Detection and quantification of natural Wolbachia in Aedes aegypti in Metropolitan Manila, Philippines using locally designed primers.

Background: The Philippines bears health and economic burden caused by high dengue cases annually. Presently, the Philippines still lack an effective and sustainable vector management. The use of Wolbachia, a maternally transmitted bacterium, that mitigate arbovirus transmission has been recommended. Cytoplasmic incompatibility and viral blocking, two characteristics that make Wolbachia suitable for vector control, depend on infection prevalence and density. There are no current Wolbachia release programs in the Philippines, and studies regarding the safety of this intervention. Here, we screened for Wolbachia in Aedes aegypti collected from Metropolitan Manila, Philippines. We designed location-specific primers for qPCR to test whether this improved Wolbachia detection in Ae. aegypti. We explored if host sex and Wolbachia strain could be potential factors affecting Wolbachia density. Methods: Ae. aegypti mosquitoes (n=429) were screened for natural Wolbachia by taqman qPCR using location-specific Wolbachia surface protein primers (wspAAML) and known 16S rRNA primers. Samples positive for wspAAML (n=267) were processed for Sanger sequencing. We constructed a phylogenetic tree using IQ-TREE 2 to further characterize Wolbachia present in the Philippine Ae. aegypti. We then compared Wolbachia densities between Wolbachia groups and host sex. Statistical analyses were done using GraphPad Prism 9.0. Results: Wolbachia prevalence for 16S rRNA (40%) and wspAAML (62%) markers were high. Wolbachia relative densities for 16S rRNA ranged from -3.84 to 2.71 and wspAAML from -4.02 to 1.81. Densities were higher in male than female mosquitoes. Wolbachia strains detected in Ae. aegypti clustered into supergroup B. Some 54% (123/226) of these sequences clustered under a group referred to here as "wAegML," that belongs to the supergroup B, which had a significantly lower density than wAegB/wAlbB, and wAlbA strains. Conclusion: Location-specific primers improved detection of natural Wolbachia in Ae. aegypti and allowed for relative quantification. Wolbachia density is relatively low, and differed between host sexes and Wolbachia strains. An economical way of confirming sporadic or transient Wolbachia in Ae. aegypti is necessary while considering host sex and bacterial strain.

Persistence of IgG and neutralizing antibodies in Crimean-Congo hemorrhagic fever survivors.

The World Health Organization classified Crimean-Congo hemorrhagic fever (CCHF) as a high-priority infectious disease and emphasized the performance of research studies and product development against it. Little information is available about the immune response due to natural CCHF virus (CCHFV) infection in humans. Here, we investigated the persistence of IgG and neutralizing antibodies in serum samples collected from 61 Iranian CCHF survivors with various time points after recovery (<12, 12-60, and >60 months after disease). The ELISA results showed IgG seropositivity in all samples while a pseudotyped based neutralization assay findings revealed the presence of neutralizing antibody in 29 samples (46.77%). For both IgG and neutralizing antibodies, a decreasing trend of titer was observed with the increase in the time after recovery. Not only the mean titer of IgG (772.80 U/mL) was higher than mean neutralizing antibody (25.64) but also the IgG persistence was longer. In conclusion, our findings provide valuable information about the long-term persistence of humoral immune response in CCHF survivors indicating that IgG antibody can be detected at least 8 years after recovery and low titers of neutralizing antibody can be detected in CCHF survivors.

Cell fusing agent virus rarely transmits vertically in artificially infected laboratory-colonized Aedes aegypti mosquitoes.

BACKGROUND: Vertical transmission (VT) of arboviruses (arthropod-borne viruses) can serve as an essential link in the transmission cycle during adverse environmental conditions. The extent of VT among mosquito-borne arboviruses can vary significantly among different virus families and even among different viruses within the same genus. For example, orthobunyaviruses exhibit a higher VT rate than orthoflaviviruses and alphaviruses. Mosquitoes are also the natural hosts of a large number of insect-specific viruses (ISV) that belong to several virus families, including Bunyaviridae, Flaviviridae, and Togaviridae. Cell fusing agent virus (CFAV), an insect-specific orthoflavivirus, displays higher VT rates than other dual-host orthoflaviviruses, such as Zika and dengue viruses. High VT rates require establishment of stabilized infections in the germinal tissues of female vectors. To delve deeper into understanding the mechanisms governing these differences in VT rates and the establishment of stabilized infections, the ovary infection patterns and VT of Zika virus (ZIKV) and CFAV were compared. METHODS: Laboratory colonized Aedes aegypti females were infected with either ZIKV or CFAV by intrathoracic injection. Ovary infection patterns were monitored by in situ hybridization using virus-specific probes, and VT was determined by detecting the presence of the virus among the progeny, using a reverse-transcription quantitative polymerase chain reaction (PCR) assay. RESULTS: Both ZIKV and CFAV infect mosquito ovaries after intrathoracic injection. Infections then become widespread following a non-infectious blood meal. VT rates of ZIKV are similar to previously reported results (3.33%). CFAV, on the contrary transmits vertically very rarely. VT was not observed in the first gonotrophic cycle following intrathoracic injection, and only rarely in the second gonotrophic cycle. VT of CFAV is mosquito population independent, since similar results were obtained with Aedes aegypti collected from two different geographic locations. CONCLUSIONS: Although CFAV infects mosquito ovaries, the occurrence of VT remains infrequent in artificially infected Ae. aegypti, despite the observation of high VT rates in field-collected mosquitoes. These results suggest that infections of insect-specific viruses are stabilized in mosquitoes by some as yet unidentified mechanisms.

Local Economic Conditions Affect Aedes albopictus Management.

Invasive mosquitoes are an emerging public health issue, as many species are competent vectors for pathogens. We assessed how multiple environmental and socio-economic factors affected the engagement of municipalities in Italy (n = 7679) in actions against Aedes albopictus, an invasive mosquito affecting human health and well-being, between 2000 and 2020. We collected information about mosquito control from official documents and municipal websites and modeled the role played by multiple environmental and socioeconomic factors characterizing each municipality through the random forest algorithm. Municipalities are more prone to manage A. albopictus if more urbanized, in lowlands and with long infestation periods. Moreover, these variables are more predictive of management in municipalities with a high median income and thus more economic resources. Only 25.5% of Italian municipalities approved regulations for managing A. albopictus, and very few of them were in Southern Italy, the most deprived area of the country. Our findings indicate that local economic conditions moderate the effect of other drivers of mosquito control and ultimately can lead to better management of A. albopictus. If the management of invasive mosquitoes, or other forms of global change, is subjected to local economic conditions, economic inequalities will jeopardize the success of large-scale policies, also raising issues of environmental and climate justice.

A brief description of the epidemiology of dengue in Dili, Timor-Leste, 2018-2022.

Abstract: Dengue virus (DENV) infection causes 390 million infections per year and 40,000 deaths globally. It is endemic in many countries in Asia, Africa, the Americas, the Caribbean, and Oceania. Dengue is endemic in Timor-Leste year-round, but peak transmission occurs during the rainy season. We briefly describe the epidemiology of DENV in the Municipality of Dili between 2018 and 2022. There were 6,234 cases notified, with a mean annual incidence rate of 330 cases per 100,000 population. There were 55 deaths (case fatality rate 0.9%). The peak annual incidence (3,904 cases) occurred in 2022 after an outbreak was declared in January of that year; this outbreak included 760 cases of dengue haemorrhagic fever and 35 deaths. The number of outbreak cases requiring hospital treatment exceeded the usual capacity, but facilities established for coronavirus disease 2019 (COVID-19) isolation and treatment were repurposed to meet this demand. Existing strategies of vector control, minimising breeding sites and promoting early presentation for treatment should continue, as should the utilisation of surveillance systems and treatment facilities established during the COVID-19 pandemic. However, dengue incidence remains high, and other dengue control strategies-including the deployment of Wolbachia-infected mosquitoes-should be considered in Timor-Leste.

The first confirmed outbreak of chikungunya reported in Timor-Leste, 2024.

Abstract: Timor-Leste is a mountainous, half-island nation with a population of 1.3 million, which shares a land border with Indonesia and is 550 km from Darwin, Australia. Since independence in 2002, Timor-Leste has achieved significant development; however, high levels of poverty remain. Chikungunya virus (CHIKV) is endemic in over 100 countries in Africa, Asia, Europe and in the Americas. It is transmitted by the bite of infected Aedes aegypti or Ae. albopictus mosquitoes, which are present in Timor-Leste and which contribute to annual rainy-season dengue virus (DENV) outbreaks. Symptomatic people typically suffer from acute onset of fever, usually accompanied by severe arthritis or arthralgia. Joint pain can be debilitating for several days, and may sometimes last for weeks, months or years. Unlike DENV infection which has significant mortality, most people recover completely. Between 2002 and 2023, there were 26 cases of CHIKV notified in Australia who acquired their infection in Timor-Leste; however, laboratory testing capability for CHIKV in Timor-Leste only became available in 2021 using polymerase chain reaction (PCR). The first locally diagnosed case was notified in November 2023. In January 2024, an outbreak of CHIKV was recognised in Timor-Leste for the first time, with 195 outbreak cases reported during 1-31 January 2024; all were PCR positive. There were no cases hospitalised, and no deaths. The median age of cases was 17 years (range 1-76 years); 51% were males. Cases were reported across the country; most (88/195) were from Dili, although the highest incidence was seen in the neighbouring municipality of Ermera (monthly incidence rate of 58.8 cases per 100,000 population). This first reported outbreak of CHIKV in Timor-Leste highlights the need for improved mosquito-borne illness control and response strategies, including minimising breeding sites and promoting early presentation for treatment and differential diagnosis from DENV, and consideration of the deployment of Wolbachia-infected mosquitoes, particularly as they have shown to reduce the transmission of CHIKV, DENV and Zika virus, all of which pose threats in Timor-Leste.

Risk of infection with arboviruses in a healthy population in Pakistan based on seroprevalence.

Infectious diseases caused by arboviruses are a public health concern in Pakistan. However, the studies on data prevalence and threats posed by arboviruses are limited. This study investigated the seroprevalence of arboviruses in a healthy population in Pakistan, including severe fever with thrombocytopenia syndrome virus (SFTSV), Crimean-Congo haemorrhagic fever virus (CCHFV), Tamdy virus (TAMV), and Karshi virus (KSIV) based on a newly established luciferase immunoprecipitation system (LIPS) assays, and Zika virus (ZIKV) by enzyme-linked immunosorbent assays (ELISA). Neutralizing activities against these arboviruses were further examined from the antibody positive samples. The results showed that the seroprevalence of SFTSV, CCHFV, TAMV, KSIV, and ZIKV was 17.37%, 7.58%, 4.41%, 1.10%, and 6.48%, respectively, and neutralizing to SFTSV (1.79%), CCHFV (2.62%), and ZIKV (0.69%) were identified, as well as to the SFTSV-related Guertu virus (GTV, 0.83%). Risk factors associated with the incidence of exposure and levels of antibody response were analysed. Moreover, co-exposure to different arboviruses was demonstrated, as thirty-seven individuals were having antibodies to multiple viruses and thirteen showed neutralizing activity. Males, individuals aged ≤ 40 years, and outdoor workers had high risk of exposure to arboviruses. All these results reveal the substantial risks of infection with arboviruses in Pakistan, and indicate the threat from co-exposure to multiple arboviruses. The findings raise the need for further epidemiologic investigation in expanded regions and populations and the necessity to improve health surveillance in Pakistan.

Phagocyte-expressed glycosaminoglycans promote capture of alphaviruses from the blood circulation in a host species-specific manner.

The magnitude and duration of vertebrate viremia are critical determinants of arbovirus transmission, geographic spread, and disease severity-yet, mechanisms determining arbovirus viremia levels are poorly defined. Previous studies have drawn associations between in vitro virion-glycosaminoglycan (GAG) interactions and in vivo clearance kinetics of virions from blood circulation. From these observations, it is commonly hypothesized that GAG-binding virions are rapidly removed from circulation due to ubiquitous expression of GAGs by vascular endothelial cells, thereby limiting viremia. Using an in vivo model for viremia, we compared the vascular clearance of low and enhanced GAG-binding viral variants of chikungunya, eastern- (EEEV), and Venezuelan- (VEEV) equine encephalitis viruses. We find GAG-binding virions are more quickly removed from circulation than their non-GAG-binding variant; however individual clearance kinetics vary between GAG-binding viruses, from swift (VEEV) to slow removal from circulation (EEEV). Remarkably, we find phagocytes are required for efficient vascular clearance of some enhanced GAG-binding virions. Moreover, transient depletion of vascular heparan sulfate impedes vascular clearance of only some GAG-binding viral variants and in a phagocyte-dependent manner, implying phagocytes can mediate vascular GAG-virion interactions. Finally, in direct contrast to mice, we find enhanced GAG-binding EEEV is resistant to vascular clearance in avian hosts, suggesting the existence of species-specificity in virion-GAG interactions. In summary, these data support a role for GAG-mediated clearance of some viral particles from the blood circulation, illuminate the potential of blood-contacting phagocytes as a site for GAG-virion binding, and suggest a role for species-specific GAG structures in arbovirus ecology.

Epidemiologic Survey of Crimean-Congo Hemorrhagic Fever Virus in Suids, Spain.

We conducted a cross-sectional study in wild boar and extensively managed Iberian pig populations in a hotspot area of Crimean-Congo hemorrhagic fever virus (CCHFV) in Spain. We tested for antibodies against CCHFV by using 2 ELISAs in parallel. We assessed the presence of CCHFV RNA by means of reverse transcription quantitative PCR protocol, which detects all genotypes. A total of 113 (21.8%) of 518 suids sampled showed antibodies against CCHFV by ELISA. By species, 106 (39.7%) of 267 wild boars and 7 (2.8%) of 251 Iberian pigs analyzed were seropositive. Of the 231 Iberian pigs and 231 wild boars analyzed, none tested positive for CCHFV RNA. These findings indicate high CCHFV exposure in wild boar populations in endemic areas and confirm the susceptibility of extensively reared pigs to CCHFV, even though they may only play a limited role in the enzootic cycle.

Molecular Epidemiology of Mayaro Virus among Febrile Patients, Roraima State, Brazil, 2018-2021.

We detected Mayaro virus (MAYV) in 3.4% (28/822) of febrile patients tested during 2018-2021 from Roraima State, Brazil. We also isolated MAYV strains and confirmed that these cases were caused by genotype D. Improved surveillance is needed to better determine the burden of MAYV in the Amazon Region.

Novel chikungunya and dengue vaccines-travel medicine applications.

Recent epidemics of dengue and chikungunya have highlighted the urgent need for vaccines to reduce the risk of infection in travellers. Given challenges tracking chikungunya outbreaks in real-time and the widespread resurgence of dengue, broader indications for the use of the new chikungunya and dengue vaccines should be considered.

Arbovirus - a threat to transfusion safety in Spain: a narrative review. / Arbovirus como amenaza para la seguridad transfusional en España: revisión narrativa.

Arboviruses represent a threat to transfusion safety for several reasons: the presence of vectors and the notification of autochthonous cases in our region, the recent increase in the number of cases transmitted through blood and/or blood component transfusion, the high prevalence rates of RNA of the main arboviruses in asymptomatic blood donors, and their ability to survive processing and storage in the different blood components. In an epidemic outbreak caused by an arbovirus in our region, transfusion centres can apply different measures: reactive measures, related to donor selection or arbovirus screening, and proactive measures, such as pathogen inactivation methods. The study of the epidemiology of the main arboviruses and understanding the effectiveness of the different measures that we can adopt are essential to ensure that our blood components remain safe.

Fever and Rash.

Infectious causes of fever and rash pose a diagnostic challenge for the emergency provider. It is often difficult to discern rashes associated with rapidly progressive and life-threatening infections from benign exanthems, which comprise the majority of rashes seen in the emergency department. Physicians must also consider serious noninfectious causes of fever and rash. A correct diagnosis depends on an exhaustive history and head-to-toe skin examination as most emergent causes of fever and rash remain clinical diagnoses. A provisional diagnosis and immediate treatment with antimicrobials and supportive care are usually required prior to the return of confirmatory laboratory testing.

Population-specific responses to developmental temperature in the arboviral vector Aedes albopictus: Implications for climate change.

The increase of environmental temperature due to current global warming is not only favouring the expansion of the distribution range of many insect species, but it is also changing their phenology. Insect phenology is tightly linked to developmental timing, which is regulated by environmental temperatures. However, the degree to which the effects of developmental temperatures extend across developmental stages and their inter-stage relationships have not been thoroughly quantified in mosquitoes. Here, we used the mosquito Aedes albopictus, which is an aggressive invasive species and an arboviral vector, to study how developmental temperature influences fitness across developmental stages, thermal traits, energy reserves, transcriptome and Wolbachia prevalence in laboratory-reared populations originally collected from either temperate or tropical regions. We show that hatchability, larval and pupal viability and developmental speed are strongly influenced by temperature, and these effects extend to wing length, body mass, longevity and content of water, protein and lipids in adults in a population-specific manner. On the contrary, neither adult thermal preference nor heat resistance significantly change with temperature. Wolbachia density was generally lower in adult mosquitoes reared at 18°C than at other tested temperatures, and transcriptome analysis showed enrichment for functions linked to stress responses (i.e. cuticle proteins and chitin, cytochrome p450 and heat shock proteins) in mosquitoes reared at both 18 and 32°C. Our data showed an overall reduced vector fitness performance when mosquitoes were reared at 32°C, and the absence of isomorphy in the relationship between developmental stages and temperature in the laboratory population deriving from larvae collected in northern Italy. Altogether, these results have important implications for reliable model projections of the invasion potentials of Ae. albopictus and its epidemiological impact.

Anodal tDCS over the motor cortex improves pain but not physical function in chronic chikungunya arthritis: Randomized controlled trial.

BACKGROUND: Chikungunya virus (CHIKV) is a globally prevalent pathogen, with outbreaks occurring in tropical regions. Chronic pain is the main symptom reported and is associated with decreased mobility and disability. Transcranial direct current stimulation (tDCS) is emerging as a new therapeutic tool for chronic arthralgia. OBJECTIVE: To evaluate the effectiveness of 10 consecutive sessions of anodal tDCS on pain (primary outcome) in participants with chronic CHIKV arthralgia. Secondary outcomes included functional status, quality of life, and mood. METHODS: In this randomized, double-blind, placebo-controlled trial, 30 participants with chronic CHIKV arthralgia were randomly assigned to receive either active (n = 15) or sham (n = 15) tDCS. The active group received 10 consecutive sessions of tDCS over M1 using the C3/Fp2 montage (2 mA for 20 min). Visual analog scale of pain (VAS), health assessment questionnaire (HAQ), short-form 36 health survey (SF-36), pain catastrophizing scale, Hamilton anxiety scale (HAS), timed up and go (TUG) test, lumbar dynamometry, 30-s arm curl and 2-min step test were assessed at baseline, day 10 and at 2 follow-up visits. RESULTS: There was a significant interaction between group and time on pain (p = 0.03; effect size 95 % CI 0.9 (-1.67 to -0.16), with a significant time interaction (p = 0.0001). There was no interaction between time and group for the 2-minute step test (p = 0.18), but the groups differed significantly at day 10 (p = 0.01), first follow-up (p = 0.01) and second follow-up (p = 0.03). HAQ and SF-36 improved but not significantly. There was no significant improvement in mental health, and physical tests. CONCLUSION: tDCS appears to be a promising intervention for reducing pain in participants with chronic CHIKV arthralgia, although further research is needed to confirm these findings and explore potential long-term benefits. TRIAL REGISTRATION: Brazilian Registry of Clinical Trials (ReBEC): RBR-245rh7.

Inflammatory Response Associated with West Nile Neuroinvasive Disease: A Systematic Review.

BACKGROUND: West Nile virus (WNV) infection is a seasonal arbovirosis with the potential to cause severe neurological disease. Outcomes of the infection from WNV depend on viral factors (e.g., lineage) and host-intrinsic factors (e.g., age, sex, immunocompromising conditions). Immunity is essential to control the infection but may also prove detrimental to the host. Indeed, the persistence of high levels of pro-inflammatory cytokines and chemokines is associated with the development of blood-brain barrier (BBB) damage. Due to the importance of the inflammatory processes in the development of West Nile neuroinvasive disease (WNND), we reviewed the available literature on the subject. METHODS: According to the 2020 updated PRISMA guidelines, all peer-reviewed articles regarding the inflammatory response associated with WNND were included. RESULTS: One hundred and thirty-six articles were included in the data analysis and sorted into three groups (in vitro on-cell cultures, in vivo in animals, and in humans). The main cytokines found to be increased during WNND were IL-6 and TNF-α. We highlighted the generally small quantity and heterogeneity of information about the inflammatory patterns associated with WNND. CONCLUSIONS: Further studies are needed to understand the pathogenesis of WNND and to investigate the extent and the way the host inflammatory response either helps in controlling the infection or in worsening the outcomes. This might prove useful both for the development of target therapies and for the development of molecular markers allowing early identification of patients displaying an inflammatory response that puts them at a higher risk of developing neuroinvasive disease and who might thus benefit from early antiviral therapies.

Comprehensive evaluation of nucleic acid amplification methods widely used for generic detection of sandfly-borne phleboviruses.

Sandfly-borne phleboviruses (SBPs), which cause sandfly fever, aseptic meningitis, encephalitis, and meningoencephalitis, are emerging pathogens of major public health concern. Virus nucleic acid testing is essential for SBP diagnosis, especially in the early stages of infection, and for the discovery of novel SBPs. The efficacy of utilizing generic primers that target conserved nucleotide sequences for the detection of both known and novel SBPs has not been extensively evaluated. We aimed to compare and evaluate the performance of five generic primer sets, widely used to detect S- and L-segments of arthropod-borne phleboviruses and designed as singleplex (n = 3) and nested (n = 2) formats, including both well-known and recently characterized 15 Old World virus strains. Furthermore, we performed in silico analysis to assess the detection capabilities of these generic primer sets. The initial evaluation of previously published generic primer sets for SBP detection yielded two singleplex primer sets with the potential to be adapted for use in real-time or high-throughput detection settings. Studies are ongoing to develop and further optimize a preliminary assay and test various hosts and vectors to assess their capacity to detect known and novel viruses. IMPORTANCE: Virus nucleic acid testing is the primary diagnostic method, particularly in the early stages of illness. Virus-specific or syndromic tests are widely used for this purpose. The use of generic primers has had a considerable impact on the discovery, identification, and detection of Old World sandfly-borne phleboviruses (OWSBP). The study is significant because it is the first to carry out a comparative evaluation of all published OWSBP generic primer sets.

Persistence of dengue serotype 2 viral RNA in blood cells of a returned traveler with dengue fever.

Dengue virus (DENV) is one of the most significant vector-borne pathogens worldwide. In this report, we describe clinical features and laboratory detection of dengue in a 45-year-old traveler to Nicaragua on return home to the United States in 2019. Clinical presentation was mild, with rash, headache, and fatigue, with only low-grade transient fever. Infection dynamics were documented by serology and PCR of serially collected body fluids. DENV serotype 2 was detected in whole blood 1 day after symptoms emerged, with viral RNA isolated to the red cell fraction, and remained detectable through day 89. DENV-2 RNA was detected in serum only on day 4, and IgM was undetectable on day 4 but evident by day 13. Viral RNA was also detected in urine. This report of DENV-2 RNA persistence in blood cells but only transient appearance in serum, supports the potential diagnostic value of whole blood over serum for PCR and opportunity of an expanded testing window. Informed testing approaches can improve diagnostic accuracy and inform strategies that preserve individual and public health.

Development of Multiplex Molecular Assays for Simultaneous Detection of Dengue Serotypes and Chikungunya Virus for Arbovirus Surveillance.

The major arboviruses mainly belong to the Bunyaviridae, Togaviridae, and Flaviviridae families, among which the chikungunya virus and dengue virus have emerged as global public health problems. The main objective of this study was to develop specific, sensitive, and cost-effective molecular multiplex RT-PCR and RT-qPCR assays for the rapid and simultaneous detection of CHIKV and the four serotypes of DENV for arbovirus surveillance. Specific primers for all viruses were designed, and one-step multiplex RT-PCR (mRT-PCR) and RT-qPCR (mRT-qPCR) were developed using reference strains of the CHIKV and DENV serotypes. The specificity of the test for all the viruses was confirmed through sequencing. The standard curves showed a high correlation coefficient, R2 = 0.99, for DENV-2 and DENV-3; R2 = 0.98, for DENV-4; and CHIKV; R2 = 0.93, for DENV-1. The limits of detection were calculated to be 4.1 × 10-1 copies/reaction for DENV-1, DENV-3, and CHIKV and 4.1 × 101 for DENV-2 and DENV-4. The specificity and sensitivity of the newly developed mRT-PCR and mRT-qPCR were validated using positive serum samples collected from India and Burkina Faso. The sensitivity of mRT-PCR and mRT-qPCR are 91%, and 100%, respectively. The specificity of both assays was 100%. mRT-PCR and mRT-qPCR assays are low-cost, and a combination of both will be a useful tool for arbovirus surveillance.

Pathophysiology of chikungunya virus infection associated with fatal outcomes.

Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes acute, subacute, and chronic human arthritogenic diseases and, in rare instances, can lead to neurological complications and death. Here, we combined epidemiological, virological, histopathological, cytokine, molecular dynamics, metabolomic, proteomic, and genomic analyses to investigate viral and host factors that contribute to chikungunya-associated (CHIK) death. Our results indicate that CHIK deaths are associated with multi-organ infection, central nervous system damage, and elevated serum levels of pro-inflammatory cytokines and chemokines compared with survivors. The histopathologic, metabolite, and proteomic signatures of CHIK deaths reveal hemodynamic disorders and dysregulated immune responses. The CHIKV East-Central-South-African lineage infecting our study population causes both fatal and survival cases. Additionally, CHIKV infection impairs the integrity of the blood-brain barrier, as evidenced by an increase in permeability and altered tight junction protein expression. Overall, our findings improve the understanding of CHIK pathophysiology and the causes of fatal infections.