{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,7]],"date-time":"2026-10-07T00:14:52Z","timestamp":1791332092553,"version":"4.3.1"},"reference-count":30,"publisher":"American Society for Microbiology","issue":"13","license":[{"start":{"date-parts":[[2010,7,1]],"date-time":"2010-07-01T00:00:00Z","timestamp":1277942400000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.asm.org\/non-commercial-tdm-license"}],"content-domain":{"domain":["journals.asm.org"],"crossmark-restriction":true},"short-container-title":["Appl Environ Microbiol"],"published-print":{"date-parts":[[2010,7]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:p>\n                    Botulinum neurotoxins are produced by the anaerobic bacterium\n                    <jats:italic>Clostridium botulinum<\/jats:italic>\n                    and are divided into seven distinct serotypes (A to G) known to cause botulism in animals and humans. In this study, a multiplexed quantitative real-time PCR assay for the simultaneous detection of the human pathogenic\n                    <jats:italic>C. botulinum<\/jats:italic>\n                    serotypes A, B, E, and F was developed. Based on the TaqMan chemistry, we used five individual primer-probe sets within one PCR, combining both minor groove binder- and locked nucleic acid-containing probes. Each hydrolysis probe was individually labeled with distinguishable fluorochromes, thus enabling discrimination between the serotypes A, B, E, and F. To avoid false-negative results, we designed an internal amplification control, which was simultaneously amplified with the four target genes, thus yielding a pentaplexed PCR approach with 95% detection probabilities between 7 and 287 genome equivalents per PCR. In addition, we developed six individual singleplex real-time PCR assays based on the TaqMan chemistry for the detection of the\n                    <jats:italic>C. botulinum<\/jats:italic>\n                    serotypes A, B, C, D, E, and F. Upon analysis of 42\n                    <jats:italic>C. botulinum<\/jats:italic>\n                    and 57 non-\n                    <jats:italic>C. botulinum<\/jats:italic>\n                    strains, the singleplex and multiplex PCR assays showed an excellent specificity. Using spiked food samples we were able to detect between 10\n                    <jats:sup>3<\/jats:sup>\n                    and 10\n                    <jats:sup>5<\/jats:sup>\n                    CFU\/ml, respectively. Furthermore, we were able to detect C.\n                    <jats:italic>botulinum<\/jats:italic>\n                    in samples from several cases of botulism in Germany. Overall, the pentaplexed assay showed high sensitivity and specificity and allowed for the simultaneous screening and differentiation of specimens for\n                    <jats:italic>C. botulinum<\/jats:italic>\n                    A, B, E, and F.\n                  <\/jats:p>","DOI":"10.1128\/aem.02490-09","type":"journal-article","created":{"date-parts":[[2010,4,30]],"date-time":"2010-04-30T21:29:02Z","timestamp":1272662942000},"page":"4387-4395","update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":65,"title":["Pentaplexed Quantitative Real-Time PCR Assay for the Simultaneous Detection and Quantification of Botulinum Neurotoxin-Producing Clostridia in Food and Clinical Samples"],"prefix":"10.1128","volume":"76","author":[{"given":"Sebastian","family":"Kirchner","sequence":"first","affiliation":[{"name":"Center for Biological Safety-Microbial Toxins (ZBS3), Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"K. Melanie","family":"Kra\u0308mer","sequence":"additional","affiliation":[{"name":"Center for Biological Safety-Microbial Toxins (ZBS3), Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Martin","family":"Schulze","sequence":"additional","affiliation":[{"name":"Viral Infections (FG12), Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Diana","family":"Pauly","sequence":"additional","affiliation":[{"name":"Center for Biological Safety-Microbial Toxins (ZBS3), Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Daniela","family":"Jacob","sequence":"additional","affiliation":[{"name":"Center for Biological Safety-Highly Pathogenic Microbial Pathogens (ZBS2), Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Frank","family":"Gessler","sequence":"additional","affiliation":[{"name":"Miprolab GmbH, Marie-Curie-Str. 7, 37079 Go\u0308ttingen, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andreas","family":"Nitsche","sequence":"additional","affiliation":[{"name":"Center for Biological Safety-Highly Pathogenic Viruses (ZBS1), Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Brigitte G.","family":"Dorner","sequence":"additional","affiliation":[{"name":"Center for Biological Safety-Microbial Toxins (ZBS3), Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Martin B.","family":"Dorner","sequence":"additional","affiliation":[{"name":"Center for Biological Safety-Microbial Toxins (ZBS3), Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"235","reference":[{"key":"e_1_3_2_2_2","doi-asserted-by":"publisher","DOI":"10.1089\/fpd.2004.1.247"},{"key":"e_1_3_2_3_2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.01192-07"},{"key":"e_1_3_2_4_2","doi-asserted-by":"publisher","DOI":"10.4315\/0362-028X-64.2.201"},{"key":"e_1_3_2_5_2","doi-asserted-by":"publisher","DOI":"10.1373\/clinchem.2008.112797"},{"key":"e_1_3_2_6_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.00784-07"},{"key":"e_1_3_2_7_2","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.00805-09"},{"key":"e_1_3_2_8_2","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.00799-09"},{"key":"e_1_3_2_9_2","doi-asserted-by":"publisher","DOI":"10.1039\/b615227a"},{"key":"e_1_3_2_10_2","doi-asserted-by":"publisher","DOI":"10.1128\/aem.61.1.389-392.1995"},{"key":"e_1_3_2_11_2","doi-asserted-by":"publisher","DOI":"10.1111\/j.1365-2672.2009.04215.x"},{"key":"e_1_3_2_12_2","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.02234-06"},{"key":"e_1_3_2_13_2","doi-asserted-by":"publisher","DOI":"10.1128\/jcm.34.4.882-885.1996"},{"key":"e_1_3_2_14_2","doi-asserted-by":"publisher","DOI":"10.1128\/jcm.32.8.1911-1917.1994"},{"key":"e_1_3_2_15_2","doi-asserted-by":"publisher","DOI":"10.1128\/CMR.3.1.66"},{"key":"e_1_3_2_16_2","doi-asserted-by":"publisher","DOI":"10.1099\/jmm.0.46802-0"},{"key":"e_1_3_2_17_2","doi-asserted-by":"publisher","DOI":"10.1128\/JB.01180-06"},{"key":"e_1_3_2_18_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.88.16.7276"},{"key":"e_1_3_2_19_2","doi-asserted-by":"publisher","DOI":"10.1046\/j.1365-2672.2003.02188.x"},{"key":"e_1_3_2_20_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.mcp.2009.02.003"},{"key":"e_1_3_2_21_2","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.67.1.206-216.2001"},{"key":"e_1_3_2_22_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.mam.2005.12.007"},{"key":"e_1_3_2_23_2","doi-asserted-by":"publisher","DOI":"10.1128\/AEM.67.12.5694-5699.2001"},{"key":"e_1_3_2_24_2","doi-asserted-by":"publisher","DOI":"10.1128\/CMR.19.2.298-314.2006"},{"key":"e_1_3_2_25_2","doi-asserted-by":"publisher","DOI":"10.1007\/s00003-007-0174-y"},{"key":"e_1_3_2_26_2","doi-asserted-by":"publisher","DOI":"10.1016\/j.jviromet.2006.12.007"},{"key":"e_1_3_2_27_2","doi-asserted-by":"publisher","DOI":"10.1016\/S0065-2911(09)05503-9"},{"key":"e_1_3_2_28_2","first-page":"137","volume":"216","year":"2003","unstructured":"Popoff, M. 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