Inhibition of the NMDA receptor/Nitric Oxide pathway in the dorsolateral periaqueductal gray causes anxiolytic-like effects in rats submitted to the Vogel conflict test

<p>Abstract</p> <p>Background</p> <p>Several studies had demonstrated the involvement of the dorsolateral portion of periaqueductal grey matter (dlPAG) in defensive responses. This region contains a significant number of neurons containing the enzyme nitric oxide synthase (NOS) and previous studies showed that non-selective NOS inhibition or glutamate NMDA-receptor antagonism in the dlPAG caused anxiolytic-like effects in the elevated plus maze.</p> <p>Methods</p> <p>In the present study we verified if the NMDA/NO pathway in the dlPAG would also involve in the behavioral suppression observed in rats submitted to the Vogel conflict test. In addition, the involvement of this pathway was investigated by using a selective nNOS inhibitor, Nω-propyl-L-arginine (N-Propyl, 0.08 nmol/200 nL), a NO scavenger, carboxy-PTIO (c-PTIO, 2 nmol/200 nL) and a specific NMDA receptor antagonist, LY235959 (4 nmol/200 nL).</p> <p>Results</p> <p>Intra-dlPAG microinjection of these drugs increased the number of punished licks without changing the number of unpunished licks or nociceptive threshold, as measure by the tail flick test.</p> <p>Conclusion</p> <p>The results indicate that activation of NMDA receptors and increased production of NO in the dlPAG are involved in the anxiety behavior displayed by rats in the VCT.</p

Co-infection between Zika and different Dengue serotypes during DENV outbreak in Brazil

Background: The recent introduction of new arboviruses in the Americas, as Zika virus (ZIKV) and Chikungunya virus (CHIKV), increased the risk of outbreaks and arboviral co-infections. Herein, we report twelve cases of co-infection of ZIKV and different DENV serotypes in a city located in the northwest region of São Paulo State, Brazil, which is hyper-endemic to Dengue. Methods: Between January and November 2016, 1254 suspected cases of arboviral infection were available by our surveillance program in São José do Rio Preto. All suspected patients were examined and, when they were arboviral disease-suspectd, had sera separated and viral RNA analyzed by PCR/qPCR assays to determine the diagnosis of DENV 1-4, ZIKV, or CHIKV in the same samples. After the molecular results, twelve patients with ZIKV-DENV coinfection were identified and their clinical and laboratory characteristics were described. Results: The mean between symptoms onset and collected sample of 3 days. DENV-1 was identified in seven co-infected patients and DEN2 in other five. Two patients presented alarm signs of Dengue and no one was hospitalized. Conclusions: The constant presence of co-circulating arboviruses increases the chance of co-infection and demonstrates the importance of the differential diagnosis, especially during periods of arboviral outbreaks. The impact of this co-infection is known individual and collectively. Keywords: Arbovirus, Arboviruses, Co-infection, Dengue, Zik

The eukaryotic translation initiation factor 3 subunit L protein interacts with Flavivirus NS5 and may modulate yellow fever virus replication

Background: Yellow fever virus (YFV) belongs to the Flavivirus genus and causes an important disease. An alarming resurgence of viral circulation and the expansion of YFV-endemic zones have been detected in Africa and South America in recent years. NS5 is a viral protein that contains methyltransferase and RNA-dependent RNA polymerase (RdRp) domains, which are essential for viral replication, and the interactions between NS5 and cellular proteins have been studied to better understand viral replication. The aim of this study was to characterize the interaction of the NS5 protein with eukaryotic translation initiation factor 3 subunit L (eIF3L) and to evaluate the role of eIF3L in yellow fever replication. Methods. To identify interactions of YFV NS5 with cellular proteins, we performed a two-hybrid screen using the YFV NS5 RdRp domain as bait with a human cDNA library, and RNApol deletion mutants were generated and analyzed using the two-hybrid system for mapping the interactions. The RNApol region involved was segmented into three fragments and analyzed using an eIF3L-expressing yeast strain. To map the NS5 residues that are critical for the interactions, we performed site-direct mutagenesis in segment 3 of the interaction domain (ID) and confirmed the interaction using in vitro assays and in vivo coimmunoprecipitation. The significance of eIF3L for YFV replication was investigated using eIF3L overexpression and RNA interference. Results: In this work, we describe and characterize the interaction of NS5 with the translation factor eIF3L. The interaction between NS5 and eIF3L was confirmed using in vitro binding and in vivo coimmunoprecipitation assays. This interaction occurs at a region (the interaction domain of the RNApol domain) that is conserved in several flaviviruses and that is, therefore, likely to be relevant to the genus. eIF3L overexpression and plaque reduction assays showed a slight effect on YFV replication, indicating that the interaction of eIF3L with YFV NS5 may play a role in YFV replication. Conclusions: Although the precise function of eIF3L on interactions with viral proteins is not entirely understood, these results indicate an interaction of eIF3L with YF NS5 and that eIF3L overexpression facilitates translation, which has potential implications for virus replication. © 2013 Morais et al.; licensee BioMed Central Ltd

Long-Term Viruria in Zika Virus–Infected Pregnant Women, Brazil, 2016

During the 2016 Zika virus outbreak in Brazil, we detected Zika virus RNA in urine samples collected from Zika virus–positive pregnant women during different stages of pregnancy. Women had positive and negative intervals of viruria; 3 newborns had adverse outcomes. Further research is needed to clarify the relationship between viruria and outcomes for newborns

Systems Biology Reveals NS4B-Cyclophilin A Interaction: A New Target to Inhibit YFV Replication

Yellow fever virus (YFV) replication is highly dependent on host cell factors. YFV NS4B is reported to be involved in viral replication and immune evasion. Here interactions between NS4B and human proteins were determined using a GST pull-down assay and analyzed using 1-DE and LC–MS/MS. We present a total of 207 proteins confirmed using Scaffold 3 Software. Cyclophilin A (CypA), a protein that has been shown to be necessary for the positive regulation of flavivirus replication, was identified as a possible NS4B partner. 59 proteins were found to be significantly increased when compared with a negative control, and CypA exhibited the greatest difference, with a 22-fold change. Fisher’s exact test was significant for 58 proteins, and the <i>p</i> value of CypA was the most significant (0.000000019). The Ingenuity Systems software identified 16 pathways, and this analysis indicated sirolimus, an mTOR pathway inhibitor, as a potential inhibitor of CypA. Immunofluorescence and viral plaque assays showed a significant reduction in YFV replication using sirolimus and cyclosporine A (CsA) as inhibitors. Furthermore, YFV replication was strongly inhibited in cells treated with both inhibitors using reporter BHK-21-rep-YFV17D-LucNeoIres cells. Taken together, these data suggest that CypA-NS4B interaction regulates YFV replication. Finally, we present the first evidence that YFV inhibition may depend on NS4B-CypA interaction

Why Did ZIKV Perinatal Outcomes Differ in Distinct Regions of Brazil? An Exploratory Study of Two Cohorts

The Zika virus (ZIKV) epidemic in Brazil occurred in regions where dengue viruses (DENV) are historically endemic. We investigated the differences in adverse pregnancy/infant outcomes in two cohorts comprising 114 pregnant women with PCR-confirmed ZIKV infection in Rio de Janeiro, Southeastern Brazil (n = 50) and Manaus, in the north region of the country (n = 64). Prior exposure to DENV was evaluated through plaque reduction neutralizing antibody assays (PRNT 80) and DENV IgG serologies. Potential associations between pregnancy outcomes and Zika attack rates in the two cities were explored. Overall, 31 women (27%) had adverse pregnancy/infant outcomes, 27 in Rio (54%) and 4 in Manaus (6%), p &lt; 0.001. This included 4 pregnancy losses (13%) and 27 infants with abnormalities at birth (24%). A total of 93 women (82%) had evidence of prior DENV exposure, 45 in Rio (90%) and 48 in Manaus (75%). Zika attack rates differed; the rate in Rio was 10.28 cases/10,000 and in Manaus, 0.6 cases/10,000, p &lt; 0.001. Only Zika attack rates (Odds Ratio: 17.6, 95% Confidence Interval 5.6-55.9, p &lt; 0.001) and infection in the first trimester of pregnancy (OR: 4.26, 95% CI 1.4-12.9, p = 0.011) were associated with adverse pregnancy and infant outcomes. Pre-existing immunity to DENV was not associated with outcomes (normal or abnormal) in patients with ZIKV infection during pregnancy

Why Did ZIKV Perinatal Outcomes Differ in Distinct Regions of Brazil? An Exploratory Study of Two Cohorts

The Zika virus (ZIKV) epidemic in Brazil occurred in regions where dengue viruses (DENV) are historically endemic. We investigated the differences in adverse pregnancy/infant outcomes in two cohorts comprising 114 pregnant women with PCR-confirmed ZIKV infection in Rio de Janeiro, Southeastern Brazil (n = 50) and Manaus, in the north region of the country (n = 64). Prior exposure to DENV was evaluated through plaque reduction neutralizing antibody assays (PRNT 80) and DENV IgG serologies. Potential associations between pregnancy outcomes and Zika attack rates in the two cities were explored. Overall, 31 women (27%) had adverse pregnancy/infant outcomes, 27 in Rio (54%) and 4 in Manaus (6%), p &lt; 0.001. This included 4 pregnancy losses (13%) and 27 infants with abnormalities at birth (24%). A total of 93 women (82%) had evidence of prior DENV exposure, 45 in Rio (90%) and 48 in Manaus (75%). Zika attack rates differed; the rate in Rio was 10.28 cases/10,000 and in Manaus, 0.6 cases/10,000, p &lt; 0.001. Only Zika attack rates (Odds Ratio: 17.6, 95% Confidence Interval 5.6–55.9, p &lt; 0.001) and infection in the first trimester of pregnancy (OR: 4.26, 95% CI 1.4–12.9, p = 0.011) were associated with adverse pregnancy and infant outcomes. Pre-existing immunity to DENV was not associated with outcomes (normal or abnormal) in patients with ZIKV infection during pregnancy