Comprehensive Characterization of a Novel Bacteriophage, vB_VhaS_MAG7 against a Fish Pathogenic Strain of Vibrio harveyi and Its In Vivo Efficacy in Phage Therapy Trials
Abstract
:1. Introduction
2. Results
2.1. vB_VhaS_MAG7 Morphology and Characteristics
2.2. Host Range
2.3. Thermal and pH Stability of vB_VhaS_MAG7
2.4. One-Step Growth of vB_VhaS_MAG7
2.5. In Vitro Cell Lysis
2.6. Genomic Analysis of vB_VhaS_MAG7
2.7. Genomic Synteny and Phylogenetic Analysis
2.8. In Vivo Phage Therapy in Gilthead Seabream Larvae
3. Discussion
4. Materials and Methods
4.1. Bacterial Strains
4.2. Antibiotic Susceptibility Testing
4.3. Isolation and Purification of Bacteriophages
4.4. Transmission Electron Microscopy
4.5. Host Range Assay
4.6. Stability of Phage in Different Temperatures and pH Values
4.7. One-Step Growth Assay
4.8. In Vitro Cell Lysis
4.9. Genomic Analysis
4.10. In Vivo Phage Therapy Trial in Gilthead Seabream Larvae
4.11. Statistical Analysis
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Strain | Species | Country | Host | Lysis |
---|---|---|---|---|
MM46 * | V. harveyi | Greece | Sparus aurata | + |
DSM 19623 | V. harveyi | USA | Talochestria capensis | - |
SA 2.1 | V. harveyi | Saudi Arabia | Sparus aurata | - |
DSM 2171 | V. alginolyticus | Japan | Trachurus trachurus | - |
Gal 90 | V. harveyi | Greece | Sparus aurata | - |
Vh No22 | V. harveyi | Greece | Dicentrarchus labrax | - |
Kef 62 | V. harveyi | Greece | Dicentrarchus labrax | - |
Kef 75 | V. harveyi | Greece | Dicentrarchus labrax | - |
Gal 56 | V. harveyi | Greece | Dicentrarchus labrax | - |
Gal 77 | V. harveyi | Greece | Sparus aurata | - |
Gal 72 | V. harveyi | Greece | Dicentrarchus labrax | - |
Gal 94 | V. harveyi | Greece | Sparus aurata | - |
L. SUSI | V. parahaemolyticus | Philippines | Shrimp | - |
V1 | V. alginolyticus | Greece | Sparus aurata | - |
LAR194 | V. mediterranei | Greece | Artemia nauplii | - |
SM1 | V. harveyi | Greece | Seriola dumerili | - |
MAN113 | V. splendidus | Greece | Seriola dumerili | - |
VarvA4 1.1 | V. harveyi | Greece | Sparus aurata | - |
VH2 | V. harveyi | Greece | Seriola dumerili | - |
VhP1 Liv | V. harveyi | Greece | Seriola dumerili | - |
VhP1 Spl | V. harveyi | Greece | Dicentrarchus labrax | - |
DY05 | V. owensii | Greece | Dicentrarchus labrax | - |
SA 6.2 | V. owensii | Saudi Arabia | Oreochromis niloticus | - |
VIB391 | V. campbellii | Thailand | Shrimp | - |
Kef 56 | V. rotiferianus | Greece | Dicentrarchus labrax | - |
VhSerFre | V. harveyi | Greece | Seriola dumerili | - |
sngr | V. harveyi | Greece | Dentex dentex | - |
ks6 | V. owensii | Greece | Dicentrarchus labrax | - |
VH5 | V. harveyi | Greece | Seriola dumerili | - |
RG1 | V. harveyi | Greece | Dicentrarchus labrax | - |
Serkid | V. harveyi | Greece | Seriola dumerili | - |
SERKID2 | V. harveyi | Greece | Seriola dumerili | - |
SERSD | V. harveyi | Greece | Seriola dumerili | - |
SA 5.1 | V. harveyi | Saudi Arabia | Sparus aurata | - |
SA 6.1 | V. harveyi | Saudi Arabia | Sparus aurata | - |
SA 9.2 | V. harveyi | Saudi Arabia | Sparus aurata | - |
SA 1.2 | V. harveyi | Saudi Arabia | Sparus aurata | - |
SA 7.1 | V. harveyi | Saudi Arabia | Sparus aurata | - |
SA 3.1 | V. harveyi | Saudi Arabia | Sparus aurata | - |
SA 4.1 | V. harveyi | Saudi Arabia | Sparus aurata | - |
SA 2.1 | V. harveyi | Saudi Arabia | Sparus aurata | - |
VH6 | V. harveyi | Greece | Dicentrarchus labrax | - |
V2 | V. alginolyticus | Greece | Dentex dentex | - |
SA 1.1 | V. owensii | Saudi Arabia | Sparus aurata | - |
SA 9.1 | V. owensii | Saudi Arabia | Sparus aurata | - |
Type | Predicted Functions | Start | End | Length | Strand |
---|---|---|---|---|---|
ORF 1 | Hypothetical protein | 401 | 629 | 72 | Forward |
ORF 2 | Hypothetical protein | 633 | 868 | 74 | Forward |
ORF 3 | Bifunctional DNA primase/polymerase | 871 | 3183 | 767 | Forward |
ORF 4 | Coil-containing protein | 3237 | 3762 | 170 | Forward |
ORF 5 | Putative helicase subunit | 3762 | 4551 | 259 | Forward |
ORF 6 | Hypothetical protein | 4542 | 5095 | 180 | Forward |
ORF 7 | PD-(D/E)XK nuclease superfamily protein | 5151 | 6229 | 355 | Forward |
ORF 8 | P-loop-containing nucleoside triphosphate hydrolase | 6212 | 7641 | 471 | Forward |
ORF 9 | Helicase superfamily 1/2 ATP-binding domain protein | 7611 | 9317 | 563 | Forward |
ORF 10 | Hypothetical protein | 9307 | 9739 | 138 | Forward |
ORF 11 | Hypothetical protein | 9870 | 10,209 | 108 | Forward |
ORF 12 | Hypothetical protein | 10,308 | 10,496 | 59 | Forward |
ORF 13 | Hypothetical protein | 10,481 | 10,903 | 136 | Forward |
ORF 14 | Hypothetical protein | 10,889 | 11,070 | 56 | Forward |
ORF 15 | Hypothetical protein | 11,055 | 11,462 | 131 | Forward |
ORF 16 | Hypothetical protein | 11,449 | 11,992 | 176 | Forward |
ORF 17 | Hypothetical protein | 11,959 | 12,164 | 65 | Forward |
ORF 18 | NTP-ppase-like protein | 12,158 | 12,601 | 144 | Forward |
ORF 19 | Yopx protein | 12,699 | 13,096 | 128 | Forward |
ORF 20 | Hypothetical protein | 13,112 | 13,233 | 36 | Forward |
ORF 21 | Hypothetical protein | 13,220 | 13,370 | 45 | Forward |
ORF 22 | Tmhelix-containing protein | 13,358 | 13,627 | 86 | Forward |
ORF 23 | Endodeoxyribonuclease I | 13,614 | 14,082 | 152 | Forward |
ORF 24 | Hypothetical protein | 14,070 | 14,594 | 169 | Forward |
ORF 25 | Hypothetical Protein | 14,582 | 14,737 | 47 | Forward |
ORF 26 | Membrane lipoprotein | 14,795 | 15,477 | 223 | Forward |
ORF 27 | Hypothetical Protein | 15,565 | 15,765 | 63 | Reverse |
ORF 28 | Intramolecular chaperone auto-processing domain protein | 15,769 | 18,027 | 750 | Reverse |
ORF 29 | Hypothetical protein | 18,117 | 18,535 | 135 | Reverse |
ORF 30 | Tail fiber protein | 18,526 | 19,525 | 329 | Reverse |
ORF 31 | Hypothetical protein | 19,566 | 19,908 | 110 | Reverse |
ORF 32 | Tmhelix-containing protein | 19,910 | 20,263 | 113 | Reverse |
ORF 33 | Hypothetical protein | 20,311 | 20,720 | 130 | Reverse |
ORF 34 | Hypothetical protein | 20,700 | 20,813 | 34 | Reverse |
ORF 35 | Hypothetical protein | 20,804 | 21,118 | 104 | Reverse |
ORF 36 | Hypothetical protein | 21,129 | 21,385 | 80 | Forward |
ORF 37 | Hypothetical protein | 21,542 | 21,658 | 34 | Forward |
ORF 38 | Hypothetical protein | 21,688 | 22,548 | 282 | Reverse |
ORF 39 | Hypothetical protein | 22,567 | 23,346 | 255 | Reverse |
ORF 40 | Baseplate J-like protein | 23,334 | 24,534 | 397 | Reverse |
ORF 41 | Hypothetical protein | 24,520 | 24,943 | 137 | Reverse |
ORF 42 | Hypothetical protein | 25,020 | 25,217 | 62 | Reverse |
ORF 43 | Hypothetical protein | 25,205 | 25,360 | 51 | Reverse |
ORF 44 | Tmhelix-containing protein | 25,390 | 25,630 | 75 | Reverse |
ORF 45 | Hypothetical protein | 25,617 | 26,541 | 303 | Reverse |
ORF 46 | Hypothetical protein | 26,612 | 27,280 | 218 | Reverse |
ORF 47 | Hypothetical Protein | 27,243 | 27,408 | 50 | Reverse |
ORF 48 | Cytosine specific methyltransferase | 27,395 | 28,586 | 392 | Reverse |
ORF 49 | Putative tail protein | 28,637 | 29,857 | 403 | Reverse |
ORF 50 | DNA circularization protein | 30,519 | 31,778 | 415 | Reverse |
ORF 51 | Phosphoesterase | 31,837 | 32,371 | 175 | Reverse |
ORF 52 | Tmhelix-containing protein | 32,364 | 32,485 | 36 | Reverse |
ORF 53 | Tmhelix-containing protein | 32,550 | 32,842 | 90 | Reverse |
ORF 54 | Hypothetical Protein | 32,876 | 33,085 | 65 | Reverse |
ORF 55 | Tail tape measure protein | 33,191 | 34,803 | 533 | Reverse |
ORF 56 | Hypothetical protein | 34,796 | 34,994 | 60 | Reverse |
ORF 57 | Tail assembly chaperone protein | 35,029 | 35,467 | 141 | Reverse |
ORF 58 | DNA binding HTH domain protein | 35,609 | 35,835 | 71 | Forward |
ORF 59 | Hypothetical protein | 35,826 | 36,023 | 61 | Forward |
ORF 60 | Hypothetical protein | 36,005 | 36,180 | 54 | Forward |
ORF 61 | Hypothetical protein | 36,153 | 36,356 | 64 | Forward |
ORF 62 | Hypothetical protein | 36,340 | 36,634 | 93 | Forward |
ORF 63 | Major tail protein | 36,673 | 37,064 | 125 | Reverse |
ORF 64 | Tail protein | 37,052 | 38,522 | 486 | Reverse |
ORF 65 | Hypothetical protein | 38,524 | 39,202 | 223 | Reverse |
ORF 66 | Hypothetical protein | 39,185 | 39,580 | 127 | Reverse |
ORF 67 | Hypothetical protein | 39,568 | 39,932 | 117 | Reverse |
ORF 68 | Major capsid protein | 39,934 | 41,028 | 360 | Reverse |
ORF 69 | Hypothetical protein | 41,031 | 41,455 | 136 | Reverse |
ORF 70 | Putative ATP dependent Clp protease | 41,441 | 42,692 | 412 | Reverse |
ORF 71 | Portal protein | 42,679 | 44,325 | 543 | Reverse |
ORF 72 | Hypothetical protein | 44,386 | 44,657 | 85 | Reverse |
ORF 73 | Terminase large subunit | 44,708 | 46,677 | 652 | Reverse |
ORF 74 | Hypothetical protein | 46,666 | 47,374 | 232 | Reverse |
ORF 75 | Putative HNH endonuclease | 47,435 | 47,948 | 167 | Reverse |
ORF 76 | Coil-containing protein | 48,283 | 49,029 | 245 | Reverse |
Antimicrobial Agent | Zone Diameter (mm) | Interpretation | |
---|---|---|---|
Flumequine | UB | 30 | S |
Tetracycline | TE | 25 | S |
Florfenicol | FFC | 25 | S |
Oxytetracycline | OT | 26 | I |
Oxolinic acid | OA | 22 | S |
Trimethoprim/sulfamethoxazole | SXT | 15 | I |
Ampicillin | AMP | - | R |
Piperacillin | PRL | - | R |
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Droubogiannis, S.; Pavlidi, L.; Skliros, D.; Flemetakis, E.; Katharios, P. Comprehensive Characterization of a Novel Bacteriophage, vB_VhaS_MAG7 against a Fish Pathogenic Strain of Vibrio harveyi and Its In Vivo Efficacy in Phage Therapy Trials. Int. J. Mol. Sci. 2023, 24, 8200. https://doi.org/10.3390/ijms24098200
Droubogiannis S, Pavlidi L, Skliros D, Flemetakis E, Katharios P. Comprehensive Characterization of a Novel Bacteriophage, vB_VhaS_MAG7 against a Fish Pathogenic Strain of Vibrio harveyi and Its In Vivo Efficacy in Phage Therapy Trials. International Journal of Molecular Sciences. 2023; 24(9):8200. https://doi.org/10.3390/ijms24098200
Chicago/Turabian StyleDroubogiannis, Stavros, Lydia Pavlidi, Dimitrios Skliros, Emmanouil Flemetakis, and Pantelis Katharios. 2023. "Comprehensive Characterization of a Novel Bacteriophage, vB_VhaS_MAG7 against a Fish Pathogenic Strain of Vibrio harveyi and Its In Vivo Efficacy in Phage Therapy Trials" International Journal of Molecular Sciences 24, no. 9: 8200. https://doi.org/10.3390/ijms24098200
APA StyleDroubogiannis, S., Pavlidi, L., Skliros, D., Flemetakis, E., & Katharios, P. (2023). Comprehensive Characterization of a Novel Bacteriophage, vB_VhaS_MAG7 against a Fish Pathogenic Strain of Vibrio harveyi and Its In Vivo Efficacy in Phage Therapy Trials. International Journal of Molecular Sciences, 24(9), 8200. https://doi.org/10.3390/ijms24098200