162a8a53c9076442eacd80ba0a52aa76 pmc_10452204.pdf d6dc31e513ecfbfe5a6dd145f8077e5e7dbbce14 pmc_10452204.pdf e313f71cf62bcb11bdc8820a3f153e24dd0e387f38167d3c308e2cf7ccf3163d pmc_10452204.pdf Title: Thrombosis and Hyperinflammation in COVID-19 Acute Phase Are Related to Anti-Phosphatidylserine and Anti-Phosphatidylinositol Antibody Positivity Subject: Antiphospholipid antibodies (APLA) are strongly associated with thrombosis seen in patients with antiphospholipid syndrome. In COVID-19, thrombosis has been observed as one of the main comorbidities. In patients hospitalised for COVID-19, we want to check whether APLA positivity is associated with COVID-19-related thrombosis, inflammation, severity of disease, or long COVID-19. We enrolled 92 hospitalised patients with COVID-19 between March and April 2020 who were tested for 18 different APLAs (IgG and IgM) with a single line-immunoassay test. A total of 30 healthy blood donors were used to set the cut-off for each APLA positivity. Of the 92 COVID-19 inpatients, 30 (32.61%; 95% CI [23.41–43.29]) tested positive for APLA, of whom 10 (33.3%; 95% CI [17.94–52.86]) had more than one APLA positivity. Anti-phosphatidylserine IgM positivity was described in 5.4% of inpatients (n = 5) and was associated with the occurrence of COVID-19-related thrombosis (p = 0.046). Anti-cardiolipin IgM positivity was the most prevalent among the inpatients (n = 12, 13.0%) and was associated with a recorded thrombosis in their clinical history (p = 0.044); however, its positivity was not associated with the occurrence of thrombosis during their hospitalisation for COVID-19. Anti-phosphatidylinositol IgM positivity, with a prevalence of 5.4% (n = 5), was associated with higher levels of interleukin (IL)-6 (p = 0.007) and ferritin (p = 0.034). Neither of these APLA positivities was a risk factor for COVID-19 severity or a predictive marker for long COVID-19. In conclusion, almost a third of COVID-19 inpatients tested positive for at least one APLA. Anti-phosphatidylserine positivity in IgM class was associated with thrombosis, and anti-phosphatidylinositol positivity in IgM class was associated with inflammation, as noticed by elevated levels of IL-6. Thus, testing for non-criteria APLA to assess the risk of clinical complications in hospitalised COVID-19 patients might be beneficial. However, they were not related to disease severity or long COVID-19. Keywords: antiphospholipid antibodies; anti-phosphatidylserine antibody; anti-phosphatidylinositol antibody; COVID-19; disease severity; thrombosis; long COVID-19 Author: Jaume Alijotas-Reig, Ariadna Anunciación-Llunell, Stephanie Morales-Pérez, Jaume Trapé, Enrique Esteve-Valverde and Francesc Miro-Mur Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Mon Aug 21 07:54:25 2023 CEST ModDate: Mon Aug 21 07:58:28 2023 CEST Custom Metadata: no Metadata Stream: no Tagged: no UserProperties: no Suspects: no Form: none JavaScript: no Pages: 16 Encrypted: no Page size: 595.276 x 841.89 pts (A4) Page rot: 0 File size: 2585937 bytes Optimized: no PDF version: 1.7 name type encoding emb sub uni object ID ------------------------------------ ----------------- ---------------- --- --- --- --------- CIQLLF+VnURWPalladioL Type 1 Custom yes yes no 10 0 LMMZGN+URWPalladioL-Bold Type 1 Custom yes yes no 15 0 VWVZTH+URWPalladioL-Roma Type 1 Custom yes yes no 20 0 JDDGNH+URWPalladioL-Ital Type 1 Custom yes yes no 24 0 HJOMHV+VnURWPalladioL-Bold Type 1 Custom yes yes no 28 0 EBFBTV+TimesNewRomanPS-BoldItalicMT Type 1C WinAnsi yes yes no 46 0 KXDJCR+CMSY10 Type 1 Builtin yes yes no 68 0 QDCWHF+EURM10 Type 1 Builtin yes yes no 72 0 QLBIEB+URWPalladioL-BoldItal Type 1 Custom yes yes no 88 0 Jhove (Rel. 1.28.0, 2023-05-18) Date: 2024-03-26 02:35:12 CET RepresentationInformation: pmc_10452204.pdf ReportingModule: PDF-hul, Rel. 1.12.4 (2023-03-16) LastModified: 2024-03-15 02:37:45 CET Size: 2585937 Format: PDF Version: 1.7 Status: Well-Formed and valid SignatureMatches: PDF-hul MIMEtype: application/pdf PDFMetadata: Objects: 365 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: Patients Destination: subsection.2.1 Item: Title: Laboratory Methods Destination: subsection.2.2 Item: Title: Statistical Analysis Destination: subsection.2.3 Item: Title: Results Destination: section.3 Children: Item: Title: Prevalence of APLA in COVID-19 Hospitalised Patients Destination: subsection.3.1 Item: Title: APLA Is Associated with COVID-19 Thrombosis and Inflammation Destination: subsection.3.2 Item: Title: Laboratory Data on Admission and Disease Outcome between APLA Positive and Negative COVID-19 Inpatients Destination: subsection.3.3 Item: Title: APLA Positivity by Sex in COVID-19 Hospitalised Patients Destination: subsection.3.4 Item: Title: APLA as Acute Markers of Long COVID-19 Destination: subsection.3.5 Item: Title: Discussion Destination: section.4 Item: Title: Conclusions Destination: section.5 Item: Title: References Destination: section.A. Info: Title: Thrombosis and Hyperinflammation in COVID-19 Acute Phase Are Related to Anti-Phosphatidylserine and Anti-Phosphatidylinositol Antibody Positivity Author: Jaume Alijotas-Reig, Ariadna Anunciación-Llunell, Stephanie Morales-Pérez, Jaume Trapé, Enrique Esteve-Valverde and Francesc Miro-Mur Subject: Antiphospholipid antibodies (APLA) are strongly associated with thrombosis seen in patients with antiphospholipid syndrome. In COVID-19, thrombosis has been observed as one of the main comorbidities. In patients hospitalised for COVID-19, we want to check whether APLA positivity is associated with COVID-19-related thrombosis, inflammation, severity of disease, or long COVID-19. We enrolled 92 hospitalised patients with COVID-19 between March and April 2020 who were tested for 18 different APLAs (IgG and IgM) with a single line-immunoassay test. A total of 30 healthy blood donors were used to set the cut-off for each APLA positivity. Of the 92 COVID-19 inpatients, 30 (32.61%; 95% CI [23.41 43.29]) tested positive for APLA, of whom 10 (33.3%; 95% CI [17.94 52.86]) had more than one APLA positivity. Anti-phosphatidylserine IgM positivity was described in 5.4% of inpatients (n = 5) and was associated with the occurrence of COVID-19-related thrombosis (p = 0.046). Anti-cardiolipin IgM positivity was the most prevalent among the inpatients (n = 12, 13.0%) and was associated with a recorded thrombosis in their clinical history (p = 0.044); however, its positivity was not associated with the occurrence of thrombosis during their hospitalisation for COVID-19. Anti-phosphatidylinositol IgM positivity, with a prevalence of 5.4% (n = 5), was associated with higher levels of interleukin (IL)-6 (p = 0.007) and ferritin (p = 0.034). Neither of these APLA positivities was a risk factor for COVID-19 severity or a predictive marker for long COVID-19. In conclusion, almost a third of COVID-19 inpatients tested positive for at least one APLA. Anti-phosphatidylserine positivity in IgM class was associated with thrombosis, and anti-phosphatidylinositol positivity in IgM class was associated with inflammation, as noticed by elevated levels of IL-6. Thus, testing for non-criteria APLA to assess the risk of clinical complications in hospitalised COVID-19 patients might be beneficial. However, they were not related to disease severity or long COVID-19. Keywords: antiphospholipid antibodies; anti-phosphatidylserine antibody; anti-phosphatidylinositol antibody; COVID-19; disease severity; thrombosis; long COVID-19 Creator: LaTeX with hyperref Producer: pdfTeX-1.40.21 CreationDate: Mon Aug 21 07:54:25 CEST 2023 ModDate: Mon Aug 21 07:58:28 CEST 2023 ID: 0x2dd68050284a76d74dfb834587d7a807, 0x2dd68050284a76d74dfb834587d7a807 Filters: FilterPipeline: FlateDecode Fonts: Type1: Font: BaseFont: VWVZTH+URWPalladioL-Roma FontSubset: true FirstChar: 2 LastChar: 252 FontDescriptor: FontName: VWVZTH+URWPalladioL-Roma Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: KXDJCR+CMSY10 FontSubset: true FirstChar: 0 LastChar: 21 FontDescriptor: FontName: KXDJCR+CMSY10 Flags: Symbolic FontBBox: -29, -960, 1116, 775 FontFile: true Font: BaseFont: JDDGNH+URWPalladioL-Ital FontSubset: true FirstChar: 3 LastChar: 121 FontDescriptor: FontName: JDDGNH+URWPalladioL-Ital Flags: Symbolic FontBBox: -170, -305, 1010, 941 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: QDCWHF+EURM10 FontSubset: true FirstChar: 11 LastChar: 12 FontDescriptor: FontName: QDCWHF+EURM10 Flags: Symbolic FontBBox: -32, -243, 1060, 720 FontFile: true Font: BaseFont: QLBIEB+URWPalladioL-BoldItal FontSubset: true FirstChar: 110 LastChar: 112 FontDescriptor: FontName: QLBIEB+URWPalladioL-BoldItal Flags: Symbolic FontBBox: -170, -300, 1073, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: CIQLLF+VnURWPalladioL FontSubset: true FirstChar: 161 LastChar: 226 FontDescriptor: FontName: CIQLLF+VnURWPalladioL Flags: Symbolic FontBBox: -166, -283, 1021, 943 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: HJOMHV+VnURWPalladioL-Bold FontSubset: true FirstChar: 178 LastChar: 226 FontDescriptor: FontName: HJOMHV+VnURWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true Font: BaseFont: LMMZGN+URWPalladioL-Bold FontSubset: true FirstChar: 3 LastChar: 122 FontDescriptor: FontName: LMMZGN+URWPalladioL-Bold Flags: Symbolic FontBBox: -152, -301, 1000, 935 FontFile: true EncodingDictionary: Differences: true Pages: Page: Sequence: 1 Annotations: Annotation: Subtype: Link Rect: 82, 281, 152, 290 Annotation: Subtype: Link Rect: 34, 270, 132, 278 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, 197, 792 Annotation: Subtype: Link Rect: 515, 756, 560, 792 Annotation: Subtype: Link Rect: 153, 642, 164, 655 Annotation: Subtype: Link Rect: 153, 629, 164, 643 Annotation: Subtype: Link Rect: 285, 629, 296, 643 Annotation: Subtype: Link Rect: 130, 17, 302, 28 Annotation: Subtype: Link Rect: 389, 17, 560, 28 Annotation: Subtype: Link Rect: 34, 402, 87, 421 Page: Sequence: 2 Annotations: Annotation: Subtype: Link Rect: 333, 736, 340, 744 ActionDest: 123 Annotation: Subtype: Link Rect: 343, 736, 350, 744 ActionDest: 123 Annotation: Subtype: Link Rect: 459, 723, 466, 732 ActionDest: 123 Annotation: Subtype: Link Rect: 469, 723, 476, 732 ActionDest: 123 Annotation: Subtype: Link Rect: 496, 710, 503, 719 ActionDest: 123 Annotation: Subtype: Link Rect: 506, 710, 518, 719 ActionDest: 123 Annotation: Subtype: Link Rect: 426, 685, 438, 694 ActionDest: 123 Annotation: Subtype: Link Rect: 439, 685, 451, 694 ActionDest: 123 Annotation: Subtype: Link Rect: 215, 635, 227, 644 ActionDest: 123 Annotation: Subtype: Link Rect: 228, 635, 240, 644 ActionDest: 123 Annotation: Subtype: Link Rect: 449, 610, 461, 619 ActionDest: 126 Annotation: Subtype: Link Rect: 464, 610, 476, 619 ActionDest: 126 Annotation: Subtype: Link Rect: 495, 597, 507, 606 ActionDest: 126 Annotation: Subtype: Link Rect: 508, 597, 520, 606 ActionDest: 126 Annotation: Subtype: Link Rect: 322, 572, 334, 581 ActionDest: 126 Annotation: Subtype: Link Rect: 314, 497, 326, 506 ActionDest: 126 Annotation: Subtype: Link Rect: 429, 484, 441, 493 ActionDest: 126 Annotation: Subtype: Link Rect: 442, 484, 454, 493 ActionDest: 126 Annotation: Subtype: Link Rect: 276, 447, 288, 456 ActionDest: 126 Annotation: Subtype: Link Rect: 426, 434, 438, 443 ActionDest: 126 Annotation: Subtype: Link Rect: 439, 434, 451, 443 ActionDest: 126 Annotation: Subtype: Link Rect: 203, 409, 215, 418 ActionDest: 126 Annotation: Subtype: Link Rect: 212, 396, 224, 406 ActionDest: 126 Annotation: Subtype: Link Rect: 405, 384, 417, 393 ActionDest: 126 Annotation: Subtype: Link Rect: 266, 359, 278, 368 ActionDest: 126 Annotation: Subtype: Link Rect: 373, 346, 385, 355 ActionDest: 126 Annotation: Subtype: Link Rect: 385, 346, 397, 355 ActionDest: 126 Annotation: Subtype: Link Rect: 315, 334, 327, 343 ActionDest: 126 Annotation: Subtype: Link Rect: 327, 334, 339, 343 ActionDest: 126 Annotation: Subtype: Link Rect: 197, 296, 209, 305 ActionDest: 126 Annotation: Subtype: Link Rect: 212, 296, 224, 305 ActionDest: 126 Page: Sequence: 3 Annotations: Annotation: Subtype: Link Rect: 316, 270, 328, 279 ActionDest: 126 Annotation: Subtype: Link Rect: 331, 270, 343, 279 ActionDest: 129 Annotation: Subtype: Link Rect: 435, 192, 442, 204 ActionDest: 79 Annotation: Subtype: Link Rect: 544, 158, 556, 166 ActionDest: 129 Page: Sequence: 4 Annotations: Annotation: Subtype: Link Rect: 242, 101, 249, 113 ActionDest: 85 Page: Sequence: 5 Annotations: Annotation: Subtype: Link Rect: 230, 458, 237, 470 ActionDest: 79 Annotation: Subtype: Link Rect: 552, 433, 559, 445 ActionDest: 92 Annotation: Subtype: Link Rect: 512, 382, 518, 394 ActionDest: 92 Annotation: Subtype: Link Rect: 517, 357, 524, 369 ActionDest: 85 Annotation: Subtype: Link Rect: 484, 320, 491, 332 ActionDest: 85 Annotation: Subtype: Link Rect: 473, 307, 480, 319 ActionDest: 85 Annotation: Subtype: Link Rect: 535, 282, 542, 294 ActionDest: 85 Annotation: Subtype: Link Rect: 318, 60, 324, 70 ActionDest: 92 Page: Sequence: 6 Page: Sequence: 7 Annotations: Annotation: Subtype: Link Rect: 279, 705, 286, 717 ActionDest: 98 Annotation: Subtype: Link Rect: 185, 407, 191, 417 ActionDest: 85 Annotation: Subtype: Link Rect: 289, 345, 296, 357 ActionDest: 98 Page: Sequence: 8 Annotations: Annotation: Subtype: Link Rect: 437, 694, 444, 706 ActionDest: 101 Annotation: Subtype: Link Rect: 430, 619, 437, 631 ActionDest: 107 Annotation: Subtype: Link Rect: 231, 594, 238, 606 ActionDest: 107 Annotation: Subtype: Link Rect: 283, 94, 289, 106 ActionDest: 107 Page: Sequence: 9 Annotations: Annotation: Subtype: Link Rect: 278, 107, 285, 119 ActionDest: 107 Annotation: Subtype: Link Rect: 400, 57, 407, 69 ActionDest: 107 Page: Sequence: 10 Annotations: Annotation: Subtype: Link Rect: 217, 622, 229, 631 ActionDest: 129 Annotation: Subtype: Link Rect: 338, 544, 345, 556 ActionDest: 98 Annotation: Subtype: Link Rect: 423, 371, 430, 380 ActionDest: 123 Annotation: Subtype: Link Rect: 431, 371, 438, 380 ActionDest: 123 Annotation: Subtype: Link Rect: 438, 371, 445, 380 ActionDest: 123 Annotation: Subtype: Link Rect: 445, 371, 457, 380 ActionDest: 126 Annotation: Subtype: Link Rect: 458, 371, 470, 380 ActionDest: 129 Annotation: Subtype: Link Rect: 253, 346, 265, 355 ActionDest: 126 Annotation: Subtype: Link Rect: 265, 346, 277, 355 ActionDest: 126 Annotation: Subtype: Link Rect: 278, 346, 290, 355 ActionDest: 126 Annotation: Subtype: Link Rect: 290, 346, 302, 355 ActionDest: 129 Annotation: Subtype: Link Rect: 201, 321, 213, 330 ActionDest: 126 Annotation: Subtype: Link Rect: 213, 321, 225, 330 ActionDest: 126 Annotation: Subtype: Link Rect: 226, 321, 238, 330 ActionDest: 129 Annotation: Subtype: Link Rect: 241, 321, 253, 330 ActionDest: 129 Annotation: Subtype: Link Rect: 312, 208, 324, 217 ActionDest: 129 Annotation: Subtype: Link Rect: 426, 158, 438, 166 ActionDest: 126 Annotation: Subtype: Link Rect: 438, 157, 450, 166 ActionDest: 129 Annotation: Subtype: Link Rect: 451, 157, 463, 166 ActionDest: 129 Annotation: Subtype: Link Rect: 344, 132, 356, 141 ActionDest: 129 Annotation: Subtype: Link Rect: 450, 120, 462, 129 ActionDest: 129 Annotation: Subtype: Link Rect: 345, 95, 357, 104 ActionDest: 126 Annotation: Subtype: Link Rect: 358, 95, 370, 104 ActionDest: 129 Annotation: Subtype: Link Rect: 500, 69, 512, 79 ActionDest: 129 Page: Sequence: 11 Annotations: Annotation: Subtype: Link Rect: 344, 735, 356, 744 ActionDest: 129 Annotation: Subtype: Link Rect: 420, 685, 432, 694 ActionDest: 129 Annotation: Subtype: Link Rect: 544, 673, 556, 682 ActionDest: 126 Annotation: Subtype: Link Rect: 397, 660, 409, 669 ActionDest: 126 Annotation: Subtype: Link Rect: 410, 660, 422, 669 ActionDest: 126 Annotation: Subtype: Link Rect: 209, 610, 221, 619 ActionDest: 129 Annotation: Subtype: Link Rect: 448, 572, 460, 581 ActionDest: 126 Annotation: Subtype: Link Rect: 543, 560, 555, 569 ActionDest: 126 Annotation: Subtype: Link Rect: 230, 497, 242, 506 ActionDest: 126 Annotation: Subtype: Link Rect: 432, 409, 444, 418 ActionDest: 129 Annotation: Subtype: Link Rect: 447, 409, 459, 418 ActionDest: 129 Annotation: Subtype: Link Rect: 350, 296, 362, 305 ActionDest: 126 Annotation: Subtype: Link Rect: 393, 271, 405, 280 ActionDest: 126 Annotation: Subtype: Link Rect: 406, 271, 418, 280 ActionDest: 126 Annotation: Subtype: Link Rect: 421, 271, 433, 280 ActionDest: 126 Annotation: Subtype: Link Rect: 302, 170, 314, 180 ActionDest: 129 Annotation: Subtype: Link Rect: 315, 170, 327, 180 ActionDest: 129 Annotation: Subtype: Link Rect: 205, 83, 217, 92 ActionDest: 126 Annotation: Subtype: Link Rect: 218, 83, 230, 92 ActionDest: 129 Page: Sequence: 12 Annotations: Annotation: Subtype: Link Rect: 291, 735, 303, 744 ActionDest: 129 Annotation: Subtype: Link Rect: 323, 685, 335, 694 ActionDest: 129 Annotation: Subtype: Link Rect: 520, 647, 532, 657 ActionDest: 129 Annotation: Subtype: Link Rect: 443, 635, 450, 644 ActionDest: 123 Annotation: Subtype: Link Rect: 254, 522, 266, 531 ActionDest: 132 Annotation: Subtype: Link Rect: 525, 522, 537, 531 ActionDest: 132 Annotation: Subtype: Link Rect: 411, 497, 422, 506 ActionDest: 132 Annotation: Subtype: Link Rect: 536, 244, 560, 255 Annotation: Subtype: Link Rect: 165, 232, 411, 243 Page: Sequence: 13 Annotations: Annotation: Subtype: Link Rect: 348, 515, 386, 525 Annotation: Subtype: Link Rect: 260, 480, 297, 491 Annotation: Subtype: Link Rect: 304, 480, 340, 491 Annotation: Subtype: Link Rect: 104, 446, 141, 456 Annotation: Subtype: Link Rect: 147, 446, 183, 456 Annotation: Subtype: Link Rect: 393, 422, 431, 433 Annotation: Subtype: Link Rect: 437, 422, 473, 433 Annotation: Subtype: Link Rect: 264, 388, 302, 399 Annotation: Subtype: Link Rect: 104, 366, 141, 376 Annotation: Subtype: Link Rect: 309, 342, 346, 353 Annotation: Subtype: Link Rect: 59, 308, 96, 318 Annotation: Subtype: Link Rect: 95, 274, 133, 284 Annotation: Subtype: Link Rect: 179, 238, 217, 249 Annotation: Subtype: Link Rect: 336, 204, 374, 215 Annotation: Subtype: Link Rect: 380, 204, 416, 215 Annotation: Subtype: Link Rect: 164, 170, 202, 180 Annotation: Subtype: Link Rect: 208, 170, 244, 180 Annotation: Subtype: Link Rect: 472, 135, 509, 146 Annotation: Subtype: Link Rect: 515, 135, 551, 146 Annotation: Subtype: Link Rect: 59, 101, 96, 111 Annotation: Subtype: Link Rect: 520, 77, 557, 88 Page: Sequence: 14 Annotations: Annotation: Subtype: Link Rect: 59, 722, 96, 733 Annotation: Subtype: Link Rect: 182, 699, 219, 710 Annotation: Subtype: Link Rect: 169, 676, 206, 687 Annotation: Subtype: Link Rect: 95, 653, 133, 664 Annotation: Subtype: Link Rect: 209, 618, 247, 629 Annotation: Subtype: Link Rect: 190, 584, 228, 594 Annotation: Subtype: Link Rect: 234, 584, 270, 594 Annotation: Subtype: Link Rect: 208, 561, 246, 571 Annotation: Subtype: Link Rect: 252, 561, 288, 571 Annotation: Subtype: Link Rect: 59, 538, 96, 548 Annotation: Subtype: Link Rect: 102, 538, 138, 548 Annotation: Subtype: Link Rect: 131, 515, 168, 525 Annotation: Subtype: Link Rect: 384, 491, 421, 502 Annotation: Subtype: Link Rect: 162, 469, 199, 479 Annotation: Subtype: Link Rect: 59, 435, 96, 445 Annotation: Subtype: Link Rect: 490, 422, 528, 433 Annotation: Subtype: Link Rect: 411, 399, 448, 410 Annotation: Subtype: Link Rect: 515, 376, 552, 387 Annotation: Subtype: Link Rect: 177, 343, 214, 353 Annotation: Subtype: Link Rect: 220, 343, 256, 353 Annotation: Subtype: Link Rect: 324, 319, 361, 330 Annotation: Subtype: Link Rect: 367, 319, 403, 330 Annotation: Subtype: Link Rect: 342, 284, 378, 295 Annotation: Subtype: Link Rect: 309, 250, 347, 261 Annotation: Subtype: Link Rect: 470, 215, 507, 226 Annotation: Subtype: Link Rect: 500, 192, 538, 203 Annotation: Subtype: Link Rect: 59, 170, 96, 180 Annotation: Subtype: Link Rect: 104, 147, 141, 157 Annotation: Subtype: Link Rect: 182, 112, 219, 123 Annotation: Subtype: Link Rect: 386, 89, 423, 100 Page: Sequence: 15 Annotations: Annotation: Subtype: Link Rect: 96, 711, 134, 721 Annotation: Subtype: Link Rect: 140, 711, 176, 721 Annotation: Subtype: Link Rect: 204, 676, 242, 687 Annotation: Subtype: Link Rect: 248, 676, 284, 687 Annotation: Subtype: Link Rect: 170, 653, 207, 664 Annotation: Subtype: Link Rect: 435, 630, 472, 640 Annotation: Subtype: Link Rect: 361, 606, 398, 617 Annotation: Subtype: Link Rect: 477, 583, 515, 594 Annotation: Subtype: Link Rect: 521, 583, 557, 594 Annotation: Subtype: Link Rect: 248, 561, 285, 571 Annotation: Subtype: Link Rect: 291, 561, 327, 571 Annotation: Subtype: Link Rect: 373, 537, 410, 548 Annotation: Subtype: Link Rect: 442, 526, 479, 537 Annotation: Subtype: Link Rect: 221, 503, 259, 514 Annotation: Subtype: Link Rect: 196, 469, 234, 479 Annotation: Subtype: Link Rect: 240, 469, 276, 479 Annotation: Subtype: Link Rect: 376, 445, 413, 456 Annotation: Subtype: Link Rect: 420, 445, 456, 456 Annotation: Subtype: Link Rect: 438, 422, 475, 433 Annotation: Subtype: Link Rect: 482, 422, 518, 433 Annotation: Subtype: Link Rect: 155, 400, 192, 410 Annotation: Subtype: Link Rect: 198, 400, 234, 410 Annotation: Subtype: Link Rect: 165, 366, 202, 376 Annotation: Subtype: Link Rect: 473, 342, 510, 353 Annotation: Subtype: Link Rect: 319, 319, 356, 330 Annotation: Subtype: Link Rect: 467, 296, 505, 307 Annotation: Subtype: Link Rect: 311, 274, 349, 284 Annotation: Subtype: Link Rect: 418, 261, 456, 272 Annotation: Subtype: Link Rect: 261, 239, 298, 249 Annotation: Subtype: Link Rect: 304, 239, 340, 249 Annotation: Subtype: Link Rect: 104, 205, 141, 215 Annotation: Subtype: Link Rect: 148, 205, 184, 215 Annotation: Subtype: Link Rect: 190, 169, 228, 180 Annotation: Subtype: Link Rect: 234, 169, 270, 180 Annotation: Subtype: Link Rect: 59, 136, 96, 146 Annotation: Subtype: Link Rect: 102, 136, 138, 146 Annotation: Subtype: Link Rect: 511, 112, 548, 123 Annotation: Subtype: Link Rect: 59, 78, 96, 88 Page: Sequence: 16 Annotations: Annotation: Subtype: Link Rect: 464, 722, 501, 733 Annotation: Subtype: Link Rect: 418, 699, 455, 710 Annotation: Subtype: Link Rect: 508, 676, 545, 687 Checksum: 55992b59 Type: CRC32 Checksum: 162a8a53c9076442eacd80ba0a52aa76 Type: MD5 Checksum: d6dc31e513ecfbfe5a6dd145f8077e5e7dbbce14 Type: SHA-1 Checksum: e313f71cf62bcb11bdc8820a3f153e24dd0e387f38167d3c308e2cf7ccf3163d Type: SHA-256