Uma aplicação numérica de modelos da dengue na presença da Wolbachia e da vacinação na Tailândia.

Dengue virus (DENV) is the mosquito-borne arbovirus with the widest impact on human health. Although its dispersal is partially conditioned by the environmental constraints that limit the distribution of its main vector (Aedes aegypti), DENV has been relentlessly taking over the planet, especially in the last decades. Despite the fact that it is mainly associated with the tropical and subtropical regions, it affects up to 1/3 of the world population. In the absence of any prophylactic vaccine or specific therapeutics, vector control remains the best alternative to restrain its circulation. However, most viral infections are either clinically silent, or expressed as a non-specific fever syndrome. Therefore, viral activity may remain undetected. Moreover, the establishment of thriving vector populations in periurban environments brings humans and viruses closer together, and opens the possibility for the occurrence of unexpected outbreaks. Such was the case of Madeira in 2012-2013. In addition to its impact on the health of the local population, health services and economy, this outbreak revealed how difficult it may be to control the circulations of pathogenic arboviruses, especially taking into considerations that Europe is already partially colonized by another DENV vector (Aedes albopictus).

ABSTRACTCertain strains of the intracellular endosymbiont Wolbachia can strongly inhibit or block the transmission of viruses such as dengue virus (DENV) by Aedes mosquitoes, and the mechanisms responsible are still not well understood. Direct infusion and liquid chromatography-Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry-based lipidomics analyses were conducted using Aedes albopictus Aa23 cells that were infected with the wMel and wMelPop strains of Wolbachia in comparison to uninfected Aa23-T cells. Substantial shifts in the cellular lipid profile were apparent in the presence of Wolbachia. Most significantly, almost all sphingolipid classes were depleted, and some reductions in diacylglycerols and phosphatidylcholines were also observed. These lipid classes have previously been shown to be selectively enriched in DENV-infected mosquito cells, suggesting that Wolbachia may produce a cellular lipid environment that is antagonistic to viral replication. The data improve our understanding of the intracellular interactions between Wolbachia and mosquitoes.IMPORTANCE Mosquitoes transmit a variety of important viruses to humans, such as dengue virus and Zika virus. Certain strains of the intracellular bacterial genus called Wolbachia found in or introduced into mosquitoes can block the transmission of viruses, including dengue virus, but the mechanisms responsible are not well understood. We found substantial shifts in the cellular lipid profiles in the presence of these bacteria. Some lipid classes previously shown to be enriched in dengue virus-infected mosquito cells were depleted in the presence of Wolbachia, suggesting that Wolbachia may produce a cellular lipid environment that inhibits mosquito-borne viruses.

BackgroundWolbachia pipientis is a common endosymbiotic bacterium of arthropods that strongly inhibits dengue virus (DENV) infection and transmission in the primary vector, the mosquito Aedes aegypti. For that reason, Wolbachia-infected Ae. aegypti are currently being released into the field as part of a novel strategy to reduce DENV transmission. However, there is evidence that DENV can be transmitted vertically from mother to progeny, and this may help the virus persist in nature in the absence of regular human transmission. The effect of Wolbachia infection on this process had not previously been examined.ResultsWe challenged Ae. aegypti with different Brazilian DENV isolates either by oral feeding or intrathoracic injection to ensure disseminated infection. We examined the effect of Wolbachia infection on the prevalence of DENV infection, and viral load in the ovaries. For orally infected mosquitoes, Wolbachia decreased the prevalence of infection by 71.29%, but there was no such effect when the virus was injected. Interestingly, regardless of the method of infection, Wolbachia infection strongly reduced DENV load in the ovaries. We then looked at the effect of Wolbachia on vertical transmission, where we observed only very low rates of vertical transmission. There was a trend towards lower rates in the presence of Wolbachia, with overall maximum likelihood estimate of infection rates of 5.04 per 1000 larvae for mosquitoes without Wolbachia, and 1.93 per 1000 larvae for Wolbachia-infected mosquitoes, after DENV injection. However, this effect was not statistically significant.ConclusionsOur data support the idea that vertical transmission of DENV is rare in nature, even in the absence of Wolbachia. Indeed, we observed that vertical transmission rates were low even when the midgut barrier was bypassed, which might help to explain why we only observed a trend towards lower vertical transmission rates in the presence of Wolbachia. Nevertheless, the low prevalence of disseminated DENV infection and lower DENV load in the ovaries supports the hypothesis that the presence of Wolbachia in Ae. aegypti would have an effect on the vertical transmission of DENV in the field.

Risk for Emergence of Dengue and Chikungunya Virus in Israel

The purpose of study was to isolate arboviruses from mosquitoes of different species in the cell culture and to identify them by using molecular and immunochemical techniques.Materials and methods. Viruses were isolated in C6/36 cell cultures. The pathogens were identified by using enzyme-linked immunosorbent assay (ELISA) kits for detection of antigens of dengue, Chikungunya, West Nile and Sindbis viruses as well as the reverse transcription polymerase chain reaction (RT-PCR) with specific primers and Sanger sequencing.Results. A total of 102 mosquitoes belonging to three genera, Culex spp, Culiseta spp., Aedes spp., were studied. Mosquitoes of each species or genus were divided into pools, each containing 4–5 mosquitoes. The study of suspensions of only 2 mosquito pools obtained from Aedes aegypti and Aedes albopictus, starting from the 3rd passage, showed changes in the C6/36 cell monolayer. Starting from the 4th passage, an antigen of Chikungunya virus was detected using ELISA test in the suspension obtained from the Aedes albopictus pool. Dengue virus was detected in the 5th passage from the materials obtained from the Aedes aegypti pool. Thus, antigens of the Chikungunya and dengue viruses were detected only in 2 of 23 examined pools of mosquitoes of different genera. Materials of the 5th passage were analyzed by RT-PCR with specific primers for dengue and Chikungunya viruses. It was confirmed that the isolate obtained from Aedes albopictus mosquitoes contained RNA of the Chikungunya virus and corresponded to the East/Central/South African genotype, while the isolate obtained from Aedes aegypti mosquitoes contained RNA of the dengue type 2 virus.Conclusion. The obtained nucleotide sequences of the Chikungunya virus were deposited in the GenBank international database under accession numbers MN271691 and MN271692.

Viral hemorrhagic fever viruses

Molecular investigations of dengue virus during outbreaks in Orissa state, Eastern India from 2010 to 2011

Dengue in the Americas—time to talk

Dengue is one of the most significant health problems that has magnified its impact globally by affecting about 390 million people annually across 110 countries. The causative agent of this life-threating disease is a positive single stranded RNA arbovirus known as dengue virus (DENV), which uses species of Aedes mosquitoes as vectors. To date, there is no effective available vaccine or cure for dengue, and the control options primarily rely on vector control strategies, mostly through the application of pesticides. However, reports of resistance in Aedes mosquitoes against pesticides has limited this option as well. Therefore, there is an urgent need for alternative approaches to control the spread of DENV. One of the novel options involves the use of the endosymbiotic bacterium Wolbachia, that has successfully limited the ability of Aedes aegypti mosquitoes to transmit a number of life-threatening mosquito-borne viruses such as DENV and Zika virus. Despite its effectiveness to inhibit replication of DENV, very little is known about the mechanism(s) that Wolbachia uses to impart this antiviral effect. In this study, we looked into Ae. aegypti host factors that affect DENV replication and their potential manipulations by Wolbachia to find molecular mechanism(s) that Wolbachia utilizes to limit DENV replication. Recently, studies have shed light on the role of chromodomain DNA binding helicases in Human Immunodeficiency virus (HIV) and Influenza A virus. In Chapter 1, we have identified three Ae. aegypti homologs of chromodomain helicase DNA binding proteins (CHD) and determined their modulation in response to Wolbachia and DENV infections. We have found that among the three CHD members, AeCHD7/Kismet levels are significantly decreased in Wolbachia infection. Further investigations demonstrated that AeCHD7 is significantly increased in the case of DENV replication suggesting that it may facilitate DENV replication. Knock down studies of AeCHD7 confirmed this assumption as it resulted in significant reductions in DENV replication and virion production. In this study, we have identified AeCHD7 as an Ae. aegypti pro-DENV host factor that is downregulated by Wolbachia which may contribute in limiting DENV replication. Vago is an insect-specific secretory protein that has been identified in Culex quinquefasciatus to play an important role in the crosstalk between the mosquito’s immune pathways and reduce West Nile virus (WNV) replication. In Chapter 2, by in-silico identification of Vago characteristic SVWC domain and secretory signal, we identified two potential homologs of the Vago protein in Ae. aegypti and looked at their expression pattern in the case of Wolbachia infection to find that AeVago1 is highly induced in Ae. aegypti upon Wolbachia infection. However, we found no induction of AeVago1 expression in Ae. aegypti mosquitoes infected with DENV. Further, AeVago1 knockdown studies demonstrated that there was a significant increase in DENV replication in Wolbachia infected cells in AeVago1 deficient cells. However, there was no effect on Wolbachia density in AeVago1 depleted cells. The outcomes of this study suggest that in the presence of Wolbachia the immune gene AeVago1 is induced, which might also contribute towards inhibition of DENV replication. Pelo has been recently reported as a positive regulator of Drosophila C virus (DCV) replication. However, its role in the case of DENV replication has not been elucidated yet. In Chapter 3, we looked into the possible involvement of pelo in the case of Wolbachia-Ae. aegypti-DENV interactions. We found that the pelo protein levels increase during DENV replication. Silencing of pelo led to severe reduction of DENV virion production, suggesting its important role in DENV replication. However, in the case of Wolbachia infection, specifically in female Ae. aegypti mosquitoes, there was a significant decrease in the transcript levels of pelo. Further experiments confirmed that Wolbachia changes the subcellular localization of the pelo protein, suggesting that it might be a novel Ae. aegypti host factor that is used by Wolbachia to limit DENV replication. In addition, we found that Wolbachia-mediated downregulation of pelo transcripts might be regulated by aae-miR-2940-5p, which is highly induced by Wolbachia. This study has identified a novel molecular mechanism that is used by Wolbachia to limit DENV replication in Ae. aegypti mosquitoes. Nevertheless, this mechanism of inhibition does not seem to be universally seen in Wolbachia-host-virus interactions. Overall, in this study we have identified three novel host genes that play important roles in DENV replication and that regulation of these genes in the presence of Wolbachia may contribute towards virus blocking in Ae. aegypti mosquitoes.

BackgroundMosquito research in Europe has a long history, primarily focused on malaria vectors. In recent years, invasive mosquito species like the Asian tiger mosquito (Aedes albopictus) and the spread of arboviruses like dengue virus, chikungunya virus or bluetongue virus have led to an intensification of research and monitoring in Europe. The risk of further dissemination of exotic species and mosquito-borne pathogens is expected to increase with ongoing globalization, human mobility, transport geography, and climate warming. Researchers have conducted various studies to understand the ecology, biology, and effective control strategies of mosquitoes and associated pathogens.Main bodyThree invasive mosquito species are established in Europe: Asian tiger mosquito (Aedes albopictus), Japanese bush mosquito (Ae. japonicus), and Korean bush mosquito (Aedes koreicus). Ae. albopictus is the most invasive species and has been established in Europe since 1990. Over the past two decades, there has been an increasing number of outbreaks of infections by mosquito-borne viruses in particular chikungunya virus, dengue virus or Zika virus in Europe primary driven by Ae. albopictus. At the same time, climate change with rising temperatures results in increasing threat of invasive mosquito-borne viruses, in particular Usutu virus and West Nile virus transmitted by native Culex mosquito species. Effective mosquito control programs require a high level of community participation, going along with comprehensive information campaigns, to ensure source reduction and successful control. Control strategies for container breeding mosquitoes like Ae. albopictus or Culex species involve community participation, door-to-door control activities in private areas. Further measures can involve integration of sterile insect techniques, applying indigenous copepods, Wolbachia sp. bacteria, or genetically modified mosquitoes, which is very unlike to be practiced as standard method in the near future.ConclusionsClimate change and globalization resulting in the increased establishment of invasive mosquitoes in particular of the Asian tiger mosquito Ae. albopictus in Europe within the last 30 years and increasing outbreaks of infections by mosquito-borne viruses warrants intensification of research and monitoring. Further, effective future mosquito control programs require increase in intense community and private participation, applying physical, chemical, biological, and genetical control activities.

Dengue is a mosquito-borne disease that has been an epidemic in China for many years. Aedes albopictus is the dominant Aedes mosquito species and the main vector of dengue in China. Epidemiologically, dengue mainly occurs in Guangdong Province; it does not occur or rarely occurs in other areas of mainland China. This distribution may be associated with climate, mosquito density, and other factors in different regions; however, the effect of temperature on the vector competence of Ae. albopictus for dengue viruses (DENV) remains unclear. In this study, Ae. albopictus was orally infected with dengue virus 2 (DENV-2) and reared at constant temperatures (18, 23, 28, and 32°C) and a fluctuating temperature (28–23–18°C). The infection status of the midguts, ovaries, and salivary glands of each mosquito was detected by polymerase chain reaction (PCR) at 0, 5, 10, and 15 days post-infection (dpi). DENV-2 RNA copies from positive tissues were quantified by quantitative real time PCR (qRT-PCR). At 18°C, DENV-2 proliferated slowly in the midgut of Ae. albopictus, and the virus could not spread to the salivary glands. At 23 and 28°C, DENV-2 was detected in the ovaries and salivary glands at 10 dpi. The rates of infection, dissemination, population transmission, and DENV-2 copies at 28°C were higher than those at 23°C at any time point. At 32°C, the extrinsic incubation period (EIP) for DENV-2 in Ae. albopictus was only 5 dpi, and the vector competence was the highest among all the temperatures. Compared with 28°C, at 28–23–18°C, the positive rate and the amount of DENV-2 in the salivary glands were significantly lower. Therefore, temperature is an important factor affecting the vector competence of Ae. albopictus for DENV-2. Within the suitable temperature range, the replication of DENV-2 in Ae. albopictus accelerated, and the EIP was shorter with a higher temperature. Our results provide a guide for vector control and an experimental basis for differences in the spatial distribution of dengue cases.

Plastic pollution and infectious diseases

[This corrects the article DOI: 10.1371/journal.pone.0171345.].

Background: Dengue hemorrhagic fever (DHF) is the most prevalent and fastest-growing vector-borne disease globally, with symptoms ranging from mild to severe and, in some cases, fatal. Quang Nam province in Vietnam can serve as a model for dengue epidemiological study, as it is an endemic region for DHF with a tropical climate, which significantly constrains the health system. However, there are very few epidemiological and microbiological reports on Dengue virus (DENV) serotypes in this region due to the limited availability of advanced surveillance infrastructure. Aims of the study: This study aims to (1) assess the PCR positivity rates among hospitalized patients with clinical Dengue presentation; (2) identify the circulating DENV serotypes; and (3) assess the impact of secondary DENV infections on outbreak severity by detecting the presence of DENV-specific IgG antibodies in the plasma of DENV-infected patients. Materials and methods: Blood samples from patients clinically diagnosed with DHF and admitted to Quang Nam General Hospital (2020-2022) were analyzed. RNA extraction was performed using the NKDNA/RNAprep MAGBEAD kit, followed by Multiplex Reverse Transcription real-time Polymerase Chain Reaction (MLP RT-rPCR) for DENV detection and serotype identification. Positive samples were further tested for DENV-specific IgG antibodies using an enzyme-linked immunosorbent assay (ELISA). Results: The PCR positivity rate among hospitalized patients was approximately 68% throughout the study period. A significant shift in DENV serotypes was observed, with DENV-2 initially dominant and later giving way to DENV-1. IgG was detected in nearly half of the MPL RT-rPCR-positive samples, indicating secondary DENV infections. Conclusions: Our study highlights persistent dengue prevalence and dynamic shifts in DENV serotypes in Quang Nam province, emphasizing the need for improved diagnostic strategies and timely sample collection. The significant serotype shifts and the presence of IgG in hospitalized patients suggest potential severe outcomes from recurrent DENV infections, possibly linked to antibody-dependent enhancement (ADE) effect, underscoring the importance of advanced surveillance, vector control, vaccination campaigns, and public education to predict and prevent future DHF epidemics.

Tsetse flies are vectors of African trypanosomiasis, a disease that affects both humans and animals. Trypanosomiasis remains a threat to lives and it is an impediment to socio-economic development in sub-Saharan Africa. In spite of decades of chemotherapy and vector control, the disease has not been eradicated. Parasitic drug resistance has been developed to existing drugs, while vector control strategies are expensive and unsustainable. Therefore, there is a need to explore other control approaches, such as the transformation of tsetse fly endosymbionts to render the fly refractory to trypanosome infection. This research focused on investigating the prevalence and triparty association of infection of trypanosomes with some endosymbionts of tsetse flies from Yankari Game Reserve. Tsetse flies were captured using biconical traps, identified morphologically, dissected and their entire guts were isolated and used for DNA extraction. Polymerase Chain Reaction (PCR) was used in confirming the identity of the tsetse flies by amplifying the cytochrome C oxidase-1 gene. PCR was also used to screen for the presence of endosymbionts (Sodalis glossinidius, Wolbachia, and Spiroplasma sp.) and trypanosomes. Glossina tachinoides was the only vector species identified. Trypanosome infection rate was 10.70% with Trypanosoma grayi being the most prevalent (9.78%) amongst the three trypanosome species detected. The prevalence of Wolbachia and Spiroplasma species were 2.80% and 40.8% respectively in flies. Sodalis glossinidius was not detected. There was an association between the presence of trypanosomes and Wolbachia, while no association was depicted between trypanosomes and Spiroplasma. It has been observed from this study that the presence of Wolbachia seems to favour trypanosome infections. Investigation on the Wolbachia genetic polymorphism in tsetse could help to better understand this association.