Malaria continues to be one of the most preeminent and pernicious global health problems affecting humans. It is one of the leading causes of morbidity across the world, especially in subtropical and tropical countries, engendering around 438,000 deaths per year (Koch et al., 2017). Despite the potency of the disease alleviating over time, the number of people at risk of being infected is increasing (from 0.8 billion in 1900 to 3.3 billion in 2010) due to growing populations (Autino et al., 2012). The need for treatment is critical.
Malaria is a caused by the Plasmodium parasite (Geleta and Ketema, 2016). It is usually carried from host to host by female Anopheles mosquitoes which release it into the victim’s bloodstream (Bartoloni and Zammarchi, 2012). Once the parasite has invaded the host’s hepatocytes, they asexually reproduce and cause hepatocytes to burst (Koch et al., 2017). The host’s immune system will mount a response, causing severe side-effects which can, in turn, lead to death. It can also be transmitted transplacentally, through transfusion and needle-sharing (Bartoloni and Zammarchi, 2012).
Thus far, malaria control has primarily focused on destroying mosquito breeding sites, using insecticides and prevention of human contact using mosquito nets and screens (White, 2004). However, malaria predominantly affects lesser economically developed countries and thus the cost of treatment serves as a barrier to disease eradication. Anti-malarial drugs are one of the most common medications provided in tropical countries. They generally work by inhibiting specific enzymes or making the environment difficult for the parasite to survive in (Basore et al., 2015). Drugs are relatively cheaper, widely available and as there is no vaccine, they currently present the most effective method of treatment (White, 2004).
One such class is Atovaquone. It works by inhibiting mitochondrial electron transport in the malarial parasite and preventing the build-up of an electrochemical gradient (Vaidya and Mather, 2000). This would prevent Plasmodium parasites from producing ATP via oxidative phosphorylation, eventually killing it. Antimalarials have been a relatively effective but their efficacy is now proving redundant (White, 2004). Parasites can evolve and gain resistance as a result of naturally occurring mutations over time via natural selection (Ashley et al., 2014). For example, the atovaquone drug can lose its efficacy due to mutations in parasitic mtDNA coding for cytochrome bc1 complex that lowers the binding affinity for the drug (Vaidya and Mather, 2000). Resistance is usually due to the misuse of drugs and not following directions properly. The rate of resistance is catching up with the rate of drug creation, with resistance materializing in nearly all classes and as such, the urgency for novel treatments has never been greater (White, 2004).
With time and advancement of scientific techniques, new solutions have presented themselves. One such avenue explored as recently as this year exploits CRISPR/Cas9 gene editing (Dong et al., 2018). It is known that the Plasmodium parasite depends on a number of agonists (receptor activators initiating biological response) when it exists in the Anopheles mosquito before being transmitted to humans (Dong et al., 2018). It was hypothesised that these could be targeted when trying to prevent transmission, potentially creating a malaria-resistant mosquito (Dong et al., 2018).
CRISPR/Cas9 technology is modified from naturally occurring type II CRISPR-Cas systems in bacteria (Doudna and Charpentier, 2014). This enables bacteria to acquire adaptive immunity to foreign DNA that invades bacterial cells, such as bacteriophage DNA. CRISPR sequences (palindromic-repeat spacer sequences within bacterial DNA) were found to be derived from invading DNA (Bolotin et al., 2005). The endonuclease, Cas9, recognises these foreign sequences and generates site-specific double-stranded breaks protecting the cell from infection by cleaving the DNA (Doudna and Charpentier, 2014).
The transmission cycle in the Anopheles gambiae mosquito was investigated as they are the primary transmitters of the most pervasive malarial strain to humans, Plasmodium falciparum (Dong et al., 2018). The fibrinogen-related protein 1 (FREP1) gene in the mosquito was identified and targeted for inactivation to see how this affected its interaction with Plasmodium and the health of the mosquito. FREP1, secreted in the midgut epithelium, is vital to parasitic infection of mosquitoes (Zhang et al., 2015). It binds Plasmodia ookinetes (zygotes) enabling it to invade the mosquito’s midgut where it can then develop and eventually be transmitted to humans (Zhang et al., 2015). Previous experiments using anti-FREP antibodies indicated that inhibiting FREP1 suppresses P. falciparum growth effectively thus blocking transmission (Niu et al., 2017). Consequently, CRISPR/Cas9 was used to create FREP1 knockout mutants and its effectiveness assessed. The mutants were fed P. falciparum gametocytes alongside wild-type, who still had FREP1, and it was observed that the mutants displayed significantly lower rates of infection than the wild-types (41.3% vs. 82.9%) (Dong et al., 2018). This profound suppression is of epidemiological significance as many of the mosquitoes are now effectively resistant, the Plasmodium cannot survive and multiply in the host, preventing transmission to humans (Dong et al., 2018).
If we can replace all wild-type mosquitoes with mutant strains, then this method seems faultless on the surface. However, it does incur some costs. FREP1 is essential for mosquito metabolism as it is required for blood feeding and digestion (Dong et al., 2018). Some health costs were observed during the experiment such as slower development which would affect their ability to take up nutrients and lay eggs. These drawbacks would make it difficult to spread this malarial-resistance into the population as the mosquitoes may not survive long enough to pass it on (Dong et al., 2018).
In conclusion, it is clear that gene targeting is key to preventing the transmission of this pernicious disease, especially with rising antimalarial drug resistance usurping new drug development. The study by (Dong et al., 2018) shows how CRISPR/Cas9 technology could provide the answer to this problem and certainly warrants further investigation. Deleting Plasmodium host factors such as FREP1 can create malaria-resistant strains, but they need to be able to compete with wild-type populations to be able to confer resistance and prevent thousands of deaths every year.