US008835117B2 (12) United States Patent (10) Patent No.: US 8,835,117 B2 Mitchell et al. (45) Date of Patent: Sep. 16, 2014 (54) NUCLEICACIDS FOR DETECTION AND OTHER PUBLICATIONS DISCRIMINATION OF GENOTYPES OF CHLAMYDOPHILA PSITTAC Nazarenko, I. Methods in Molecular Biology (2006) 335: 95-114.* Jeffrey et al. Microbiology (2007) 153: 2679-2688.* 75) Inventors: Stephaniep L. Mitchell, Somerville, MA Geens et al. Journal of Clinical Microbiology (2005) 43(5): 2456 (US); Jonas M. Winchell, Lilburn, GA 2461. (US) Geens et al., “Development of a Chlamydophila psittaci species specific and genotype-specific real-time PCR.” Vet. Res., 36: 787 (73) Assignee: The United States of America as 797, 2005. represented by the Secretary of the Geens et al., “Sequencing of the Chlamydophila psittaci OmpA Gene Department of Health and Human Reveals a New Genotype, E/B, and the Need for a Rapid Discrimi Services, Centers for Disease Control natory Genotyping Method.” J. Clin. Microbiol. 43(5): 2456-2461, and Prevention, Washington, DC (US) 2005. Heddema, “Genotyping of Chlamydophila psittaci in Human (*) Notice: Subject to any disclaimer, the term of this Samples.” Emerging Infectious Diseases, 12(12): 1989-1990, 2006. patent is extended or adjusted under 35 Menard, “Development of a real-time PCR for the detection of U.S.C. 154(b) by 82 days. Chlamydia psittaci,” J. Med. Microbiol. 55(Pt. 4): 471-473, 2006. Mitchellet al., “Genotyping of Chlamydophilapsittaci by Real-Time (21) Appl. No.: 13/322,787 PCR and High-Resolution Melt Analysis,” Journal of Clinical Microbiology, 47(1): 175-181, 2009. (22) PCT Filed: May 28, 2010 Sachse et al., “Genotyping of Chlamydophila psittaci using a new DNA microarray assay based on sequence analysis of ompA genes.” (86). PCT No.: PCT/US2O10/036742 BMC Microbiology, 8:63, pp. 1-12, 2008. S371 (c)(1), (2), (4) Date: Nov. 28, 2011 * cited by examiner (87) PCT Pub. No.: WO2010/138926 Primary Examiner — Angela MBertagna PCT Pub. Date: Dec. 2, 2010 (74) Attorney, Agent, or Firm — Klarduist Sparkman, LLP (65) Prior Publication Data US 2012/0082984 A1 Apr. 5, 2012 (57) ABSTRACT Methods of detecting Chlamydophila, including differentiat Related U.S. Application Data ing between species of Chlamydophila and/or strains of (60) Provisional application No. 61/182,628, filed on May Chlamydophila psittaci are disclosed, for example to detect 29, 2009. and genotype a Chlamydophila psittaci infection. A sample Suspected of containing a nucleic acid of a Chlamydophila, is (51) Int. Cl. screened for the presence of that nucleic acid. The presence of CI2O I/68 (2006.01) the Chlamydophila nucleic acid indicates the presence of the C7H 2L/04 (2006.01) Chlamydophila bacterium. Determining whether a Chlamy (52) U.S. Cl. dophila nucleic acid is present in a sample can be accom CPC ...................................... CI2O I/689 (2013.01) plished by detecting hybridization between a Chlamydophila USPC ... 435/6.12: 435/6.15:536/24.32:536/24.33 specific primer, a Chlamydophila psittaci specific primer, (58) Field of Classification Search and/or a Chlamydophila psittaci genotype-specific primer None and the Chlamydophila nucleic acid containing sample. Thus, See application file for complete search history. primers for the detection, species-specific and/or genotype specific identification of Chlamydophila psittaci are dis (56) References Cited closed. Kits that contain the disclosed primers also are dis U.S. PATENT DOCUMENTS closed. 6,884,784 B1* 4/2005 Mitchell et al. ................. 514/31 2006, OOO8828 A1 1/2006 Vanrompay 9 Claims, 10 Drawing Sheets U.S. Patent Sep. 16, 2014 Sheet 1 of 10 US 8,835,117 B2 s Eo: . S . o is Esee .. e o rt v. ise aoueoSejon entee U.S. Patent Sep. 16, 2014 Sheet 2 of 10 US 8,835,117 B2 FIG. 2 . six later test k g site. ior ... s ... lossessessssssssssssss U.S. Patent Sep. 16, 2014 Sheet 3 of 10 US 8,835,117 B2 sae mae De íbol Bouaission pazeltic N U.S. Patent Sep. 16, 2014 Sheet 4 of 10 US 8,835,117 B2 ”: sg) se ,! s: s ae addeosauori pezetion U.S. Patent US 8,835,117 B2 |0:0 30 sease. On pazieutoN U.S. Patent Sep. 16, 2014 Sheet 6 of 10 US 8,835,117 B2 FIG. 4A festyge: * * sies: sers: sse-assass seas). x & sesse yxxxx x axes sea-ww.we w w w w 8. E. * -X X xx xxx xx. 8w x w, ... we w w w w x. xxx xxx xxx x X x w w w w w www.xxxx xxx xxx x. $xx xxx. $$ x x 8 8x X x w8 saw 8.8% we w w xx aggers . .'"Ce actersectice. Accer C3 '''''C' Ceras. 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