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Capsular polysaccharide synthesis gene cluster- what’s correct?
Posted by lixuerui2002 on 13 Jan 2010 at 01:47 GMT
red by the
S. suis 2 isolates from China. Five of 12 structural genes, including
cps2B, cps2D, cps2F, cps2K, and
cps2L exactly matched those of
S. suis P1/7. In contrast, seven structural genes showed some point variations. The divergent genes included
cps2A (G163A, F273I, and F302L),
cps2C (I160V and Q195K),
cps2E (G190E, W264L, and S281C),
Capsular polysaccharide synthesis gene cluster- what’s correct?
Xuerui-Li Juan-Xin, Jixing-Liu
Animal Infectious Diseases Research Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences; State Key Laboratory of Veterinary etiological Biology; Key Laboratory of Grazing Animal Diseases of Ministry of Agriculture; Key Laboratory of Veterinary Public Health of Ministry of Agriculture, Lanzhou, Gansu, 730046, China
Streptococcus suis (S. suis) is an important pathogen that causes meningitis, encephalitis, pneumonia, endocarditis, polyserositis, arthritis, septicemia and abortions in pigs. This microorganism has also been described as a pathogen for people in contact with swine or/and swine by-products, as well as for other mammalian and bird species. Up to now, there are 35 S. suis serotypes have been described based on the capsular polysaccharides, of which, capsular serotype 2 is the most common one in the world. The first case of human S. suis infection was reported in Denmark in 1968, Since then, human infections caused by S. suis have been reported in countries such as Netherlands; France; the United Kingdom; China; Thailand; Vietnam ; the United States and other region. This pathogen has emerged as an important public health issue in some countries (China CDC, http://www.chinacdc.net.cn ). It seems the human infection of S. suis in China is much more serious than other countries. During an episode in Nantong, Jiangsu province (China) in 1998, 14 patients died of the disease. S. suis type 2 strains 98HAH was responsible for the outbreak. During another episode in Ziyang, Sichuan province(China) in 2005, 39 patients died of the disease. S. suis type 2 strains 05ZYH was associated with this outbreak.
To compare the S. suis isolates of the Chinese outbreak with European invasive isolates, S. suis isolates from the 2005 outbreak in Sichuan Province were compared at a molecular level with a European serotype-2 strain (P1/7) and with some isolates from the 1998 outbreak in Jiangsu Province by Tang and his colleagues（Jiaqi Tang et al. Streptococcal Toxic Shock Syndrome Caused by Streptococcus suis Serotype 2. Plos medicine, May 2006. 3 (5 ):0668-0676）. They reported that the 2005 isolates are indistinguishable from the isolates that caused the 1998 outbreak in China. But the serotype-2 strain associated with 2005 outbreak is quite different to European invasive isolate S. suis P1/7 with regard to the capsular polysaccharide synthesis gene cluster pattern. They reported that five of 12 structural genes of S. suis 05ZYH33, including cps2B, cps2D, cps2F, cps2K, and cps2L were exactly matched those of S. suis P1/7. In contrast, seven structural genes showed some point variations. The divergent genes included cps2A (G163A, F273I, and F302L), cps2C (I160V and Q195K), cps2E (G190E, W264L, and S281C), cps2G (F183Y), cps2H (M268I and K300E), cps2I (K145M), and cps2J (Q175K, R201S, S213G, L214I, andY223K).(see fig 1).
This is interesting. However, when we compared the capsular polysaccharide synthesis gene cluster of S. suis 05ZYH33, S. suis 98HAH33, S. suis P1/7, and S. suis SC84, which was also isolated in Sichuan in 2005 outbreak, our results are not in agreement with this result.
We collected capsular polysaccharide synthesis gene cluster of S. suis 05ZYH33, S. suis SC84, S. suis 98HAH33 and S. suis P1/7 from the Genbank and used DNAstar software to analysis these genes. In our results, cps2B, cps2D, cps2F, cps 2H, cps 2J, cps2K of S. suis 05ZYH33, S. suis SC84 and S. suis 98HAH33 were exactly matched those of S. suis P1/7 and cps2C gene of S. suis 05ZYH33, S. suis SC84 and S. suis 98HAH33 was dismatched those of S. suis P1/7
For cps2A, this gene of S. suis SC84 and S. suis 98HAH33 was exactly matched those of S. suis P1/7, the point variations only exist in S. suis 05ZYH33 ( T 53 G, T 104 G, C 670 G).( See Fig 2)
For cps 2E , this gene of S. suis SC84 was exactly matched that of S. suis P1/7. the point variations only exist in S. suis 98HAH33( G1118A ). It seems there missed a base at 1118 for S. suis 05ZYH33, as a result, we don't know exactly the base of cps 2E gene for S. suis 05ZYH33 at this position. ( See Fig 3)
For cps 2G , this gene of S. suis 05ZYH33 and S. suis SC84 was exactly matched that of S. suis P1/7. the point variations only exist in S. suis 98HAH33 (A 7 C, T 11 C). ( See Fig 4)
For cps 2I , this gene of S. suis 05ZYH33 and S. suis SC84 was exactly matched that of S. suis P1/7. the point variations was also only exist in S. suis 98HAH33 ( A 550 G , T 729 G). ( See Fig 5)
However , we could not find the cps 2L gene of these strains in Genbank.
From the analysis we can see that the genes of cps2B, cps2D, cps2F, cps2G, cps 2H, cps2I, cps 2J and cps2K of S. suis 05ZYH33 are exactly matched those of S. suis P1/7 .The genes of cps2A, cps2B, cps2D, cps2E ,cps2F, cps2G, cps 2H, cps2I, cps 2J and cps2K of S. suis SC84 are exactly matched those of S. suis P1/7 and the genes of cps2A, cps2B, cps2D, cps2F, cps 2H, cps 2J and cps2K of S. suis 98HAH33 are exactly matched those of S. suis P1/7 . The gene of cps 2C of Streptococcus suis SC84, S. suis 05ZYH33, and S. suis 98HAH33 was all dismatched with S. suis P1/7 and showed the same point variations.(See Fig 6 ).
When considering S. suis 05ZYH33, S. suis SC84 and S. suis s 98HAH33 were all isolated from china and S. suis 05ZYH33 and S. suis SC84 wre all isolated from the same outbreak, we can hardly believe S. suis SC84 has more closer relationship with S. suis P1/7 than S. suis 05ZYH33 and S. suis 98HAH33. Also, we can hardly believe S. suis SC84, S. suis 05ZYH33 have more closer relationship with S. suis P1/7 than S. suis 98HAH33.
It seems that the capsular polysaccharide synthesis gene cluster of S. suis 98HAH33 showed more point variations. This may caused by sequencing error, Wether some of the point variations caused by sequencing error or not need further research.
Nucleotide sequence accession numbers. The nucleotide sequences for CPS gene of s.suis have been deposited in GenBank under the following respective accession numbers:
For Streptococcus suis P1/7: Accession number:AM946016 Reigion 552842..554287 for cps2A; Accession number NC_012925 Reigion: 554305..554994 for cps2B; Region 555004..555681 for cps2C; Region: 555720..556451 for cps2D; Region: 556476..557855 for cps2E; Region: 557890..559059 for cps2F; Region 559063..560220 for cps2G; Region: 560609..561979 for cps2H; Region 562047..563174 for cps2I; Region 563385..564383 for cps2J; Region: 564376..565380 for cps2K.
For Streptococcus suis 05ZYH33 Accession number: NC_009442 Region 552507..553958 for cps2A; Region: 553976..554665 for cps2B; Region: 554675..555370 for cps 2C; Region 555396..556124 for cps 2D; region: 556149..557387 for cps 2E; Region : 557562..558731 for cps 2F; region 558735..559892 for cps 2G; region: 560206..561651 for cps 2H; Region 561686..562918 for cps 2I; region: 563057..564055 for cps 2J; Region: 564048..565052 for cps 2K.
Streptococcus suis 98HAH33 Accession number: NC_009443 Region 552366..553358 for cps2A; Region: 553839..554528 for cps 2B; Region: 554538..555215 for cps 2C; Region 555254..555985 for cps 2D; region 556010..557389 for cps 2E; region: 558596..559753 for cps 2G; region: 561549..562781 for cps 2I; region: 562920..563918 for cps 2J.
For Streptococcus suis SC84, Accession number: NC_012924 Region 552409..553854 for cps2A; Region: 553872..554561 for cps 2B； Region: 554571..555248 for cps 2C; Region 555287..556018 for cps 2D; Region: 556043..557422 for cps 2E; Region: 557457..558626 for cps 2F; Region: 558630..559787 for cps 2G; region: 560176..561546 for cps 2H; region: 561614..562813 for cps 2I; Region: 562952..563950 for cps 2J; region : 562952..563950 for cps 2K.
The following reply to the comments by Li et al was provided by email to PLoS Medicine by Dr. Youjun Feng, one of the co-authors of the study.
Molecular epidemiology of human Streptococcus suis outbreak in China, 2005
Jiaqi Tang1*, Changjun Wang1, Youjun Feng2, Weizhong Yang3, Huaidong Song4, 9, Zhihai Chen5, Hongjie Yu3, Xiuzhen Pan1, Xiaojun Zhou6, Huaru Wang1, Bo Wu6, Haili Wang1, Huamei Zhao1, Ying Lin7, Jianhua Yue1, Zhenqiang Wu7, Xiaowei He7, Feng Gao2, Abdul Hamid Khan2, Jian Wang8, Guo-Ping Zhao9, Yu Wang3, Xiaoning Wang7, Zhu Chen4,9, George F. Gao2,*
1 Department of Epidemiology, Research Institute for Medicine of Nanjing Command, Nanjing, China
2 Center for Molecular Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China
3 Chinese Center for Disease Control and Prevention (China CDC), Beijing, China
4 State Key Laboratory of Medical Genomics, Ruijin Hospital Affiliated to Medical School of Shanghai Jiao-Tong University, Shanghai, China
5 Beijing Ditan Hospital, Beijing, China
6 Department of Pathology, Jinling Hospital of Nanjing, Nanjing, China
7 School of Biosciences and Bioengineering, South China University of Technology, Guangzhou, China
8 Beijing Genomics Institute, Chinese Academy of Sciences, Beijing, China
9 Chinese National Human Genome Center, Shanghai, China
Correspondence: Prof. Jiaqi Tang (tjq85@ hotmail.com) or George F. Gao (firstname.lastname@example.org).
We are pleased to read Dr. Liu and coworkers’ comments on our epidemiological study of Streptococcus suis published in PLoS Medicine five years ago. For the concerns raised by Li et al, we will address as follows:
1) The sequences of capsular polysaccharide (CPS) synthesis gene cluster we reported in PLoS Medicine is acquired by direct sequencing of the clinical samples from diseased piglets and STSS patients. However, the cps sequences (available in Gebank, cited by Li et al.) actually are from the Streptococcus suis 05ZYH33, and SC84, which have been sub-cultured for many generations. As we know, during the process of bacteria propagation, it is common that mutations occur esp., those genes encoding bacterial components (e.g., CPS) under the selection pressure of host and environment. Thereafter, it should be noted that the two laboratory strains (05ZYH33, SC84) can not completely match or reflect the original and field causative agent of the SS2 epidemic we reported. Additionally, the difference of different sequencer’s sequencing capability, and the essence of Streptococcus suis at low GC% both determine the quality/confidence/exactness of bacterial genomes, and it is of probably that minor variation/difference exists in the different genomes (even the same strain) from different sequencing facility. In light of the knowledge mentioned above, it is extremely hard to assess which genome sequence is correct, which one is wrong.
2) Li et al.’ comments to our PLoS Medicine is based not on the repeated experimental data from their own laboratory, but only four sequence online (they are separately from three different laboratory, and also are four different isolate). Indeed, it is hard to figure out 1) which sequencing facility is best, 2) which genome is absolutely exact/ reliable. However, it sounds like that Liu and coworker have supposed the two premises (1) those genomes deposited in GenBank are completely correct; 2) SC84 and 05ZYH33 are fully same as our clinical agent we reported) before they criticized our paper. Frankly, we still can not believe the 05ZYH33 genome is 100% correct, although it is sequenced in our group, because we can not thus far rule out the possibility that it is a revertant of the originally clinical isolate or not. Therefore, we believed the general conclusion drew by Li et al. cannot be supported by these data. Actually, the only one way that can probably provide us a relatively more convincing information thus far, is re-sequencing of multiple strains with the aid of more advanced different sequencing tools/strategies (including 454, comparative genomics hybridization (CGS), Solida, Pyro-sequencing, etc.). By the way, our group have joined a joint SS2 research and employed CGS to carry out extensive comparative genomics of over 20 Streptococcus suis strains.
3) To our surprise, Li et al. concluded that SC84 is much more closely to P1/7, which only based on 1-2 point mutations in CPS gene cluster. It is common sense for a bacteriologist that RFLP and MLST (at genomic level) and comparative genomics are generally used to address bacterial phylogenetic relationship. That is why we do not agree with Li et al.’s comments at this point.
4) Li et al. claimed that Streptococcus suis 2 does not possess cps2L gene in CPS synthesis cluster. Our 5-year follow-up study together with preliminary result in PLoS Medicine have pointed out that cps2L does exist in the genome of 05ZYH33 (05SSU0577), and also completely match to that of P1/7, SC84 and 98HAH12. It is proposed to be a membrane protein involved in the export of O-antigen and teichoic acid of Streptococcus suis. Apparently, the cartoon depiction of CPS2 cluster showed by Li et al. is not correct.
In conclusion, we appreciate Li et al’s interest in our study, and also welcome their comments.