Sexual reproduction in trypanosomes

Trypanosomiasis is a major livestock disease that historically crippled the development of agriculture in sub-Saharan Africa by destroying both draught and production animals. This tsetse fly-transmitted disease continues to severely constrain livestock production in many countries in Africa, because it is very widespread and affects most of the major livestock species, including cattle, sheep, goats, pigs, horses, donkeys and camels. Vets have few drugs available for prophylaxis or treatment, and drug resistance is increasing and widespread; there is no vaccine. Climate change is likely to exacerbate the problem of animal trypanosomiasis, which already affects temperate as well as tropical and subtropical regions.

The disease is caused by microscopic, single-celled organisms called trypanosomes, which are found in the blood of affected livestock. The goal of our research is to find out how trypanosomes swap their genetic material among themselves and whether this generates novel types of parasite with new features. Genetic exchange enables the rapid spread of genes through a cell population by transfer from cell to cell. This is particularly important in the case of pathogenic microbes, because once a gene for drug resistance has arisen, it can quickly spread, rendering a previously effective drug useless. A microbe with a sexual cycle is therefore potentially more difficult to control than one that reproduces asexually.

Here we will investigate sexual reproduction in the livestock pathogens Trypanosoma brucei and T. congolense. Both these trypanosomes are carried by tsetse flies, but with significant differences: T. brucei develops in the fly salivary glands and this is where it is known to undergo sexual reproduction with formation of novel hybrid strains; T. congolense develops in the fly's biting and feeding apparatus, the proboscis, and in our recent work we have shown it also undergoes genetic exchange there, but as yet have no information on the details.

In humans, the gametes - egg and sperm - are produced by a special form of cell division called meiosis. Egg and sperm combine to form a single hybrid cell called the zygote, which then develops into the embryo. The genome of the zygote consists of one set of maternal and paternal chromosomes in the cell nucleus, plus maternal mitochondrial DNA. We will investigate how trypanosomes produce gametes and how the gametes combine to form a zygote. We have identified trypanosomes undergoing meiosis by searching for cells expressing meiosis-specific proteins tagged by fluorescence and will follow these meiotic cells to see how they produce gametes. We have shown that the gametes fuse together and exchange cytoplasm, but do not yet understand how they combine their DNA. Like humans, trypanosomes have both nuclear DNA consisting of paired chromosomes and mitochondrial DNA; however trypanosome mitochondrial DNA is tightly packaged into a cellular compartment called the kinetoplast. We know from analysis of hybrid progeny that both the nuclear and kinetoplast genomes are inherited from both parental trypanosomes.

Our preliminary experiments show that the approaches are feasible and will yield interpretable data. By the end of the project we will have elucidated the process by which trypanosomes exchange genetic material. In the long term, this will inform strategies to control livestock trypanosomiasis, because we will understand the limitations to spread of harmful genes through the parasite population and the likelihood of new strains arising that have never been encountered previously by livestock.

It is now recognised that the kinetoplastid protozoa, a group that includes many important human and livestock pathogens, are capable of sexual as well as asexual reproduction. Sex provides the opportunity for gene flow, generating new recombinant pathogen strains never previously encountered by the host organisms and enabling the rapid spread of harmful traits through a pathogen population. Recombination between pathogen genomes can have devastating consequences as evidenced by the last three influenza pandemics, all caused by recombinant animal and human viruses.

Trypanosomiasis is a major livestock disease that causes vast economic losses and is widespread in tsetse-infested regions of sub-Saharan Africa. It is caused by trypanosomes, protozoan parasites found in the blood of affected livestock and transmitted by tsetse. There is no vaccine and few drugs are available for treatment, while reports of drug resistance are increasing. Here we will investigate sexual reproduction in Trypanosoma brucei and T. congolense, gaining comparative and complementary data from these two important livestock pathogens. We will determine how these trypanosomes undergo meiosis and produce haploid gametes, and how genetic material is transferred from the gametes to the zygote using fluorescent reporters to track the inheritance of organelles such as the nucleus and kinetoplast (= mitochondrial DNA).

By the end of the project we will have elucidated the process by which trypanosomes exchange genetic material. In the long term, this will inform strategies to control livestock trypanosomiasis, because we will understand the limitations to spread of harmful genes through the parasite population, and the likelihood of new, potentially more virulent, strains arising. Crossing drug resistant trypanosome strains in the lab will elucidate the genetic basis of drug resistance enabling us to develop molecular markers for drug resistance in field isolates

Summary
Investigating how pathogen strains recombine and mix their genetic material informs our understanding of the disease we are trying to control. With increasing reports of drug resistance in livestock trypanosomiasis, we need to understand the genetic basis of drug resistance and determine how fast it is likely to spread in the trypanosome population. We also need to understand the likelihood of new, more virulent trypanosome strains arising that have not been previously encountered by livestock. These factors have long term impact on approaches to control livestock disease and thereby improve livestock health. Therefore, in the long term, the research benefits livestock farmers and governmental bodies responsible for livestock health in countries in sub-Saharan Africa, and international bodies developing strategies for disease control. Understanding the genetic basis of drug resistance enables the rational deployment of existing drugs with common resistance mechanisms, and informs the development of new drugs.

Who might benefit?
The results of the research will be disseminated to veterinary practitioners and personnel of government departments and non-governmental organisations involved in control of livestock diseases, and to those with affected or at risk livestock such as farmers, ranchers and small-holders in sub-Saharan Africa. To reach this audience, we will report our work in both academic and non-academic publications, and on specialist animal health websites. The World Organization for Animal Health (OIE; http://www.oie.int/) acts as a central authority and source of information on animal diseases worldwide. The annual ISCTRC (International Scientific Council for Trypanosomiasis Research and Control) meeting held in sub-Saharan Africa is an important forum to reach those engaged in research and control of animal trypanosomiasis in the endemic regions.

We will engage with the general public and school children about research in parasitology via events co-ordinated by the Public Engagement team at University of Bristol, such as the annual Festival of Nature and Bristol Open Doors day. We will develop parasitology-themed displays relevant to this project.

How might they benefit?
Beneficiaries will gain increased awareness and understanding of animal trypanosomiasis and parasitology in general. The research highlights the fact that pathogen strains are not static and that it is possible for new pathogen strains to be generated by recombination between different pathogen strains. The resulting strain diversity may have impacts on the epidemiology and pathogenicity of the disease observed in the field. The research also provides information on how genes, such as those for drug resistance, are spread through pathogen populations by genetic exchange. This emphasizes the need to administer drugs correctly to avoid the development of drug resistance and feeds into strategies to control disease by rational deployment of existing drugs.