Suxiang Tong

A distinct lineage of influenza A virus from bats

Influenza A virus reservoirs in animals have provided novel genetic elements leading to the emergence of global pandemics in humans. Most influenza A viruses circulate in waterfowl, but those that infect mammalian hosts are thought to pose the greatest risk for zoonotic spread to humans and the generation of pandemic or panzootic viruses. We have identified an influenza A virus from little yellow-shouldered bats captured at two locations in Guatemala. It is significantly divergent from known influenza A viruses. The HA of the bat virus was estimated to have diverged at roughly the same time as the known subtypes of HA and was designated as H17. The neuraminidase (NA) gene is highly divergent from all known influenza NAs, and the internal genes from the bat virus diverged from those of known influenza A viruses before the estimated divergence of the known influenza A internal gene lineages. Attempts to propagate this virus in cell cultures and chicken embryos were unsuccessful, suggesting distinct requirements compared with known influenza viruses. Despite its divergence from known influenza A viruses, the bat virus is compatible for genetic exchange with human influenza viruses in human cells, suggesting the potential capability for reassortment and contributions to new pandemic or panzootic influenza A viruses.

New World Bats Harbor Diverse Influenza A Viruses

Aquatic birds harbor diverse influenza A viruses and are a major viral reservoir in nature. The recent discovery of influenza viruses of a new H17N10 subtype in Central American fruit bats suggests that other New World species may similarly carry divergent influenza viruses. Using consensus degenerate RT-PCR, we identified a novel influenza A virus, designated as H18N11, in a flat-faced fruit bat (Artibeus planirostris) from Peru. Serologic studies with the recombinant H18 protein indicated that several Peruvian bat species were infected by this virus. Phylogenetic analyses demonstrate that, in some gene segments, New World bats harbor more influenza virus genetic diversity than all other mammalian and avian species combined, indicative of a long-standing host-virus association. Structural and functional analyses of the hemagglutinin and neuraminidase indicate that sialic acid is not a ligand for virus attachment nor a substrate for release, suggesting a unique mode of influenza A virus attachment and activation of membrane fusion for entry into host cells. Taken together, these findings indicate that bats constitute a potentially important and likely ancient reservoir for a diverse pool of influenza viruses.

Case Definitions, Diagnostic Algorithms, and Priorities in Encephalitis: Consensus Statement of the International Encephalitis Consortium

BACKGROUND Encephalitis continues to result in substantial morbidity and mortality worldwide. Advances in diagnosis and management have been limited, in part, by a lack of consensus on case definitions, standardized diagnostic approaches, and priorities for research. METHODS In March 2012, the International Encephalitis Consortium, a committee begun in 2010 with members worldwide, held a meeting in Atlanta to discuss recent advances in encephalitis and to set priorities for future study. RESULTS We present a consensus document that proposes a standardized case definition and diagnostic guidelines for evaluation of adults and children with suspected encephalitis. In addition, areas of research priority, including host genetics and selected emerging infections, are discussed. CONCLUSIONS We anticipate that this document, representing a synthesis of our discussions and supported by literature, will serve as a practical aid to clinicians evaluating patients with suspected encephalitis and will identify key areas and approaches to advance our knowledge of encephalitis.

Identification of a Novel Astrovirus (Astrovirus VA1) Associated with an Outbreak of Acute Gastroenteritis

ABSTRACT The etiology of a large proportion of gastrointestinal illness is unknown. In this study, random Sanger sequencing and pyrosequencing approaches were used to analyze fecal specimens from a gastroenteritis outbreak of unknown etiology in a child care center. Multiple sequences with limited identity to known astroviruses were identified. Assembly of the sequences and subsequent reverse transcription-PCR (RT-PCR) and rapid amplification of cDNA ends generated a complete genome of 6,586 nucleotides. Phylogenetic analysis demonstrated that this virus, named astrovirus VA1 (AstV-VA1), is highly divergent from all previously described astroviruses. Based on RT-PCR, specimens from multiple patients in this outbreak were unequivocally positive for Ast-VA1.

Sensitive and Broadly Reactive Reverse Transcription-PCR Assays To Detect Novel Paramyxoviruses

ABSTRACT We have developed a set of reverse transcription-PCR assays for the detection and identification of known and novel paramyxoviruses in clinical specimens. Primers were designed from the conserved motifs of the polymerase pol gene sequences to detect members of the Paramyxovirinae or Pneumovirinae subfamily or groups of genera within the Paramyxovirinae subfamily. The consensus-degenerate hybrid oligonucleotide primer design and seminested or nested PCR assay design were used to enhance the breadth of reactivity and sensitivity of the respective assays. Using expressed RNA and 10-fold dilution series of virus-infected tissue culture isolates from different members of the family or genera, these assays were able to detect on average between 100 and 500 copies of template RNA. The assays were specific to the respective group of genera or subfamily viruses. This set of primers enhances our ability to look for novel viruses in outbreaks and diseases of unknown etiology.

Hemagglutinin homologue from H17N10 bat influenza virus exhibits divergent receptor-binding and pH-dependent fusion activities

Bat influenza virus H17N10 represents a distinct lineage of influenza A viruses with gene segments coding for proteins that are homologs of the surface antigens, hemagglutinin (HA) and neuraminidase (NA). Our recent study of the N10 NA homolog revealed an NA-like structure, but with a highly divergent putative active site exhibiting little or no NA activity, and provided strong motivation for performing equivalent structural and functional analyses of the H17 HA protein. The overall structure of the H17 HA homolog from A/little yellow-shouldered bat/Guatemala/060/2010 at 3.18 Å resolution is very similar to other influenza HAs, with a putative receptor-binding site containing some conserved aromatic residues that form the base of the sialic acid binding site. However, the rest of the H17 receptor-binding site differs substantially from the other HA subtypes, including substitution of other conserved residues associated with receptor binding. Significantly, electrostatic potential analyses reveal that this putative receptor-binding site is highly acidic, making it unfavorable to bind any negatively charged sialylated receptors, consistent with the recombinant H17 protein exhibiting no detectable binding to sialylated glycans. Furthermore, the fusion mechanism is also distinct; trypsin digestion with recombinant H17 protein, when exposed to pH 4.0, did not degrade the HA1 and HA2, in contrast to other HAs. These distinct structural features and functional differences suggest that the H17 HA behaves very differently compared with other influenza HAs.

Real-time reverse transcription-polymerase chain reaction assay for SARS-associated coronavirus.

A real-time reverse transcription–polymerase chain reaction (RT-PCR) assay was developed to rapidly detect the severe acute respiratory syndrome–associated coronavirus (SARS-CoV). The assay, based on multiple primer and probe sets located in different regions of the SARS-CoV genome, could discriminate SARS-CoV from other human and animal coronaviruses with a potential detection limit of <10 genomic copies per reaction. The real-time RT-PCR assay was more sensitive than a conventional RT-PCR assay or culture isolation and proved suitable to detect SARS-CoV in clinical specimens. Application of this assay will aid in diagnosing SARS-CoV infection.

Detection of Novel SARS-like and Other Coronaviruses in Bats from Kenya

Diverse coronaviruses have been identified in bats from several continents but not from Africa. We identified group 1 and 2 coronaviruses in bats in Kenya, including SARS-related coronaviruses. The sequence diversity suggests that bats are well-established reservoirs for and likely sources of coronaviruses for many species, including humans.

Laboratory Diagnosis of Four Recent Sporadic Cases of Community-acquired SARS, Guangdong Province, China

Four sporadic cases of SARS-associated coronavirus infection were identified through collaboration of four laboratories.

Descriptive epidemiology of fatal respiratory outbreaks and detection of a human-related metapneumovirus in wild chimpanzees (Pan troglodytes) at Mahale Mountains National Park, Western Tanzania

Over the past several years, acute and fatal respiratory illnesses have occurred in the habituated group of wild chimpanzees at the Mahale Mountains National Park, Tanzania. Common respiratory viruses, such as measles and influenza, have been considered possible causative agents; however, neither of these viruses had been detected. During the fatal respiratory illnesses in 2003, 2005 and 2006, regular observations on affected individuals were recorded. Cause‐specific morbidity rates were 98.3, 52.4 and 33.8%, respectively. Mortality rates were 6.9, 3.2 and 4.6%; all deaths were observed in infants 2 months–2 years 9 months of age. Nine other chimpanzees have not been seen since the 2006 outbreak and are presumed dead; hence, morbidity and mortality rates for 2006 may be as high as 47.7 and 18.5%, respectively. During the 2005 and 2006 outbreaks, 12 fecal samples were collected from affected and nonaffected chimpanzees and analyzed for causative agents. Analysis of fecal samples from 2005 suggests the presence of paramyxovirus, and in 2006 a human‐related metapneumovirus was detected and identified in an affected chimpanzee whose infant died during the outbreak. Our findings provide preliminary evidence that the causative agent associated with these illnesses is viral and contagious, possibly of human origin; and that, possibly more than one agent may be circulating in the population. We recommend that baseline health data be acquired and food wadge and fecal samples be obtained and bio‐banked as early as possible when attempting to habituate new groups of chimpanzees or other great apes. For already habituated populations, disease prevention strategies, ongoing health monitoring programs and reports of diagnostic findings should be an integral part of managing these populations. In addition, descriptive epidemiology should be a major component of disease outbreak investigations. Am. J. Primatol. 70: 755–765, 2008.