Irene Bosch - Academia.edu (original) (raw)

Papers by Irene Bosch

TRAIL is a novel antiviral protein against dengue virus

This article cites 47 articles, 25 of which can be accessed free

Nature Neuroscience, 2020

Zika virus (ZIKV) is a flavivirus linked to multiple birth defects including microcephaly, known ... more Zika virus (ZIKV) is a flavivirus linked to multiple birth defects including microcephaly, known as congenital ZIKV syndrome. The identification of host factors involved in ZIKV replication may guide efficacious therapeutic interventions. In genome-wide transcriptional studies, we found that ZIKV infection triggers aryl hydrocarbon receptor (AHR) activation. Specifically, ZIKV infection induces kynurenine (Kyn) production, which activates AHR, limiting the production of type I interferons (IFN-I) involved in antiviral immunity. Moreover, ZIKV-triggered AHR activation suppresses intrinsic immunity driven by the promyelocytic leukemia (PML) protein, which limits ZIKV replication. AHR inhibition suppressed the replication of multiple ZIKV strains in vitro and also suppressed replication of the related flavivirus dengue. Finally, AHR inhibition with a nanoparticle-delivered AHR antagonist or an inhibitor developed for human use limited ZIKV replication and ameliorated newborn microcephaly in a murine model. In summary, we identified AHR as a host factor for ZIKV replication and PML protein as a driver of anti-ZIKV intrinsic immunity. NATuRE NEuRoSCIENCE | www.nature.com/natureneuroscience Articles Nature NeuroscieNce Results AHR signaling is activated by ZIKV infection. The liver is a primary target of human ZIKV infection 10. To identify molecular mechanisms associated with ZIKV replication, we analyzed, using RNA-sequencing (RNA-seq) messenger RNA expression in liver-derived human HepG2 cells infected with ZIKV. We detected 179 differentially expressed genes related to cellular processes such as protein translation, cell cycle control, energy metabolism and autophagy (Fig. 1a-c and see also Supplementary Table 1). These processes have been shown to be targeted by ZIKV and other flaviviruses to promote their replication 11,12. In addition, using Ingenuity Pathway Analysis (IPA), we detected significant effects of ZIKV infection on signaling by the transcription factor AHR (Fig. 1c). Indeed, AHR was identified as a regulator of the transcriptional response of HepG2 cells to ZIKV infection (Fig. 1d and see also Supplementary Table 2). In validation studies on independent samples we detected increased expression of the AHR transcriptional targets CYP1A1 and CYP1B1 in ZIKV-infected HepG2 cells (Fig. 1e). These observations were further validated by analyzing RNA-seq data available in public databases, which detected upregulated AHR signaling in ZIKV-infected human brain organoids 13 and also in full-term placentas from ZIKV-infected women 14 (see Extended Data Fig. 1a-c). Based on the potential for therapeutic modulation of AHR signaling by small molecules, and our own interest in the role of AHR in immunity, we explored the role of AHR in ZIKV replication. AHR is a ligand-activated transcription factor with transcriptional activity that requires both AHR expression and the presence of AHR agonists 15. Kyn is a tryptophan (Trp)-derived AHR ligand produced in the context of cancer and inflammation by the enzymes indoleamine 2,3-dioxygenase 1 and 2 (IDO1 and IDO2) and tryptophan 2,3-dioxygenase (TDO2) (Fig. 1f) 16. We did not detect IDO1 or IDO2 expression in ZIKV-infected HepG2 cells. However, ZIKV infection upregulated TDO2 expression and increased Kyn levels in culture supernatants, whereas it decreased the levels of Trp used as a substrate by TDO2 to generate Kyn (Fig. 1g,h). ZIKV replicates in neural progenitor cells (NPCs) 12 as part of the disease mechanisms thought to promote brain abnormalities in Zika congenital syndrome 17. Indeed, we detected 49% of infection in human primary NPCs exposed to ZIKV (see Extended Data Fig. 2a). NPCs express AHR (Fig. 1i and see also Extended Data Fig. 2b); the expression of AHR, its transcriptional target CYP1A1, and IDO1 and TDO2, which promote the synthesis of the AHR agonist Kyn, was upregulated by ZIKV infection (Fig. 1i). Of note, AHR mRNA expression was upregulated in ZIKV-infected NPCs but not in HepG2 cells (Fig. 1e,i), potentially reflecting intrinsic differences between these two cell types. Finally, the increase in AHR, CYP1A1, IDO1 and TDO2 expression in NPCs was dependent on the multiplicity of infection (MOI) (Fig. 1j), and it was also reflected as an increase in Kyn levels in culture supernatants (Fig. 1k). Taken together, these findings suggest that ZIKV-triggered Kyn production activates AHR.

Annual Review of Biomedical Engineering, 2020

Rapid diagnostic tests (point-of-care devices) are critical components of informed patient care a... more Rapid diagnostic tests (point-of-care devices) are critical components of informed patient care and public health monitoring (surveillance applications). We propose that among the many rapid diagnostics platforms that have been tested or are in development, lateral flow immunoassays and synthetic biology–based diagnostics (including CRISPR-based diagnostics) represent the best overall options given their ease of use, scalability for manufacturing, sensitivity, and specificity. This review describes the identification of lateral flow immunoassay monoclonal antibody pairs that detect and distinguish between closely related pathogens and that are used in combination with functionalized multicolored nanoparticles and computational methods to deconvolute data. We also highlight the promise of synthetic biology–based diagnostic tests, which use synthetic genetic circuits that activate upon recognition of a pathogen-associated nucleic acid sequence, and discuss how the combined or parallel...

PLOS Neglected Tropical Diseases, 2020

Nature Neuroscience, 2020

In the version of this article initially published, two plasmids were listed incorrectly in the M... more In the version of this article initially published, two plasmids were listed incorrectly in the Methods section. Plasmid phDLX1-N174 (#66859) should be #60859, and phDLX2-N174 (#66860) should be 60860. The error has been corrected in the PDF and HTML versions of this article.

International Journal of Infectious Diseases, 2019

To evaluate the clinical, laboratory, and immune characteristics of Zika virus (ZIKV)-associated ... more To evaluate the clinical, laboratory, and immune characteristics of Zika virus (ZIKV)-associated encephalitis in pediatric patients after the epidemic in Huila, southern Colombia. Methods: A pediatric neuro-surveillance hospital study was conducted in a referral health center in southern Colombia, from October 2016 to October 2017. Cases of encephalitis were confirmed by nucleic acid amplification tests and serological methods in cerebrospinal fluid (CSF), plasma, and/or urine. Levels of six cytokines were evaluated by flow cytometry. Patients underwent daily clinical and laboratory follow-up. Results: Twenty children with probable encephalitis were included for further studies and 16 of them were confirmed. Four cases of bacterial meningoencephalitis (Streptococcus pneumoniae, group B Streptococcus, Staphylococcus epidermidis, and Escherichia coli) and 12 cases of viral encephalitis were identified, six of them associated with ZIKV infection. Other viral encephalitis cases were caused by herpes viruses (n = 3), enterovirus (n = 2), and dengue virus type 2 (DENV-2; n = 1) infections. ZIKVassociated encephalitis symptoms subsided faster than those of patients with encephalitis caused by other agents. CSF analysis revealed lymphocytic pleocytosis. Compared to healthy controls, children with ZIKV-associated encephalitis presented modest plasma interleukin (IL)-10 but not IL-2, IL-4, IL-6, interferon gamma (IFN-g), or tumor necrosis factor alpha (TNF-α). Cytokine expression was differentially regulated, as dramatically elevated IL-6, IL-10, and IFN-g levels were observed in CSF but not in paired plasma samples in one of the patients with ZIKV detectable in CSF. Conclusions: This study provides evidence that ZIKV is responsible for pediatric encephalitis in endemic areas, and the local presence of the virus may induce cephalic but not systemic expression of cytokines.

Proceedings of the National Academy of Sciences of the United States of America, Jul 3, 2018

Maternal Zika virus (ZIKV) infection during pregnancy is recognized as the cause of an epidemic o... more Maternal Zika virus (ZIKV) infection during pregnancy is recognized as the cause of an epidemic of microcephaly and other neurological anomalies in human fetuses. It remains unclear how ZIKV accesses the highly vulnerable population of neural progenitors of the fetal central nervous system (CNS), and which cell types of the CNS may be viral reservoirs. In contrast, the related dengue virus (DENV) does not elicit teratogenicity. To model viral interaction with cells of the fetal CNS in vitro, we investigated the tropism of ZIKV and DENV for different induced pluripotent stem cell-derived human cells, with a particular focus on microglia-like cells. We show that ZIKV infected isogenic neural progenitors, astrocytes, and microglia-like cells (pMGLs), but was only cytotoxic to neural progenitors. Infected glial cells propagated ZIKV and maintained ZIKV load over time, leading to viral spread to susceptible cells. DENV triggered stronger immune responses and could be cleared by neural an...

Science (New York, N.Y.), Apr 27, 2018

Mitigating global infectious disease requires diagnostic tools that are sensitive, specific, and ... more Mitigating global infectious disease requires diagnostic tools that are sensitive, specific, and rapidly field deployable. In this study, we demonstrate that the Cas13-based SHERLOCK (specific high-sensitivity enzymatic reporter unlocking) platform can detect Zika virus (ZIKV) and dengue virus (DENV) in patient samples at concentrations as low as 1 copy per microliter. We developed HUDSON (heating unextracted diagnostic samples to obliterate nucleases), a protocol that pairs with SHERLOCK for viral detection directly from bodily fluids, enabling instrument-free DENV detection directly from patient samples in <2 hours. We further demonstrate that SHERLOCK can distinguish the four DENV serotypes, as well as region-specific strains of ZIKV from the 2015-2016 pandemic. Finally, we report the rapid (<1 week) design and testing of instrument-free assays to detect clinically relevant viral single-nucleotide polymorphisms.

Science translational medicine, Jan 27, 2017

The recent Zika virus (ZIKV) outbreak demonstrates that cost-effective clinical diagnostics are u... more The recent Zika virus (ZIKV) outbreak demonstrates that cost-effective clinical diagnostics are urgently needed to detect and distinguish viral infections to improve patient care. Unlike dengue virus (DENV), ZIKV infections during pregnancy correlate with severe birth defects, including microcephaly and neurological disorders. Because ZIKV and DENV are related flaviviruses, their homologous proteins and nucleic acids can cause cross-reactions and false-positive results in molecular, antigenic, and serologic diagnostics. We report the characterization of monoclonal antibody pairs that have been translated into rapid immunochromatography tests to specifically detect the viral nonstructural 1 (NS1) protein antigen and distinguish the four DENV serotypes (DENV1-4) and ZIKV without cross-reaction. To complement visual test analysis and remove user subjectivity in reading test results, we used image processing and data analysis for data capture and test result quantification. Using a 30-μ...

Maternal Zika virus (ZIKV) infection during pregnancy is increasingly recognized as the cause of ... more Maternal Zika virus (ZIKV) infection during pregnancy is increasingly recognized as the cause of an epidemic of microcephaly and other neurological anomalies in human fetuses. However, it remains unclear how ZIKV gains access to the highly vulnerable population of neural progenitors of the fetal central nervous system (CNS), and which cell types of the CNS may serve as viral reservoirs. To model viral interaction with cells of the fetal CNS invitro, we investigated the tropism of ZIKV for different iPS-derived human cells, with a particular focus on microglia-like cells derived from human pluripotent stem cells. We show that ZIKV infected isogenic neural progenitors, astrocytes and microglia-like cells, but was only cytotoxic to neural progenitors. Infected glial cells propagated the virus and maintained viral load over time, leading to viral spread to susceptible cells. ZIKV-infected microglia, when co-cultured with pre-established neural spheroids, invaded the tissue and initiated...

Host Gene Responses to Infections, 2009

PLoS ONE, 2013

Global dengue virus spread in tropical and subtropical regions has become a major international p... more Global dengue virus spread in tropical and subtropical regions has become a major international public health concern. It is evident that DENV genetic diversity plays a significant role in the immunopathology of the disease and that the identification of polymorphisms associated with adaptive responses is important for vaccine development. The investigation of naturally occurring genomic variants may play an important role in the comprehension of different adaptive strategies used by these mutants to evade the human immune system. In order to elucidate this role we sequenced the complete polyprotein-coding region of thirty-three DENV-3 isolates to characterize variants circulating under high endemicity in the city of São José de Rio Preto, Brazil, during the onset of the 2006-07 epidemic. By inferring the evolutionary history on a local-scale and estimating rates of synonymous (dS) and nonsynonimous (dN) substitutions, we have documented at least two different introductions of DENV-3 into the city and detected 10 polymorphic codon sites under significant positive selection (dN/dS. 1) and 8 under significant purifying selection (dN/dS , 1). We found several polymorphic amino acid coding sites in the envelope (15), NS1 (17), NS2A (11), and NS5 (24) genes, which suggests that these genes may be experiencing relatively recent adaptive changes. Furthermore, some polymorphisms correlated with changes in the immunogenicity of several epitopes. Our study highlights the existence of significant and informative DENV variability at the spatio-temporal scale of an urban outbreak.

PLoS ONE, 2013

Global dengue virus spread in tropical and subtropical regions has become a major international p... more Global dengue virus spread in tropical and subtropical regions has become a major international public health concern. It is evident that DENV genetic diversity plays a significant role in the immunopathology of the disease and that the identification of polymorphisms associated with adaptive responses is important for vaccine development. The investigation of naturally occurring genomic variants may play an important role in the comprehension of different adaptive strategies used by these mutants to evade the human immune system. In order to elucidate this role we sequenced the complete polyprotein-coding region of thirty-three DENV-3 isolates to characterize variants circulating under high endemicity in the city of São José de Rio Preto, Brazil, during the onset of the 2006-07 epidemic. By inferring the evolutionary history on a local-scale and estimating rates of synonymous (dS) and nonsynonimous (dN) substitutions, we have documented at least two different introductions of DENV-3 into the city and detected 10 polymorphic codon sites under significant positive selection (dN/dS. 1) and 8 under significant purifying selection (dN/dS , 1). We found several polymorphic amino acid coding sites in the envelope (15), NS1 (17), NS2A (11), and NS5 (24) genes, which suggests that these genes may be experiencing relatively recent adaptive changes. Furthermore, some polymorphisms correlated with changes in the immunogenicity of several epitopes. Our study highlights the existence of significant and informative DENV variability at the spatio-temporal scale of an urban outbreak.

Memórias do Instituto Oswaldo Cruz, 2012

Journal of General Virology, 2009

The depletion ofl-tryptophan (L-Trp) has been associated with the inhibition of growth of micro-o... more The depletion ofl-tryptophan (L-Trp) has been associated with the inhibition of growth of micro-organisms and also has profound effects on T cell proliferation and immune tolerance. The enzyme indoleamine 2,3-dioxygenase (IDO) catalyses the rate-limiting step in the catabolic pathway of L-Trp. Gene expression analysis has shown upregulation of genes involved in L-Trp catabolism inin vitromodels of dengue virus (DENV) infection. To understand the role of IDO during DENV infection, we measured IDO activity in sera from control and DENV-infected patients. We found increased IDO activity, lower levels of L-Trp and higher levels ofl-kynurenine in sera from DENV-infected patients during the febrile days of the disease compared with patients with other febrile illnesses and healthy donors. Furthermore, we confirmed upregulation of IDO mRNA expression in response to DENV infectionin vitro, using a dendritic cell (DC) model of DENV infection. We found that the antiviral effect of gamma inter...

International Journal of Infectious Diseases, 2012

International Journal of Infectious Diseases, 2012

TRAIL is a novel antiviral protein against dengue virus

This article cites 47 articles, 25 of which can be accessed free

Nature Neuroscience, 2020

Zika virus (ZIKV) is a flavivirus linked to multiple birth defects including microcephaly, known ... more Zika virus (ZIKV) is a flavivirus linked to multiple birth defects including microcephaly, known as congenital ZIKV syndrome. The identification of host factors involved in ZIKV replication may guide efficacious therapeutic interventions. In genome-wide transcriptional studies, we found that ZIKV infection triggers aryl hydrocarbon receptor (AHR) activation. Specifically, ZIKV infection induces kynurenine (Kyn) production, which activates AHR, limiting the production of type I interferons (IFN-I) involved in antiviral immunity. Moreover, ZIKV-triggered AHR activation suppresses intrinsic immunity driven by the promyelocytic leukemia (PML) protein, which limits ZIKV replication. AHR inhibition suppressed the replication of multiple ZIKV strains in vitro and also suppressed replication of the related flavivirus dengue. Finally, AHR inhibition with a nanoparticle-delivered AHR antagonist or an inhibitor developed for human use limited ZIKV replication and ameliorated newborn microcephaly in a murine model. In summary, we identified AHR as a host factor for ZIKV replication and PML protein as a driver of anti-ZIKV intrinsic immunity. NATuRE NEuRoSCIENCE | www.nature.com/natureneuroscience Articles Nature NeuroscieNce Results AHR signaling is activated by ZIKV infection. The liver is a primary target of human ZIKV infection 10. To identify molecular mechanisms associated with ZIKV replication, we analyzed, using RNA-sequencing (RNA-seq) messenger RNA expression in liver-derived human HepG2 cells infected with ZIKV. We detected 179 differentially expressed genes related to cellular processes such as protein translation, cell cycle control, energy metabolism and autophagy (Fig. 1a-c and see also Supplementary Table 1). These processes have been shown to be targeted by ZIKV and other flaviviruses to promote their replication 11,12. In addition, using Ingenuity Pathway Analysis (IPA), we detected significant effects of ZIKV infection on signaling by the transcription factor AHR (Fig. 1c). Indeed, AHR was identified as a regulator of the transcriptional response of HepG2 cells to ZIKV infection (Fig. 1d and see also Supplementary Table 2). In validation studies on independent samples we detected increased expression of the AHR transcriptional targets CYP1A1 and CYP1B1 in ZIKV-infected HepG2 cells (Fig. 1e). These observations were further validated by analyzing RNA-seq data available in public databases, which detected upregulated AHR signaling in ZIKV-infected human brain organoids 13 and also in full-term placentas from ZIKV-infected women 14 (see Extended Data Fig. 1a-c). Based on the potential for therapeutic modulation of AHR signaling by small molecules, and our own interest in the role of AHR in immunity, we explored the role of AHR in ZIKV replication. AHR is a ligand-activated transcription factor with transcriptional activity that requires both AHR expression and the presence of AHR agonists 15. Kyn is a tryptophan (Trp)-derived AHR ligand produced in the context of cancer and inflammation by the enzymes indoleamine 2,3-dioxygenase 1 and 2 (IDO1 and IDO2) and tryptophan 2,3-dioxygenase (TDO2) (Fig. 1f) 16. We did not detect IDO1 or IDO2 expression in ZIKV-infected HepG2 cells. However, ZIKV infection upregulated TDO2 expression and increased Kyn levels in culture supernatants, whereas it decreased the levels of Trp used as a substrate by TDO2 to generate Kyn (Fig. 1g,h). ZIKV replicates in neural progenitor cells (NPCs) 12 as part of the disease mechanisms thought to promote brain abnormalities in Zika congenital syndrome 17. Indeed, we detected 49% of infection in human primary NPCs exposed to ZIKV (see Extended Data Fig. 2a). NPCs express AHR (Fig. 1i and see also Extended Data Fig. 2b); the expression of AHR, its transcriptional target CYP1A1, and IDO1 and TDO2, which promote the synthesis of the AHR agonist Kyn, was upregulated by ZIKV infection (Fig. 1i). Of note, AHR mRNA expression was upregulated in ZIKV-infected NPCs but not in HepG2 cells (Fig. 1e,i), potentially reflecting intrinsic differences between these two cell types. Finally, the increase in AHR, CYP1A1, IDO1 and TDO2 expression in NPCs was dependent on the multiplicity of infection (MOI) (Fig. 1j), and it was also reflected as an increase in Kyn levels in culture supernatants (Fig. 1k). Taken together, these findings suggest that ZIKV-triggered Kyn production activates AHR.

Annual Review of Biomedical Engineering, 2020

Rapid diagnostic tests (point-of-care devices) are critical components of informed patient care a... more Rapid diagnostic tests (point-of-care devices) are critical components of informed patient care and public health monitoring (surveillance applications). We propose that among the many rapid diagnostics platforms that have been tested or are in development, lateral flow immunoassays and synthetic biology–based diagnostics (including CRISPR-based diagnostics) represent the best overall options given their ease of use, scalability for manufacturing, sensitivity, and specificity. This review describes the identification of lateral flow immunoassay monoclonal antibody pairs that detect and distinguish between closely related pathogens and that are used in combination with functionalized multicolored nanoparticles and computational methods to deconvolute data. We also highlight the promise of synthetic biology–based diagnostic tests, which use synthetic genetic circuits that activate upon recognition of a pathogen-associated nucleic acid sequence, and discuss how the combined or parallel...

PLOS Neglected Tropical Diseases, 2020

Nature Neuroscience, 2020

In the version of this article initially published, two plasmids were listed incorrectly in the M... more In the version of this article initially published, two plasmids were listed incorrectly in the Methods section. Plasmid phDLX1-N174 (#66859) should be #60859, and phDLX2-N174 (#66860) should be 60860. The error has been corrected in the PDF and HTML versions of this article.

International Journal of Infectious Diseases, 2019

To evaluate the clinical, laboratory, and immune characteristics of Zika virus (ZIKV)-associated ... more To evaluate the clinical, laboratory, and immune characteristics of Zika virus (ZIKV)-associated encephalitis in pediatric patients after the epidemic in Huila, southern Colombia. Methods: A pediatric neuro-surveillance hospital study was conducted in a referral health center in southern Colombia, from October 2016 to October 2017. Cases of encephalitis were confirmed by nucleic acid amplification tests and serological methods in cerebrospinal fluid (CSF), plasma, and/or urine. Levels of six cytokines were evaluated by flow cytometry. Patients underwent daily clinical and laboratory follow-up. Results: Twenty children with probable encephalitis were included for further studies and 16 of them were confirmed. Four cases of bacterial meningoencephalitis (Streptococcus pneumoniae, group B Streptococcus, Staphylococcus epidermidis, and Escherichia coli) and 12 cases of viral encephalitis were identified, six of them associated with ZIKV infection. Other viral encephalitis cases were caused by herpes viruses (n = 3), enterovirus (n = 2), and dengue virus type 2 (DENV-2; n = 1) infections. ZIKVassociated encephalitis symptoms subsided faster than those of patients with encephalitis caused by other agents. CSF analysis revealed lymphocytic pleocytosis. Compared to healthy controls, children with ZIKV-associated encephalitis presented modest plasma interleukin (IL)-10 but not IL-2, IL-4, IL-6, interferon gamma (IFN-g), or tumor necrosis factor alpha (TNF-α). Cytokine expression was differentially regulated, as dramatically elevated IL-6, IL-10, and IFN-g levels were observed in CSF but not in paired plasma samples in one of the patients with ZIKV detectable in CSF. Conclusions: This study provides evidence that ZIKV is responsible for pediatric encephalitis in endemic areas, and the local presence of the virus may induce cephalic but not systemic expression of cytokines.

Proceedings of the National Academy of Sciences of the United States of America, Jul 3, 2018

Maternal Zika virus (ZIKV) infection during pregnancy is recognized as the cause of an epidemic o... more Maternal Zika virus (ZIKV) infection during pregnancy is recognized as the cause of an epidemic of microcephaly and other neurological anomalies in human fetuses. It remains unclear how ZIKV accesses the highly vulnerable population of neural progenitors of the fetal central nervous system (CNS), and which cell types of the CNS may be viral reservoirs. In contrast, the related dengue virus (DENV) does not elicit teratogenicity. To model viral interaction with cells of the fetal CNS in vitro, we investigated the tropism of ZIKV and DENV for different induced pluripotent stem cell-derived human cells, with a particular focus on microglia-like cells. We show that ZIKV infected isogenic neural progenitors, astrocytes, and microglia-like cells (pMGLs), but was only cytotoxic to neural progenitors. Infected glial cells propagated ZIKV and maintained ZIKV load over time, leading to viral spread to susceptible cells. DENV triggered stronger immune responses and could be cleared by neural an...

Science (New York, N.Y.), Apr 27, 2018

Mitigating global infectious disease requires diagnostic tools that are sensitive, specific, and ... more Mitigating global infectious disease requires diagnostic tools that are sensitive, specific, and rapidly field deployable. In this study, we demonstrate that the Cas13-based SHERLOCK (specific high-sensitivity enzymatic reporter unlocking) platform can detect Zika virus (ZIKV) and dengue virus (DENV) in patient samples at concentrations as low as 1 copy per microliter. We developed HUDSON (heating unextracted diagnostic samples to obliterate nucleases), a protocol that pairs with SHERLOCK for viral detection directly from bodily fluids, enabling instrument-free DENV detection directly from patient samples in <2 hours. We further demonstrate that SHERLOCK can distinguish the four DENV serotypes, as well as region-specific strains of ZIKV from the 2015-2016 pandemic. Finally, we report the rapid (<1 week) design and testing of instrument-free assays to detect clinically relevant viral single-nucleotide polymorphisms.

Science translational medicine, Jan 27, 2017

The recent Zika virus (ZIKV) outbreak demonstrates that cost-effective clinical diagnostics are u... more The recent Zika virus (ZIKV) outbreak demonstrates that cost-effective clinical diagnostics are urgently needed to detect and distinguish viral infections to improve patient care. Unlike dengue virus (DENV), ZIKV infections during pregnancy correlate with severe birth defects, including microcephaly and neurological disorders. Because ZIKV and DENV are related flaviviruses, their homologous proteins and nucleic acids can cause cross-reactions and false-positive results in molecular, antigenic, and serologic diagnostics. We report the characterization of monoclonal antibody pairs that have been translated into rapid immunochromatography tests to specifically detect the viral nonstructural 1 (NS1) protein antigen and distinguish the four DENV serotypes (DENV1-4) and ZIKV without cross-reaction. To complement visual test analysis and remove user subjectivity in reading test results, we used image processing and data analysis for data capture and test result quantification. Using a 30-μ...

Maternal Zika virus (ZIKV) infection during pregnancy is increasingly recognized as the cause of ... more Maternal Zika virus (ZIKV) infection during pregnancy is increasingly recognized as the cause of an epidemic of microcephaly and other neurological anomalies in human fetuses. However, it remains unclear how ZIKV gains access to the highly vulnerable population of neural progenitors of the fetal central nervous system (CNS), and which cell types of the CNS may serve as viral reservoirs. To model viral interaction with cells of the fetal CNS invitro, we investigated the tropism of ZIKV for different iPS-derived human cells, with a particular focus on microglia-like cells derived from human pluripotent stem cells. We show that ZIKV infected isogenic neural progenitors, astrocytes and microglia-like cells, but was only cytotoxic to neural progenitors. Infected glial cells propagated the virus and maintained viral load over time, leading to viral spread to susceptible cells. ZIKV-infected microglia, when co-cultured with pre-established neural spheroids, invaded the tissue and initiated...

Host Gene Responses to Infections, 2009

PLoS ONE, 2013

Global dengue virus spread in tropical and subtropical regions has become a major international p... more Global dengue virus spread in tropical and subtropical regions has become a major international public health concern. It is evident that DENV genetic diversity plays a significant role in the immunopathology of the disease and that the identification of polymorphisms associated with adaptive responses is important for vaccine development. The investigation of naturally occurring genomic variants may play an important role in the comprehension of different adaptive strategies used by these mutants to evade the human immune system. In order to elucidate this role we sequenced the complete polyprotein-coding region of thirty-three DENV-3 isolates to characterize variants circulating under high endemicity in the city of São José de Rio Preto, Brazil, during the onset of the 2006-07 epidemic. By inferring the evolutionary history on a local-scale and estimating rates of synonymous (dS) and nonsynonimous (dN) substitutions, we have documented at least two different introductions of DENV-3 into the city and detected 10 polymorphic codon sites under significant positive selection (dN/dS. 1) and 8 under significant purifying selection (dN/dS , 1). We found several polymorphic amino acid coding sites in the envelope (15), NS1 (17), NS2A (11), and NS5 (24) genes, which suggests that these genes may be experiencing relatively recent adaptive changes. Furthermore, some polymorphisms correlated with changes in the immunogenicity of several epitopes. Our study highlights the existence of significant and informative DENV variability at the spatio-temporal scale of an urban outbreak.

PLoS ONE, 2013

Global dengue virus spread in tropical and subtropical regions has become a major international p... more Global dengue virus spread in tropical and subtropical regions has become a major international public health concern. It is evident that DENV genetic diversity plays a significant role in the immunopathology of the disease and that the identification of polymorphisms associated with adaptive responses is important for vaccine development. The investigation of naturally occurring genomic variants may play an important role in the comprehension of different adaptive strategies used by these mutants to evade the human immune system. In order to elucidate this role we sequenced the complete polyprotein-coding region of thirty-three DENV-3 isolates to characterize variants circulating under high endemicity in the city of São José de Rio Preto, Brazil, during the onset of the 2006-07 epidemic. By inferring the evolutionary history on a local-scale and estimating rates of synonymous (dS) and nonsynonimous (dN) substitutions, we have documented at least two different introductions of DENV-3 into the city and detected 10 polymorphic codon sites under significant positive selection (dN/dS. 1) and 8 under significant purifying selection (dN/dS , 1). We found several polymorphic amino acid coding sites in the envelope (15), NS1 (17), NS2A (11), and NS5 (24) genes, which suggests that these genes may be experiencing relatively recent adaptive changes. Furthermore, some polymorphisms correlated with changes in the immunogenicity of several epitopes. Our study highlights the existence of significant and informative DENV variability at the spatio-temporal scale of an urban outbreak.

Memórias do Instituto Oswaldo Cruz, 2012

Journal of General Virology, 2009

The depletion ofl-tryptophan (L-Trp) has been associated with the inhibition of growth of micro-o... more The depletion ofl-tryptophan (L-Trp) has been associated with the inhibition of growth of micro-organisms and also has profound effects on T cell proliferation and immune tolerance. The enzyme indoleamine 2,3-dioxygenase (IDO) catalyses the rate-limiting step in the catabolic pathway of L-Trp. Gene expression analysis has shown upregulation of genes involved in L-Trp catabolism inin vitromodels of dengue virus (DENV) infection. To understand the role of IDO during DENV infection, we measured IDO activity in sera from control and DENV-infected patients. We found increased IDO activity, lower levels of L-Trp and higher levels ofl-kynurenine in sera from DENV-infected patients during the febrile days of the disease compared with patients with other febrile illnesses and healthy donors. Furthermore, we confirmed upregulation of IDO mRNA expression in response to DENV infectionin vitro, using a dendritic cell (DC) model of DENV infection. We found that the antiviral effect of gamma inter...

International Journal of Infectious Diseases, 2012

International Journal of Infectious Diseases, 2012