worm burden reduction (%) continue trials with rSh28GST as a potential vaccine candidate against urogenital schistosomiasis (with a possibly useful de- Complete Freunds and Mouse 30–37 Mouse 44–60 Mouse 22–45 Mouse 23–55 gree of cross reaction against S. mansoni). Phase-2 clin- Mouse 43 Baboon 17 ical testing in Senegal showed that Bilhvax in combination with PZQ treatment was safe for infected adults and children Host (unpublished). A phase-3 trial (2009 to 2012) in 250 children incomplete Freunds (125 randomly assigned to receive the vaccine and 125 the placebo) aged 6–9 years in Senegal showed the Bilhvax vac- cine to be immunogenic and well tolerated by all subjects Adjuvant investigated but, disappointingly, producing an insufficient level of protection, i.e. there was no significant delay in schis- tosomiasis recurrence between the vaccinated and the placebo – – – – groups as the primary endpoint. Modifying the trial de- sign and/or utilising an alternative adjuvant to provide a more balanced isotypic response, while at the same time reducing the number of vaccine administrations, may improve the effi- Recombinant protein cacy of the rSh28GST vaccine and encourage further clinical Immunisation testing. UV-attenuated DNA vaccine DNA vaccine DNA vaccine strategy Sm14 Dr. Miriam Tendler and her team in Rio de Janeiro, Brazil pioneered over many years the progression to clinical trials Scaffold supporting protein of a recombinant 14 kDa (rSm14) S. mansoni fatty acid- binding protein (FABP) vaccine candidate. The antigen was Identity/function developed under a Brazilian platform led by the Oswaldo Cruz Foundation, the Health Ministry in Brazil. Like the S. mansoni homologue of Sh28GST (Sm28GST), Sm14 was a member of the original group of potential S. mansoni vac- – – – – cine antigens, promoted by WHO/TDR in the late 1980s. The safety and immunogenicity of the rSm14 vaccine were evalu- ated in 2011 through an open, non-placebo-controlled, dose- standardised phase-1a trial, performed at a single site [56, 57]. Expressed in Pichia pastoris, the rSm14 vaccine was formu- lated with glucopyranosyl lipid A (GLA) adjuvant produced Intestinal tract adult worm Location in by the International Disease Research Institute (IDRI) Glyceraldehyde 3-phosphate dehydrogenase (Seattle, WA, USA) in an oil-in-water emulsion (SE). Tested in 20 male volunteers from a non-endemic area for schistoso- – – -Signal peptide-containing superoxide – – miasis in the state of Rio de Janeiro, Brazil, no serious adverse events were observed. Although the vaccine was shown to be -Glutathione peroxidase enzymes immunogenic, no specific IgE response was generated, a S. mansoni Syntenin (SmSynt) Radiation-attenuated cercariae -Cu/Zn cytosolic superoxide major advantage of the product. A phase-1b clinical study, to evaluate the safety and immunogenicity of the rSm14 vac- Table 1 (continued) cine in 10 healthy female volunteers, was successfully con- Peroxiredoxin cluded in 2012 , paving the way for a phase-2a clinical Antioxidants dismutase dismutase trial undertaken in 2015–2017. This trial was undertaken Antigen in 30 adult males living in a high-endemic area for both S. mansoni and S. haematobium in the Senegal River Basin b a Semin Immunopathol (2020) 42:355–371 365 where the safety and strong long-lasting immunogenicity of japonicum and S. haematobium [47, 66] Additionally, Sm- the rSm14 vaccine were confirmed. Based on these outcomes p80-specific IgE was not detected in infected populations in adults, a phase-2b trial design and protocol were defined in from Africa and South America, thus potentially minimizing 2018 involving 95 Senegalese school children, aged 7– the risk of a hypersensitivity reaction following vaccination 11 years and living in the same area endemic for both schis-. Furthermore, Sm-p80 was shown to have a therapeutic tosome species. Subsequent phase-2c and phase-2d (in Brazil) effect in vaccinated baboons through suppression of the num- and phase-3 (in Senegal) clinical trials are planned. bers of established worms, reducing the retention of eggs in tissues and decreasing the number of eggs excreted by the host Sm-TSP-2 in faeces. It is anticipated that the recombinant Sm-p80/ GLA-SE vaccine, “SchistoShield” will soon move forward to Highly abundant at the outer-most tegumental membrane of phase-1 and phase-2 human clinical trials. the intra-mammalian stages of schistosomes, the tetraspanins constitute a group of proteins continually exposed to the host immune system. Required for tegument biogenesis and A transmission blocking vaccine integrity, Sm-TSP-1 and Sm-TSP-2 constitute the major for schistosomiasis japonica S. mansoni tetraspanins [60, 61]. Furthermore, Sm-TSP-2 confers a high level of protection in the mouse vaccine model Schistosomiasis control in the People’s Republic of China and with the corresponding IgG antibodies correlating positively the Philippines is complicated by the zoonotic nature of the with protective immunity in naturally resistant people [62, disease with bovines (water buffalos and cattle) acting as ma- 63]. As a result, Sm-TSP-2 was selected for further process jor reservoir hosts for S. japonicum being responsible for up to development as a prime schistosomiasis vaccine candidate 90% of environmental contamination of parasite eggs. and is currently being advanced by the Sabin Vaccine From the point of view of vaccine development, this permits Institute Product Development Partnership as a 9-kDa recom- a step-wise tactic that would start with the use of a “transmis- binant Sm-TSP-2/Alhydrogel® vaccine in combination with sion-blocking” veterinary vaccine before moving on to a prod- GLA-SE as adjuvant. The vaccine candidate can be readily uct aimed at human vaccination should that be necessary. In scaled up in the Pichia pink (yeast) system, has undergone light of this, vaccination of animals, particularly bovines, is required toxicology studies and shown good pre-clinical re- considered useful with respect to Asian schistosomiasis as it sults, while several clinical trials of the vaccine have been would assist in long-term prevention of human (and animal) completed or are in progress in non-endemic and endemic infection, a concept supported by mathematical modelling communities. These include an initial phase-1 study test- [17, 39, 68, 69]. ing its safety and immunogenicity undertaken at the Baylor Vaccination can reduce egg excretion from buffalo and College of Medicine, Houston, TX, USA, a phase-1b trial cattle, thereby interrupting transmission from bovines to underway in an endemic area of Brazil to assess the vaccine’s snails, and its use would be particularly applicable in areas safety and immunogenicity in a group of healthy adults who deemed unsuitable for the replacement of bovines by may have previously been exposed to schistosomiasis, with mechanised farming, one of the interventions featuring in further field trials in Uganda planned. the current integrated schistosomiasis control strategy in the People’s Republic of China. Indeed, the implementation Sm-p80 of an animal-based transmission-blocking vaccine as part of a package of integrated control measures sits well with the One A second S. mansoni surface protein, Sm-p80 (the large Health concept synergizing human and animal health, a con- calcium-activated neutral cysteine protease subunit of clusion reached at two workshops on schistosomiasis elimina- calpain), expressed in all schistosome life cycle stages and tion strategies and the potential role of a vaccine co-sponsored likely to play critical role in the worm’s tegumental biogenesis by the National Institute of Allergy and Infectious Diseases and renewal , is another molecule rapidly moving towards (NIAID) and the Bill & Melinda Gates Foundation [34, 71]. In clinical testing under the auspices of a consortium led by Dr. the discussion at one of these workshops on the question of a Afzal A Siddiqui, Texas Tech University Health Sciences clinical versus a veterinary vaccine, the latter was seen as the Center at Lubbock, TX, USA. Sm-p80 has been tested for preferred target product profile (TPP) with respect to schisto- its vaccine efficacy using different vaccine formulations and somiasis japonica as the less rigorous safety concerns required approaches, including naked DNA, recombinant protein and would permit a more rapid implementation [34, 71]. Whereas prime/boost in three experimental animal models of infection this characteristic could potentially reduce the costs of and disease (mouse, hamster and baboon) and shown to deploying a schistosomiasis vaccine for use in livestock com- generate significant protection against S. mansoni challenge pared with one for human use, a comparative cost–benefit infections in addition to cross-species protection against S. evaluation of applying the two different types of vaccine 366 Semin Immunopathol (2020) 42:355–371 needs to be undertaken. A number of S. japonicum SjTPI transmission-blocking vaccine candidates have been identi- fied [33, 34, 53, 69, 71]) with Sj-GST26, Sj-GST28, Sj-97 Triose-phosphate isomerase (TPI) is another lead anti- (paramyosin) and Sj-TPI (triose-phosphate isomerase) the S. japonicum vaccine candidate; this compound is located on most tested (see Table 1). the surface membranes of newly transformed schistosomula and is present in most cells of adult schistosomes. TPI has The Sj-GSTs been tested in mice, pigs and bovines in various vaccine for- mulations [69, 71], such as with a plasmid-generated (Chinese Considerable early work showed the most repeatable host strain of S. japonicum) DNA vaccine (SjCTPI- heat shock protective effect generated against the GSTs of all schistosome protein (hsp)-70), which induced very good protective effica- species was the significant inhibition of female worm fecun- cy against S. japonicum in Chinese water buffaloes when co- dity resulting in decreased egg output and viability and, con- administered with IL-12 as adjuvant. This vaccine was sequently, reduced transmission and a reduction in egg- subsequently field-tested in bovines against natural induced pathology ; these important characteristics result- S. japonicum challenge in Hunan Province in the People’s ed in the clinical testing of the Bilhvax (rSh28GST) vaccine Republic of China in a double-blind, phase-3 cluster described earlier. Protective efficacy (the characteristic anti- randomised controlled trial (RCT). The trial was designed fecundity effect) was reported in Chinese bovines (and sheep) to assess the impact on schistosomiasis transmission of a vaccinated with S. japonicum 28 kDa GST (Sj-28GST) and in multi-component, integrated control strategy, including hu- water buffaloes, cattle, sheep and pigs following vaccination man treatment, mollusciciding against the intermediate snail with the 26 kDa GST isoform (Sj-26GST). Field testing host (Oncomelania hupensis) and the bovine vaccine. The showed the protective effect of Sj26GST against S. japonicum regimen involved heterologous (DNA-protein) prime-boost is, encouragingly, maintained in cattle and water buffaloes for delivery of the SjCTPI vaccine; bovines were primed with at least 12 months post-vaccination but, disappointingly, the SjTPI-hsp-70 DNA vaccine and booster vaccinated with there have been no subsequent trials undertake, which is likely the recombinant SjTPI protein using the VacSIM (vaccine due to the considerable associated costs. self-assembling immune matrix) delivery method but without adjuvant [76, 77]. Sj-97 Although some of the outcomes of this large and complex trial were positive, the effect of the SjTPI vaccine in Paramyosin is a large (97-kDa) coiled coil myofibrillar protein preventing human infection was inconclusive due mainly to found in the secretory glands of schistosome cercariae, in the the treatment, removal or sacrifice of bovines over the course muscles of adult worms and larvae, and on the surface tegu- of the trial by the Chinese authorities. Indeed, most ani- ment of lung stage schistosomula; it first caught interest as a mals in the previously high schistosomiasis-endemic areas of vaccine candidate following encouraging results of vaccine/ Dongting and Poyang lakes have now been removed and re- challenge experiments in mice targeting S. mansoni. placed by mechanised tractors as a strategy that may provide Then, early trials in sheep, pigs and water buffaloes with na- the final element required for disease elimination from China tive or recombinant (Chinese strain) S. japonicum paramyosin [70, 78]. A similar RCT of schistosomiasis bovine vaccination (rSj-97) resulted in significant, albeit partial protection. using a multi-factorial design has been completed in the There was a pause in the progression of Sj-97 as a priority Philippines; it is anticipated the outcomes of the trial will be vaccine candidate in the early 2000s owing to its size, its poor reported soon (Allen G. Ross, personal communication). As expression, low protein yield and insolubility, the high cost with the African schistosomes, the development of a vaccine associated of up-scaling production and the expense of com- for schistosomiasis japonica has proven highly challenging pleting large animal vaccine testing. However, a robust meth- but the deployment of a transmission blocking vaccine, in od for pilot-scale expression and purification of rSj-97 was tandem with other interventions (e.g. snail control plus im- later developed (2008) which signified a major advance on proved water, sanitation and hygiene and health education), the road towards its pre-clinical assessment as a vaccine [69, for the prevention of S. japonicum would be invaluable if the 73]. Strengthened by the adjuvant Montanide ISA 206, rSj-97 goal of elimination is to be reached. proved safe, well tolerated and highly immunogenic in water buffaloes both in a highly endemic area for S. japonicum in the Schistosomiasis vaccines: the way forward Philippines [69, 73] and in the People’s Republic of China [69, 73], thereby reinforcing its status as a promising anti- Genomics, post-genomics and new vaccine antigen discovery S. japonicum vaccine candidate. Phase-2 trials of rSj97/ ISA206 in bovines and phase-1 trials in humans now need Despite the recent encouraging progress in schistosomiasis to be undertaken and should be encouraged. vaccine development, especially the clinical testing of lead Semin Immunopathol (2020) 42:355–371 367 molecules, the antigens currently used may still not provide targeting specific protein-encoding genes implicated in dis- the required level of immunological protective potency, so it is ease due to schistosomiasis but also for the identification of important to continue the search for novel target candidates. novel anti-schistosome vaccine (and drug) candidates. The application of new technologies will be key in this endeavour, and there is no doubt that pivotal to many of the A new paradigm—a controlled human challenge recent (and future) advances in antigen discovery are the avail- infection model for testing schistosomiasis vaccines ability of draft nuclear genomic sequences from S. japonicum and S. mansoni, published in 2009 [79, 80], and Introducing a paradigm shift, Meta Roestenberg and her team S. haematobium, which appeared 3 years later. at the Leiden University Medical Centre in the Netherlands Improved genome assemblies are now available for are pioneering the use of human skin explants showing that S. mansoni , S. japonicum and S. haematobium natural schistosome cercariae induce a predominantly regula-. These seminal reports provide a wealth of invaluable tory immune response whereas radiation-attenuated parasites data opening new avenues for the design and development do not. Along with this approach, accelerating schistoso- of novel targeted anti-schistosomiasis interventions including miasis vaccine development, the Roestenberg research group vaccination, as well as new insights with regard to schisto- is in the process of establishing an altogether novel some development, host-parasite interactions. Coupled with approach—the S. mansoni controlled human infection (CHI- the availability of large Schistosoma transcriptomes and ex- S) model. Thanks to the use of skin explants, this road panded genome databases , post-genomic technologies may now be open, not for morbidity manipulation or interfer- such as proteomics, glycomics, metabolomics (metabolic fin- ence with parasite fecundity, but clearly for evaluation of how gerprinting), proteome microarray profiling and exosomics to stop cercarial skin penetration. There has been a recent that provide the necessary technology platforms for exploiting resurgence in the popularity of CHI application, e.g. for the this treasure trove of information [35, 87–92], we are at a testing of vaccines against Helicobacter pylori, Norovirus and vantage point that promises rapid progress. As schistosomes Plasmodium falciparum [94–96]. With respect to the most are equipped with small RNA interference machinery, gene promising schistosomiasis vaccine candidates, the CHI-S manipulation has already been used to target different life model would indeed provide estimates of early protective ef- cycle stages of the three major schistosome species and this ficacy to be used to guide further clinical development, there- approach may guide the future development of novel interven- by reducing the risk of downstream efficacy failure and, at the tion tools. Indeed, deciphering important biological func- same time, advance understanding of the protective immunity tions, such as those involved in nutrient uptake, the mainte- generated by vaccination [93, 95]. nance of tegumental integrity, immune modulation mecha- The CHI-S work underway is especially noteworthy as the nisms, gene regulation, reproductive biology and fecundity, production of the schistosome cercariae required for challenge apoptosis and self-renewal, through the use of RNA interfer- has to strictly comply with all regulatory requirements and ence (RNAi), can reveal important genes encoding proteins good manufacturing practice (GMP) principles, a daunting, essential for parasite survival which can be targeted for immu- demanding and difficult endeavour. Safety issues are para- nological attack. mount in CHI trials, and to ensure safety of volunteers, the Although still in its infancy in relation to helminth para- Leiden group conceptualised the propagation of single sex sites, the novel clustered regulatory interspaced short palin- male schistosome cercariae that can infect human beings and dromic repeats (CRISPR) and CRISPR-associated protein mature to adult worms without mating so that egg-associated (Cas9) genome editing tool is attracting considerable attention morbidity due to granuloma formation and fibrosis is as a potent method to precisely target and then deactivate the circumvented. The CHI-S model utilises the laboratory genetic information a cell needs to produce a given protein maintained lifecycle of S. mansoni, whereby individual. The first (and, to date, only) application of this technol- Biomphalaria snails are carefully infected with a single mira- ogy for the study of schistosomes is that reported by Ittiprasert cidium which undergoes asexual reproduction in the mollus- et al. who harnessed CRISPR/Cas9 to deactivate the gene can host, producing after 5 weeks thousands of cercariae of a that encodes S. mansoni egg omega-1 ribonuclease and create single clone, and hence single sex. Following a series of parasites that either do not produce this protein, or only very quality control steps and determination of male or female sex little of it. Mice infected with this kind of gene-edited eggs by PCR, male cercariae are used for the controlled infection of have been shown to produce markedly reduced granuloma- volunteers which is performed by taping a chamber of water tous inflammation in the lungs and less pathology in general containing a predetermined number of male parasites onto the compared to those carrying non-edited parasites. This pivotal subject’s arm for an interval of 30 min. Successful infec- study provides a blueprint for editing other key schistosome tions can be detected (usually after 6–12 weeks) and quanti- genes in the future. Indeed, the application of this powerful, fied by measuring circulating anodic antigen (CAA)—a pro- but affordable, technology may prove of value not only for tein which is secreted in large quantities by adult worms into 368 Semin Immunopathol (2020) 42:355–371 the blood; this highly sensitive diagnostic assay was crucial (APP1102926) to DPM from the National Health and Medical Research Council of Australia. for the model’s development, allowing for accurate quantifi- cation of worm loads despite the absence of eggs. The CHI-S model would be suitable for testing new drugs Compliance with ethical standards and the currently available Sm14 and Sm-TSP-2 vaccines, Conflict of interest The authors declare that they have no conflict of although many more identified vaccine candidates could interest. be evaluated, either individually or in combination; molecules that predominantly generate anti-fecundity effects, such as the Sm-p80 vaccine, however, would unlikely be tested in this References single sex infection model. The current CHI-S model from Leiden provides a blueprint for future development of infec- 1. Ross AG, Bartley PB, Sleigh AC, Olds GR, Li Y, Williams GM, tion models using female cercariae avoiding the morbidity McManus DP (2002) Schistosomiasis. N Engl J Med 346:1212– 1220 otherwise following down the road. However, this will require 2. Gryseels BC, Polman K, Clerinx J, Kestens L (2006) Human schis- evaluation from the full vaccine pipeline perspective. tosomiasis. 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