# US 2019/0316167 A1 · Methods for Treating and Detecting Johne's Disease in Cattle

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- Number: US 2019/0316167 A1
- Document ID: US20190316167A1
- Status: Published application
- Kind: A1
- Application number: 16/385,970
- Filed: 2019-04-16
- Published: 2019-10-17
- Earliest claimed: 2018-04-16
- Assignees: CNA Diagnostics Inc.
- Applicants: CNA Diagnostics Inc.
- Inventors: Christoph W. Sensen; Jung Soh; Petra Heidinger; Laura Villanova; Stefan Grabuschnig
- Web page: https://patentgrep.com/patent/US20190316167A1

## CPC

- C12Q 1/04: Determining presence or kind of microorganism; Use of selective media for testing antibiotics or bacteriocides; Compositions containing a chemical indicator therefor {(C12Q1/6897 takes precedence)}
- A61K 31/7048: having oxygen as a ring hetero atom, e.g. leucoglucosan, hesperidin, erythromycin, nystatin {, digitoxin or digoxin}
- A61P 31/04: Antibacterial agents
- C12Q 1/6806: Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence)
- C12Q 1/6837: using probe arrays or probe chips (C12Q1/6874 takes precedence)
- C12Q 1/686: Polymerase chain reaction [PCR]
- C12Q 1/6874: involving nucleic acid arrays, e.g. sequencing by hybridisation
- C12Q 1/6876: Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q 1/6883: for diseases caused by alterations of genetic material
- C12Q 2600/112: Disease subtyping, staging or classification
- C12Q 2600/158: Expression markers
- C12Q 2600/16: Primer sets for multiplex assays
- C12Q 2600/166: Oligonucleotides used as internal standards, controls or normalisation probes
- G01N 2333/36: from Actinomyces; from Streptomyces (G)
- 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
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

Biomarkers for identifying Johne's disease in cattle are presented herein, as are related methods, uses, agents, and kits comprising same. Methods for treating, detecting, quarantining, and diagnosing Johne's disease in cattle are presented herein.

## Claims

### Claim 1

A method for treating a bovine animal suspected of having Johne's disease, the method comprising treating the bovine animal identified as having Johne's disease with a therapeutically effective amount of at least one agent used to treat Johne's disease, wherein the bovine animal is identifiable as having Johne's disease by
analyzing a biological sample isolated from the bovine animal for over-representation or under-representation of at least one polynucleotide relative to an internal standard region, wherein the at least one polynucleotide comprises any one of SEQ ID NOs: 1-16 or 134-164 and wherein the over-representation or under-representation of the at least one polynucleotide in the biological sample is a positive indicator of Johne's disease.

### Claim 2 (depends on 1)

The method of claim 1, wherein the at least one polynucleotide comprising any one of SEQ ID NOs: 1-16 or 134-164 over-represented or under-represented relative to the internal standard region is at least two, at least three, at least four, or at least five of the polynucleotides comprising any one of SEQ ID NOs: 1-16 or 134-164.

### Claim 3 (depends on 1)

The method of claim 1, wherein the biological sample is blood, a product derived from blood, or a fraction derived from blood.

### Claim 4 (depends on 3)

The method of claim 3, wherein the product derived from blood is plasma or serum.

### Claim 5 (depends on 1)

The method of claim 1, wherein detecting the over-representation or under-representation of the at least one polynucleotide relative to an internal standard region comprises at least one of a polymerase chain reaction (PCR)-based detection method, a hybridization-based method, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (MA), solid-phase enzyme immunoassay (EIA), mass spectrometry, and microarray analysis.

### Claim 6 (depends on 5)

The method of claim 5, wherein the PCR-based detection method comprises amplifying nucleic acid sequences in the biological sample using primers that are specific for and capable of amplifying any one of SEQ ID NOs: 1-16 or 134-164, wherein the amplifying generates amplification products corresponding to any one of SEQ ID NOs: 1-16 or 134-164 when the biological sample comprises any one of SEQ ID NOs: 1-16 or 134-164.

### Claim 7 (depends on 5)

The method of claim 5, wherein the PCR-based detection method is performed using at least one primer pair, wherein each primer pair of the at least one primer pair is specific for any one of SEQ ID NOs: 1-16 or 134-164.

### Claim 8 (depends on 7)

The method of claim 7, wherein the primer pair specific for any one of SEQ ID NOs: 1-16 or 134-164 is any one of the primer pairs presented in Table 1.

### Claim 9 (depends on 6)

The method of claim 6, further comprising sequencing the amplification products corresponding to any one of SEQ ID NOs: 1-16 or 134-164.

### Claim 10 (depends on 6)

The method of claim 6, wherein the nucleic acid sequences comprise circulating nucleic acid.

### Claim 11 (depends on 1)

The method of claim 1, wherein the at least one agent used to treat Johne's disease comprises at least one of antibiotic.

### Claim 12 (depends on 11)

The method of claim 11, wherein the at least antibiotic is in a genus comprising rifabutin.

### Claim 13 (depends on 11)

The method of claim 11, wherein the at least antibiotic is in a genus comprising clarithromycin.

### Claim 14 (depends on 11)

The method of claim 11, wherein the at least antibiotic comprises rifabutin and clarithromycin.

### Claim 15 (depends on 1)

The method of claim 1, wherein the bovine animal is monitored for Johne's disease.

### Claim 16

A probe comprising a manmade nucleotide sequence capable of binding specifically to a polynucleotide comprising any one of SEQ ID NOs: 1-16 or 134-164 and at least one manmade tag conjugated thereto, wherein the manmade nucleotide sequence is complementary to the polynucleotide comprising any one of SEQ ID NOs: 1-16 or 134-164.

### Claim 17 (depends on 16)

The probe of claim 16, wherein the manmade nucleotide sequence capable of binding specifically to a polynucleotide comprising any one of SEQ ID NOs: 1-16 or 134-164 exhibits at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, or at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% complementarity to any one of SEQ ID NOs: 1-16 or 134-164.

### Claim 18 (depends on 16)

The probe of claim 16, wherein the manmade tag is a detectable marker.

### Claim 19

An array comprising at least one probe comprising a manmade nucleotide sequence capable of binding specifically to a polynucleotide comprising any one of SEQ ID NOs: 1-16 or 134-164, wherein the manmade nucleotide sequence is complementary to the polynucleotide comprising any one of SEQ ID NOs: 1-16 or 134-164, wherein the at least one probe is bound to a solid surface.

### Claim 20 (depends on 19)

The array of claim 19, comprising a microarray, gene chip, DNA chip, or a FILMARRAY®.