Abstract:
Trichomonas vaginalis (TV) is a pathogenic protozoan that inhabits the human urogenital tract and causes Trichomonasis. This is a very common pathogen with an estimated 174 million new cases every year. This makes it the most common curable sexually transmitted infection (STI) globally, and is almost as common as chlamydia, gonorrhoea and syphilis combined. Infection with TV has been associated with many health concerns such as cervical cancer, HIV, infertility, and adverse pregnancy outcomes. Despite this, Trichomonas vaginalis remains largely ignored. In 2007 the draft genome of this parasite was published, which has allowed for many new insights into its pathogenicity. In particular this may be useful for the development of new drugs for the treatment of TV. This is important as there is currently only one family of drugs for TV and drug resistance is on the rise. While the genome is published, the majority of the genes found are described as hypothetical. Molecular studies are needed to elucidate the nature and function of these genes and their proteins. Knock out studies would be particularly useful in this regard. However, studies of this kind are notoriously difficult in TV and new techniques are needed. The CRISPR/Cas system is a bacterial immune system that has recently been adapted for use in genetic engineering. The CRISPR/Cas system is currently untried in this organism. In order to easily genetically modify TV, the components of the CRISPR/Cas system, and a fluorescent reporter protein needs to be expressed. This work compares the two fluorescent proteins iLOV and eGFP, and evaluates their effectiveness for use in TV. Two components of the CRISPR/Cas system, the gRNA and Cas9, are also described and their expression in this trichomonad attempted. This study shows that iLOV is superior to eGFP as a fluorescent marker protein in TV. The effectiveness of the expression of Cas9 and other CRISPR/Cas components in TV remains uncertain.