{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,1,8]],"date-time":"2026-01-08T08:34:13Z","timestamp":1767861253330,"version":"3.49.0"},"reference-count":58,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2023,5,5]],"date-time":"2023-05-05T00:00:00Z","timestamp":1683244800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Microbiol."],"abstract":"<jats:sec><jats:title>Introduction<\/jats:title><jats:p><jats:italic>Balamuthia<\/jats:italic> (<jats:italic>B.<\/jats:italic>) <jats:italic>mandrillaris<\/jats:italic> is a free-living amoeba that can cause rare yet fatal granulomatous amoebic encephalitis (GAE). However, efficacious treatment for GAE is currently unavailable, especially when genomic studies on <jats:italic>B. mandrillaris<\/jats:italic> are limited.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>In this study, <jats:italic>B. mandrillaris<\/jats:italic> strain KM-20 was isolated from the brain tissue of a GAE patient, and its mitochondrial genome was <jats:italic>de novo<\/jats:italic> assembled using high-coverage Nanopore long reads and Illumina short reads.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results and Discussion<\/jats:title><jats:p>Phylogenetic and comparative analyses revealed a range of diversification in the mitochondrial genome of KM-20 and nine other <jats:italic>B. mandrillaris<\/jats:italic> strains. According to the mitochondrial genome alignment, one of the most variable regions was observed in the ribosomal protein S3 (<jats:italic>rps3<\/jats:italic>), which was caused by an array of novel protein tandem repeats. The repeating units in the <jats:italic>rps3<\/jats:italic> protein tandem region present significant copy number variations (CNVs) among <jats:italic>B. mandrillaris<\/jats:italic> strains and suggest KM-20 as the most divergent strain for its highly variable sequence and highest copy number in <jats:italic>rps3<\/jats:italic>. Moreover, mitochondrial heteroplasmy was observed in strain V039, and two genotypes of <jats:italic>rps3<\/jats:italic> are caused by the CNVs in the tandem repeats. Taken together, the copy number and sequence variations of the protein tandem repeats enable <jats:italic>rps3<\/jats:italic> to be a perfect target for clinical genotyping assay for <jats:italic>B. mandrillaris<\/jats:italic>. The mitochondrial genome diversity of <jats:italic>B. mandrillaris<\/jats:italic> paves the way to investigate the phylogeny and diversification of pathogenic amoebae.<\/jats:p><\/jats:sec>","DOI":"10.3389\/fmicb.2023.1162963","type":"journal-article","created":{"date-parts":[[2023,5,5]],"date-time":"2023-05-05T06:10:56Z","timestamp":1683267056000},"update-policy":"https:\/\/doi.org\/10.3389\/crossmark-policy","source":"Crossref","is-referenced-by-count":4,"title":["Mitochondrial genome diversity of Balamuthia mandrillaris revealed by a fatal case of granulomatous amoebic encephalitis"],"prefix":"10.3389","volume":"14","author":[{"given":"Cherie Tsz-Yiu","family":"Law","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Thirapa","family":"Nivesvivat","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Qing","family":"Xiong","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kasem","family":"Kulkeaw","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ling","family":"Shi","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Pichet","family":"Ruenchit","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Detchvijitr","family":"Suwanpakdee","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Piradee","family":"Suwanpakdee","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nongnat","family":"Tongkrajang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Patsharaporn T.","family":"Sarasombath","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Stephen Kwok-Wing","family":"Tsui","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1965","published-online":{"date-parts":[[2023,5,5]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","first-page":"e116","DOI":"10.7717\/peerj-cs.116","article-title":"AliTV\u2014interactive visualization of whole genome comparisons","volume":"3","author":"Ankenbrand","year":"2017","journal-title":"PeerJ Comput. Sci."},{"key":"ref2","doi-asserted-by":"publisher","first-page":"447","DOI":"10.1097\/01.inf.0000066540.18671.f8","article-title":"Amebic encephalitis caused by Balamuthia mandrillaris: report of four cases","volume":"22","author":"Bakardjiev","year":"2003","journal-title":"Pediatr. Infect. Dis. J."},{"key":"ref3","doi-asserted-by":"publisher","first-page":"313","DOI":"10.1016\/j.ympev.2012.08.023","article-title":"MITOS: improved de novo metazoan mitochondrial genome annotation","volume":"69","author":"Bernt","year":"2013","journal-title":"Mol. Phylogenet. Evol."},{"key":"ref4","doi-asserted-by":"publisher","first-page":"D344","DOI":"10.1093\/nar\/gkaa977","article-title":"The InterPro protein families and domains database: 20 years on","volume":"49","author":"Blum","year":"2021","journal-title":"Nucleic Acids Res."},{"key":"ref5","doi-asserted-by":"publisher","first-page":"453","DOI":"10.1128\/JCM.41.1.453-455.2003","article-title":"Identification of Balamuthia mandrillaris by PCR assay using the mitochondrial 16S rRNA Gene as a target","volume":"41","author":"Booton","year":"2003","journal-title":"J. Clin. Microbiol."},{"key":"ref6","doi-asserted-by":"publisher","first-page":"391","DOI":"10.1007\/s11908-012-0266-4","article-title":"Balamuthia Mandrillaris amoebic encephalitis: an emerging parasitic infection","volume":"14","author":"Bravo","year":"2012","journal-title":"Curr. Infect. Dis. Rep."},{"key":"ref7","doi-asserted-by":"publisher","first-page":"6192","DOI":"10.1128\/jb.176.20.6192-6198.1994","article-title":"Ribosomal protein gene sequence changes in erythromycin-resistant mutants of Escherichia coli","volume":"176","author":"Chittum","year":"1994","journal-title":"J. Bacteriol."},{"key":"ref8","doi-asserted-by":"publisher","first-page":"1815","DOI":"10.1093\/cid\/ciy813","article-title":"The epidemiology and clinical features of Balamuthia mandrillaris disease in the United States, 1974\u20132016","volume":"68","author":"Cope","year":"2019","journal-title":"Clin. Infect. Dis."},{"key":"ref9","doi-asserted-by":"publisher","first-page":"1188","DOI":"10.1101\/gr.849004","article-title":"WebLogo: a sequence logo generator","volume":"14","author":"Crooks","year":"2004","journal-title":"Genome Res."},{"key":"ref10","doi-asserted-by":"publisher","first-page":"251406","DOI":"10.1155\/2009\/251406","article-title":"Diagnosis of infections caused by pathogenic free-living amoebae","volume":"2009","author":"da Rocha-Azevedo","year":"2009","journal-title":"Interdiscip. Perspect. Infect. Dis."},{"key":"ref11","doi-asserted-by":"publisher","first-page":"giab008","DOI":"10.1093\/gigascience\/giab008","article-title":"Twelve years of SAMtools and BCFtools","volume":"10","author":"Danecek","year":"2021","journal-title":"Gigascience"},{"key":"ref12","doi-asserted-by":"publisher","first-page":"2069","DOI":"10.1016\/j.biocel.2009.04.007","article-title":"Particularities of mitochondrial structure in parasitic protists (Apicomplexa and Kinetoplastida)","volume":"41","author":"de Souza","year":"2009","journal-title":"Int. J. Biochem. Cell Biol."},{"key":"ref13","doi-asserted-by":"publisher","first-page":"e01013","DOI":"10.1128\/genomeA.01013-15","article-title":"First draft genome sequence of Balamuthia mandrillaris, the causative agent of amoebic encephalitis","volume":"3","author":"Detering","year":"2015","journal-title":"Genome Announc."},{"key":"ref14","doi-asserted-by":"publisher","first-page":"2837","DOI":"10.1099\/mic.0.27218-0","article-title":"Balamuthia mandrillaris from soil samples","volume":"150","author":"Dunnebacke","year":"2004","journal-title":"Microbiology (Society for General Microbiology)"},{"key":"ref15","doi-asserted-by":"publisher","first-page":"W297","DOI":"10.1093\/nar\/gkab408","article-title":"IUPred3: prediction of protein disorder enhanced with unambiguous experimental annotation and visualization of evolutionary conservation","volume":"49","author":"Erd\u0151s","year":"2021","journal-title":"Nucleic Acids Res."},{"key":"ref16","doi-asserted-by":"publisher","first-page":"1239","DOI":"10.1111\/j.1365-2958.1993.tb01253.x","article-title":"Molecular basis of streptomycin resistance in Mycobacterium tuberculosis: alterations of the ribosomal protein S12 gene and point mutations within a functional 16S ribosomal RNA pseudoknot","volume":"9","author":"Finken","year":"1993","journal-title":"Mol. Microbiol."},{"key":"ref17","doi-asserted-by":"publisher","first-page":"1024","DOI":"10.1093\/clind\/15.6.1024","article-title":"Culture isolation of Acanthamoeba species and Leptomyxid Amebas from patients with amebic meningoencephalitis, including two patients with AIDS","volume":"15","author":"Gordon","year":"1992","journal-title":"Clin. Infect. Dis."},{"key":"ref18","doi-asserted-by":"publisher","first-page":"W181","DOI":"10.1093\/nar\/gkn179","article-title":"CGView server: a comparative genomics tool for circular genomes","volume":"36","author":"Grant","year":"2008","journal-title":"Nucleic Acids Res."},{"key":"ref19","doi-asserted-by":"publisher","first-page":"865","DOI":"10.1093\/nar\/26.4.865","article-title":"Genome structure and gene content in protist mitochondrial DNAs","volume":"26","author":"Gray","year":"1998","journal-title":"Nucleic Acids Res."},{"key":"ref20","doi-asserted-by":"publisher","first-page":"113","DOI":"10.1186\/s13073-015-0235-2","article-title":"Clinical metagenomic identification of Balamuthia mandrillaris encephalitis and assembly of the draft genome: the continuing case for reference genome sequencing","volume":"7","author":"Greninger","year":"2015","journal-title":"Genome Med."},{"key":"ref21","doi-asserted-by":"publisher","first-page":"1857","DOI":"10.1093\/nar\/26.8.1857","article-title":"The protein splicing domain of the homing endonuclease PI-SceI is responsible for specific DNA binding","volume":"26","author":"Grindl","year":"1998","journal-title":"Nucleic Acids Res."},{"key":"ref22","volume-title":"Improved pairwise alignment of genomic DNA","author":"Harris","year":"2007"},{"key":"ref23","doi-asserted-by":"publisher","first-page":"159","DOI":"10.1186\/1471-2148-7-159","article-title":"Cyanobacterial ribosomal RNA genes with multiple, endonuclease-encoding group I introns","volume":"7","author":"Haugen","year":"2007","journal-title":"BMC Evol. Biol."},{"key":"ref24","doi-asserted-by":"publisher","first-page":"1440","DOI":"10.1093\/molbev\/msn088","article-title":"Mitochondrial genome evolution in the social amoebae","volume":"25","author":"Heidel","year":"2008","journal-title":"Mol. Biol. Evol."},{"key":"ref25","doi-asserted-by":"publisher","first-page":"666","DOI":"10.4269\/ajtmh.2004.70.666","article-title":"Balamuthia Mandrillaris meningoencephalitis: the first case in Southeast Asia","volume":"70","author":"Intalapaporn","year":"2004","journal-title":"Am. J. Trop. Med. Hyg."},{"key":"ref26","doi-asserted-by":"publisher","first-page":"622","DOI":"10.1016\/j.sbi.2013.06.011","article-title":"Tandem-repeat proteins: regularity plus modularity equals design-ability","volume":"23","author":"Javadi","year":"2013","journal-title":"Curr. Opin. Struct. Biol."},{"key":"ref27","doi-asserted-by":"publisher","first-page":"e732","DOI":"10.7717\/peerj.732","article-title":"Tandem-repeat protein domains across the tree of life","volume":"3","author":"Jernigan","year":"2015","journal-title":"PeerJ (San Francisco, CA)"},{"key":"ref28","doi-asserted-by":"publisher","first-page":"1236","DOI":"10.1093\/bioinformatics\/btu031","article-title":"InterProScan 5: genome-scale protein function classification","volume":"30","author":"Jones","year":"2014","journal-title":"Bioinformatics"},{"key":"ref29","doi-asserted-by":"publisher","first-page":"275","DOI":"10.1093\/bioinformatics\/8.3.275","article-title":"The rapid generation of mutation data matrices from protein sequences","volume":"8","author":"Jones","year":"1992","journal-title":"Bioinformatics"},{"key":"ref30","doi-asserted-by":"publisher","first-page":"2943","DOI":"10.1016\/j.bbamcr.2013.07.015","article-title":"Cytoplasmic ribosomal protein S3 (rpS3) plays a pivotal role in mitochondrial DNA damage surveillance","volume":"1833","author":"Kim","year":"2013","journal-title":"Biochim. Biophys. Acta Mol. Cell Res."},{"key":"ref32","doi-asserted-by":"publisher","first-page":"540","DOI":"10.1038\/s41587-019-0072-8","article-title":"Assembly of long, error-prone reads using repeat graphs","volume":"37","author":"Kolmogorov","year":"2019","journal-title":"Nat. Biotechnol."},{"key":"ref34","doi-asserted-by":"publisher","first-page":"1547","DOI":"10.1093\/molbev\/msy096","article-title":"MEGA X: molecular evolutionary genetics analysis across computing platforms","volume":"35","author":"Kumar","year":"2018","journal-title":"Mol. Biol. Evol."},{"key":"ref35","doi-asserted-by":"publisher","first-page":"W22","DOI":"10.1093\/nar\/gkt389","article-title":"aLeaves facilitates on-demand exploration of metazoan gene family trees on MAFFT sequence alignment server with enhanced interactivity","volume":"41","author":"Kuraku","year":"2013","journal-title":"Nucleic Acids Res."},{"key":"ref36","doi-asserted-by":"publisher","first-page":"119","DOI":"10.1016\/j.tig.2007.01.006","article-title":"Mitochondrial introns: a critical view","volume":"23","author":"Lang","year":"2007","journal-title":"Trends Genet."},{"key":"ref37","doi-asserted-by":"publisher","first-page":"11","DOI":"10.1093\/nar\/gkh152","article-title":"ARAGORN, a program to detect tRNA genes and tmRNA genes in nucleotide sequences","volume":"32","author":"Laslett","year":"2004","journal-title":"Nucleic Acids Res."},{"key":"ref38","doi-asserted-by":"publisher","first-page":"2896","DOI":"10.1093\/gbe\/evw195","article-title":"Animal mitochondrial DNA as we do not know it: mt-genome organization and evolution in nonbilaterian lineages","volume":"8","author":"Lavrov","year":"2016","journal-title":"Genome Biol. Evol."},{"key":"ref39","doi-asserted-by":"publisher","first-page":"W293","DOI":"10.1093\/nar\/gkab301","article-title":"Interactive tree of life (iTOL) v5: an online tool for phylogenetic tree display and annotation","volume":"49","author":"Letunic","year":"2021","journal-title":"Nucleic Acids Res."},{"key":"ref40","doi-asserted-by":"publisher","first-page":"3094","DOI":"10.1093\/bioinformatics\/bty191","article-title":"Minimap2: pairwise alignment for nucleotide sequences","volume":"34","author":"Li","year":"2018","journal-title":"Bioinformatics"},{"key":"ref41","doi-asserted-by":"publisher","first-page":"84","DOI":"10.1186\/1471-2164-12-84","article-title":"Potential efficacy of mitochondrial genes for animal DNA barcoding: a case study using eutherian mammals","volume":"12","author":"Luo","year":"2011","journal-title":"BMC Genom."},{"key":"ref42","doi-asserted-by":"publisher","first-page":"151","DOI":"10.1006\/jmbi.1999.3136","article-title":"A census of protein repeats","volume":"293","author":"Marcotte","year":"1999","journal-title":"J. Mol. Biol."},{"key":"ref43","doi-asserted-by":"publisher","first-page":"435","DOI":"10.1128\/CMR.00056-07","article-title":"Increasing importance of Balamuthia mandrillaris","volume":"21","author":"Matin","year":"2008","journal-title":"Clin. Microbiol. Rev."},{"key":"ref45","doi-asserted-by":"publisher","first-page":"6097","DOI":"10.1093\/nar\/18.20.6097","article-title":"Sequence logos: a new way to display consensus sequences","volume":"18","author":"Schneider","year":"1990","journal-title":"Nucleic Acids Res."},{"key":"ref47","doi-asserted-by":"publisher","first-page":"342","DOI":"10.1128\/CMR.15.3.342-354.2002","article-title":"Cultivation of pathogenic and opportunistic free-living Amebas","volume":"15","author":"Schuster","year":"2002","journal-title":"Clin. Microbiol. Rev."},{"key":"ref48","doi-asserted-by":"publisher","first-page":"3175","DOI":"10.1128\/JCM.41.7.3175-3180.2003","article-title":"Environmental isolation of Balamuthia mandrillaris associated with a case of amebic encephalitis","volume":"41","author":"Schuster","year":"2003","journal-title":"J. Clin. Microbiol."},{"key":"ref49","doi-asserted-by":"publisher","first-page":"879","DOI":"10.1086\/597260","article-title":"Under the radar: Balamuthia amebic encephalitis","volume":"48","author":"Schuster","year":"2009","journal-title":"Clin. Infect. Dis."},{"key":"ref50","doi-asserted-by":"publisher","first-page":"89","DOI":"10.1016\/j.micpath.2007.06.008","article-title":"Balamuthia amoebic encephalitis: an emerging disease with fatal consequences","volume":"44","author":"Siddiqui","year":"2008","journal-title":"Microb. Pathog."},{"key":"ref51","doi-asserted-by":"publisher","first-page":"539","DOI":"10.1038\/msb.2011.75","article-title":"Fast, scalable generation of high-quality protein multiple sequence alignments using Clustal omega","volume":"7","author":"Sievers","year":"2011","journal-title":"Mol. Syst. Biol."},{"key":"ref52","doi-asserted-by":"publisher","first-page":"1611","DOI":"10.1101\/gr.361602","article-title":"The Bioperl toolkit: Perl modules for the life sciences","volume":"12","author":"Stajich","year":"2002","journal-title":"Genome Res."},{"key":"ref53","doi-asserted-by":"publisher","first-page":"537","DOI":"10.1093\/bioinformatics\/bti054","article-title":"Circular genome visualization and exploration using CGView","volume":"21","author":"Stothard","year":"2005","journal-title":"Bioinformatics"},{"key":"ref54","doi-asserted-by":"publisher","first-page":"W6","DOI":"10.1093\/nar\/gkx391","article-title":"GeSeq\u2014versatile and accurate annotation of organelle genomes","volume":"45","author":"Tillich","year":"2017","journal-title":"Nucleic Acids Res."},{"key":"ref55","first-page":"153","article-title":"Infections with free-living amebae","volume-title":"Handb. Clin. Neurol.","author":"Visvesvara","year":"2013"},{"key":"ref56","doi-asserted-by":"crossref","first-page":"683","DOI":"10.1016\/B978-0-7020-5101-2.00051-0","article-title":"Pathogenic and opportunistic free-living amoebae: agents of human and animal disease","volume-title":"Anonymous Manson\u2019s tropical diseases","author":"Visvesvara","year":"2014"},{"key":"ref57","doi-asserted-by":"publisher","first-page":"2750","DOI":"10.1128\/JCM.28.12.2750-2756.1990","article-title":"Leptomyxid ameba, a new agent of amebic meningoencephalitis in humans and animals","volume":"28","author":"Visvesvara","year":"1990","journal-title":"J. Clin. Microbiol."},{"key":"ref58","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1111\/j.1574-695X.2007.00232.x","article-title":"Pathogenic and opportunistic free-living amoebae: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri, and Sappinia diploidea","volume":"50","author":"Visvesvara","year":"2007","journal-title":"FEMS Immunol. Med. Microbiol."},{"key":"ref59","doi-asserted-by":"publisher","first-page":"e005031","DOI":"10.1099\/jmmcr.0.005031","article-title":"A Balamuthia survivor","volume":"3","author":"Vollmer","year":"2016","journal-title":"JMM Case Rep."},{"key":"ref60","doi-asserted-by":"publisher","first-page":"e112963","DOI":"10.1371\/journal.pone.0112963","article-title":"Pilon: an integrated tool for comprehensive microbial variant detection and genome assembly improvement","volume":"9","author":"Walker","year":"2014","journal-title":"PLoS One"},{"key":"ref61","doi-asserted-by":"publisher","first-page":"2348","DOI":"10.1080\/22221751.2020.1835447","article-title":"Balamuthia mandrillaris infection in China: a retrospective report of 28 cases","volume":"9","author":"Wang","year":"2020","journal-title":"Emerg. Microbes Infect."},{"key":"ref62","doi-asserted-by":"publisher","first-page":"18","DOI":"10.1038\/nrm3920","article-title":"Intrinsically disordered proteins in cellular signalling and regulation","volume":"16","author":"Wright","year":"2015","journal-title":"Nat. Rev. Mol. Cell Biol."}],"container-title":["Frontiers in Microbiology"],"original-title":[],"link":[{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fmicb.2023.1162963\/full","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,5,5]],"date-time":"2023-05-05T06:11:00Z","timestamp":1683267060000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.frontiersin.org\/articles\/10.3389\/fmicb.2023.1162963\/full"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2023,5,5]]},"references-count":58,"alternative-id":["10.3389\/fmicb.2023.1162963"],"URL":"https:\/\/doi.org\/10.3389\/fmicb.2023.1162963","relation":{},"ISSN":["1664-302X"],"issn-type":[{"value":"1664-302X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2023,5,5]]},"article-number":"1162963"}}