	US 12,404,558 B2
	Methods and assays for detection and subtyping of microbial pathogens
James Schupp, Flagstaff, AZ (US); Jason Sahl, Phoenix, AZ (US); Paul Keim, Flagstaff, AZ (US); David Engelthaler, Flagstaff, AZ (US); David Wagner, Phoenix, AZ (US); and Viacheslav Fofanov, Phoenix, AZ (US)
Assigned to The Translational Genomics Research Institute, Phoenix, AZ (US); and Arizona Board of Regents on Behalf of Northern Arizona University, Flagstaff, AZ (US)
Appl. No. 17/426,600
Filed by The Translational Genomics Research Institute, Phoenix, AZ (US); and Arizona Board of Regents on Behalf of Northern Arizona University, Flagstaff, AZ (US)
PCT Filed Jan. 28, 2020, PCT No. PCT/US2020/015395 § 371(c)(1), (2) Date Jul. 28, 2021, PCT Pub. No. WO2020/159977, PCT Pub. Date Aug. 6, 2020.
Claims priority of provisional application 62/798,463, filed on Jan. 29, 2019.
Prior Publication US 2022/0090172 A1, Mar. 24, 2022
Int. Cl. C12Q 1/689 (2018.01); C12Q 1/04 (2006.01); C12Q 1/6844 (2018.01); C12Q 1/686 (2018.01); C12R 1/07 (2006.01)

CPC C12Q 1/689 (2013.01) [C12Q 1/04 (2013.01); C12Q 1/6844 (2013.01); C12Q 1/686 (2013.01); C12Q 2600/156 (2013.01); C12Q 2600/16 (2013.01); C12R 2001/07 (2021.05)]

20 Claims

1. A method of detecting Bacillus anthracis in a sample, comprising:

generating one or more amplicons from the sample using at least one primer pair comprising a forward and reverse primer in at least one amplification reaction;

sequencing the one or more amplicons using next-generation sequencing; and

detecting at least one B. anthracis-specific amplicon in the sequenced amplicons, wherein each forward and reverse primer comprise a forward and reverse sequence selected from the group consisting of:

SEQ ID NO: 1 and SEQ ID NO: 2;

SEQ ID NO: 3 and SEQ ID NO: 4;

SEQ ID NO: 5 and SEQ ID NO: 6;

SEQ ID NO: 7 and SEQ ID NO: 8;

SEQ ID NO: 9 and SEQ ID NO: 10;

SEQ ID NO: 11 and SEQ ID NO: 12;

SEQ ID NO: 13 and SEQ ID NO: 14;

SEQ ID NO: 15 and SEQ ID NO: 16; and

SEQ ID NO: 17 and SEQ ID NO: 18;

wherein the presence of the at least one B. anthracis-specific amplicon indicates the presence of B. anthracis in the sample, and the absence of the at least one B. anthracis-specific amplicon indicates the absence of B. anthracis from the sample.