# US 2011/0045018 A1 · Astrovirus species

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- Number: US 2011/0045018 A1
- Document ID: US20110045018A1
- Status: Published application
- Kind: A1
- Application number: 12/859,068
- Filed: 2010-08-18
- Published: 2011-02-24
- Earliest claimed: 2009-08-19
- Assignees: Blood Systems, Inc.
- Inventors: ERIC DELWART; AMIT KAPOOR
- Web page: https://patentgrep.com/patent/US20110045018A1

## CPC

- C12Q 1/70: involving virus or bacteriophage {(immunoassay for viruses G01N33/56983)}
- A61P 31/14: for RNA viruses
- C07K 14/005: from viruses
- C12N 7/00: Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof (preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00)
- C12N 2770/12022: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2770/12051: Methods of production or purification of viral material
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

Provided herein are sequences of the genomes and encoded proteins of new astrovirus species, and variants thereof. Also provided are methods of detecting the new astrovirus species and diagnosing astrovirus infection, methods of treating or preventing astrovirus infection, and methods for identifying anti-astrovirus compounds. Provided are two new astrovirus species named HMOAstV-A and HMOAstV-B, and both are distantly related to known astroviruses. Also provided is a new method of classifying astroviruses, where there are three groups of human astroviruses, including HAstV, AstV-MLB, and HMOAstV.

## Claims

### Claim 1

An isolated nucleic acid molecule comprising a nucleotide sequence having at least 75% identity to SEQ ID NO:1, SEQ ID NO:5, or a complement thereof, wherein the nucleotide sequence is at least 12, 20, 25, 30, 40, 50, 75, 100, or 200 nucleotides in length.

### Claim 2

An isolated nucleic acid molecule comprising a nucleotide sequence that hybridizes under highly stringent conditions to at least 12, 25, 50, 100, or 150 contiguous nucleotides of a nucleotide sequence of SEQ ID NO:1, SEQ ID NO:5, or a complement thereof, wherein the hybridization reaction is incubated at 42° C. in a solution comprising 50% formamide, 5×SSC, and 1% SDS and washed at 65° C. in a solution comprising 0.2×SSC and 0.1% SDS.

### Claim 3 (depends on 1)

The nucleic acid molecule of claim 1, wherein the nucleotide sequence is at least 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to SEQ ID NO:1, SEQ ID NO:5, or a complement thereof.

### Claim 4 (depends on 1)

The nucleic acid of claim 1, wherein the nucleotide sequence comprises an open reading frame.

### Claim 5 (depends on 1)

A substantially purified protein encoded by a nucleotide sequence of claim 1.

### Claim 6

A substantially purified protein comprising an amino acid sequence at least 80% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

### Claim 7 (depends on 6)

The protein of claim 6, comprising an amino acid sequence at least 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% identical to an amino acid sequence selected from the group consisting of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

### Claim 8 (depends on 6)

An isolated antibody that specifically binds to the protein of claim 6.

### Claim 9 (depends on 6)

A purified serum composition comprising a polyclonal antibody that specifically binds to the protein of claim 6.

### Claim 10 (depends on 1)

An isolated astrovirus comprising the nucleic acid molecule of claim 1.

### Claim 11 (depends on 1)

An expression vector comprising the nucleic acid molecule of claim 1.

### Claim 12 (depends on 11)

A host cell comprising the expression vector of claim 11.

### Claim 13

A method of detecting an astrovirus nucleic acid comprising:
(a) contacting a sample suspected of containing an astrovirus nucleic acid with a nucleotide sequence that hybridizes under highly stringent conditions to a nucleotide sequence of SEQ ID NO:1, SEQ ID NO:5, or a complement thereof; and
(b) detecting the presence or absence of hybridization.

### Claim 14

A method of detecting an astrovirus nucleic acid comprising:
(a) amplifying the nucleic acid of a sample suspected of containing astrovirus nucleic acid with at least one primer that hybridizes to a nucleotide sequence of SEQ ID NO:1, SEQ ID NO:5, or a complement thereof, to produce an amplification product; and
(b) detecting the presence of an amplification product, thereby detecting the presence of the astrovirus nucleic acid.

### Claim 15

A method of detecting an astrovirus infection in a sample comprising:
(a) contacting a sample suspected of containing an astrovirus protein with an antibody that specifically binds to a polypeptide encoded by SEQ ID NO:1 or SEQ ID NO:5 to form a protein/antibody complex; and
(b) detecting the presence of the protein/antibody complex, thereby detecting the presence of the astrovirus protein.

### Claim 16

A kit for detecting an anstrovirus nucleic acid comprising at least one oligonucleotide primer that hybridizes to a nucleotide sequence of SEQ ID NO:1, SEQ ID NO:5, or a complement thereof, under highly stringent PCR conditions.

### Claim 17

A method of assaying for an anti-astrovirus compound comprising:
(a) contacting a sample containing an astrovirus with a test compound, the astrovirus comprising a genome that hybridizes under highly stringent conditions to a nucleotide sequence of SEQ ID NO:1, SEQ ID NO:5, or a complement thereof; and
(b) determining whether the test compound inhibits astrovirus replication, wherein inhibition of astrovirus replication indicates that the test compound is an anti-astrovirus compound.

### Claim 18

A method of treating or preventing an astrovirus infection in a subject comprising: administering to the subject an antigen encoded by an astrovirus, the astrovirus comprising a genome that hybridizes under highly stringent conditions to a nucleotide sequence of SEQ ID NO:1, SEQ ID NO:5, or a complement thereof; thereby treating or preventing infection in the subject.

### Claim 19

A method for detecting and serotyping astrovirus in a sample comprising:
(a) contacting a first portion of the sample with a first pair of primers in a first amplification protocol, wherein the first pair of primers have an associated first characteristic amplification product if an astrovirus is present in the sample;
(b) determining whether or not the first characteristic amplification product is present;
(c) contacting a second portion of the sample with a second pair of primers in a second amplification protocol, wherein the second pair of primers have an associated second characteristic amplification product if an astrovirus is present in the sample and wherein the second pair of primers are different from the first pair of primers;
(d) determining whether or not the second characteristic amplification product is present;
(e) based on whether or not the first and second characteristic amplification product are present, selecting one or more subsequent pair of primers and contacting the one or more subsequent pair of primers with additional portions of the sample in subsequent amplification protocols, wherein each subsequent pair of primers is different from each pair of primers already used and wherein each subsequent pair of primers has an associated subsequent characteristic amplification product if an astrovirus is present in the sample;
(f) determining whether or not the associated characteristic amplification product for each subsequent pair of primers used is present;
(g) repeating steps (e) and (f) for one or more subsequent pairs of primers if the astrovirus cannot be serotyped based on the determinations of steps (b), (d), and (f) until the astrovirus can be serotyped, wherein the one or more subsequent pairs of primers are different from all pairs of primers used in earlier amplification protocols; and
(h) determining the serotype or groups of serotypes of the astrovirus that may be present in the sample.

### Claim 20 (depends on 19)

The method of claim 19, wherein the astrovirus has a genome comprising a nucleic acid sequence of SEQ ID NO:1, SEQ ID NO:5, or a complement thereof.

### Claim 21 (depends on 19)

The method of claim 19, wherein the astrovirus has a genome comprising a nucleic acid sequence encoding an amino acid sequence selected from the group consisting of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

### Claim 22 (depends on 19)

The method of claim 19, wherein the first, second, and any subsequent amplification protocols are polymerase chain reactions or reverse-transcription polymerase chain reactions.

### Claim 23 (depends on 19)

The method of claim 19, wherein the detecting and serotyping of the astrovirus in the sample is used to diagnose a viral disease or medical condition.

### Claim 24 (depends on 23)

The method of claim 23, wherein the viral disease or medical condition is a gastrointestinal tract infection.

### Claim 25

A method for detecting the presence of an astrovirus in a sample comprising:
(a) purifying RNA contained in the sample;
(b) reverse transcribing the RNA with primers effective to reverse transcribe astrovirus RNA to provide a cDNA;
(c) contacting at least a portion of the cDNA with (i) a composition that promotes amplification of a nucleic acid and (ii) an oligonucleotide mixture wherein the mixture comprises at least one oligonucleotide that hybridizes to a highly conserved sequence of the sense strand of an astrovirus nucleic acid and at least one oligonucleotide that hybridizes to a highly conserved sequence of the antisense strand of an astrovirus nucleic acid;
(d) carrying out an amplification procedure on the amplification mixture such that, if an astrovirus is present in the sample, an astrovirus amplicon is produced whose sequence comprises a nucleotide sequence of at least a portion of the astrovirus genome; and
(e) detecting whether an amplicon is present; wherein the presence of the amplicon indicates that an astrovirus is present in the sample.

### Claim 26 (depends on 25)

The method of claim 25, wherein the astrovirus has a genome comprising a nucleic acid sequence of SEQ ID NO:1 SEQ ID NO:5, or a complement thereof.

### Claim 27 (depends on 25)

The method of claim 25, wherein the astrovirus has a genome comprising a nucleic acid sequence encoding an amino acid sequence selected from the group consisting of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

### Claim 28 (depends on 25)

The method of claim 25, wherein the detecting of the astrovirus in the sample is used to diagnose a viral disease or medical condition.

### Claim 29 (depends on 28)

The method of claim 28, wherein the viral disease or medical condition is an gastrointestinal tract infection.