Public health concerns over gene-drive mosquitoes: will future use of gene-drive snails for schistosomiasis control gain increased level of community acceptance?

ABSTRACT

With the advent of CRISPR (clustered regularly interspaced short palindromic repeat)-based gene drive, present genetic research in schistosomiasis vector control envisages the breeding and release of transgenic schistosome-resistant (TSR) snail vectors to curb the spread of the disease. Although this approach is still in its infancy, studies focussing on production of genetically modified (GM) mosquitoes (including gene-drive mosquitoes) are well advanced and set the pace for other transgenic vector research. Unfortunately, as with other GM mosquitoes, open field release of gene-drive mosquitoes is currently challenged in part by some concerns such as gene drive failure and increased transmission potential for other mosquito-borne diseases among others, which might have adverse effects on human well-being. Therefore, not only should we learn from the GM mosquito protocols, frameworks and guidelines but also appraise the applicability of its current hurdles to other transgenic vector systems, such as the TSR snail approach. Placing these issues in a coherent comparative perspective, I argue that although the use of TSR snails may face similar technical, democratic and diplomatic challenges, some of the concerns over gene-drive mosquitoes may not apply to gene-drive snails, proposing a theory that community consent will be no harder and possibly easier to obtain for TSR snails than the experience with GM mosquitoes. In the future, these observations may help public health practitioners and policy makers in effective communication with communities on issues regarding the use of TSR snails to interrupt schistosomiasis transmission, especially in sub-Saharan Africa.

Introduction

Mosquitoes and freshwater snails are important vectors of human diseases, especially in the tropical and subtropical countries; however, mosquitoes are the most important disease vectors that significantly impact global health [1]. Many species of freshwater snails serve as vectors for trematode-causative neglected tropical diseases (NTDs), of which schistosomiasis, caused by the intravascular blood-dwelling trematodes of the genus Schistosoma, is the most important and widespread. Schistosomiasis causes both acute and chronic disease manifestations such as diarrhea, hematuria, hepatosplenomegaly, periportal fibrosis (Symmer’s pipestem fibrosis) with portal hypertension, hydronephrosis and bladder cancer [2]. Out of the 779 million people thought to be at risk of schistosomiasis infection [2], approximately 220 million people [3] or many more [4] are currently infected, and over 2.5 million human life-years were lost to consequent disabilities and mortality in 2016 [5]. Apart from the 2013 outbreak in Corsica (France) [6], schistosomiasis transmission has been reported from 78 countries, out of which large-scale preventive chemotherapy is still required in 52 endemic countries, most of which are clustered in the World Health Organization (WHO) African region [7,8]. Some countries including Japan, China, Thailand and Morocco have successfully achieved schistosomiasis control and are progressing toward the disease elimination [7]. Unfortunately, many projected national goals to control and/or eliminate schistosomiasis have failed or become infeasible and despite that WHO has slated schistosomiasis for global elimination as a public health problem by the year 2025 [7], there is no immediate prospect of meeting all requirements within the time frame [9,10]. While monochemotherapy with praziquantel (PZQ) has been commendably useful for schistosomiasis treatment and morbidity management since the past 40 years, implementation of more holistic approach including but not limited to sanitation, access to clean water, health education, high-sensitive diagnostic techniques and enhanced snail control is needed to make global elimination of schistosomiasis an achievable goal. Integrated control approach is especially needed for schistosomiasis since reliance on PZQ is flawed by its (i) ineffectiveness against juvenile schistosomes (schistosomula) [11], (ii) waning efficacy against adult schistosomes [12,13] and (iii) inability to prevent parasite reinfection [14]. Given the demonstrated effectiveness of artemisinin against schistosomula, combination of artemether and PZQ seems to be a more effective combination therapy but may adversely increase the risk for the spread of artemisinin-resistant malaria parasites, especially in areas that are co-endemic for schistosomiasis and malaria [15].

Human schistosomiasis is transmitted by four genera of freshwater snails: Bulinus, Biomphalaria, Oncomelania and Neotricula. Much of the early focus on snail control has been shifted on mass drug administration (MDA), initially since the late 1970s to the early 1980s after PZQ met the WHO’s requirements for population-based chemotherapy [reviewed in 16] and increasingly since 2001 when the World Health Assembly (WHA) officially endorsed preventive chemotherapy as the principal strategy for schistosomiasis morbidity and mortality reduction [17]. Nevertheless, it is clearly beyond doubt that effective snail control is pivotal to the targeted global elimination of schistosomiasis [18]. Unfortunately, however, there are still some challenges complicating the present snail control strategies. For example, chemical mollusciciding with niclosamide is expensive, toxic to other aquatic organisms (therefore may threaten aquatic biodiversity), does not prevent snail recolonization and may not be practically effective against Neotricula. Chemical mollusciciding is in fact becoming increasingly unwanted by communities due to the toxicity of niclosamide to important freshwater animals (such as fishes) and yellowing of treated water [19,20]. In the Philippines for instance, the use of niclosamide has been disallowed under the Clean Water Act [20]. It has also been recently shown that cadavers of molluscicide-treated snails are repulsive to decomposer flies and therefore may show slower decomposition in the environment [21]. Moreover, introduction of molluscivores or biological competitors is not reliable and could impact agricultural and ecological systems; whereas, environmental alteration of the vector habitats, apart from putting workers at risk of infection, is usually not cost-effective and may be difficult to implement in resource-constrained areas or in areas where the snail habitats are important horticultural or agricultural sites such as aquacultural and rice fields [19,20]. These suggest that although the current snail control methods, especially the niclosamide-based mollusciciding, may drastically reduce vector population, their promise toward achieving global schistosomiasis elimination and subsequent eradication may be diminishing. To overcome these challenges, snail control research is being revisited with renewed interest. Notwithstanding some documented concerns about genetically modified (GM) mosquito approach as well as its complexity for obtaining community acceptance, scientists in molecular genetics of the molluskan vectors of schistosomiasis envisage the breeding and release of transgenic schistosome-resistant (TSR) snails to curb the spread of the disease [reviewed in 22]. Indeed, successful breeding and eventual use of TSR snails will be a monumental achievement in the history of schistosomiasis vector control and will expedite sustainable global elimination of the competent vectors and therefore, of the disease. This conceived approach is nevertheless still in its infancy, but advancements in the genetic engineering of mosquito vectors set the pace for other transgenic vector research. The advances and potential benefits notwithstanding, release of transgenic mosquitoes in nature is in part challenged by some human health concerns which are subjects for ongoing debate. Therefore, while we learn from the GM mosquito protocols, guidelines and frameworks, it is also worth assessing the applicability of its current hurdles to other transgenic vector systems.

Major approaches to developing gene-drive vectors

Gene drives are systems of biased inheritance in which, through sexual reproduction, specific genes (termed payloads) are spread throughout a population from one generation to another in a super-Mendelian fashion [23]. Depending on their specific genetic configuration, gene drives such as the natural homing endonuclease genes (HEGs) [24] and Medea (Maternal Effect Dominant Embryonic Arrest) [25] may be self-sustaining (i.e. spread indefinitely among generations of target population); whereas, others such as daisy-chain drive [26] and underdominance system [27,28] may be self-limiting (i.e. lose their ability to spread among subsequent generations over time). Although the use of self-sustaining approaches has been supported for humanitarian objectives as they could ultimately provide more lasting and cost-effective public health solutions, reversibility of their unforeseen effects may be more difficult [29]. Whether self-sustaining or self-limiting, the two major approaches to developing gene-drive vectors are: (i) population suppression, which is intended to reduce or crash the population of target vector and (ii) population replacement (or modification), which would render population of target vector refractory to specific pathogen. Using several species of mosquito vectors, excellent progress has been made in the laboratory experimentation of gene drive-based vector control approaches, either for population suppression [30,31] or replacement [32–34]. Each of these approaches however has its peculiar advantages and disadvantages. For instance, population suppression strategy such as that based on the release of GM male mosquitoes may pose minimal human health risk (since male mosquitoes neither bite nor transmit diseases) [35] while on the other hand, population modification may be less disruptive to the ecosystem (since the population of the target vector will not be decimated or eliminated) [36,37].

A brief conception of the TSR snail approach

As noted previously [22,38], developing schistosome-resistant snails is a more promising approach for transgenic snail design for schistosomiasis control. This is in part due to greater extent of genetic research into the immunological basis of snail/schistosome systems which has amplified knowledge in this area but more importantly, TSR approach proves a more viable option since the snail vectors are either hermaphroditic (with many species capable of cross-fertilization) or when dioecious, both sexes could equally transmit schistosomiasis [22]. As a consequence, release of transgenic yet susceptible snails carrying population-suppressing payload may rather temporarily increase the chances for disease transmission. This may also apply to arthropod vectors with blood-sucking and disease-transmitting males (e.g. tse-tse flies, fleas, lice, ticks and triatomine bugs). The aptness of this population replacement approach for hermaphroditic (but cross-fertilizing) vectors and those with disease-transmitting males may also be an additional advantage it has over the population suppression approach. In line with the approach that best applies to the genetic control of schistosomiasis vectors, this article will focus more on the current public health concerns over the use of gene-drive resistant mosquitoes and snails.

Current public health concerns over gene-drive resistant vectors: a mosquito-versus-snail appraisal

Concerns over potential environmental, ecological and health-related impacts of the proposed release of gene-drive modified organisms have been expressed [39–41]. Some perceived ecological risks generated many of the current public health concerns. Generally, public health concerns raised over the release of GM mosquitoes center majorly on their potential and unpredictable adverse effects on the human population [37]. These issues have been enumerated and discussed in detail [35,37,42,43]. However, those within the purview of this article, which also cover the majority of the most important issues, relate to: (1) failure of the GM mosquito techniques to work expectedly (2) evolution of more virulent pathogen strains (3) alteration of the GM mosquito blood-feeding behavior (4) susceptibility of GM mosquitoes to other mosquito-borne pathogens (5) GM mosquito resistance to insecticides and (6) spread of GM mosquitoes beyond release sites (Figure 1).

Figure 1.

Comparative public health concerns over the use of gene-drive resistant mosquitoes and snails and proposed attitude of communities toward acceptance. SA: strongly agree (unconditional acceptance); A: agree (may be conditional upon meeting certain requirements); D: disagree (may be due to current unresolved issues); SD: strongly disagree (unconditional rejection). Notes: (i) Human population decreases with ascending level of the pyramid (ii) Since no gene-drive vector has been released in an open field anywhere, for the purpose of this article, a currently high public resistance to the use of gene-drive vectors is assumed and was used as a basis for the proposed reaction of communities toward acceptance. Further differential levels of community attitude toward acceptance were presumably and fairly based on the present discussion on comparative human safety concerns over both gene-drive vectors.

The advent of CRISPR/Cas9 (clustered regularly interspaced short palindromic repeat/CRISPR-associated protein 9), an endonuclease-based system, in genome editing provided a renewed momentum for effective establishment of gene drives in the laboratory for eventual release in the field [44,45]. However, the mushrooming interest in the application of CRISPR/Cas9-mediated gene drive also stokes concerns about its potential public health risks. In the recent past, these concerns triggered an inter-organizational collaboration and convention of an expert committee on scientific advances in gene drive technology and considerations for its responsible application [40]. Among other perplexing yet fundamental questions that were raised are: will gene drive approaches be effective? Will they have unintended consequences on the public health? And if yes, what can be done to reduce the risks?

During an expert workshop organized by the Foundation for the National Institutes of Health (FNIH) and the International Life Sciences Institute Research Foundation (ILSIRF) on problem formulation for the use of gene drive in mosquitoes [41], some current concerns related to human health were examined and few consensuses were reached, two of which placed limits on some existing concerns: first, incidental exposure to GM mosquitoes through inhalation or ingestion is not likely to provide significant levels of harmful exposure; second, horizontal gene flow from GM mosquitoes to humans is biologically impossible. These might as well limit similar concerns about the use of TSR snails but having stated these, other aforementioned public health concerns remain very pertinent. From this point forward, this discussion will be tailored more specifically toward current concerns over potential adverse consequences of the use of gene-drive pathogen-resistant mosquitoes on human safety, also juxtaposing these concerns with possible scenarios in the case of gene drive-based TSR snails (Figure 1).

The most daunting concern about the use of gene-drive vectors is the possibility of gene drive failure. In this context, CRISPR/Cas9-based knockout of susceptible genes and/or knockin of resistant genes may fail under certain conditions. Failure may occur inherently in the CRISPR/Cas9 system itself, when the Cas9-mediated double-strand incision in the DNA sequence is sewn back together via nonhomologous end joining (NHEJ) repair mechanism which is error-prone [24,46]. Failure may also ensue as a result of (i) natural genetic variation of the target gene sequence within a population [47,48] or (ii) genetic mutation in the target gene sequence [46]. In the first circumstance, it is possible that the modified vectors work effectively somewhere (such as where they are developed) and not elsewhere (such as in areas with different geographic variants of the vector or target pathogen). For example, a recent genome analysis of wild Anopheles mosquitoes from across Africa found extreme genetic diversity among populations of the same species [49]. Such heterogeneity, as observed by Ogola et al., [50] underlies variation in vector competence and Plasmsodium infectivity, and could have practical implications on the development of gene-drive mosquitoes. In the same vein, compatibility polymorphism is not an uncommon phenomenon in snail/schistosome interactions [51]. A transcriptomic analysis by Galinier et al., [52] showed that some known molecular determinants of Schistosoma mansoni infectivity in Biomphalaria glabrata are variable genetically and in their level of expression among different combinations of the vector/parasite strains [52]. In the second circumstance, pathogens generally demonstrate tenacious ability to evolve resistance and circumvent every control stratagem devised against them, not excluding the transgene-driven vector immunity. The evolutionary counteractive response in the target pathogen could result in disease resurgence after a period of effective control [53]. There is also a concern over possible evolution of new pathogen strains or variants due to natural selection against the anti-pathogen payload effects. It has been speculated that this occurrence may be accompanied by an unforeseen event. For instance, a new pathogen variant might be more difficult to control than the previous version [54]. Considering possible increased virulence of the evolved pathogen variant, James et al., [39] suggested an assessment of the genotypic or phenotypic changes in pathogens after passage through gene-drive vectors. Some potential pathways to mitigate the risk of gene drive resistance [as postulated by 24, and 46] have been considered experimentally by Buchman et al., [55] and Champer et al [56]. This however remains a part of the research yet to be concluded in the development of an efficient gene drive technology. Taken together, since developing these ideas for vector/pathogen targets will require a lot of research effort, experimental advancement and convincing proofs of the concept; and since there is currently no effective way to prevent or suppress the repair of CRISPR/Cas9 gene drive-induced cleavage through the NHEJ pathway, the issue of gene drive failure remains a formidable barrier.

It has also been identified that introgression of transgenes into the natural mosquito populations might alter mosquito blood-feeding behavior which could have a negative impact on human health, such as nuisance biting and allergic reaction [29]. Unarguably, the most likely route of exposure to GM mosquitoes (thus the most direct human health risk) is via biting. The saliva of all mosquitoes naturally elicits an immunological or even strong allergic response in humans. It is therefore often recommended that possible expression of a transgene product in the saliva of a genetically engineered blood-sucking mosquito and potential toxicity and allergenicity of the introduced proteins to human should be considered [29,39,41]. With or without the understanding of this, people may show stronger opposition to the use of any genetically engineered blood-sucking mosquito. For instance, many participants in an opinion survey in Mali opted for the genetic mosquito reduction/elimination strategy rather than the mosquito modification strategy [43]. Only knowing that they would directly be exposed to bites from these genetically engineered mosquitoes may be enough to generate significant level of fear. Therefore, one fundamental factor that may prompt the disapproval of gene-drive resistant mosquitoes by many individuals and communities is the fear of the natural intimacy between mosquitoes and humans (Figure 1). Generally, snail vectors of human schistosomiasis do not have or require direct association with human at any stage of their lifespan. In this case, potential changes in the feeding behavior of transgenic snails would not pose any direct threat to human health.

Furthermore, in many cases, a single species of mosquito may efficiently transmit two or more human-infecting pathogens [see 35, for examples]. Another concern that has been raised is that conferment of resistance to target infections in such mosquito vectors might simultaneously render them more susceptible to other infections [37], therefore increasing the prevalence of other mosquito-borne diseases among human populations. For instance, antagonistic co-occurrence of Plasmodium and filarial worms in Anopheles mosquitoes has been evidenced, suggesting that elimination of filarial worms within Anopheles mosquito vectors in a malaria/filariasis co-endemic locale may enhance the vector competence for malaria transmission [57]. In the recent problem formulation workshops on the use of gene-drive mosquitoes in Africa, organized by the New Partnership for Africa’s Development (NEPAD) in four different African regions (in Ghana, Kenya, Gabon and Botswana), the potential increase in incidence rates of other mosquito-borne diseases was also identified as a major health concern to be addressed [58]. In snail/trematode associations, other than the species of Schistosoma, the snail vectors of human schistosomiasis almost never transmit any other human-infecting trematode [59,60]; only the secondary vectorial role of Oncomelania hupensis nosophora in the transmission of Paragonimus spp. (lung fluke) appears unequivocal [60,61]. Empirical evidence also suggests that Oncomelania and Biomphalaria snails may potentially carry Angiostrongylus cantonensis (the rat lungworm) [62,63], a zoonotic snail-vectored nematode that is broad in its choice of snail host [59,64]. Apart from these clarifications, no other known human pathogen is transmitted by the schistosomiasis vectors. Moreover, among the few species of Schistosoma known to infect humans, S. mansoni, S. haematobium and S. japonicum are responsible for most cases of human schistosomiasis. S. mekongi is also a main human-infecting schistosome in several districts in Cambodia and along Mekong River in Lao People’s Democratic Republic. Biologically, no two species of the aforementioned human-infecting Schistosoma are transmitted by the same genus of snail vector, even when they both occur in the same geographical location or freshwater body. The single exception is that S. haematobium, S. intercalatum and S. guineensis are transmitted by Bulinus snails; the two latter species are geographically limited to West and Central Africa, overlapping with other occurring Schistosoma species, and are less pathogenic [2]. Considering the stringent snail/schistosome interactions, one would agree that although increased susceptibility of gene-drive resistant mosquitoes to another mosquito-borne human-infecting pathogen has been emphasized, such occurrence would raise minimal concern, if any, in the use of TSR snails.

The emergence of mosquito resistance to a range of available insecticides has been a considerable cause for concern in mosquito control effort [65,66]. The occurrence leads to persistently high vector densities, thus perpetuating biting nuisance and causing the spread of mosquito-borne diseases to continue unabated. Mosquitoes that are genetically configured to resist pathogens may abate disease spread but will retain their ability to develop resistance to insecticides and participate in spreading insecticide-resistance genes within the target vector population. On the contrary, niclosamide (Bayluscide®) is a potent chemical molluscicide that is active against snail vectors. Despite that the knowledge of its mechanisms of action remains sparsely understood [67], the molluscicide has officially been in use for snail control for nearly three decades and up till now, there is no evidence of snail resistance to niclosamide [68–70]. Although, over 30 years ago, Sullivan et al., [71] demonstrated the eventuality of B. glabrata selection for niclosamide tolerance, it was recently stated by WHO [72] that evolution of snail resistance to niclosamide is of little concern, especially with the current limited use of molluscicides which keeps selective pressure for resistance below threshold. Nevertheless, the WHO recommends a 2-year-interval resistance monitoring in areas where niclosamide is being frequently used [72]. A major relief and advantage here is that chemical mollusciciding may still be applied successfully to complement or mitigate the TSR snail strategy without the threat of vector resistance to the molluscicide.

In a checklist formulated by Brandt et al., [73] one important element for risk identification is the ability of the gene-drive organisms to spread, either globally or locally, also taking into consideration the speed of spread and distance covered by the organisms. Interboundary or cross-country spread of gene-drive resistant vectors may create more problems if they spread to other communities or areas where the idea is not welcomed (perhaps due to any fear or yet-to-be-cleared concern among residents of the new host areas). Here, it is important to briefly mention some possibilities on the spread of TSR snails beyond target areas, either as a consequence of flooding or of snail release into contiguous water bodies that extend from a community to other neighboring communities. In addition, similar to some major pathways by which mosquitoes could passively disperse farther through transportation by humans [74,75], it is also possible that freshwater snail vectors intentionally or inadvertently get transported into new areas, especially by agricultural and horticultural industries [76,77]. Yet, it may be argued that, unlike the GM mosquitoes which may exhibit active rapid dispersal mechanism to spread extensively within and perhaps beyond the release area, TSR snails would not spread actively as such, since gastropods generally exhibit sluggish movement and active dispersal of snail vectors of schistosomiasis is normally confined within or around freshwater sites (Figure 1). This factor may localize the spread of TSR snails or make physical containment more practicable post-release in some settings.

Further considerations

Schistosomiasis and malaria are among the human scourges that CRISPR-based gene drive technology can most likely help to eradicate [78,79]. However, deployment of gene-drive organisms in nature cannot be actualized solely by scientific elegance. It is generally evident that obtaining public approval is more decisive but unfortunately more challenging in the release of GM organisms. Moral consideration for the application of CRISPR technology in genome engineering and control of vector-borne diseases requires that a particular transgenic vector design should be used only when the potential benefits outweigh the risks [40,78]. To maximize the benefits of TSR snails, field release will have to be conducted in areas or communities where schistosomiasis is a major public health problem, such as in many developing countries. In sub-Saharan Africa, the impact of schistosomiasis is second only to that of malaria. The region bears the highest global burden (up to 93%) of schistosomiasis, especially in rural areas and among the poor human populations [80]. This implies that attitude of the people living in sub-Saharan Africa toward the use of TSR snails will play a critical role as to the extent to which this approach will influence global elimination or eradication of schistosomiasis. Notwithstanding the opposition to the recent (non-gene-drive) GM mosquito release in Burkina Faso [81,82], experience derived from the release initiative forms an important precedent that will inform the development and subsequent release of GM vectors such as TSR snails in Africa [83,84]. It has been emphasized that the ethically challenging execution of trials of high-tech, controversial and potentially irreversible innovations in African settings requires careful public engagement, regardless of the scientific and operational rationality behind executing such control initiatives in a region with pronounced burden of diseases [83]. Indeed, public engagement (a long term, multidirectional, iterative process of communication that seeks and facilitates the sharing and exchange of knowledge, perspectives and preferences between or among groups with varying expertise, power and values) is one of the key areas of responsible science and application of gene drive technology [40], and is essential to meeting ethical obligations of informed consent, building trust and gaining acceptance of research [39].

As described in the NASEM report [40], Communities are group of people who live in or near enough to the gene drive technology release site such that they have tangible and immediate interests in the project. These may include those who reside within the flight range of the mosquito vectors [85] or nearby residents who visit the snail-colonized water bodies for domestic or occupational purposes. Important pathways to an effective community engagement have been discussed elsewhere [39,86–88] and if the issues highlighted can be successfully addressed and the ideas provided can be practically applied, increased number of local populations may be motivated to support gene drive technology for the control of vector-borne diseases. Encouragingly, inferences from previous public opinion surveys on the use of GM mosquitoes in some sub-Saharan African countries [43,89,90] coupled with the discussions in this article and with a substantive community engagement show potential for an improved level of community support in the future deployment of TSR snails for schistosomiasis control (Figure 1).

Concluding remarks

Placing the current public health issues on the use of gene-drive vectors in coherent perspective, it is becoming increasingly clear that amidst the present technical, democratic and diplomatic challenges of gene drive technology, the future use of gene drive-based TSR snails may be more appealing in terms of human safety (due to potentially fewer perceived risks) and may not raise as many public health debates as raised by the current proposed use of gene-drive mosquitoes. This might however not be the case with the anti-GM players who will oppose any form of genetic strategy. Although the theory cannot be immediately proven that the release of gene-drive snails will be more acceptable to communities than the gene-drive mosquitoes would be, there are some factors that are in its favor. This presents the use of TSR snails as a more fascinating and promising control approach that may be pursued. It should be noted, however, that the key issues discussed herein may not be exhaustive; there may be other adverse human safety issues that are presently insufficiently understood, and as they emerge, further comparative assessment will be worthwhile. Nevertheless, in the future, the human safety issues highlighted in this review may help public health practitioners and policy makers in effective communication with communities on issues regarding the use of TSR snails to block schistosomiasis transmission. Ultimately, this process of learning from GM mosquitoes may inform future programs on best practices, just as more recent programs have learnt from the criticism against earlier non-gene-drive mosquito releases. Therefore, those planning TSR snail release should build on successes of other genetic strategies and amplify the difference that would favor TSR snails over GM mosquitoes from the perspective of public concern.

Disclosure statement

No potential conflict of interest was reported by the author.

Correction Statement

This article has been republished with minor changes. These changes do not impact the academic content of the article.