Sán lá phổi Paragonimus heterotremus và Paragonimus westermani ở Việt Nam hình thái, phân tử, sinh học và chuẩn đoán miễn dịchSán lá phổi Paragonimus heterotremus và Paragonimus westermani ở Việt Nam hình thái, phân tử, sinh học và chuẩn đoán miễn dịchSán lá phổi Paragonimus heterotremus và Paragonimus westermani ở Việt Nam hình thái, phân tử, sinh học và chuẩn đoán miễn dịchSán lá phổi Paragonimus heterotremus và Paragonimus westermani ở Việt Nam hình thái, phân tử, sinh học và chuẩn đoán miễn dịchSán lá phổi Paragonimus heterotremus và Paragonimus westermani ở Việt Nam hình thái, phân tử, sinh học và chuẩn đoán miễn dịchSán lá phổi Paragonimus heterotremus và Paragonimus westermani ở Việt Nam hình thái, phân tử, sinh học và chuẩn đoán miễn dịchSán lá phổi Paragonimus heterotremus và Paragonimus westermani ở Việt Nam hình thái, phân tử, sinh học và chuẩn đoán miễn dịchSán lá phổi Paragonimus heterotremus và Paragonimus westermani ở Việt Nam hình thái, phân tử, sinh học và chuẩn đoán miễn dịch
i - Paragonimus heterotremus VÀ Paragonimus westermani SINH H 2018 ii - Paragonimus heterotremus VÀ Paragonimus westermani Chuyên ngành: Ký sinh trùn 62.42.01.05 TS Bùi Khánh Linh H 2018 i L I CAM O N u c a Các s li u k t qu nêu lu n án trung th c s d ng lu n án khác Hà N i, ngày tháng n m 2018 Tác gi ii L IC M N c bày t l i c Doanh TS Bùi Khánh Linh - gi ng d n khoa h n d y ki n th c, kinh nghi ng th c t i TS Ph m Ng c u ki n t i ng d n th c hi tài vi t lu n án, ng viên tơi su t q trình th c hi n lu n án Xin trân tr ng c Phát tri n khoa h c công ngh qu c gia tài mã s 106.12-2012.52 106-NN.052016.17 c th c hi n n i dung nghiên c u Tôi xin trân tr ng c c Vi n Khoa h c Công ngh o Vi n Sinh thái Tài nguyên sinh v t, Phòng Qu n lý t ng h p, th y cô Vi n ng viên t o m u ki n thu n l i giúp tơi hồn thành lu n án Tôi xin trân tr ng c n t p th cán b Phòng Ký sinh trùng h c, Vi t u ki n cho th c hi n thí nghi m Tơi xin trân tr ng c o B nh vi n Ph , ng viên tơi su t q trình th c hi n lu n án Tôi xin g i l i c n th y giáo, cô giáo, nhà khoa h c, ng nghi p, b n bè Vi n sinh thái Tài ngun sinh v ng viên ch d tơi hồn thành lu n án Cu i cùng, xin bày t lòng bi ct d a v ng ch c cho tơi su t q trình th c hi n lu c cm ng l t qua m Hà N i, ngày tháng n m 2018 Tác gi iii DANH M C T VI T T T Vi t t t Vi ng Vi t CO1 Cytochrome c oxidase subunit mitochondrial gene Gien ty th CO1 ELISA Enzyme-linked immunosorbent assay K thu t h p th mi n d ch liên k t enzym dot-ELISA dot-ELISA m gi y DNA Deoxyribonucleic acid ITS2 Internal transcribed spacer n chèn h gen nhân ITS2 16S rDNA 16S ribosomal DNA/ Gien ty th 16S PCR Polymerase Chain Reaction Ph n ng khuy MC Metacercaria LC Lào Cai YB Yên Bái QT Qu ng Tr SLP Sán ph i i gien iv M CL C L i L IC ii DANH M C T VI T T T iii M C L C iv DANH M C B NG vii DANH M C HÌNH viii M U NG QUAN 1.1 Khái quát chung v sán ph i 1.1.1 1.1.2 1.1.3 11 1.1.4 13 1.1.5 14 .16 1.1.6 17 1.1.7 18 1.2 westermani Paragonimus heterotremus Paragonimus 19 onimus heterotremus 19 20 .21 25 25 25 2.1.3 .25 26 .26 v 27 27 29 29 2.3.4 onimus heterotremus Paragonimus westermani 30 2.3.5 31 2.3.6 31 2.3.7 32 2.3.8 .32 2.3.9 .35 2.3.10 - 36 2.3.10.1 36 2.3.10.2 -ELISA 36 40 .40 40 .47 3.2.1 47 3.2.2 54 3.3 Paragonimus heterotremus Paragonimus westermani .59 3.3.1 59 69 3.3.3 .72 3.3.4 81 3.4 - 85 .85 ° C 87 vi 3.4.3 ° 3.4.4 ° C .88 C .89 o C 90 94 94 94 Paragonimus heterotremus Paragonimus westermani 94 Paragonimus heterotremus Paragonimus westermani .94 - .95 .95 .97 98 .99 vii DANH M C CÁC B NG B ng 1.1 Nh ng v c gi i quy t c a loài sán ph i B ng 2.1 Các c p m i s d nhân b n trình t u ng c B -l c cua m cua B ng 3.4 T l nhi m metacercaria sán ph i B ng 3.5 T l nhi m loài sán ph i B ng 3.6 Hình d m nghiên c u cua su i c d ng metacercaria c a loài P westermani B ng 3.7 Sai khác v trí nucleotide c a trình t gen 16S gi a d ng diploid triploid P westermani B ng 3.8 Kho ng cách di truy n gi a m u c thu t Vi t Nam so v i loài thu c gi ng Gammatricula d a vào trình t gen CO1 B ng 3.9 c u trùng sán ph c B ng 3.10 T l nhi m u trùng sán ph i B ng 3.11 V t ch ct m nghiên c u c c a loài sán ph i c Vi t Nam K t qu gây nhi m metacercaria P westermani cho chu t b ch chu t b B ng 3.14 K t qu gây nhi m P westermani B ng 3.15 c sán thu t ph B ng 3.16 S c s ng c a metacercaria ng v t nuôi phòng (25-30o u ki n nhi c mu i sinh lý B ng 3.17 S c s ng c a metacercaria dung d m B ng 3.18 c mu i sinh lý t i 4oC khác nh c hi u c a ph n ng dot-ELISA B ng 3.19 Th i gian ph n ng ELISA u ki n nhi khác viii DANH M TH Hình 1.1 Hình d ng chung c a sán ph i Paragonimus Hình 1.2 Hình d ng chung c a metacercaria sán ph i Paragonimus Hình 1.3 Hình d ng chung cercaria c a sán ph i Paragonimus Hình 1.4 Hình d ng tr ng sán ph i Paragonimus Hình 1.5 i phát tri n c a lồi Paragonimus westermani Hình 2.1 m nghiên c u Hình 2.2 n i dung nghiên c u Hình 2.3 thí nghi nh v t ch trung gian th nh t Hình 2.4 thí nghi nh v t ch Hình 2.5 Chu n b nh huy Hình 3.1 Các lồi cua su i b i ch ng kháng nguyên lên gi y nitrocellulose c m nghiên c u Hình 3.2 Metacercaria sán ph i tìm th y cua su i thu t i Qu ng Tr Hình 3.3 Metacercaria sán ph i tìm th y cua su i thu t i Yên Bái Lào Cai Hình 3.4 Paragonimus westermani metacercaria thu t cua su i b t t i Yên Bái Hình 3.5 Các d ng P westermani metacercaria thu t cua su i t i t nh Qu ng Tr Hình 3.6 M i quan h ti n hóa phân t c a qu n th P westermani d a trình t c xây d ng b Hình 3.7 M i quan h ti n hóa phân t c a qu n th P westermani d a trình t c xây d ng b Hình 3.8 Metacercaria lồi P heterotremus thu t Yên Bái Lào Cai Hình 3.9 Metacercaria lồi P heterotremus thu t Qu ng Tr Hình 3.10 M i quan h ti n hóa phân t c a qu n th P heterotremus d a trình t c xây d ng b Hình 3.11 M i quan h ti n hóa phân t c a qu n th P heterotremus d a trình t c xây d ng b Hình 3.12 c nhi m u trùng Microcercaria Hình 3.13 Trình t CO1 c a loài c nhi m microcercaria thu t Qu ng Tr cao nh t (99%) v i loài S quangtriensis ng 97 I C A LU N ÁN NH tài lu n án l u tiên th c hi n n i dung nghiên c m i k t qu sau: Phát hi n loài P heterotremus mi n Trung P westermani mi n B c c metacercaria c a loài P westermani nh s P heterotremus Tuy nhiên, qu n th c a lồi mi n Trung có ng cao v di truy n mi n B c nh qu n th P westermani Vi t Nam thu c nhóm 2n nh xác v t ch trung gian th nh t c a loài SLP P westermani, X P heterotremus P proliferus c S quangtriensis, Triculinae gen sp.1, Triculinae gen ng chi u dài ru t s ng cercaria c a redia nh mèo r ng (Prionailurus bengalensis X P skrjabini Da Krong, Qu ng Tr nh chó nhà khơng b nhi m P westermani Mèo Gây nhi m thí nghi nhà b nhi i P heterotremus m n c m th nh chu t nh t tr t ch ch a c a loài P westermani P heterotremus v t ch ch a có th phát tri n ti v t ch nhiên c a loài sán ph i P westermani, P heterotremus t u trùng c a loài SLP có th phân bi t b ng i phát tri n Vi ng thành nh sán non v t ch nh s c s ng c a metacercaria sán ph i không ch ph thu c vào dung d ch ni, nhi , mà ph thu c vào m metacercaria M metacercaria th p metacercaria s sinh lý nhi Trong c mu i 4oC, metacercaria có th s ng t i 12 tháng nh nh th i gian ph n ng c hi u c a k thu t dotu ki n nhi b nh SLP ngồi th khác nhau, có th s d a, k c th i ti t l nh ng th i xác ch n 98 DANH M n, Bùi Khánh Linh, Ph m Ng c Doanh, 2017 So -l c cua tìm u trùng sán ph i H i ngh Sinh thái tài nguyên sinh v t l n th 7, tr 2000-2004 , Ph m Ng Th H p, 2016 S n phát tri n kh Paragonimus westermani y b nh c a lồi sán ph i ng v t thí nghi m T p chí Sinh h c, 38(2):133- 139 Doanh PN, Hien HV, Tu LA, Nonaka N, Horii Y, Nawa Y., 2016 Molecular identification of the trematode Paragonimus in faecal samples from the wild cat Prionailurus bengalensis in the Da Krong Nature Reserve, Vietnam Journal of Helminthology 90(6):658-662 Doanh PN, Tu LA, Bui TD, Loan TH, Nonaka N, Horii Y, Blair D, Nawa Y., 2016 Molecular and morphological variation of Paragonimus westermani in Vietnam with records of new second intermediate crab hosts and a new locality in a 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M TH Hình 1.1 Hình d ng chung c a sán ph i Paragonimus Hình 1.2 Hình d ng chung c a metacercaria sán ph i Paragonimus Hình 1.3 Hình d ng chung cercaria c a sán ph i Paragonimus Hình 1.4 Hình. .. ng k thu t phân t [22] Hình 1.4 Hình d ng tr ng sán ph i Paragonimus [25] 1.1.1.5 Phân lo i sán ph i l n loài phân b thái sán c a tr i thu c gi ng Paragonimus c mô t , ph n châu Á [1] Phân lo i... bengalensis Hình 3.28 S phát tri n c a sán ph i P westermani ng v t thí nghi m Hình 3.29 Paragonimus westermani P westermani Hình 3.31 Khác v c sán non thu a chu t b ch sau gây nhi m tháng Hình 3.32