NASA Technical Reports Server (NTRS) 20160001291: Methods for Detecting and Quantifying Viable Bacterial Endo-Spores

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(12) United States Patent
(1o) Patent No.: (cid:9) US 9,228,240 B2
Venkateswaran et al. (45) Date of Patent: (cid:9) Jan. 5, 2016
(54) METHODS FOR DETECTING AND removal of DNA from dead cells, Journal of Microbiological Meth-
QUANTIFYING VIABLE BACTERIAL ods, 67:310-320, 2006.*
ENDO-SPORES Riedy et al., Use of a Photolabeling Technique to Identify Nonviable
Cells in Fixed Homologous or Heterologous Cell Populations,
(75) Inventors: Kasthuri J. Venkateswaran, Azusa, CA Cytometry, 12:133-139, 1991.*
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Bacterial Endospore, attached, Oct. 23, 2008.*
(73) Assignee: CALIFORNIA INSTITUTE OF Henriques et al. (Structure and Assembly of the Bacterial Endospore
TECHNOLOGY, Pasadena, CA (US) Coat, Methods, 20:95-110, 2000).*
Rawsthorne et al., PCR-Based Method Using Propidium Monoazide
(*) Notice: (cid:9) Subject to any disclaimer, the term of this to Distinguish Viable from Nonviable Bacillus subtilis Spores,
patent is extended or adjusted under 35 Applied and Environmental Microbiology, May 2009, p. 2936-
U.S.C. 154(b) by 53 days. 2939.*
Liu et al., Formation and Composition of the Bacillus anthracis
(21) Appl. No.: 13/152,213 Endospore, Journal of Bacteriology, Jan. 2004, p. 164-178.*
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(22) Filed: (cid:9) Jun. 2, 2011 fluorescent dyes, Journal of Applied Microbiology 2004, 96, 684-
692).*
(65) (cid:9) Prior Publication Data Kelly et al. (Use of the direct epifluorescent filter technique for the
enumeration of bacterial spores, Journal of Applied Bacteriology
US 2011/0318750 Al (cid:9) Dec. 29, 2011 1987,63, 545-550).*
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pling, Sterilization Validation to Astrobiology, dissertation, attached,
Related U.S. Application Data
available May 9, 2008).*
(60) Provisional application No. 61/351,238, filed on Jun. Kroll (The Direct Epifluorescent Filter Technique (DEFT), in Meth-
3, 2010, provisional application No. 61/441,820, filed ods in Molecular Biology, vol. 46: Diagnostic Bacteriology Proto-
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CPC (cid:9) .............. C12Q 11689 (2013.01); C12Q 116848
Amann, R.I., Krumholz, L., and Stahl, D.A. (1990) Fluorescent-
(2013.01)
Oligonucleotide Probing of Whole Cells for Determinative,
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tive analysis of methods for the purification of DNA from low-
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8,021,848 B2 * 9/2011 (cid:9) Straus (cid:9) ............................ (cid:9) 435/7.1 Primary Examiner Stephanie K Mummert
8,771,940 B2 7/2014 (cid:9) Andersen et al.
Assistant Examiner Aaron Priest
2011/0053790 Al * 3/2011 (cid:9) Yoshida et al . (cid:9) ................... (cid:9) 506/9
(74) Attorney, Agent, or Firm Steinfl & Bruno LLP
FOREIGN PATENT DOCUMENTS
(57) (cid:9) ABSTRACT
WO (cid:9) 2010/151842 (cid:9) 12/2010
Methods and systems for detecting viable bacterial
OTHER PUBLICATIONS endospores in a sample and related methods to quantify
viable bacterial endospores in a sample.
Nocker et al. (Comparison of propidium monoazide with ethidium
monoazide for differentiation of live vs. dead bacteria by selective 14 Claims, 7 Drawing Sheets
US 9,228,240 B2
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652-654. * cited by examiner
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