Whole-genome enrichment and sequencing of Chlamydia trachomatis directly from clinical samples

被引：66

作者：

Christiansen, Mette T. ^{[1
]}

Brown, Amanda C. ^{[2
]}

Kundu, Samit ^{[1
,8
]}

Tutill, Helena J. ^{[1
]}

Williams, Rachel ^{[1
]}

Brown, Julianne R. ^{[3
]}

Holdstock, Jolyon ^{[2
]}

Holland, Martin J. ^{[4
]}

Stevenson, Simon ^{[5
]}

Dave, Jayshree ^{[6
]}

Tong, C. Y. William ^{[6
]}

Einer-Jensen, Katja ^{[7
]}

Depledge, Daniel P. ^{[1
]}

Breuer, Judith ^{[1
]}

机构：

[1] UCL, Div Infect & Immun, London WC1E 6BT, England

[2] Oxford Gene Technol, Begbroke OX5 1PF, Oxon, England

[3] Great Ormond St Hosp Sick Children, London WC1N 3JH, England

[4] London Sch Hyg & Trop Med, London WC1E 7HT, England

[5] Univ Coll London Hosp, London WC1E 6DE, England

[6] Barts Hlth NHS Trust, London E1 2ES, England

[7] QIAGEN AAR, DK-8200 Aarhus N, Denmark

[8] Canterbury Christchurch Univ, Sch Human & Life Sci, Canterbury CT1 1QU, Kent, England

来源：

BMC INFECTIOUS DISEASES | 2014年 / 14卷

关键词：

Whole-genome enrichment; Whole-genome sequencing; Chlamydia trachomatis; Clinical samples; PHYLOGENETIC ANALYSIS; BACTERIAL VAGINOSIS; MEMBRANE-PROTEIN; IN-VITRO; RESISTANCE; MUTATIONS; RECOMBINATION; INFECTIONS; DISEASE; YOLK;

D O I：

10.1186/s12879-014-0591-3

中图分类号：

R51 [传染病];

学科分类号：

100401 ;

摘要：

Background: Chlamydia trachomatis is a pathogen of worldwide importance, causing more than 100 million cases of sexually transmitted infections annually. Whole-genome sequencing is a powerful high resolution tool that can be used to generate accurate data on bacterial population structure, phylogeography and mutations associated with antimicrobial resistance. The objective of this study was to perform whole-genome enrichment and sequencing of C. trachomatis directly from clinical samples. Methods: C. trachomatis positive samples comprising seven vaginal swabs and three urine samples were sequenced without prior in vitro culture in addition to nine cultured C. trachomatis samples, representing different serovars. A custom capture RNA bait set, that captures all known diversity amongst C. trachomatis genomes, was used in a whole-genome enrichment step during library preparation to enrich for C. trachomatis DNA. All samples were sequenced on the MiSeq platform. Results: Full length C. trachomatis genomes (> 95-100% coverage of a reference genome) were successfully generated for eight of ten clinical samples and for all cultured samples. The proportion of reads mapping to C. trachomatis and the mean read depth across each genome were strongly linked to the number of bacterial copies within the original sample. Phylogenetic analysis confirmed the known population structure and the data showed potential for identification of minority variants and mutations associated with antimicrobial resistance. The sensitivity of the method was > 10-fold higher than other reported methodologies. Conclusions: The combination of whole-genome enrichment and deep sequencing has proven to be a non-mutagenic approach, capturing all known variation found within C. trachomatis genomes. The method is a consistent and sensitive tool that enables rapid whole-genome sequencing of C. trachomatis directly from clinical samples and has the potential to be adapted to other pathogens with a similar clonal nature.

引用

页数：11

共 45 条

[1]

Buriti FCA, 2007, ARCH LATINOAM NUTR, V57, P373

[2] Sequences of multiple bacterial genomes and a Chlamydia trachomatis genotype from direct sequencing of DNA derived from a vaginal swab diagnostic specimen [J].

Andersson, P. ;

Klein, M. ;

Lilliebridge, R. A. ;

Giffard, P. M. .

CLINICAL MICROBIOLOGY AND INFECTION, 2013, 19 (09) :E405-E408

[3] Non-coding nucleotides and amino acids near the active site regulate peptide deformylase expression and inhibitor susceptibility in Chlamydia trachomatis [J].

Bao, Xiaofeng ;

Pachikara, Niseema D. ;

Oey, Christopher B. ;

Balakrishnan, Amit ;

Westblade, Lars F. ;

Tan, Ming ;

Chase, Theodore, Jr. ;

Nickels, Bryce E. ;

Fan, Huizhou .

MICROBIOLOGY-SGM, 2011, 157 :2569-2581

[4] Frequency of development and associated physiological cost of azithromycin resistance in Chlamydia psittaci 6BC and C-trachomatis L2 [J].

Binet, Rachel ;

Maurelli, Anthony T. .

ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, 2007, 51 (12) :4267-4275

[5] Productivity losses attributable to untreated chlamydial infection and associated pelvic inflammatory disease in reproductive-aged women [J].

Blandford, John M. ;

Gift, Thomas L. .

SEXUALLY TRANSMITTED DISEASES, 2006, 33 (10) :S117-S121

[6] The Global Burden of Trachoma: A Review [J].

Burton, Matthew J. ;

Mabey, David C. W. .

PLOS NEGLECTED TROPICAL DISEASES, 2009, 3 (10)

[7]

Caller V, 2013, WHITE PAPER PROBALIT, P1

[8] jModelTest 2: more models, new heuristics and parallel computing [J].

Darriba, Diego ;

Taboada, Guillermo L. ;

Doallo, Ramon ;

Posada, David .

NATURE METHODS, 2012, 9 (08) :772-772

[9] Deep Sequencing of Viral Genomes Provides Insight into the Evolution and Pathogenesis of Varicella Zoster Virus and Its Vaccine in Humans [J].

Depledge, Daniel P. ;

Kundu, Samit ;

Jensen, Nancy J. ;

Gray, Eleanor R. ;

Jones, Meleri ;

Steinberg, Sharon ;

Gershon, Anne ;

Kinchington, Paul R. ;

Schmid, D. Scott ;

Balloux, Francois ;

Nichols, Richard A. ;

Breuer, Judith .

MOLECULAR BIOLOGY AND EVOLUTION, 2014, 31 (02) :397-409

[10] Specific Capture and Whole-Genome Sequencing of Viruses from Clinical Samples [J].

Depledge, Daniel P. ;

Palser, Anne L. ;

Watson, Simon J. ;

Lai, Imogen Yi-Chun ;

Gray, Eleanor R. ;

Grant, Paul ;

Kanda, Ravinder K. ;

Leproust, Emily ;

Kellam, Paul ;

Breuer, Judith .

PLOS ONE, 2011, 6 (11)

← 1 2 3 4 5 →