Abstract
We have used HiSeq 2000 technology to generate a draft genome sequence of Streptococcus pneumoniae strain A66. This is a common study strain used in investigations of pneumococcal bacterium-host interactions and was used in the seminal genetic studies of Avery et al.
GENOME ANNOUNCEMENT
Streptococcus pneumoniae is a prominent human pathogen throughout the world, particularly as a cause of pneumonia, meningitis, bacteremia, and otitis media. S. pneumoniae strain A66 (NCTC 7978) and derivatives, such as A66.1, are virulent in mice and have been used by many laboratories for in vivo studies examining host immune responses (1, 2), preclinical vaccination evaluation (3–5), the development of novel therapies (6, 7), studies on virulence mechanisms (8, 9), and the use of a bioluminescent form to track infection in vivo (10, 11). It has also been used in a variety of in vitro studies, notably including the work of Avery et al. (12). To facilitate the use of A66 as a study strain, we present here its draft genome sequence. DNA was harvested from a minimally passaged culture from the National Collection of Type Cultures, where the strain was deposited in 1949 from the Rockefeller Institute for Medical Research, and sequenced using Illumina HiSeq 2000 technology.
Assembly was performed using Velvet software (13), and the assembled contigs were reordered and oriented by alignment using Mauve software (14) with S. pneumoniae OXC141 as a reference. The assembly consisted of 159 contigs, with an N50 of 53,248 bp. It was automatically annotated using RAST (15) and compared to strains TIGR4 and OXC141 using the Artemis Comparison Tool. The A66 draft genome was 1,983,415 bp in length with a GC content of 39.7%. Annotation found 2,087 coding sequences, including the genes for pneumolysin, the serotype 3 capsular locus, neuraminidase A, choline binding protein A, hyluronidase, ZmpA, and zinc metalloprotease B. However, A66 lacked the rlrA pilus locus, psrP locus, and zmpC. The genome-derived multilocus sequence type is ST387 (16). No acquired antimicrobial resistance genes were identified in the genome sequence using ResFinder version 2.1 (17), and the strain was phenotypically susceptible to all antibiotics tested using Vitek 2 (card AST-ST01).
The availability of a draft genome sequence for A66 will greatly facilitate its value as a model strain for investigating pneumococcal biology.
Nucleotide sequence accession numbers.
This whole-genome shotgun project has been deposited in DDBJ/ENA/GenBank under the accession number LN847353. The version described in this paper is the first version, LN847353.1.
ACKNOWLEDGMENTS
The help of the core sequencing and informatics team at the Wellcome Trust Sanger Institute is gratefully acknowledged.
This project was supported by the Wellcome Trust (grant 098051).
Footnotes
Citation Hahn C, Harrison EM, Parkhill J, Holmes MA, Paterson GK. 2015. Draft genome sequence of the Streptococcus pneumoniae Avery strain A66. Genome Announc 3(3):e00697-15. doi:10.1128/genomeA.00697-15.
REFERENCES
- 1.Weber SE, Tian H, Pirofski LA. 2011. CD8+ cells enhance resistance to pulmonary serotype 3 Streptococcus pneumoniae infection in mice. J Immunol 186:432–442. doi: 10.4049/jimmunol.1001963. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 2.Seyoum B, Yano M, Pirofski LA. 2011. The innate immune response to Streptococcus pneumoniae in the lung depends on serotype and host response. Vaccine 29:8002–8011. doi: 10.1016/j.vaccine.2011.08.064. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 3.Oliveira ML, Miyaji EN, Ferreira DM, Moreno AT, Ferreira PC, Lima FA, Santos FL, Sakauchi MA, Takata CS, Higashi HG, Raw I, Kubrusly FS, Ho PL. 2010. Combination of pneumococcal surface protein A (PspA) with whole cell pertussis vaccine increases protection against pneumococcal challenge in mice. PLoS One 5:e10863. doi: 10.1371/journal.pone.0010863. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4.Yamamoto M, McDaniel LS, Kawabata K, Briles DE, Jackson RJ, McGhee JR, Kiyono H. 1997. Oral immunization with PspA elicits protective humoral immunity against Streptococcus pneumoniae infection. Infect Immun 65:640–644. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5.Khan MN, Shukla D, Bansal A, Mustoori S, Ilavazhagan G. 2009. Immunogenicity and protective efficacy of GroEL (hsp60) of Streptococcus pneumoniae against lethal infection in mice. FEMS Immunol Med Microbiol 56:56–62. doi: 10.1111/j.1574-695X.2009.00548.x. [DOI] [PubMed] [Google Scholar]
- 6.Srivastava A, Casey H, Johnson N, Levy O, Malley R. 2007. Recombinant bactericidal/permeability-increasing protein rBPI21 protects against pneumococcal disease. Infect Immun 75:342–349. doi: 10.1128/IAI.01089-06. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7.Weeks JN, Boyd KL, Rajam G, Ades EW, McCullers JA. 2011. Immunotherapy with a combination of intravenous immune globulin and P4 peptide rescues mice from postinfluenza pneumococcal pneumonia. Antimicrob Agents Chemother 55:2276–2281. doi: 10.1128/AAC.00057-11. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Paterson GK, Mitchell TJ. 2006. The role of Streptococcus pneumoniae sortase A in colonisation and pathogenesis. Microbes Infect 8:145–153. doi: 10.1016/j.micinf.2005.06.009. [DOI] [PubMed] [Google Scholar]
- 9.Iannelli F, Chiavolini D, Ricci S, Oggioni MR, Pozzi G. 2004. Pneumococcal surface protein C contributes to sepsis caused by Streptococcus pneumoniae in mice. Infect Immun 72:3077–3080. doi: 10.1128/IAI.72.5.3077-3080.2004. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 10.Francis KP, Yu J, Bellinger-Kawahara C, Joh D, Hawkinson MJ, Xiao G, Purchio TF, Caparon MG, Lipsitch M, Contag PR. 2001. Visualizing pneumococcal infections in the lungs of live mice using bioluminescent Streptococcus pneumoniae transformed with a novel Gram-positive lux transposon. Infect Immun 69:3350–3358. doi: 10.1128/IAI.69.5.3350-3358.2001. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11.Orihuela CJ, Gao G, Mcgee M, Yu J, Francis KP, Tuomanen E. 2003. Organ-specific models of Streptococcus pneumoniae disease. Scand J Infect Dis 35:647–652. doi: 10.1080/00365540310015854. [DOI] [PubMed] [Google Scholar]
- 12.Avery OT, MacLeod CM, McCarty M. 1944. Studies on the chemical nature of the substrate inducing natural transformation of pneumococcal types: induction of transformation by a deoxyribonucleic acid fraction acid fraction from pneumococcus type III. J Exp Med 79:137–158. doi: 10.1084/jem.79.2.137. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 13.Zerbino DR, Birney E. 2008. Velvet: Algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 18:821–829. doi: 10.1101/gr.074492.107. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 14.Darling AC, Mau B, Blattner FR, Perna NT. 2004. Mauve: multiple alignment of conserved genomic sequence with rearrangements. Genome Res 14:1394–1403. doi: 10.1101/gr.2289704. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 15.Aziz RK, Bartels D, Best AA, DeJongh M, Disz T, Edwards RA, Formsma K, Gerdes S, Glass EM, Kubal M, Meyer F, Olsen GJ, Olson R, Osterman AL, Overbeek RA, McNeil LK, Paarmann D, Paczian T, Parrello B, Pusch GD, Reich C, Stevens R, Vassieva O, Vonstein V, Wilke A, Zagnitko O. 2008. The RAST server: rapid annotations using subsystems technology. BMC Genomics 9:75. doi: 10.1186/1471-2164-9-75. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16.Larsen MV, Cosentino S, Rasmussen S, Friis C, Hasman H, Marvig RL, Jelsbak L, Sicheritz-Pontén T, Ussery DW, Aarestrup FM, Lund O. 2012. Multilocus sequence typing of total-genome-sequenced bacteria. J Clin Microbiol 50:1355–1361. doi: 10.1128/JCM.06094-11. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 17.Zankari E, Hasman H, Cosentino S, Vestergaard M, Rasmussen S, Lund O, Aarestrup FM, Larsen MV. 2012. Identification of acquired antimicrobial resistance genes. J Antimicrob Chemother 67:2640–2644. doi: 10.1093/jac/dks261. [DOI] [PMC free article] [PubMed] [Google Scholar]