Characterization of CRESS-DNA viruses in human vaginal secretions: An exploratory metagenomic investigation.

The Phylum Cressdnaviricota consists of a large number of circular Rep-encoding single-stranded (CRESS)-DNA viruses. Recently, metagenomic analyzes revealed their ubiquitous distribution in a diverse range of eukaryotes. Data relating to CRESS-DNA viruses in humans remains scarce. Our study investigated the presence and genetic diversity of CRESS-DNA viruses in human vaginal secretions. Vaginal swabs were collected from 28 women between 29 and 43 years old attending a fertility clinic in New York City. An exploratory metagenomic analysis was performed and detection of CRESS-DNA viruses was confirmed through analysis of near full-length sequences of the viral isolates. A phylogenetic tree was based on the REP open reading frame sequences of the CRESS-DNA virus genome. Eleven nearly complete CRESS-DNA viral genomes were identified in 16 (57.1%) women. There were no associations between the presence of these viruses and any demographic or clinical parameters. Phylogenetic analysis indicated that one of the sequences belonged to the genus Gemycircularvirus within the Genomoviridae family, while ten sequences represented previously unclassified species of CRESS-DNA viruses. Novel species of CRESS-DNA viruses are present in the vaginal tract of adult women. Although they be transient commensal agents, the potential clinical implications for their presence at this site cannot be dismissed.

T cell-mediated Immune response and correlates of inflammation and their relationship with COVID-19 clinical severity: not an intuitive guess.

BACKGROUND: Predictors of the outcome of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection remain to be fully determined. We evaluated selected viral characteristics and immunological responses that might predict and/or correlate to the clinical outcome of COVID-19. METHODS: For individuals developing divergent clinical outcomes, the magnitude and breadth of T cell-mediated responses were measured within 36 h of symptom onset. Peripheral Blood Mononuclear Cells (PBMCs) were subjected to in vitro stimulation with SARS-CoV-2-based peptides. In addition, SARS-CoV-2 sequences were generated by metagenome, and HLA typing was performed using Luminex technology. FINDINGS: CD4+ T cell activation was negatively correlated with SARS-CoV-2 basal viral load in patients with severe COVID-19 (p = 0·043). The overall cellular immune response, as inferred by the IFN-γ signal, was higher at baseline for patients who progressed to mild disease compared to patients who progressed to severe disease (p = 0·0044). Subjects with milder disease developed higher T cell responses for MHC class I and II-restricted peptides (p = 0·033). INTERPRETATION: Mounting specific cellular immune responses in the first days after symptom onset, as inferred by IFN-γ magnitude in the ELISPOT assay, may efficiently favor a positive outcome. In contrast, progression to severe COVID-19 was accompanied by stronger cellular immune responses, higher CD4 + T cell activation, and a higher number of in silico predicted high-affinity class I HLA alleles.

Binding Evolution of the Dengue Virus Envelope Against DC-SIGN: A Combined Approach of Phylogenetics and Molecular Dynamics Analyses Over 30 Years of Dengue Virus in Brazil.

The Red Queen Hypothesis (RQH), derived from Lewis Carroll's "Through the Looking-Glass", postulates that organisms must continually adapt in response to each other to maintain relative fitness. Within the context of host-pathogen interactions, the RQH implies an evolutionary arms race, wherein viruses evolve to exploit hosts and hosts evolve to resist viral invasion. This study delves into the dynamics of the RQH in the context of virus-cell interactions, specifically focusing on virus receptors and cell receptors. We observed multiple virus-host systems and noted patterns of co-evolution. As viruses evolved receptor-binding proteins to effectively engage with cell receptors, cells countered by altering their receptor genes. This ongoing mutual adaptation cycle has influenced the molecular intricacies of receptor-ligand interactions. Our data supports the RQH as a driving force behind the diversification and specialization of both viral and host cell receptors. Understanding this co-evolutionary dance offers insights into the unpredictability of emerging viral diseases and potential therapeutic interventions. Future research is crucial to dissect the nuanced molecular changes and the broader ecological consequences of this ever-evolving battle. Here, we combine phylogenetic inferences, structural modeling, and molecular dynamics analyses to describe the epidemiological characteristics of major Brazilian DENV strains that circulated from 1990 to 2022 from a combined perspective, thus providing us with a more detailed picture on the dynamics of such interactions over time.

Genomoviruses in Liver Samples of Molossus molossus Bats.

CRESS-DNA encompasses a broad spectrum of viruses documented across diverse organisms such as animals, plants, diatoms, fungi, and marine invertebrates. Despite this prevalence, the full extent of these viruses' impact on the environment and their respective hosts remains incompletely understood. Furthermore, an increasing number of viruses within this category lack detailed characterization. This investigation focuses on unveiling and characterizing viruses affiliated with the Genomoviridae family identified in liver samples from the bat Molossus molossus. Leveraging viral metagenomics, we identified seven sequences (MmGmV-PA) featuring a circular DNA genome housing two ORFs encoding replication-associated protein (Rep) and capsid protein (Cap). Predictions based on conserved domains typical of the Genomoviridae family were established. Phylogenetic analysis revealed the segregation of these sequences into two clades aligning with the genera Gemycirculavirus (MmGmV-06-PA and MmGmV-07-PA) and Gemykibivirus (MmGmV-01-PA, MmGmV-02-PA, MmGmV-03-PA, MmGmV-05-PA, and MmGmV-09-PA). At the species level, pairwise comparisons based on complete nucleotide sequences indicated the potential existence of three novel species. In summary, our study significantly contributes to an enhanced understanding of the diversity of Genomoviridae within bat samples, shedding light on previously undiscovered viral entities and their potential ecological implications.

Metagenomic of Liver Tissue Identified at Least Two Genera of Totivirus-like Viruses in Molossus molossus Bats.

The Totiviridae family of viruses has a unique genome consisting of double-stranded RNA with two open reading frames that encode the capsid protein (Cap) and the RNA-dependent RNA polymerase (RdRpol). Most virions in this family are isometric in shape, approximately 40 nm in diameter, and lack an envelope. There are five genera within this family, including Totivirus, Victorivirus, Giardiavirus, Leishmaniavirus, and Trichomonasvirus. While Totivirus and Victorivirus primarily infect fungi, Giardiavirus, Leishmaniavirus, and Trichomonasvirus infect diverse hosts, including protists, insects, and vertebrates. Recently, new totivirus-like species have been discovered in fish and plant hosts, and through metagenomic analysis, a novel totivirus-like virus (named Tianjin totivirus) has been isolated from bat guano. Interestingly, Tianjin totivirus causes cytopathic effects in insect cells but cannot grow in mammalian cells, suggesting that it infects insects consumed by insectivorous bats. In this study, we used next-generation sequencing and identified totivirus-like viruses in liver tissue from Molossus molossus bats in the Amazon region of Brazil. Comparative phylogenetic analysis based on the RNA-dependent RNA polymerase region revealed that the viruses identified in Molossus bats belong to two distinct phylogenetic clades, possibly comprising different genera within the Totiviridae family. Notably, the mean similarity between the Tianjin totivirus and the totiviruses identified in Molossus bats is less than 18%. These findings suggest that the diversity of totiviruses in bats is more extensive than previously recognized and highlight the potential for bats to serve as reservoirs for novel toti-like viruses.

Genetic variability of Leishmania (Leishmania) infantum causing human visceral leishmaniasis in the Southeastern Brazil.

Leishmania infantum is a protozoan that causes visceral leishmaniasis (VL) in the Americas and some regions of Europe. The disease is mainly characterized by hepatosplenomegaly and fever, and can be fatal. Factors related to the host and parasite can contribute to the transmission of Leishmania and the clinical outcome. The intraspecific genetic variability of L. infantum strains may be one of these factors. In this study, we evaluated the genetic variability of L. infantum obtained from bone marrow smear slides from patients in the Sao Paulo State, Brazil. For this, the minicircle of the kDNA hypervariable region was used as target by Sanger sequencing. By analyzing the similarity of the nucleotides and the maximum likelihood tree (Fasttree), we observed a high similarity (98%) among samples. Moreover, we identified four different profiles of L. infantum. In conclusion, L. infantum strains from Sao Paulo State, Brazil, showed low diversity measured by minicircle of the kDNA hypervariable region.

Characterization of Torquetenovirus in amniotic fluid at the time of in utero fetal surgery: correlation with early premature delivery and respiratory distress.

Torquetenovirus (TTV) is a commensal virus present in many healthy individuals. Although considered to be non-pathogenic, its presence and titer have been shown to be indicative of altered immune status in individuals with chronic infections or following allogeneic transplantations. We evaluated if TTV was present in amniotic fluid (AF) at the time of in utero surgery to correct a fetal neurological defect, and whether its detection was predictive of adverse post-surgical parameters. AF was collected from 27 women by needle aspiration prior to a uterine incision. TTV titer in the AF was measured by isolation of viral DNA followed by gene amplification and analysis. The TTV genomes were further characterized and sequenced by metagenomics. Pregnancy outcome parameters were subsequently obtained by chart review. Three of the AFs (11.1%) were positive for TTV at 3.36, 4.16, and 4.19 log10 copies/mL. Analysis of their genomes revealed DNA sequences similar to previously identified TTV isolates. Mean gestational age at delivery was >2 weeks earlier (32.5 vs. 34.6 weeks) and the prevalence of respiratory distress was greater (100% vs. 20.8%) in the TTV-positive pregnancies. TTV detection in AF prior to intrauterine surgery may indicate elevated post-surgical risk for earlier delivery and newborn respiratory distress.

Detailed characterization of Redondovirus in saliva of SARS-CoV-2-infected individuals in Sao Paulo, Brazil.

BACKGROUND: Redondovirus (ReDoV) is a DNA virus present in the respiratory tract of many healthy individuals. Since SARS-CoV-2, the virus responsible for COVID-19, also primarily infects the same site, we evaluated whether ReDoV was present at increased frequency in patients with COVID-19 and influenced infection parameters. METHODS: Saliva samples were collected weekly from 59 individuals with COVID-19 and from 132 controls. ReDoV was detected by polymerase chain reaction and the genotypes were identified by metagenomics. Torque Teno Virus (TTV) in these samples were previously reported. RESULTS: ReDoV was detected in saliva more frequently from COVID-19 patients (72.9%) than from controls (50.0%) (p = 0.0015). There were no associations between ReDoV detection and either continuous or intermittent SARS-CoV-2 shedding, the duration of SARS-CoV-2 detection in saliva, patients' sex or if infection was by the B1 or Gamma strain. The two ReDoV strains, Brisavirus and Vientovirus, were present in equivalent frequencies in ReDoV-positive COVID-19 patients and controls. Phylogenetic analysis suggested that the two ReDoV strains in Brazil were similar to strains previously detected on other continents. CONCLUSION: ReDoV expression in saliva is increased in males and females in Brazil with mild COVID-19 but its presence does not appear to influence properties of the SARS-CoV-2 infection.

Characterization of Phietavirus Henu 2 in the virome of individuals with acute gastroenteritis.

There is a growing interest in phages as potential biotechnological tools in human health owing to the antibacterial activity of these viruses. In this study, we characterized a new member (named PhiV_005_BRA/2016) of the recently identified phage species Phietavirus Henu 2. PhiV_005_BRA/2016 was detected through metagenomic analysis of stool samples of individuals with acute gastroenteritis. PhiV_005_BRA/2016 contains double-stranded linear DNA (dsDNA), it has a genome of 43,513 base pairs (bp), with a high identity score (99%) with phage of the genus Phietavirus, species of Phietavirus Henu 2. Life style prediction indicated that PhiV_005_BRA/2016 is a lysogenic phage whose the main host is methicillin-resistant Staphylococcus aureus (MRSA). Indeed, we found PhiV_005_BRA/2016 partially integrated in the genome of distinct MRSA strains. Our findings highlights the importance of large-scale screening of bacteriophages to better understand the emergence of multi-drug resistant bacterial.

Novel Chaphamaparvovirus in Insectivorous Molossus molossus Bats, from the Brazilian Amazon Region.

Chaphamaparvovirus (CHPV) is a recently characterized genus of the Parvoviridae family whose members can infect different hosts, including bats, which constitute the second most diverse order of mammals and are described worldwide as important transmitters of zoonotic diseases. In this study, we identified a new CHPV in bat samples from the municipality of Santarém (Pará state, North Brazil). A total of 18 Molossus molossus bats were analyzed using viral metagenomics. In five animals, we identified CHPVs. These CHPV sequences presented the genome with a size ranging from 3797 to 4284 bp. Phylogenetic analysis-based nucleotide and amino acid sequences of the VP1 and NS1 regions showed that all CHPV sequences are monophyletic. They are also closely related to CHPV sequences previously identified in bats in southern and southeast Brazil. According to the International Committee on Taxonomy of Viruses (ICTV) classification criteria for this species (the CHPV NS1 gene region must have 85% identity to be classified in the same species), our sequences are likely a new specie within the genus Chaphamaparvovirus, since they have less than 80% identity with other CHPV described earlier in bats. We also make some phylogenetic considerations about the interaction between CHPV and their host. We suggest a high level of specificity of CPHV and its hosts. Thus, the findings contribute to improving information about the viral diversity of parvoviruses and show the importance of better investigating bats, considering that they harbor a variety of viruses that may favor zoonotic events.

Aedes aegypti Totivirus identified in mosquitoes in the Brazilian Amazon region.

The totiviridae family contains viruses with double-stranded RNA genomes of 4.6-7.0 kpb, which encode a capsid protein (CP) and RNA-dependent RNA polymerase (RdRp), and they are approximately 40 nm in diameter with icosahedral symmetry. Totiviruses were first isolated from mosquitoes collected in Shaanxi Province (China). Here, we report a new Aedes aegypti Totivirus (AaTV) identified in mosquitoes from the Amazon rainforest. Mosquitoes (Diptera: Culicidae) were collected from a forest reserve belonging to the Amazon forest in the city of Macapá, Amapá state, Northern Brazil. A viral sequence with a 5748 nucleotide length that was nearly identical to Aedes aegypti Totivirus (AaTV), here named Aedes aegypti Totivirus BR59AP, was detected. A detailed molecular analysis was performed and shows that AaTV-BR59AP is highly related to the AaTV strain from the Caribbean region. We emphasize the importance of the characterization of new viruses in mosquitoes to deepen our understanding of viral diversity in insects and their potential role in disease.

Pegivirus Detection in Cerebrospinal Fluid from Patients with Central Nervous System Infections of Unknown Etiology in Brazil by Viral Metagenomics.

Metagenomic next-generation sequencing (mNGS) methodology serves as an excellent supplement in cases where diagnosis is challenging to establish through conventional laboratory tests, and its usage is increasingly prevalent. Examining the causes of infectious diseases in the central nervous system (CNS) is vital for understanding their spread, managing outbreaks, and effective patient care. In a study conducted in the state of São Paulo, Brazil, cerebrospinal fluid (CSF) samples from 500 patients with CNS diseases of indeterminate etiology, collected between 2017 and 2021, were analyzed. Employing a mNGS approach, we obtained the complete coding sequence of Pegivirus hominis (HPgV) genotype 2 in a sample from a patient with encephalitis (named IAL-425/BRA/SP/2019); no other pathogen was detected. Subsequently, to determine the extent of this virus's presence, both polymerase chain reaction (PCR) and/or real-time PCR assays were utilized on the entire collection. The presence of the virus was identified in 4.0% of the samples analyzed. This research constitutes the first report of HPgV detection in CSF samples in South America. Analysis of the IAL-425 genome (9107 nt) revealed a 90% nucleotide identity with HPgV strains from various countries. Evolutionary analyses suggest that HPgV is both endemic and extensively distributed. The direct involvement of HPgV in CNS infections in these patients remains uncertain.

Genetic variability of Leishmania (Leishmania) infantum causing human visceral leishmaniasis in the Southeastern Brazil

ABSTRACT Leishmania infantum is a protozoan that causes visceral leishmaniasis (VL) in the Americas and some regions of Europe. The disease is mainly characterized by hepatosplenomegaly and fever, and can be fatal. Factors related to the host and parasite can contribute to the transmission of Leishmania and the clinical outcome. The intraspecific genetic variability of L. infantum strains may be one of these factors. In this study, we evaluated the genetic variability of L. infantum obtained from bone marrow smear slides from patients in the Sao Paulo State, Brazil. For this, the minicircle of the kDNA hypervariable region was used as target by Sanger sequencing. By analyzing the similarity of the nucleotides and the maximum likelihood tree (Fasttree), we observed a high similarity (98%) among samples. Moreover, we identified four different profiles of L. infantum. In conclusion, L. infantum strains from Sao Paulo State, Brazil, showed low diversity measured by minicircle of the kDNA hypervariable region.

Is the Intergenic Region of Aedes aegypti Totivirus a Recombination Hotspot?

The genus totivirus in the family Totiviridae contains double-stranded RNA viruses. Their genome has two open reading frames (ORFs) that encode capsid protein (CP) and RNA-dependent RNA polymerase (RdRp). The toti-like viruses recently identified in Anopheles sp. and Aedes aegypti mosquitoes (AaTV) share the same genome organization as other totiviruses. The AaTVs that have been described in distinct geographical regions are monophyletic. In this study, we show that AaTV sequences can be grouped into at least three phylogenetic clades (named A, B, and C). Clades A and B are composed of AaTV sequences from mosquitoes collected in the Caribbean region (Guadeloupe), and clade C contains sequences from the USA. These clades may represent AaTV lineages that are locally adapted to their host populations. We also identified three recombinant AaTV strains circulating in mosquitoes in Guadeloupe. Although these strains have different chimeric patterns, the position of the recombination breakpoint was identical in all strains. Interestingly, this breakpoint is located in a hairpin-like structure in the intergenic region of the AaTV genome. This RNA structure may stall RNA polymerase processivity and consequently induce template switching. In vitro studies should be conducted to further investigate the biological significance of AaTV's intergenic region as a recombination hotspot.

One-year surveillance of SARS-CoV-2 in wastewater from vulnerable urban communities in metropolitan São Paulo, Brazil.

The current COVID-19 pandemic has emphasized the vulnerability of communities living in the urban outskirts and informal settlements. The lack of reliable COVID-19 case data highlights the importance and application of wastewater-based epidemiology. This study aimed to monitor the COVID-19 trends in four vulnerable urban communities (slums and low-income neighborhoods) in metropolitan São Paulo by assessing the SARS-CoV-2 RNA viral load in wastewater. We analyzed 160 samples from May 2020 to June 2021 with weekly or fortnightly samplings. The samples were ultracentrifuged with glycine elution and quantified by N1/N2 SARS-CoV-2 RT-qPCR. The results of positivity were 100% (Paraisópolis, Heliópolis and Cidade Tiradentes) and 76.9% (Vila Brasilândia). The new case numbers of COVID-19, counted from the onset of symptoms, positively correlated with SARS-CoV-2 N1 viral loads from the two largest communities (p<0.001). SARS-CoV-2 infectivity was tested in Vero E6 cells after concentration with the two techniques, ultrafiltration (Centricon® Plus-70 10 kDa) and sucrose cushion ultracentrifugation, but none of the evaluated samples presented positive results. Next-generation sequencing (NGS) analysis from samples collected in March and August 2021 revealed the presence of the clade 20 J (lineage P.1) belonging to the most prevalent circulating variant in the country. Our results showed that wastewater surveillance data can be used as complementary indicators to monitor the dynamics and temporal trends of COVID-19. The infectivity test results strengthened the evidence of low risk of infection associated with SARS-CoV-2 in wastewater.

Detection of human feces pecovirus in newly diagnosed HIV patients in Brazil.

Circular single stranded DNA viruses (CRESS DNA) encoding a homologous replication-associated protein (REP) have been identified in most of eukaryotic groups. It is not clear yet the role in human diseases or details of the life cycle of these viruses. Recently, much interest has been raised in the evolutionary history of CRESS DNA owing to the increasing number of new sequences obtained by Next-Generation Sequencing (NGS) in distinct host species. In this study we describe two full-length CRESS DNA genomes obtained of two newly diagnosed HIV patients from São Paulo State, Brazil. The initial BLASTx search indicated that both sequences (named SP-FFB/2020 and SP-MJMS/2020) are highly similar (98%) to a previous CRESS DNA sequence detected in human fecal sample from Peru in 2016 and designated as pecovirus (Peruvian stool-associated circo-like virus). This study reported for the first time the Human feces pecovirus in the feces of two newly diagnosed HIV patients in Brazil. Our comparative analysis showed that although pecoviruses in South America share an identical genome structure they diverge and form distinct clades. Thus, we suggest the circulation of different species of pecoviruses in Latin America. Nevertheless, further studies must be done to examine the pathogenicity of this virus.

Chikungunya Virus Asian Lineage Infection in the Amazon Region Is Maintained by Asiatic and Caribbean-Introduced Variants.

The simultaneous transmission of two lineages of the chikungunya virus (CHIKV) was discovered after the pathogen's initial arrival in Brazil. In Oiapoque (Amapá state, north Brazil), the Asian lineage (CHIKV-Asian) was discovered, while in Bahia state, the East-Central-South-African lineage (CHIKV-ECSA) was discovered (northeast Brazil). Since then, the CHIKV-Asian lineage has been restricted to the Amazon region (mostly in the state of Amapá), whereas the ECSA lineage has expanded across the country. Despite the fact that the Asian lineage was already present in the Amazon region, the ECSA lineage brought from the northeast caused a large outbreak in the Amazonian state of Roraima (north Brazil) in 2017. Here, CHIKV spread in the Amazon region was studied by a Zika-Dengue-Chikungunya PCR assay in 824 serum samples collected between 2013 and 2016 from individuals with symptoms of viral infection in the Amapá state. We found 11 samples positive for CHIKV-Asian, and, from these samples, we were able to retrieve 10 full-length viral genomes. A comprehensive phylogenetic study revealed that nine CHIKV sequences came from a local transmission cluster related to Caribbean strains, whereas one sequence was related to sequences from the Philippines. These findings imply that CHIKV spread in different ways in Roraima and Amapá, despite the fact that both states had similar climatic circumstances and mosquito vector frequencies.

A New Circular Single-Stranded DNA Virus Related with Howler Monkey Associated Porprismacovirus 1 Detected in Children with Acute Gastroenteritis.

Putative replication-associated protein (REP) and capsid-like (CAP) proteins are encoded by circular single-stranded DNA viruses (CRESS DNA), which have been found in samples from most eukaryotic groups. However, the details of these viruses' life cycles and their significance in diseases have yet to be established. We presented and analyzed two full-length CRESS DNA genomes acquired from two children diagnosed with acute gastroenteritis (GI) in the northeast state of Tocantins, Brazil, using next-generation sequencing and a virus-like filtration approach. Both sequences (named SmaCV3BR08 and SmaCV3BR291) are closely similar to a prior CRESS DNA sequence discovered in the feces of a new world monkey (Alouatta caraya) from the United States in 2009 and termed Howler monkey-associated porprismacovirus 1 (Genbank ID: NC 026317). According to our comparative study, these porprismacovirus genomes deviate by 10% at the nucleotide level. For comparative reasons, the divergence between our sequences (SmaCV3BR08 and SmaCV3BR291) and a porprismacovirus recently identified in a human fecal sample from Peru is 37%. These data suggest that there is a great diversity of porprismacoviruses in South America, perhaps more than two species. In addition, the finding of closely related sequences of porprismacoviruses in humans and native monkeys highlights the zoonotic potential of these viruses.

Coxsackievirus A6 strains causing an outbreak of hand-foot-and-mouth disease in Northeastern Brazil in 2018.

Hand-foot-and-mouth disease (HFMD) is a highly contagious viral disease commonly associated to Enteroviruses (EV). During 2018, Brazil faced massive HFMD outbreaks spread across the country. This study aimed to characterize the EV responsible for the HFMD outbreak that occurred in Paraiba State, Brazilian Northeastern region, in 2018, followed by a phylogenetic analysis to detail information on its genetic diversity. A total of 49 serum samples (one from each patient) collected from children ≤ 15 years old, clinically diagnosed with HFMD were tested for EV using conventional RT-PCR and RT-qPCR. EV infection was confirmed in 71.4% (35/49) of samples. The mean and median ages were 1.83 years and one year old, respectively. Twenty-two EV-positive samples were successfully sequenced and classified as EV-A species; 13 samples were also identified with the CV-A6 genotype. The phylogenetic analysis (VP1 region) of three samples revealed that the detected CV-A6 strains belonged to sub-lineage D3. The CV-A6 strains detected here clustered with strains from South America, Europe and West Asia strains that were also involved in HFMD cases during the 2017-2018 seasons, in addition to the previously detected Brazilian CV-A6 strains from 2012 to 2017, suggesting a global co-circulation of a set of different CV-A6 strains introduced in the country at different times. The growing circulation of the emerging CV-A6 associated with HFMD, together with the detection of more severe cases worldwide, suggests the need for a more intense surveillance system of HFMD in Brazil. In addition, this investigation was performed exclusively on serum samples, and the analysis of whole blood samples should be considered and could have shown advantages when employed in the diagnosis of enteroviral HFMD outbreaks.

Coxsackievirus A6 strains causing an outbreak of hand-foot-and-mouth disease in Northeastern Brazil in 2018

ABSTRACT Hand-foot-and-mouth disease (HFMD) is a highly contagious viral disease commonly associated to Enteroviruses (EV). During 2018, Brazil faced massive HFMD outbreaks spread across the country. This study aimed to characterize the EV responsible for the HFMD outbreak that occurred in Paraiba State, Brazilian Northeastern region, in 2018, followed by a phylogenetic analysis to detail information on its genetic diversity. A total of 49 serum samples (one from each patient) collected from children ≤ 15 years old, clinically diagnosed with HFMD were tested for EV using conventional RT-PCR and RT-qPCR. EV infection was confirmed in 71.4% (35/49) of samples. The mean and median ages were 1.83 years and one year old, respectively. Twenty-two EV-positive samples were successfully sequenced and classified as EV-A species; 13 samples were also identified with the CV-A6 genotype. The phylogenetic analysis (VP1 region) of three samples revealed that the detected CV-A6 strains belonged to sub-lineage D3. The CV-A6 strains detected here clustered with strains from South America, Europe and West Asia strains that were also involved in HFMD cases during the 2017-2018 seasons, in addition to the previously detected Brazilian CV-A6 strains from 2012 to 2017, suggesting a global co-circulation of a set of different CV-A6 strains introduced in the country at different times. The growing circulation of the emerging CV-A6 associated with HFMD, together with the detection of more severe cases worldwide, suggests the need for a more intense surveillance system of HFMD in Brazil. In addition, this investigation was performed exclusively on serum samples, and the analysis of whole blood samples should be considered and could have shown advantages when employed in the diagnosis of enteroviral HFMD outbreaks.