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50 kDa

Maltose. 2%. 0%. NaCl. Bile salts. 1%. 3%. 0.3%. 0%. MW. 14.4. 20.1. 30.0. 43.0. 67.0. 94.0. 90 kDa. 70 kDa. 65 kDa. 50 kDa. 45 kDa. 40 kDa. 35 kDa. 25 kDa. 20 kDa. 15 kDa. 1a.

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50 kDa

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  1. Maltose 2% 0% NaCl Bile salts 1% 3% 0.3% 0% MW 14.4 20.1 30.0 43.0 67.0 94.0 90 kDa 70 kDa 65 kDa 50 kDa 45 kDa 40 kDa 35 kDa 25 kDa 20 kDa 15 kDa 1a. Supplemental Figure 1a. SDS-PAGE analysis showing OMPs expressed by V. tubiashii strain 19105. From left to right, Lane 1, cells grown in the presence of 3% NaCl compared to those grown with 1% NaCl in Lane 2; Lane 3 cells grown in the presence of 0.3% bile salts compared to those grown without bile salts Lane 4; Lane 5 cells grown in the presence of 2% maltose compared to those grown without maltose Lane 6; Lane 7 represents the molecular markers, sizes are reported in kDa. Note the sequenced and uniquely expressed OMPs at each environmental condition are identified with an arrow.

  2. Temperature pH 37° 8.5 6 30° 7 MW 14.4 20.1 30.0 43.0 67.0 94.0 90 kDa 70 kDa 65 kDa 50 kDa 45 kDa 40 kDa 35 kDa 25 kDa 20 kDa 15 kDa 1b. Supplemental Figure 1b. SDS-PAGE analysis showing OMPs expressed by V. tubiashii strain 19105. From left to right, Lane 1-2, cells grown at different temperatures; and Lane 3-5, cells grown at various pH conditions; Lane 6 represents the molecular markers, sizes are reported in kDa. Note the sequenced and uniquely expressed OMPs at each environmental condition are identified with an arrow.

  3. NaCl Bile salts Maltose 1% 0.3% 2% 3% 0% 0% MW 14.4 90 kDa 70 kDa 20.1 65 kDa 50 kDa 45 kDa 30.0 40 kDa 35 kDa 25 kDa 43.0 20 kDa 67.0 15 kDa 94.0 2a. Supplemental Figure 2a. SDS-PAGE analysis showing OMPs expressed by V. tubiashii strain 19109. From left to right, Lane 1, cells grown in the presence of 3% NaCl compared to those grown with 1% NaCl Lane 2; Lane 3, cells grown in the presence of 0.3% bile salts compared to those grown without bile salts Lane 4; Lane 5, cells grown in the presence of 2% maltose compared to those grown without maltose, Lane 6; Lane 7 represents the molecular markers, sizes are reported in kDa. Note the sequenced and uniquely expressed OMPs at each environmental condition are identified with an arrow.

  4. pH Temperature 37° 8.5 7 6 43° 30° MW 14.4 90 kDa 70 kDa 20.1 65 kDa 50 kDa 45 kDa 30.0 40 kDa 35 kDa 25 kDa 43.0 20 kDa 67.0 15 kDa 94.0 2b. Supplemental Figure 2b. SDS-PAGE analysis showing OMPs expressed by V. tubiashii strain 19109. From left to right, Lane 1-3, cells grown at various temperatures; and Lane 4-6, cells grown at various pH conditions; Lane 7 represents the molecular markers, sizes are reported in kDa. Note the sequenced and uniquely expressed OMPs at each environmental condition are identified with an arrow.

  5. a. Vv4965-T1 CTCTAGAAGTTGGTGGCCGTGCTGAAGCTCGCCTATCTCTGAAAGATGGTAAGGCACAAG Vc395 CTCTAGAAGTTGGTGGCCGTGCTGAAGCTCGCCTATCTCTGAAAGATGGTAAGGCACAAG Vt109 CTCTAGAGATCGGCGGTCGTGCTGAAGCTCGTATGTCTCTAAAAGATGGTAAAGCTGAAG Vt105 CTATCGAAATGGGTGGTCGCGCGGAAGCACGTCTAGCTCTTAAAGATGGNAAGGCAGAAN ** * ** * ** ** ** ** ***** ** * **** ******** ** ** ** Vv4965-T1 ACAACTCTCGCGTACGTCTAAACTTCTTGGGTAAAGCAGAAATCAATGACAGCCTATACG Vc0395 ACAACTCTCGCGTACGTCTAAACTTCTTGGGTAAAGCAGAAATCAATGACAGCCTATACG Vt109 ACAAAACACGTGTACGTCTAAACTTCCTAGGTAAGACTCAAATCACTGATAGCCTATATG Vt105 ACAAATCTCGCGTACGTCTAAACTTCCTAGGTACAGCTCAAATCACTGACAGCCTATACG **** * ** *************** * **** * ****** *** ******** * Vv4965-T1 GTGTTGGTTTCTACGAAGGCGAGTTTACTACTAATGATCA------------AGGTAAAA Vc395 GTGTTGGTTTCTACGAAGGCGAGTTTACTACTAATGATCA------------AGGTAAAA Vt109 GTCTAGGTTTCTACGAAGCTGAGTACACTACAGCAGACAACACTGTAACTGTAGACGGCA Vt105 GTGTGGGTTTCTACGAAGGTGAGTTCACTACTAATGACAATGGCGGCGCTACAGATGGCA ** * ************* **** ***** ** * ** * Vv4965-T1 ACG-CGTCTAACA---ACAGCCTANACAACCGTTATACCTACGCTGGTATCGGTGGCACT Vc395 ACG-CGTCTAACA---ACAGCCTAGACAACCGTTATACCTACGCTGGTATCGGTGGCACT Vt109 AAA-CAACTGACGGTGATAACATTGAGCACCGTTACACTTACGCTGGTATTGGTGGTGAC Vt105 ACAACAGCTCATC----TGATCTAACTAACCGTTACCTATACGGTGGTATCGGCGGAGCT * * ** * * ******* **** ****** ** ** Vv4965-T1 TACGGTGAAGTGACTTACGGTAAAAACGATGGCGCATTGGGCGTAATCACTGACTTCACC Vc395 TACGGTGAAGTGACTTACGGTAAAAACGATGGCGCATTGGGCGTAATCACTGACTTCACC Vt109 TTTGGTGAAGTAACTTACGGTAAAAACGATGGTGCACTANGCGTTATCACTGACTTTACT Vt105 TTCGGTGAAGTAACTTACGGTAAGAACGACGGTGCGCTANGTGTTATCACTGATTTCACT * ******** *********** ***** ** ** * * ** ******** ** ** Supplemental Figure 3. Clustal X alignment of nucleotide sequence of ompU gene from V. tubiashii strains 19105 and 19109, V. vulnificus 4965-T1 and V. cholerae 395, respectively. ompU gene sequences were obtained in this study. Indicated by underlying are the PCR primer sequences used in the study. a. Forward sequence, b. Reverse sequence. The symbol “*” represents conserved nucleotides among the sequences.

  6. b. Vv4965-T1 NCNNTGGACTGACC-CAGTTCGTC-TTCTGATGCTACTTTACCAA----------CTTTG Vc395 GCNNNGNANTTANG-CAATACNTNGTTCTGATGCTACTTTACCAAA---------CTTTG Vt109 GCNNNGAACTGAANATAGCTCGTC-TTCTGCACCAGCTTTACCTACAGTACCAGTTGCTG Vt105 NCCNTNGACNNAAA-AAGCTCGTC-TTCTGCATCTGCTTTAGAGATG----TAAGATTTG * * * * * * ***** * ***** * ** Vv4965-T1 TCTGA--ATCT------AGCAGATTGAACTGGTAAGAGATGTAAGAGCGNNAGTTTGGCT Vc395 GCTGA--ATCT------AGCAGATTGAACTGGTAAGAGATGTAAGAGCGNAAGTTTGGCT Vt109 TCTTATCATCTTTACTAAGTAGGTTGAAGTTGTAAGAGATGTAAGTACGGAAGTTTGGCT Vt105 TCGTC--GTCT------AGCAGGTTAAAGTTGTACGATACGTAGNCACGGAAGTTAGGCT * *** ** ** ** ** * *** ** * *** ** **** **** Vv4965-T1 TGAAGTAGTAAGTTGCGTCGATAGCAAAATTATCTGCTGAAGT--------T-TCTTTCG Vc395 TGAAGTAGTAAGTTGCGTCGATAGCAAAATTATCTGCTGAAGT--------TCTCTTTCG Vt109 TGAAGTAGTATGTTGCATCGATCGCTACATTGTCAGCTGCATCATACGTAGTACCTGAAG Vt105 TAAAGTAGTATGTTGCGTCGATAGCGATGTTATCTGCAGCATCTTCTTTAGTTGAACCTG * ******** ***** ***** ** * ** ** ** * * * * Vv4965-T1 CTGTTTCTGCA---TTGTTGTATGTCGCAGTAAACGCAGCTTGACCTAGTTTGTAACCAG Vc395 CTGTTTCTGCA---TTGTTGTATGTCGCAGTAAACGCAGCTTGACCTAGTTTGTANCCAG Vt109 TTTTCTCTTCTAGCTTATTGTATGTTGTAGTGAATGCTACTTTTTCTAGAGTGTAAGCTG Vt105 TTTTGTTTTCTAGGTAGTTGTAAGACGTTGTGAATACAGTCTGGCCCATAGTGTAAGCTG * * * * * ***** * * ** ** * * * * ***** * * Vv4965-T1 CAGCTAGCTCGTAGCCAGTGTAGTCAACGT-----CTTTCGC---TAGCTC-------AC Vc395 CANCTANCTCGTAGCCAGTGTAGTCAACGT-----CTTNCGC---TAGCTC-------AC Vt109 CTGCGAATTCGTAACCAGTTTTGTCTGTTTT---GCTTGAGC---CGTCTTTGTCTTTAC Vt105 CTGCTAGTTCGTAACCTGTCTTATCAGTTGTTGAGCTAGAACCTTTCACTTGGTCTTCGC * * * ***** ** ** * ** ** * ** * Supplemental Figure 3. Clustal X alignment of nucleotide sequence of ompU gene from V. tubiashii strains 19105 and 19109, V. vulnificus 4965-T1 and V. cholerae 395, respectively. ompU gene sequences were obtained in this study. Indicated by underlying are the PCR primer sequences used in the study. a. Forward sequence, b. Reverse sequence. The symbol “*” represents conserved nucleotides among the sequences.

  7. a. Vt109 NCNNNNTCCNNNNCTTACCCAGCCTGACTCAAGCCATCTCAACCCTACGTAAAATGCTGA Vt105 ----------NCNNNTTCAAAACCTGACTCAGGCCATCTCATCCCTACGTAAAATGCTGA Vf33809 ---------------TGGTGGAATGCACACCGGCGGACTCGGCCCCAAAGAAGTCATCGC * ** * ** * *** *** * ** * Vt109 AAGACTCGACCAAGTCGCC--TCAGTTTGTGAAA-ACGGTTCCAAAACGTGGTTATCAGT Vt105 AAGACTCGACTAAATCCCCCATCAATCCGTGGAAGACGGTTCCAAAACGTGGTTATCAGT Vf33809 TACCGGCGGTCAAAAC-------------------------------------------- * ** ** Vt109 TGATCGCGACCGTCGAAAGCGTTCAGCTCGATGCAAGTAAAGATCCTGATGCCATCGAGC Vt105 TGATCGCGACCGTCGAAACCCTGGAGCTCGATGCAAGTAAAGATCCTGATGCCATCGAGC Vf33809 ------------------------------------------------------------ Vt109 AGTCAGATGAGTCTTACTCAAGTGCCGTATCCTCTTCAGAACTTGTCAGTCAAGCTGCTG Vt105 AATCATATGAGTCTTACTCAAGTGCCGTATCCTCTTCACAACTTGTCAGTCAAGCTGCTG Vf33809 ------------------------------------------------------------ Vt109 TTTCTGAAGCAGTTATTACTCAGACGTCGCCTTCTGTAGACTCGACACCTGCGGCACCAA Vt105 TTTCTGAATCAGTTATTACTCAGACGTCGCCTTCTGTAGACTCGACACCTGCAGCACCAA Vf33809 ------------------------------------------------------------ Vt109 AACCACGAATGGATCTGTTCACTAAATTGATTCTGTTGGTTGCCGTCATACTGCCAT--- Vt105 AACCACGAATGGATCTGTTCACTAAATTGATTCTGTTGGTTGCCGTCATACTGCCATCNN Vf33809 ------------------------------------------------------------ Supplemental Figure 4. Clustal X alignment of nucleotide sequence of toxR gene from V. tubiashii strains 19105 and 19109 compared to that of V. cholerae 395 and V. fluvialis strain ATCC 33809. toxR gene sequences were obtained in this study and gene sequence for V. fluvialis strain ATCC 33809 was obtained from the NCBI accession number AF170885. a. Forward sequence, b. Reverse sequence. The symbol “*” represents conserved nucleotides among the sequences.

  8. b. Vt109 TGAACAGATCCATTCGTGGTTTTGGTGCCGCAGGTGTCGAGT-CTACAGAAGGCGACGTC Vt105 TAAAGGATAACCGTAAGGATGTCGGAGTTCCAATTATTACGCACTCCAGCAGTCGATGTG Vf33809 TGGACGACTCCAGCCTGACTCAAGCCATCTCAACC--------CTACGTAAAATGCTGAA * * * * * ** ** * * * * Vt109 TGAGTAATAACTGCTTCAGAAACA-------GCAGCTTG-ACTGACAAGTTCTGAAGAGG Vt105 TGAGCCAAGAGGACTCCATGAACACTTAGNCGCGGTTTCTGCGGACCGATTCCGTGAAGG Vf33809 AGACT--CGACCAAGTC---GCCT--------CAGTTTGTGAAAAC-GGTTCCAAAACGT ** * * * * ** * *** * Vt109 AT------------ACGGTACTTGAGTAAGACTCATCTGACTGCTCGATG--GCATCAGG Vt105 ATGGATAGGGCCGGACGATAATTGTGCAAAATTAACTTGACTGCTCGATTATGCATCAGG Vf33809 GGTTATCAGTT--GATCGCGACCGTCGAAAGCGTTCAGCTCGATGCAAGTAAAGATCCTG * * ** * * * ** * Vt109 ATCT-TTACTT--GCATCGAGC-TGAACGCTT-TCGACGGTCGCGATC--AACTGATAAC Vt105 ATCTATTACCTATGCATCGAGCCTGAACGCTTATTAACGCTCGCTACCCNGAGTGATAAC Vf33809 ATGC--CATCGA-GCAGTCAGA-TGAGTCTTACTCAAGTGCCGTATCC--TCTTCAGAAC ** * *** ** *** * * * ** * * * *** Vt109 CAC---GTTTTGGAACCG--TTT-----TCACAAACTGAGGCGACTTGGTCGAGT----- Vt105 CACAGTGGTTGGTAACCGGATTTGNCGATCATAAATGGGTGCGACTTGGTGGTCTAGCTC Vf33809 TTGTCAGTCAAGCTGCTG---TT-----TCTGAAGCAGTTATTACTCAGACGTCG----- * * * * ** ** ** * *** * * Vt109 -CTTTCAGCATTTTACGTA------GGGTTGAG-ATGGCTTGAGTCAGGCTGGA--GTCG Vt105 GCTCTCAAACTTTTACAAACGTTAGGGGTTGAACATGGCTTGACTCAGGCTGCAACGTCT Vf33809 -CCTTCTGTAGACTCGACACCTGCGGCACCAAA---ACCACGAATGGATCTGTT----CA * ** * * * ** *** * Vt109 TCCACCT-CAAACCCCTGAG---AATACGTCTCCCNCNCTCTTNCAATCTCACAGTTCCA Vt105 TCCACCTTCAAACCCCTGAACCAAATACCTGTACTNCNCTCTTNCGTTTCCATAGTCACA Vf33809 CTAAATT--GATTCTGTTGGT--TGCCGTCATACTGCCTATTTGTGTCATTTTGTTTACG * * * * * * ** * * Supplemental Figure 4. Clustal X alignment of nucleotide sequence of toxR gene from V. tubiashii strains 19105 and 19109 compared to that of V. cholerae 395 and V. fluvialis strain ATCC 33809. toxR gene sequences were obtained in this study and gene sequence for V. fluvialis strain ATCC 33809 was obtained from the NCBI accession number AF170885. a. Forward sequence, b. Reverse sequence. The symbol “*” represents conserved nucleotides among the sequences.

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