Incursion of Highly Pathogenic Avian Influenza A(H5N1) Clade 2.3.4.4b Virus, Brazil, 2023.

We report 4 highly pathogenic avian influenza A(H5N1) clade 2.3.4.4.b viruses in samples collected during June 2023 from Royal terns and Cabot's terns in Brazil. Phylodynamic analysis revealed viral movement from Peru to Brazil, indicating a concerning spread of this clade along the Atlantic Americas migratory bird flyway.

Mortality in sea lions is associated with the introduction of the H5N1 clade 2.3.4.4b virus in Brazil October 2023: whole genome sequencing and phylogenetic analysis.

Clade 2.3.4.4b highly pathogenic avian influenza (HPAI) H5N1 virus was detected in the South American sea lions found dead in Santa Catarina, Brazil, in October 2023. Whole genome sequencing and comparative phylogenetic analysis were conducted to investigate the origin, genetic diversity, and zoonotic potentials of the H5N1 viruses. The H5N1 viruses belonged to the genotype B3.2 of clade 2.3.4.4b H5N1 virus, which was identified in North America and disseminated to South America. They have acquired new amino acid substitutions related to mammalian host affinity. Our study provides insights into the genetic landscape of HPAI H5N1 viruses in Brazil, highlighting the continuous evolutionary processes contributing to their possible adaptation to mammalian hosts.

SARS-CoV-2 Spillback to Wild Coatis in Sylvatic-Urban Hotspot, Brazil.

We tested coatis (Nasua nasua) living in an urban park near a densely populated area of Brazil and found natural SARS-CoV-2 Zeta variant infections by using quantitative reverse transcription PCR, genomic sequencing, and serologic surveillance. We recommend a One Health strategy to improve surveillance of and response to COVID-19.

Gas6 drives Zika virus-induced neurological complications in humans and congenital syndrome in immunocompetent mice.

Zika virus (ZIKV) has the ability to cross placental and brain barriers, causing congenital malformations in neonates and neurological disorders in adults. However, the pathogenic mechanisms of ZIKV-induced neurological complications in adults and congenital malformations are still not fully understood. Gas6 is a soluble TAM receptor ligand able to promote flavivirus internalization and downregulation of immune responses. Here we demonstrate that there is a correlation between ZIKV neurological complications with higher Gas6 levels and the downregulation of genes associated with anti-viral response, as type I IFN due to Socs1 upregulation. Also, Gas6 gamma-carboxylation is essential for ZIKV invasion and replication in monocytes, the main source of this protein, which was inhibited by warfarin. Conversely, Gas6 facilitates ZIKV replication in adult immunocompetent mice and enabled susceptibility to transplacental infection. Our data indicate that ZIKV promotes the upregulation of its ligand Gas6, which contributes to viral infectivity and drives the development of severe adverse outcomes during ZIKV infection.

Bovine respiratory syncytial virus seroprevalence and risk factors in non-vaccinated dairy cattle herds in Brazil.

BACKGROUND: The cattle industry is one of the most important Brazilian agribusiness sectors and is a strong contributor to the national economy. Annually about 44.6 million calves are bred, which makes the optimal management of these animals extremely important. Several diseases can affect the initial stages of the bovine production chain, being the bovine respiratory syncytial virus (BRSV) one of the most relevant pathogens. This study aimed to characterize the epidemiology of BRSV infection in dairy cattle herds of São Paulo State, Brazil, using serological and risk factors analyses. For that, 1243 blood samples were collected of animals from 26 farms and a questionnaire about possible risk factors for BRSV prevalence was performed. The obtained blood sera were analyzed using virus neutralization test (VNT). RESULTS: VNT results showed high BRSV prevalence in dairy cattle herds, reaching 79.5% of seropositivity. The BRSV seroprevalence among studied farms ranged from 40 to 100%. The analysis of risk factors indicated that the age group and the occurrence of coinfection with bovine herpesvirus 1 (BoHV-1) and bovine viral diarrhea virus 1 (BVDV-1) should be associated with a higher prevalence of BRSV, while natural suckling was considered a protective factor. CONCLUSIONS: The study showed that adult animals over 1 year old are an important risk factor for the high seroprevalence of BRSV in herds. The high BRSV prevalence associated with BoHV-1 and BVDV-1 suggests that biosecurity measures should be applied in order to reduce viral dissemination. Additionally, the natural suckling may be an important management to protect calves from high BRSV seroprevalence.

Specific Biomarkers Associated With Neurological Complications and Congenital Central Nervous System Abnormalities From Zika Virus-Infected Patients in Brazil.

Background: Zika virus (ZIKV) infections have been linked to different levels of clinical outcomes, ranging from mild rash and fever to severe neurological complications and congenital malformations. Methods: We investigated the clinical and immunological response, focusing on the immune mediators profile in 95 acute ZIKV-infected adult patients from Campinas, Brazil. These patients included 6 pregnant women who later delivered during the course of this study. Clinical observations were recorded during hospitalization. Levels of 45 immune mediators were quantified using multiplex microbead-based immunoassays. Results: Whereas 11.6% of patients had neurological complications, 88.4% displayed mild disease of rash and fever. Several immune mediators were specifically higher in ZIKV-infected patients, and levels of interleukin 10, interferon gamma-induced protein 10 (IP-10), and hepatocyte growth factor differentiated between patients with or without neurological complications. Interestingly, higher levels of interleukin 22, monocyte chemoattractant protein 1, TNF-α, and IP-10 were observed in ZIKV-infected pregnant women carrying fetuses with fetal growth-associated malformations. Notably, infants with congenital central nervous system deformities had significantly higher levels of interleukin 18 and IP-10 but lower levels of hepatocyte growth factor than those without such abnormalities born to ZIKV-infected mothers. Conclusions: This study identified several key markers for the control of ZIKV pathogenesis. This will allow a better understanding of the molecular mechanisms of ZIKV infection in patients.

Whole genome sequencing of low pathogenicity avian influenza virus (H6N2) detected from a Brazilian teal (Amazonnetta brasiliensis) in Brazil, 2023.

The whole genome sequence of a low pathogenicity avian influenza virus (H6N2) was sequenced from a Brazilian teal (Amazonetta brasiliensis) in Brazil, 2023. Phylogenetic analysis of the whole genome revealed a distinct genome pertaining to South American LPAIV from 2014 to 2016, indicating extensive circulation among South American wild birds.

No molecular evidence for influenza A virus and coronavirus in bats belonging to the families Phyllostomidae, Vespertilionidae, and Molossidae in the state of São Paulo, Brazil.

This study aimed to evaluate, by molecular methods, the presence of influenza A virus (IAV) and coronavirus in non-hematophagous bats collected in the state of São Paulo, Brazil. Samples of lung tissue and small intestine from 105 bats belonging to three families (Phyllostomidae, Vespertilionidae, and Molossidae) were collected in 22 municipalities in the state of São Paulo. Genetic identification of bats species was performed by amplification and sequencing of a fragment of 710 bp of the mitochondrial COI gene. In the detection of IAV, genomes were performed by RT-PCR, aiming at the amplification of a 245-bp fragment of the IAV matrix (M) protein gene. For coronaviruses, two fragments of 602 and 440 bp corresponding to segments along the gene encoding the RNA-dependent RNA polymerase (RdRp) were targeted. The detection limit for each of the PCRs was also determined. All samples analyzed here were negative for both viruses, and the lower limit of detection of the PCRs for the amplification of influenza virus A and coronavirus was estimated at 3.5 × 103 and 4.59 genomic copies per microliter, respectively. Although bats have been shown to harbor a large number of pathogens, the results of the present study support the theory that virus circulation in bats in the wild often occurs at low viral loads and that our understanding of the complex infectious dynamics of these viruses in wild conditions is still limited.

Genetic diversity of adenovirus in neotropical bats from Brazil.

Bats can harbor a diversity of viruses, such as adenovirus. Ten different species of bat adenoviruses (BtAdV A to J) have been previous described worlwide. In Brazil, BtAdV was described in three species of phyllostomid species: Artibeus lituratus, Desmodus rotundus, and Sturnira lilium. There are around 180 bat species in Brazil, with 67% inhabiting the Atlantic Forest, with few information about the circulation of BtAdV in this biome. We aimed to describe the molecular detection and the phylogenetic characterization and suggest a classification of BtAdVs circulating in bats from the Brazilian Atlantic Forest. We collected 382 oral and rectal swabs from 208 bats between 2014-2015 and 2020-2021 from São Paulo, Pernambuco, and Santa Catarina Brazilian states. The adenovirus detection was done by a nested PCR targeting the DNA polymerase gene, and all positive samples were sequenced by the Sanger method. The phylogenetic analyses were based on the amino acid sequences using the MEGA 7 and BEAST software. We obtained 16 positive animals (detection rate 7.7%) belonging to seven bat species: Artibeus lituratus, Carollia perspicillata, Sturnira lilium, Molossus molossus, and the first record of Phyllostomus discolor, Eptesicus diminutus, and Myotis riparius. The phylogenetic analysis based on partial amino acid sequences showed that all obtained AdV sequences belong to the Mastadenovirus genus. We observed a high genetic diversity of BtAdV and identified eleven potential BtAdV species circulating in Brazil (BtAdV K to U). Our results contribute to the epidemiological surveillance of adenovirus, increasing the knowledge about the viral diversity and the distribution of AdV in bats from the Atlantic Forest.

Partial VP1 sequencing of Brazilian infectious bursal disease virus strains.

Infectious bursal disease virus (IBDV) is classified according to the antigenicity and virulence into classical virulent (cv), very virulent (vv), and antigenic variant strains. The molecular basis for the IBDV antigenic variation is well established and is associated to the capsid protein, VP2 (gene VP2 of segment A), whereas both VP2 and the RNA-dependent RNA polymerase, VP1 (gene VP1 of segment B), have been correlated with the virulence. In this study, seventeen Brazilian IBDV samples previously characterized by the VP2 gene as cv (three) and vv (fourteen) strains were genetically and molecularly analyzed for their VP1 gene. All of the strains kept with the same cv or vv classification except one sample, Br/03/DR. This sample was classified as vv by its VP2 gene, but it was most closely related to the cv strains by its VP1 partial sequence and phylogeny. Studies on the phylogeny of VP1 have suggested a possible reassortment event that originated the vvVP1. In this case, the sample carrying vvVP2 and cvVP1 could be a descendant of IBDV ancestors prior to the reassortment of vvVP1; alternatively, it could be the result of a genetic exchange between the segments of different strains or with a live attenuated vaccine. Nevertheless, this is the first report of natural genetic reassortment of IBDV in Brazil.

Deep phylogenetic-based clustering analysis uncovers new and shared mutations in SARS-CoV-2 variants as a result of directional and convergent evolution.

Nearly two decades after the last epidemic caused by a severe acute respiratory syndrome coronavirus (SARS-CoV), newly emerged SARS-CoV-2 quickly spread in 2020 and precipitated an ongoing global public health crisis. Both the continuous accumulation of point mutations, owed to the naturally imposed genomic plasticity of SARS-CoV-2 evolutionary processes, as well as viral spread over time, allow this RNA virus to gain new genetic identities, spawn novel variants and enhance its potential for immune evasion. Here, through an in-depth phylogenetic clustering analysis of upwards of 200,000 whole-genome sequences, we reveal the presence of previously unreported and hitherto unidentified mutations and recombination breakpoints in Variants of Concern (VOC) and Variants of Interest (VOI) from Brazil, India (Beta, Eta and Kappa) and the USA (Beta, Eta and Lambda). Additionally, we identify sites with shared mutations under directional evolution in the SARS-CoV-2 Spike-encoding protein of VOC and VOI, tracing a heretofore-undescribed correlation with viral spread in South America, India and the USA. Our evidence-based analysis provides well-supported evidence of similar pathways of evolution for such mutations in all SARS-CoV-2 variants and sub-lineages. This raises two pivotal points: (i) the co-circulation of variants and sub-lineages in close evolutionary environments, which sheds light onto their trajectories into convergent and directional evolution, and (ii) a linear perspective into the prospective vaccine efficacy against different SARS-CoV-2 strains.

Prompt Inactivation of Coronavirus Using a 280 nm Light-Emitting Diode Cluster Device.

Objective: We report on the development and characterization of a UV-C light-emitting diode (LED) 280 nm cluster prototype device designed for the rapid disinfection of SARS-CoV-2 coronaviruses. The device was evaluated against the Betacoronavirus mouse hepatitis virus-3 strain, and its virucidal capacity was probed as a function of different applied UV-C doses versus different situations concerning irradiation distances. Background: UV-C LEDs are light emitters that offer advantages over low-pressure mercury lamps, such as quasimonochromaticity, lower electrical power consumption, instant on/off with the instant full-power operation, unlimited on/off cycles for disinfection schemes, and a much longer lifetime operation, in addition to portability aspects, as well as UV-C LEDs do not contain heavy metal in its composition such as mercury, found in ultraviolet germicidal irradiation (UVGI) lamps. Results: This novel device reached a 99.999% elimination rate at a distance of 9 cm at all the tested irradiation times (dose dependence), demonstrating that it took only 30 sec to achieve this inactivation rate. Its virucidal effectivity in rapid virus inactivation was demonstrated. Conclusions: We conclude that the HHUVCS cluster device (λp = 280 nm) provides a rapid virucidal effect against the SARS-CoV-2 coronavirus. The current research should encourage further advances in UV-C LED-based devices designed for the inactivation of SARS-CoV-2 virus on surfaces, in air, and in liquids.

Fast inactivation of coronavirus in filtering-facepiece respirators in a reflective cylindrical UV-C chamber.

Objective: We report on the development and characterization of a UV-C (λ  =  200 - 280 nm, λpeak = 254 nm) chamber designed for the rapid disinfection of N95 class filtering-facepiece respirators contaminated with SARS-CoV-2 coronaviruses. The device was evaluated against Betacoronavirus strain MHV-3 and its virucidal capacity was evaluated as a function of different applied UV-C doses (UV-C exposure times of 60 s, 120 s, 180 s, and 240 s) using two types of respirators geometry (shell and two-panel shapes, 3M 8801 H and 9920 H, respectively), at eight points of the respirators. Background: Most chemical disinfection methods are not recommended for N95 masks. UV-C light provided by UVGI lamps (254 nm) is an effective physical agent against viruses and bacteria due to direct photochemical harming effect on DNA/RNA, and can provide rapid disinfection for personal protective equipment such as N95/PFF2 masks. Results: The device reached a mean elimination rate of 99.9999% of MHV-3 inoculated into all the assessed different points on the tested PFF2 respirator models in a UV-C cycle of just 60 s. Statistical analysis performed through Person´s chi-square test showed no correlation between the viral infectivity reduction and the viral inoculation point (p = 0.512) and the tested respirator models (p = 0.556). However, a correlation was found between the exposure time and the viral infectivity reduction (p = 0.000*), between UV-C and no UV-C exposure. All the tested UV-C exposure times (60 s, 120 s, 180 s, and 240 s) provided the same reduction in infection rates. Therefore, 60 s was confirmed as the minimum exposure time to achieve a 99.9999% or 6 Log reduction in MHV-3 coronavirus infection rates in the PFF2 samples tested in the device. Conclusions: We conclude that the assessed UV-C chamber for the inactivation of MHV-3 coronavirus in N95/PFF2 standard masks can be a promising tool for effective and rapid disinfection of coronaviruses, including SARS-CoV-2 virus.

Rhinoviruses as critical agents in severe bronchiolitis in infants.

OBJECTIVES: To detect RSV or other thirteen respiratory viruses as possible causer agent of bronchiolitis in infants. METHOD: This is an epidemiological analytical study, conducted using a nasopharyngeal aspirate of 173 hospitalized children younger than two years old with severe bronchiolitis in three hospitals in the Campinas Metropolitan Region (CMR) during 2013-14. The data was statically evaluated by Pearson's chi-squared test with statistical significance of 0.05 and 95% confidence level. RESULTS: As expected, the most prevalent viruses detected were RSV A and B in 47% and 16% of the samples, respectively. However, almost a third of severe bronchiolitis cases there were no detection of RSV, and the viruses more commonly detected were rhinoviruses, which were identified in almost a quarter of all positive samples for at least a viral agent. CONCLUSIONS: Although nothing could be concluded from the disease severity and clinical-epidemiological data, the present study's results indicate that severe bronchiolitis is not always related to RSV infections in children younger than two years old, and the rhinoviruses were more prevalent in these cases. These findings reinforce the need to carry out a viral diagnosis in the hospital emergency would be very appropriate for all cases of respiratory infections in children, even for diseases in which the primary etiological agent seems to be well known.

Genomic Epidemiology of SARS-CoV-2 in Tocantins State and the Diffusion of P.1.7 and AY.99.2 Lineages in Brazil.

Tocantins is a state in the cross-section between the Central-West, North and Northeast regions of Brazilian territory; it is a gathering point for travelers and transportation from the whole country. In this study, 9493 genome sequences, including 241 local SARS-CoV-2 samples (collected from 21 December 2020, to 16 December 2021, and sequenced in the MinION platform) were analyzed with the following aims: (i) identify the relative prevalence of SARS-CoV-2 lineages in the state of Tocantins; (ii) analyze them phylogenetically against global SARS-CoV-2 sequences; and (iii) hypothesize the viral dispersal routes of the two most abundant lineages found in our study using phylogenetic and phylogeographic approaches. The performed analysis demonstrated that the majority of the strains sequenced during the period belong to the Gamma P.1.7 (32.4%) lineage, followed by Delta AY.99.2 (27.8%), with the first detection of VOC Omicron. As expected, there was mainly a dispersion of P.1.7 from the state of São Paulo to Tocantins, with evidence of secondary spreads from Tocantins to Goiás, Mato Grosso, Amapá, and Pará. Rio de Janeiro was found to be the source of AY.99.2 and from then, multiple cluster transmission was observed across Brazilian states, especially São Paulo, Paraiba, Federal District, and Tocantins. These data show the importance of trade routes as pathways for the transportation of the virus from Southeast to Northern Brazil.

Virucidal PVP-Copper Salt Composites against Coronavirus Produced by Electrospinning and Electrospraying.

Electrospinning technology was used to produced polyvinylpyrrolidone (PVP)-copper salt composites with structural differences, and their virucidal activity against coronavirus was investigated. The solutions were prepared with 20, 13.3, 10, and 6.6% w/v PVP containing 3, 1.0, 0.6, and 0.2% w/v Cu (II), respectively. The rheological properties and electrical conductivity contributing to the formation of the morphologies of the composite materials were observed by scanning electron microscopy (SEM). SEM images revealed the formation of electrospun PVP-copper salt ultrafine composite fibers (0.80 ± 0.35 µm) and electrosprayed PVP-copper salt composite microparticles (1.50 ± 0.70 µm). Energy-dispersive X-ray spectroscopy (EDS) evidenced the incorporation of copper into the produced composite materials. IR spectra confirmed the chemical composition and showed an interaction of Cu (II) ions with oxygen in the PVP resonant ring. Virucidal composite fibers inactivated 99.999% of coronavirus within 5 min of contact time, with moderate cytotoxicity to L929 cells, whereas the virucidal composite microparticles presented with a virucidal efficiency of 99.999% within 1440 min of exposure, with low cytotoxicity to L929 cells (mouse fibroblast). This produced virucidal composite materials have the potential to be applied in respirators, personal protective equipment, self-cleaning surfaces, and to fabric coat personal protective equipment against SARS-CoV-2, viral outbreaks, or pandemics.

High genetic diversity of alphacoronaviruses in bat species (Mammalia: Chiroptera) from the Atlantic Forest in Brazil.

Bat coronaviruses (Bat-CoVs) represent around 35% of all virus genomes described in bats. Brazil has one of the highest mammal species diversity, with 181 species of bats described so far. However, few Bat-CoV surveillance programmes were carried out in the country. Thus, our aim was to jevaluate the Bat-CoV diversity in the Atlantic Forest, the second biome with the highest number of bat species in Brazil. We analysed 456 oral and rectal swabs and 22 tissue samples from Atlantic Forest bats, detecting Alphacoronavirus in 44 swab samples (9.6%) targeting the RdRp gene from seven different bat species, three of which have never been described as Bat-CoV hosts. Phylogenetic analysis of the amino acid (aa) sequences coding the RdRp gene grouped the sequences obtained in our study with Bat-CoV previously detected in identical or congeneric bat species, belonging to four subgenera, with high aa identity (over 90%). The RdRp gene was also detected in three tissue samples from Diphylla ecaudata and Sturnira lilium, and the partial S gene was successfully sequenced in five tissues and swab samples of D. ecaudata. The phylogenetic analysis based on the partial S gene obtained here grouped the sequence of D. ecaudata with CoV from Desmodus rotundus previously detected in Peru and Brazil, belonging to the Amalacovirus subgenus, with aa identity ranging from 73.6% to 88.8%. Our data reinforce the wide distribution of Coronaviruses in bats from Brazil and the novelty of three bats species as Bat-CoV hosts and the co-circulation of four Alphacoronavirus subgenera in Brazil.

Detection of and phylogenetic studies with avian metapneumovirus recovered from feral pigeons and wild birds in Brazil.

The aim of the present study was to determine whether avian metapneumovirus (aMPV)-related viruses were present in wild and synanthropic birds in Brazil. Therefore, we analysed samples from wild birds, feral pigeons and domestic chickens in order to perform a phylogenetic comparison. To detect the presence of aMPV, a nested reverse transcriptase-polymerase chain reaction was performed with the aim of amplifying a fragment of 270 bases for subtype A and 330 bases for subtype B, comprising the gene coding the G glycoprotein. Positive samples for aMPV subtypes A and B were found in seven (13.2%) different asymptomatic wild birds and pigeons (50%) that had been received at the Bosque dos Jequitibás Zoo Triage Center, Brazil. Also analysed were positive samples from 15 (12.9%) domestic chickens with swollen head syndrome from several regions of Brazil. The positive samples from wild birds, pigeons and domestic chickens clustered in two major phylogenetic groups: some with aMPV subtype A and others with subtype B. The similarity of the G fragment nucleotide sequence of aMPV isolated from chickens and synanthropic and wild avian species ranged from 100 to 97.5% (from 100 to 92.5% for the amino acids). Some positive aMPV samples, which were obtained from wild birds classified in the Orders Psittaciformes, Anseriformes and Craciformes, clustered with subtype A, and others from the Anas and Dendrocygma genera (Anseriformes Order) with subtype B. The understanding of the epizootiology of aMPV is very important, especially if this involves the participation of non-domestic bird species, which would add complexity to their control on farms and to implementation of vaccination programmes for aMPV.

Coding-Complete Genome Sequence of Murine Hepatitis Virus Strain 3 from Brazil.

Murine hepatitis virus (MHV) strain 3, one of the most important inducers of viral hepatitis, has been extensively studied as an organism to gain a better understanding of coronavirus biology and pathogenesis. Only one sequence is currently available. Another representative isolate has now been sequenced and added to the arsenal of MHV-3 variants.

Ultraviolet germicidal irradiation is effective against SARS-CoV-2 in contaminated makeup powder and lipstick.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is mainly transmitted by airborne droplets generated by infected individuals. Since this and many other pathogens are able to remain viable on inert surfaces for extended periods of time, contaminated surfaces play an important role in SARS-CoV-2 fomite transmission. Cosmetic products are destined to be applied on infection-sensitive sites, such as the lips and eyelids. Therefore, special biosafety precautions should be incorporated into the routine procedures of beauty parlors and shops. Indeed, innovative cosmetics companies are currently searching for disinfection protocols that ensure the customers' safety in makeup testing. Here, we propose an ultraviolet germicidal irradiation (UVGI) strategy that can be used to reduce the odds of COVID-19 fomite transmission by makeup testers. It is well-known that UVGI effectively inactivates pathogens on flat surfaces and clear fluids. However, ultraviolet-C (UVC) radiation at 254 nm penetrates poorly in turbid and porous materials, such as makeup and lipstick formulations. Thus, we investigated the virucidal effect of UVGI against SARS-CoV-2 deposited on such substrates and compared their performance to that of flat polystyrene surfaces, used as controls. Concentrated infectious SARS-CoV-2 inoculum (106 PFU/mL) deposited on lipstick and makeup powder was completely inactivated (>5log10 reduction) following UVC exposures at 1,260 mJ/cm2, while flat plastic surfaces required 10 times less exposure (126 mJ/cm2) to reach the same microbicidal performance. We conclude that UVGI comprises an effective disinfection strategy to promote biosafety for cosmetics testers. However, appropriate UVC dosimetry must be implemented to overcome inefficiencies caused by the optical properties of turbid materials in lipsticks and makeup powders.