Correlation between structure, protein composition, morphogenesis and cytopathology of Glossina pallidipes salivary gland hypertrophy virus

Abstract

The Glossina pallidipes salivary gland hypertrophy virus (GpSGHV) is a dsDNA virus with rodshaped, enveloped virions. Its 190 kb genome contains 160 putative protein-coding ORFs. Here, the structural components, protein composition and associated aspects of GpSGHV morphogenesis and cytopathology were investigated. Four morphologically distinct structures: the nucleocapsid, tegument, envelope and helical surface projections, were observed in purified GpSGHV virions by electron microscopy. Nucleocapsids were present in virogenic stroma within the nuclei of infected salivary gland cells, whereas enveloped virions were located in the cytoplasm. The cytoplasm of infected cells appeared disordered and the plasma membranes disintegrated. Treatment of virions with 1 % NP-40 efficiently partitioned the virions into envelope and nucleocapsid fractions. The fractions were separated by SDS-PAGE followed by in-gel trypsin digestion and analysis of the tryptic peptides by liquid chromatography coupled to electrospray and tandem mass spectrometry. Using the MaxQuant program with Andromeda as a database search engine, a total of 45 viral proteins were identified. Of these, ten and 15 were associated with the envelope and the nucleocapsid fractions, respectively, whilst 20 were detected in both fractions, most likely representing tegument proteins. In addition, 51 host-derived proteins were identified in the proteome of the virus particle, 13 of which were verified to be incorporated into the mature virion using a proteinase K protection assay. This study provides important information about GpSGHV biology and suggests options for the development of future anti-GpSGHV strategies by interfering with virus-host interactions.

Comprehensive annotation of Glossina pallidipes salivary gland hypertrophy virus from Ethiopian tsetse flies: a proteogenomics approach

“…We used a combination of proteogenomic and transcriptomic mapping approaches (Nesvizhskii, 2014) to complement the annotation of ORFs in the GpSGHVEth genome. In addition to using peptides identified from flies infected by GpSGHV-Eth, we also included data obtained from previous proteomics of the GpSGHVUga (Kariithi et al, , 2011(Kariithi et al, , 2013b(Kariithi et al, , 2016. Moreover, we compared the GpSGHV-Eth genome to the GpSGHVUga genome, which was annotated in silico (Abd-Alla et al, 2008).…”

“…Out of the 86 proteins, 10, 15, 20 and 12 were classified as envelope, nucleocapsid, tegument and virion proteins, respectively, based on the detection of their homologues in highly purified GpSGHV-Uga virions by LC-MS/MS (Kariithi et al, 2013b). The remaining 29 of the 86 proteins without specific localization were designated infected cell-specific viral proteins (ICSVPs).…”

“…Taking into account the proteomic datasets from GpSGHV-Uga (Kariithi et al, , 2011(Kariithi et al, , 2013b(Kariithi et al, , 2016 and the strong homologies between the two virus isolates (Table S1), additional homologous ORFs could be functionally annotated in the GpSGHV-Eth genome (Table 1). Furthermore, of these 79 ORFs, 68 had appropriately upstream positioned TATAA-like box and/or (G/T/A) TAAG transcriptional signals of early and late baculovirus genes, respectively (Chen et al, 2013), whilst 70 ORFs had poly(A) signals.…”

“…Preparation of G. pallidipes salivary gland proteins and MS analyses were performed as described by Kariithi et al (2013b). Briefly, proteins were extracted from HSG extracts, fractionated using 12 % SDS-PAGE and subjected to in-gel trypsin digestion.…”

Antiviral drug valacyclovir treatment combined with a clean feeding system enhances the suppression of salivary gland hypertrophy in laboratory colonies of Glossina pallidipes

“…Following these basic studies, several virus management approaches were suggested [14] and the effect of two of these strategies on the prevalence of SGH in colonies of G. pallidipes was recently published i.e. adding the antiviral drug valacyclovir to the blood diet of the flies [15,16] and the application of a clean feeding strategy (each fly received a clean fresh blood meal each time) [17,18]. Briefly, the clean feeding system can be initiated in a large-scale colony by selecting as many cages of teneral flies from the main colony as can be fed at one time on the available clean membranes and thereafter these flies and their progeny should always be fed first on the fresh, clean blood (that has not been supplied to other flies).…”