By David N. Leff

Half a million people in central Africa are under death sentences with virtually no appeal. Their implacable verdict doesn¿t come from AIDS or malaria, but from incurable sleeping sickness.

These victims represent collateral damage in biowarfare between humans (Homo sapiens) and the tsetse fly (Diptera:Glossinidae). The word tsetse derives from the term an African tribe coined, related to blood-sucking and death among people and cattle.

Then there¿s nagana, a Zulu term describing domestic livestock and wild hoofed mammals fatally bitten by the tsetse fly.

Tsetse is a brownish, half-inch-long airborne vector, which lives exclusively on mammalian blood. To obtain this sanguine meal ticket, the fly penetrates its victim¿s skin with a sawtooth nose cone that lacerates open a freely bleeding wound. As payback for this sanguine fix, tsetse squirts from its saliva a swarm of pathogenic parasites called trypanosomes.

¿African trypanosomes cover a fairly broad range of organisms,¿ observed molecular entomologist Sarap Aksoy, at Yale University School of Medicine. ¿For example, we have Trypanosoma brucei, which transmits the sickness to humans, and T. congolense, the cause of devastating diseases in animals.¿

In human populations, the sleeping-sickness trend is on the rise.

¿It¿s absolutely devastating,¿ Aksoy stated. ¿In June we had our meeting at the World Health Organization to re-evaluate the trypanosomiasis program,¿ she recalled, ¿and many of our colleagues cited the various epidemics going on that are unknown to the health systems. Conservatively,¿ Aksoy continued, ¿there are 500,000 people actively infected with the disease symptoms, and many more millions at risk who probably are not diagnosed currently. There¿s no vaccine or therapy for the disease, so unless they are treated immediately, these people will die. And nagana is such a scourge to the economy and livelihood of people that whole areas have given up on raising livestock. There are some trypanosome-resistant or -tolerant cattle. It¿s an ongoing field of research to develop trypano-tolerant breeds.¿

Blowing Tsetse¿s Immune System Cover

Aksoy is senior author of a paper in the Proceedings of the National Academy of Sciences (PNAS) dated Oct. 9, 2001. Its title: ¿Tsetse immune responses and trypanosome transmission: Implications for the development of tsetse-based strategies to reduce trypanosomiasis.¿

¿This article reports for the first time that we have gained some understanding of tsetse immune responses,¿ Aksoy told BioWorld Today. ¿Previously, nothing was known from an immune perspective about the basis for trypanosome transmission.

¿Most insects have a pretty robust immune system of their own,¿ she pointed out. ¿They can clear most infectious agents that they acquire either through ingesting blood or encountering something that somehow invades their body. But parasites for some reason ¿ in this case African trypanosomes ¿ have been able to evade these natural immune responses. To understand this, we first had to discover what some of those responses are in tsetse.

¿We have now shown that the insect actually is eliciting a very strong immune reaction,¿ Aksoy went on, ¿but somehow this response is not able to clear the infectious trypanosomes. Either the parasite is resistant to these immune molecules, or is hiding in specific tissues or organs of the insect, where they¿re not affected by its immune mechanism. In our paper we show some tentative data that localization of the parasites is key. That if we can synthesize these immune products in the vicinity of where the parasites are hiding, we can possibly affect parasite transmission in the flies.¿

Oddly, tsetse makes life difficult for the parasites it harbors to continue their life cycle of transmission to the next mammalian target. The trypanosoma sulk for a time, while replicating, in the insect¿s midgut on their way to the salivary download. In the insect¿s immune-system fat body, adjacent to that gut, three genes ¿ defensin, attacin and diptericin ¿ express antibacterial peptides.

¿Typically, insects will not harbor these peptides unless there is an invader,¿ Aksoy observed. ¿But in tsetse flies,¿ she recounted, ¿we¿re finding that diptericin ¿ which is specific against Gram-negative bacteria ¿ is synthesized in the insects, even in the absence of any immune challenge. And we think this might be one of the responses the fly has against its own symbiotic bacteria. When we looked at the activity of those bacteria against this peptide we found them to be significantly more resistant than their closely related, free-living organisms, such as E. coli.

¿The general dogma has been that these symbiotic bacteria are somehow not causing any reaction in insects,¿ Aksoy went on. ¿Rather, they are regarded as a part of the insect. But maybe not so,¿ she added, ¿because we clearly saw that there were high expression levels of diptericins. We haven¿t conclusively shown that that is due to these symbiotic bacteria. It¿s a direction we¿re going to be taking in the lab, asking: What is the cost to the fly of harboring these symbiotic organisms?¿

Antibacterial Genes Rise To E. coli Challenge

¿We cloned the genes for diptericins and attacins, isolated them from tsetse, then used them as probes to look at their expression under a variety of conditions, compared with the normal fly. We microinjected pathogens into tsetse¿s hemolymph ¿ the equivalent of blood ¿ then looked at their expression. We also fed these parasites to the fly, which would be the more natural route. Trypanosomes are acquired by ingestion. Lastly, we examined flies that had thousands of trypanosome infections. Initially, we didn¿t see expression of these immune response genes in tsetse.

¿But when we gave tsetse very high levels of E. coli,¿ Aksoy narrated, ¿immediately we saw very high expression of these antibacterial genes in the fly. So that,¿ she pointed out, ¿is how trypanosomes do not initially turn on the immune response of their host.¿

The coauthors¿ goal is to come up with a strategy that cuts tsetse¿s parasite transmission off at the knees.

¿When we immune-challenged flies before giving them an infected blood meal,¿ Aksoy related, ¿we reduced infections drastically. When we challenged tsetse by microinjections of E. coli, followed by an infectious blood meal, we could drastically reduce parasite infections. So that gives us additional confidence that if we can express these molecules in response to the immune challenge, then we can block parasite transmission therapeutically.¿

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