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Complete Genome Sequence of Fish Pathogen Aeromonas hydrophila JBN2301 Wuming Yang,a Ningqiu Li,b Ming Li,a Defeng Zhang,b Guannan Ana School of Animal Science and Nutritional Engineering, Wuhan Polytechnic University, Wuhan, Chinaa; Pearl River Fishery Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangzhou, Chinab
Aeromonas hydrophila is one of the most important fish pathogens in China. Here, we report complete genome sequence of a virulent strain, A. hydrophila JBN2301, which was isolated from diseased crucian carp.
Citation Yang W, Li N, Li M, Zhang D, An G. 2016. Complete genome sequence of fish pathogen Aeromonas hydrophila JBN2301. Genome Announc 4(1):e01615-15. doi: 10.1128/genomeA.01615-15. Copyright © 2016 Yang et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 3.0 Unported license. Address correspondence to Ningqiu Li, [email protected].
A
eromonas hydrophila, belonging to the family Aeromonadaceae, is an autochthonous species in freshwater environments and a member of the normal microflora in the fish intestinal tract (1). However, under stress conditions, some virulent strains of A. hydrophila can invade the most majority of freshwater fish species and infect them with hemorrhagic septicemia. In China, the fish disease caused by this microorganism has become the most important bacterial disease of fish to date and leads to great economic losses periodically per year (2–4). In July 2009, an epidemic septicemia with continuous fish mortality broke out in a crucian carp pond of Guanqiao Fishery surrounded by Donghu Lake, Wuhan, China. A. hydrophila JBN2301, isolated from a diseased crucian carp, is highly virulent to crucian carp and zebrafish, with a 50% lethal dose (LD50) of ⬍3 ⫻ 105 CFU per fish. Whole-genome sequencing was performed employing Illumina genome analyzer (BGI, Shenzhen, China) using a shotgun strategy, which produced paired-ends totaling about 724 Mb with about 180-fold coverage of the genome. Genome sequence data were processed and assembled into 28 contigs and 6 scaffolds with the software SOAPdenovo version 2.04 (5). Gaps between contigs were closed by combinatorial PCR and sequencing amplicons by primer walking. Finally, this assembly process produced a complete genome with one circular chromosome and three circular plasmids. Gene prediction was performed using the NCBI Prokaryotic Genome Annotation Pipeline (2013 release). The genome of strain JBN2301 includes a chromosome with a length of 5,127,362 bp and three plasmids with lengths of 6,318 bp, 6,162 bp, and 6,045 bp. This genome contains 4,438 proteincoding genes, with a G⫹C content of 60.77%. A total of 129 tRNA genes were predicted by tRNAscan-SE (6), while 10 rRNA operons were predicted by RNAmmer (7). The A. hydrophila JBN2301 genome was also annotated by RAST (8) to facilitate a comparison with A. hydrophila ATCC 7966. Among 95 genes involving virulence, disease, and defense, there are two genes involving bile hydrolysis and lysozyme inhibitor in strain JBN2301 only, which enables this strain to survive in the fish immune system. Compared to ATCC 7966, JBN2301
January/February 2016 Volume 4 Issue 1 e01615-15
uniquely encodes 17 proteins, which are entirely associated with bacteriophages. The plasmids from A. hydrophila JBN2301 have unique features. Plasmid1 (6,318 bp) contains five genes, including those encoding the YdcE-YdcD toxin-antitoxin system, whose role is to prevent programming death of host cells. Plasmid2 (6,162 bp) carries seven genes, including those encoding ferric enterobactin uptake protein FepE and the toxin-antitoxin replicon stabilization system. Plasmid3 (6,045 bp) contains genes encoding three hypothetical proteins with unknown functions. In summary, the genome sequence of A. hydrophila JBN2301 contributes to the understanding of its mechanism of survival in the pathogen-host interaction environment and plasmid-carried genes encoding the toxin-antitoxin system. Further comparison with the genomes of many virulent and avirulent strains will help search for conserved virulence genes of this bacterial pathogen. Nucleotide sequence accession numbers. The complete genome sequence of A. hydrophila JBN2301 was deposited at DDBJ/EMBL/ GenBank under the accession numbers CP013178 to CP013181. ACKNOWLEDGMENTS This study was supported by KLFDD/KAAIT project (201303), Pearl River Fishery Research Institute (C.A.F.S.) and XLXM project (2014RZ), Wuhan Polytechnic University.
FUNDING INFORMATION Key Laboratory of Fishery Drug Development, Ministry of Agriculture provided funding to Wuming Yang under grant number 201303. Wuhan Polytechnic University provided funding to Ming Li under grant number 2014RZ.
REFERENCES 1. Cipriano RC. 2001. Aeromonas hydrophila and motile aeromonad septicemias of fish. U.S. Department of the Interior, Fish and Wildlife Service, Washington, DC. http://digitalcommons.unl.edu/cgi/viewcontent .cgi?article⫽1133&context⫽usfwspubs. 2. Lu CP. 1992. Pathogenic Aeromonas hydrophila and the fish diseases caused by it. J Fish China 16:282–288 (In Chinese). 3. Zhang X, Yang W, Wu H, Gong X, Li A. 2014. Multilocus sequence typing revealed a clonal lineage of Aeromonas hydrophila caused motile Aeromonas
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Received 26 November 2015 Accepted 7 December 2015 Published 28 January 2016
Yang et al.
septicemia outbreaks in pond-cultured cyprinid fish in an epidemic area in central China. Aquaculture 432:1– 6. http://dx.doi.org/10.1016/ j.aquaculture.2014.04.017. 4. Zhang XJ, Yang WM, Zhang DF, Li TT, Gong XN, Li AH. 2013. Does the gastrointestinal tract serve as the infectious route of Aeromonas hydrophila in crucian carp (Carassius carassius)? Aquacult Res 45:1–14. 5. Luo R, Liu B, Xie Y, Li Z, Huang W, Yuan J, He G, Chen Y, Pan Q, Liu Y, Tang J, Wu G, Zhang H, Shi Y, Liu Y, Yu C, Wang B, Lu Y, Han C, Cheung DW, Yiu SM, Peng S, Zhu X, Liu G, Liao X, Li Y, Yang H, Wang J, Lam TW, Wang J. 2012. SOAPdenovo2: an empirically improved memory-efficient short-read de novo assembler. GigaScience 1:18. http:// dx.doi.org/10.1186/2047-217X-1-18.
6. Lowe TM, Eddy SR. 1997. tRNAscan-SE: a program for improved detection of transfer RNA genes in genomic sequence. Nucleic Acids Res 25: 955–964. http://dx.doi.org/10.1093/nar/25.5.0955. 7. Lagesen K, Hallin P, Rødland EA, Staerfeldt HH, Rognes T, Ussery DW. 2007. RNAmmer: consistent annotation of rRNA genes in genomic sequences. Nucleic Acids Res 35:3100 –3108. 8. 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. http://dx.doi.org/10.1186/1471-2164-9-75.
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Genome Announcements
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Complete Genome Sequence of Fish Pathogen Aeromonas hydrophila JBN2301 Wuming Yang,a Ningqiu Li,b Ming Li,a Defeng Zhang,b Guannan Ana School of Animal Science and Nutritional Engineering, Wuhan Polytechnic University, Wuhan, Chinaa; Pearl River Fishery Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangzhou, Chinab
Aeromonas hydrophila is one of the most important fish pathogens in China. Here, we report complete genome sequence of a virulent strain, A. hydrophila JBN2301, which was isolated from diseased crucian carp.
Citation Yang W, Li N, Li M, Zhang D, An G. 2016. Complete genome sequence of fish pathogen Aeromonas hydrophila JBN2301. Genome Announc 4(1):e01615-15. doi: 10.1128/genomeA.01615-15. Copyright © 2016 Yang et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 3.0 Unported license. Address correspondence to Ningqiu Li, [email protected].
A
eromonas hydrophila, belonging to the family Aeromonadaceae, is an autochthonous species in freshwater environments and a member of the normal microflora in the fish intestinal tract (1). However, under stress conditions, some virulent strains of A. hydrophila can invade the most majority of freshwater fish species and infect them with hemorrhagic septicemia. In China, the fish disease caused by this microorganism has become the most important bacterial disease of fish to date and leads to great economic losses periodically per year (2–4). In July 2009, an epidemic septicemia with continuous fish mortality broke out in a crucian carp pond of Guanqiao Fishery surrounded by Donghu Lake, Wuhan, China. A. hydrophila JBN2301, isolated from a diseased crucian carp, is highly virulent to crucian carp and zebrafish, with a 50% lethal dose (LD50) of ⬍3 ⫻ 105 CFU per fish. Whole-genome sequencing was performed employing Illumina genome analyzer (BGI, Shenzhen, China) using a shotgun strategy, which produced paired-ends totaling about 724 Mb with about 180-fold coverage of the genome. Genome sequence data were processed and assembled into 28 contigs and 6 scaffolds with the software SOAPdenovo version 2.04 (5). Gaps between contigs were closed by combinatorial PCR and sequencing amplicons by primer walking. Finally, this assembly process produced a complete genome with one circular chromosome and three circular plasmids. Gene prediction was performed using the NCBI Prokaryotic Genome Annotation Pipeline (2013 release). The genome of strain JBN2301 includes a chromosome with a length of 5,127,362 bp and three plasmids with lengths of 6,318 bp, 6,162 bp, and 6,045 bp. This genome contains 4,438 proteincoding genes, with a G⫹C content of 60.77%. A total of 129 tRNA genes were predicted by tRNAscan-SE (6), while 10 rRNA operons were predicted by RNAmmer (7). The A. hydrophila JBN2301 genome was also annotated by RAST (8) to facilitate a comparison with A. hydrophila ATCC 7966. Among 95 genes involving virulence, disease, and defense, there are two genes involving bile hydrolysis and lysozyme inhibitor in strain JBN2301 only, which enables this strain to survive in the fish immune system. Compared to ATCC 7966, JBN2301
January/February 2016 Volume 4 Issue 1 e01615-15
uniquely encodes 17 proteins, which are entirely associated with bacteriophages. The plasmids from A. hydrophila JBN2301 have unique features. Plasmid1 (6,318 bp) contains five genes, including those encoding the YdcE-YdcD toxin-antitoxin system, whose role is to prevent programming death of host cells. Plasmid2 (6,162 bp) carries seven genes, including those encoding ferric enterobactin uptake protein FepE and the toxin-antitoxin replicon stabilization system. Plasmid3 (6,045 bp) contains genes encoding three hypothetical proteins with unknown functions. In summary, the genome sequence of A. hydrophila JBN2301 contributes to the understanding of its mechanism of survival in the pathogen-host interaction environment and plasmid-carried genes encoding the toxin-antitoxin system. Further comparison with the genomes of many virulent and avirulent strains will help search for conserved virulence genes of this bacterial pathogen. Nucleotide sequence accession numbers. The complete genome sequence of A. hydrophila JBN2301 was deposited at DDBJ/EMBL/ GenBank under the accession numbers CP013178 to CP013181. ACKNOWLEDGMENTS This study was supported by KLFDD/KAAIT project (201303), Pearl River Fishery Research Institute (C.A.F.S.) and XLXM project (2014RZ), Wuhan Polytechnic University.
FUNDING INFORMATION Key Laboratory of Fishery Drug Development, Ministry of Agriculture provided funding to Wuming Yang under grant number 201303. Wuhan Polytechnic University provided funding to Ming Li under grant number 2014RZ.
REFERENCES 1. Cipriano RC. 2001. Aeromonas hydrophila and motile aeromonad septicemias of fish. U.S. Department of the Interior, Fish and Wildlife Service, Washington, DC. http://digitalcommons.unl.edu/cgi/viewcontent .cgi?article⫽1133&context⫽usfwspubs. 2. Lu CP. 1992. Pathogenic Aeromonas hydrophila and the fish diseases caused by it. J Fish China 16:282–288 (In Chinese). 3. Zhang X, Yang W, Wu H, Gong X, Li A. 2014. Multilocus sequence typing revealed a clonal lineage of Aeromonas hydrophila caused motile Aeromonas
Genome Announcements
genomea.asm.org 1
Downloaded from http://genomea.asm.org/ on January 30, 2016 by guest
Received 26 November 2015 Accepted 7 December 2015 Published 28 January 2016
Yang et al.
septicemia outbreaks in pond-cultured cyprinid fish in an epidemic area in central China. Aquaculture 432:1– 6. http://dx.doi.org/10.1016/ j.aquaculture.2014.04.017. 4. Zhang XJ, Yang WM, Zhang DF, Li TT, Gong XN, Li AH. 2013. Does the gastrointestinal tract serve as the infectious route of Aeromonas hydrophila in crucian carp (Carassius carassius)? Aquacult Res 45:1–14. 5. Luo R, Liu B, Xie Y, Li Z, Huang W, Yuan J, He G, Chen Y, Pan Q, Liu Y, Tang J, Wu G, Zhang H, Shi Y, Liu Y, Yu C, Wang B, Lu Y, Han C, Cheung DW, Yiu SM, Peng S, Zhu X, Liu G, Liao X, Li Y, Yang H, Wang J, Lam TW, Wang J. 2012. SOAPdenovo2: an empirically improved memory-efficient short-read de novo assembler. GigaScience 1:18. http:// dx.doi.org/10.1186/2047-217X-1-18.
6. Lowe TM, Eddy SR. 1997. tRNAscan-SE: a program for improved detection of transfer RNA genes in genomic sequence. Nucleic Acids Res 25: 955–964. http://dx.doi.org/10.1093/nar/25.5.0955. 7. Lagesen K, Hallin P, Rødland EA, Staerfeldt HH, Rognes T, Ussery DW. 2007. RNAmmer: consistent annotation of rRNA genes in genomic sequences. Nucleic Acids Res 35:3100 –3108. 8. 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. http://dx.doi.org/10.1186/1471-2164-9-75.
Downloaded from http://genomea.asm.org/ on January 30, 2016 by guest
2 genomea.asm.org
Genome Announcements
January/February 2016 Volume 4 Issue 1 e01615-15
