37d578eca0472628f10360122c306cf6 animals-14-02352-v2.pdf f71090e9de41dc593f17710af36aa4e4edf71e9a animals-14-02352-v2.pdf 5d43a24a77211a0488a034defd5900f246b7d811f3aaa1a166296b8394290751 animals-14-02352-v2.pdf Title: Limited Emergence of Salmonella enterica Serovar Infantis Variants with Reduced Phage Susceptibility in PhagoVet-Treated Broilers Subject: Salmonella enterica serovar Infantis (S. Infantis) poses a growing issue in the poultry sector, with phage-based products emerging as a safe and effective control measure. This study investigated the emergence of reduced-phage-susceptibility variants (RPSV) of S. Infantis in PhagoVet-treated broilers, given that RPSV could undermine phage treatment efficacy. The bacteriophages in the PhagoVet product were characterized using transmission electron microscopy (TEM), genome sequencing, and infection profiling. Furthermore, two broiler trials were conducted: a challenge group (T1) and a challenge-and-treated group (T2). The S. Infantis infective dose was set at 104 and 106 colony-forming units (CFUs) per animal, with PhagoVet administration at 106 and 108 plaque-forming units (PFUs) per animal, in Trials 1 and 2, respectively. The results revealed that the four PhagoVet bacteriophages belonged to different genera. PhagoVet evidenced broad-spectrum efficacy against 271 strains representing 18 Salmonella serovars. In Trial 1, PhagoVet reduced bacterial counts in feces to nearly undetectable levels by day 42, with no RPSV detected. However, in Trial 2, three and five RPSVs were detected in feces and ceca, respectively. Consequently, PhagoVet demonstrated efficacy against S. Infantis in broilers, and the potential impact of RPSV is deemed unlikely to compromise its efficacy. Keywords: Salmonella Infantis; broilers; bacteriophage; PhagoVet; reduced phage susceptibility Author: Sandra Sevilla-Navarro, Jennifer Otero, Júlia López-Pérez, Jan Torres-Boncompte, Tiago Prucha, Maarten De Gussem, Daniela Silva, Julia Burgan, Pablo Catalá-Gregori, Pilar Cortés and Montserrat Llagostera Creator: LaTeX with hyperref Producer: pdfTeX-1.40.25 CreationDate: Thu Aug 15 10:40:53 2024 CEST ModDate: Thu Aug 15 10:55:32 2024 CEST Custom Metadata: no Metadata Stream: no Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 12 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 978517 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- YUQXUW+VnURWPalladioL-Bold Type 1 Custom yes yes yes 10 0 YADGVR+URWPalladioL-Bold Type 1 Custom yes yes yes 16 0 BHVYSX+URWPalladioL-Roma Type 1 Custom yes yes yes 22 0 KFCVHH+URWPalladioL-Ital Type 1 Custom yes yes yes 27 0 RPEHUW+URWPalladioL-BoldItal Type 1 Custom yes yes yes 32 0 ARIQJE+VnURWPalladioL Type 1 Custom yes yes yes 37 0 SZMRXJ+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 53 0 IBIKHZ+CMSY10 Type 1 Builtin yes yes yes 73 0 OVNEXW+EURM10 Type 1 Builtin yes yes yes 78 0 GDFKLG+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 105 0 GDFKLH+PalatinoLinotype CID TrueType Identity-H yes yes yes 108 0 GDFKLI+PalatinoLinotype TrueType WinAnsi yes yes no 114 0 GDFKMJ+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 117 0 Jhove (Rel. 1.28.0, 2023-05-18) Date: 2024-10-23 02:40:10 CEST RepresentationInformation: animals-14-02352-v2.pdf ReportingModule: PDF-hul, Rel. 1.12.4 (2023-03-16) LastModified: 2024-10-10 01:35:47 CEST Size: 978517 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 308 FreeObjects: 1 IncrementalUpdates: 0 DocumentCatalog: PageLayout: SinglePage PageMode: UseNone Outlines: Item: Title: Introduction Destination: section.1 Item: Title: Materials and Methods Destination: section.2 Children: Item: Title: Bacterial Strains Destination: subsection.2.1 Item: Title: PhagoVet Product Destination: subsection.2.2 Item: Title: Host Range Determination of the PhagoVet Product Destination: subsection.2.3 Item: Title: Bacteriophage Characterization and Genome Sequencing Destination: subsection.2.4 Item: Title: Isolation of S. Infantis Variants with Reduced Phage Susceptibility Destination: subsection.2.5 Item: Title: Statistical Analysis Destination: subsection.2.6 Item: Title: Results Destination: section.3 Children: Item: Title: Characteristics of PhagoVet Product Destination: subsection.3.1 Item: Title: Identification of S. Infantis Variants with Reduced PhagoVet-Susceptibility Destination: subsection.3.2 Item: Title: Discussion Destination: section.4 Item: Title: Conclusions Destination: section.5 Item: Title: References Destination: appendix.A. Info: Title: Limited Emergence of Salmonella enterica Serovar Infantis Variants with Reduced Phage Susceptibility in PhagoVet-Treated Broilers Author: Sandra Sevilla-Navarro, Jennifer Otero, Júlia López-Pérez, Jan Torres-Boncompte, Tiago Prucha, Maarten De Gussem, Daniela Silva, Julia Burgan, Pablo Catalá-Gregori, Pilar Cortés and Montserrat Llagostera Subject: Salmonella enterica serovar Infantis (S. Infantis) poses a growing issue in the poultry sector, with phage-based products emerging as a safe and effective control measure. This study investigated the emergence of reduced-phage-susceptibility variants (RPSV) of S. Infantis in PhagoVet-treated broilers, given that RPSV could undermine phage treatment efficacy. The bacteriophages in the PhagoVet product were characterized using transmission electron microscopy (TEM), genome sequencing, and infection profiling. Furthermore, two broiler trials were conducted: a challenge group (T1) and a challenge-and-treated group (T2). The S. Infantis infective dose was set at 104 and 106 colony-forming units (CFUs) per animal, with PhagoVet administration at 106 and 108 plaque-forming units (PFUs) per animal, in Trials 1 and 2, respectively. The results revealed that the four PhagoVet bacteriophages belonged to different genera. PhagoVet evidenced broad-spectrum efficacy against 271 strains representing 18 Salmonella serovars. In Trial 1, PhagoVet reduced bacterial counts in feces to nearly undetectable levels by day 42, with no RPSV detected. However, in Trial 2, three and five RPSVs were detected in feces and ceca, respectively. Consequently, PhagoVet demonstrated efficacy against S. Infantis in broilers, and the potential impact of RPSV is deemed unlikely to compromise its efficacy. Keywords: Salmonella Infantis; broilers; bacteriophage; PhagoVet; reduced phage susceptibility Creator: LaTeX with hyperref Producer: pdfTeX-1.40.25 CreationDate: Thu Aug 15 10:40:53 CEST 2024 ModDate: Thu Aug 15 10:55:32 CEST 2024 ID: 0x64f9cff7eb64434b2eb11bd6f322a648, 0x64f9cff7eb64434b2eb11bd6f322a648 Filters: FilterPipeline: FlateDecode Fonts: Type1: Font: BaseFont: YADGVR+URWPalladioL-Bold FontSubset: true FirstChar: 3 LastChar: 134 FontDescriptor: FontName: YADGVR+URWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true ToUnicode: true Font: BaseFont: RPEHUW+URWPalladioL-BoldItal FontSubset: true FirstChar: 83 LastChar: 116 FontDescriptor: FontName: RPEHUW+URWPalladioL-BoldItal Flags: Symbolic FontBBox: -170, -300, 1073, 935 FontFile: true EncodingDictionary: Differences: true ToUnicode: true Font: BaseFont: ARIQJE+VnURWPalladioL FontSubset: true FirstChar: 129 LastChar: 243 FontDescriptor: FontName: ARIQJE+VnURWPalladioL Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: Differences: true ToUnicode: true Font: BaseFont: BHVYSX+URWPalladioL-Roma FontSubset: true FirstChar: 2 LastChar: 252 FontDescriptor: FontName: BHVYSX+URWPalladioL-Roma Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: Differences: true ToUnicode: true Font: BaseFont: IBIKHZ+CMSY10 FontSubset: true FirstChar: 2 LastChar: 20 FontDescriptor: FontName: IBIKHZ+CMSY10 Flags: Symbolic FontBBox: -29, -960, 1116, 775 FontFile: true ToUnicode: true Font: BaseFont: YUQXUW+VnURWPalladioL-Bold FontSubset: true FirstChar: 161 LastChar: 243 FontDescriptor: FontName: YUQXUW+VnURWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true ToUnicode: true Font: BaseFont: KFCVHH+URWPalladioL-Ital FontSubset: true FirstChar: 2 LastChar: 122 FontDescriptor: FontName: KFCVHH+URWPalladioL-Ital Flags: Symbolic FontBBox: -170, -305, 1010, 941 FontFile: true EncodingDictionary: Differences: true ToUnicode: true Font: BaseFont: OVNEXW+EURM10 FontSubset: true FirstChar: 22 LastChar: 22 FontDescriptor: FontName: OVNEXW+EURM10 Flags: Symbolic FontBBox: -32, -243, 1060, 720 FontFile: true ToUnicode: true Pages: Page: Sequence: 1 Annotations: Annotation: Subtype: Link Rect: 34, 305, 152, 314 Annotation: Subtype: Link Rect: 34, 295, 74, 302 Annotation: Subtype: Link Rect: 34, 164, 96, 186 Annotation: Subtype: Link Rect: 125, 90, 154, 99 Annotation: Subtype: Link Rect: 34, 78, 149, 87 Annotation: Subtype: Link Rect: 34, 756, 165, 792 Annotation: Subtype: Link Rect: 515, 756, 560, 792 Annotation: Subtype: Link Rect: 154, 642, 165, 655 Annotation: Subtype: Link Rect: 452, 642, 463, 655 Annotation: Subtype: Link Rect: 208, 629, 219, 643 Annotation: Subtype: Link Rect: 394, 629, 405, 643 Annotation: Subtype: Link Rect: 475, 629, 486, 643 Annotation: Subtype: Link Rect: 116, 17, 251, 28 Annotation: Subtype: Link Rect: 409, 17, 560, 28 Annotation: Subtype: Link Rect: 34, 426, 87, 445 Page: Sequence: 2 Annotations: Annotation: Subtype: Link Rect: 469, 715, 476, 724 ActionDest: 132 Annotation: Subtype: Link Rect: 255, 678, 262, 687 ActionDest: 132 Annotation: Subtype: Link Rect: 377, 627, 384, 636 ActionDest: 132 Annotation: Subtype: Link Rect: 363, 577, 370, 586 ActionDest: 132 Annotation: Subtype: Link Rect: 371, 577, 378, 586 ActionDest: 132 Annotation: Subtype: Link Rect: 381, 577, 388, 586 ActionDest: 132 Annotation: Subtype: Link Rect: 500, 527, 507, 536 ActionDest: 132 Annotation: Subtype: Link Rect: 508, 527, 515, 536 ActionDest: 132 Annotation: Subtype: Link Rect: 518, 527, 525, 536 ActionDest: 132 Annotation: Subtype: Link Rect: 198, 414, 205, 423 ActionDest: 132 Annotation: Subtype: Link Rect: 544, 364, 556, 373 ActionDest: 132 Annotation: Subtype: Link Rect: 220, 326, 232, 335 ActionDest: 135 Annotation: Subtype: Link Rect: 240, 289, 252, 298 ActionDest: 135 Annotation: Subtype: Link Rect: 252, 288, 264, 298 ActionDest: 135 Page: Sequence: 3 Annotations: Annotation: Subtype: Link Rect: 498, 593, 510, 602 ActionDest: 135 Annotation: Subtype: Link Rect: 292, 504, 304, 513 ActionDest: 135 Annotation: Subtype: Link Rect: 377, 454, 389, 463 ActionDest: 135 Annotation: Subtype: Link Rect: 446, 429, 458, 438 ActionDest: 135 Annotation: Subtype: Link Rect: 291, 353, 303, 362 ActionDest: 135 Annotation: Subtype: Link Rect: 168, 326, 277, 338 Annotation: Subtype: Link Rect: 400, 328, 412, 337 ActionDest: 135 Annotation: Subtype: Link Rect: 319, 275, 447, 287 Annotation: Subtype: Link Rect: 289, 265, 301, 275 ActionDest: 135 Annotation: Subtype: Link Rect: 543, 228, 555, 237 ActionDest: 135 Annotation: Subtype: Link Rect: 217, 213, 473, 225 Annotation: Subtype: Link Rect: 255, 203, 267, 212 ActionDest: 135 Annotation: Subtype: Link Rect: 335, 175, 470, 187 Annotation: Subtype: Link Rect: 196, 165, 208, 174 ActionDest: 135 Annotation: Subtype: Link Rect: 470, 165, 482, 174 ActionDest: 135 Annotation: Subtype: Link Rect: 210, 153, 222, 162 ActionDest: 135 Annotation: Subtype: Link Rect: 338, 140, 350, 149 ActionDest: 135 Page: Sequence: 4 Annotations: Annotation: Subtype: Link Rect: 273, 618, 285, 627 ActionDest: 135 Annotation: Subtype: Link Rect: 195, 317, 207, 326 ActionDest: 135 Annotation: Subtype: Link Rect: 516, 291, 528, 301 ActionDest: 135 Page: Sequence: 5 Annotations: Annotation: Subtype: Link Rect: 202, 681, 214, 690 ActionDest: 135 Annotation: Subtype: Link Rect: 486, 617, 498, 627 ActionDest: 135 Annotation: Subtype: Link Rect: 531, 478, 543, 487 ActionDest: 135 Annotation: Subtype: Link Rect: 543, 478, 555, 487 ActionDest: 135 Annotation: Subtype: Link Rect: 246, 451, 252, 463 ActionDest: 89 Annotation: Subtype: Link Rect: 418, 353, 430, 362 ActionDest: 135 Annotation: Subtype: Link Rect: 523, 262, 530, 274 ActionDest: 86 Page: Sequence: 6 Page: Sequence: 7 Annotations: Annotation: Subtype: Link Rect: 280, 741, 287, 754 ActionDest: 123 Annotation: Subtype: Link Rect: 456, 665, 463, 677 ActionDest: 123 Annotation: Subtype: Link Rect: 296, 362, 303, 374 ActionDest: 123 Annotation: Subtype: Link Rect: 299, 349, 306, 361 ActionDest: 123 Annotation: Subtype: Link Rect: 501, 349, 508, 361 ActionDest: 123 Annotation: Subtype: Link Rect: 239, 287, 246, 299 ActionDest: 126 Page: Sequence: 8 Annotations: Annotation: Subtype: Link Rect: 401, 394, 408, 406 ActionDest: 89 Annotation: Subtype: Link Rect: 415, 396, 427, 405 ActionDest: 135 Annotation: Subtype: Link Rect: 277, 371, 289, 380 ActionDest: 135 Annotation: Subtype: Link Rect: 290, 334, 302, 343 ActionDest: 135 Annotation: Subtype: Link Rect: 529, 318, 536, 330 ActionDest: 86 Annotation: Subtype: Link Rect: 286, 256, 293, 268 ActionDest: 123 Annotation: Subtype: Link Rect: 426, 230, 433, 242 ActionDest: 123 Annotation: Subtype: Link Rect: 242, 208, 254, 217 ActionDest: 135 Annotation: Subtype: Link Rect: 255, 208, 267, 217 ActionDest: 135 Annotation: Subtype: Link Rect: 283, 108, 295, 117 ActionDest: 135 Annotation: Subtype: Link Rect: 529, 70, 541, 79 ActionDest: 139 Annotation: Subtype: Link Rect: 544, 70, 556, 79 ActionDest: 139 Page: Sequence: 9 Annotations: Annotation: Subtype: Link Rect: 531, 718, 543, 727 ActionDest: 135 Annotation: Subtype: Link Rect: 543, 718, 555, 727 ActionDest: 135 Annotation: Subtype: Link Rect: 480, 640, 487, 652 ActionDest: 126 Annotation: Subtype: Link Rect: 524, 530, 536, 539 ActionDest: 135 Annotation: Subtype: Link Rect: 425, 467, 437, 476 ActionDest: 139 Annotation: Subtype: Link Rect: 247, 430, 259, 439 ActionDest: 139 Annotation: Subtype: Link Rect: 388, 404, 400, 414 ActionDest: 135 Annotation: Subtype: Link Rect: 272, 367, 283, 376 ActionDest: 135 Annotation: Subtype: Link Rect: 284, 367, 296, 376 ActionDest: 135 Annotation: Subtype: Link Rect: 297, 367, 308, 376 ActionDest: 139 Annotation: Subtype: Link Rect: 309, 367, 321, 376 ActionDest: 139 Annotation: Subtype: Link Rect: 289, 342, 301, 351 ActionDest: 139 Annotation: Subtype: Link Rect: 451, 317, 463, 326 ActionDest: 139 Annotation: Subtype: Link Rect: 525, 266, 537, 275 ActionDest: 135 Annotation: Subtype: Link Rect: 290, 254, 302, 263 ActionDest: 135 Annotation: Subtype: Link Rect: 256, 241, 268, 250 ActionDest: 139 Annotation: Subtype: Link Rect: 268, 241, 280, 250 ActionDest: 139 Annotation: Subtype: Link Rect: 295, 229, 307, 238 ActionDest: 139 Page: Sequence: 10 Annotations: Annotation: Subtype: Link Rect: 536, 741, 560, 752 Annotation: Subtype: Link Rect: 165, 729, 369, 740 Annotation: Subtype: Link Rect: 230, 357, 268, 368 Annotation: Subtype: Link Rect: 275, 334, 312, 345 Annotation: Subtype: Link Rect: 251, 299, 288, 310 Annotation: Subtype: Link Rect: 400, 276, 437, 287 Annotation: Subtype: Link Rect: 443, 276, 480, 287 Annotation: Subtype: Link Rect: 151, 242, 188, 253 Annotation: Subtype: Link Rect: 244, 207, 282, 218 Annotation: Subtype: Link Rect: 131, 173, 168, 183 Annotation: Subtype: Link Rect: 175, 173, 211, 183 Annotation: Subtype: Link Rect: 462, 138, 500, 149 Annotation: Subtype: Link Rect: 211, 103, 248, 114 Annotation: Subtype: Link Rect: 271, 80, 308, 91 Page: Sequence: 11 Annotations: Annotation: Subtype: Link Rect: 238, 695, 276, 706 Annotation: Subtype: Link Rect: 282, 695, 318, 706 Annotation: Subtype: Link Rect: 259, 673, 297, 683 Annotation: Subtype: Link Rect: 303, 673, 339, 683 Annotation: Subtype: Link Rect: 419, 660, 457, 671 Annotation: Subtype: Link Rect: 463, 660, 499, 671 Annotation: Subtype: Link Rect: 333, 637, 370, 648 Annotation: Subtype: Link Rect: 376, 637, 412, 648 Annotation: Subtype: Link Rect: 59, 592, 96, 602 Annotation: Subtype: Link Rect: 102, 592, 138, 602 Annotation: Subtype: Link Rect: 109, 558, 147, 568 Annotation: Subtype: Link Rect: 143, 535, 181, 545 Annotation: Subtype: Link Rect: 59, 512, 96, 522 Annotation: Subtype: Link Rect: 102, 512, 138, 522 Annotation: Subtype: Link Rect: 350, 476, 388, 487 Annotation: Subtype: Link Rect: 216, 454, 253, 464 Annotation: Subtype: Link Rect: 259, 454, 295, 464 Annotation: Subtype: Link Rect: 519, 430, 557, 441 Annotation: Subtype: Link Rect: 59, 420, 95, 430 Annotation: Subtype: Link Rect: 145, 385, 182, 395 Annotation: Subtype: Link Rect: 180, 362, 218, 372 Annotation: Subtype: Link Rect: 56, 327, 249, 338 Annotation: Subtype: Link Rect: 436, 304, 474, 315 Annotation: Subtype: Link Rect: 59, 270, 96, 280 Annotation: Subtype: Link Rect: 148, 235, 324, 246 Annotation: Subtype: Link Rect: 353, 212, 391, 223 Annotation: Subtype: Link Rect: 234, 190, 272, 200 Annotation: Subtype: Link Rect: 519, 166, 557, 177 Annotation: Subtype: Link Rect: 59, 155, 95, 165 Annotation: Subtype: Link Rect: 95, 121, 133, 131 Annotation: Subtype: Link Rect: 323, 85, 361, 96 Annotation: Subtype: Link Rect: 367, 85, 403, 96 Page: Sequence: 12 Annotations: Annotation: Subtype: Link Rect: 443, 730, 480, 741 Annotation: Subtype: Link Rect: 486, 730, 523, 741 Annotation: Subtype: Link Rect: 384, 706, 421, 718 Annotation: Subtype: Link Rect: 427, 706, 463, 718 Annotation: Subtype: Link Rect: 287, 683, 325, 694 Annotation: Subtype: Link Rect: 331, 683, 367, 694 Annotation: Subtype: Link Rect: 520, 672, 557, 683 Annotation: Subtype: Link Rect: 59, 638, 96, 648 Annotation: Subtype: Link Rect: 112, 604, 150, 614 Checksum: f1e5439d Type: CRC32 Checksum: 37d578eca0472628f10360122c306cf6 Type: MD5 Checksum: f71090e9de41dc593f17710af36aa4e4edf71e9a Type: SHA-1 Checksum: 5d43a24a77211a0488a034defd5900f246b7d811f3aaa1a166296b8394290751 Type: SHA-256