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Hlístice Vybraných Druhů Studenokrevných Obratlovců Západní Afriky Diplomová Práce

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Hlístice Vybraných Druhů Studenokrevných Obratlovců Západní Afriky Diplomová Práce MASARYKOVA UNIVERZITA Přírodovědecká fakulta Ústav botaniky a zoologie Hlístice vybraných druhů studenokrevných obratlovců západní Afriky Diplomová práce Brno 2008 autor: Bc. Šárka Mašová Vedoucí DP: RNDr. Božena Koubková, Ph.D. PROHLÁŠENÍ Souhlasím s uložením této diplomové práce v knihovně Ústavu botaniky a zoologie PřF MU v Brně, případně v jiné knihovně MU, s jejím veřejným půjčováním a využitím pro vědecké, vzdělávací nebo jiné veřejně prospěšné účely, a to za předpokladu, že převzaté informace budou řádně citovány a nebudou využívány komerčně. Brno, 19. května 2008 …………………………….. PODĚKOVÁNÍ Ráda bych poděkovala vedoucí mé diplomové práce RNDr. Boženě Koubkové, Ph.D. za její odborné vedení a praktické rady. Velice děkuji svému konzultantovi prof. Ing. Vlastimilu Barušovi, DrSc. za cenné rady a pomoc při zpracování problematiky taxonomie nematod, prof. RNDr. Františkovi Tenorovi, DrSc. za pomoc s determinací tasemnic, dále Mgr. Ivetě Matějusové, Ph.D. za molekulární analýzy, Mgr. Ivetě Hodové za zasvěcení do SEM a Mgr. Radimovi Sonnekovi za zasvěcení do CLSM a v neposlední řadě Doc. RNDr. Petru Koubkovi, CSc. za poskytnutí studijního materiálu. Také děkuji všem, kteří mi jakýmkoliv způsobem pomohli při zpracování této diplomové práce a svým nejbližším za podporu. Práce byla finančně podporována Grantovou agenturou AV ČR, grant číslo IAA6093404 a výzkumným záměrem Masarykovy university v Brně číslo MSM 0021622416. ABSTRAKT Za účelem studia parazitických hlístic ryb Senegalu bylo v letech 2004 – 2006 vyšetřeno 330 jedinců náležejících ke 49 sladkovodním druhům ryb. Většina vyšetřených ryb pocházela z národního parku Nikolo Koba ve východním Senegalu. Celkem byly determinovány 3 rody parazitických hlístic ve 24 druzích ryb (prevalence 71%) z 9 čeledí. Nalezená fauna hlístic se skládala většinou ze zástupců čeledi Camallanidae. Nálezy parazitů byly zastoupeny následujícími rody: Cithariniella Khalil, 1964; Labeonema Puylaert, 1970; Camallanus Railliet et Henry, 1915. Převážná část nalezených hlístic byla lokalizována v plynovém měchýři a ve střevě. Jeden druh obojživelníka (Bufo maculatus Hallowell, 1854) a 6 západoafrických druhů plazů [Pelomedusa subrufa Lacépède, 1788; Agama agama (Linnaeus, 1758); Chamaeleo senegalensis Daudin, 1802; Trachylepis perrotetii (Duméril et Bibron, 1839); Tarentola parvicarinata Joger, 1980; Bitis arietans (Merren, 1820)] bylo odchyceno či odloveno v Senegalu a bylo podrobeno parazitologické pitvě. Byli nalezeni tito helminté: Oochoristica truncata (Krabbe 1879), Oochoristica sp. (Linstowiidae) [Cestoda]; Parapharyngodon echinatus (Rudolphi, 1819), Parapharyngodon micipsae (Seurat, 1917), Parapharyngodon sp. (Pharyngodonidae); Strongyluris brevicaudata (Mueller, 1894), Strongyluris sp. (Heterakidae); Cosmocerca sp. (Cosmocercidae); Spirocerca lupi (Spirocercidae) a nedeterminované hlístice řádu Spirurida [Nematoda]. T. parvicarinata, ulovená v národním parku Niokolo Koba ve východním Senegalu, je novým hostitelským druhem hlístic P. echinatus a P. micipsae. Jedinci byli studováni optickým mikroskopem a P. echinatus a P. micipsae poprvé také skanovacím elektronovým mikroskopem. Za účelem získání genetické charakteristiky druhu byla použita částečná sekvence malé podjednotky ribozomálního RNA genu (SSU) ze vzorků P. echinatus. Jedná se o první publikaci sekvence druhu z čeledi Pharyngodonidae. ABSTRACT A total of 49 species (330 specimens) of freshwater fishes from the Senegal Republic were examined for nematode parasites during 2004 – 2006. Majority of the fish were from Niokolo Koba National Park in east Senegal. The survey revealed the presence of 3 determined genera of helminth parasites in 234 fish (prevalence 71%) of 24 species belonging to 9 families. The nematode fauna found was composed predominantly of members of the family Camallanidae. The foundings of parasites were represented by the following genera: Cithariniella Khalil, 1964; Labeonema Puylaert, 1970; Camallanus Railliet et Henry, 1915. The majority of the nematode species found were situated in gas bladder and intestine. One species of amphibian (Bufo maculatus Hallowell, 1854) and 6 west African reptiles species [Pelomedusa subrufa Lacépède, 1788; Agama agama (Linnaeus, 1758); Chamaeleo senegalensis Daudin, 1802; Trachylepis perrotetii (Duméril et Bibron, 1839); Tarentola parvicarinata Joger, 1980; Bitis arietans (Merren, 1820)] were collected in the Senegal and they were underwent parasitological dissection. These helminth species were found in them: Oochoristica truncata (Krabbe 1879), Oochoristica sp. (Linstowiidae) [Cestoda]; Parapharyngodon echinatus (Rudolphi, 1819), Parapharyngodon micipsae (Seurat, 1917), Parapharyngodon sp. (Pharyngodonidae); Strongyluris brevicaudata (Mueller, 1894), Strongyluris sp. (Heterakidae); Cosmocerca sp. (Cosmocercidae); Spirocerca lupi (Spirocercidae) and not determinated nematodes of order Spirurida [Nematoda]. T. parvicarinata, collected in the Niokolo Koba National Park in East Senegal, is a new host species recorded for the nematodes P. echinatus and P. micipsae. Specimens were studied under an optical microscope and P. echinatus and P. micipsae for the first time also by a scanning electron microscope. For the purposes of DNA barcoding, a partial sequence of small subunit ribosomal RNA gene (SSU) was obtained from specimens of P. echinatus. This is the first published sequence of species from the family Pharyngodonidae. OBSAH 1. Úvod............................................................................................................................. 7 1.1 Cíl práce............................................................................................................ 7 1.2 Přehled literatury............................................................................................... 8 1.2.1 Studenokrevní obratlovci jako hostitelé hlístic............................... 8 1.2.2 Ryby ................................................................................................ 8 1.2.3 Obojživelníci................................................................................. 13 1.2.4 Plazi............................................................................................... 13 2. Materiál a metodika ................................................................................................... 15 2.1 Charakteristika studovaného území ................................................................ 15 2.2 Determnace a počet hostitelských organismů................................................. 17 2.3 Odchyt hostitelů, parazitologická pitva a fixace hlístic.................................. 20 2.4 Morfometrie .................................................................................................... 23 2.5 Skanovací elektronová mikroskopie – SEM................................................... 25 2.6 Konfokální laserová skanovací mikroskopie – CLSM ................................... 25 2.7 Analýza DNA.................................................................................................. 26 2.8 Uložení studovaného materiálu....................................................................... 26 3. Výsledky .................................................................................................................... 27 3.1 Nálezy parazitů u vyšteřených druhů hostitelů............................................... 27 3.1.1 Nálezy hlístic u ryb v letech 2004 – 2006..................................... 30 3.1.2 Systematické zařazení jednotlivých druhů hostitelů (plazů a obojživelníka) ........................................................................................ 34 3.2 Přehled determinovaných druhů hlístic a tasemnic ........................................ 35 3.3 Abstrakty rukopisů.......................................................................................... 40 4. Diskuze ...................................................................................................................... 42 5. Závěr .......................................................................................................................... 45 6. Použitá literatura ........................................................................................................ 46 7. Přílohy........................................................................................................................ 58 1. Úvod 1. ÚVOD Nematoda (hlístice) jsou jedním z nejpočetnějších, nejrozšířenějších a nejúspěšnějších kmenů vícebuněčných živočichů. Osídlují obrovské spektrum habitatů a vytvářejí řadu fascinujících životních strategií (Holovachov 2006). Mohou se dělit podle jejich potravní ekologie na bakteriovorní, algivorní – omnivorní – predační (algivore-omnivore-predator), fungivorní, parazité rostlin, entomopatogení a parazité bezobratlých i obratlovců (Martins-Neto 2003). Blaxter et al. (1998) usuzují, že parazitace u živočichů vznikla nezávisle na sobě nejméně čtyřikrát a u rostlin třikrát. Prozatím bylo popsáno přibližně 20 000 druhů hlístic, ale předpokládá se, že jejich počet se může pohybovat mezi 100 000 až několika miliony druhů. Převažují volně žijící půdní, humusové a bentosové druhy, jež hrají důležitou roli ve správném fungování ekosystémů. Nejsou jen konzumenty (bakterií, prvoků, řas, malých máloštětinatců a rostlin), ale jsou i potravním zdrojem jiných zvířat či hub (Holovachov 2006). Parazitace nematody má často silný vliv na populace hostitelů, ať už jsou to rostliny či živočichové. Díky mnoha různým obývaným habitatům,
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