Presence Of Zika Virus Research Articles

Evaluation of the effects of the Zika Virus-Immunoglobulin G+ complex on murine microglial cells.

After the Zika virus (ZIKV) epidemic in Brazil, ZIKV infections were linked to damage to the central nervous system (CNS) and congenital anomalies. Due to the virus's ability to cross the placenta and reach brain tissue, its effects become severe, leading to Congenital Zika Syndrome (CZS) and resulting in neuroinflammation, microglial activation, and secretion of neurotoxic factors. The presence of ZIKV triggers an inadequate fetal immune response, as the fetus only has the protection of maternal antibodies of the Immunoglobulin G (IgG) class, which are the only antibodies capable of crossing the placenta. Because of limited understanding regarding the long term consequences of ZIKV infection and the involvement of maternal antibodies, this study sought to assess the impact of the ZIKV + IgG⁺complex on murine microglial cells. The cells were exposed to ZIKV, IgG antibodies, and the ZIKV + IgG⁺complex for 24 and 72h. Treatment-induced cytotoxic effects were evaluated using the cell viability assay, oxidative stress, and mitochondrial membrane potential. The findings indicated that IgG antibodies exhibit cytotoxic effects on microglia, whether alone or in the presence of ZIKV, leading to compromised cell viability, disrupted mitochondrial membrane potential, and heightened oxidative damage. Our conclusion is that IgG antibodies exert detrimental effects on microglia, triggering their activation and potentially disrupting the creation of a neurotoxic environment. Moreover, the presence of antibodies may correlate with an elevated risk of ZIKV-induced neuroinflammation, contributing to long-term CNS damage.

Negevirus Piura Suppresses Zika Virus Replication in Mosquito Cells.

We investigated the interaction between the insect-specific virus, Piura virus (PIUV), and the arbovirus Zika virus (ZIKV) in Aedes albopictus cells. We performed coinfection experiments in C6/36 cells. Piura virus (Cor 33 strain, Colombia) and ZIKV (PRVABC58 strain, Puerto Rico) were co-inoculated into C6/36 cells using two multiplicity of infection (MOI) combinations: 0.1 for both viruses and 1.0 for ZIKV, 0.1 for PIUV. Wells were infected in triplicate with either PIUV and ZIKV coinfection, ZIKV-only, or PIUV-only. Mock infected cells served as control wells. The cell suspension was collected daily 7 days post-infection. Zika virus load was titrated by TCID50 on Vero 76 cells. The ZIKV-only infection and PIUV and ZIKV coinfection experiments were also quantified by RT-qPCR. We also investigated whether ZIKV interfered in the PIUV replication. PIUV suppressed the replication of ZIKV, resulting in a 10,000-fold reduction in ZIKV titers within 3 days post-infection. PIUV viral loads were not reduced in the presence of ZIKV. We conclude that, when concurrently infected, PIUV suppresses ZIKV in C6/36 cells while ZIKV does not interfere in PIUV replication.

Association of Viral Control and Maternal Antibody Titers in Congenital Zika Virus Infection

Abstract Background Exposure to Zika virus (ZIKV) in utero can result in a wide spectrum of outcomes, ranging from asymptomatic to severe neurological defects or fetal demise. The presence of virus in the fetus or in the mother’s blood for a prolonged period of time may lead to worse outcomes, which means that limiting the duration of fetal exposure to virus may improve outcomes. One factor that may be related to control of maternal viremia and presence of virus in the fetus is the maternal IgG antibody response. We utilized a rhesus macaque model of prenatal ZIKV to quantify the viral RNA present in the maternal blood and maternal-fetal interface (MFI) tissues and determine the association of viral RNA loads with maternal IgG antibody titers over time. Methods Dams were infected with either African lineage ZIKV-DAKAR (n=9) or Asian lineage ZIKV-PRVABC59 (PR, n=10) isolate in the first trimester, then all fetal and MFI tissues were removed either 7 or 14 days post-infection (dpi) and viral RNA loads were measured. Maternal plasma was tested for viral RNA and IgG throughout pregnancy. Results A greater proportion of animals infected with the DAKAR isolate had viral RNA present in the MFI tissues compared with the animals infected with the PR isolate, at both 7 and 14 dpi. DAKAR animals also tested positive for ZIKV-specific IgG at 14 dpi or earlier, while IgG was typically negative at 14 dpi for PR animals. Viral RNA was detected in the fetal and MFI tissues in a higher proportion of dams that still had ZIKV RNA positive plasma at 14 dpi had viral RNA. We found no difference in IgG antibody titers between dams that were ZIKV positive or negative at 14 dpi. Conclusions The presence of ZIKV in the MFI tissues and earlier detection of ZIKV-specific maternal IgG responses were both associated with the DAKAR isolate, suggesting that it may be a more virulent isolate. A higher proportion of dams that had virus in their plasma at 14 dpi also had virus present in the fetal and MFI tissues, but there was no difference in long term IgG response within either isolate between animals that tested positive or negative at 14 dpi. This suggests that the dam clearing viral RNAfrom their plasma by 14 dpi may impact whether or not the virus infects the MFI tissues, but that maternal IgG response likely does not play a role in this difference.

Zika virus E protein modulates functions of human brain microvascular endothelial cells and astrocytes: implications on blood-brain barrier properties.

Neurotropic viruses can cross the otherwise dynamically regulated blood-brain barrier (BBB) and affect the brain cells. Zika virus (ZIKV) is an enveloped neurotropic Flavivirus known to cause severe neurological complications, such as encephalitis and fetal microcephaly. In the present study, we employed human brain microvascular endothelial cells (hBMECs) and astrocytes derived from human progenitors to establish a physiologically relevant BBB model. We used this model to investigate the effects of ZIKV envelope (E) protein on properties of cells comprising the BBB. E protein is the principal viral protein involved in interaction with host cell surface receptors, facilitating the viral entry. Our findings show that the presence of ZIKV E protein leads to activation of both hBMECs and astrocytes. In hBMECs, we observed a decrease in the expression of crucial endothelial junction proteins such as ZO-1, Occludin and VE-Cadherin, which are vital in establishment and maintenance of the BBB. Consequently, the ZIKV E protein induced changes in BBB integrity and permeability. We also found upregulation of genes involved in leukocyte recruitment along with increased proinflammatory chemokines and cytokines upon exposure to E protein. Additionally, the E protein also led to astrogliosis, evident from the elevated expression of GFAP and Vimentin. Both cell types comprising the BBB exhibited inflammatory response upon exposure to E protein which may influence viral access into the central nervous system (CNS) and subsequent infection of other CNS cells. Overall, our study provides valuable insights into the transient changes that occur at the site of BBB upon ZIKV infection.

Japanese Encephalitis Vaccine Generates Cross-Reactive Memory T Cell Responses to Zika Virus in Humans.

Zika virus (ZIKV) and Japanese encephalitis virus (JEV) are mosquito-borne flaviviruses with sequence homology. ZIKV circulates in some regions where JEV also circulates, or where JE vaccination is used. Cross-immunity between flaviviruses exists, but the precise mechanisms remain unclear. We previously demonstrated that T cell immunity induced by the live-attenuated Japanese encephalitis (JE) SA14-14-2 vaccine conferred protective immunity against ZIKV infection in mice, which could even bypass antibody-dependent enhancement. However, the role of T cell immune, especially memory T cell subsets, in cross-reactive immune responses between JE vaccine and ZIKV in humans has not been reported. We examined central and effector memory CD4+ and CD8+ T cell (TCM and TEM) responses (including degranulation, cytokines, and chemokines) in the presence of JEV and ZIKV, respectively, by using qualified peripheral blood mononuclear cell samples from 18 children who had recently received a two-dose course of JE vaccine SA14-14-2 as well as seven children without JE vaccination. Cross-reactive CD8+ TCM in response to ZIKV was characterized by secretion of IFN-γ, whereas CD8+ TEM did not show significant upregulation of functional factors. In the presence of ZIKV, IFN-γ and TNF-α expression was upregulated by CD4+ TEM, and the expression signature of CD4+ TCM is more cytotoxic potential. We profiled the cross-reactive memory T cell responses to ZIKV in JE vaccine recipients. These data will provide evidence for the mechanism of cross-reactive memory T cell immune responses between JEV and ZIKV and a more refined view of bivalent vaccine design strategy.

Identification of viral RNA sequences in vampire bats (Desmodus rotundus) from central Mexico

Bats are important reservoirs of viral entities that cause diverse economical and health problems in several sectors. The purpose of this study was to determine the prevalence of the rabies virus in vampire bats. We also tested for 3 other zoonotic viruses in the collected tissue. We captured 45 vampire bats (Desmodus rotundus), which were measured and sexed under standard parameters. We followed an RT-PCR reaction to amplify the viral products of four virus types: chikungunya (CHIKV), dengue (DENV), rabies (RABV) and Zika virus (ZIKV) from cerebral tissue. We obtained an amplicon of 100 bp for ZIKV in 2 samples, representing a prevalence of 4.4%. An amplicon of 581 bp for RABV was observed in 9 individuals, which is equivalent to 20% of our sample. We obtained 4 different haplotypes for RABV and a single haplotype for ZIKV. We did not record the presence of DENV and CHIKV. We corroborated the presence of the family Rhabdoviridae in the vampire bat and recorded the presence of ZIKV for the first time in this bat species.

Evidence of Spreading Zika Virus Infection Caused by Males of Different Species.

Zika virus (ZIKV) is a positive-sense single-stranded RNA flavivirus and is mainly transmitted by Aedes mosquitoes. This arbovirus has had a significant impact on health in recent years by causing malformations, such as microcephaly in babies and Guillain–Barré syndrome in adults. Some evidence indicates that ZIKV can be sexually transmitted and may persist in the male reproductive tract for an extended period in humans. Knockout and vasectomized mice have been used as models to reveal ZIKV infection in the male reproductive tract as a virus source. ZIKV presence in male and female mosquito reproductive tracts and eggs point to venereal and vertical/transovarian transmission, again demonstrating that the reproductive tract can be involved in the spread of ZIKV. Moreover, eggs protected by eggshells have the potential to be a ZIKV reservoir. Given the +-lack of vaccines and therapies for Zika fever and the underestimated prevalence rate, an understanding of ZIKV infection and its spread from the reproductive tract, which is protected from the immune system and potentially active for virus transmission, is imperative. We must also develop cheaper, more efficient techniques for virological surveillance inside vectors and humans, control vectors with ecofriendly insecticides, and promote condom use to avoid ZIKV contamination during sexual intercourse, as recommended by the World Health Organization.

The Extrinsic Incubation Period of Zika Virus in Florida Mosquitoes Aedes aegypti and Ae. albopictus.

The Asian genotype of Zika virus (ZIKV) emerged in Brazil in 2015 and subsequently spread throughout the Americas. In July 2016, Florida experienced its first locally acquired ZIKV infection in the continental U.S. Concerns about health risks from ZIKV infection have increased the need to investigate the interactions between potential mosquito vectors and ZIKV. The time it takes for an arbovirus to propagate within a mosquito, and become transmissible, is the extrinsic incubation period (EIP). The EIP for potential mosquito vectors in Florida is unknown. To address this gap in the understanding of ZIKV epidemiology, Florida Aedes aegypti (L.) and Ae. albopictus (Skuse) were orally exposed to ZIKV infected blood meals and fully engorged mosquitoes were held at a constant temperature of 28 °C through the duration of the experiment. Saliva expectorates were collected from cohorts of mosquitoes and tested for the presence of ZIKV at three-day intervals over a period of 24 days to allow for an evaluation of the EIP of the emergent Asian lineage of ZIKV. High rates of infected bodies in Ae. albopictus (75–94%) and Ae. aegypti (68–86%) were observed throughout the incubation period, which did not differ by species. Higher rates of disseminated infection were observed later during the incubation period but did not differ between species. We calculated the 50% EIP to be shorter in Ae. albopictus than Ae. aegypti (16.2 and 18.2 days post infection, respectively). The competence for ZIKV observed in both species may contribute to high rates of ZIKV transmission in Florida populations.

The clinical spectrum and immunopathological mechanisms underlying ZIKV-induced neurological manifestations.

Since the 2015 to 2016 outbreak in America, Zika virus (ZIKV) infected almost 900,000 patients. This international public health emergency was mainly associated with a significant increase in the number of newborns with congenital microcephaly and abnormal neurologic development, known as congenital Zika syndrome (CZS). Furthermore, Guillain–Barré syndrome (GBS), a neuroimmune disorder of adults, has also been associated with ZIKV infection. Currently, the number of ZIKV-infected patients has decreased, and most of the cases recently reported present as a mild and self-limiting febrile illness. However, based on its natural history of a typical example of reemerging pathogen and the lack of specific therapeutic options against ZIKV infection, new outbreaks can occur worldwide, demanding the attention of researchers and government authorities. Here, we discuss the clinical spectrum and immunopathological mechanisms underlying ZIKV-induced neurological manifestations. Several studies have confirmed the tropism of ZIKV for neural progenitor stem cells by demonstrating the presence of ZIKV in the central nervous system (CNS) during fetal development, eliciting a deleterious inflammatory response that compromises neurogenesis and brain formation. Of note, while the neuropathology of CZS can be due to a direct viral neuropathic effect, adults may develop neuroimmune manifestations such as GBS due to poorly understood mechanisms. Antiganglioside autoantibodies have been detected in multiple patients with ZIKV infection–associated GBS, suggesting a molecular mimicry. However, further additional immunopathological mechanisms remain to be uncovered, paving the way for new therapeutic strategies.

Forebrain Neural Precursor Cells Are Differentially Vulnerable to Zika Virus Infection.

Prenatal exposure to Zika virus (ZIKV) can result in microencephaly and congenital Zika syndrome, although some brain cells and structures are spared by the virus for unknown reasons. Here, a novel murine model of fetal ZIKV infection incorporating intraventricular infection and cell type-specific in utero electroporation (IUE) was used to identify the time course of ZIKV infection and to determine the identity of cells that are initially infected or spared during neocortical neurogenesis. In vivo time course studies revealed the presence of ZIKV in apical radial glial cells (aRGCs) at early time points following virus exposure, while basal intermediate progenitor cells (bIPCs) became maximally (ZIKV+) after 3 d of virus exposure. ZIKV-infected fetal brains exhibited microencephaly as early as 1 d following infection, regardless of developmental age. This change in brain size was caused in part by apoptosis and reduced proliferation that persisted until birth. While 60% of aRGC basal fibers were perturbed during infection, 40% retained normal morphology, indicating that aRGCs are not uniformly vulnerable to ZIKV infection. To investigate this heterogeneous vulnerability, we performed genetic fate mapping using cell type-specific probes derived from a mouse embryonic day (E)15.5 neocortical wall single-cell RNA sequencing (scRNAseq) dataset. The results indicate that one class of aRGCs preferentially express the putative ZIKV entry receptor AXL and that these cells are more vulnerable to ZIKV infection than other aRGC subtypes with low AXL expression. Together, these data uncover crucial temporal and cellular details of ZIKV fetal brain infection for prevention strategies and for management of congenital Zika syndrome.

Spatiotemporal Heterogeneity of Zika Virus Transmission in Indonesia: Serosurveillance Data from a Pediatric Population.

.The presence of Zika virus (ZIKV) in Indonesia has been recognized since the 1970s, but its transmission dynamics there have been poorly understood. To understand more fully the geographic distribution and burden of ZIKV infection, we performed retrospective serological tests on specimens collected from asymptomatic children age 5 to 9 years old living at 30 sites in 14 provinces. Of 870 serum samples tested, 9.2% were found to be positive for anti-ZIKV antibodies, as confirmed by plaque reduction neutralization assays. This was the same overall prevalence reported previously for 1- to 4-year-old children collected at the same sites at the same time. Together with geographic differences in seroprevalence between the age groups, these data suggest that, although ZIKV might be endemic in Indonesia, its occurrence has been focal and episodic.

Microcephaly Outcomes among Zika Virus-Infected Pregnant Women in Honduras.

The impact of Zika virus (ZIKV) infection on pregnancies shows regional variation emphasizing the importance of studies in different geographical areas. We conducted a prospective study in Tegucigalpa, Honduras, recruiting 668 pregnant women between July 20, 2016, and December 31, 2016. We performed Trioplex real-time reverse transcriptase-PCR (rRT-PCR) in 357 serum samples taken at the first prenatal visit. The presence of ZIKV was confirmed in seven pregnancies (7/357, 2.0%). Nine babies (1.6%) had microcephaly (head circumference more than two SDs below the mean), including two (0.3%) with severe microcephaly (head circumference [HC] more than three SDs below the mean). The mothers of both babies with severe microcephaly had evidence of ZIKV infection. A positive ZIKV Trioplex rRT-PCR was associated with a 33.3% (95% CI: 4.3-77.7%) risk of HC more than three SDs below the mean.

Pregnant Women Infected with Zika Virus Show Higher Viral Load and Immunoregulatory Cytokines Profile with CXCL10 Increase.

Background: Zika virus (ZIKV) infection during pregnancy usually shows only mild symptoms and is frequently subclinical. However, it can be vertically transmitted to the fetus, causing microcephaly and other congenital defects. During pregnancy, the immune environment modifications can alter the response to viruses in general and ZIKV in particular. Objective: To describe the role of pregnancy in the systemic pro- and anti-inflammatory response during symptomatic ZIKV infection. Materials and Methods: A multiplex assay was used to measure 25 cytokines, chemokines, and receptors in 110 serum samples from pregnant and nonpregnant women with and without ZIKV infection with and without symptoms. Samples were collected through an epidemiological surveillance system. Results: Samples from pregnant women with ZIKV infection showed a higher viral load but had similar profiles of inflammatory markers as compared with nonpregnant infected women, except for CXCL10 that was higher in infected pregnant women. Notably, the presence of ZIKV in pregnancy favored a regulatory profile by significantly increasing anti-inflammatory cytokines such as interleukin (IL)-10, receptors IL-1RA, and IL-2R, but only those pro-inflammatory cytokines such as IL-6, interferon (IFN)-α, IFN-γ and IL-17 that are essential for the antiviral response. Interestingly, there were no differences between symptomatic and weakly symptomatic ZIKV-infected groups. Conclusion: Our results revealed a systemic anti-inflammatory cytokine and chemokine profile that could participate in the control of the virus. The anti-inflammatory response in pregnant women infected with ZIKA was characterized by high CXCL10, a cytokine that has been correlated with congenital malformations.

Clinico-epidemiological and genomic profile of first Zika Virus outbreak in India at Jaipur city of Rajasthan state

BackgroundFirst Zika virus (ZIKV) positive case from North India was detected on routine surveillance of Dengue-Like Illness in an 85-year old female. Objective of the study was to conduct an investigation for epidemiological, clinical and genomic analysis of first ZIKV outbreak in Rajasthan, North India and enhance routine ZIKV surveillance. MethodOutbreak investigation was performed in 3 Km radius of the index case among patient contacts, febrile cases, and pregnant women. Routine surveillance was enhanced to include samples from various districts of Rajasthan. Presence of ZIKV in serum and urine samples was detected by real time PCR test and CDC trioplex kit. Few ZIKV positive samples were sequenced using the next-generation sequencing method for genomic analysis. ResultOn outbreak investigation 153/2043 (7.48%) cases were found positive: 1/153 (0.65%) among contacts, 90/153 (58.8%) in fever cases, 62/153(40.5%) in pregnant females. In routine surveillance, 6/4722 (0.12%) serum samples were ZIKV positive.Majority of patients had mild signs and symptoms, no case of microcephaly and Guillain- Barre Syndrome was seen, 25 (40.3%) pregnant females delivered healthy babies, four (6.4%) reported abortion and three (4.8%) had intrauterine death, one (1.6%) child had colorectal malformation and died after few days of birth. ZIKV was found to belong to Asian lineage, mutation related to enhanced neuro-virulence and transmission in animal models was not found. ConclusionZIKV was endogenous to India belonging to Asian Lineage. Disease profile of the ZIKV was asymptomatic to mild. No major anomaly was observed in infants born to ZIKV positive mothers; however, long term follow up of these children is required. There is need to scale up surveillance in the virology lab network of India for early detection and control. Summary lineZika virus infection was endogenous due to Asian Lineage with mild disease, no case of microcephaly or Guillain- Barre Syndrome was seen but children need to be followed for anomalies and surveillance of ZIKV needs to be enhanced in the country.

Zika virus in rhesus macaque semen and reproductive tract tissues: a pilot study of acute infection†.

Although sexual transmission of Zika virus (ZIKV) is well-documented, the viral reservoir(s) in the male reproductive tract remains uncertain in humans and immune-intact animal models. We evaluated the presence of ZIKV in a rhesus macaque pilot study to determine persistence in semen, assess the impact of infection on sperm functional characteristics, and define the viral reservoir in the male reproductive tract. Five adult male rhesus monkeys were inoculated with 105 PFU of Asian-lineage ZIKV isolate PRVABC59, and two males were inoculated with the same dose of African-lineage ZIKV DAKAR41524. Viremia and viral RNA (vRNA) shedding in semen were monitored, and a cohort of animals were necropsied for tissue collection to assess tissue vRNA burden and histopathology. All animals exhibited viremia for limited periods (1-11days); duration of shedding did not differ significantly between viral isolates. There were sporadic low levels of vRNA in the semen from some, but not all animals. Viral RNA levels in reproductive tract tissues were also modest and present in the epididymis in three of five cases, one case in the vas deferens, but not detected in testis, seminal vesicles or prostate. ZIKV infection did not impact semen motility parameters as assessed by computer-assisted sperm analysis. Despite some evidence of prolonged ZIKV RNA shedding in human semen and high tropism of ZIKV for male reproductive tract tissues in mice deficient in Type 1 interferon signaling, in the rhesus macaques assessed in this pilot study, we did not consistently find ZIKV RNA in the male reproductive tract.

Surveillance of Zika virus in field-caught Aedes aegypti and Aedes albopictus suggests important role of male mosquitoes in viral populations maintenance in Medellín, Colombia

Due to the rapid spread of Zika virus (ZIKV) infection after its emergence in the Americas in 2015 and its relationship with birth defects, it became declared a Public Health Emergency of International Concern (WHO). The main mechanism by which this virus circulates in nature is horizontal transmission between vectors and humans. However, it has been suggested that vertical transmission (parent to offspring infection) or venereal mosquito-mosquito transmission may have an important role in viral populations maintenance during inter-epidemic periods. In this study we evaluate the presence of ZIKV in males and females of Aedes aegypti and Ae. albopictus in Medellín, Colombia, throughout the post-epidemic period of 2017 and 2018. A total of 7986 mosquitoes Aedes sp. resting within houses were captured and grouped in 2768 pools; 146 of these were RT-PCR positive for ZIKV, of which 38 (26%) were male mosquito pools (36 of Ae. aegypti and 2 of Ae. albopictus). The partial NS5 gene was sequenced in all ZIKV PCR-positive pools to confirm the ZIKV presence throughout spatial and temporal sampling. The results suggest a vector role of ZIKV by Ae. Albopictus; and because it is well known that male mosquitoes are not hematophagous, the high rate detection of ZIKV in male Aedes mosquitoes pools supports the existence of vertical or venereal transmission in Medellín, which can contribute to ZIKV maintenance during low transmission periods. This study provides a better understanding of the population dynamics of ZIKV in an endemic region during an inter-epidemic period and supports alternative transmission pathways as a mechanism to maintain endemism of this arbovirus.

A preliminary survey of Zika virus infection by nucleic acid test in the volunteer blood donor samples in Shenzhen China.

Although Zika virus (ZIKV) infection is mainly transmitted through mosquito bite, it can also be transmitted through blood transfusion. More than 500 000 cases of ZIKV infection were reported in the Americas from 2015 to 2016. Up till now, over 10 cases of imported ZIKV infection have been reported due to frequent international exchanges in the Shenzhen city of Guangdong Province, China. Unfortunately, there were no data on ZIKV infection in Chinese blood donors because it has not been included in routine screening for volunteer blood donors. As such, we performed a preliminary survey of the prevalence of ZIKV infection among volunteer blood donors in Shenzhen, China, to assess the potential risk of ZIKV infection through transfusion. A total of 9626 blood donor samples were collected and ZIKA RNA was detected by transcription-mediated amplification (TMA) nucleic acid amplification method with the Panther nucleic acid automatic analysis system of Grifols,Spain, including Procleix ZIKV Assay reagent. All the experiments in this study were conducted in accordance with the standard operating procedure of the blood center. Of the 9626 donor blood samples tested, none of these samples was Zika RNA reactive. There was no positive case from ZIKV RNA screening in this preliminary survey. There was no ZIKV presence in blood donors in Shenzhen, China, from this preliminary survey. The potential risk of ZIKV infection by transfusion is low in Shenzhen at this moment. Therefore, there is no need to add ZIKV nucleic acid test as a routine screening for blood donors.

First report on the transmission of Zika virus by Aedes (Stegomyia) aegypti (L.) (Diptera: Culicidae) during the 2018 Zika outbreak in India

In a recent outbreak of Zika virus (ZIKV) infection in Jaipur city (Rajasthan, India), a total of 159 cases were reported in September 2018. In order to identify vector responsible for Zika transmission, mosquitoes were collected from houses with reported Zika cases and nearby houses. A total of 108 pools containing 522 mosquitoes were tested for presence of ZIKV using RT-PCR and Real Time RT-PCR. We detected presence of ZIKV in three pools of Aedes (Stegomyia) aegypti (L.), out of a total of 79 pools with 383 Ae. aegypti through RT-PCR as well as real-time RT-PCR. The presence of ZIKV in Ae. aegypti was further confirmed by DNA sequencing of the partial envelope region of ZIKV. Homology search of DNA sequence revealed highest identity (100%) with a ZIKV isolate from human from the study area which support the role of Ae. aegypti acting as a ZIKV vector. All other mosquitoes (Aedes vittatus and Culex quinquefasciatus) were negative for ZIKV. None of the F1 generation mosquito pools (276 mosquitoes in 43 pools) were found positive. This is the first report of presence of ZIKV in Ae. aegypti from the Indian subcontinent.

Vertical transmission of Zika virus in Culex quinquefasciatus Say and Aedes aegypti (L.) mosquitoes

Several mosquito species have been described as vectors for the Zika virus (ZIKV), such as those in the Aedes, Anopheles, Mansonia and Culex genera. Our previous survey studies were found the ZIKV RNA positive in both male, female and larvae of Culex quinquefasciatus Say and Aedes aegypti (L.) mosquitoes collected from active ZIKV infected patients’ homes in Thailand. Therefore, the aims of this study were to investigate whether ZIKV could be vertically transmitted in Cx. quinquefasciatus, Ae. aegypti and Ae. albopictus. Laboratory and field colonies of these mosquito species were maintained and artificially fed with ZIKV in human blood. Fully engorged mosquitoes (F0) were selected and reared for the vertical transmission study. The subsequent mosquito generations were fed with human blood without the virus. ZIKV in the mosquitoes was detected by hemi-nested RT-PCR and sequencing. C6/36 cells were used to isolate ZIKV from samples that tested positive by hemi-nested RT-PCR. Moreover, ZIKV was identified by immunocytochemical staining 7 days after infection in several organs of infected F0 females, including the salivary glands, midguts, yoke granules and facet cells of the eye. The localization of the ZIKV antigen was identified by the presence of the specific antibody in the salivary glands, midguts, yoke granules and facet cells. ZIKV was detected in female and male Cx. quinquefasciatus until the F6 and F2 generations, respectively. The isolated virus showed cytopathic effects in C6/36 cells by 5 days postinfection. The results suggested that the vertical transmission of ZIKV occurs in Cx. quinquefasciatus in the laboratory. However, we were able to detect the presence of ZIKV in Ae. aegypti in only the F1 generation in both male and female mosquitoes, and Ae. albopictus mosquitoes were not able to vertically transmit the virus at all. Data obtained from this study could be valuable for developing a better understanding of the role of Cx. quinquefasciatus as a potential vector for ZIKV transmission in Thailand and may be useful in creating more effective mosquito vector control strategies in the future.

Methods for detecting Zika virus in feces: A case study in captive squirrel monkeys (Saimiri boliviensis boliviensis).

A strain of Zika virus (ZIKV) of Asian origin associated with birth defects and neurological disorders has emerged and spread through the Americas. ZIKV was first isolated in the blood of nonhuman primates in Africa and has been detected in the blood, saliva, and urine of a few catarrhine species in both Africa and Asia, suggesting that nonhuman primates may serve as both a source and a reservoir of the virus. The recent introduction of ZIKV to human populations in the Americas presents the potential for the virus to spread into nonhuman primate reservoirs. Thus, it is critical to develop efficient and noninvasive detection methods to monitor the spread of the virus in wild nonhuman primate populations. Here, we describe a method for ZIKV detection in noninvasively collected fecal samples of a Neotropical primate. Fecal samples were collected from two captive squirrel monkeys (Saimiri boliviensis boliviensis) that were experimentally infected with ZIKV (Strain Mexico_1_44) and an additional two uninfected squirrel monkeys. Nucleic acids were extracted from these samples, and RT-qPCR was used to assay for the presence of ZIKV using primers flanking a 101 bp region of the NS5 gene. In both ZIKV-inoculated animals, ZIKV was detected 5–11 days post-infection, but was not detected in the uninfected animals. We compare the fecal results to ZIKV detection in serum, saliva, and urine samples from the same individuals. Our results indicate that fecal detection is a cost-effective, noninvasive method for monitoring wild populations of Neotropical primates as possible ZIKV reservoirs.