Comprehensive Quantitative Proteome Analysis of Aedes aegypti Identifies Proteins and Pathways Involved in Wolbachia pipientis and Zika Virus Interference Phenomenon

Abstract

Zika virus (ZIKV) is a global public health emergency due to its association with microcephaly, Guillain-Barré syndrome, neuropathy, and myelitis in children and adults. A total of 87 countries have had evidence of autochthonous mosquito-borne transmission of ZIKV, distributed across four continents, and no antivirus therapy or vaccines are available. Therefore, several strategies have been developed to target the main mosquito vector, Aedes aegypti, to reduce the burden of different arboviruses. Among such strategies, the use of the maternally-inherited endosymbiont Wolbachia pipientis has been applied successfully to reduce virus susceptibility and decrease transmission. However, the mechanisms by which Wolbachia orchestrate resistance to ZIKV infection remain to be elucidated. In this study, we apply isobaric labeling quantitative mass spectrometry (MS)-based proteomics to quantify proteins and identify pathways altered during ZIKV infection; Wolbachia infection; co-infection with Wolbachia/ZIKV in the A. aegypti heads and salivary glands. We show that Wolbachia regulates proteins involved in reactive oxygen species production, regulates humoral immune response, and antioxidant production. The reduction of ZIKV polyprotein in the presence of Wolbachia in mosquitoes was determined by MS and corroborates the idea that Wolbachia helps to block ZIKV infections in A. aegypti. The present study offers a rich resource of data that may help to elucidate mechanisms by which Wolbachia orchestrate resistance to ZIKV infection in A. aegypti, and represents a step further on the development of new targeted methods to detect and quantify ZIKV and Wolbachia directly in complex tissues.

Interspecies isobaric labeling-based quantitative proteomics reveals protein changes in the ovary of Aedes aegypti co-infected with ZIKV and Wolbachia

“…At 14 days post-infection (dpi), each mosquito ovary was extracted from the body according to the previous method (Charlwood et al, 2018). Protein extraction, digestion, iTRAQ labeling and LC-MS/MS protocol were performed as described previously (Martins et al, 2021). Proteins were extracted by lysis with buffer (7 M urea, 2 M thiourea, 50 mM HEPES pH 8, 75 mM NaCl, 1 mM EDTA, 1 mM PMSF) with the addition of a protease/phosphatase inhibitor cocktail (Roche).…”

“…After transinfection, Wolbachia is currently located in several tissues and organs of Ae. aegypti mosquitoes and the mechanisms resulting in viral blocking are under investigation Most likely it involves the activation of the immune system by oxidative stress and down-regulation of host proteins involved in pathways that would help to produce resources essential to the virus life cycle (Ford et al, 2020; Martins et al, 2021; Ogunlade et al, 2021; Pimentel et al, 2021).…”

“…A diversity of quantitative methods have already been successfully applied in insects to analyze differentially regulated proteins (De Mandal et al, 2020; García-Robles et al, 2020; Serteyn et al, 2020), including a previous study from our group, where it was able to elucidate different mechanisms responses to ZIKV and Wolbachia infection in Ae. aegypti head and salivary glands (Martins et al, 2021).…”

Impacts of fungal entomopathogens on survival and immune responses of Aedes albopictus and Culex pipiens mosquitoes in the context of native Wolbachia infections

“…albopictus into Aedes aegypti, impaired the mosquito's ability to transmit dengue [9,16]. Although the main mechanism of Wolbachia-derived viral suppression is not clearly established [17,18], changes in the mosquito immune system [19,20], Wolbachia-virus competition and Wolbachia modulation of the mosquito methylation patterns have been suggested as potential mechanisms of viral suppression [17,[21][22][23][24]. In addition to interfering with the replication and transmission of several arboviruses [9], Wolbachia has been shown to reduce filarial [25] and Plasmodium infection in mosquitoes [9,26] and to interact with the host native microbiota [27], resulting in increased host fitness [28,29].…”

Evaluation of Total Female and Male Aedes aegypti Proteomes Reveals Significant Predictive Protein–Protein Interactions, Functional Ontologies, and Differentially Abundant Proteins

“…Proteomics also identified notable proteins differentiated during Zika infections when analysing Wolbachia/Zika coinfection in Ae. aegypti heads and salivary glands [16]. Moreover, proteomics has deciphered some of the possible mechanisms involved in the occurrence of insecticide resistance in mosquitoes, leading to the potential discovery of biomarkers for detecting resistance in field mosquito populations [17][18][19][20][21].…”