

@article {11005,
	title = {A {Real}-{Time} {PCR} {Method} for {Quantifying} {Viable} {Ascaris} {Eggs} {Using} the {First} {Internally} {Transcribed} {Spacer} {Region} of {Ribosomal} {DNA}},
	journal = {Applied and Environmental Microbiology},
	volume = {72},
	number = {12},
	year = {2006},
	month = {dec},
	pages = {7864{\textendash}7872},
	abstract = {Worldwide, 1.4 billion people are infected with the intestinal worm Ascaris lumbricoides. As a result, Ascaris eggs are commonly found in wastewater and sludges. The current microscopy method for detecting viable Ascaris eggs is time- and labor-intensive. The goal of this study was to develop a real-time quantitative PCR (qPCR) method to determine the levels of total and viable Ascaris eggs in laboratory solutions using the first internally transcribed spacer (ITS-1) region of ribosomal DNA (rDNA) and rRNA. ITS-1 rDNA levels were proportional to Ascaris egg cell numbers, increasing as eggs developed from single cells to mature larvae and ultimately reaching a constant level per egg. Treatments causing {\textgreater}99\% inactivation (high heat, moderate heat, ammonia, and UV) eliminated this increase in ITS-1 rDNA levels and caused decreases that were dependent on the treatment type. By taking advantage of this difference in ITS-1 rDNA level between viable, larvated eggs and inactivated, single-celled eggs, qPCR results were used to develop inactivation profiles for the different treatments. No statistical difference from the standard microscopy method was found in 75\% of the samples (12 of 16). ITS-1 rRNA was detected only in samples containing viable eggs, but the levels were more variable than rDNA levels and ITS-1 rRNA could not be used for quantification. The detection limit of the rDNA-based method was approximately one larvated egg or 90 single-celled eggs; the detection limit for the rRNA-based method was several orders of magnitude higher. The rDNA qPCR method is promising for both research and regulatory applications.},
	issn = {0099-2240},
	doi = {10.1128/AEM.01983-06},
	url = {http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1694259/},
	author = {Pecson, Brian M and Barrios, Jos{\'e} Antonio and Johnson, David R and Nelson, Kara L}
}
