Sand flies, which transmit diseases like leishmaniases, bartonellosis, and certain viruses, pose a significant public health threat. Our research focuses on the immune responses of Lutzomyia longipalpis, the primary vector for visceral leishmaniasis in the Americas. We use L. longipalpis LL5 cells as a model to study how sand flies respond to pathogens. These cells exhibit robust immune reactions, producing molecules mainly regulated by the Toll, IMD, Jak-STAT, and RNAi pathways. In previous studies, we detected a non-specific antiviral response in LL5 cells following double-stranded RNAs (dsRNAs) transfection. A previous complete secretome of these cells showed molecules resembling an interferon-like antiviral response when transfected with polyinosinic-polycytidylic acid (poly I:C), a synthetic dsRNA analog. In the current study, we analyzed soluble proteins secreted by LL5 cells after poly I:C transfection. Using comparative mass spectrometry, we examined protein composition of conditioned media depleted of exosomes at 24 h and 48 h. Most proteins uniquely expressed in the transfected groups had low abundance compared to the overall expressed proteins. Interactome prediction analysis revealed that at 24 h, the proteins uniquely found in the secretome of the transfected group were involved in RNA degradation and purine metabolism, while at 48 h they were linked to ribosomal proteins and signaling pathways such as Hedgehog, Transforming Growth Factor-beta (TGF-β), and Wingless/integrated (Wnt). We highlight increased abundance of the TGF-β-induced protein ig-h3 (24 h and 48 h), a Toll-like receptor 3 (48 h), and a hemocytin (48 h) in the secretion of transfected groups compared to the controls. We also performed an interaction analysis of proteins more secreted by the treated group at 24 h and 48 h. Unlike the interactome of uniquely identified proteins, few interactions were observed at 24 h, with a predominance of extracellular matrix and cell adhesion proteins. The set of proteins more secreted at 48 h presented more interactions than at 24 h, with emphasis on catabolic processes, including RNA degradation. These findings indicate that poly I:C transfection in LL5 cells induces the secretion of proteins involved in cellular defense and repair, revealing molecules involved in the LL5 non-specific antiviral response.

• Klíčová slova
• RNA degradation, cell repair, non-specific antiviral response, poly I:C, sand fly cell line, secreted proteins,
• MeSH
• buněčné linie MeSH
• dvouvláknová RNA * genetika   MeSH
• hmyzí proteiny * metabolismus   MeSH
• poly I-C MeSH
• proteom * analýza   MeSH
• proteomika MeSH
• Psychodidae * imunologie   metabolismus   genetika   MeSH
• sekretom * MeSH
• signální transdukce MeSH
• transfekce MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• dvouvláknová RNA * MeSH
• hmyzí proteiny * MeSH
• poly I-C MeSH
• proteom * MeSH

Traditionally, insects have been thought to be entirely dependent on their innate immune system, which has little capacity for the acquisition of experience from previous infections. However, much experimental evidence has challenged this view, showing that insects can develop long-term, pathogen-specific immune memory, which in some cases can be transmitted to offspring. Although significant progress has been made in this area, the underlying mechanism is still not fully understood, and a number of fundamental questions remain unanswered. In this review, we present an overview of documented cases of insect immune memory and summarize the experimental evidence in support of the prevailing hypotheses on the mechanism of antiviral and antibacterial immune memory in insects. We also highlight key questions that remain unanswered and discuss Drosophila melanogaster as a powerful model organism for investigating the mechanisms of innate immune memory formation. Finally, we evaluate the significance of this research and explore the potential for insect vaccination.

• Klíčová slova
• Dscam, evolutionary biology, hemocytes, immune memory, immunology, inflammation, insect immunity, insect vaccination, trained immunity,
• MeSH
• Drosophila melanogaster * imunologie   MeSH
• hmyz * imunologie   MeSH
• imunologická paměť * MeSH
• přirozená imunita * MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Up to 170 tick-borne viruses (TBVs) have been identified to date. However, there is a paucity of information regarding TBVs and their interaction with respective vectors, limiting the development of new effective and urgently needed control methods. To overcome this gap of knowledge, it is essential to reproduce transmission cycles under controlled laboratory conditions. In this study we assessed an artificial feeding system (AFS) and an immersion technique (IT) to infect Ixodes ricinus ticks with tick-borne encephalitis (TBE) and Kemerovo (KEM) virus, both known to be transmitted predominantly by ixodid ticks. Both methods permitted TBEV acquisition by ticks and we further confirmed virus trans-stadial transmission and onward transmission to a vertebrate host. However, only artificial feeding system allowed to demonstrate both acquisition by ticks and trans-stadial transmission for KEMV. Yet we did not observe transmission of KEMV to mice (IFNAR-/- or BALB/c). Artificial infection methods of ticks are important tools to study tick-virus interactions. When optimally used under laboratory settings, they provide important insights into tick-borne virus transmission cycles.

• MeSH
• arachnida jako vektory fyziologie   virologie   MeSH
• interakce hostitele a patogenu MeSH
• klíště fyziologie   virologie   MeSH
• klíšťová encefalitida přenos   virologie   MeSH
• lidé MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• Orbivirus fyziologie   MeSH
• reovirové infekce přenos   virologie   MeSH
• virologie metody   MeSH
• viry klíšťové encefalitidy fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• práce podpořená grantem MeSH

The Drosophila melanogaster sigma virus, a member of the Rhabdoviridae family, specifically propagates itself in D. melanogaster. It contains six genes in the order of 3'-N-P-X-M-G-L-5'. The sigma virus is the only arthropod-specific virus of the Rhabdoviridae family. Sigma-virus-infected Drosophila may suffer from irreversible paralysis when exposed to a high CO2 concentration, but generally, no other symptoms are reported. A recent study reported that host gene expression in immune pathways was not changed in sigma-virus-infected Drosophila, which does not necessarily suggest that they are not involved in virus-host interactions. The present study aimed to identify host genes associated with sigma virus replication. Immune pathways JAK-STAT and IMD were selected for detailed study. The results showed that the genome copy number of the sigma virus increased after knocking down the immune pathway genes domeless and PGRP-LC in Drosophila S2 cells. The knocking down of domeless and PGRP-LC significantly up-regulated the expression of the L gene compared to the other viral genes. We propose that the immune pathways respond to sigma virus infection by altering L expression, hence suppressing viral replication. This effect was further tested in vivo, when D. melanogaster individuals injected with dsdome and dsPGRP-LC showed not only an increase in sigma virus copy number, but also a reduced survival rate when treated with CO2. Our study proved that host immunity influences viral replication, even in persistent infection. Knocking down the key components of the immune process deactivates immune controls, thus facilitating viral expression and replication. We propose that the immunity system of D. melanogaster regulates the replication of the sigma virus by affecting the L gene expression. Studies have shown minimal host-virus interaction in persistent infection. However, our study demonstrated that the immunity continued to affect viral replication even in persistent infection because knocking down the key components of the immune process disabled the relevant immune controls and facilitated viral expression and replication.

• Klíčová slova
• IMD pathway, JAK-STAT pathway, RNA interference, sigma virus,
• Publikační typ
• časopisecké články MeSH

BACKGROUND: Ixodid ticks are important vectors of a wide variety of viral, bacterial and protozoan pathogens of medical and veterinary importance. Although several studies have elucidated tick responses to bacteria, little is known about the tick response to viruses. To gain insight into the response of tick cells to flavivirus infection, the transcriptomes and proteomes of two Ixodes spp cell lines infected with the flavivirus tick-borne encephalitis virus (TBEV) were analysed. METHODS: RNA and proteins were isolated from the Ixodes scapularis-derived cell line IDE8 and the Ixodes ricinus-derived cell line IRE/CTVM19, mock-infected or infected with TBEV, on day 2 post-infection (p.i.) when virus production was increasing, and on day 6 p.i. when virus production was decreasing. RNA-Seq and mass spectrometric technologies were used to identify changes in abundance of, respectively, transcripts and proteins. Functional analyses were conducted on selected transcripts using RNA interference (RNAi) for gene knockdown in tick cells infected with the closely-related but less pathogenic flavivirus Langat virus (LGTV). RESULTS: Differential expression analysis using DESeq resulted in totals of 43 and 83 statistically significantly differentially-expressed transcripts in IDE8 and IRE/CTVM19 cells, respectively. Mass spectrometry detected 76 and 129 statistically significantly differentially-represented proteins in IDE8 and IRE/CTVM19 cells, respectively. Differentially-expressed transcripts and differentially-represented proteins included some that may be involved in innate immune and cell stress responses. Knockdown of the heat-shock proteins HSP90, HSP70 and gp96, the complement-associated protein Factor H and the protease trypsin resulted in increased LGTV replication and production in at least one tick cell line, indicating a possible antiviral role for these proteins. Knockdown of RNAi-associated proteins Argonaute and Dicer, which were included as positive controls, also resulted in increased LGTV replication and production in both cell lines, confirming their role in the antiviral RNAi pathway. CONCLUSIONS: This systems biology approach identified several molecules that may be involved in the tick cell innate immune response against flaviviruses and highlighted that ticks, in common with other invertebrate species, have other antiviral responses in addition to RNAi.

• MeSH
• arachnida jako vektory * genetika   metabolismus   virologie   MeSH
• buněčné linie MeSH
• genový knockdown MeSH
• klíště * genetika   imunologie   metabolismus   virologie   MeSH
• přirozená imunita MeSH
• proteomika * MeSH
• RNA interference MeSH
• stanovení celkové genové exprese * MeSH
• viry klíšťové encefalitidy imunologie   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH