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Objectives: In Pakistan, cutaneous leishmaniasis is an emerging tropical disease and a very high number (>70%) of children are afflicted by this marring infection. This study aimed to scrutinise the prevalence, spatial distribution and socio‐demographic and behavioural risk factors associated with cutaneous leishmaniasis in children aged <5–15 years in Khyber Pakhtunkhwa. Methods: A total of 1, 559 clinically confirmed records of children diagnosed with cutaneous leishmaniasis (January–December) from 2020 and 2022 were obtained from selected district hospitals. In addition, a risk factors‐related questionnaire was administered to 1, 011 households (400 in 2020 and 611 in 2022) in nine districts during a household survey. Results: The maximum number of cutaneous leishmaniasis cases was recorded in 2022 (n = 877, 56.25%) as compared to 2020 (n = 682, 43.75%). The hospital records showed a greater number of male patients in the 2022 cohort (n = 603, 68.76%). The highest number of cases were observed in children aged 5–9 years in 2022 (n = 282, 32.16%) and 2020 (n = 255, 37.39%). In 2020 and 2022, cutaneous leishmaniasis cases showed peak aggregation in March (n = 118, 17.3%) and January (n = 322, 36.72%). From a spatial analysis, the maximum number of cutaneous leishmaniasis cases was recorded at 59–1700 m elevation in various land‐use/land‐cover and climatic regions with quaternary alluvium rock formations. A multivariate logistic regression model analysis of risk factors from the households survey suggested that age group, socio‐economic status, construction materials of the house, use of insect repellents, Afghan refugee camps in the village/district, knowledge and biting times of sand flies, frequent use of mosquito bed nets, presence of domestic animals in the house, knowledge of the transmission period and peak month of leishmaniasis infection increased the risk of acquiring cutaneous leishmaniasis (p value < 0.05). Conclusion: Our analysis demonstrated that cutaneous leishmaniasis in children is influenced by a variety of environmental, socio‐demographic and behavioural risk factors in Khyber Pakhtunkhwa. The increase in recorded cases of cutaneous leishmaniasis in children in 2022 compared to 2020 suggests that the infection likely extended to new foci in the province. [ABSTRACT FROM AUTHOR]

Background: Chikungunya is a viral disease caused by a mosquito-borne alphavirus. The acute phase of the disease includes symptoms such as fever and arthralgia and lasts 7–10 days. However, debilitating symptoms can persist for months or years. Despite the substantial impact of this disease, a comprehensive assessment of its clinical picture is currently lacking. Methods: We conducted a systematic literature review on the clinical manifestations of chikungunya, their prevalence and duration, and related hospitalization. Embase and MEDLINE were searched with no time restrictions. Subsequently, meta-analyses were conducted to quantify pooled estimates on clinical outcomes, the symptomatic rate, the mortality rate, and the hospitalization rate. The pooling of effects was conducted using the inverse-variance weighting methods and generalized linear mixed effects models, with measures of heterogeneity reported. Results: The systematic literature review identified 316 articles. Out of the 28 outcomes of interest, we were able to conduct 11 meta-analyses. The most prevalent symptoms during the acute phase included arthralgia in 90% of cases (95% CI: 83–94%), and fever in 88% of cases (95% CI: 85–90%). Upon employing broader inclusion criteria, the overall symptomatic rate was 75% (95% CI: 63–84%), the chronicity rate was 44% (95% CI: 31–57%), and the mortality rate was 0.3% (95% CI: 0.1–0.7%). The heterogeneity between subpopulations was more than 92% for most outcomes. We were not able to estimate all predefined outcomes, highlighting the existing data gap. Conclusion: Chikungunya is an emerging public health concern. Consequently, a thorough understanding of the clinical burden of this disease is necessary. Our study highlighted the substantial clinical burden of chikungunya in the acute phase and a potentially long-lasting chronic phase. Understanding this enables health authorities and healthcare professionals to effectively recognize and address the associated symptoms and raise awareness in society. Author summary: Chikungunya disease is an emerging public health concern. The disease is transmitted by mosquitoes and characterized by arthralgia and fever in the acute phase, lasting 7–10 days. Additionally, some individuals experience chronic symptoms such as arthralgia and tiredness that can last from months to years. Chikungunya is mainly present in the Americas and Asian countries, but the mosquitoes transmitting the disease are spreading to other regions due to climate change, amongst others. This increased disease threat highlights the importance of understanding chikungunya symptoms. However, there are currently no precise estimates on the prevalence of chikungunya symptoms. Therefore, we analysed the available literature on the clinical manifestations of chikungunya. We found that 75% of infected people develop symptoms, primarily characterized by arthralgia in 90% and fever in 88% of cases. Chronic symptoms affect 44% of symptomatic people, and 0.3% of patients with chikungunya die. Unfortunately, we were not able to estimate all predefined outcomes of interest because we did not find enough studies publishing on some of these, demonstrating that there is still much unknown around the clinical manifestations of chikungunya. However, the results can help healthcare workers early identifying chikungunya and raise awareness of this debilitating disease. [ABSTRACT FROM AUTHOR]

Venezuelan (VEE), eastern (EEE), and western (WEE) equine encephalitis viruses are encephalitic New World alphaviruses that cause periodic epizootic and epidemic outbreaks in horses and humans that may cause severe morbidity and mortality. Currently there are no FDA-licensed vaccines or effective antiviral therapies. Each year, there are a limited number of human cases of encephalitic alphaviruses; thus, licensure of a vaccine or therapeutic would require approval under the FDA animal rule. Approval under the FDA animal rule requires the disease observed in the animal model to recapitulate what is observed in humans. Currently, initial testing of vaccines and therapeutics is performed in the mouse model. Unfortunately, alphavirus disease manifestations in a mouse do not faithfully recapitulate human disease; the VEEV mouse model is lethal whereas in humans VEEV is rarely lethal. In an effort to identify a more appropriate small animal model, we evaluated hamsters in an aerosol exposure model of encephalitic alphavirus infection. The pathology, lethality, and viremia observed in the infected hamsters was inconsistent with what is observed in NHP models and humans. These data suggest that hamsters are not an appropriate model for encephalitic alphaviruses to test vaccines or potential antiviral therapies. [ABSTRACT FROM AUTHOR]

Japanese Encephalitis (JE) poses a significant public health threat across Asia and the Western Pacific, leading to considerable mortality and morbidity if not promptly diagnosed and treated. This flaviviral infection, transmitted by Culex mosquitoes, primarily affects children but can impact individuals of all ages. Prompt diagnosis relies on detecting Japanese Encephalitis Virus (JEV) IgM antibodies in serum or Cerebrospinal Fluid (CSF), alongside characteristic Magnetic Resonance Imaging (MRI) findings. In this MRI case series, three confirmed cases of JE are presented, showcasing the typical neuroimaging manifestations observed in affected individuals. All cases exhibited bilateral thalamic hyperintensities on T2-weighted and Fluid Attenuated Inversion Recovery (FLAIR) images, a hallmark feature of JE. Additionally, one patient demonstrated involvement of the substantia nigra and bilateral frontal cortex. The clinical implications of present study findings underscore the importance of considering JE as a differential diagnosis in patients presenting with symptoms of encephalitis, especially when MRI reveals bi-thalamic signal alterations. Early recognition and initiation of appropriate treatment, including antiviral agents, are crucial for improving outcomes and reducing mortality. This study contributes to the existing literature by reinforcing the importance of neuroimaging in diagnosing JE and highlighting the distinct MRI patterns associated with the disease. Recognising these characteristic imaging features can aid clinicians in promptly identifying and managing JE cases, thereby mitigating the associated morbidity and mortality. [ABSTRACT FROM AUTHOR]

Alphaviruses are vector-borne, medically relevant, positive-stranded RNA viruses that cause disease in animals and humans worldwide. Of this group, chikungunya virus (CHIKV) is the most significant human pathogen, responsible for generating millions of infections leading to severe febrile illness and debilitating chronic joint pain. Currently, there are limited treatments to protect against alphavirus disease; thus, there is a tremendous need to generate safe and effective vaccines. Live-attenuated vaccines (LAVs) are cost-effective and potent immunization strategies capable of generating long-term protection in a single dose. However, LAVs often produce systemic viral replication, which can lead to unwanted post-vaccination side effects and pose a risk of reversion to a pathogenic phenotype and transmission to mosquitoes. Here, we utilized a chimeric infectious clone of CHIKV engineered with the domain C of the E2 gene of Semliki Forest virus (SFV) to express IFNγ and IL-21—two potent antiviral and immunomodulatory cytokines—in order to improve the LAV's attenuation while maintaining immunogenicity. The IFNγ- and IL-21-expressing vaccine candidates were stable during passage and significantly attenuated post-vaccination, as mice experienced reduced footpad swelling with minimal systemic replication and dissemination capacity compared to the parental vaccine. Additionally, these candidates provided complete protection to mice challenged with WT CHIKV. Our dual attenuation strategy represents an innovative way to generate safe and effective alphavirus vaccines that could be applied to other viruses. [ABSTRACT FROM AUTHOR]

Mayaro virus (MAYV) is a mosquito-borne Alphavirus that is widespread in South America. MAYV infection often presents with non-specific febrile symptoms but may progress to debilitating chronic arthritis or arthralgia. Despite the pandemic threat of MAYV, its true distribution remains unknown. The objective of this study was to clarify the geographic distribution of MAYV using an established risk mapping framework. This consisted of generating evidence consensus scores for MAYV presence, modeling the potential distribution of MAYV in select countries across Central and South America, and estimating the population residing in areas suitable for MAYV transmission. We compiled a georeferenced compendium of MAYV occurrence in humans, animals, and arthropods. Based on an established evidence consensus framework, we integrated multiple information sources to assess the total evidence supporting ongoing transmission of MAYV within each country in our study region. We then developed high resolution maps of the disease's estimated distribution using a boosted regression tree approach. Models were developed using nine climatic and environmental covariates that are related to the MAYV transmission cycle. Using the output of our boosted regression tree models, we estimated the total population living in regions suitable for MAYV transmission. The evidence consensus scores revealed high or very high evidence of MAYV transmission in several countries including Brazil (especially the states of Mato Grosso and Goiás), Venezuela, Peru, Trinidad and Tobago, and French Guiana. According to the boosted regression tree models, a substantial region of South America is suitable for MAYV transmission, including north and central Brazil, French Guiana, and Suriname. Some regions (e.g., Guyana) with only moderate evidence of known transmission were identified as highly suitable for MAYV. We estimate that approximately 58.9 million people (95% CI: 21.4–100.4) in Central and South America live in areas that may be suitable for MAYV transmission, including 46.2 million people (95% CI: 17.6–68.9) in Brazil. Our results may assist in prioritizing high-risk areas for vector control, human disease surveillance and ecological studies. Author summary: We used a comprehensive, georeferenced compendium of Mayaro virus (MAYV) occurrence to model the suitability for MAYV in Central and South America. Our boosted regression tree model incorporated 195 MAYV occurrence locations and 9 gridded environmental covariates to generate a 5 x 5 km continuous surface of MAYV suitability across Central and South America. Using this distribution model, we estimated the total population residing in areas that are suitable for MAYV transmission. Furthermore, we developed evidence consensus scores for each country in our study region that synthesized a variety of sources to assess the overall evidence of known MAYV transmission. Our study provides a contemporary estimate of MAYV distribution using a well-established disease mapping framework. This information provides an evidence base that can guide disease surveillance (including human cases and ecological studies) and vector control efforts in Central and South America. This is especially useful in regions with high predicted MAYV suitability but only moderate evidence of current MAYV transmission (e.g., Guyana and Suriname). [ABSTRACT FROM AUTHOR]

Mayaro virus (MAYV) is a mosquito-transmitted alphavirus that causes debilitating and persistent arthritogenic disease. While MAYV was previously reported to infect non-human primates (NHP), characterization of MAYV pathogenesis is currently lacking. Therefore, in this study we characterized MAYV infection and immunity in rhesus macaques. To inform the selection of a viral strain for NHP experiments, we evaluated five MAYV strains in C57BL/6 mice and showed that MAYV strain BeAr505411 induced robust tissue dissemination and disease. Three male rhesus macaques were subcutaneously challenged with 105 plaque-forming units of this strain into the arms. Peak plasma viremia occurred at 2 days post-infection (dpi). NHPs were taken to necropsy at 10 dpi to assess viral dissemination, which included the muscles and joints, lymphoid tissues, major organs, male reproductive tissues, as well as peripheral and central nervous system tissues. Histological examination demonstrated that MAYV infection was associated with appendicular joint and muscle inflammation as well as presence of perivascular inflammation in a wide variety of tissues. One animal developed a maculopapular rash and two NHP had viral RNA detected in upper torso skin samples, which was associated with the presence of perivascular and perifollicular lymphocytic aggregation. Analysis of longitudinal peripheral blood samples indicated a robust innate and adaptive immune activation, including the presence of anti-MAYV neutralizing antibodies with activity against related Una virus and chikungunya virus. Inflammatory cytokines and monocyte activation also peaked coincident with viremia, which was well supported by our transcriptomic analysis highlighting enrichment of interferon signaling and other antiviral processes at 2 days post MAYV infection. The rhesus macaque model of MAYV infection recapitulates many of the aspects of human infection and is poised to facilitate the evaluation of novel therapies and vaccines targeting this re-emerging virus. Author summary: Mayaro virus (MAYV) is an arbovirus capable of causing debilitating arthritis and myalgia in humans and the virus is currently circulating in Central and South America. With several factors supporting viral emergence, MAYV is a public health risk due to the lack of FDA-approved countermeasures. Although non-human primate (NHP) infection models are well established for chikungunya virus (CHIKV) and the equine encephalitic alphaviruses, there is currently no well-characterized NHP model of MAYV pathogenesis. With evidence of well-established mouse models of MAYV infection and a report from 1967 demonstrating that MAYV infection of NHPs in a laboratory setting was feasible, we aimed to further characterize MAYV infection in three rhesus macaques. Following precursor studies in mice to identify an optimal viral strain for NHP infection, we subcutaneously challenged rhesus macaques and characterized viral pathogenesis and immunity over the course of 10 days. Our study establishes a framework for future evaluation of MAYV-specific treatments in this relevant animal model. [ABSTRACT FROM AUTHOR]

Zika virus (ZIKV) is an arthropod-borne flavivirus that presents with acute febrile illness associated with rash, arthralgia and conjunctivitis. After years of sporadic reports in Africa, the three major outbreaks of this disease occurred in Yap Islands (2007), French Polynesia (2013–14) and South Americas (2015–16). Although, serological surveys suggested the presence of ZIKV in India in 1950s, cross-reactivity could not be ruled out. The first four proven cases of ZIKV from India were reported in 2017. This was followed by major outbreaks in the states of Rajasthan and Madhya Pradesh in 2018. Fortunately, the outbreaks in India were not associated with neurological complications. These outbreaks in India highlighted the spread of this disease beyond geographical barriers owing to the growing globalization, increased travel and ubiquitous presence of its vector, the Aedes mosquito. In this review, we discuss the epidemiology, clinical features and management of ZIKV in India. [ABSTRACT FROM AUTHOR]

Zika virus is a mosquito‐borne flavivirus which caused major epidemics in the Pacific and the Americas between 2013 and 2015. International travellers have previously acted as a sentinel population for Zika virus transmission in endemic areas, where local transmission may be incompletely captured by local surveillance systems. We report five recent European travellers returning from Thailand with Zika virus infection, highlighting the risk of ongoing endemic transmission in this popular tourist destination. [ABSTRACT FROM AUTHOR]

Background Zika virus (ZIKV) infection during pregnancy can cause severe birth defects in the fetus and is associated with neurodevelopmental abnormalities in childhood. Our objective was to describe ZIKV knowledge and attitudes among pregnant women in Colombia while ZIKV was circulating and whether they predicted the adoption of behaviors to prevent ZIKV mosquito-borne and sexual transmission. Methods We used self-reported data from Zika en Embarazadas y Niños (ZEN), a cohort study of women in early pregnancy across three regions of Colombia during 2017–2018. We used Poisson regression to estimate associations between knowledge, attitudes and previous experience with mosquito-borne infection and preventative behaviors. Results Among 1519 women, knowledge of mosquito-borne transmission was high (1480; 97.8%) and 1275 (85.5%) participants were worried about ZIKV infection during pregnancy. The most common preventive behavior was wearing long pants (1355; 89.4%). Regular mosquito repellent use was uncommon (257; 17.0%). While ZIKV knowledge and attitudes were not associated with the adoption of ZIKV prevention behaviors, previous mosquito-borne infection was associated with increased condom use (prevalence ratio 1.4, 95% CI 1.1 to 1.7). Conclusions Participants were well informed about ZIKV transmission and its health consequences. However, whether this knowledge resulted in behavior change is less certain. [ABSTRACT FROM AUTHOR]

Introduction: Chikungunya fever is an infection transmitted by the Chikungunya virus (CHIKV), which is an arbovirus that is transmitted by the mosquitoes Aedes aegypti and Aedes albopictus. The most common sequelae caused by CHIKV are chronic musculoskeletal pain, nerve damage, joint deformation and functional impairment. Objective: To systematically identify the literature on the contributions of physiotherapy in the treatment of patients with CHIKV sequelae. Materials and methods: Systematic review of the literature, guided by the recommendations of the Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA). The databases used were PUBMED, LILACS, Scielo and PEDro. Experimental studies and/or full case studies published without language restriction or publication data were included, in which they stood out as contributions of musculoskeletal functional rehabilitation in the treatment of patients with the condition in question. Analytical observational studies, editorial letters, review protocols, reflective studies, literature reviews and articles that do not have an abstract and/or full text available online were excluded. Results: The search in the databases was carried out between July and August 2022. A total of 4,782 articles were found on the platforms used and 10 articles from the gray literature search. After the duplicate analysis, 2,027 studies were excluded, leaving 2,755 articles that had their titles and abstracts read, of which 600 articles were selected for full reading. After this step, a final sample of 13 articles was eligible for this review. Final considerations: The most consolidated approaches used in the literature demonstrate that kinesiotherapy, associated or not with electrothermophototherapy, the pilates method and auriculotherapy are useful resources in the treatment of these individuals, significantly inspired by pain relief, improved quality of life and of functionality. [ABSTRACT FROM AUTHOR]

Human monoclonal antibodies might offer an important new approach to reduce malaria morbidity and mortality. In the first two parts of a three-part clinical trial, the antimalarial monoclonal antibody CIS43LS conferred high protection against parasitaemia at doses of 20 mg/kg or 40 mg/kg administered intravenously followed by controlled human malaria infection. The ability of CIS43LS to confer protection at lower doses or by the subcutaneous route is unknown. We aimed to provide data on the safety and optimisation of dose and route for the human antimalaria monoclonal antibody CIS43LS. VRC 612 Part C was the third part of a three-part, first-in-human, phase 1, adaptive trial, conducted at the University of Maryland, Baltimore Center for Vaccine Development and Global Health, Baltimore, MD, USA. We enrolled adults aged 18–50 years with no previous malaria vaccinations or infections, in a sequential, dose-escalating manner. Eligible participants received the monoclonal antibody CIS43LS in a single, open-label dose of 1 mg/kg, 5 mg/kg, or 10 mg/kg intravenously, or 5 mg/kg or 10 mg/kg subcutaneously. Participants underwent controlled human malaria infection by the bites of five mosquitoes infected with Plasmodium falciparum 3D7 strain approximately 8 weeks after their monoclonal antibody inoculation. Six additional control participants who did not receive CIS43LS underwent controlled human malaria infection simultaneously. Participants were followed-up daily on days 7–18 and day 21, with qualitative PCR used for P falciparum detection. Participants who tested positive for P falciparum were treated with atovaquone-proguanil and those who remained negative were treated at day 21. Participants were followed-up until 24 weeks after dosing. The primary outcome was safety and tolerability of CIS43LS at each dose level, assessed in the as-treated population. Secondary outcomes included protective efficacy of CIS43LS after controlled human malaria infection. This trial is now complete and is registered with ClinicalTrials.gov , NCT04206332. Between Sept 1, 2021, and Oct 29, 2021, 47 people were assessed for eligibility and 31 were enrolled (one subsequently withdrew and was replaced) and assigned to receive doses of 1 mg/kg (n=7), 5 mg/kg (n=4), and 10 mg/kg (n=3) intravenously and 5 mg/kg (n=4) and 10 mg/kg (n=4) subcutaneously, or to the control group (n=8). CIS43LS administration was safe and well tolerated; no serious adverse events occurred. CIS43LS protected 18 (82%) of 22 participants who received a dose. No participants developed parasitaemia following dosing at 5 mg/kg intravenously or subcutaneously, or at 10 mg/kg intravenously or subcutaneously. All six control participants and four of seven participants dosed at 1 mg/kg intravenously developed parasitaemia after controlled human malaria infection. CIS43LS was safe and well tolerated, and conferred protection against P falciparum at low doses and by the subcutaneous route, providing evidence that this approach might be useful to prevent malaria across several clinical use cases. National Institute of Allergy and Infectious Diseases, National Institutes of Health. [ABSTRACT FROM AUTHOR]

Venezuelan and eastern equine encephalitis viruses (VEEV and EEEV, respectively) are mosquito-borne, neuroinvasive human pathogens for which no FDA-approved therapeutic exists. Besides the biothreat posed by these viruses when aerosolized, arthropod transmission presents serious health risks to humans, as demonstrated by the 2019 outbreak of EEE disease in the United States that resulted in 38 confirmed cases, 19 deaths, and neurological effects in survivors. Here, we describe the discovery of a 2-pyrrolidinoquinazolinone scaffold, efficiently synthesized in two to five steps, whose structural optimization resulted in profound antiviral activity. The lead quinazolinone, BDGR-49, potently reduced cellular VEEV and EEEV titers by >7 log at 1 μM and exhibited suitable intravenous and oral pharmacokinetic profiles in BALB/c mice to achieve excellent brain exposure. Outstanding in vivo efficacy was observed in several lethal, subcutaneous infection mouse models using an 8-day dosing regimen. Prophylactically administered BDGR-49 at 25 mg kg−1 per day fully protected against a 10× LD50 VEEV Trinidad donkey (TrD) challenge in BALB/c mice. Similarly, we observed 70% protection when 10× LD50 EEEV FL93-939–infected C57BL/6 mice were treated prophylactically with BDGR-49 at 50 mg kg−1 per day. Last, we observed 100% therapeutic efficacy when mice, challenged with 10× LD50 VEEV TrD, were dosed at 48 hours after infection with BDGR-49 at 25 mg kg−1 per day. Mouse brain viral titers at 96 hours after infection were reduced to values near the limit of detection. Collectively, these results underscore the substantial development potential of a well-tolerated, brain-penetrant lead compound that shows promise in preventing and treating encephalitic alphavirus disease. Antiviral advance: Venezuelan and eastern equine encephalitis viruses (VEEV and EEEV, respectively) are mosquito-borne alphaviruses that can cause severe neurological and systemic disease in humans and equids. There is no approved vaccine or therapeutic available to treat human alphavirus infections. Cao et al. now describe the identification and optimization of a quinazolinone, BDGR-49, that showed potent antiviral activity against VEEV and EEEV in vitro and outstanding prophylactic and therapeutic efficacy in mouse models of VEEV and EEEV. BDGR-49 also showed promising pharmacokinetic profiles and evidence of brain penetration. Together these findings highlight BDGR-49 as a promising compound for treating encephalitic alphavirus infection. —CF [ABSTRACT FROM AUTHOR]

Mosquito saliva, which contains proteins with antihemostatic, anti-inflammatory, and immunomodulatory activities, is inoculated into the host during malaria parasite transmission. Some saliva proteins associate with the Plasmodium sporozoite surface and may interfere in host–pathogen interactions. Early work on a few Anopheles proteins indicates that specific saliva factors can negatively or positively affect Plasmodium transmission and infection. Following a mosquito bite, components of its saliva remain in the skin for many hours and may alter the innate and adaptive immune response in the skin. Some salivary proteins such as Anopheles gambiae sporozoite-associated protein (AgSAP), sporozoite-associated mosquito saliva protein 1 (SAMSP1), and AgTRIO can inhibit T cell responses and overall inflammatory reactions that may directly impact the survival of Plasmodium sporozoites. Next-generation vaccines and monoclonal antibodies targeting salivary proteins may lead to the development of broad-spectrum therapeutics that can block the transmission of Plasmodium and other mosquito-borne pathogens. Malaria begins when anopheline mosquitoes inject Plasmodium sporozoites, along with saliva when taking a blood meal, into the dermis of a vertebrate host. Mosquito saliva has pleiotropic effects that can influence local host immune responses. Although the role of mosquito saliva in malaria transmission has been speculated on for more than two decades, recent studies identified the role of salivary proteins in host–pathogen interactions. Elucidating the role of these proteins in modulating immune responses in the skin may help design new approaches to control the establishment of Plasmodium infection. Malaria is caused by Plasmodium protozoa that are transmitted by anopheline mosquitoes. Plasmodium sporozoites are released with saliva when an infected female mosquito takes a blood meal on a vertebrate host. Sporozoites deposited into the skin must enter a blood vessel to start their journey towards the liver. After migration out of the mosquito, sporozoites are associated with, or in proximity to, many components of vector saliva in the skin. Recent work has elucidated how Anopheles saliva, and components of saliva, can influence host–pathogen interactions during the early stage of Plasmodium infection in the skin. Here, we discuss how components of Anopheles saliva can modulate local host responses and affect Plasmodium infectivity. We hypothesize that therapeutic strategies targeting mosquito salivary proteins can play a role in controlling malaria and other vector-borne diseases. [ABSTRACT FROM AUTHOR]

Background: To date, no specific therapy or vaccination is available for West Nile virus (WNV) infections in humans; preventive strategies represent the only possibility to control transmission. To focus these strategies, detailed knowledge of the virus dynamics is of paramount importance. However, several aspects of WNV transmission are still unclear, especially regarding the role of potential vertebrate host species. Whereas mosquitoes' intrinsic characteristics cause them to favour certain hosts (host preference), absolute selection is impossible in natural settings. Conversely, the selection carried out among available hosts and influenced from hosts' availability and other ecological/environmental factors is defined as host selection. Methodology/Principal findings: In July 2022, we searched PubMed database for original articles exploring host selection among WNV-transmitting Culex mosquitoes, the main WNV vector. We considered only original field studies estimating and reporting forage ratio. This index results from the ratio between the proportion of blood meals taken by mosquitoes on potential host species and the hosts' relative abundance. From the originally retrieved 585 articles, 9 matched the inclusion criteria and were included in this review. All but one of the included studies were conducted in the Americas, six in the United States, and one each in Mexico and Colombia. The remaining study was conducted in Italy. American Robin, Northern Cardinal, and House Finch were the most significantly preferred birds in the Americas, Common Blackbird in Italy. Conclusions/Significance: Although ornithophilic, all observed WNV-transmitting mosquitoes presented opportunistic feeding behaviour. All the observed species showed potential to act as bridges for zoonotic diseases, feeding also on humans. All the observed mosquitoes presented host selection patterns and did not feed on hosts as expected by chance alone. The articles observe different species of mosquitoes in different environments. In addition, the way the relative host abundance was determined differed. Finally, this review is not systematic. Therefore, the translation of our results to different settings should be conducted cautiously. Author summary: West Nile virus (WNV) is a mosquito-borne virus that can cause a neuroinvasive and potentially deadly disease in vertebrates (including humans). The disease is transmitted from vectors (mostly Culex mosquitoes) during the blood meal. As no specific therapy or vaccination are available, the control of human cases centres on mosquitoes–humans contact prevention. For these reasons, deep knowledge of WNV transmission dynamics is considered paramount to focus preventive strategies. Nevertheless, WNV transmission dynamics are still unclear, especially regarding the role of potential hosts. Mosquitoes are intrinsically attracted from one host over another. However, in natural settings, absolute selection is often impossible. Therefore, in this review, we aimed at summarising knowledge on WNV-transmitting mosquitoes' host selection in natural settings. We specifically considered original scientific publications calculating forage ratio for Culex mosquitoes. The forage ratio is the proportion of the relative abundance of blood meals taken from a potential host in trapped mosquitoes and that host's relative abundance. Our results suggest that all observed Culex mosquitoes presented an opportunistic feeding behaviour and the potential to transmit zoonotic pathogens, since they all fed also on humans. Moreover, all the observed mosquitoes fed on hosts with a certain degree of selection, rather than what might be expected by host abundance alone. [ABSTRACT FROM AUTHOR]

St. Louis encephalitis virus (SLEV) is an endemic flavivirus in the western and southeastern United States, including California. From 1938 to 2003, the virus was detected annually in California, but after West Nile virus (WNV) arrived in 2003, SLEV was not detected again until it re-emerged in Riverside County in 2015. The re-emerging virus in California and other areas of the western US is SLEV genotype III, which previously had been detected only in Argentina, suggesting a South American origin. This study describes SLEV activity in California since its re-emergence in 2015 and compares it to WNV activity during the same period. From 2015 to 2020, SLEV was detected in 1,650 mosquito pools and 26 sentinel chickens, whereas WNV was detected concurrently in 18,108 mosquito pools and 1,542 sentinel chickens from the same samples. There were 24 reported human infections of SLEV in 10 California counties, including two fatalities (case fatality rate: 8%), compared to 2,469 reported human infections of WNV from 43 California counties, with 143 fatalities (case fatality rate: 6%). From 2015 through 2020, SLEV was detected in 17 (29%) of California's 58 counties, while WNV was detected in 54 (93%). Although mosquitoes and sentinel chickens have been tested routinely for arboviruses in California for over fifty years, surveillance has not been uniform throughout the state. Of note, since 2005 there has been a steady decline in the use of sentinel chickens among vector control agencies, potentially contributing to gaps in SLEV surveillance. The incidence of SLEV disease in California may have been underestimated because human surveillance for SLEV relied on an environmental detection to trigger SLEV patient screening and mosquito surveillance effort is spatially variable. In addition, human diagnostic testing usually relies on changes in host antibodies and SLEV infection can be indistinguishable from infection with other flaviviruses such as WNV, which is more prevalent. Author summary: St. Louis encephalitis virus is a mosquito-borne virus that can cause human disease and is found in California, where it was detected every year from 1938 to 2003. However, after West Nile virus arrived in 2003, St. Louis encephalitis virus was not detected again until 2015, when it re-emerged in Riverside County. From 2015 through 2020, St. Louis encephalitis virus has been detected in mosquito pools and sentinel chicken sera samples in 16 counties and a total of 24 human disease cases have been reported. However, during that same time-period, West Nile virus has been detected in 10 times as many mosquito pools and 60 times as many chicken sera samples across 58 counties and over 2,400 human disease cases have been reported. Although mosquitoes are tested routinely for both viruses, surveillance is not uniform throughout the state, and there has been a steady decline in the use of sentinel chickens. Since St. Louis encephalitis virus patient screening is dependent upon environmental detection, California may be underestimating the incidence of human disease due to this virus. [ABSTRACT FROM AUTHOR]

• For asymptomatic Plasmodium falciparum infections, median duration of infection was 37 days. • For asymptomatic Plasmodium vivax cases, median duration of infection was 60 days. • Clinical P. vivax cases and mixed species infections were infectious to mosquitoes. • A single asymptomatic mixed species infection was highly infectious to mosquitoes. • Low-density asymptomatic infections were sparsely infectious to mosquitoes. A 15-month longitudinal study was conducted to determine the duration and infectivity of asymptomatic qPCR-detected Plasmodium falciparum and Plasmodium vivax infections in Ethiopia. Total parasite and gametocyte kinetics were determined by molecular methods; infectivity to Anopheles arabiensis mosquitoes by repeated membrane feeding assays. Infectivity results were contrasted with passively recruited symptomatic malaria cases. For P. falciparum and P. vivax infections detected at enrolment, median durations of infection were 37 days (95% confidence interval [CI], 15-93) and 60 days (95% CI, 18-213), respectively. P. falciparum and P. vivax parasite densities declined over the course of infections. From 47 feeding assays on 22 asymptomatic P. falciparum infections, 6.4% (3/47) were infectious and these infected 1.8% (29/1579) of mosquitoes. No transmission was observed in feeding assays on asymptomatic P. vivax mono-infections (0/56); one mixed-species infection was highly infectious. Among the symptomatic cases, 4.3% (2/47) of P. falciparum and 73.3% (53/86) of P. vivax patients were infectious to mosquitoes. The majority of asymptomatic infections were of short duration and low parasite density. Only a minority of asymptomatic individuals were infectious to mosquitoes. This contrasts with earlier findings and is plausibly due to the low parasite densities in this population. [ABSTRACT FROM AUTHOR]

Arthritogenic alphaviruses, such as Ross River virus, chikungunya virus and O'nyong-nyong virus, cause endemic disease globally and are a major public health concern. The hallmarks of arthritogenic alphavirus disease are debilitating pain, and potentially chronic inflammation of the muscles, thus influencing quality of life. The type I IFN response is a major component of the innate immune response against arthritogenic alphaviruses, and is essential in inhibiting viral replication and dissemination. Type I IFNs are induced during early stages of infection and are essential for the activation of the antiviral innate immune response. They also link the innate immune response and the activation of adaptive immunity. This review focuses on the host immune response, particularly that involving type I IFN, in arthritogenic alphavirus disease. [ABSTRACT FROM AUTHOR]

Alphavirus capsid proteins (CPs) have two domains: the N-terminal domain (NTD), which interacts with the viral RNA, and the C-terminal domain (CTD), which forms CP-CP interactions and interacts with the cytoplasmic domain of the E2 spike protein (cdE2). In this study, we examine how mutations in the CP NTD affect CP CTD interactions with cdE2. We changed the length and/or charge of the NTD of Ross River virus CP and found that changing the charge of the NTD has a greater impact on core and virion assembly than changing the length of the NTD. The NTD CP insertion mutants are unable to form cytoplasmic cores during infection, but they do form cores or core-like structures in virions. Our results are consistent with cdE2 having a role in core maturation during virion assembly and rescuing core formation when cytoplasmic cores are not assembled. We go on to find that the isolated cores from some mutant virions are now assembly competent in that they can be disassembled and reassembled back into cores. These results show how the two domains of CP may have distinct yet coordinated roles. [ABSTRACT FROM AUTHOR]

Dengue (DENV) and chikungunya (CHIKV) viral infections trigger high patient morbidity and mortality. Mono-/co-infection of these viruses activates innate immune response, triggering Toll-like receptor (TLR) pathways. The present study investigated the differential role of TLR3, 7 and 8 single-nucleotide polymorphisms (SNPs) between mono- and co-infected Eastern Indian patients. Interaction of TLR polymorphic variants with signal peptidase complex (SPC18) was explored which might affect immune signalling against DENV/CHIKV infections. Out of 550 febrile symptomatic patients, 128 DENV-CHIKV co-infected samples were genotyped for eight SNPs of TLR3 (rs3775290-chr4:186083063), TLR7 (rs179008-chrX:12885540, rs5741880-chrX:12869297, rs179010-chrX:12884766, rs3853839-chrX:12889539) and TLR8 (rs5744080-chrX:12919685, rs3764879-chrX:12906578, rs3764880-chrX:12906707) by PCR-RFLP along with 157 healthy individuals. Statistical analysis established genotypic association of TLR SNPs with DENV-CHIKV co-infection, and difference between mono- and co-infected patients and their role in determining high viral load (HVL) during competitive viral replication among co-infected patients. In silico protein-protein docking evaluated interactive effect of TLR variants with SPC18. The findings revealed patients with CC genotypes of TLR7 and 8 SNPs were significantly susceptible towards co-infection, whereas specific genotypes of TLR7 and 8 imparted protection against co-infection. Differential analysis between mono-/co-infected patients revealed distinct genotypic distribution of TLR3, 7 and 8 SNPs. Co-infected patients with TT-rs179010 exhibited DENV-HVL, whereas CHIKV-HVL was detected among patients with other genotypes. Molecular docking of TLR7-rs179008 Q variant and TLR8-rs3764880 V variant with SPC18 generated better free binding energy. This study underlined the importance of TLR7 and 8 SNPs towards mono-/co-infection of DENV/CHIKV, with certain genotypes associated with co-infection susceptibility. Moreover, it suggested a probable role of specific genotypes of TLR7 and 8 polymorphisms imparting high dengue/chikungunya viral load among co-infected patients. [ABSTRACT FROM AUTHOR]

Chikungunya virus (CHIKV) infection is endemic in many different countries. CHIKV outbreaks are emerging in new areas and re-emerging in previously exposed geographical regions, thus making it a significant public health concern. CHIKV infections are often clinically inapparent, especially in children, which poses a challenge to testing and evaluating any vaccine. During CHIKV infection, CHIKV-specific antibodies are produced, and some of these antibodies can neutralize viruses released from infected cells before they can enter uninfected cells. In this study, we evaluated IgG binding and neutralizing antibody responses in paired serum samples from CHIKV-infected children and those with other febrile illness, using a recombinant truncated E2 protein and whole CHIKV particles as test antigens. Antibody detection using the truncated E2 protein showed a significant overlap between CHIKV-infected subjects and those with other febrile illnesses. This overlap was greater when binding antibody titers were determined using fixed CHIKV particles as the test antigen. Acute- and convalescent-phase sera collected from children after CHIKV infection showed significant differences in their neutralizing capacity. The neutralizing and binding antibody response showed a significant positive correlation. We detected IgG antibodies in most cases during the acute phase of infection. This was observed at two different geographical locations, one of which is not considered highly endemic. Conventional wisdom would suggest this to be a marker of re-infection (secondary infection). However, dissenting opinions have been voiced in other viral diseases (such as Ebola) where studies have detected IgG in acute illness. In the absence of any significant body of work documenting secondary CHIKV infections, we believe further work is needed to understand the early IgG response that we observed. [ABSTRACT FROM AUTHOR]

Chikungunya virus (CHIKV) is an arthropod-borne virus that has re-emerged recently and has spread to previously unaffected regions, resulting in millions of infections worldwide. The genus Alphavirus , in the family Togaviridae, contains several members with a similar potential for epidemic emergence. In order for CHIKV to replicate in targeted cell types it is essential for the virus to enter these cells. In this review, we summarize our current understanding of the versatile and promiscuous steps in CHIKV binding and entry into human and mosquito host cells. We describe the different entry pathways, receptors, and attachment factors so far described for CHIKV and other mosquito-borne alphaviruses and discuss them in the context of tissue tropism and potential therapeutic targeting. In the replication cycle of alphaviruses, the entry step is the first essential step to allow efficient production of new viral particles. The main entry pathway of alphaviruses is clathrin-mediated endocytosis. Fusion requires a low pH and takes place in the endosome. However, other, clathrin-independent, pathways that facilitate alphavirus entry have also been described. Some host factors that can mediate alphavirus entry have been identified but entry in the absence of these host factors is still possible. Alphavirus entry appears to be a promiscuous process following multiple routes where various pathways, attachment factors, and receptors can be used, assuring a broad tissue tropism. [ABSTRACT FROM AUTHOR]

Chikungunya virus (CHIKV) is a vector (mosquito)-transmitted alphavirus (family Togaviridae). CHIKV can cause fever and febrile illness associated with severe arthralgia and rash. Genotypic and phylogenetic analysis are important to understand the spread of CHIKV during epidemics and the diversity of circulating strains for the prediction of effective control measures. Molecular epidemiologic analysis of CHIKV is necessary to understand the complex interaction of vectors, hosts and environment that influences the genotypic evolution of epidemic strains. In this study, different works published during 1950s to 2020 concerning CHIKV evolution, epidemiology, vectors, phylogeny, and clinical outcomes were analyzed. Outbreaks of CHIKV have been reported from Bangladesh, Bhutan, India, Pakistan, Sri Lanka, Nepal, and Maldives in South Asia during 2007–2020. Three lineages- Asian, East/Central/South African (ECSA), and Indian Ocean Lineage (IOL) are circulating in South Asia. Lineage, ECSA and IOL became predominant over Asian lineage in South Asian countries during 2011–2020 epidemics. Further, the mutant E1-A226V is circulating in abundance with Aedes albopictus in India, Bangladesh, Nepal, and Bhutan. CHIKV is underestimated as clinical symptoms of CHIKV infection merges with the symptoms of dengue fever in South Asia. Failure to inhibit vector mediated transmission and predict epidemics of CHIKV increase the risk of larger global epidemics in future. To understand geographical spread of CHIKV, most of the studies focused on CHIKV outbreak, biology, pathogenesis, infection, transmission, and treatment. This updated study will reveal the collective epidemiology, evolution and phylogenies of CHIKV, supporting the necessity to investigate the circulating strains and vectors in South Asia. [ABSTRACT FROM AUTHOR]

During the dengue epidemic in Yunnan Province, China, during 2019, a concurrent outbreak of chikungunya occurred in the city of Ruili, which is located in the southwest of the province, adjacent to Myanmar. As part of this outbreak, three neonatal cases of infection with indigenous chikungunya virus from mother-to-child (vertical) transmission were observed. Isolates of chikungunya virus were obtained from 37 serum samples of patients with chikungunya during this outbreak, and a phylogenetic analysis of these isolates revealed that they belong to the Indian Ocean subclade of the East/Central/South African genotype. The E1 genes of these viruses did not harbor the A226V mutation. [ABSTRACT FROM AUTHOR]

Chikungunya virus (CHIKV, family Togaviridae) is a mosquito-transmitted alphavirus. The positive-sense RNA genome of CHIKV encodes four nonstructural proteins (nsP1 to nsP4) that are virus-specific subunits of the RNA replicase. Among nsP functions, those of nsP3 are the least understood. The C-terminal hypervariable domain (HVD) in nsP3 is disordered and serves as a platform for interactions with multiple host proteins. For Sindbis virus (SINV) and Semliki Forest virus (SFV), the nsP3 HVD has been shown to be phosphorylated. Deletion of phosphorylated regions has a mild effect on the growth of SFV and SINV in vertebrate cells. Using radiolabeling, we demonstrated that nsP3 in CHIKV and o'nyong-nyong virus is also phosphorylated. We showed that the phosphorylated residues in CHIKV nsP3 are not clustered at the beginning of the HVD. The substitution of 20 Ser/Thr residues located in the N-terminal half of the HVD or 26 Ser/Thr residues located in its C-terminal half with Ala residues reduced the activity of the CHIKV replicase and the infectivity of CHIKV in mammalian cells. Furthermore, the substitution of all 46 potentially phosphorylated residues resulted in the complete loss of viral RNA synthesis and infectivity. The mutations did not affect the interaction of the HVD in nsP3 with the host G3BP1 protein; interactions with CD2AP, BIN1, and FHL1 proteins were significantly reduced but not abolished. Thus, CHIKV differs from SFV and SINV both in the location of the phosphorylated residues in the HVD in nsP3 and, significantly, in their effect on replicase activity and virus infectivity. [ABSTRACT FROM AUTHOR]

Chikungunya virus (CHIKV) is an arthropod-borne virus transmitted by mosquitoes of the genus Aedes. CHIKV infection causes various rheumatic symptoms, including enthesitis; however, these effects are rarely investigated. The aim of this study was to describe the rheumatic manifestations in CHIKV infection, estimate the prevalence of enthesitis in CHIKV-infected patients, and determine the factors associated with CHIKV-induced enthesitis. We conducted a prospective, observational study in patients with CHIKV infection confirmed by positive RT-PCR or IgM assay from October 2019 to March 2020. Patients with pre-existing inflammatory rheumatic diseases were excluded. A rheumatologist evaluated the demographic and clinical characteristics of the patients, including the number of inflamed joints, enthesitis sites, tendinitis, and tenosynovitis. The Leeds enthesitis index (LEI) and the Maastricht ankylosing spondylitis enthesis score (MASES) were used to evaluate enthesitis sites. Factors associated with enthesitis were determined using logistic regression analysis. One hundred and sixty-four participants diagnosed with CHIKV infection were enrolled. The mean (SD) age of the patients was 48.2 (14) years. The most common pattern of rheumatic manifestations was polyarthritis with or without enthesitis. Enthesitis was observed in 63 patients (38.4%). The most common site of enthesitis was the left lateral epicondyle as assessed by LEI and the posterior superior iliac spine as assessed by MASES. Multivariate analysis indicated that the number of actively inflamed joints and Thai-HAQ score at the initial evaluation were significantly associated with the presence of enthesitis. The main rheumatic manifestations of CHIKV infection were arthritis/arthralgia, with enthesitis as a prominent extraarticular feature. CHIKV infection can cause enthesitis at peripheral and axial sites. We found that enthesitis was associated with a high number of inflamed joints and reduced physical function. These results indicate that the assessment of enthesitis should be considered when monitoring disease activity and as a treatment response parameter in CHIKV-infected patients. [ABSTRACT FROM AUTHOR]

Between 2018 and 2019, the incidence of chikungunya was approximately 15,000 cases across 60 provinces in Thailand. Here, the clinical presentations in chikungunya, emergent pattern, and genomic diversity of the chikungunya virus (CHIKV) causing this massive outbreak were demonstrated. A total of 1,806 sera samples from suspected cases of chikungunya were collected from 13 provinces in Thailand, and samples were tested for the presence of CHIKV RNA, IgG, and IgM using real-time PCR, enzyme-linked immunoassay (ELISA), commercial immunoassay (rapid test). The phylogenetic tree of CHIKV whole-genome and CHIKV E1 were constructed using the maximum-likelihood method. CHIKV infection was confirmed in 547 (42.2%) male and 748 (57.8%) female patients by positive real-time PCR results and/or CHIKV IgM antibody titers. Unsurprisingly, CHIKV RNA was detected in >80% of confirmed cases between 1 and 5 days after symptom onset, whereas anti-CHIKV IgM was detectable in >90% of cases after day 6. Older age was clearly one of the risk factors for the development of arthralgia in infected patients. Although phylogenetic analysis revealed that the present CHIKV Thailand strain of 2018–2020 belongs to the East, Central, and Southern African (ECSA) genotype similar to the CHIKV strains that caused outbreaks during 2008–2009 and 2013, all present CHIKV Thailand strains were clustered within the recent CHIKV strain that caused an outbreak in South Asia. Interestingly, all present CHIKV Thailand strains possess two mutations, E1-K211E, and E2-V264A, in the background of E1-226A. These mutations are reported to be associated with virus-adapted Aedes aegypti. Taken together, it was likely that the present CHIKV outbreak in Thailand occurred as a result of the importation of the CHIKV strain from South Asia. Understanding with viral genetic diversity is essential for epidemiological study and may contribute to better disease management and preventive measures. [ABSTRACT FROM AUTHOR]

In this paper, a novel age-structured delayed mathematical model to control Aedes aegypti mosquitoes via Wolbachia-infected mosquitoes is introduced. To eliminate the deadly mosquito-borne diseases such as dengue, chikungunya, yellow fever, and Zika virus, the Wolbachia infection is introduced into the wild mosquito population at every stage. This method is one of the promising biological control strategies. To predict the optimal amount of Wolbachia release, the time varying delay is considered. Firstly, the positiveness of the solution and existence of both Wolbachia present and Wolbachia free equilibrium were discussed. Through linearization, construction of suitable Lyapunov–Krasovskii functional, and linear matrix inequality theory (LMI), the exponential stability is also analyzed. Finally, the simulation results are presented for the real-world data collected from the existing literature to show the effectiveness of the proposed model. [ABSTRACT FROM AUTHOR]

Background: Chikungunya virus is an alphavirus, primarily transmitted by Aedes aegypti and Ae. albopictus. In late 2017–2018, an outbreak of chikungunya occurred in Mombasa county, Kenya, and investigations were conducted to establish associated entomological risk factors. Methods: Homes were stratified and water-filled containers inspected for immature Ae. aegypti, and larval indices were calculated. Adult mosquitoes were collected in the same homesteads using BG-Sentinel and CDC light traps and screened for chikungunya virus. Experiments were also conducted to determine the ability of Culex quinquefasciatus to transmit chikungunya virus. Results: One hundred thirty-one houses and 1637 containers were inspected; 48 and 128 of them, respectively, were positive for immature Ae. aegypti, with the house index (36.60), container index (7.82) and Breteau index (97.71) recorded. Jerry cans (n = 1232; 72.26%) and clay pots (n = 2; 0.12%) were the most and least inspected containers, respectively, while drums, the second most commonly sampled (n = 249; 15.21%), were highly positive (65.63%) and productive (60%). Tires and jerry cans demonstrated the highest and lowest breeding preference ratios, 11.36 and 0.2, respectively. Over 6900 adult mosquitoes were collected and identified into 15 species comprising Cx. quinquefasciatus (n = 4492; 65.04%), Aedes vittatus (n = 1137; 16.46%) and Ae. aegypti (n = 911; 13.19%) and 2 species groups. Simpson's dominance and Shannon-Wiener diversity indices of 0.4388 and 1.1942 were recorded, respectively. Chikungunya virus was isolated from pools of Ae. aegypti (1) and Cx. quinquefasciatus (4), two of which were males. Minimum infection rates of 3.0 and 0.8 were observed for female Ae. aegypti and Cx. quinquefasciatus, respectively. Between 25 and 31.3% of exposed mosquitoes became infected with CHIKV 7, 14 and 21 days post-exposure. For the experimentally infected Cx. quinquefasciatus mosquitoes, between 13 and 40% had the virus disseminated, with 100% transmission being observed among those with disseminated infection. Conclusions: These results demonstrated high risk of chikungunya transmission for residents in the sampled areas of Mombasa. Transmission data confirmed the probable role played by Cx. quinquefasciatus in the outbreak while the role of Ae. vittatus in the transmission of chikungunya virus remains unknown. [ABSTRACT FROM AUTHOR]

• RNA repeated sequence elements are a common trait in the 3'UTR of arboviruses. • Alphavirus sequence duplications assure high viral fitness in mosquitoes. • Coevolution of viruses and their mosquito hosts shapes the viral 3'UTRs. • RNA recombination is a source of alphavirus 3'UTR diversity. • 3'UTR plasticity is associated to enhanced transmission and epidemic potential. Alphaviruses such as chikungunya and western equine encephalitis viruses are important human pathogens transmitted by mosquitoes that have recently caused large epidemic and epizootic outbreaks. The epidemic potential of alphaviruses is often related to enhanced mosquito transmission. Tissue barriers and antiviral responses impose bottlenecks to viral populations in mosquitoes. Substitutions in the envelope proteins and the presence of repeated sequence elements (RSEs) in the 3'UTR of epidemic viruses were proposed to be specifically associated to efficient replication in mosquito vectors. Here, we discuss the molecular mechanisms that originated RSEs, the evolutionary forces that shape the 3'UTR of alphaviruses, and the significance of RSEs for mosquito transmission. Finally, the presence of RSEs in the 3'UTR of viral genomes appears as evolutionary trait associated to mosquito adaptation and emerges as a common feature among viruses from the alphavirus and flavivirus genera. [ABSTRACT FROM AUTHOR]

The monitoring of Aedes urban species is usually conducted by ovitraps, which can provide a good estimate of population density at low cost and relatively easy management. When the monitoring network is managed by many operators, as in the case of the Emilia-Romagna regional plan, it becomes necessary to develop a quality control methodology capable of highlighting the possible data incoherency and ensuring that the monitoring system provides reliable data. This paper presents the procedure applied in the egg counting phase as developed in Emilia-Romagna in order to check the data quality and validate the data before being included in the database. Several steps have been identified and protocols developed to serve quality control such as training of technicians and intralaboratory quality check, daily random double counting of Masonite paddles and interlaboratory periodic comparison. The qualitative test showed that all operators were able to distinguish Aedes albopictus eggs from other mosquito species eggs. The quantitative test showed significant differences between the 11 operators, with a relative error resulting in the range of 0.1–25.8%. The daily internal double counting of randomly extracted Masonite slides was performed by the coordinator showing a relative error in the range 2.47–2.63% without statistical differences, confirming a good alignment of the operators with the coordinator throughout the monitoring period. The interlaboratory comparison trial produced an average relative error of 7.20% showing a sufficient alignment between the three laboratories. Moreover a new time-saving methods in egg counting was developed and tested in real condition. [ABSTRACT FROM AUTHOR]

Chikungunya virus (CHIKV) is a reemerging and rapidly spreading pathogen transmitted by mosquitoes. The emergence of new epidemic variants of the virus is associated with genetic evolutionary traits, including duplication of repeated RNA elements in the 3= untranslated region (UTR) that seemingly favor transmission by mosquitoes. The transmission potential of a given variant results from a complex interplay between virus populations and anatomical tissue barriers in the mosquito. Here, we used the wild-type CHIKV Caribbean strain and an engineered mutant harboring a deletion in the 3= UTR to dissect the interactions of virus variants with the anatomical barriers that impede transmission during the replication cycle of the virus in Aedes mosquitoes. Compared to the 3=-UTR mutant, we observed that the wild-type virus had a short extrinsic incubation period (EIP) after an infectious blood meal and was expectorated into mosquito saliva much more efficiently. We found that high viral titers in the midgut are not sufficient to escape the midgut escape barrier. Rather, viral replication kinetics play a crucial role in determining midgut escape and the transmission ability of CHIKV. Finally, competition tests in mosquitoes coinfected with wild-type and mutant viruses revealed that both viruses successfully colonized the midgut, but wild-type viruses effectively displaced mutant viruses during systemic infection due to their greater efficiency of escaping from the midgut into secondary tissues. Overall, our results uncover a link between CHIKV replication kinetics and the effect of bottlenecks on population diversity, as slowly replicating variants are less able to overcome the midgut escape barrier. IMPORTANCE It is well established that selective pressures in mosquito vectors impose population bottlenecks for arboviruses. Here, we used a CHIKV Caribbean lineage mutant carrying a deletion in the 3= UTR to study host-virus interactions in vivo in the epidemic mosquito vector Aedes aegypti. We found that the mutant virus had a delayed replication rate in mosquitoes, which lengthened the extrinsic incubation period (EIP) and reduced fitness relative to the wild-type virus. As a result, the mutant virus displayed a reduced capacity to cross anatomical barriers during the infection cycle in mosquitoes, thus reducing the virus transmission rate. Our findings show how selective pressures act on CHIKV noncoding regions to select variants with shorter EIPs that are preferentially transmitted by the mosquito vector. [ABSTRACT FROM AUTHOR]

This study presents a novel plasmonic fiber optic sandwich immunobiosensor for the detection of chikungunya, an infectious mosquito-borne disease with chronic musculoskeletal pain and acute febrile illness, by exploiting non-structural protein 3 (CHIKV-nsP3) as a biomarker. A plasmonic sandwich immunoassay for CHIKV-nsP3 was realized on the surface of a compact U-bent plastic optical fiber (POF, 0.5 mm core diameter) with gold nanoparticles (AuNPs) as labels. The high evanescent wave absorbance (EWA) sensitivity of the U-bent probes allows the absorption of the light passing through the fiber by the AuNP labels, upon the formation of a sandwich immunocomplex of CHIKV-nsP3 on the core surface of the U-bent probe region. A simple optical set-up with a low-cost green LED and a photodetector on either end of the U-bent probe gave rise to a detection limit of 0.52 ng mL−1 (8.6 pM), and a linear range of 1–104 ng mL−1 with a sensitivity of 0.1043A530 nm/log(CnsP3). In addition, the plasmonic POF biosensor shows strong specificity towards the CHIKV-nsP3 analyte in comparison with Pf-HRP2, HIgG, and dengue whole virus. The results illustrate the potential of plasmonic POF biosensors for direct and sensitive point-of-care detection of the chikungunya viral disease. [ABSTRACT FROM AUTHOR]

Chikungunya virus (CHIKV), a virus that induces pathogenic inflammatory host immune responses, is re-emerging worldwide, and there are currently no established antiviral control measures. Transient receptor potential vanilloid 1 (TRPV1), a non-selective Ca2+-permeable ion channel, has been found to regulate various host inflammatory responses including several viral infections. Immune responses to CHIKV infection in host macrophages have been reported recently. However, the possible involvement of TRPV1 during CHIKV infection in host macrophages has not been studied. Here, we investigated the possible role of TRPV1 in CHIKV infection of the macrophage cell line RAW 264.7. It was found that CHIKV infection upregulates TRPV1 expression in macrophages. To confirm this observation, the TRPV1-specific modulators 5ˊ-iodoresiniferatoxin (5ˊ-IRTX, a TRPV1 antagonist) and resiniferatoxin (RTX, a TRPV1 agonist) were used. Our results indicated that TRPV1 inhibition leads to a reduction in CHIKV infection, whereas TRPV1 activation significantly enhances CHIKV infection. Using a plaque assay and a time-of-addition assay, it was observed that functional modulation of TRPV1 affects the early stages of the viral lifecycle in RAW 264.7 cells. Moreover, CHIKV infection was found to induce of pNF-κB (p65) expression and nuclear localization. However, both activation and inhibition of TRPV1 were found to enhance the expression and nuclear localization of pNF-κB (p65) and production of pro-inflammatory TNF and IL-6 during CHIKV infection. In addition, it was demonstrated by Ca2+ imaging that TRPV1 regulates Ca2+ influx during CHIKV infection. Hence, the current findings highlight a potentially important regulatory role of TRPV1 during CHIKV infection in macrophages. This study might also have broad implications in the context of other viral infections as well. [ABSTRACT FROM AUTHOR]

Chronic joint pain is the most common pathology found in chikungunya virus (CHIKV)-infected patients. Eight cytokines were compared in CHIKV patients with and without joint pain. IL-4 and IL-13 levels were significantly lower in patients with joint pain (p = 0.006 and p < 0.0001, respectively). IL-18 levels were higher in the group of patients with joint pain (p < 0.0001) and were significantly higher on days 3 and 4 after onset (p = 0.0012 and p = 0.003, respectively). Moreover, TNF-α levels were significantly higher in patients with joint pain on day 3 (p = 0.028). This study demonstrated that cytokines, particularly IL-18, may be candidates for immunomodulation. [ABSTRACT FROM AUTHOR]

Chikungunya fever is a disease caused by the Chikungunya virus (CHIKV) that is transmitted by the bite of the female of Aedes sp. mosquito. The symptoms include fever, muscle aches, skin rash, and severe joint pains. The disease may develop into a chronic condition and joint pain for months or years. Currently, there is no effective antiviral treatment against CHIKV infection. Treatments based on natural compounds have been widely studied, as many drugs were produced by using natural molecules and their derivatives. Alpha-phellandrene (α-Phe) is a naturally occurring organic compound that is a ligand for ruthenium, forming the organometallic complex [Ru2Cl4(p-cymene)2] (RcP). Organometallic complexes have shown promising as candidate molecules to a new generation of compounds that presented relevant biological properties, however, there is a lack of knowledge concerning the anti-CHIKV activity of these complexes. The present work evaluated the effects of the RcP and its precursors, the hydrate ruthenium(III) chloride salt (RuCl3⋅xH2O) (Ru) and α-Phe, on CHIKV infection in vitro. To this, BHK-21 cells were infected with CHIKV- nanoluciferase (CHIKV -nanoluc), a viral construct harboring the nanoluciferase reporter gene, at the presence or absence of the compounds for 16 h. Cytotoxicity and impact on infectivity were analyzed. The results demonstrated that RcP exhibited a strong therapeutic potential judged by the selective index > 40. Antiviral effects of RcP on different stages of the CHIKV replicative cycle were investigated; the results showed that it affected early stages of virus infection reducing virus replication by 77% at non-cytotoxic concentrations. Further assays demonstrated the virucidal activity of the compound that completely blocked virus infectivity. In silico molecular docking calculations suggested different binding interactions between aromatic rings of RcP and the loop of amino acids of the E2 envelope CHIKV glycoprotein mainly through hydrophobic interactions. Additionally, infrared spectroscopy spectral analysis indicated interactions of RcP with CHIKV glycoproteins. These data suggest that RcP may act on CHIKV particles, disrupting virus entry to the host cells. Therefore, RcP may represent a strong candidate for the development of anti-CHIKV drugs. [ABSTRACT FROM AUTHOR]

The mosquito-borne chikungunya virus (CHIKV) causes an acute febrile illness with rash, joint and muscle pain.A realtime RT-PCR assay for CHIKV detecting non-structural protein (nsP2; CHIKV nsP2-RT-qPCR) was set up. All the serodiagnosed CHIKV cases detected during 2009-2019 in Finland were screened with the assay, followed by isolations attempts and sequencing using Sanger and next generation sequencing (NGS). To validate the assay external and in-house quality control samples were used and all were correctly identified. Specificity of the assay was 100%. Assay was sensitive to detect CHIKV RNA in dilution of 10-8.During years 2009-2019 34 patients were diagnosed for acute CHIKV infection. Twelve out of 34 cases were positive by CHIKV nsP2-RT-qPCR.Two CHIKV isolations succeeded from two individuals infected originally in Thailand, 2019. From 12 CHIKV nsP2-RT-qPCR positive samples, five (42%) CHIKVs were successfully sequenced. In this study, CHIKVs from year 2019 clustered with CHIKV ECSA-lineage forming sub-cluster with strains from ones detected in Bangladesh 2017, and the ones from Jamaica (2014) within Asian lineage showing highest similarity to strains detected in Caribbean outbreak 2013-15. Majority of the CHIKV infections detected in Finland originates from Asia and virus lineages reflect the global circulation of the pathogen. [ABSTRACT FROM AUTHOR]

Chikungunya is a mosquito-borne disease, caused by the member of the Togaviridae family belongs to the genus alphavirus, making it a major threat in all developing countries as well as some developed countries. The mosquito acts as a vector for the disease and carries the CHIK-Virus. To date there is no direct treatment available and that demands the development of more effective vaccines. In this study author employed Immune Epitope Database and Analysis Resource, a machine learning-based algorithm principally working on the Artificial Neural Network (ANN) algorithm, also known as (IEDB-ANN) for the prediction and analysis of Epitopes. A total of 173 epitopes were identified on the basis of IC50 values, among them 40 epitopes were found, sharing part with the linear B-cell epitopes and exposed to the cTAP1protein, and out of 40, 6 epitopes were noticed to show interactions with the cTAP with their binding energy ranging from − 3.61 to − 1.22 kcal/mol. The six epitopes identified were exposed to the HLA class I alleles and from this all revealed interaction with the HLA alleles and minimum binding energy that ranges from − 4.12 to − 5.88 kcal/mol. Besides, two T cell epitopes i.e. 145KVFTGVYPE153 and 395STVPVAPPR403 were found most promiscuous candidates. These promiscuous epitopes-HLA complexes were further analyzed by the molecular dynamics simulation to check the stability of the complex. Results obtained from this study suggest that the identified epitopes i.e. and 395STVPVAPPR403, are likely to be capable of passing through the lumen of ER to bind withthe HLA class I allele and provide new insights and potential application in the designing and development of peptide-based vaccine candidate for the treatment of chikungunya. [ABSTRACT FROM AUTHOR]

Alphaviruses (genus Alphavirus ; family Togaviridae) are a medically relevant family of viruses that include chikungunya virus and Mayaro virus. Infectious cDNA clones of these viruses are necessary molecular tools to understand viral biology. Traditionally, rescuing virus from an infectious cDNA clone requires propagating plasmids in bacteria, which can result in mutations in the viral genome due to bacterial toxicity or recombination and requires specialized equipment and knowledge to propagate the bacteria. Here, we present an alternative- rolling circle amplification (RCA), an in vitro technology. We demonstrate that the viral yield of transfected RCA product is comparable to midiprepped plasmid, albeit with a slight delay in kinetics. RCA, however, is cheaper and less time-consuming. Further, sequential RCA did not introduce mutations into the viral genome, subverting the need for glycerol stocks and retransformation. These results indicate that RCA is a viable alternative to traditional plasmid-based approaches to viral rescue. [ABSTRACT FROM AUTHOR]

Chikungunya virus (CHIKV), a mosquito-transmitted alphavirus, enters a cell through endocytosis, followed by viral and cell membrane fusion. The fusion protein, E1, undergoes an acid pH-induced pre- to postfusion conformation change during membrane fusion. As part of the conformation change, E1 dissociates from the receptorbinding protein, E2, and swivels its domains I and II over domain III to form an extended intermediate and then eventually to form a postfusion hairpin homotrimer. In this study, we tested if the domain I-III linker acts as a "hinge" for the swiveling motion of E1 domains. We found a conserved spring-twisted structure in the linker, stabilized by a salt bridge between a conserved arginine-aspartic acid pair, as a "hinge point" for domain swiveling. Molecular dynamics (MD) simulation of the CHIKV E1 or E2-E1 structure predicted that the spring-twisted region untwists at pH 5.5. Corroborating the prediction, introduction of a "cystine staple" at the hinge point, replacing the conserved arginine-aspartic acid pair with cysteine residues, resulted in loss of fusion activity of E1. MD simulation also predicted domain I-III swiveling at acidic pH. We tested if breaking the His 331-Lys 16 H bond between domains I and III, seen only in the prefusion conformation, is important for domain swiveling. When domains I and III are "stapled" by introducing a disulfide bond in between, E1 showed loss of fusion activity, implying that domain I and III dissociation is a critical acid pH-induced step in membrane fusion. However, replacement of His 331 with an acidic residue did not affect the pH threshold for fusion, suggesting His 331 is not an acid-sensing residue. [ABSTRACT FROM AUTHOR]

The first autochthonous case of chikungunya virus (CHIKV) infection in Brazil was in September 2014 in the State of Amapá, and from there it rapidly spread across the country. The present study was conducted in 2016 in the state of Rio Grande do Norte, and the aims were to describe the epidemiological and the clinical aspects of the CHIKV outbreak. Biological samples from 284 chikungunya suspected cases were screened for CHIKV and Flavivirus (FV) RNA using qRT-PCR. Negative PCR samples were also screened for anti-CHIKV and anti-FVIgM by ELISA. CHIKV RNA were detected in 125 samples mostly occurring from January through March (46%), mainly affecting adults and older adults. We found a gradual decrease in viral RNA over the disease time. Anti-CHIKV IgM was found in 47.5% after negative CHIKV qRT-PCR. Interestingly, 45.0% simultaneously had positive results for CHIKV and FV IgM, suggesting the occurrence of virus co-circulation. The most frequent symptom was fever (91%). Women presented more chance to develop nausea and abdominal pain compared to men. Our data described and allows us to better understand the clinical and epidemiological aspects of the 2016 chikungunya outbreak in Rio Grande do Norte and can help in the early clinical diagnosis of the virus. [ABSTRACT FROM AUTHOR]

Superinfection exclusion (SIE) is a process by which a virally infected cell is protected from subsequent infection by the same or a closely related virus. By preventing cell coinfection, SIE favors preservation of genome integrity of a viral strain and limits its recombination potential with other viral genomes, thereby impacting viral evolution. Although described in virtually all viral families, the precise step(s) impacted by SIE during the viral life cycle have not been systematically explored. Here, we describe for the first time SIE triggered by chikungunya virus (CHIKV), an alphavirus of public health importance. Using single-cell technologies, we demonstrate that CHIKV excludes subsequent infection with: CHIKV; Sindbis virus, a related alphavirus; and influenza A, an unrelated RNA virus. We further demonstrate that SIE does not depend on the action of type I interferon, nor does it rely on host cell transcription. Moreover, exclusion is not mediated by the action of a single CHIKV protein; in particular, we observed no role for non-structural protein 2 (nsP2), making CHIKV unique among characterized alphaviruses. By stepping through the viral life cycle, we show that CHIKV exclusion occurs at the level of replication, but does not directly influence virus binding, nor viral structural protein translation. In sum, we characterized co-infection during CHIKV replication, which likely influences the rate of viral diversification and evolution. [ABSTRACT FROM AUTHOR]

Blood safety is a major global issue. Transfusion transmitted parasitic infections (TTPI) like malaria are rare and possibly under-reported, a situation which could be attributed to lack of awareness of the mosquito-borne transmission of infection. Such infections are still considered potential health hazards, as they can pose a significant threat especially in immunocompromised patients, where they have proven to be fatal. Prevention of the transmission depends solely on the donor's questionnaire which addresses previous or current infection with aetiologic agents. Donor deferral is effective however clear guidelines are needed. This case report features the transfusion-transmitted of Plasmodium Falciparum in a 15-year-old splenectomised patient with underlying beta thalassaemia major. [ABSTRACT FROM AUTHOR]

Zika virus is an arthropod-borne flavivirus mainly transmitted by the bite of infected mosquitoes. However, alternative transmission routes can occur. In this study, we show the accidental transmission of virus from an infected mouse to a human during the experimental manipulation. This study describes the patient clinical manifestations and virus genome identification. [ABSTRACT FROM AUTHOR]

Japanese encephalitis virus (JEV) is a viral zoonosis that can cause viral encephalitis, death and disability. Although the Culex mosquito is the primary vector of JEV, little is known about JEV transmission by this kind of mosquito. Here, we found that mosquito defensin facilitated the adsorption of JEV on target cells via the defensin/lipoprotein receptor-related protein 2 (LRP2) axis. Mosquito defensin bound the ED III domain of the viral envelope (E) protein and directly mediated efficient virus adsorption on the target cell surface; the receptor LRP2, which is expressed on the cell surface, affected defensin-dependent adsorption. As a result, mosquito defensin enhanced JEV infection in the salivary gland, increasing the possibility of viral transmission by mosquitoes. These findings demonstrate the novel role of mosquito defensin in JEV infection and the mechanisms through which the virus exploits mosquito defensin for infection and transmission. [ABSTRACT FROM AUTHOR]

A generalized chikungunya virus (CHIKV) infection model with nonlinear incidence functions and two time delays is proposed and investigated. The model takes into account both modes of transmission that are virus-to-cell infection and cell-to-cell transmission. Furthermore, the local and global stabilities of the disease-free equilibrium and the chronic infection equilibrium are established by using the linearization and Lyapunov functional methods. Moreover, the existence of Hopf bifurcation is also analyzed. Finally, an application is presented in order to support the analytical results. [ABSTRACT FROM AUTHOR]

Aedes aegypti is the primary vector of several infectious viruses that cause yellow, dengue, chikungunya, and Zika fevers. Recently, plant-derived products have been tested as safe and eco-friendly larvicides against Ae. aegypti. The present study aimed to improve QSAR models for 62 larvicidal phytocompounds against Ae. aegypti via the Monte Carlo method based on the index of the ideality of correlation (IIC) criterion. The representation of structures was done with SMILES. Three splits were prepared randomly and three QSAR models were constructed using IIC target function. The molecular descriptors were selected from SMILES descriptors and the hydrogen-filled molecular graphs. The predictability of three models was evaluated on the validation sets, the r2 of which was 0.9770, 0.8660, and 0.8565 for models 1 to 3, respectively. The statistical results of three randomized splits indicated that robust, simple, predictive, and reliable models were obtained for different sets. From the modelling results, important descriptors were identified to enhance and reduce the larvicidal activity of compounds. Based on the identified important descriptors, some new structures of larvicidal compounds were proposed. The larvicidal activity of novel molecules designed further was supported by docking studies. Using the simple QSAR model, one can predict pLC50 of new similarity larvicidal phytocompounds. [ABSTRACT FROM AUTHOR]