{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,4]],"date-time":"2026-05-04T07:51:15Z","timestamp":1777881075837,"version":"3.51.4"},"reference-count":49,"publisher":"SAGE Publications","license":[{"start":{"date-parts":[[2025,12,1]],"date-time":"2025-12-01T00:00:00Z","timestamp":1764547200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"},{"start":{"date-parts":[[2025,12,1]],"date-time":"2025-12-01T00:00:00Z","timestamp":1764547200000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/journals.sagepub.com\/page\/policies\/text-and-data-mining-license"}],"funder":[{"name":"Thammasat University Research Fund","award":["Contract No. TUFT 048\/2568"],"award-info":[{"award-number":["Contract No. TUFT 048\/2568"]}]}],"content-domain":{"domain":["journals.sagepub.com"],"crossmark-restriction":true},"short-container-title":["Sage Open Medicine"],"published-print":{"date-parts":[[2025,12]]},"abstract":"<jats:sec>\n                    <jats:title>Objectives:<\/jats:title>\n                    <jats:p>\n                      Legionellosis, primarily caused by\n                      <jats:italic toggle=\"yes\">Legionella pneumophila<\/jats:italic>\n                      serogroup 1, remains a public health concern in Thailand, particularly in tourism-driven regions. Conventional culture-based surveillance is hindered by limited sensitivity and prolonged turnaround times. This study aimed to develop and validate an optimized multiplex TaqMan real-time PCR assay for simultaneous detection of\n                      <jats:italic toggle=\"yes\">Legionella<\/jats:italic>\n                      spp.,\n                      <jats:italic toggle=\"yes\">Legionella pneumophila<\/jats:italic>\n                      , and\n                      <jats:italic toggle=\"yes\">Legionella pneumophila<\/jats:italic>\n                      serogroup 1 in environmental water, while also investigating antimicrobial resistance genes.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods:<\/jats:title>\n                    <jats:p>\n                      A modified primer\u2013probe set targeting the\n                      <jats:italic toggle=\"yes\">23S\u20135S rRNA<\/jats:italic>\n                      ,\n                      <jats:italic toggle=\"yes\">mip<\/jats:italic>\n                      , and\n                      <jats:italic toggle=\"yes\">wzm<\/jats:italic>\n                      genes was combined with optimized DNA extraction and PCR conditions. Analytical sensitivity was assessed using colony suspensions and spiked water, and specificity was tested against 24 bacterial strains. The assay was validated on 202 environmental water samples collected from four provinces in southern Thailand. Samples positive by real-time PCR were further screened for antimicrobial resistance genes (\n                      <jats:italic toggle=\"yes\">lpeAB<\/jats:italic>\n                      and\n                      <jats:italic toggle=\"yes\">tet56<\/jats:italic>\n                      ) using conventional PCR.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results:<\/jats:title>\n                    <jats:p>\n                      The optimized assay achieved a detection limit of 1\u2009CFU\/mL in suspensions and 1\u2009CFU\/L in filtered water, with 100% specificity. Among 202 field samples, PCR detected\n                      <jats:italic toggle=\"yes\">Legionella<\/jats:italic>\n                      in 48.51%, significantly higher than culture (24.26%;\n                      <jats:italic toggle=\"yes\">p<\/jats:italic>\n                      \u2009&lt;\u20090.001). Detection was particularly enhanced in cold-water systems from premise plumbing (41.61% versus 9.49%;\n                      <jats:italic toggle=\"yes\">p<\/jats:italic>\n                      \u2009&lt;\u20090.001). Both methods identified\n                      <jats:italic toggle=\"yes\">Legionella pneumophila<\/jats:italic>\n                      serogroup 1 in 14.29% of samples. Spatial mapping identified Patong subdistrict, Phuket, as a contamination hotspot. Antimicrobial resistance surveillance showed\n                      <jats:italic toggle=\"yes\">lpeAB<\/jats:italic>\n                      in 6.1% of samples and\n                      <jats:italic toggle=\"yes\">tet56<\/jats:italic>\n                      in one\n                      <jats:italic toggle=\"yes\">Legionella pneumophila<\/jats:italic>\n                      isolate, representing the first report of antimicrobial resistance in environmental\n                      <jats:italic toggle=\"yes\">Legionella<\/jats:italic>\n                      from Thailand.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions:<\/jats:title>\n                    <jats:p>\n                      This optimized multiplex real-time PCR assay offers a rapid, sensitive, specific, and cost-effective alternative to culture for\n                      <jats:italic toggle=\"yes\">Legionella<\/jats:italic>\n                      surveillance. The detection of antimicrobial resistance genes highlights potential emerging resistance risks, underscoring the need for integrated molecular and resistance monitoring. Implementation of this approach can strengthen outbreak preparedness and improve water safety management in high-risk settings.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1177\/20503121251403893","type":"journal-article","created":{"date-parts":[[2025,12,12]],"date-time":"2025-12-12T06:55:22Z","timestamp":1765522522000},"update-policy":"https:\/\/doi.org\/10.1177\/sage-journals-update-policy","source":"Crossref","is-referenced-by-count":0,"title":["Two-stage surveillance of\n                    <i>Legionella<\/i>\n                    in environmental water: Optimized multiplex TaqMan qPCR for pathogen detection followed by antimicrobial resistance gene identification"],"prefix":"10.1177","volume":"13","author":[{"given":"Watcharee","family":"Thongkhaw","sequence":"first","affiliation":[{"name":"Graduate Program in Medical Technology, Faculty of Allied Health Sciences, Thammasat University, Rangsit Campus, Pathum Thani, Thailand"},{"name":"Department of Medical Sciences, Regional Medical Sciences Center, Phuket, Thailand"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Iriyaporn","family":"Kongthap","sequence":"additional","affiliation":[{"name":"Department of Medical Sciences, Regional Medical Sciences Center, Phuket, Thailand"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Watcharaporn","family":"Kamjumphol","sequence":"additional","affiliation":[{"name":"Department of Medical Sciences, National Institute of Health, Ministry of Public Health, Nonthaburi, Thailand"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hiranya","family":"Sritart","sequence":"additional","affiliation":[{"name":"Department of Medical Technology, Faculty of Allied Health Sciences, Thammasat University, Rangsit Campus, Pathum Thani, Thailand"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5364-6716","authenticated-orcid":false,"given":"Nattamon","family":"Niyomdecha","sequence":"additional","affiliation":[{"name":"Department of Medical Technology, Faculty of Allied Health Sciences, Thammasat University, Rangsit Campus, Pathum Thani, Thailand"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"179","published-online":{"date-parts":[[2025,12,11]]},"reference":[{"key":"e_1_3_3_2_2","unstructured":"Department of Disease Control Ministry of Public Health Thailand. 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