A hepatitis B virus causes chronic infections in equids worldwide.

Preclinical testing of novel therapeutics for chronic hepatitis B (CHB) requires suitable animal models. Equids host homologs of hepatitis C virus (HCV). Because coinfections of hepatitis B virus (HBV) and HCV occur in humans, we screened 2,917 specimens from equids from five continents for HBV. We discovered a distinct HBV species (Equid HBV, EqHBV) in 3.2% of donkeys and zebras by PCR and antibodies against EqHBV in 5.4% of donkeys and zebras. Molecular, histopathological, and biochemical analyses revealed that infection patterns of EqHBV resembled those of HBV in humans, including hepatotropism, moderate liver damage, evolutionary stasis, and potential horizontal virus transmission. Naturally infected donkeys showed chronic infections resembling CHB with high viral loads of up to 2.6 × 109 mean copies per milliliter serum for >6 mo and weak antibody responses. Antibodies against Equid HCV were codetected in 26.5% of donkeys seropositive for EqHBV, corroborating susceptibility to both hepatitis viruses. Deltavirus pseudotypes carrying EqHBV surface proteins were unable to infect human cells via the HBV receptor NTCP (Na+/taurocholate cotransporting polypeptide), suggesting alternative viral entry mechanisms. Both HBV and EqHBV deltavirus pseudotypes infected primary horse hepatocytes in vitro, supporting a broad host range for EqHBV among equids and suggesting that horses might be suitable for EqHBV and HBV infections in vivo. Evolutionary analyses suggested that EqHBV originated in Africa several thousand years ago, commensurate with the domestication of donkeys. In sum, EqHBV naturally infects diverse equids and mimics HBV infection patterns. Equids provide a unique opportunity for preclinical testing of novel therapeutics for CHB and to investigate HBV/HCV interplay upon coinfection.

Development of a New Multispecies Protein A-Based ELISA for Serodiagnosis of Hepatitis E Infection with Validation in Domestic Swine.

Hepatitis E virus (HEV) infects roughly 20 million people worldwide, causing self-limiting acute hepatic disease that can evolve into a chronic course. HEV-3, HEV-4, and HEV-7 genotypes are zoonotic and transmitted to humans by consuming raw or undercooked meat. Here, we developed an indirect ELISA based on the recombinant HEV-3 capsid and performed a seroprevalence study on domestic swine in northeastern Brazil. Our in-house ELISA was initially validated using a subset of 79 sera characterized by concordant results for two distinct commercial ELISA kits. Our ELISA exhibited excellent sensitivity (94%) and specificity (100%), with an area under the curve of 0.99 Further testing, including 212 swine sera, revealed a seroprevalence of 57.5% (95% confidence interval, 50.6-64.3%). Our findings indicate that the novel ELISA test could accurately detect specific anti-HEV antibodies in domestic pigs and should be further validated in humans and other mammals.

Divergent Genotype of Hepatitis A Virus in Alpacas, Bolivia, 2019.

Hepatitis A virus (HAV) is a common human pathogen found exclusively in primates. In a molecular and serologic study of 64 alpacas in Bolivia, we detected RNA of distinct HAV in ≈9% of animals and HAV antibodies in ≈64%. Complete-genome analysis suggests a long association of HAV with alpacas.

Mammalian deltavirus without hepadnavirus coinfection in the neotropical rodent Proechimys semispinosus.

Hepatitis delta virus (HDV) is a human hepatitis-causing RNA virus, unrelated to any other taxonomic group of RNA viruses. Its occurrence as a satellite virus of hepatitis B virus (HBV) is a singular case in animal virology for which no consensus evolutionary explanation exists. Here we present a mammalian deltavirus that does not occur in humans, identified in the neotropical rodent species Proechimys semispinosus The rodent deltavirus is highly distinct, showing a common ancestor with a recently described deltavirus in snakes. Reverse genetics based on a tandem minus-strand complementary DNA genome copy under the control of a cytomegalovirus (CMV) promoter confirms autonomous genome replication in transfected cells, with initiation of replication from the upstream genome copy. In contrast to HDV, a large delta antigen is not expressed and the farnesylation motif critical for HBV interaction is absent from a genome region that might correspond to a hypothetical rodent large delta antigen. Correspondingly, there is no evidence for coinfection with an HBV-related hepadnavirus based on virus detection and serology in any deltavirus-positive animal. No other coinfecting viruses were detected by RNA sequencing studies of 120 wild-caught animals that could serve as a potential helper virus. The presence of virus in blood and pronounced detection in reproductively active males suggest horizontal transmission linked to competitive behavior. Our study establishes a nonhuman, mammalian deltavirus that occurs as a horizontally transmitted infection, is potentially cleared by immune response, is not focused in the liver, and possibly does not require helper virus coinfection.

Effectiveness of Naturally Acquired and Vaccine-Induced Immune Responses to SARS-CoV-2 Mu Variant.

SARS-CoV-2 Mu variant emerged in Colombia in 2021 and spread globally. In 49 serum samples from vaccinees and COVID-19 survivors in Colombia, neutralization was significantly lower (p<0.0001) for Mu than a parental strain and variants of concern. Only the Omicron variant of concern demonstrated higher immune evasion.

Highly Diverse Arenaviruses in Neotropical Bats, Brazil.

We detected arenavirus RNA in 1.6% of 1,047 bats in Brazil that were sampled during 2007-2011. We identified Tacaribe virus in 2 Artibeus sp. bats and a new arenavirus species in Carollia perspicillata bats that we named Tietê mammarenavirus. Our results suggest that bats are an underrecognized arenavirus reservoir.

Emergence of SARS-CoV-2 Delta Variant, Benin, May-July 2021.

Severe acute respiratory syndrome coronavirus 2 Delta variant epidemiology in Africa is unknown. We found Delta variant was introduced in Benin during April-May 2021 and became predominant within 2 months, after which a steep increase in reported coronavirus disease incidence occurred. Benin might require increased nonpharmaceutical interventions and vaccination coverage.

ICTV Virus Taxonomy Profile: Hepeviridae 2022.

The family Hepeviridae includes enterically transmitted small quasi-enveloped or non-enveloped positive-sense single-stranded RNA viruses infecting mammals and birds (subfamily Orthohepevirinae) or fish (Parahepevirinae). Hepatitis E virus (genus Paslahepevirus) is responsible for self-limiting acute hepatitis in humans; the infection may become chronic in immunocompromised individuals and extrahepatic manifestations have been described. Avian hepatitis E virus (genus Avihepevirus) causes hepatitis-splenomegaly syndrome in chickens. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Hepeviridae, which is available at www.ictv.global/report/hepeviridae.

Evidence against Zika virus infection of pets and peri-domestic animals in Latin America and Africa.

Decades after its discovery in East Africa, Zika virus (ZIKV) emerged in Brazil in 2013 and infected millions of people during intense urban transmission. Whether vertebrates other than humans are involved in ZIKV transmission cycles remained unclear. Here, we investigate the role of different animals as ZIKV reservoirs by testing 1723 sera of pets, peri-domestic animals and African non-human primates (NHP) sampled during 2013-2018 in Brazil and 2006-2016 in Côte d'Ivoire. Exhaustive neutralization testing substantiated co-circulation of multiple flaviviruses and failed to confirm ZIKV infection in pets or peri-domestic animals in Côte d'Ivoire (n=259) and Brazil (n=1416). In contrast, ZIKV seroprevalence was 22.2% (2/9, 95% CI, 2.8-60.1) in West African chimpanzees (Pan troglodytes verus) and 11.1% (1/9, 95% CI, 0.3-48.3) in king colobus (Colobus polycomos). Our results indicate that while NHP may represent ZIKV reservoirs in Africa, pets or peri-domestic animals likely do not play a role in ZIKV transmission cycles.

Sensitivity of Detection and Variant Typing of SARS-CoV-2 in European Laboratories.

The molecular detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is key for clinical management and surveillance. Funded by the European Centre for Disease Prevention and Control, we conducted an external quality assessment (EQA) on the molecular detection and variant typing of SARS-CoV-2 that included 59 European laboratories in 34 countries. The EQA panel consisted of 12 lyophilized inactivated samples, 10 of which were SARS-CoV-2 variants (Alpha, Beta, Gamma, Delta, Epsilon, Eta, parental B.1 strain) ranging from 2.5 to 290.0 copies/µL or pooled respiratory viruses (adenovirus, enterovirus, influenza virus A, respiratory syncytial virus, or human coronaviruses 229E and OC43). Of all participants, 72.9% identified the presence of SARS-CoV-2 RNA correctly. In samples containing 25.0 or more genome copies/µL, SARS-CoV-2 was detected by 98.3% of the participating laboratories. Laboratories applying commercial tests scored significantly better (P < 0.0001, Kruskal-Wallis test) than those using in-house assays. Both the molecular detection and the typing of the SARS-CoV-2 variants were associated with the RNA concentrations (P < 0.0001, Kruskal-Wallis test). On average, only 5 out of the 10 samples containing different SARS-CoV-2 variants at different concentrations were correctly typed. The identification of SARS-CoV-2 variants was significantly more successful among EQA participants who combined real-time reverse transcription polymerase chain reaction (RT-PCR)-based assays for mutation detection and high-throughput genomic sequencing than among those who used a single methodological approach (P = 0.0345, Kruskal-Wallis test). Our data highlight the high sensitivity of SARS-CoV-2 detection in expert laboratories as well as the importance of continuous assay development and the benefits of combining different methodologies for accurate SARS-CoV-2 variant typing.

Genomic Characterization and Antimicrobial Susceptibility of Dromedary-Associated Staphylococcaceae from the Horn of Africa.

Members of the Staphylococcaceae family, particularly those of the genus Staphylococcus, encompass important human and animal pathogens. We collected and characterized Staphylococcaceae strains from apparently healthy and diseased camels (n = 84) and cattle (n = 7) in Somalia and Kenya. We phenotypically characterized the strains, including their antimicrobial inhibitory concentrations. Then, we sequenced their genomes using long-read sequencing, closed their genomes, and subsequently compared and mapped their virulence- and resistance-associated gene pools. Genome-based phylogenetics revealed 13 known Staphylococcaceae and at least two novel species. East African strains of different species encompassed novel sequence types and phylogenetically distant clades. About one-third of the strains had non-wild-type MICs. They were resistant to at least one of the following antimicrobials: tetracycline, benzylpenicillin, oxacillin, erythromycin, clindamycin, trimethoprim, gentamicin, or streptomycin, encoded by tet(K), blaZ/blaARL, mecA/mecA1, msrA/mphC, salA, dfrG, aacA-aphD, and str, respectively. We identified the first methicillin- and multidrug-resistant camel S. epidermidis strain of sequence type (ST) 1136 in East Africa. The pool of virulence-encoding genes was largest in the S. aureus strains, as expected, although other rather commensal strains contained distinct virulence-encoding genes. We identified toxin-antitoxin (TA) systems such as the hicA/hicB and abiEii/abiEi families, reported here for the first time for certain species of Staphylococcaceae. All strains contained at least one intact prophage sequence, mainly belonging to the Siphoviridae family. We pinpointed potential horizontal gene transfers between camel and cattle strains and also across distinct Staphylococcaceae clades and species. IMPORTANCE Camels are a high value and crucial livestock species in arid and semiarid regions of Africa and gain importance giving the impact of climate change on traditional livestock species. Our current knowledge with respect to Staphylococcaceae infecting camels is very limited compared to that for other livestock species. Better knowledge will foster the development of specific diagnostic assays, guide promising antimicrobial treatment options, and inform about potential zoonotic risks. We characterized 84 Staphylococcaceae strains isolated from camels with respect to their antimicrobial resistance and virulence traits. We detected potentially novel Staphylococcus species, resistances to different classes of antimicrobials, and the first camel multidrug-resistant S. epidermidis strain of sequence type 1136.

Highly diversified shrew hepatitis B viruses corroborate ancient origins and divergent infection patterns of mammalian hepadnaviruses.

Shrews, insectivorous small mammals, pertain to an ancient mammalian order. We screened 693 European and African shrews for hepatitis B virus (HBV) homologs to elucidate the enigmatic genealogy of HBV. Shrews host HBVs at low prevalence (2.5%) across a broad geographic and host range. The phylogenetically divergent shrew HBVs comprise separate species termed crowned shrew HBV (CSHBV) and musk shrew HBV (MSHBV), each containing distinct genotypes. Recombination events across host orders, evolutionary reconstructions, and antigenic divergence of shrew HBVs corroborated ancient origins of mammalian HBVs dating back about 80 million years. Resurrected CSHBV replicated in human hepatoma cells, but human- and tupaia-derived primary hepatocytes were resistant to hepatitis D viruses pseudotyped with CSHBV surface proteins. Functional characterization of the shrew sodium taurocholate cotransporting polypeptide (Ntcp), CSHBV/MSHBV surface peptide binding patterns, and infection experiments revealed lack of Ntcp-mediated entry of shrew HBV. Contrastingly, HBV entry was enabled by the shrew Ntcp. Shrew HBVs universally showed mutations in their genomic preCore domains impeding hepatitis B e antigen (HBeAg) production and resembling those observed in HBeAg-negative human HBV. Deep sequencing and in situ hybridization suggest that HBeAg-negative shrew HBVs cause intense hepatotropic monoinfections and low within-host genomic heterogeneity. Geographical clustering and low MSHBV/CSHBV-specific seroprevalence suggest focal transmission and high virulence of shrew HBVs. HBeAg negativity is thus an ancient HBV infection pattern, whereas Ntcp usage for entry is not evolutionarily conserved. Shrew infection models relying on CSHBV/MSHBV revertants and human HBV will allow comparative assessments of HBeAg-mediated HBV pathogenesis, entry, and species barriers.

Venezuelan Equine Encephalitis Complex Alphavirus in Bats, French Guiana.

Although essential for control strategies, knowledge about transmission cycles is limited for Venezuelan equine encephalitis complex alphaviruses (VEEVs). After testing 1,398 bats from French Guiana for alphaviruses, we identified and isolated a new strain of the encephalitogenic VEEV species Tonate virus (TONV). Bats may contribute to TONV spread in Latin America.

Ecologic Determinants of West Nile Virus Seroprevalence among Equids, Brazil.

Among 713 equids sampled in northeastern Brazil during 2013-2018, West Nile virus seroprevalence was 4.5% (95% CI 3.1%-6.3%). Mathematical modeling substantiated higher seroprevalence adjacent to an avian migratory route and in areas characterized by forest loss, implying increased risk for zoonotic infections in disturbed areas.

Limited Specificity of Serologic Tests for SARS-CoV-2 Antibody Detection, Benin.

We used commercially available ELISAs to test 68 samples from coronavirus disease cases and prepandemic controls from Benin. We noted <25% false-positive results among controls, likely due to unspecific immune responses elicited by acute malaria. Serologic tests must be carefully evaluated to assess coronavirus disease spread and immunity in tropical regions.

Mutations Associated with SARS-CoV-2 Variants of Concern, Benin, Early 2021.

Intense transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in Africa might promote emergence of variants. We describe 10 SARS-CoV-2 lineages in Benin during early 2021 that harbored mutations associated with variants of concern. Benin-derived SARS-CoV-2 strains were more efficiently neutralized by antibodies derived from vaccinees than patients, warranting accelerated vaccination in Africa.

Variable Sensitivity of SARS-CoV-2 Molecular Detection in European Expert Laboratories: External Quality Assessment, June and July 2020.

During the ongoing coronavirus disease 2019 (COVID-19) outbreak, robust detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a key element for clinical management and to interrupt transmission chains. We organized an external quality assessment (EQA) of molecular detection of SARS-CoV-2 for European expert laboratories. An EQA panel composed of 12 samples, containing either SARS-CoV-2 at different concentrations to evaluate sensitivity or other respiratory viruses to evaluate specificity of SARS-CoV-2 testing, was distributed to 68 laboratories in 35 countries. Specificity samples included seasonal human coronaviruses hCoV-229E, hCoV-NL63, and hCoV-OC43, as well as Middle East respiratory syndrome coronavirus (MERS-CoV), SARS-CoV, and human influenza viruses A and B. Sensitivity results differed among laboratories, particularly for low-concentration SARS-CoV-2 samples. Results indicated that performance was mostly independent of the selection of specific extraction or PCR methods.

Zika Virus Circulation in Mali.

The circulation of Zika virus (ZIKV) in Mali has not been clearly characterized. Therefore, we conducted a serologic survey of 793 asymptomatic volunteers >15 years of age (2016), and 637 blood donors (2013) to assess the seroprevalence of ZIKV infection in 2 ecoclimatic regions of Mali, tropical savannah and warm semiarid region, using ELISA and seroneutralization assays. The overall seroprevalence was ≈12% and increased with age, with no statistical difference between male and female participants. In the warm semiarid study sites we detected immunological markers of an outbreak that occurred in the late 1990s in 18% (95% CI 13%-23%) of participants. In tropical savannah sites, we estimated a low rate of endemic transmission, with 2.5% (95% CI 2.0%-3.1%) of population infected by ZIKV annually. These data demonstrate the circulation of ZIKV in Mali and provide evidence of a previously unidentified outbreak that occurred in the late 1990s.

Typical epidemiology of respiratory virus infections in a Brazilian slum.

Host population size, density, immune status, age structure, and contact rates are critical elements of virus epidemiology. Slum populations stand out from other settings and may present differences in the epidemiology of acute viral infections. We collected nasopharyngeal specimens from 282 children aged ≤5 years with acute respiratory tract infection (ARI) during 2005 to 2006 in one of the largest Brazilian slums. We conducted real-time reverse transcription-polymerase chain reaction (RT-PCR) for 16 respiratory viruses, nested RT-PCR-based typing of rhinoviruses (HRVs), and collected clinical symptoms. Viruses were common causes of respiratory disease; with ≥1 virus being detected in 65.2% of patients. We detected 15 different viruses during 1 year with a predominance of HRV (33.0%) and human respiratory syncytial virus (hRSV, 12.1%) infections, and a high rate of viral coinfections (28.3%). We observed seasonality of hRSV, HRV and human coronavirus infections, more severe symptoms in hRSV and influenza virus (FLU) infections and prolonged circulation of seven HRV clusters likely representing distinct serotypes according to genomic sequence distances. Potentially unusual findings included the absence of human metapneumovirus detections and lack of typical FLU seasonal patterns, which may be linked to the population size and density of the slum. Nonetheless, most epidemiological patterns were similar to other studies globally, suggesting surprising similarities of virus-associated ARI across highly diverse settings and a complex impact of population characteristics on respiratory virus epidemiology.

Rapid decline of Zika virus NS1 antigen-specific antibody responses, northeastern Brazil.

Zika virus (ZIKV) is a positive-stranded RNA virus within the Flaviviridae family. After decades of circulation in Asia, ZIKV was introduced to Brazil in 2014-2015, associated with a rise in congenital malformations. Unlike the genetically related dengue virus (DENV), ZIKV constitutes only one serotype. Although assumed that ZIKV infection may engender lifelong immunity, the long-term kinetics of ZIKV antibody responses are unclear. We assessed long-term kinetics of ZIKV NS1-IgG response in 144 individuals from 3 different subpopulations: HIV patients, tuberculosis patients and healthy individuals first tested in 2016 and retested 1.5-2 years after the 2015-2016 ZIKV epidemic in Salvador de Bahia, Brazil, using a widely distributed NS1-based commercial ELISA. The seropositivity in 2016 reached 59.0% (85/144, 95% confidence interval (CI) 50.7-66.7%), and decreased to 38.6% (56/144, CI 31.3-47.0%) 1.5-2 years later. In addition, the median ZIKV NS1-ELISA reactivity for individuals that remained positive in both timepoints significantly decreased from a ratio of 4.4 (95% CI 3.8-5.0) to 1.6 (95% CI 1.6-1.9) over the 2-year interval (Z: - 6.1; p < 0.001) irrespective of the subpopulation analyzed. Initial 2016 DENV antibody response was non-significant between groups, suggesting comparable DENV background. The high 20.6% seroreversion suggest that widely used serologic tests may fail to account a considerable proportion of past ZIKV infections in flavivirus endemic countries. In addition, ZIKV immunity might be shorter-lived than previously thought, which may contribute to local ZIKV resurgence once individual immune responses wane sufficiently to reduce community protective immunity in addition to birth and migration.