Tsetse fly tolerance to T. brucei infection: transcriptome analysis of trypanosome-associated changes in the tsetse fly salivary gland

Abstract

BackgroundFor their transmission, African trypanosomes rely on their blood feeding insect vector, the tsetse fly (Glossina sp.). The ingested Trypanosoma brucei parasites have to overcome a series of barriers in the tsetse fly alimentary tract to finally develop into the infective metacyclic forms in the salivary glands that are transmitted to a mammalian host by the tsetse bite. The parasite population in the salivary gland is dense with a significant number of trypanosomes tightly attached to the epithelial cells. Our current knowledge on the impact of the infection on the salivary gland functioning is very limited. Therefore, this study aimed to gain a deeper insight into the global gene expression changes in the salivary glands of Glossina morsitans morsitans in response to an infection with the T. brucei parasite. A detailed whole transcriptome comparison of midgut-infected tsetse with and without a mature salivary gland infection was performed to study the impact of a trypanosome infection on different aspects of the salivary gland functioning and the mechanisms that are induced in this tissue to tolerate the infection i.e. to control the negative impact of the parasite presence. Moreover, a transcriptome comparison with age-matched uninfected flies was done to see whether gene expression in the salivary glands is already affected by a trypanosome infection in the tsetse midgut.ResultsBy a RNA-sequencing (RNA-seq) approach we compared the whole transcriptomes of flies with a T. brucei salivary gland/midgut infection versus flies with only a midgut infection or versus non-infected flies, all with the same age and feeding history. More than 7500 salivary gland transcripts were detected from which a core group of 1214 differentially expressed genes (768 up- and 446 down-regulated) were shared between the two transcriptional comparisons. Gene Ontology enrichment analysis and detailed gene expression comparisons showed a diverse impact at the gene transcript level. Increased expression was observed for transcripts encoding for proteins involved in immunity (like several genes of the Imd-signaling pathway, serine proteases, serpins and thioester-containing proteins), detoxification of reactive species, cell death, cytoskeleton organization, cell junction and repair. Decreased expression was observed for transcripts encoding the major secreted proteins such as 5′-nucleotidases, adenosine deaminases and the nucleic acid binding proteins Tsals. Moreover, expression of some gene categories in the salivary glands were found to be already affected by a trypanosome midgut infection, before the parasite reaches the salivary glands.ConclusionsThis study reveals that the T. brucei population in the tsetse salivary gland has a negative impact on its functioning and on the integrity of the gland epithelium. Our RNA-seq data suggest induction of a strong local tissue response in order to control the epithelial cell damage, the ROS intoxication of the cellular environment and the parasite infection, resulting in the fly tolera...

Thioester‐containing proteins in the tsetse fly (Glossina) and their response to trypanosome infection

“…In contrast to T. congolense, T. brucei parasites do not establish in the tsetse mouthparts but migrate to the salivary glands, where they attach to the gland epithelium and complete their development into the infective metacyclic form. Consistent with our previously reported RNA-seq results (Matetovici et al, 2016), there is strong up-regulation of different TEPs (especially TEP2) in the T. brucei-infected salivary gland. None of these TEP genes were found to be differentially expressed in the salivary glands from T. congolense-infected flies (data not shown), suggesting that the TEP response is locally induced by the parasite in the gland.…”

“…In T. brucei ‐infected salivary glands, the parasites tightly attach with their flagellum to the epithelial lining (Vickerman et al ., ), thereby possibly damaging the septate junctions. Increased expression of Gmm_TEP4 , together with the other eight genes encoding for septate junction proteins (Matetovici et al ., ), would consequently be needed to repair and maintain the structural integrity of this tissue. Gmm _ TEP4 up‐regulation in salivary glands was not observed in T. congolense ‐infected flies (data not shown), again demonstrating the specificity of this parasite‐related response.…”

“…In the tsetse fly ( Glossina spp.) we have recently demonstrated an increased expression of some TEPs in Trypanosoma brucei ‐infected salivary glands (Matetovici et al ., ), which suggests a possible role of this protein family in controlling the parasite population in this micro‐environment. Tsetse flies ( Glossina spp.)…”

“…The Gmm_TEP7 gene was identified de novo by querying the scaffold sequence (Gmor-Y1:scf7180000652159) for open reading frames (ORFs) with the ORF FINDER program available from the National Center for Biotechnology Information (NCBI; Supporting Information Figs S1 and S2). Exon-intron structures were predicted using GENSCAN software and validated by visual inspection in INTEGRATIVE GENOMICS VIEWER software (Thorvaldsdottir et al, 2013) of tsetse fly RNA sequencing (RNA-seq) mapped reads (Matetovici et al, 2016). The coordinates of the seven G. m. morsitans genes encoding thioester-containing proteins are listed in Supporting Information Table S1.…”

The crystal structure and localization ofTrypanosoma bruceiinvariant surface glycoproteins suggest a more permissive VSG coat in the tsetse-transmitted metacyclic stage

“…During a blood meal, the infected tsetse inoculates MCFs into the host along with saliva 24,32 to inhibit the host hemostatic response to the cutaneous trauma 33,34 . However, during parasite development in the tsetse SGs, there is a drastic (~80%) transcriptional down regulation of genes encoding for salivary proteins, which induces a feeding phenotype that may amplify disease transmission 32,35,36 . Although the proteome of tsetse saliva from T. brucei -infected flies has been recently determined 34 , soluble parasite factors have not been characterized in detail and thus, the molecular mechanism by which these parasites manipulate the vector to promote transmission is largely unknown.…”