caa a22 4500 445340029 CHVBK 20180317142809.0 cr unu---uuuuu 170323e20110901xx s 000 0 eng 10.1007/s00436-011-2312-9 doi (NATIONALLICENCE)springer-10.1007/s00436-011-2312-9 Use of PCR to detect Entamoeba gingivalis in diseased gingival pockets and demonstrate its absence in healthy gingival sites [Elektronische Daten] [Robert Trim, Michael Skinner, Mary Farone, John DuBois, Anthony Newsome] Investigators using light microscopy have identified the protozoan parasite Entamoeba gingivalis from diseased gingival pockets for nearly 100years. The objective of the present investigation was to develop a molecular biology approach for determining the presence of E. gingivalis in both diseased gingival pockets and healthy gingival sites. For this, a previously developed conventional polymerase chain reaction (PCR) was evaluated and a real-time polymerase chain reaction assay was developed. Paper points were inserted into the base of the sulcus of both diseased gingival pockets and healthy gingival sites. DNA was extracted using the QIAamp DNA mini kit, and subsequently analyzed using conventional and real-time PCR analysis. A previously described primer set specific for the small subunit ribosomal RNA gene (SSU rDNA) of E. gingivalis was used for the conventional PCR. For the real-time PCR, a primer set was designed to amplify a 135-bp fragment inside the SSU rDNA of E. gingivalis. A conventional PCR assay detected E. gingivalis in 27% of diseased gingival pockets. The real-time PCR using a different primer set detected protozoa in 69% of diseased pocket sites. Thus, the latter technique proved more sensitive for detection of E. gingivalis. No E. gingivalis were detected in any of the healthy gingival pocket sites using either type of PCR assay. Results support a concept that the presence of E. gingivalis is associated only with diseased gingival pocket sites. The newly described methodology may also serve to provide a novel eukaryotic cell marker of disease status in gingival pockets. Springer-Verlag, 2011 Trim Robert University of Tennessee School of Dentistry, 875 Union Ave, Memphis, TN, USA aut Skinner Michael Advanced Institute for Oral Health, 213 Overlook Circle Suite B4, Brentwood, TN, USA aut Farone Mary Department of Biology, Middle Tennessee State University, Box XO33, Murfreesboro, TN, USA aut DuBois John Department of Biology, Middle Tennessee State University, Box XO33, Murfreesboro, TN, USA aut Newsome Anthony Department of Biology, Middle Tennessee State University, Box XO33, Murfreesboro, TN, USA aut Parasitology Research Springer-Verlag 109/3(2011-09-01), 857-864 0932-0113 109:3<857 2011 109 436 https://doi.org/10.1007/s00436-011-2312-9 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s00436-011-2312-9 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Trim Robert University of Tennessee School of Dentistry, 875 Union Ave, Memphis, TN, USA aut NATIONALLICENCE 700 1- Skinner Michael Advanced Institute for Oral Health, 213 Overlook Circle Suite B4, Brentwood, TN, USA aut NATIONALLICENCE 700 1- Farone Mary Department of Biology, Middle Tennessee State University, Box XO33, Murfreesboro, TN, USA aut NATIONALLICENCE 700 1- DuBois John Department of Biology, Middle Tennessee State University, Box XO33, Murfreesboro, TN, USA aut NATIONALLICENCE 700 1- Newsome Anthony Department of Biology, Middle Tennessee State University, Box XO33, Murfreesboro, TN, USA aut NATIONALLICENCE 773 0- Parasitology Research Springer-Verlag 109/3(2011-09-01), 857-864 0932-0113 109:3<857 2011 109 436 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer