**4. Past, present, and future of transmission-blocking intervention strategies**

Over the last two decades, the use of long-lasting insecticide-treated nets (LLINs) and indoor residual spraying (IRS) have been major contributors to gains in malaria eradication efforts [35–38]. Even with the routine use of these key malaria interventions as well as effective malaria treatment with artemisinin-based combination therapies (ACTs), malaria-related mortality and morbidity remain unacceptably high with about half a million people losing their lives to the disease annually. Further, the development of resistance by the vector to insecticides as well as generation of parasite resistance to antimalarial drugs highlights the need for sustaining current gains and developing additional innovative control measures. Novel transmissionblocking intervention by vaccines or genetically engineered mosquitoes may provide a promising approach.

In 2013, the Malaria Vaccine Initiative (MVI) was rolled out. One of its key goals is the development of vaccines capable of interrupting transmission. Transmission- blocking vaccines (TBVs) are the result of efforts put in place by researchers to understand the interaction between the parasite and mosquito. Some TBVs are currently undergoing trials for efficacy and other key measures of success (**Table 1**). Alanyl aminopeptidase


#### **Table 1.**

*Transmission-blocking vaccine candidates.*

N (AnAPN1) is the leading midgut TBV immunogen [8]. (AnAPN1) is highly immunogenic and conserved between different anophelines. This makes it very attractive for vaccine development as vaccines prepared with this antigen should be active against all human malaria vectors hence saving the resources needed to develop specific targets for different Anopheles/Plasmodium species combinations [54]. Other midgut candidate molecules include carboxypeptidase [40], calreticulin [41], Croquemort SCRBQ2 [42] and myosin [43]. Candidate parasite molecules include those found on the surface of gametocytes and gametes (Pfs 2400, Pfs230, Pfs48/45) as well as zygotes and ookinetes (Pfs25, Pfs28) [55]. Vaccine against Pvs25 and Pfs25, which blocks *P. vivax* and *P.falciparum* respectively, are currently the leading molecule for a TBV [45, 56].

More recently, scientists have shifted attention from regular vector-control strategies (LLINs and IRS) to engaging advanced molecular tools such as CRISPRcas9 to re-program the vector genome so as to make them refractory to the parasite (**Figure 1**). Gene drives skew the pattern of inheritance of genes creating mosquitoes that will either reduce mosquito populations or make mosquitoes less likely to spread the malaria parasite. Using such gene-drive systems in the laboratory, researchers have been able to transfer antimalarial effector genes to mosquitoes [57, 58]. Intriguing using similar allelic drive system in a *Drosophila melanogasta* model, Kaduskar et al. [59] were successful at reversing the most widely spread insecticide resistance mutation in anophelines (L1014F) (a mutation in the

**Figure 1.** *Breaking the cycle of malaria transmission.*

## *Vector-Parasite Interactions and Malaria Transmission DOI: http://dx.doi.org/10.5772/intechopen.105025*

voltage-gated sodium channels of mosquitoes which make them resistant to pyrethroids) to susceptible wild-type genes (1014 L). The report of these researchers opens new vistas for vector control as it demonstrated that not only can insecticide resistance (IR) be reversed but that there is a relative negative fitness cost for the (L1014F) insecticide-conferring mutation as compared to the wild-type allelic variant. Thus, offering the opportunity to synergize the use of a gene drive that confers a bias inheritance of the preferred wild-type allelic variant with regular vector control methods. They went on to suggest that the identification of target site variants that would make the vector hypersensitive to insecticides will hold an even better promise. Further, they suggested the possibility of combining such gene drives for reversing insecticide resistance with other systems promoting refractoriness in mosquitoes [59].

Mosquito picks up gametocytes from the blood of an infected person. The sexual cycle is initiated due to the presence of xanthurenic acid (XA) as well as


#### **Table 2.**

*Pros and cons of transmission-blocking intervention strategies.*

the low temperature and high pH of the mosquito stomach. The parasite then progresses through different stages and eventually forms sporozoites which are infective to humans. As depicted in the image above, parasite development can be interrupted at any stage of its sexual cycle either by using TBVs (candidate molecules are in boxes bordered by red broken lines) or genetic-based tools could be used to alter the expression levels of proteins crucial for mosquito infection (see boxes with broken black lines). Such tools capable of breaking the transmission chain could be incorporated as part of an integrated antimalarial strategy to eradicate the disease.

Although successful transgenic manipulation of mosquitoes has been achieved in the laboratories, their relative negative fitness in relation to wild-type populations is an important limitation for their relevance for large-scale use. For example, in a mark-release-recapture study in Burkina Faso recently, hemizygous geneticallymodified (GM) sterile and non-transgenic sibling males of *Anopheles coluzzii* were released into a field in a controlled study. Recovered carriers of the GM trait had lower survival and were less mobile than their wild-type siblings [60]. Another shortcoming to the use of genetic-based vector control tools is that employing methods such as transposon-mediated transformation which modify only one allele of the desired gene, would spread the desired trait only to half of the offspring, and would eventually get eliminated in the wild population. This can however be overcome by employing gene drive systems such as CRISPR nuclease Cas9 which are capable of copying themselves to both gene alleles that will be inherited by all offspring, and thus spread more efficiently through a wild population [59]. Despite their great promise, scientists are wary of gene drive because they could cause irreparable damage since they permanently alter an entire population (**Table 2**). Further, issues bothering on their safety, governance, affordability, and cost-effectiveness need to be addressed (**Table 2**).

#### **5. Conclusion and future perspectives**

The prolonged and repeated use of insecticides of a limited chemical class is a major contributor for the acquisition of resistance mutations in insecticide-target genes in insects. On the part of the malaria parasite, it also constantly evades antimalarial drugs by generating resistance mutations. Hence, the need to identify additional control measures that TBVs and genetics-based vector control tools may offer. Broad-spectrum malaria transmission-blocking vaccine antigens such as FREP1 [66] and (AnAPN1) offer attractive targets for the development of TBVs. However, due to the lack of industrial partners, the development and production of TBVs have stalled [61].

To generate transgenic mosquitoes, mosquito proteins that cause complete refractoriness in the vector upon silencing may be the target of choice. An ideal target molecule would be one that does not impose a relative negative fitness cost on the insect and one which could be used in combination with others targeting different stages of the parasite life cycle [67]. However, a major challenge with genome editing techniques is devising means to safely drive effector genes into mosquito populations in the field without causing harm to other organisms, including humans. Once safety concerns are addressed, such tools could be integrated with traditional vector control strategies, in addition to effective malaria treatment and good sanitation practices for the actualization of the eradication goal.
