{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,27]],"date-time":"2026-08-27T23:54:42Z","timestamp":1787874882883,"version":"build-2784847793"},"reference-count":58,"publisher":"Oxford University Press (OUP)","issue":"9","license":[{"start":{"date-parts":[[2021,12,1]],"date-time":"2021-12-01T00:00:00Z","timestamp":1638316800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/academic.oup.com\/journals\/pages\/open_access\/funder_policies\/chorus\/standard_publication_model"}],"funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["AI146454"],"award-info":[{"award-number":["AI146454"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2022,1,12]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>Mycoplasma contamination of cell culture represents a serious problem in research and decontamination from cell-propagated obligate intracellular bacteria has proven challenging. Here, we presented an optimized protocol to remove Mycoplasma from contaminated Chlamydia trachomatis culture. A stepwise procedure of Mycoplasma removal entails (i) incubation in nonionic detergent-containing solution and (ii) separation of viable chlamydial organisms by fluorescence-activated cell sorting (FACS), followed by subcloning using a focus-forming assay. We also adapted a polymerase chain reaction (PCR) assay using paired universal and Mycoplasma-specific primers, which are distinguishable from the C. trachomatis counterparts, in combination with Sanger sequencing to determine the presence of mycoplasmas\u2019 16S rRNA genes. These integrated approaches allow for full removal of Mycoplasma, as verified by the improved PCR assay, without compromising the capacity of viable C. trachomatis to adapt to new infection in epithelial cells. Some pitfalls during the Mycoplasma decontamination process are discussed.<\/jats:p>","DOI":"10.1093\/femspd\/ftab056","type":"journal-article","created":{"date-parts":[[2021,12,14]],"date-time":"2021-12-14T20:14:58Z","timestamp":1639512898000},"source":"Crossref","is-referenced-by-count":10,"title":["<i>Mycoplasma<\/i>decontamination in<i>Chlamydia trachomatis<\/i>culture: a curative approach"],"prefix":"10.1093","volume":"79","author":[{"given":"Madison","family":"Greer","sequence":"first","affiliation":[{"name":"Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA 70112, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jacob H","family":"Elnaggar","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA 70112, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christopher M","family":"Taylor","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA 70112, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9869-7317","authenticated-orcid":false,"given":"Li","family":"Shen","sequence":"additional","affiliation":[{"name":"Department of Microbiology, Immunology, and Parasitology, Louisiana State University Health Sciences Center, New Orleans, LA 70112, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2021,12,16]]},"reference":[{"key":"2022011513582285900_bib1","doi-asserted-by":"crossref","first-page":"201","DOI":"10.1016\/j.mimet.2006.07.012","article-title":"Clonal isolation of Chlamydia-infected cells using flow cytometry","volume":"68","author":"Alzhanov","year":"2007","journal-title":"J Microbiol Methods"},{"key":"2022011513582285900_bib2","doi-asserted-by":"crossref","first-page":"736","DOI":"10.1111\/1523-1747.ep12470407","article-title":"Effect of mupirocin on the growth and lifespan of human fibroblasts","volume":"88","author":"Balin","year":"1987","journal-title":"J Invest Dermatol"},{"key":"2022011513582285900_bib3","doi-asserted-by":"crossref","first-page":"149","DOI":"10.1038\/nri1551","article-title":"Immunology of Chlamydiainfection: implications for a Chlamydia trachomatis vaccine","volume":"5","author":"Brunham","year":"2005","journal-title":"Nat Rev Immunol"},{"key":"2022011513582285900_bib4","doi-asserted-by":"crossref","first-page":"8","DOI":"10.1016\/j.jri.2011.08.002","article-title":"Innate immune mediator profiles and their regulation in a novel polarized immortalized epithelial cell model derived from human endocervix","volume":"92","author":"Buckner","year":"2011","journal-title":"J Reprod Immunol"},{"key":"2022011513582285900_bib5","doi-asserted-by":"crossref","first-page":"1161","DOI":"10.1128\/iai.31.3.1161-1176.1981","article-title":"Purification and partial characterization of the major outer membrane protein of Chlamydiatrachomatis","volume":"31","author":"Caldwell","year":"1981","journal-title":"Infect Immun"},{"key":"2022011513582285900_bib6","doi-asserted-by":"crossref","first-page":"861","DOI":"10.1128\/JCM.38.2.861-862.2000","article-title":"Effect of a Mycoplasmahominis-like mycoplasma on the infection of HEp-2 cells by the TW-183 strain of Chlamydiapneumoniae","volume":"38","author":"Castilla","year":"2000","journal-title":"J Clin Microbiol"},{"key":"2022011513582285900_bib7","doi-asserted-by":"crossref","first-page":"330","DOI":"10.1016\/j.mimet.2007.02.005","article-title":"A detailed analysis of 16S ribosomal RNA gene segments for the diagnosis of pathogenic bacteria","volume":"69","author":"Chakravorty","year":"2007","journal-title":"J Microbiol Methods"},{"key":"2022011513582285900_bib8","doi-asserted-by":"crossref","first-page":"41","DOI":"10.32607\/20758251-2014-6-3-41-51","article-title":"Mycoplasma contamination of cell cultures: vesicular traffic in bacteria and control over infectious agents","volume":"6","author":"Chernov","year":"2014","journal-title":"Acta Naturae"},{"key":"2022011513582285900_bib9","doi-asserted-by":"crossref","first-page":"1682","DOI":"10.3389\/fmicb.2018.01682","article-title":"The role of lipoproteins in mycoplasma-mediated immunomodulation","volume":"9","author":"Christodoulides","year":"2018","journal-title":"Front Microbiol"},{"key":"2022011513582285900_bib10","doi-asserted-by":"crossref","first-page":"27244","DOI":"10.1038\/srep27244","article-title":"Quantifying promoter activity during the developmental cycle of Chlamydiatrachomatis","volume":"6","author":"Cong","year":"2016","journal-title":"Sci Rep"},{"key":"2022011513582285900_bib11","doi-asserted-by":"crossref","DOI":"10.1007\/0-306-47606-1_11","article-title":"Comparative genome analysis of Mollicutes","volume-title":"Molecular Biology and Pathogenicity of Mycoplasmas","author":"Dandekar","year":"2002"},{"key":"2022011513582285900_bib12","doi-asserted-by":"crossref","first-page":"75","DOI":"10.1023\/A:1022913015916","article-title":"Mycoplasma contamination of cell cultures: incidence, sources, effects, detection, elimination, prevention","volume":"39","author":"Drexler","year":"2002","journal-title":"Cytotechnology"},{"key":"2022011513582285900_bib14","doi-asserted-by":"crossref","first-page":"73","DOI":"10.3389\/fcimb.2014.00073","article-title":"Membrane vesicle production by Chlamydiatrachomatis as an adaptive response","volume":"4","author":"Frohlich","year":"2014","journal-title":"Front Cell Infect Microbiol"},{"key":"2022011513582285900_bib13","doi-asserted-by":"crossref","first-page":"222","DOI":"10.1016\/j.mimet.2012.08.012","article-title":"Isolation of Chlamydiatrachomatis and membrane vesicles derived from host and bacteria","volume":"91","author":"Frohlich","year":"2012","journal-title":"J Microbiol Methods"},{"key":"2022011513582285900_bib15","doi-asserted-by":"crossref","first-page":"5827","DOI":"10.1128\/IAI.70.10.5827-5834.2002","article-title":"Isolation of Chlamydia pneumoniae clonal variants by a focus-forming assay","volume":"70","author":"Gieffers","year":"2002","journal-title":"Infect Immun"},{"key":"2022011513582285900_bib16","doi-asserted-by":"crossref","first-page":"3819","DOI":"10.1128\/JB.00511-13","article-title":"Characterization of Chlamydiatrachomatis plasmid-encoded open reading frames","volume":"195","author":"Gong","year":"2013","journal-title":"J Bacteriol"},{"key":"2022011513582285900_bib17","doi-asserted-by":"crossref","first-page":"25","DOI":"10.1128\/jb.161.1.25-31.1985","article-title":"Disulfide-mediated interactions of the chlamydial major outer membrane protein: role in the differentiation of chlamydiae?","volume":"161","author":"Hackstadt","year":"1985","journal-title":"J Bacteriol"},{"key":"2022011513582285900_bib18","doi-asserted-by":"crossref","first-page":"1115","DOI":"10.1099\/00222615-47-12-1115","article-title":"Comparative susceptibilities of various aids-associated and human urogenital tract mycoplasmas and strains of Mycoplasma pneumoniae to 10 classes of antimicrobial agent in vitro","volume":"47","author":"Hannan","year":"1998","journal-title":"J Med Microbiol"},{"key":"2022011513582285900_bib19","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1128\/jb.178.1.1-5.1996","article-title":"Disulfide cross-linked envelope proteins: the functional equivalent of peptidoglycan in chlamydiae?","volume":"178","author":"Hatch","year":"1996","journal-title":"J Bacteriol"},{"key":"2022011513582285900_bib20","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1093\/femspd\/ftu022","article-title":"Andrographolide inhibits intracellular Chlamydiatrachomatis multiplication and reduces CXCL-8 secretion produced by human epithelial cells","volume":"73","author":"Hua","year":"2015","journal-title":"Pathog Dis"},{"key":"2022011513582285900_bib21","doi-asserted-by":"crossref","first-page":"4765","DOI":"10.1128\/JB.180.18.4765-4774.1998","article-title":"Impact of culture-independent studies on the emerging phylogenetic view of bacterial diversity","volume":"180","author":"Hugenholtz","year":"1998","journal-title":"J Bacteriol"},{"key":"2022011513582285900_bib22","doi-asserted-by":"crossref","first-page":"e0172358","DOI":"10.1371\/journal.pone.0172358","article-title":"Assessing mycoplasma contamination of cell cultures by qPCR using a set of universal primer pairs targeting a 1.5\u00a0kb fragment of 16S rRNA genes","volume":"12","author":"Jean","year":"2017","journal-title":"PLoS One"},{"key":"2022011513582285900_bib23","doi-asserted-by":"crossref","first-page":"261","DOI":"10.1128\/JCM.41.1.261-266.2003","article-title":"Detection ofMycoplasma genitalium by PCR amplification of the 16S rRNA gene","volume":"41","author":"Jensen","year":"2003","journal-title":"J Clin Microbiol"},{"key":"2022011513582285900_bib24","doi-asserted-by":"crossref","first-page":"145","DOI":"10.1385\/0-89603-525-5:145","article-title":"Characterization of mycoplasmas by PCR and sequence analysis with universal 16S rDNA primers","volume-title":"Mycoplasma Protocols","author":"Johansson","year":"1998"},{"key":"2022011513582285900_bib26","doi-asserted-by":"crossref","first-page":"19","DOI":"10.1007\/PL00008253","article-title":"Frequent contamination of Chlamydia trachomatis and Chlamydia pneumoniae strains with mycoplasma. Biological relevance and selective eradication of mycoplasma from chlamydial cultures with mupirocin","volume":"189","author":"Krau\u00dfe-Opatz","year":"2000","journal-title":"Med Microbiol Immunol"},{"key":"2022011513582285900_bib25","doi-asserted-by":"crossref","first-page":"3910","DOI":"10.1128\/JCM.38.10.3910-3911.2000","article-title":"Mycoplasma fermentans, M. hominis, and M. hyorhinis inhibit infectivity and growth of Chlamydiatrachomatis and C. pneumoniae in HEp-2 cells","volume":"38","author":"Krau\u00dfe-Opatz","year":"2000","journal-title":"J Clin Microbiol"},{"key":"2022011513582285900_bib27","doi-asserted-by":"crossref","first-page":"208","DOI":"10.1128\/CMR.00110-14","article-title":"Current and past strategies for bacterial culture in clinical microbiology","volume":"28","author":"Lagier","year":"2015","journal-title":"Clin Microbiol Rev"},{"key":"2022011513582285900_bib28","doi-asserted-by":"crossref","first-page":"1150","DOI":"10.1093\/jac\/dkab016","article-title":"Identification of 16S rRNA mutations in Mycoplasma genitalium potentially associated with tetracycline resistance in vivo but not selectedin vitro in M. genitalium and Chlamydia trachomatis","volume":"76","author":"Le\u00a0Roy","year":"2021","journal-title":"J Antimicrob Chemother"},{"key":"2022011513582285900_bib29","doi-asserted-by":"crossref","first-page":"3681","DOI":"10.1128\/JCM.01092-11","article-title":"Sequence homologies between Mycoplasma and Chlamydia spp. lead to false-positive results in chlamydial cell cultures tested for mycoplasma contamination with a commercial PCR assay","volume":"49","author":"Maass","year":"2011","journal-title":"J Clin Microbiol"},{"key":"2022011513582285900_bib30","doi-asserted-by":"crossref","first-page":"W636","DOI":"10.1093\/nar\/gkz268","article-title":"The EMBL-EBI search and sequence analysis tools APIs in 2019","volume":"47","author":"Madeira","year":"2019","journal-title":"Nucleic Acids Res"},{"key":"2022011513582285900_bib31","doi-asserted-by":"crossref","first-page":"S802","DOI":"10.1093\/cid\/civ785","article-title":"Efficacy of antimicrobial therapy for Mycoplasma genitalium infections","volume":"61","author":"Manhart","year":"2015","journal-title":"Clin Infect Dis"},{"key":"2022011513582285900_bib32","doi-asserted-by":"crossref","first-page":"3013","DOI":"10.1128\/JCM.36.10.3013-3019.1998","article-title":"Plaque formation by and plaque cloning of Chlamydia trachomatis biovar trachoma","volume":"36","author":"Matsumoto","year":"1998","journal-title":"J Clin Microbiol"},{"key":"2022011513582285900_bib33","doi-asserted-by":"crossref","first-page":"e00105","DOI":"10.1128\/IAI.00105-20","article-title":"Human fallopian tube epithelial cell culture model to study host responses to Chlamydia trachomatis infection","volume":"88","author":"McQueen","year":"2020","journal-title":"Infect Immun"},{"key":"2022011513582285900_bib34","doi-asserted-by":"crossref","first-page":"695","DOI":"10.1128\/aem.59.3.695-700.1993","article-title":"Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes coding for 16S rRNA","volume":"59","author":"Muyzer","year":"1993","journal-title":"Appl Environ Microbiol"},{"key":"2022011513582285900_bib35","first-page":"203","article-title":"Prevention and detection of mycoplasma contamination in cell culture","volume":"13","author":"Nikfarjam","year":"2012","journal-title":"Cell J"},{"key":"2022011513582285900_bib38","doi-asserted-by":"crossref","first-page":"1044","DOI":"10.1128\/IAI.01118-10","article-title":"Toll-like receptor 2 activation by Chlamydia trachomatis is plasmid dependent, and plasmid-responsive chromosomal loci are coordinately regulated in response to glucose limitation by C. trachomatis but not by C. muridarum","volume":"79","author":"O'Connell","year":"2011","journal-title":"Infect Immun"},{"key":"2022011513582285900_bib37","doi-asserted-by":"crossref","first-page":"4027","DOI":"10.4049\/jimmunol.179.6.4027","article-title":"Plasmid-deficient Chlamydia muridarum fail to induce immune pathology and protect against oviduct disease","volume":"179","author":"O'Connell","year":"2007","journal-title":"J Immunol"},{"key":"2022011513582285900_bib36","doi-asserted-by":"crossref","first-page":"1652","DOI":"10.1074\/jbc.M510182200","article-title":"Localization of TLR2 and myd88 to Chlamydia trachomatis inclusions. Evidence for signaling by intracellular TLR2 during infection with an obligate intracellular pathogen","volume":"281","author":"O'Connell","year":"2006","journal-title":"J Biol Chem"},{"key":"2022011513582285900_bib39","doi-asserted-by":"crossref","first-page":"2535","DOI":"10.1093\/nar\/gkv136","article-title":"Assessing the prevalence of mycoplasma contamination in cell culture via a survey of NCBI's RNA-seq archive","volume":"43","author":"Olarerin-George","year":"2015","journal-title":"Nucleic Acids Res"},{"key":"2022011513582285900_bib41","doi-asserted-by":"crossref","first-page":"328","DOI":"10.1128\/aem.62.2.328-331.1996","article-title":"Application of a mycoplasma group-specific PCR for monitoring decontamination of mycoplasma-infected Chlamydia sp. strains","volume":"62","author":"Ossewaarde","year":"1996","journal-title":"Appl Environ Microbiol"},{"key":"2022011513582285900_bib42","doi-asserted-by":"crossref","first-page":"72","DOI":"10.1186\/s12915-019-0690-0","article-title":"A systematic sequencing-based approach for microbial contaminant detection and functional inference","volume":"17","author":"Park","year":"2019","journal-title":"BMC Biol"},{"key":"2022011513582285900_bib43","doi-asserted-by":"crossref","first-page":"534","DOI":"10.1128\/IAI.02764-14","article-title":"Transcriptional profiling of human epithelial cells infected with plasmid-bearing and plasmid-deficient Chlamydia trachomatis","volume":"83","author":"Porcella","year":"2015","journal-title":"Infect Immun"},{"key":"2022011513582285900_bib44","doi-asserted-by":"crossref","first-page":"77","DOI":"10.1172\/JCI119136","article-title":"Secretion of proinflammatory cytokines by epithelial cells in response to Chlamydia infection suggests a central role for epithelial cells in chlamydial pathogenesis","volume":"99","author":"Rasmussen","year":"1997","journal-title":"J Clin Invest"},{"key":"2022011513582285900_bib45","doi-asserted-by":"crossref","first-page":"217","DOI":"10.1016\/0014-5793(69)80140-7","article-title":"Solubilization of mycoplasma membranes by the nonionic detergent triton X-100","volume":"3","author":"Razin","year":"1969","journal-title":"FEBS Lett"},{"key":"2022011513582285900_bib46","doi-asserted-by":"crossref","first-page":"609","DOI":"10.1128\/jb.91.2.609-616.1966","article-title":"Influence of lipid components of Mycoplasma laidlawii membranes on osmotic fragility of cells","volume":"91","author":"Razin","year":"1966","journal-title":"J Bacteriol"},{"key":"2022011513582285900_bib47","doi-asserted-by":"crossref","first-page":"491","DOI":"10.1177\/2472555220911451","article-title":"Keeping it clean: the cell culture quality control experience at the national center for advancing translational sciences","volume":"25","author":"Roth","year":"2020","journal-title":"SLAS Discov"},{"key":"2022011513582285900_bib48","doi-asserted-by":"crossref","first-page":"417","DOI":"10.1152\/physrev.00030.2002","article-title":"Interaction of mycoplasmas with host cells","volume":"83","author":"Rottem","year":"2003","journal-title":"Physiol Rev"},{"key":"2022011513582285900_bib49","doi-asserted-by":"crossref","first-page":"165","DOI":"10.3389\/fcimb.2017.00165","article-title":"Manipulation of host cholesterol by obligate intracellular bacteria","volume":"7","author":"Samanta","year":"2017","journal-title":"Front Cell Infect Microbiol"},{"key":"2022011513582285900_bib50","first-page":"11A.1.1","article-title":"Cultivation and laboratory maintenance of Chlamydia trachomatis","author":"Scidmore","year":"2006","journal-title":"Curr Protoc Microbiol"},{"key":"2022011513582285900_bib51","doi-asserted-by":"crossref","first-page":"361","DOI":"10.1186\/s13104-018-3455-x","article-title":"Elimination of mycoplasma contamination in Chlamydiastocks as a result of in vivo passage or plaque isolation","volume":"11","author":"Sigar","year":"2018","journal-title":"BMC Res Notes"},{"key":"2022011513582285900_bib52","doi-asserted-by":"crossref","first-page":"636","DOI":"10.1128\/IAI.01305-12","article-title":"Chlamydia trachomatis plasmid-encoded pgp4 is a transcriptional regulator of virulence-associated genes","volume":"81","author":"Song","year":"2013","journal-title":"Infect Immun"},{"key":"2022011513582285900_bib53","doi-asserted-by":"crossref","first-page":"58","DOI":"10.1016\/j.biologicals.2019.12.007","article-title":"A highly sensitive internally-controlled real-time PCR assay for mycoplasma detection in cell cultures","volume":"64","author":"Sung","year":"2020","journal-title":"Biologicals"},{"key":"2022011513582285900_bib54","doi-asserted-by":"crossref","first-page":"2759","DOI":"10.1099\/mic.0.044917-0","article-title":"Altered protein secretion of Chlamydiatrachomatis in persistently infected human endocervical epithelial cells","volume":"157","author":"Wang","year":"2011","journal-title":"Microbiology"},{"key":"2022011513582285900_bib55","doi-asserted-by":"crossref","first-page":"5261","DOI":"10.1128\/AEM.00062-07","article-title":"Naive Bayesian classifier for rapid assignment of rRNA sequences into the new bacterial taxonomy","volume":"73","author":"Wang","year":"2007","journal-title":"Appl Environ Microbiol"},{"key":"2022011513582285900_bib56","doi-asserted-by":"crossref","first-page":"e1002258","DOI":"10.1371\/journal.ppat.1002258","article-title":"Development of a transformation system for Chlamydiatrachomatis: restoration of glycogen biosynthesis by acquisition of a plasmid shuttle vector","volume":"7","author":"Wang","year":"2011","journal-title":"PLoS Pathog"},{"key":"2022011513582285900_bib57","doi-asserted-by":"crossref","first-page":"697","DOI":"10.1128\/jb.173.2.697-703.1991","article-title":"16S ribosomal DNA amplification for phylogenetic study","volume":"173","author":"Weisburg","year":"1991","journal-title":"J Bacteriol"},{"key":"2022011513582285900_bib40","volume-title":"Sexually Transmitted Infections (STIs), Fact Sheet","author":"World Health Organization","year":"2016"},{"key":"2022011513582285900_bib58","doi-asserted-by":"crossref","first-page":"141","DOI":"10.1016\/j.tim.2016.09.006","article-title":"Chlamydial plasmid-dependent pathogenicity","volume":"25","author":"Zhong","year":"2017","journal-title":"Trends Microbiol"}],"container-title":["Pathogens and Disease"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/academic.oup.com\/femspd\/advance-article-pdf\/doi\/10.1093\/femspd\/ftab056\/41787347\/ftab056.pdf","content-type":"application\/pdf","content-version":"am","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/femspd\/article-pdf\/79\/9\/ftab056\/42198770\/ftab056.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/femspd\/article-pdf\/79\/9\/ftab056\/42198770\/ftab056.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,1,18]],"date-time":"2023-01-18T13:05:56Z","timestamp":1674047156000},"score":1,"resource":{"primary":{"URL":"https:\/\/academic.oup.com\/femspd\/article\/doi\/10.1093\/femspd\/ftab056\/6464140"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,12]]},"references-count":58,"journal-issue":{"issue":"9","published-print":{"date-parts":[[2022,1,12]]}},"URL":"https:\/\/doi.org\/10.1093\/femspd\/ftab056","relation":{},"ISSN":["2049-632X"],"issn-type":[{"value":"2049-632X","type":"electronic"}],"subject":[],"published-other":{"date-parts":[[2021,12]]},"published":{"date-parts":[[2021,12]]},"article-number":"ftab056"}}