A J da Silva1, F J Bornay-Llinares, I N Moura, S B Slemenda, J L Tuttle, N J Pieniazek. 1. Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Public Health Service, U.S. Department of Health and Human Services, Atlanta, Georgia 30341-3724, USA.
Abstract
BACKGROUND: Polymerase chain reaction (PCR) detection of intestinal protozoa in fecal specimens is hampered by poor recovery of DNA and by the presence of PCR inhibitors. In this study we describe a novel method for DNA extraction from such specimens containing spores and oocysts of Enterocytozoon bieneusi and Cryptosporidium parvum, respectively. METHODS AND RESULTS: Extraction was done using commercial kits modified to maximize the recovery and purity of extracted DNA. In comparison with a procedure we previously reported, we estimate that this method may increase the sensitivity of parasite DNA detection in fecal specimens up to tenfold. An additional advantage of this method is that up to 12 samples may be processed simultaneously within 2 hours. CONCLUSIONS: By using this method, we were able to increase reproducibility of PCR amplification on fecal specimens and significantly reduce the hands-on time required to process the samples.
BACKGROUND: Polymerase chain reaction (PCR) detection of intestinal protozoa in fecal specimens is hampered by poor recovery of DNA and by the presence of PCR inhibitors. In this study we describe a novel method for DNA extraction from such specimens containing spores and oocysts of Enterocytozoon bieneusi and Cryptosporidium parvum, respectively. METHODS AND RESULTS: Extraction was done using commercial kits modified to maximize the recovery and purity of extracted DNA. In comparison with a procedure we previously reported, we estimate that this method may increase the sensitivity of parasite DNA detection in fecal specimens up to tenfold. An additional advantage of this method is that up to 12 samples may be processed simultaneously within 2 hours. CONCLUSIONS: By using this method, we were able to increase reproducibility of PCR amplification on fecal specimens and significantly reduce the hands-on time required to process the samples.
Authors: Yvonne Qvarnstrom; James J Sullivan; Henry S Bishop; Robert Hollingsworth; Alexandre J da Silva Journal: Appl Environ Microbiol Date: 2006-12-28 Impact factor: 4.792
Authors: Adriana S Lopez; Jean M Bendik; Jean Y Alliance; Jacquelin M Roberts; Alexandre J da Silva; Iaci N S Moura; Michael J Arrowood; Mark L Eberhard; Barbara L Herwaldt Journal: J Clin Microbiol Date: 2003-05 Impact factor: 5.948
Authors: Yvonne Qvarnstrom; Theresa Benedict; Paula L Marcet; Ryan E Wiegand; Barbara L Herwaldt; Alexandre J da Silva Journal: Parasitology Date: 2017-11-08 Impact factor: 3.234
Authors: Thaddeus K Graczyk; Michael A Johansson; Leena Tamang; Govinda S Visvesvara; Laci S Moura; Alexandre J DaSilva; Autumn S Girouard; Olga Matos Journal: J Clin Microbiol Date: 2007-02-07 Impact factor: 5.948
Authors: Alice Y Ho; Adriana S Lopez; Michael G Eberhart; Robert Levenson; Bernard S Finkel; Alexandre J da Silva; Jacquelin M Roberts; Palmer A Orlandi; Caroline C Johnson; Barbara L Herwaldt Journal: Emerg Infect Dis Date: 2002-08 Impact factor: 6.883