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Comparative genome analysis identifies two large deletions in the genome of highly-passaged attenuated Streptococcus agalactiae strain YM001 compared to the parental pathogenic strain HN016.

Wang R, Li L, Huang Y, Luo F, Liang W, Gan X, Huang T, Lei A, Chen M, Chen L - BMC Genomics (2015)

Bottom Line: The global outbreaks of Streptococcus disease in tilapia cause huge economic losses and threaten human food hygiene safety as well.Comparative genome analysis indicated that compared with HN016, YM001 genome had two significant large deletions, at the sizes of 5832 and 11,116 bp respectively, resulting in the deletion of three rRNA and ten tRNA genes, as well as the deletion and functional damage of ten genes related to metabolism, transport, growth, anti-stress, etc.The deleted and mutated functional genes all encode metabolism- and growth-related proteins, not the known virulence proteins, indicating that the metabolism- and growth-related genes are important for the pathogenesis of S. agalactiae.

View Article: PubMed Central - PubMed

Affiliation: Guangxi Key Laboratory for Aquatic Genetic Breeding and Healthy Aquaculture, Guangxi Academy of Fishery Sciences, Nanning, 530021, People's Republic of China. raywongxx@163.com.

ABSTRACT

Background: Streptococcus agalactiae (S. agalactiae), also known as group B Streptococcus (GBS), is an important pathogen for neonatal pneumonia, meningitis, bovine mastitis, and fish meningoencephalitis. The global outbreaks of Streptococcus disease in tilapia cause huge economic losses and threaten human food hygiene safety as well. To investigate the mechanism of S. agalactiae pathogenesis in tilapia and develop attenuated S. agalactiae vaccine, this study sequenced and comparatively analyzed the whole genomes of virulent wild-type S. agalactiae strain HN016 and its highly-passaged attenuated strain YM001 derived from tilapia.

Methods: We performed Illumina sequencing of DNA prepared from strain HN016 and YM001. Sequencedreads were assembled and nucleotide comparisons, single nucleotide polymorphism (SNP) , indels were analyzed between the draft genomes of HN016 and YM001. Clustered regularly interspaced short palindromic repeats (CRISPRs) and prophage were detected and analyzed in different S. agalactiae strains.

Results: The genome of S. agalactiae YM001 was 2,047,957 bp with a GC content of 35.61 %; it contained 2044 genes and 88 RNAs. Meanwhile, the genome of S. agalactiae HN016 was 2,064,722 bp with a GC content of 35.66 %; it had 2063 genes and 101 RNAs. Comparative genome analysis indicated that compared with HN016, YM001 genome had two significant large deletions, at the sizes of 5832 and 11,116 bp respectively, resulting in the deletion of three rRNA and ten tRNA genes, as well as the deletion and functional damage of ten genes related to metabolism, transport, growth, anti-stress, etc. Besides these two large deletions, other ten deletions and 28 single nucleotide variations (SNVs) were also identified, mainly affecting the metabolism- and growth-related genes.

Conclusions: The genome of attenuated S. agalactiae YM001 showed significant variations, resulting in the deletion of 10 functional genes, compared to the parental pathogenic strain HN016. The deleted and mutated functional genes all encode metabolism- and growth-related proteins, not the known virulence proteins, indicating that the metabolism- and growth-related genes are important for the pathogenesis of S. agalactiae.

No MeSH data available.


Related in: MedlinePlus

Diversity of the CRISPR1 locus in 5 S. agalactiae strains. Spacers were identified by the CRISPRtionary program, with numbers assigned to each spacer [51]. The names of strains are given on the left. R stands for Repeat, and S stands for Spacer
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Fig3: Diversity of the CRISPR1 locus in 5 S. agalactiae strains. Spacers were identified by the CRISPRtionary program, with numbers assigned to each spacer [51]. The names of strains are given on the left. R stands for Repeat, and S stands for Spacer

Mentions: CRISPRs are a bacterial adaptive immune defense mechanism against the invasion of foreign genes. When foreign gene invades bacteria, the CRISPRs integrate and save the intruding gene fragment. Under the re-invasion of the same genes, mediated by specific RNA, CRISPRs and CRISPR-associated proteins (Cas proteins) will cut and destroy the invading foreign genes, which may include bacterial phages, plasmids, and mobile genetic elements (MGEs) [31, 32]. S. agalactiae has 2 CRISPR/Cas systems, type 1-C CRISPR2 and type 2-A CRISPR1; while the latter is ubiquitous, the former is only present in a few strains [33]. The CRISPR sequences were analyzed among the 8 S. agalactiae strains in Table 2 using the CRISPRs web server (http://crispr.u-psud.fr/Server/). The results indicated that 3 tilapia-derived S. agalactiae serotype Ib strains did not contain any CRISPR sequences, while 5 S. agalactiae serotype Ia strains all had CRISPR1 but did not contain CRISPR2. Further analysis of the CRISPR1 from the 5 S. agalactiae serotype Ia strains showed that the CRISPR sequence in S. agalactiae strain HN016 derived from tilapia in China was same as that in GD201008-001 and ZQ0910 and had eight spacers, but the attenuated strain YM001 only contained seven spacers; all other sequences were the same between both strains (Fig. 3). During the process of foreign nucleic acid invasion and bacterial evolution, to avoid overly long locus of CRISPRs, bacteria may choose to insert or remove spacer sequences between CRISPRs, and the insertion or removal of spacer is polarized, i.e., a new spacer is always inserted between the leader sequence and the following repetitive sequence, while the removed spacer is usually located at the 3’ end of CRISPRs [34]. Lopez-Sanchez et al. analyzed the CRISPRs of more than 200 wild-type S. agalactiae strains but did not find the 3’ terminal deletion [33]. Although Liu et al. showed that two spacer sequences were deleted at the 3’ end of CRISPRs in ZQ0910 stain [12], we found that the assembling of the published sequence of this fragment in ZQ0910 had certain mistake. After reanalysis and alignment of this sequence, we confirmed that the CRISPRs of ZQ0910 were exactly the same as those of HN016 and GD201008-001. Compared to other tilapia-derived wild-type virulent strains, the CRISPRs of YM001 had a deletion of 1 spacer at the 3’ end. Philippe et al. studied the CRISPR loci of S. thermophilus and found that the selective removal of spacer sequence may be caused by that these spacer sequences have little value for the survival of bacteria in the environment at the time [35]. Therefore, normal natural growth and passage are hard to cause the removal of CRISPR spacer sequence in S. agalactiae, whereas in the absence of the threat of foreign nucleic acid invasion, highly intensified continuous passage in laboratory may lead to the loss or removal of spacer sequence in CRISPRs.Table 2


Comparative genome analysis identifies two large deletions in the genome of highly-passaged attenuated Streptococcus agalactiae strain YM001 compared to the parental pathogenic strain HN016.

Wang R, Li L, Huang Y, Luo F, Liang W, Gan X, Huang T, Lei A, Chen M, Chen L - BMC Genomics (2015)

Diversity of the CRISPR1 locus in 5 S. agalactiae strains. Spacers were identified by the CRISPRtionary program, with numbers assigned to each spacer [51]. The names of strains are given on the left. R stands for Repeat, and S stands for Spacer
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4634907&req=5

Fig3: Diversity of the CRISPR1 locus in 5 S. agalactiae strains. Spacers were identified by the CRISPRtionary program, with numbers assigned to each spacer [51]. The names of strains are given on the left. R stands for Repeat, and S stands for Spacer
Mentions: CRISPRs are a bacterial adaptive immune defense mechanism against the invasion of foreign genes. When foreign gene invades bacteria, the CRISPRs integrate and save the intruding gene fragment. Under the re-invasion of the same genes, mediated by specific RNA, CRISPRs and CRISPR-associated proteins (Cas proteins) will cut and destroy the invading foreign genes, which may include bacterial phages, plasmids, and mobile genetic elements (MGEs) [31, 32]. S. agalactiae has 2 CRISPR/Cas systems, type 1-C CRISPR2 and type 2-A CRISPR1; while the latter is ubiquitous, the former is only present in a few strains [33]. The CRISPR sequences were analyzed among the 8 S. agalactiae strains in Table 2 using the CRISPRs web server (http://crispr.u-psud.fr/Server/). The results indicated that 3 tilapia-derived S. agalactiae serotype Ib strains did not contain any CRISPR sequences, while 5 S. agalactiae serotype Ia strains all had CRISPR1 but did not contain CRISPR2. Further analysis of the CRISPR1 from the 5 S. agalactiae serotype Ia strains showed that the CRISPR sequence in S. agalactiae strain HN016 derived from tilapia in China was same as that in GD201008-001 and ZQ0910 and had eight spacers, but the attenuated strain YM001 only contained seven spacers; all other sequences were the same between both strains (Fig. 3). During the process of foreign nucleic acid invasion and bacterial evolution, to avoid overly long locus of CRISPRs, bacteria may choose to insert or remove spacer sequences between CRISPRs, and the insertion or removal of spacer is polarized, i.e., a new spacer is always inserted between the leader sequence and the following repetitive sequence, while the removed spacer is usually located at the 3’ end of CRISPRs [34]. Lopez-Sanchez et al. analyzed the CRISPRs of more than 200 wild-type S. agalactiae strains but did not find the 3’ terminal deletion [33]. Although Liu et al. showed that two spacer sequences were deleted at the 3’ end of CRISPRs in ZQ0910 stain [12], we found that the assembling of the published sequence of this fragment in ZQ0910 had certain mistake. After reanalysis and alignment of this sequence, we confirmed that the CRISPRs of ZQ0910 were exactly the same as those of HN016 and GD201008-001. Compared to other tilapia-derived wild-type virulent strains, the CRISPRs of YM001 had a deletion of 1 spacer at the 3’ end. Philippe et al. studied the CRISPR loci of S. thermophilus and found that the selective removal of spacer sequence may be caused by that these spacer sequences have little value for the survival of bacteria in the environment at the time [35]. Therefore, normal natural growth and passage are hard to cause the removal of CRISPR spacer sequence in S. agalactiae, whereas in the absence of the threat of foreign nucleic acid invasion, highly intensified continuous passage in laboratory may lead to the loss or removal of spacer sequence in CRISPRs.Table 2

Bottom Line: The global outbreaks of Streptococcus disease in tilapia cause huge economic losses and threaten human food hygiene safety as well.Comparative genome analysis indicated that compared with HN016, YM001 genome had two significant large deletions, at the sizes of 5832 and 11,116 bp respectively, resulting in the deletion of three rRNA and ten tRNA genes, as well as the deletion and functional damage of ten genes related to metabolism, transport, growth, anti-stress, etc.The deleted and mutated functional genes all encode metabolism- and growth-related proteins, not the known virulence proteins, indicating that the metabolism- and growth-related genes are important for the pathogenesis of S. agalactiae.

View Article: PubMed Central - PubMed

Affiliation: Guangxi Key Laboratory for Aquatic Genetic Breeding and Healthy Aquaculture, Guangxi Academy of Fishery Sciences, Nanning, 530021, People's Republic of China. raywongxx@163.com.

ABSTRACT

Background: Streptococcus agalactiae (S. agalactiae), also known as group B Streptococcus (GBS), is an important pathogen for neonatal pneumonia, meningitis, bovine mastitis, and fish meningoencephalitis. The global outbreaks of Streptococcus disease in tilapia cause huge economic losses and threaten human food hygiene safety as well. To investigate the mechanism of S. agalactiae pathogenesis in tilapia and develop attenuated S. agalactiae vaccine, this study sequenced and comparatively analyzed the whole genomes of virulent wild-type S. agalactiae strain HN016 and its highly-passaged attenuated strain YM001 derived from tilapia.

Methods: We performed Illumina sequencing of DNA prepared from strain HN016 and YM001. Sequencedreads were assembled and nucleotide comparisons, single nucleotide polymorphism (SNP) , indels were analyzed between the draft genomes of HN016 and YM001. Clustered regularly interspaced short palindromic repeats (CRISPRs) and prophage were detected and analyzed in different S. agalactiae strains.

Results: The genome of S. agalactiae YM001 was 2,047,957 bp with a GC content of 35.61 %; it contained 2044 genes and 88 RNAs. Meanwhile, the genome of S. agalactiae HN016 was 2,064,722 bp with a GC content of 35.66 %; it had 2063 genes and 101 RNAs. Comparative genome analysis indicated that compared with HN016, YM001 genome had two significant large deletions, at the sizes of 5832 and 11,116 bp respectively, resulting in the deletion of three rRNA and ten tRNA genes, as well as the deletion and functional damage of ten genes related to metabolism, transport, growth, anti-stress, etc. Besides these two large deletions, other ten deletions and 28 single nucleotide variations (SNVs) were also identified, mainly affecting the metabolism- and growth-related genes.

Conclusions: The genome of attenuated S. agalactiae YM001 showed significant variations, resulting in the deletion of 10 functional genes, compared to the parental pathogenic strain HN016. The deleted and mutated functional genes all encode metabolism- and growth-related proteins, not the known virulence proteins, indicating that the metabolism- and growth-related genes are important for the pathogenesis of S. agalactiae.

No MeSH data available.


Related in: MedlinePlus