ab6fdf7202a1a104e667db2067610249 pmc_10255704.pdf 473522635c54ee0b29e852e330cf9dca59834f1f pmc_10255704.pdf 81d1ef88321d2644e3f54df031dabf3599b9b7040f77435198dc3b11ba2895bc pmc_10255704.pdf Title: Repeatedly Northwards and Upwards: Southern African Grasslands Fuel the Colonization of the African Sky Islands in Helichrysum (Compositae) Subject: The Afromontane and Afroalpine areas constitute some of the main biodiversity hotspots of Africa. They are particularly rich in plant endemics, but the biogeographic origins and evolutionary processes leading to this outstanding diversity are poorly understood. We performed phylogenomic and biogeographic analyses of one of the most species-rich plant genera in these mountains, Helichrysum (Compositae-Gnaphalieae). Most previous studies have focused on Afroalpine elements of Eurasian origin, and the southern African origin of Helichrysum provides an interesting counterexample. We obtained a comprehensive nuclear dataset from 304 species (50% of the genus) using target-enrichment with the Compositae1061 probe set. Summary-coalescent and concatenation approaches combined with paralog recovery yielded congruent, well-resolved phylogenies. Ancestral range estimations revealed that Helichrysum originated in arid southern Africa, whereas the southern African grasslands were the source of most lineages that dispersed within and outside Africa. Colonization of the tropical Afromontane and Afroalpine areas occurred repeatedly throughout the Miocene–Pliocene. This timing coincides with mountain uplift and the onset of glacial cycles, which together may have facilitated both speciation and intermountain gene flow, contributing to the evolution of the Afroalpine flora.+12pt Keywords: Afroalpine; Afromontane; biogeography; Asteraceae; long-distance dispersal; evolution; Helichrysum; phylogeny; target-enrichment Author: Carme Blanco-Gavaldà, Mercè Galbany-Casals, Alfonso Susanna, Santiago Andrés-Sánchez, Randall J. Bayer, Christian Brochmann, Glynis V. Cron, Nicola G. Bergh, Núria Garcia-Jacas, Abel Gizaw, Martha Kandziora, Filip Kolář, Javier López-Alvarado, Frederik Leliaert, Rokiman Letsara, Lucía D. Moreyra, Sylvain G. Razafimandimbison, Roswitha Schmickl and Cristina Roquet Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Thu Jun 8 13:47:50 2023 CEST ModDate: Thu Jun 8 13:50:37 2023 CEST Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 24 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 3034381 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- IWTZGU+VnURWPalladioL Type 1 Custom yes yes no 10 0 RYVCUX+VnURWPalladioL-Italic Type 1 Custom yes yes no 15 0 MFOSSD+URWPalladioL-Bold Type 1 Custom yes yes no 19 0 GINKHZ+URWPalladioL-Roma Type 1 Custom yes yes no 24 0 ZGZDAB+URWPalladioL-Ital Type 1 Custom yes yes no 28 0 ZHDWOE+URWPalladioL-BoldItal Type 1 Custom yes yes no 32 0 EMTPGF+CMSY10 Type 1 Builtin yes yes no 36 0 CKPGFH+VnURWPalladioL-Bold Type 1 Custom yes yes no 40 0 VTLKDP+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 59 0 NLLKAB+PalatinoLinotype TrueType MacRoman yes yes no 102 0 NLLJDN+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 105 0 NLLJDO+PalatinoLinotype CID TrueType Identity-H yes yes yes 108 0 NLLJDP+PalatinoLinotype TrueType WinAnsi yes yes no 114 0 NLLJEB+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 117 0 OVNEXW+EURM10 Type 1 Builtin yes yes no 124 0 NLLKAB+PalatinoLinotype TrueType MacRoman yes yes no 141 0 NLLJDN+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 144 0 NLLJDO+PalatinoLinotype CID TrueType Identity-H yes yes yes 147 0 NLLJDP+PalatinoLinotype TrueType WinAnsi yes yes no 153 0 NLLJEB+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 156 0 EVTBZF+ArialMT CID TrueType Identity-H yes yes yes 367 0 IBRZDX+MyriadVariableConcept-Roman Type 1C Custom yes yes no 375 0 SDPHND+MyriadPro-Regular Type 1C WinAnsi yes yes no 379 0 EVTBZF+ArialMT TrueType WinAnsi yes yes no 382 0 EVTBZF+ArialNarrow TrueType WinAnsi yes yes no 385 0 SDPHND+Arial-ItalicMT TrueType WinAnsi yes yes no 388 0 EVTBZF+ArialMT TrueType WinAnsi yes yes no 391 0 SDPHND+Arial-BoldMT TrueType WinAnsi yes yes no 392 0 EVTBZF+ArialNarrow TrueType WinAnsi yes yes no 395 0 NLLKAB+PalatinoLinotype TrueType MacRoman yes yes no 478 0 NLLJDN+PalatinoLinotype,Bold TrueType WinAnsi yes yes no 481 0 NLLJDO+PalatinoLinotype CID TrueType Identity-H yes yes yes 484 0 NLLJDP+PalatinoLinotype TrueType WinAnsi yes yes no 490 0 NLLJEB+PalatinoLinotype,Italic TrueType WinAnsi yes yes no 493 0 Jhove (Rel. 1.28.0, 2023-05-18) Date: 2024-02-02 02:33:58 CET RepresentationInformation: pmc_10255704.pdf ReportingModule: PDF-hul, Rel. 1.12.4 (2023-03-16) LastModified: 2023-09-16 02:17:19 CEST Size: 3034381 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 923 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: Taxon Sampling Destination: subsection.2.1 Item: Title: DNA Extraction, Library Preparation, Target Capture, and Sequencing Destination: subsection.2.2 Item: Title: Molecular Data Processing and Phylogenetic Analyses Destination: subsection.2.3 Item: Title: Divergence Time Estimation Destination: subsection.2.4 Item: Title: Ancestral Range Estimation Destination: subsection.2.5 Item: Title: Results Destination: section.3 Children: Item: Title: Alignment Processing and Filtering Destination: subsection.3.1 Item: Title: Phylogenetic Analyses Destination: subsection.3.2 Item: Title: Divergence Time and Ancestral Range Estimation Destination: subsection.3.3 Item: Title: Number, Type, and Directionality Estimation of Biogeographical Events Destination: subsection.3.4 Item: Title: Discussion Destination: section.4 Children: Item: Title: Utility of Target-Enrichment Strategies in Reconstructing the Radiation of Helichrysum Destination: subsection.4.1 Item: Title: The Early History of Helichrysum and Colonization of Madagascar Destination: subsection.4.2 Item: Title: Repeatedly Northwards Destination: subsection.4.3 Item: Title: Repeatedly Upwards Destination: subsection.4.4 Item: Title: Conclusions Destination: section.5 Item: Title: References Destination: section.A. Info: Title: Repeatedly Northwards and Upwards: Southern African Grasslands Fuel the Colonization of the African Sky Islands in Helichrysum (Compositae) Author: ぃちひねづ〠あぬちのっは〭ぇちぶちぬつム〬〠きづひっヨ〠ぇちぬぢちのべ〭ぃちびちぬび〬〠ぁぬてはのびは〠こふびちののち〬〠こちのぴどちでは〠ぁのつひラび〭こメのっとづぺ〬〠げちのつちぬぬ〠お〮〠あちべづひ〬〠ぃとひどびぴどちの〠あひはっとねちのの〬〠ぇぬべのどび〠ざ〮〠ぃひはの〬〠ぎどっはぬち〠ぇ〮〠あづひでと〬〠ぎヺひどち〠ぇちひっどち〭おちっちび〬〠ぁぢづぬ〠ぇどぺちぷ〬〠きちひぴとち〠かちのつぺどはひち〬〠うどぬどば〠かはぬメㅙ〬〠おちぶどづひ〠がンばづぺ〭ぁぬぶちひちつは〬〠うひづつづひどに〠がづぬどちづひぴ〬〠げはにどねちの〠がづぴびちひち〬〠がふっロち〠い〮〠きはひづべひち〬〠こべぬぶちどの〠ぇ〮〠げちぺちてどねちのつどねぢどびはの〬〠げはびぷどぴとち〠こっとねどっにぬ〠ちのつ〠ぃひどびぴどのち〠げはぱふづぴ Subject: The Afromontane and Afroalpine areas constitute some of the main biodiversity hotspots of Africa. They are particularly rich in plant endemics, but the biogeographic origins and evolutionary processes leading to this outstanding diversity are poorly understood. We performed phylogenomic and biogeographic analyses of one of the most species-rich plant genera in these mountains, Helichrysum (Compositae-Gnaphalieae). Most previous studies have focused on Afroalpine elements of Eurasian origin, and the southern African origin of Helichrysum provides an interesting counterexample. We obtained a comprehensive nuclear dataset from 304 species (50% of the genus) using target-enrichment with the Compositae1061 probe set. Summary-coalescent and concatenation approaches combined with paralog recovery yielded congruent, well-resolved phylogenies. Ancestral range estimations revealed that Helichrysum originated in arid southern Africa, whereas the southern African grasslands were the source of most lineages that dispersed within and outside Africa. Colonization of the tropical Afromontane and Afroalpine areas occurred repeatedly throughout the Miocene Pliocene. This timing coincides with mountain uplift and the onset of glacial cycles, which together may have facilitated both speciation and intermountain gene flow, contributing to the evolution of the Afroalpine flora.+12pt Keywords: Afroalpine; Afromontane; biogeography; Asteraceae; long-distance dispersal; evolution; Helichrysum; phylogeny; target-enrichment Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Thu Jun 08 13:47:50 CEST 2023 ModDate: Thu Jun 08 13:50:37 CEST 2023 ID: 0x6ccce3cbfef16b2f4c4505b75d424708, 0x6ccce3cbfef16b2f4c4505b75d424708 Filters: FilterPipeline: FlateDecode Fonts: Type1: Font: BaseFont: ZHDWOE+URWPalladioL-BoldItal FontSubset: true FirstChar: 72 LastChar: 121 FontDescriptor: FontName: ZHDWOE+URWPalladioL-BoldItal Flags: Symbolic FontBBox: -170, -300, 1073, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: MFOSSD+URWPalladioL-Bold FontSubset: true FirstChar: 2 LastChar: 134 FontDescriptor: FontName: MFOSSD+URWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: EMTPGF+CMSY10 FontSubset: true FirstChar: 0 LastChar: 25 FontDescriptor: FontName: EMTPGF+CMSY10 Flags: Symbolic FontBBox: -29, -960, 1116, 775 FontFile: true Font: BaseFont: CKPGFH+VnURWPalladioL-Bold FontSubset: true FirstChar: 160 LastChar: 243 FontDescriptor: FontName: CKPGFH+VnURWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: GINKHZ+URWPalladioL-Roma FontSubset: true FirstChar: 2 LastChar: 252 FontDescriptor: FontName: GINKHZ+URWPalladioL-Roma Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: IWTZGU+VnURWPalladioL FontSubset: true FirstChar: 161 LastChar: 254 FontDescriptor: FontName: IWTZGU+VnURWPalladioL Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: ZGZDAB+URWPalladioL-Ital FontSubset: true FirstChar: 3 LastChar: 150 FontDescriptor: FontName: ZGZDAB+URWPalladioL-Ital Flags: Symbolic FontBBox: -170, -305, 1010, 941 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: OVNEXW+EURM10 FontSubset: true FirstChar: 22 LastChar: 22 FontDescriptor: FontName: OVNEXW+EURM10 Flags: Symbolic FontBBox: -32, -243, 1060, 720 FontFile: true Font: BaseFont: RYVCUX+VnURWPalladioL-Italic FontSubset: true FirstChar: 243 LastChar: 243 FontDescriptor: FontName: RYVCUX+VnURWPalladioL-Italic Flags: Symbolic FontBBox: -170, -305, 1010, 941 FontFile: true EncodingDictionary: Differences: true Pages: Page: Sequence: 1 Annotations: Annotation: Subtype: Link Rect: 82, 292, 152, 301 Annotation: Subtype: Link Rect: 34, 280, 110, 289 Annotation: Subtype: Link Rect: 34, 164, 96, 186 Annotation: Subtype: Link Rect: 126, 90, 154, 99 Annotation: Subtype: Link Rect: 34, 78, 147, 87 Annotation: Subtype: Link Rect: 34, 756, 142, 792 Annotation: Subtype: Link Rect: 515, 756, 560, 792 Annotation: Subtype: Link Rect: 153, 641, 164, 655 Annotation: Subtype: Link Rect: 366, 641, 377, 655 Annotation: Subtype: Link Rect: 503, 641, 514, 655 Annotation: Subtype: Link Rect: 224, 629, 235, 642 Annotation: Subtype: Link Rect: 500, 629, 511, 642 Annotation: Subtype: Link Rect: 102, 616, 113, 630 Annotation: Subtype: Link Rect: 392, 616, 403, 630 Annotation: Subtype: Link Rect: 492, 616, 503, 630 Annotation: Subtype: Link Rect: 213, 604, 224, 617 Annotation: Subtype: Link Rect: 380, 604, 391, 617 Annotation: Subtype: Link Rect: 109, 17, 255, 28 Annotation: Subtype: Link Rect: 415, 17, 560, 28 Annotation: Subtype: Link Rect: 34, 425, 87, 444 Page: Sequence: 2 Annotations: Annotation: Subtype: Link Rect: 256, 632, 263, 641 ActionDest: 515 Annotation: Subtype: Link Rect: 334, 582, 341, 591 ActionDest: 515 Annotation: Subtype: Link Rect: 344, 582, 351, 591 ActionDest: 515 Annotation: Subtype: Link Rect: 369, 557, 376, 565 ActionDest: 515 Annotation: Subtype: Link Rect: 379, 556, 386, 565 ActionDest: 515 Annotation: Subtype: Link Rect: 296, 544, 303, 553 ActionDest: 515 Annotation: Subtype: Link Rect: 296, 506, 303, 515 ActionDest: 515 Annotation: Subtype: Link Rect: 303, 506, 310, 515 ActionDest: 515 Annotation: Subtype: Link Rect: 474, 506, 481, 515 ActionDest: 515 Annotation: Subtype: Link Rect: 291, 418, 303, 427 ActionDest: 515 Annotation: Subtype: Link Rect: 536, 406, 543, 415 ActionDest: 515 Annotation: Subtype: Link Rect: 544, 406, 556, 415 ActionDest: 515 Annotation: Subtype: Link Rect: 271, 381, 283, 390 ActionDest: 518 Annotation: Subtype: Link Rect: 496, 305, 503, 314 ActionDest: 515 Annotation: Subtype: Link Rect: 185, 268, 197, 277 ActionDest: 518 Annotation: Subtype: Link Rect: 236, 243, 248, 252 ActionDest: 518 Annotation: Subtype: Link Rect: 402, 230, 409, 239 ActionDest: 515 Annotation: Subtype: Link Rect: 524, 205, 531, 214 ActionDest: 515 Annotation: Subtype: Link Rect: 531, 205, 543, 214 ActionDest: 518 Annotation: Subtype: Link Rect: 544, 205, 556, 214 ActionDest: 518 Annotation: Subtype: Link Rect: 446, 167, 458, 176 ActionDest: 518 Annotation: Subtype: Link Rect: 461, 167, 473, 176 ActionDest: 518 Annotation: Subtype: Link Rect: 480, 155, 492, 164 ActionDest: 518 Annotation: Subtype: Link Rect: 461, 117, 473, 126 ActionDest: 518 Annotation: Subtype: Link Rect: 474, 117, 486, 126 ActionDest: 518 Annotation: Subtype: Link Rect: 314, 92, 326, 101 ActionDest: 518 Annotation: Subtype: Link Rect: 326, 92, 338, 101 ActionDest: 518 Page: Sequence: 3 Annotations: Annotation: Subtype: Link Rect: 543, 736, 555, 744 ActionDest: 518 Annotation: Subtype: Link Rect: 435, 723, 447, 732 ActionDest: 518 Annotation: Subtype: Link Rect: 503, 698, 515, 707 ActionDest: 518 Annotation: Subtype: Link Rect: 516, 698, 527, 707 ActionDest: 518 Annotation: Subtype: Link Rect: 288, 673, 300, 682 ActionDest: 518 Annotation: Subtype: Link Rect: 401, 635, 413, 644 ActionDest: 518 Annotation: Subtype: Link Rect: 255, 610, 267, 619 ActionDest: 518 Annotation: Subtype: Link Rect: 434, 544, 441, 556 ActionDest: 92 Annotation: Subtype: Link Rect: 231, 522, 243, 531 ActionDest: 518 Annotation: Subtype: Link Rect: 246, 522, 258, 531 ActionDest: 518 Annotation: Subtype: Link Rect: 395, 497, 407, 506 ActionDest: 518 Annotation: Subtype: Link Rect: 247, 434, 259, 443 ActionDest: 518 Annotation: Subtype: Link Rect: 450, 397, 462, 406 ActionDest: 518 Annotation: Subtype: Link Rect: 463, 396, 475, 406 ActionDest: 518 Annotation: Subtype: Link Rect: 475, 396, 487, 406 ActionDest: 518 Annotation: Subtype: Link Rect: 441, 346, 453, 355 ActionDest: 518 Annotation: Subtype: Link Rect: 454, 346, 465, 355 ActionDest: 518 Annotation: Subtype: Link Rect: 193, 321, 205, 330 ActionDest: 518 Annotation: Subtype: Link Rect: 191, 271, 203, 280 ActionDest: 518 Annotation: Subtype: Link Rect: 545, 183, 557, 192 ActionDest: 518 Annotation: Subtype: Link Rect: 323, 170, 335, 180 ActionDest: 518 Annotation: Subtype: Link Rect: 455, 158, 467, 167 ActionDest: 518 Page: Sequence: 4 Annotations: Annotation: Subtype: Link Rect: 464, 138, 476, 146 ActionDest: 518 Annotation: Subtype: Link Rect: 194, 112, 206, 121 ActionDest: 518 Annotation: Subtype: Link Rect: 207, 112, 219, 121 ActionDest: 518 Annotation: Subtype: Link Rect: 377, 75, 389, 84 ActionDest: 518 Annotation: Subtype: Link Rect: 389, 75, 401, 84 ActionDest: 518 Page: Sequence: 5 Annotations: Annotation: Subtype: Link Rect: 290, 673, 302, 682 ActionDest: 518 Annotation: Subtype: Link Rect: 273, 647, 285, 657 ActionDest: 518 Annotation: Subtype: Link Rect: 285, 647, 297, 657 ActionDest: 518 Annotation: Subtype: Link Rect: 298, 647, 310, 657 ActionDest: 518 Annotation: Subtype: Link Rect: 313, 647, 325, 657 ActionDest: 521 Annotation: Subtype: Link Rect: 426, 258, 438, 267 ActionDest: 521 Annotation: Subtype: Link Rect: 438, 258, 450, 267 ActionDest: 521 Annotation: Subtype: Link Rect: 416, 245, 428, 254 ActionDest: 521 Annotation: Subtype: Link Rect: 489, 233, 501, 242 ActionDest: 518 Annotation: Subtype: Link Rect: 165, 217, 367, 230 Annotation: Subtype: Link Rect: 269, 157, 281, 166 ActionDest: 521 Annotation: Subtype: Link Rect: 340, 155, 560, 167 Annotation: Subtype: Link Rect: 326, 107, 338, 116 ActionDest: 521 Annotation: Subtype: Link Rect: 545, 107, 557, 116 ActionDest: 521 Annotation: Subtype: Link Rect: 209, 82, 221, 91 ActionDest: 518 Page: Sequence: 6 Annotations: Annotation: Subtype: Link Rect: 360, 597, 371, 606 ActionDest: 521 Annotation: Subtype: Link Rect: 324, 585, 336, 594 ActionDest: 518 Annotation: Subtype: Link Rect: 238, 572, 250, 581 ActionDest: 521 Annotation: Subtype: Link Rect: 466, 522, 478, 531 ActionDest: 521 Annotation: Subtype: Link Rect: 187, 484, 199, 493 ActionDest: 521 Annotation: Subtype: Link Rect: 412, 472, 424, 481 ActionDest: 521 Annotation: Subtype: Link Rect: 189, 422, 201, 431 ActionDest: 521 Annotation: Subtype: Link Rect: 395, 422, 407, 431 ActionDest: 521 Annotation: Subtype: Link Rect: 366, 396, 378, 406 ActionDest: 521 Annotation: Subtype: Link Rect: 313, 371, 325, 380 ActionDest: 521 Annotation: Subtype: Link Rect: 545, 359, 556, 368 ActionDest: 521 Annotation: Subtype: Link Rect: 339, 334, 351, 343 ActionDest: 525 Annotation: Subtype: Link Rect: 448, 321, 460, 330 ActionDest: 525 Annotation: Subtype: Link Rect: 380, 283, 392, 292 ActionDest: 525 Annotation: Subtype: Link Rect: 192, 271, 204, 280 ActionDest: 525 Annotation: Subtype: Link Rect: 374, 258, 386, 267 ActionDest: 525 Annotation: Subtype: Link Rect: 396, 220, 408, 229 ActionDest: 518 Annotation: Subtype: Link Rect: 531, 183, 543, 192 ActionDest: 525 Annotation: Subtype: Link Rect: 543, 183, 555, 192 ActionDest: 525 Annotation: Subtype: Link Rect: 543, 158, 555, 167 ActionDest: 525 Annotation: Subtype: Link Rect: 342, 133, 354, 142 ActionDest: 518 Page: Sequence: 7 Annotations: Annotation: Subtype: Link Rect: 444, 707, 456, 716 ActionDest: 518 Annotation: Subtype: Link Rect: 456, 707, 468, 716 ActionDest: 518 Annotation: Subtype: Link Rect: 469, 707, 481, 716 ActionDest: 518 Annotation: Subtype: Link Rect: 481, 707, 493, 716 ActionDest: 518 Annotation: Subtype: Link Rect: 496, 707, 508, 716 ActionDest: 521 Annotation: Subtype: Link Rect: 234, 667, 241, 679 ActionDest: 131 Annotation: Subtype: Link Rect: 211, 657, 218, 666 ActionDest: 515 Annotation: Subtype: Link Rect: 471, 190, 477, 203 ActionDest: 163 Annotation: Subtype: Link Rect: 399, 160, 406, 169 ActionDest: 515 Annotation: Subtype: Link Rect: 447, 147, 454, 156 ActionDest: 515 Page: Sequence: 8 Annotations: Annotation: Subtype: Link Rect: 255, 685, 262, 694 ActionDest: 515 Annotation: Subtype: Link Rect: 263, 685, 275, 694 ActionDest: 518 Annotation: Subtype: Link Rect: 345, 673, 357, 682 ActionDest: 525 Annotation: Subtype: Link Rect: 357, 673, 369, 682 ActionDest: 525 Annotation: Subtype: Link Rect: 255, 660, 267, 669 ActionDest: 525 Annotation: Subtype: Link Rect: 267, 660, 279, 669 ActionDest: 525 Annotation: Subtype: Link Rect: 267, 623, 274, 631 ActionDest: 515 Annotation: Subtype: Link Rect: 315, 547, 327, 556 ActionDest: 525 Annotation: Subtype: Link Rect: 327, 547, 339, 556 ActionDest: 525 Annotation: Subtype: Link Rect: 218, 419, 413, 431 Annotation: Subtype: Link Rect: 545, 422, 557, 431 ActionDest: 525 Annotation: Subtype: Link Rect: 442, 371, 454, 380 ActionDest: 525 Annotation: Subtype: Link Rect: 455, 371, 467, 380 ActionDest: 525 Annotation: Subtype: Link Rect: 523, 359, 535, 368 ActionDest: 525 Annotation: Subtype: Link Rect: 232, 346, 244, 355 ActionDest: 525 Annotation: Subtype: Link Rect: 196, 283, 208, 293 ActionDest: 525 Annotation: Subtype: Link Rect: 181, 258, 193, 267 ActionDest: 525 Page: Sequence: 9 Page: Sequence: 10 Annotations: Annotation: Subtype: Link Rect: 282, 279, 289, 291 ActionDest: 163 Annotation: Subtype: Link Rect: 287, 193, 299, 202 ActionDest: 525 Annotation: Subtype: Link Rect: 197, 168, 209, 177 ActionDest: 525 Annotation: Subtype: Link Rect: 386, 115, 393, 127 ActionDest: 163 Page: Sequence: 11 Annotations: Annotation: Subtype: Link Rect: 544, 698, 556, 707 ActionDest: 525 Annotation: Subtype: Link Rect: 519, 685, 531, 694 ActionDest: 525 Annotation: Subtype: Link Rect: 455, 557, 462, 569 ActionDest: 163 Annotation: Subtype: Link Rect: 225, 394, 233, 406 ActionDest: 468 Annotation: Subtype: Link Rect: 278, 344, 285, 356 ActionDest: 163 Page: Sequence: 12 Page: Sequence: 13 Annotations: Annotation: Subtype: Link Rect: 493, 544, 500, 556 ActionDest: 497 Annotation: Subtype: Link Rect: 165, 417, 325, 429 Page: Sequence: 14 Annotations: Annotation: Subtype: Link Rect: 202, 710, 213, 719 ActionDest: 525 Annotation: Subtype: Link Rect: 324, 673, 336, 682 ActionDest: 525 Annotation: Subtype: Link Rect: 405, 673, 417, 682 ActionDest: 525 Annotation: Subtype: Link Rect: 485, 673, 497, 682 ActionDest: 525 Annotation: Subtype: Link Rect: 199, 660, 210, 669 ActionDest: 525 Annotation: Subtype: Link Rect: 526, 647, 538, 657 ActionDest: 518 Annotation: Subtype: Link Rect: 318, 522, 330, 531 ActionDest: 518 Annotation: Subtype: Link Rect: 268, 246, 280, 255 ActionDest: 525 Annotation: Subtype: Link Rect: 337, 208, 349, 217 ActionDest: 525 Annotation: Subtype: Link Rect: 409, 196, 420, 205 ActionDest: 525 Annotation: Subtype: Link Rect: 544, 183, 556, 192 ActionDest: 528 Annotation: Subtype: Link Rect: 425, 80, 433, 92 ActionDest: 163 Annotation: Subtype: Link Rect: 350, 70, 362, 79 ActionDest: 528 Page: Sequence: 15 Annotations: Annotation: Subtype: Link Rect: 484, 723, 496, 732 ActionDest: 528 Annotation: Subtype: Link Rect: 496, 723, 513, 732 ActionDest: 528 Annotation: Subtype: Link Rect: 471, 698, 483, 707 ActionDest: 518 Annotation: Subtype: Link Rect: 488, 510, 500, 519 ActionDest: 521 Annotation: Subtype: Link Rect: 500, 509, 517, 519 ActionDest: 528 Annotation: Subtype: Link Rect: 520, 509, 537, 519 ActionDest: 528 Annotation: Subtype: Link Rect: 190, 484, 207, 493 ActionDest: 528 Annotation: Subtype: Link Rect: 238, 472, 255, 481 ActionDest: 528 Annotation: Subtype: Link Rect: 327, 472, 344, 481 ActionDest: 528 Annotation: Subtype: Link Rect: 394, 447, 411, 456 ActionDest: 528 Annotation: Subtype: Link Rect: 448, 344, 455, 356 ActionDest: 468 Annotation: Subtype: Link Rect: 420, 309, 432, 318 ActionDest: 518 Annotation: Subtype: Link Rect: 386, 268, 393, 280 ActionDest: 468 Annotation: Subtype: Link Rect: 543, 246, 555, 255 ActionDest: 525 Annotation: Subtype: Link Rect: 320, 221, 337, 230 ActionDest: 528 Annotation: Subtype: Link Rect: 346, 92, 353, 104 ActionDest: 468 Page: Sequence: 16 Annotations: Annotation: Subtype: Link Rect: 250, 735, 267, 744 ActionDest: 528 Annotation: Subtype: Link Rect: 531, 735, 543, 744 ActionDest: 518 Annotation: Subtype: Link Rect: 544, 735, 556, 744 ActionDest: 518 Annotation: Subtype: Link Rect: 378, 710, 390, 719 ActionDest: 518 Annotation: Subtype: Link Rect: 393, 710, 405, 719 ActionDest: 518 Annotation: Subtype: Link Rect: 543, 685, 555, 694 ActionDest: 518 Annotation: Subtype: Link Rect: 203, 635, 210, 644 ActionDest: 515 Annotation: Subtype: Link Rect: 341, 610, 348, 619 ActionDest: 515 Annotation: Subtype: Link Rect: 349, 610, 361, 619 ActionDest: 525 Annotation: Subtype: Link Rect: 463, 597, 474, 606 ActionDest: 525 Annotation: Subtype: Link Rect: 475, 597, 492, 606 ActionDest: 528 Annotation: Subtype: Link Rect: 461, 572, 478, 581 ActionDest: 528 Annotation: Subtype: Link Rect: 185, 484, 197, 493 ActionDest: 515 Annotation: Subtype: Link Rect: 335, 409, 347, 418 ActionDest: 515 Annotation: Subtype: Link Rect: 531, 271, 543, 280 ActionDest: 518 Annotation: Subtype: Link Rect: 544, 271, 556, 280 ActionDest: 518 Annotation: Subtype: Link Rect: 411, 130, 418, 142 ActionDest: 468 Page: Sequence: 17 Annotations: Annotation: Subtype: Link Rect: 283, 710, 294, 719 ActionDest: 518 Annotation: Subtype: Link Rect: 295, 710, 312, 719 ActionDest: 528 Annotation: Subtype: Link Rect: 538, 597, 555, 606 ActionDest: 528 Annotation: Subtype: Link Rect: 226, 585, 243, 594 ActionDest: 528 Annotation: Subtype: Link Rect: 305, 584, 317, 594 ActionDest: 515 Annotation: Subtype: Link Rect: 354, 585, 361, 594 ActionDest: 515 Annotation: Subtype: Link Rect: 361, 584, 373, 594 ActionDest: 515 Annotation: Subtype: Link Rect: 430, 584, 447, 594 ActionDest: 528 Annotation: Subtype: Link Rect: 502, 584, 519, 594 ActionDest: 528 Annotation: Subtype: Link Rect: 213, 572, 230, 581 ActionDest: 528 Annotation: Subtype: Link Rect: 289, 547, 301, 556 ActionDest: 518 Annotation: Subtype: Link Rect: 392, 469, 399, 481 ActionDest: 92 Annotation: Subtype: Link Rect: 420, 371, 437, 380 ActionDest: 528 Annotation: Subtype: Link Rect: 326, 296, 338, 305 ActionDest: 525 Annotation: Subtype: Link Rect: 339, 296, 356, 305 ActionDest: 528 Annotation: Subtype: Link Rect: 251, 258, 268, 267 ActionDest: 528 Annotation: Subtype: Link Rect: 269, 258, 286, 267 ActionDest: 528 Annotation: Subtype: Link Rect: 360, 245, 377, 255 ActionDest: 528 Annotation: Subtype: Link Rect: 380, 245, 397, 255 ActionDest: 528 Annotation: Subtype: Link Rect: 246, 221, 253, 229 ActionDest: 515 Annotation: Subtype: Link Rect: 385, 158, 397, 167 ActionDest: 518 Annotation: Subtype: Link Rect: 398, 158, 410, 167 ActionDest: 518 Annotation: Subtype: Link Rect: 540, 120, 552, 129 ActionDest: 518 Annotation: Subtype: Link Rect: 520, 107, 532, 117 ActionDest: 518 Annotation: Subtype: Link Rect: 232, 82, 239, 91 ActionDest: 515 Annotation: Subtype: Link Rect: 239, 82, 246, 91 ActionDest: 515 Annotation: Subtype: Link Rect: 247, 82, 259, 91 ActionDest: 525 Page: Sequence: 18 Annotations: Annotation: Subtype: Link Rect: 382, 723, 394, 732 ActionDest: 515 Annotation: Subtype: Link Rect: 395, 723, 407, 732 ActionDest: 518 Annotation: Subtype: Link Rect: 192, 685, 203, 694 ActionDest: 518 Annotation: Subtype: Link Rect: 204, 685, 221, 694 ActionDest: 528 Annotation: Subtype: Link Rect: 222, 685, 238, 694 ActionDest: 528 Annotation: Subtype: Link Rect: 354, 660, 366, 669 ActionDest: 518 Annotation: Subtype: Link Rect: 515, 647, 532, 657 ActionDest: 531 Annotation: Subtype: Link Rect: 538, 635, 555, 644 ActionDest: 528 Annotation: Subtype: Link Rect: 258, 610, 270, 619 ActionDest: 525 Annotation: Subtype: Link Rect: 270, 610, 287, 619 ActionDest: 531 Annotation: Subtype: Link Rect: 392, 371, 404, 380 ActionDest: 515 Annotation: Subtype: Link Rect: 404, 371, 416, 380 ActionDest: 518 Annotation: Subtype: Link Rect: 435, 359, 442, 368 ActionDest: 515 Annotation: Subtype: Link Rect: 442, 358, 454, 368 ActionDest: 515 Annotation: Subtype: Link Rect: 405, 296, 417, 305 ActionDest: 518 Annotation: Subtype: Link Rect: 418, 296, 430, 305 ActionDest: 528 Annotation: Subtype: Link Rect: 525, 269, 560, 280 Annotation: Subtype: Link Rect: 165, 257, 379, 268 Page: Sequence: 19 Annotations: Annotation: Subtype: Link Rect: 196, 487, 398, 498 Annotation: Subtype: Link Rect: 250, 251, 288, 262 Annotation: Subtype: Link Rect: 95, 206, 132, 216 Annotation: Subtype: Link Rect: 480, 193, 517, 204 Annotation: Subtype: Link Rect: 138, 137, 175, 147 Annotation: Subtype: Link Rect: 342, 113, 379, 124 Annotation: Subtype: Link Rect: 144, 91, 182, 101 Annotation: Subtype: Link Rect: 442, 67, 480, 78 Page: Sequence: 20 Annotations: Annotation: Subtype: Link Rect: 104, 711, 141, 721 Annotation: Subtype: Link Rect: 59, 688, 96, 698 Annotation: Subtype: Link Rect: 439, 664, 476, 675 Annotation: Subtype: Link Rect: 95, 642, 133, 652 Annotation: Subtype: Link Rect: 299, 618, 336, 629 Annotation: Subtype: Link Rect: 196, 584, 234, 594 Annotation: Subtype: Link Rect: 240, 584, 276, 594 Annotation: Subtype: Link Rect: 410, 560, 448, 571 Annotation: Subtype: Link Rect: 364, 537, 401, 548 Annotation: Subtype: Link Rect: 338, 514, 375, 525 Annotation: Subtype: Link Rect: 339, 491, 376, 502 Annotation: Subtype: Link Rect: 289, 469, 326, 479 Annotation: Subtype: Link Rect: 420, 434, 457, 445 Annotation: Subtype: Link Rect: 131, 412, 168, 422 Annotation: Subtype: Link Rect: 200, 342, 238, 353 Annotation: Subtype: Link Rect: 186, 319, 224, 330 Annotation: Subtype: Link Rect: 131, 262, 168, 272 Annotation: Subtype: Link Rect: 276, 238, 313, 249 Annotation: Subtype: Link Rect: 187, 204, 225, 215 Annotation: Subtype: Link Rect: 190, 170, 228, 180 Annotation: Subtype: Link Rect: 358, 146, 395, 157 Annotation: Subtype: Link Rect: 356, 123, 393, 134 Annotation: Subtype: Link Rect: 154, 100, 283, 111 Page: Sequence: 21 Annotations: Annotation: Subtype: Link Rect: 125, 699, 247, 710 Annotation: Subtype: Link Rect: 476, 687, 514, 698 Annotation: Subtype: Link Rect: 312, 652, 350, 664 Annotation: Subtype: Link Rect: 164, 629, 341, 640 Annotation: Subtype: Link Rect: 164, 606, 286, 617 Annotation: Subtype: Link Rect: 195, 583, 340, 594 Annotation: Subtype: Link Rect: 164, 560, 304, 571 Annotation: Subtype: Link Rect: 124, 537, 509, 548 Annotation: Subtype: Link Rect: 125, 457, 266, 468 Annotation: Subtype: Link Rect: 140, 412, 177, 422 Annotation: Subtype: Link Rect: 184, 412, 220, 422 Annotation: Subtype: Link Rect: 59, 377, 96, 387 Annotation: Subtype: Link Rect: 102, 377, 138, 387 Annotation: Subtype: Link Rect: 329, 353, 367, 364 Annotation: Subtype: Link Rect: 171, 331, 209, 341 Annotation: Subtype: Link Rect: 59, 308, 96, 318 Annotation: Subtype: Link Rect: 378, 284, 416, 295 Annotation: Subtype: Link Rect: 192, 262, 229, 272 Annotation: Subtype: Link Rect: 59, 239, 96, 249 Annotation: Subtype: Link Rect: 361, 215, 399, 226 Annotation: Subtype: Link Rect: 333, 192, 370, 203 Annotation: Subtype: Link Rect: 519, 181, 557, 192 Annotation: Subtype: Link Rect: 59, 170, 95, 180 Annotation: Subtype: Link Rect: 396, 146, 434, 157 Annotation: Subtype: Link Rect: 440, 146, 476, 157 Annotation: Subtype: Link Rect: 104, 124, 141, 134 Annotation: Subtype: Link Rect: 195, 100, 232, 111 Annotation: Subtype: Link Rect: 221, 77, 259, 88 Page: Sequence: 22 Annotations: Annotation: Subtype: Link Rect: 118, 722, 155, 733 Annotation: Subtype: Link Rect: 161, 722, 197, 733 Annotation: Subtype: Link Rect: 487, 710, 560, 721 Annotation: Subtype: Link Rect: 56, 699, 155, 710 Annotation: Subtype: Link Rect: 172, 676, 209, 687 Annotation: Subtype: Link Rect: 140, 653, 177, 664 Annotation: Subtype: Link Rect: 171, 630, 209, 640 Annotation: Subtype: Link Rect: 148, 606, 441, 617 Annotation: Subtype: Link Rect: 131, 573, 168, 583 Annotation: Subtype: Link Rect: 125, 491, 308, 502 Annotation: Subtype: Link Rect: 255, 468, 293, 479 Annotation: Subtype: Link Rect: 299, 468, 335, 479 Annotation: Subtype: Link Rect: 158, 445, 195, 456 Annotation: Subtype: Link Rect: 201, 445, 237, 456 Annotation: Subtype: Link Rect: 95, 423, 132, 433 Annotation: Subtype: Link Rect: 171, 399, 209, 410 Annotation: Subtype: Link Rect: 171, 376, 209, 387 Annotation: Subtype: Link Rect: 215, 376, 251, 387 Annotation: Subtype: Link Rect: 344, 353, 382, 364 Annotation: Subtype: Link Rect: 297, 330, 335, 341 Annotation: Subtype: Link Rect: 209, 307, 246, 318 Annotation: Subtype: Link Rect: 453, 285, 490, 295 Annotation: Subtype: Link Rect: 256, 261, 294, 272 Annotation: Subtype: Link Rect: 450, 238, 487, 249 Annotation: Subtype: Link Rect: 494, 238, 530, 249 Annotation: Subtype: Link Rect: 59, 205, 96, 215 Annotation: Subtype: Link Rect: 102, 205, 138, 215 Annotation: Subtype: Link Rect: 278, 169, 315, 180 Annotation: Subtype: Link Rect: 95, 136, 133, 146 Annotation: Subtype: Link Rect: 458, 112, 495, 123 Annotation: Subtype: Link Rect: 472, 100, 509, 111 Annotation: Subtype: Link Rect: 173, 78, 211, 88 Annotation: Subtype: Link Rect: 344, 66, 381, 77 Page: Sequence: 23 Annotations: Annotation: Subtype: Link Rect: 383, 722, 420, 733 Annotation: Subtype: Link Rect: 144, 688, 182, 698 Annotation: Subtype: Link Rect: 301, 664, 339, 675 Annotation: Subtype: Link Rect: 140, 642, 177, 652 Annotation: Subtype: Link Rect: 59, 619, 96, 629 Annotation: Subtype: Link Rect: 135, 596, 173, 606 Annotation: Subtype: Link Rect: 135, 561, 173, 571 Annotation: Subtype: Link Rect: 179, 561, 215, 571 Annotation: Subtype: Link Rect: 359, 526, 397, 537 Annotation: Subtype: Link Rect: 252, 491, 289, 502 Annotation: Subtype: Link Rect: 420, 480, 560, 491 Annotation: Subtype: Link Rect: 56, 468, 198, 479 Annotation: Subtype: Link Rect: 108, 435, 146, 445 Annotation: Subtype: Link Rect: 194, 411, 231, 422 Annotation: Subtype: Link Rect: 365, 399, 402, 410 Annotation: Subtype: Link Rect: 135, 377, 173, 387 Annotation: Subtype: Link Rect: 175, 353, 212, 364 Annotation: Subtype: Link Rect: 270, 330, 307, 341 Annotation: Subtype: Link Rect: 228, 308, 265, 318 Annotation: Subtype: Link Rect: 174, 285, 212, 295 Annotation: Subtype: Link Rect: 291, 250, 329, 261 Annotation: Subtype: Link Rect: 335, 250, 371, 261 Annotation: Subtype: Link Rect: 494, 215, 532, 226 Annotation: Subtype: Link Rect: 132, 182, 170, 192 Annotation: Subtype: Link Rect: 367, 158, 404, 169 Annotation: Subtype: Link Rect: 454, 135, 492, 146 Annotation: Subtype: Link Rect: 173, 113, 210, 123 Annotation: Subtype: Link Rect: 217, 113, 253, 123 Annotation: Subtype: Link Rect: 262, 89, 300, 100 Page: Sequence: 24 Annotations: Annotation: Subtype: Link Rect: 239, 711, 277, 721 Annotation: Subtype: Link Rect: 325, 676, 362, 687 Annotation: Subtype: Link Rect: 368, 676, 405, 687 Checksum: 079e7cbf Type: CRC32 Checksum: ab6fdf7202a1a104e667db2067610249 Type: MD5 Checksum: 473522635c54ee0b29e852e330cf9dca59834f1f Type: SHA-1 Checksum: 81d1ef88321d2644e3f54df031dabf3599b9b7040f77435198dc3b11ba2895bc Type: SHA-256