Arboviruses Research Articles

Arbovirus in Solid Organ Transplants: A Narrative Review of the Literature

The incidence of arbovirus infections has increased in recent decades. Other than dengue, chikungunya, and West Nile viruses, the data on arbovirus in solid organ transplant (SOT) are limited to case reports, and infections in renal transplant recipients account for most of the reported cases. Dengue and West Nile infections seem to be more severe with higher mortality in SOT patients than in the general population. Acute kidney injury is more frequent in patients with dengue and chikungunya although persistent arthralgia with the latter is less frequent. There is no clear relationship between arboviral infection and acute cellular rejection. Pre-transplant screening of donors should be implemented during increased arboviral activity but, despite donor screening and negative donor nucleic acid amplification test (NAT), donor derived infection can occur. NAT may be transiently positive. IgM tests lack specificity, and neutralizing antibody assays are more specific but not readily available. Other tests, such as immunohistochemistry, antigen tests, PCR, metagenomic assays, and viral culture, can also be performed. There are a few vaccines available against some arboviruses, but live vaccines should be avoided. Treatment is largely supportive. More data on arboviral infection in SOT are needed to understand its epidemiology and clinical course.

Vaccine candidates based on MVA viral vectors expressing VP2 or VP7 confer full protection against Epizootic hemorrhagic disease virus in IFNAR(-/-) mice.

Epizootic hemorrhagic disease (EHD), caused by Epizootic hemorrhagic disease virus (EHDV), is an emerging and severe livestock disease. Recent incursion and distribution of EHDV in Europe have outlined the need for vaccine research against this viral disease. In this work, we report modified vaccinia virus Ankara (MVA)-vectored vaccines designed to express protein VP2 of EHDV-8 or protein VP7 of EHDV-2. Prime boost immunization of adult IFNAR(-/-) mice with the MVA-VP2 vaccine candidate induced high titers of EHDV-8-specific neutralizing antibodies (NAbs) and conferred full protection against homologous lethal challenge with EHDV-8. However, no heterologous protection was observed after lethal challenge with EHDV-6. In contrast, the MVA-VP7 vaccine candidate elicited strong cytotoxic CD8+ T-cell responses against VP7 and conferred complete protection against lethal challenge with either EHDV-8 or EHDV-6 in IFNAR(-/-) mice in the absence of NAbs, being the first multiserotype vaccine candidate against EHDV. Moreover, we expressed recombinant proteins VP2 and VP7 of EHDV in the baculovirus expression system, which were used to analyze the potential DIVA (differentiating infected from vaccinated animals) character of these vaccine candidates.IMPORTANCEEmergence and re-emergence of arthropod-borne viruses are major concerns for both human and animal health. The most recent example is the fast expansion of EHDV-8 through Europe. Besides, EHDV-8 relates with a high prevalence of pathologic cases in cattle populations. No vaccine is currently available in Europe, and vaccine research against this arboviral disease is negligible. In this work, we present novel DIVA vaccine candidates against EHDV, and most importantly, we identified the protein VP7 of EHDV as an antigen capable of inducing multiserotype protection, one of the major challenges in vaccine research against orbiviruses.

Experimental arboviral infection of mosquito larvae: A novel approach for vector competence studies

Vector competence studies in mosquitoes present valuable opportunities to explore arboviral transmission and virus-vector interactions. However, oral infection studies in mosquitoes can be challenging. An alternative approach is to infect mosquitoes during their aquatic larval stage, resulting in the emergence of infected adults. To investigate the potential of this method, Culex pipiens biotype molestus larvae were infected with Usutu virus (USUV, Orthoflavivirus usutuense). For this purpose, larvae were exposed to USUV-infected mammalian and mosquito cell cultures for 24 h before being reared to adults. Subsequent analysis via RT-qPCR revealed that the Culex larvae successfully acquired USUV from the infected cells and exhibited high susceptibility to infection. Immediately after emergence, 32.10 % (26/81) of male and 41.03 % (16/39) of female mosquitoes tested positive for USUV RNA. Notably, females that were incubated for 15 days post-emergence demonstrated even higher infection rates, reaching 100.00 % (23/23). In addition, viral RNA and infectious particles were detected in some saliva samples, indicating the potential for transmission. This experimental infection of mosquito larvae thus offers the opportunity to produce infected adult mosquitoes for studies enhancing our understanding of virus-vector interactions, co-infections, and transmission routes. Such research contributes to better public health strategies addressing arboviral diseases.

Distribution and abundance of Aedes caspius (Pallas, 1771) and Aedes vexans (Meigen, 1830) in the Po Plain (northern Italy).

Knowledge of the distribution and abundance of disease-causing mosquito vectors is fundamental for assessing the risk of disease circulation and introduction. Aedes caspius (Pallas, 1771) and Aedes vexans (Meigen, 1830) have been implicated, to different extents, in the circulation of several arthropod-borne viruses (arboviruses). These two mosquitoes are vectors of Tahyna virus in Europe and are considered potential vectors of Rift Valley fever virus, a virus not present but at risk of introduction on the continent. In this work, we analysed abundance data collected during West Nile virus (WNV) surveillance in northern Italy (Po Plain) via 292 CO2-baited traps to evaluate the distribution and density of these two non-target mosquitoes. We modelled the distribution and abundance of these two mosquito species in the surveyed area using two distinct spatial analysis approaches (geostatistical and machine learning), which yielded congruent results. Both species are more abundant close to the Po River than elsewhere, but Ae. caspius is present in the eastern and western parts of the plain, linked with the occurrence of rice fields and wetlands, while Ae. vexans is observed in the middle area of the plain. Presence and abundance data at the municipality level were obtained and made available through this work. This work demonstrates the importance of maintaining and improving entomological surveillance programs with an adequate sampling effort.

Tissue specific innate immune responses impact viral infection in Drosophila.

All organisms sense and respond to pathogenic challenge. Tissue-specific responses are required to combat pathogens infecting distinct cell types. Cyclic dinucleotides (CDNs) are produced endogenously downstream of pathogen recognition or by pathogens themselves which bind to STING to activate NF-kB-dependent antimicrobial gene expression programs. It remains unknown whether there are distinct immune responses to CDNs in Drosophila tissues. Here, we investigated tissue specific CDN-STING responses and uncovered differences in gene-induction patterns across tissues that play important roles in viral infections. Using tissue-and cell-specific genetic studies we found that dSTING in the fat body controls CDN-induced expression of dSTING-regulated gene 1 (Srg1) but not dSTING-regulated gene 2 (Srg2) or 3 (Srg3). In contrast, the gastrointestinal tract largely controls expression of Srg2 and Srg3. We found that Srg3 is antiviral against the natural fly pathogen Drosophila C virus and the human arthropod-borne Rift Valley Fever virus (RVFV), but not other arthropod-borne viruses including Sindbis virus and dengue virus. Furthermore, we found that Srg3 has an important role in controlling RVFV infection of the ovary which has important implications in understanding vertical transmission of viruses and RVFV in mosquitoes. Overall, our study underscores the importance of tissue-specific responses in antiviral immunity and highlights the complex tissue regulation of the CDN-STING pathway.

Dicer-2 mutations in Aedes aegypti cells lead to a diminished antiviral function against Rift Valley fever virus and Bunyamwera virus infection.

Mosquitoes are known to transmit different arthropod-borne viruses belonging to various virus families. The exogenous small interfering RNA pathway plays an important role in the mosquito defence against such virus infections, with Dicer-2 (Dcr2) as one of the key proteins that initiates the cleavage of viral dsRNAs into 21 nt long virus-derived small interfering RNAs. Previous data identified the importance of various motifs in Dcr2 for its small interfering RNA (siRNA)-mediated antiviral activity. However, all these data focus on positive-strand RNA viruses, although negative-strand RNA viruses, like Bunyaviricetes, include several important mosquito-borne viruses. Here, we aim to investigate the importance of different domains of Dcr2 for antiviral activity against viruses of the Bunyaviricetes. For this, we used the Aedes aegypti-derived Dcr2 knock-out cell line Aag2-AF319 to study the importance of the helicase, RNase III and PIWI-Argonaute-Zwille domains of Dcr2 on the antiviral activity of two viruses belonging to different families of the Bunyaviricetes: the Rift Valley fever virus (RVFV) vaccine strain MP12 (Phenuiviridae, Phlebovirus) and the Bunyamwera orthobunyavirus (BUNV; Peribunyaviridae, Orthobunyavirus). All three domains were determined to be critical for the antiviral activity against both RVFV and BUNV. Interestingly, one specific mutation in the helicase domain (KN) did not result in a loss of antiviral activity for RVFV, but for BUNV, despite losing the ability to produce 21 nt siRNAs.

Toscana virus - an emerging Mediterranean arbovirus transmitted by sand flies.

Toscana virus (TOSV) is an emerging arthropod-borne virus (arbovirus) of medical importance that is increasing its range across much of the Mediterranean Basin, Europe and the Middle East. Transmitted by Phlebotomus spp. sand flies, it is the most clinically relevant sand fly-borne phlebovirus. Initially isolated in the Tuscany region of Central Italy, it has now been detected in multiple countries that surround this geographical area. Infection of the vertebrate host can cause fever and neurological disease, following the dissemination of the virus to the brain. The prevalence is high in some regions, with a notable percentage of individuals showing seroconversion. TOSV can be a leading cause of acute meningitis and encephalitis (AME) during the summer months. In this comprehensive review, we will focus on several key topics. We discuss how TOSV has spread to establish outbreaks of infection in both humans and animals around the Mediterranean and the wider region. Clinical aspects of TOSV infection in humans are described, along with the best standards in diagnosis. Finally, we focus our discussion on the role of the sand fly vector, describing their biology, vector competency, implications for putative vertebrate reservoirs, the effect of the climate emergency on sand fly distribution and the putative role that sand fly-derived salivary factors may have on modulating host susceptibility to TOSV infection.

Auto-Abs neutralizing type I IFNs in patients with severe Powassan, Usutu, or Ross River virus disease.

Arboviral diseases are a growing global health concern. Pre-existing autoantibodies (auto-Abs) neutralizing type I interferons (IFNs) can underlie encephalitis due to West Nile virus (WNV) (∼40% of patients) and tick-borne encephalitis (TBE, due to TBE virus [TBEV]) (∼10%). We report here that these auto-Abs can also underlie severe forms of rarer arboviral infections. Auto-Abs neutralizing high concentrations of IFN-α2, IFN-β, and/or IFN-ω are present in the single case of severe Powassan virus (POWV) encephalitis studied, two of three cases of severe Usutu virus (USUV) infection studied, and the most severe of 24 cases of Ross River virus (RRV) disease studied. These auto-Abs are not found in any of the 137 individuals with silent or mild infections with these three viruses. Thus, auto-Abs neutralizing type I IFNs underlie an increasing list of severe arboviral diseases due to Flaviviridae (WNV, TBEV, POWV, USUV) or Togaviridae (RRV) viruses transmitted to humans by mosquitos (WNV, USUV, RRV) or ticks (TBEV, POWV).

Detection and evolutionary characterization of arboviruses in mosquitoes and biting midges on Hainan Island, China, 2019-2023.

We conducted a large-scale survey of arboviruses in mosquitoes and biting midges to assess the presence and spread of mosquito-borne pathogens currently circulating on Hainan Island, China. RT-PCR assays were used to detect the arbovirus species, distribution, and infection rates in mosquitoes and biting midges. Cell inoculation and high throughput sequencing were performed to isolate the viruses and assemble full viral genomes. Phylogenetic analysis was conducted to identify the viral genotypes and evolutionary relationships with known viruses. During 2019-2023, 32,632 mosquitoes and 21,000 biting midges were collected from 14 of 18 cities/counties on Hainan Island. Japanese encephalitis virus (JEV) was detected in Culex mosquitoes from five cities/counties, where the minimum infection rate (MIR) was 1.6 (0.6-2.6) per 1,000 females tested. Tembusu virus (TMUV) was detected in Culex mosquitoes from three cities/counties with MIR1.0 (0-2.2) per 1,000. Getah virus (GETV) was detected in Armigeres mosquitoes from Qionghai city with MIR 7.1 (0-15.2) per 1,000. Oya virus (OYAV) and Bluetongue virus (BTV) were detected in biting midges from Wanning city with MIRs of 0.4 (0-1.2) and 0.1 (0-10.2) per 1,000, respectively. Three JEV strains were isolated and clustered within the genotype I group, which is presently the dominant genotype in China. Three TMUV strains were isolated for the first time on Hainan Island that belonged to Cluster 3. Three isolated GETVs were identified as Group 3. BTV was reported for the first time on Hainan Island, and the complete genome for one BTV strain was successfully assembled, which was classified as serotype 1 based on the sequences of segment 2. These results stress the need to develop adequate surveillance plan measures to better control the public health threat of arboviruses carried by mosquitoes and biting midges in local regions.

A Growing Public Health Problem in Europe with Potential Severe Renal Involvement.

Dengue is an arboviral infection transmitted by the mosquito of the Aedes genus, widespread especially in tropical and subtropical regions but now with worldwide involvement. Cases of contagion are also progressively increasing in Europe, and the differential diagnosis with other infections is not always easy. Renal involvement with acute renal failure is possible and caused by the direct action of the virus, hemodynamic instability, rhabdomyolysis, or acute glomerular damage. In patients most at risk of renal involvement, there is high morbidity and mortality, with more extended hospital stays and follow-ups over time, which increases the burden on healthcare systems. Knowledge of this infection by nephrologists is essential for reducing morbidity, mortality, and, therefore, healthcare costs.

Psychological barriers in children: an exploratory study on Dengue transmission using an adapted DIPB scale.

Dengue is an arboviral infection found in tropical and subtropical regions transmitted by hematophagous mosquitoes from the genus Aedes spp. and responsible for millions of cases every year. Public campaigns and educational curriculum are designed to educate people, including children. However, what has been reported is that many decide not to follow these guidelines, even though they allegedly know what has to be done. To understand this phenomenon, this study aims to identify psychological barriers behind the adoption of pro-environmental behaviors that seek to reduce Aedes aegypti's population. For that, middle school students participated on two studies responsible for (1) adapting the Dragons of Inaction Psychological Barrier (DIPB) scale to the target group (n = 150) and then (2) testing it on a larger group (n = 449). In the exploratory factor analysis, Bartlett correlation (p < 0.001), Cronbach's alpha (0.83), and KMO analysis (overall MSA = 0.84) showed that data was suited for factor analysis. Five factors were retained by Kaiser Criterion and scree test (i.e., Conflicting goals and unnecessary changes-α 0.76, Interpersonal relations-α 0.72, Conflicting goals and lacking knowledge-α 0.58, Tokenism-α 0.73, and Tokenism toward the government-α 0.66). After that, the scale was tested across 11 different schools, where students also answered a questionnaire about the mosquito. Results suggested that the factors Conflicting goals and lacking knowledge and Tokenism toward the government presented a higher level of agreement for all students (means: 2.6 and 2.12 out of five, respectively). Those who scored higher in the mosquito's questionnaire had factors Conflicting goals and unnecessary change and Interpersonal relations inhibited when compared to others (p < 0.05). These results suggests that future educational campaigns should build different actions that focuses on addressing both internal and external factors, creating a mosaic of projects, with different goals, each aiming different environmental challenges.

Identification of a newly discovered virus from Culex and Armigeres mosquitoes in China

Mosquito associated virus have always been a significant threat to global health. Metagenomics offers a straightforward and quantitative means to acquire the information of novel virus and has greatly enriched the content of mosquito associated virus databases. During an entomological surveillance for arthropod-borne viruses in China, we identified a previously unrecognized virus from mosquitoes, temporarily named Huajieling virus. In this study, a total of 3,960 mosquitoes were collected and then divided into 91 pools, according to location and species. QRT-PCR and nested PCR were performed to confirm the presence of Huajieling virus. Its genomic features and phylogenetic relationships were further analyzed. Our results showed that Huajieling virus was detected in 7 of the 91 mosquito pools and that the minimum infection rate (MIR) was 0.18% (7/3,960). One complete genome sequence and 2 viral partial sequences were obtained from the Huajieling virus-positive pools. Pairwise distances analysis indicated that these amplified sequences shared high nucleotide identity. Phylogenetic analysis demonstrated that Huajieling virus is most closely related to Wufeng shrew picorna-like virus 43, which belonging to Picornavirales. Further analyses indicated that Huajieling virus is a new member of unclassified Picornavirales, and is intermediate between the family Caliciviridae and Secoviridae in taxonomic status.

Prevalence of Emerging Arboviral Infections: A Tertiary Care Hospital-Based Study from Kashmir, Northern India.

Background: Arboviruses are becoming a global public health menace. The common diseases worldwide caused by arboviruses are dengue, chikungunya, and Zika. This study aims to determine the prevalence of these three arboviral infections in patients with acute febrile illness at a tertiary care hospital in Kashmir, North India. Materials and Methods: A total of 812 blood samples were collected and tested for anti-dengue, anti-chikungunya immunoglobulin M (IgM), and dengue nonstructural protein 1 antigen by enzyme linked immunosorbent assay. Molecular testing of these samples was also done to detect dengue, chikungunya, and Zika viruses by using the CDC Trioplex real-time PCR assay. Results: The prevalence of diseases found among the studied patients was dengue 105/812 (12.93%) followed by chikungunya 17/812 (2.09%), and 3 cases (0.37%) were positive for both dengue and chikungunya; however, no case of Zika was detected. Interestingly, we found that only individuals that had a history of travel to different destinations within the country were positive for these viruses. Most affected cases were males 105/812 (12.93%) compared with females 20/812 (2.46%). Dengue serotyping results indicate that Dengue virus-1 was the most commonly found serotype. The most common symptoms in patients positive for dengue and chikungunya were fever, intense fatigue, myalgia, arthralgia, retro-orbital pain, anorexia, conjunctivitis, and skin rash. Conclusion: This study showed that dengue, chikungunya, and Zika viruses are not prevalent in the indigenous population of Kashmir. However, screening for these agents is required in people who have recently traveled outside Kashmir and have symptoms of acute febrile illness.

Rapid review: effectiveness of the Wolbachia strategy for arbovirus controlRevisión sistemática rápida: efectividad de la estrategia de control con Wolbachia en la lucha contra las arbovirosis

To identify and synthesize evidence of the effectiveness and cost-effectiveness of the Wolbachia-Aedes methodology for reducing cases of arboviral infection. Rapid systematic review. A search of five online portals and bibliographic databases was conducted in February 2022. Two reviewers conducted the selection stages, with any disagreements resolved by a third investigator. Data were extracted by one reviewer and subsequently checked by another. The inclusion criteria were studies describing the cost-effectiveness and effectiveness of the Wolbachia-Aedes methodology, based on field interventions, published in Portuguese, English, and Spanish, with no restrictions on date of publication. Four studies were included: one cluster randomized trial, two quasi-experimental studies, and one pre-post study, published between 2019 and 2021 in four countries. The included studies reported effectiveness for some of the outcomes of interest, such as a reduction in the incidence of dengue, Zika, and chikungunya cases. Although no experimental studies addressing the cost-effectiveness of the Wolbachia-Aedes strategy were identified, some modeling studies have demonstrated potential cost-benefit of this methodology. Considering the results of the included studies, Wolbachia has the potential to be an economically effective strategy that leads to reductions in the incidence of dengue, Zika, and chikungunya. Nevertheless, this strategy cannot yet be recommended as a public policy; additional large-scale studies with high methodological quality are still needed to inform political decision-making.

High-Sensitivity RT-LAMP for Molecular Detection of O'nyong-nyong (Alphavirus onyong).

Mosquitoes serve as vectors for many arthropod-borne viruses (arboviruses) that are responsible for millions of human infections and thousands of deaths each year. Among these arboviruses, O'nyong-nyong virus (ONNV) is an African alphavirus mainly transmitted by Anopheles mosquitoes. ONNV can be detected through serological or molecular tests, the first showing cross-reactivity to co-circulating alphaviruses and requiring technically demanding confirmation, while the latter, usually based on real-time PCR, are costly and demand specific equipment. Isothermal amplification approaches, such as Loop-Mediated Isothermal Amplification (LAMP), should therefore provide a cost-effective, sensitive, and specific alternative for virus detection, suitable for the resource-limited regions where ONNV circulates up to the present time. Here, we describe the development and optimization of a rapid and highly sensitive (10 pfu/reaction) RT-LAMP assay for ONNV detection. Additionally, we demonstrate that it is possible to bypass the RNA extraction step, reducing sample handling time and costs. The final RT-LAMPONNV is a promising field detection tool for ONNV, enabling a better understanding of its impact and serving as a point-of-care diagnostic method.

Dengue Fever in Urban Bangladesh: Atypical Presentations and Platelet Count Dynamics

Dengue fever, an arboviral infection transmitted by Aedes mosquitoes, remains a significant public health challenge in urban Bangladesh, especially in densely populated areas like Dhaka. This retrospective study investigates atypical clinical manifestations and platelet count dynamics among dengue patients, analyzing the association between thrombocytopenia and disease severity. Twenty patients treated in Dhaka were analyzed for clinical and hematological markers, with a specific focus on atypical presentations and platelet-hematocrit correlations. The findings highlight that while thrombocytopenia is a well-established indicator of dengue severity, several patients with critically low platelet counts did not exhibit typical warning signs, necessitating a more nuanced clinical approach.

Prevalence of Toscana Virus Antibodies in Residents of Bulgaria: A Nationwide Study Following the Pandemic of COVID-19.

Introduction: Toscana virus (TOSV) is an arthropod-borne virus transmitted by sandflies and is mainly found in countries around the Mediterranean basin. In this article, we present the findings of a seroprevalence study on TOSV in Bulgaria. We aim to assess the current epidemiological situation regarding TOSV in the country and raise clinical awareness. Methods: Serum samples (n = 1892) were collected in December 2023. Serological tests were performed using a commercial anti-TOSV ELISA kit. Results: Specific immunoglobulin G (IgG) antibodies were detected in 6.4% (121/1892) of the participants. A significantly higher seropositivity rate was found in the age group over 65 years compared with the age groups 18-29 and 40-64 (11.8% vs. 3.9% vs. 3.4%), as well as in males compared with females (8.0% vs. 5.3%). The seroprevalence rates in districts ranged from 0% to 18.5%. Higher seropositivity was found in the southern and northern regions. Conclusions: The seroprevalence rate of TOSV in Bulgaria, found in this study, is a significant decrease compared with the seropositivity rate of 24.5% reported in the country in 2018. The reasons for this are unknown and could possibly be related to the COVID-19 pandemic and the constantly changing environmental conditions. There is also a possibility that the higher seropositivity detected in 2018 together with the rise in clinical cases reported from endemic countries around that time might have been due to an unrecognized TOSV outbreak taking place in this period. Continued clinical awareness and surveillance are necessary for recognition and management of potential cases of TOSV neuroinfection, especially during summer.

Cerebellar Involvement in Dengue Fever

Abstract Dengue fever is a common and important arboviral infection prevalent in tropical countries, contributing to significant morbidity and mortality. The clinical spectrum of dengue fever ranges from simple febrile illness to life-threatening complications such as dengue hemorrhagic fever and dengue shock syndrome. Among the many unusual manifestations of this fever, a rare entity is its cerebellar involvement. This report aims to highlight this uncommon presentation.

Evaluation of the In2care Mosquito Station against Culex quinquefasciatus mosquitoes (Diptera: Culicidae)under semifield conditions.

Culex quinquefasciatus is an important mosquito vector responsible for the transmission of filarial worms, arthropod-borne viruses like Oropouche, St. Louis encephalitis, andWest Nile and protozoans that cause avian malaria. Due to insecticide resistance documented in Cx. quinquefasciatus populations worldwide, integrated vector management programs can benefit from new strategies to control this species. The In2Care Mosquito Station (In2Care station), a commercially available dissemination station containing pyriproxyfen (PPF) and Beauveria bassiana spores, has been shown to be effective against skip-ovipositing Aedes aegypti and Aedes albopictus in previously conducted semifield and field trials. To determine the potential of Cx. quinquefasciatus adult females to autodisseminate PPF and if the In2Care station could be used for Cx. quinquefasciatus control, we assessed its efficacy in a semifield setting against wild Cx. quinquefasciatus. We found that the In2Care station was attractive to gravid Cx. quinquefasciatus females, with a significantly higher percentage of egg rafts laid in the In2Care station compared to alternative ovipots. Adult females successfully autodisseminated PPF from the In2Care station to surrounding ovipots, leading to a significant increase in mosquito emergence inhibition. Additionally, adult Cx. quinquefasciatus exposure to B. bassiana spores significantly reduced mosquito survivorship. These results suggest that the In2Care station may be effective against Cx. quinquefasciatus in addition to Ae. aegypti and Ae. albopictus. Additional field evaluations are needed to assess impacts at the population level.

Spatio-temporal distribution of Dengue, Zika and Chikungunya in Cali, Colombia: 2014-2016

Introduction: Emerging and re-emerging arboviral infections have become a public health challenge in the Americas due to their epidemic potential. Objective: To determine the spatio-temporal distribution of the dengue, Zika, and chikungunya viruses during an epidemic period in Cali, Colombia. Materials and methods: Multi-method descriptive ecological and exploratory study of confirmed and suspected cases reported to the epidemiological surveillance system between 2014 and 2016. Results: 40,168 cases were analyzed, and it was found that dengue was the most frequent arboviral infection (59.2 %). The most affected individuals were women (65 %) and those with a mean age of 34.5 years. Although arboviral infections spread out throughout the city, the three diseases were concentrated in significant groups located at the center-east and northeast areas of Cali (p&lt;0.01; z=-203.7). Conclusions: This study identified critical zones for the three arboviral infections, which are located in areas with low socioeconomic status. Likewise, the results suggest that in addition to eco-epidemiological and bio-psychosocial factors, temperature, precipitation, and the aedic index may play an important role in the spatio-temporal behavior of these diseases. A multidisciplinary and collaborative approach is necessary, which must involve communities and authorities to implement effective control strategies, especially during epidemic periods.