The Effects of High Temperature, Low Dissolved Oxygen, and Asian Tapeworm Pengaruh Suhu Tinggi, Oksigen terlarut

rendah, dan Asia cacing pita Infection on Growth and Survival of the Topeka shiner, Notropis topeka Infeksi pada Pertumbuhan dan Survival dari berlian Topeka, Notropis Topeka A THESIS Sebuah Tesis SUBMITTED TO THE FACULTY OF THE GRADUATE SCHOOL Disampaikan kepada FAKULTAS SEKOLAH SARJANA OF THE UNIVERSITY OF MINNESOTA DARI University of Minnesota BY OLEH Jessica Jill Koehle Jessica Jill Koehle IN PARTIAL FULFILLMENT OF THE REQUIREMENTS DALAM PEMENUHAN PARSIAL DARI PERSYARATAN FOR THE DEGREE OF UNTUK TINGKAT MASTERS SCIENCE MASTER ILMU March 2006 Mar 2006 i aku Acknowledgments Ucapan Terima Kasih I am grateful to many people who have contributed to this research and to my Saya berterima kasih kepada banyak orang yang telah memberikan kontribusi untuk penelitian dan untuk saya graduate school experience. lulusan sekolah pengalaman. First, I would like to thank my advisor, Dr. Ira Adelman, Pertama, saya ingin mengucapkan terima kasih kepada penasehat saya, Dr Ira Adelman, who is an exceptional mentor and role model. yang merupakan mentor yang luar biasa dan model peran. Also, thanks to my committee members, Juga, terima kasih kepada anggota komite saya, Dr. Bruce Vondracek and Dr. Jay Hatch for their invaluable guidance. Dr Bruce Vondracek dan Dr Jay Hatch atas bimbingan mereka. For funding this project, I thank the Minnesota Department of Natural Resources, Untuk pendanaan proyek ini, saya mengucapkan terima kasih kepada Departemen Sumber Daya Alam Minnesota, Division of Ecological Services, especially Rich Baker, and the United States Fish and Divisi Layanan Ekologis, terutama Rich Baker, dan Amerika Serikat dan Ikan Wildlife Service, Endangered Species Program, for a grant provided under Section 6 of Wildlife Service, Endangered Species Program, untuk hibah yang diberikan berdasarkan Bagian 6 dari the Endangered Species Act. Endangered Species Act. I thank Rich Cook and others at the Lost Valley Fish Saya berterima kasih Rich Cook dan lain-lain di Ikan Lost Valley Hatchery (Missouri Department of Conservation), and Scott Campbell with the Hatchery (Departemen Konservasi Missouri), dan Scott Campbell dengan University of Kansas for providing Topeka shiners. University of Kansas untuk memberikan shiners Topeka. Thanks also to Andrew Goodwin of Terima kasih juga kepada Andrew Goodwin dari the University of Arkansas for identifying tapeworms, and to Drew Mitchell of Stuttgart University of Arkansas untuk mengidentifikasi cacing pita, dan Drew Mitchell dari Stuttgart National Aquaculture Research Center, for recommending worming treatment. Budidaya Nasional Research Center, untuk merekomendasikan pengobatan cacing.

Thanks to several professors at the University of Minnesota, including Dr. Peter Terima kasih kepada beberapa dosen di University of Minnesota, termasuk Dr Peter Sorensen, Dr. Ray Newman, Dr. George Spangler, Dr. Andrew Simons, Dr. Anne Sorensen, Dr Newman Ray, Dr George Spangler, Dr Andrew Simons, Dr Anne Kapuscinski, Dr. Yossi Cohen, and Dr. Sandy Weisberg, who have offered lab space, Kapuscinski, Dr Yossi Cohen, dan Dr Sandy Weisberg, yang menawarkan ruang laboratorium, statistical advice, and other guidance. statistik saran, dan bimbingan lainnya. Also, thanks to the departmental office staff, Juga, terima kasih kepada staf kantor departemen, including Nancy Rothman, Abby Lloyd, Rachel Jones, Mary Barfield, Rosalyn Zippa, termasuk Nancy Rothman, Abby Lloyd, Rachel Jones, Mary Barfield, Rosalyn Zippa, and Jo Schroeder. dan Jo Schroeder. Many thanks to Jay Maher for his great attitude, ingenuity, and Banyak terima kasih untuk Jay Maher untuk sikap yang besar, kecerdikan, dan selfless commitment to fisheries research at the University of Minnesota, and to Mark komitmen tanpa pamrih untuk penelitian perikanan di University of Minnesota, dan untuk Mark Hove, Rachel Jones, Matt Hennen, and Thor Eide for their assistance in the laboratory. Hove, Rachel Jones, Matt Hennen, dan Thor Eide atas bantuan mereka di laboratorium. Thanks also to my fellow graduate students! Terima kasih juga kepada sesama siswa saya lulus! I am grateful to many of you, but Saya berterima kasih kepada banyak dari Anda, tetapi particularly to Dr. Dalma Martinovic, Dr. Marilyn Sherman, Jared Fine, Lance Vrieze, terutama untuk Dr Dalma Martinovic, Dr Marilyn Sherman, Jared Baik, Lance Vrieze, Sean Sisler, Ben Thwaits, Branda Long, Leah Sharpe, Mike Simone, and Nick Schlesser. Sean Sisler, Thwaits Ben, Branda Long, Sharpe Leah, Simone Mike, dan Nick Schlesser. Also, thanks to Carl Cabak, Nick Gidmark, and Kristan Maccaroni for lab assistance. Juga, berkat Carl Cabak, Nick Gidmark, dan Kristan Maccaroni untuk bantuan lab. Finally, I would like to thank my friends and family. Akhirnya, saya ingin mengucapkan terima kasih kepada teman-teman dan keluarga. My parents, Alan and Orangtuaku, Alan dan Patricia Davis, showed me the beauty of the natural world which is the inspiration for my Patricia Davis, menunjukkan keindahan alam yang merupakan inspirasi bagi saya career, and helped to give me the opportunity for higher education. karir, dan membantu memberikan saya kesempatan untuk pendidikan yang lebih tinggi. Also thanks to my Juga terima kasih kepada saya three sisters, Jennie Kelly, Alita Miller, and Laura Strader, who have supported and tiga bersaudara, Jennie Kelly, Alita Miller, dan Laura Strader, yang telah mendukung dan encouraged me constantly. mendorong saya terus-menerus. Last but not least, thanks to my husband, Karl Koehle, for Last but not least, terima kasih kepada suami saya, Karl Koehle, untuk being my sounding board and best friend, and for always making life interesting. papan yang terdengar saya dan teman terbaik, dan untuk selalu membuat hidup menarik. ii ii Abstract Abstrak The Topeka shiner ( Notropis topeka ) is an endangered fish species, historically described The berlian Topeka (Notropis Topeka) adalah spesies ikan terancam punah, historis dijelaskan as inhabiting cool, headwater prairie streams. sebagai mendiami dingin, padang rumput hulu sungai. However, Topeka shiners recently have Namun, baru-baru ini telah shiners Topeka been found in off-channel habitats with high temperatures and low dissolved oxygen ditemukan di-channel habitat off dengan suhu tinggi dan rendah oksigen terlarut

levels. tingkat. To determine if Topeka shiners can tolerate conditions in these off-channel Untuk menentukan apakah Topeka shiners bisa mentolerir kondisi dalam saluran-off habitats for extended periods of time, I determined their critical thermal maximum habitat untuk waktu yang lama, saya memutuskan mereka termal maksimum kritis (CTM), optimum temperature for growth, and lower lethal dissolved oxygen level (96hr (CTM), suhu optimum untuk pertumbuhan, dan mematikan lebih rendah tingkat oksigen terlarut (96hr LC LC 50 50 ). ). Also, I studied the effects of reduced oxygen as well as Asian tapeworm infection Juga, saya mempelajari efek oksigen berkurang serta infeksi cacing pita Asia ( Bothriocephalus acheilognathi ) on growth. (Bothriocephalus acheilognathi) pada pertumbuhan. Topeka shiners have a CTM of 39 C at a 31 shiners Topeka memiliki CTM 39 C pada 31 C acclimation temperature and their optimum temperature for growth is about 27 C. C aklimatisasi suhu dan suhu optimum untuk pertumbuhan adalah sekitar 27 C. Topeka shiners are capable of growth at dissolved oxygen concentrations as low as 2 mg Topeka shiners mampu pertumbuhan pada konsentrasi oksigen terlarut serendah 2 mg LL -1 -1 , but in some circumstances at a considerably lower rate than at dissolved oxygen , Tapi dalam beberapa keadaan pada tingkat yang jauh lebih rendah dari pada oksigen terlarut concentrations at or above 4 mg L konsentrasi pada atau di atas 4 mg L -1 -1 . . Their 96-hour LC Mereka 96-jam LC 50 50 for dissolved oxygen at 26 C is untuk oksigen terlarut di 26 C 1.2 mg L 1,2 mg L -1 -1 . . Finally, growth is reduced by the presence of Asian tapeworms. Akhirnya, pertumbuhan berkurang dengan adanya cacing pita Asia. Asian Asia tapeworm is not only a threat to fish health but use of infected fish in restoration cacing pita tidak hanya menjadi ancaman bagi kesehatan ikan tetapi ikan terinfeksi penggunaan restorasi programs is prohibited. program adalah dilarang. Overall, temperature and oxygen are probably not responsible for Secara keseluruhan, suhu dan oksigen mungkin tidak bertanggung jawab atas Topeka shiner population declines and are not limiting factors in most off-channel Topeka penurunan populasi mata bengkak dan tidak membatasi faktor-faktor di kebanyakan channel offhabitats. habitat. Judging by the abundance of Topeka shiners in off-channel habitats, these Dilihat oleh kelimpahan shiners Topeka di-channel habitat off, ini habitats may be population sources, rather than sinks, and thus may be important to mungkin habitat populasi sumber, bukan sink, dan dengan demikian mungkin penting Topeka shiner populations. Topeka mata bengkak populasi. iii iii Table of Contents Daftar isi Acknowledgments Ucapan Terima Kasih i aku

Abstract Abstrak ii ii List of Tables and Figures Daftar Tabel dan Angka iv iv Introduction Pengenalan 11 Methods Metode 44 General Procedures Prosedur Umum 44 Optimum Temperature for Growth Suhu optimum untuk Pertumbuhan 66 Critical Thermal Maximum Kritis Thermal Maksimum 99 Effect of Dissolved Oxygen on Growth Pengaruh Oksigen terlarut terhadap Pertumbuhan 10 10 Lower Lethal Dissolved Oxygen Concentration Konsentrasi Oksigen terlarut rendah Lethal 13 13 Data Analysis Analisis Data 15 15 Results Hasil 17 17 Optimum Temperature and Effect of Asian Tapeworm on Growth Suhu optimum dan cacing pita Pengaruh Asia Terhadap Pertumbuhan 17 17 Critical Thermal Maximum Kritis Thermal Maksimum 21 21 Effect of Dissolved Oxygen on Growth Pengaruh Oksigen terlarut terhadap Pertumbuhan 22 22 Lower Lethal Dissolved Oxygen Concentration Konsentrasi Oksigen terlarut rendah Lethal 26 26 Discussion Diskusi 26 26 References Referensi 34 34 iv iv List of Tables and Figures Daftar Tabel dan Angka Table 1 Tabel 1 Means and standard deviations for all treatment temperatures Sarana dan deviasi standar untuk semua perlakuan suhu 77 in experiments T-1, T-2, and T-3 dalam percobaan T-1, T-2, dan T-3 Table 2 Tabel 2 Means and standard deviations for all treatment dissolved oxygen 11 Sarana dan deviasi standar untuk semua perawatan oksigen terlarut 11

concentrations in experiments O-1 and O-2 konsentrasi dalam percobaan O-1 dan O-2 Table 3 Tabel 3 Means and standard deviations for all treatment dissolved oxygen 14 Sarana dan deviasi standar untuk semua perawatan oksigen terlarut 14 concentrations in experiments LC konsentrasi dalam eksperimen LC 50 50 -1 and LC -1 Dan LC 50 50 -2 -2 Table 4 Tabel 4 Summary of specific growth rate results for experiments Ringkasan hasil laju pertumbuhan spesifik untuk eksperimen 18 18 T-1, T-2, and T-3 T-1, T-2, dan T-3 Table 5 Tabel 5 Summary of results for experiments CTM-1, CTM-2, and CTM-3 Ringkasan hasil untuk percobaan CTM-1, CTM-2, dan CTM-3 23 23 Table 6 Tabel 6 CTM values for other species of Notropis , adapted from CTM nilai untuk spesies lain dari Notropis, diadaptasi dari Beitinger et al . (2000) Beitinger et al. (2000) 29 29 Figure 1 Gambar 1 Relationship of specific growth rate to temperature Hubungan laju pertumbuhan spesifik ke suhu 19 19 for weight and length in Experiment T-1 untuk berat dan panjang dalam Percobaan T-1 Figure 2 Gambar 2 Observations of behavior during experiment T-1 for both Pengamatan perilaku selama percobaan T-1 untuk kedua 19 19 swimming activity and reproductive behavior Aktivitas berenang dan perilaku reproduksi Figure 3 Gambar 3 Relationship of specific growth rate to temperature Hubungan laju pertumbuhan spesifik ke suhu 20 20 For weight and length in Experiment T-2 Untuk berat dan panjang dalam Percobaan T-2 Figure 4 Gambar 4 Relationship of specific growth rate to temperature Hubungan laju pertumbuhan spesifik ke suhu 20 20 for weight and length in Experiment T-3, for fish both untuk berat dan panjang dalam Percobaan T-3, untuk ikan baik with and without Asian tapeworm infection dengan dan tanpa infeksi cacing pita Asia Figure 5 Gambar 5 Mean specific growth rates for weight and length of 19-month-old Berarti tingkat pertumbuhan spesifik berat dan panjang 19-month-old 24 24

fish (A)and 7-month-old fish (B) in relation to dissolved oxygen ikan (A) dan bulan ikan berusia 7 (B) dalam hubungannya dengan oksigen terlarut concentration in experiment O-1 konsentrasi dalam percobaan O-1 Figure 6 Gambar 6 Mean specific growth rates for weight and length of fish in relation Berarti tingkat pertumbuhan spesifik berat dan panjang ikan sehubungan 25 25 to dissolved oxygen concentration in experiment O-2 terhadap konsentrasi oksigen terlarut dalam percobaan O-2 Figure 7 Gambar 7 Summary of behavior observations during experiment O-2 Ringkasan dari pengamatan perilaku selama percobaan O-2 25 25 Page 6 Page 6 11 Introduction Pengenalan The Topeka shiner ( Notropis topeka ) is an endangered minnow species native to The berlian Topeka (Notropis Topeka) adalah spesies ikan kecil terancam asli North American prairie streams in Minnesota, South Dakota, Iowa, Nebraska, Missouri, Padang rumput Amerika Utara sungai di Minnesota, South Dakota, Iowa, Nebraska, Missouri, and Kansas (Hatch 2001). dan Kansas (Hatch 2001). Over the past century the Topeka shiner's range has decreased Selama abad terakhir Topeka's berlian kisaran menurun by 80%, with more than 50% of this decline within the past 25 years (Tabor 1998). dengan 80%, dengan lebih dari 50% dari penurunan dalam tahun terakhir 25 (Tabor 1998). Habitat loss may be one of the main reasons for their decline, but habitat suitability Habitat kerugian mungkin salah satu alasan utama untuk penurunan mereka, tetapi kesesuaian habitat information for Topeka shiners is limited. informasi untuk shiners Topeka terbatas. Topeka shiners have recently been found in Topeka shiners baru saja ditemukan abundance in off-channel habitats in Minnesota. kelimpahan di-channel habitat off di Minnesota. These off-channel habitats may be Ini-saluran habitat off mungkin sources, rather than sinks, for shiner populations (Pulliam, 1988; Dahle, 2001) and it is sumber, bukan tenggelam, untuk mata bengkak populasi (Pulliam, 1988; Dahle, 2001) dan merupakan vital to understand how Topeka shiners use these habitats. penting untuk memahami bagaimana menggunakan Topeka shiners habitat. The continued existence of Kelangsungan hidup this species is important to the larger goal of protecting native biodiversity in Midwest spesies ini penting untuk tujuan yang lebih besar untuk melindungi keanekaragaman hayati asli di Midwest prairie stream ecosystems. padang rumput ekosistem sungai. Information is now available about some aspects of Topeka shiner life history, but Informasi kini tersedia tentang beberapa aspek sejarah kehidupan berlian Topeka, tetapi questions still remain about their habitat requirements. pertanyaan masih tetap tentang persyaratan habitat mereka. Topeka shiners are small (<75 Topeka shiners kecil (<75 mm) omnivorous cyprinids (Hatch and Besaw 2001). mm) ikan Cyprinid omnivora (Hatch dan Besaw 2001). Spawning occurs between early Pemijahan terjadi antara awal May and late August, and is often associated with sunfish ( Lepomis spp. ) nests (Dahle Mei dan akhir Agustus, dan sering dikaitkan dengan mola-mola (sarang Lepomis spp.) (Dahle

2001; Kerns and Bonneau 2002; Stark et al. 2001; Kerns dan 2002 Bonneau; Stark et al. 2002). 2002). Historically, Topeka shiners were Secara historis, shiners Topeka adalah reported to live in cool, clear headwater stream pools with substrates of gravel, rubble, dilaporkan tinggal di dingin, jelas kolam hulu sungai dengan substrat kerikil, puing-puing, clay hardpan, or bedrock, sometimes with a thin layer of silt. hardpan tanah liat, atau batuan dasar, kadang-kadang dengan lapisan tipis lumpur. Many of these streams Banyak dari sungaisungai ini were intermittent during dry summers, but were maintained by groundwater seepage yang berselang selama musim panas kering, namun dikelola oleh rembesan airtanah (Minckley and Cross 1959). (Minckley dan Cross 1959). Current descriptions of Topeka shiner habitat in Kansas and Deskripsi saat ini habitat berlian di Topeka Kansas dan Missouri agree with historical descriptions. Missouri setuju dengan deskripsi historis. Topeka shiners are found in pools often Topeka shiners ditemukan di kolam sering connected by water seeping through the gravel (Hrabik 1996; Kerns and Bonneau 2002; dihubungkan dengan air merembes melalui kerikil (Hrabik 1996; Kerns dan Bonneau 2002; Page 7 Page 7 22 Stark et al. Stark et al. 2002) with generally good water quality (Bayless et al. 2003). 2002) dengan umumnya kualitas air yang baik (Bayless et al). 2003. However, in Namun, dalam addition to those habitats, populations of Topeka shiners in Minnesota have been reported Selain habitat mereka, populasi shiners Topeka di Minnesota telah dilaporkan to inhabit and spawn in off-channel oxbow lakes and excavated pools, as well as in untuk tinggal dan bertelur di saluran oxbow danau-off dan kolam digali, serta streams carrying high sediment loads (Dahle 2001; Hatch 2001). sungai yang membawa beban sedimen yang tinggi (Dahle 2001; Hatch 2001). Kuitunen (2001) Kuitunen (2001) reported that Topeka shiners in Minnesota streams were more often collected in clearer, melaporkan bahwa Topeka shiners di Minnesota sungai lebih sering dikumpulkan dalam lebih jelas, vegetated backwater areas than in more turbid stream channels. tumbuhan daerah terpencil di lebih dari saluran sungai keruh. The presence of Topeka Kehadiran Topeka shiners in these off-channel habitats has been found to depend on a variety of factors, shiners dalam habitat off-channel telah ditemukan tergantung pada berbagai faktor, including flooding events (Berg et al. 2004). termasuk peristiwa banjir (Berg et al). 2004. Offchannel habitat is often maintained by Off-saluran habitat sering dikelola oleh groundwater or seepage from the main stream channel (Berg et al. 2004), but it is unclear air tanah atau rembesan dari saluran sungai utama (Berg et al). 2004, tetapi tidak jelas whether temperature and dissolved oxygen levels in degraded streams and off-channel apakah tingkat oksigen terlarut dan suhu di sungai terdegradasi dan off-channel habitat are suitable for Topeka shiners. habitat yang cocok untuk shiners Topeka. Because these factors may be limiting in off- Karena faktor-faktor ini dapat membatasi di offchannel habitats, this study was undertaken to determine tolerance of Topeka shiners to habitat saluran, penelitian ini dilakukan untuk menentukan toleransi dari shiners Topeka untuk high temperature and low oxygen. tinggi suhu dan oksigen rendah. A few anecdotal observations have been made on Topeka shiner temperature Sebuah pengamatan beberapa anekdot yang telah dibuat pada suhu bintang Topeka

tolerance, but no information has been published for Topeka shiner low dissolved oxygen toleransi, tetapi tidak ada informasi yang telah dipublikasikan untuk mata bengkak Topeka oksigen terlarut rendah tolerance. toleransi. In Kansas, Kerns and Bonneau (2002) reported that Topeka shiners were Dalam Kansas, Kerns dan Bonneau (2002) melaporkan bahwa Topeka shiners adalah usually the last fish to succumb to deteriorating conditions in drying pools; temperatures biasanya ikan terakhir untuk menyerah memburuknya kondisi di pengeringan kolam; suhu reached 28.5 C during their study, and more extreme temperatures likely occurred. mencapai 28,5 C selama studi mereka, dan lebih ekstrim suhu mungkin terjadi. Also Juga in Kansas, Topeka shiners were observed in temperatures up to 29 C (Schrank et al. di Kansas, Topeka shiners diamati pada suhu sampai 29 C (Schrank et al. 2001). 2001). In Minnesota, Hatch (2001) reported observing Topeka shiner spawning behavior Di Minnesota, Hatch (2001) melaporkan melihat perilaku pemijahan berlian Topeka at 31 C. Specifically in off-channel habitat in Minnesota, Dahle (2001) reported finding Pada tanggal 31 C. Khususnya di-channel habitat off di Minnesota, Dahle (2001) melaporkan temuan shiners in water that was 24 C in August 1998. shiners dalam air yang 24 C pada bulan Agustus 1998. Page 8 Page 8 33 During the course of the experiments to determine temperature and oxygen Selama percobaan untuk menentukan suhu dan oksigen tolerance, I discovered that some fish were infected with the Asian tapeworm toleransi, saya menemukan bahwa ikan yang terinfeksi dengan cacing pita Asia ( Bothriocephalus acheilognathi ) as a result of exposure at the hatchery where they were (Bothriocephalus acheilognathi) sebagai akibat dari eksposur pada pembenihan di mana mereka spawned. melahirkan. Asian tapeworms are intestinal parasites of teleosts (Hoffman 1998). cacing pita Asia adalah parasit usus teleosts (Hoffman 1998). These Ini tapeworms were probably introduced to the United States in the 1960s with grass carp cacing pita itu mungkin diperkenalkan ke Amerika Serikat pada 1960-an dengan mas rumput ( Cyprinus idella ), imported for aquatic vegetation control (Mitchell 2004). (Cyprinus idella), diimpor untuk pengendalian vegetasi akuatik (Mitchell 2004). The life cycle Siklus hidup of Asian tapeworms involves an intermediate stage with a copepod host, so a population dari cacing pita Asia melibatkan tahap intermediate dengan host copepoda, sehingga populasi of infected copepods was probably the mechanism of exposure in the hatchery ponds. copepoda terinfeksi mungkin mekanisme pemaparan di kolam pembenihan. Although Asian tapeworm infection has not yet been observed in wild populations of Meskipun infeksi cacing pita Asia belum diamati dalam populasi liar Topeka shiners, hatchery infections could threaten rehabilitation efforts. shiners Topeka, infeksi hatchery dapat mengancam upaya rehabilitasi. Widespread Luas introduction of Asian tapeworms into a threatened population could be devastating. pengenalan cacing pita Asia menjadi penduduk terancam bisa menghancurkan. Further study is needed because few studies have quantified the effects of Asian studi lebih lanjut diperlukan karena beberapa studi telah diukur efek Asia tapeworm infection on fish growth. infeksi cacing pita pada pertumbuhan ikan.

Information about Topeka shiner tolerance of high temperature, low dissolved Informasi tentang toleransi berlian Topeka temperatur tinggi, rendah terlarut oxygen, and tapeworm infection need to be better understood in order to protect critical oksigen, dan infeksi cacing pita perlu dipahami dengan lebih baik untuk melindungi kritis habitats and ensure the survival of this species. habitat dan menjamin kelangsungan hidup spesies ini. Therefore, the objectives of this study Oleh karena itu, tujuan penelitian ini were: 1) to determine the optimum temperature for growth, 2) to estimate the upper lethal adalah: 1) untuk menentukan suhu optimum untuk pertumbuhan, 2) untuk memperkirakan mematikan atas temperature [as critical thermal maximum (CTM)] after acclimation to high temperature, suhu [semaksimal termal kritis (CTM)] aklimasi setelah ke suhu tinggi, 3) to determine the effect of reduced dissolved oxygen on growth, 4) to estimate the 3) untuk menentukan efek mengurangi oksigen terlarut pada pertumbuhan, 4) untuk memperkirakan lower lethal concentration of dissolved oxygen, and 5) to determine the effect of Asian mematikan konsentrasi rendah oksigen terlarut, dan 5) untuk menentukan pengaruh Asia tapeworm infection on growth and CTM. infeksi cacing pita pada pertumbuhan dan CTM. Page 9 Page 9 44 Methods Metode General procedures Prosedur Umum Three experiments were conducted to determine the effect of temperature on Tiga percobaan yang dilakukan untuk menentukan pengaruh temperatur terhadap growth of Topeka shiners (T-1, T-2, and T-3) and two experiments were conducted to pertumbuhan shiners Topeka (T-1, T-2, dan T-3) dan dua percobaan telah dilakukan untuk determine the effect of oxygen on growth (O-1 and O-2). menentukan pengaruh oksigen terhadap pertumbuhan (O-1 dan O-2). Also, experiments to determine Selain itu, percobaan untuk menentukan the critical thermal maximum (CTM) (Becker and Genoway 1979) were conducted on termal maksimum kritis (CTM) (Becker dan Genoway 1979) dilakukan pada fish used in T-1, T-2, and T-3 (CTM-1, -2, and -3). ikan yang digunakan dalam T-1, T-2, dan T-3 (CTM-1, -2, dan -3). The effects of Asian tapeworm Pengaruh cacing pita Asia infection on growth and CTM were determined in experiments T-3 and CTM-3. infeksi pada pertumbuhan dan CTM ditentukan dalam percobaan T-3 dan CTM-3. Finally, Akhirnya, experiments were conducted on fish used in experiments O-1 and O-2 to determine lethal Percobaan dilakukan pada ikan yang digunakan dalam percobaan O-1 dan O-2 untuk menentukan mematikan oxygen concentrations for fish previously acclimated to low dissolved oxygen (LC konsentrasi oksigen untuk ikan sebelumnya menyesuaikan diri oksigen terlarut rendah (LC 50 50 -1 -1 and LC dan LC 50 50 -2). -2). All experiments except the CTM determinations were conducted using a flow- Semua percobaan kecuali CTM penentuan dilakukan dengan menggunakan aliran-

through system with water supplied by a deep dedicated well. melalui sistem dengan air yang disediakan oleh berdedikasi dalam sumur. Topeka shiners used in Topeka shiners digunakan dalam experiments T-1, CTM-1, O-1, and LC percobaan T-1, CTM-1, O-1, LC 50 50 -1 were obtained from the Lost Valley Fish -1 Diperoleh dari Ikan Lost Valley Hatchery (LVFH) Missouri, and Topeka shiners used in experiments T-2, CTM-2, T-3, Hatchery (LVFH) Missouri, dan shiners Topeka digunakan dalam percobaan T-2, CTM-2, T-3, CTM-3, O-2, and LC CTM-3, O-2, dan LC 50 50 -2 were from the University of Kansas (KU). -2 Berasal dari University of Kansas (KU). All fish came from ikan Semua berasal dari an original breeding population collected from Deep Creek at Pillsbury Crossing (T11S, sebuah peternakan penduduk asli yang dikumpulkan dari Deep Creek di Pillsbury Crossing (T11S, R9E, Sec 5), Riley County, Kansas. R9E, Sec 5), Riley County, Kansas. No tapeworms were ever detected in the fish from LVFH. Tidak ada cacing pita yang pernah terdeteksi dalam ikan dari LVFH. The fish from KU Ikan dari KU were infected with the Asian tapeworm because grass carp had previously been held in terinfeksi dengan cacing pita ikan mas rumput Asia karena sebelumnya telah diadakan di the rearing ponds and thus infected the intermediate host copepods. kolam pembesaran dan dengan demikian menginfeksi host copepoda menengah. For experiments T-3, Untuk percobaan T-3, CTM-3, and O-2, and LC CTM-3, dan O-2, dan LC 50 50 -2, Praziquantel (manufactured by Bayer as Droncit) was used -2, Praziquantel (diproduksi oleh Bayer sebagai Droncit) digunakan to rid the fish of the tapeworms. untuk membersihkan ikan dari cacing pita. Praziquantel dosage was 1.5 mg L Praziquantel dosis adalah 1,5 mg L -1 -1 for 24 hours as selama 24 jam sebagai Page 10 Page 10 55 recommended by Mitchell (2004) with fish density of about 0.15 g L direkomendasikan oleh Mitchell (2004) dengan kepadatan ikan sekitar 0,15 g L -1 -1 . . Microscopic Mikroskopis examination of the gastrointestinal track of over 60 fish treated with Praziquantel pemeriksaan jalur gastrointestinal lebih dari 60 ikan diperlakukan dengan Praziquantel indicated 100% removal of tapeworms with no fish mortality and no observed stress. menunjukkan 100% dari cacing pita penghapusan tanpa kematian ikan dan tidak stres diamati. Post-experiment dissection to determine the presence or absence of tapeworms was Posteksperimen pembedahan untuk menentukan ada atau tidak adanya cacing pita itu performed on all fish from experiments T-1, T-2, T-3 and CTM-3, and on randomly dilakukan pada semua ikan dari percobaan T-1, T-2, T-3 dan CTM-3, dan secara acak

sampled fish from experiment O-1. sampel ikan dari percobaan O-1. Fish from experiment O-2 were not dissected for Ikan dari percobaan O-2 tidak dibedah untuk tapeworms because by that time the Praziquantel treatment was assumed to be 100% cacing pita karena pada saat itu adalah perlakuan Praziquantel diasumsikan 100% effective. efektif. At the beginning of the acclimation period of all growth experiments, Topeka Pada awal periode aklimatisasi semua percobaan pertumbuhan, Topeka shiners of both sexes were distributed to test aquaria in a stratified-random manner so that shiners dari kedua jenis kelamin itu didistribusikan untuk menguji akuarium-acak dengan cara bertingkat sehingga every aquarium received one fish before any aquarium received a second fish, and so on. setiap akuarium menerima satu ikan akuarium yang diterima sebelum ikan kedua, dan seterusnya. Growth was measured as specific growth rate (SGR) over 28 to 30 days as percent Pertumbuhan diukur sebagai laju pertumbuhan spesifik (SGR) selama 28 sampai 30 hari sebagai persen growth per day (SGR = [ ln (final measure initial measure ) ] / [number of days elapsed pertumbuhan per hari (SGR = [ln (Ukuran akhir - awal mengukur)] / [jumlah hari berlalu between measurements]). antara ukuran]). At the beginning and end of each experiment, weight was Pada awal dan akhir setiap percobaan, berat itu recorded to the nearest 0.1 g and total length was recorded to the nearest 0.5 mm. dicatat ke 0,1 g terdekat dan panjang total tercatat ke pusat 0,5 mm. Before Sebelum these measurements were taken, food was withheld for 24 hours. diambil ukuran ini, makanan ditahan selama 24 jam. During the measuring Selama mengukur procedure, fish were anaesthetized with 100 mg L prosedur, ikan dibius dengan 100 mg L -1 -1 tricaine methanesulfate (MS-222). tricaine methanesulfate (MS-222). At the beginning of some experiments, individual fish were identified using either one or Pada awal dari beberapa percobaan, ikan individu diidentifikasi dengan menggunakan salah satu atau two fin clips. dua sirip klip. Preliminary experiments not reported here showed that fin clips did not Awal percobaan tidak dilaporkan di sini menunjukkan bahwa sirip klip tidak significantly lower Topeka shiner growth rate over 30 days. secara signifikan lebih rendah Topeka berlian laju pertumbuhan lebih dari 30 hari. Shiners were fed frozen Shiners diberi beku brine shrimp ( Artemia salina ) once per day during holding periods, two to three times per air garam udang (Artemia salina) sekali per hari selama periode memegang, dua atau tiga kali per day during acclimation periods, and three times per day to satiation during test periods. hari selama periode aklimatisasi, dan tiga kali per hari untuk pemuas selama periode uji. Deaths during growth experiments were recorded as missing data points. Kematian selama percobaan pertumbuhan dicatat sebagai titik data hilang. Photoperiod for Fotoperiodik untuk Page 11 Page 11 66 experiment T-1 was 16h light: 8h dark, but for all other experiments photoperiod was 9h percobaan T-1 adalah 16h terang: gelap 8h, tetapi untuk semua percobaan lainnya fotoperiodik adalah 9h light: 15h dark. terang: gelap 15h. A shorter daylength was used in an attempt to suppress reproductive Sebuah daylength lebih pendek digunakan dalam upaya untuk menekan reproduksi

behavior. perilaku. Fluorescent lights provided illumination during daylight hours. pencahayaan lampu fluorescent yang disediakan pada siang hari. During all experiments except CTM determinations, flow rate to each aquarium Selama semua percobaan penentuan kecuali CTM, laju alir untuk setiap akuarium was regulated with pipette tips. telah diatur dengan tips pipet. Desired test temperatures were achieved by mixing warm suhu uji yang diinginkan dicapai dengan mencampur hangat and cold water, and by using 200- and 300-watt Marineland Visi-Therm Deluxe dan air dingin, dan dengan menggunakan 200 - dan 300-watt Marineland Visi-Therm Deluxe submersible heaters in mixing chambers. submersible pemanas di ruang pencampuran. Foam insulation was placed around aquaria and Insulasi busa ditempatkan di akuarium dan tubing to maintain high water temperatures. tabung untuk menjaga suhu air yang tinggi. Onset Stowaway Tidbit temperature loggers Mulai berita menarik penumpang gelap suhu penebang were placed in each aquarium to monitor temperature, and records were later downloaded ditempatkan di setiap akuarium untuk memonitor suhu, dan catatan kemudian download to Microsoft Excel using BoxCar Pro software. ke Microsoft Excel dengan menggunakan software Pro gerbong. Temperature and dissolved oxygen were Suhu dan oksigen terlarut also monitored daily during experiments using digital thermometers and a YSI dissolved juga dipantau tiap hari selama percobaan menggunakan termometer digital dan YSI terlarut oxygen meter. oksigen meter. The oxygen meter was calibrated using the manufacturer's air calibration Oksigen meter dikalibrasi dengan menggunakan produsen udara kalibrasi procedure and periodically checked against dissolved oxygen concentrations using the prosedur dan diperiksa secara berkala terhadap konsentrasi oksigen terlarut dengan menggunakan Winkler method (APHA 1998). Metode Winkler (APHA 1998). Optimum Temperature for Growth Suhu optimum untuk Pertumbuhan The goals of experiments T-1, T-2, and T-3 were to determine the Topeka Tujuan percobaan T-1, T-2, dan T-3 adalah untuk menentukan Topeka shiner's optimum temperature for growth. berlian's temperatur optimum untuk pertumbuhan. Optimum temperature for growth is important suhu optimum untuk pertumbuhan adalah penting information because a high level of somatic fish growth indicates that the fish is able to informasi karena tingkat tinggi pertumbuhan ikan somatik menunjukkan bahwa ikan dapat put large amounts of energy towards growth. menempatkan sejumlah besar energi terhadap pertumbuhan. Optimum temperature for growth may not Suhu optimum untuk pertumbuhan mungkin tidak indicate optimum temperature for all aspects of the fish's life, but it does indicate that menunjukkan suhu optimum untuk semua aspek kehidupan ikan, tetapi tidak menunjukkan bahwa metabolism is efficient; the fish is at a temperature high enough for growth, yet low metabolisme yang efisien; ikan tersebut berada pada temperatur cukup tinggi untuk pertumbuhan, namun rendah enough to avoid thermal stress. cukup untuk menghindari stres termal. Page 12 Page 12 77 To determine the optimum temperature for growth, growth rate was measured Untuk menentukan suhu optimal untuk pertumbuhan, laju pertumbuhan diukur

over a range of five treatment temperatures with two replicates of each treatment (Table rentang lima suhu pengobatan dengan dua kali ulangan setiap perlakuan (Tabel 1). 1). Each replicate consisted of one 20-liter aquarium for a total of 10 aquaria. Setiap replikasi terdiri dari satu liter akuarium 20 untuk total 10 akuarium. Flow rate Laju alir to each aquarium was 600 ml min untuk masing-masing akuarium minimal 600 ml -1 -1 .. Experiment T-1. In this 30-day experiment, 14 Topeka shiners (13 months old) Percobaan T-1 .- Dalam percobaan 30 hari, 14 Topeka shiners (13 bulan) were placed in each replicate treatment, for totals of 10 aquaria and 140 fish. ditempatkan di setiap mereplikasi perawatan, untuk total 10 akuarium dan 140 ikan. Before the Sebelum experiment began, water in all aquaria was at the fish's acclimation temperature (22 C), percobaan dimulai, air di akuarium semua berada di's aklimatisasi suhu ikan (22 C), and temperatures were changed daily over 10 days to reach desired treatment levels. dan suhu yang berubah setiap hari selama 10 hari untuk mencapai tingkat perawatan yang diinginkan. Fish Ikan were not individually marked for identification; therefore, specific growth rate was tidak individual ditandai untuk identifikasi, sehingga, laju pertumbuhan spesifik calculated from the mean initial and final weights and lengths of fish in each replicate dihitung dari berat awal dan akhir mean dan panjang ikan di setiap ulangan treatment. pengobatan. At the conclusion of the experiment, all fish from the two highest temperature Di akhir percobaan, semua ikan dari temperatur tertinggi dua treatments were used in experiment CTM-1. perlakuan percobaan yang digunakan adalah CTM1. TABLE 1 TABEL 1 .Mean and standard deviation (in parenthesis) of at least 50 randomly sampled .- Rata-rata dan deviasi standar (dalam kurung) minimal 50 sampel secara acak temperature (C) logger data points for experiments T-1, T-2, and T-3, for all treatments suhu (C) titik data logger untuk eksperimen T-1, T-2, dan T-3, untuk semua perawatan and replications (Rep). dan ulangan (Rep). Treatment Pengobatan 11 22 33 44 55 Experiment Percobaan Rep 1 Rep 2 Rep 1 Rep 2 Rep 1 Rep 2 Rep 1 Rep 2 Rep 1 Rep 2 Rep 1 Rep 2 Rep 1 Rep 2 Rep 1 Rep 2 Rep 1 Rep 2 Rep 1 Rep 2 T-1 T-1 12.7 12,7 (0.24) (0,24) 12.8 12,8 (0.26) (0,26) 18.0 18,0 (0.24) (0,24)

18.2 18,2 (0.24) (0,24) 23.3 23,3 (0.24) (0,24) 23.4 23,4 (0.24) (0,24) 27.8 27,8 (0.18) (0,18) 27.7 27,7 (0.21) (0,21) 33.0 33,0 (0.35) (0,35) 32.9 32,9 (0.41) (0,41) T-2 T-2 12.3 12,3 (0.26) (0,26) 13.0 13,0 (0.53) (0,53) 17.1 17,1 (0.81) (0,81) 17.9 17,9 (0.88) (0,88) 22.9 22,9 (0.63) (0,63) 22.4 22,4 (0.67) (0,67) 26.8 26,8 (0.41) (0,41) 27.0 27,0 (0.41) (0,41) 31.1 31,1 (0.40) (0,40) 31.3 31,3 (0.34) (0,34) T-3 T-3 14.5 14,5 (0.43) (0,43) 14.7 14,7 (0.52) (0,52) 19.4 19,4 (0.53) (0,53) 19.9 19,9 (0.50) (0,50) 24.4 24,4 (0.48) (0,48)

24.5 24,5 (0.50) (0,50) 29.6 29,6 (0.17) (0,17) 29.6 29,6 (0.26) (0,26) 34.4 34,4 (0.39) (0,39) 34.8 34,8 (0.25) (0,25) Behavior was observed once a day on nine different days, and aquaria were Perilaku diamati sekali sehari pada sembilan hari yang berbeda, dan wadah observed in random order for 10 seconds each. diamati secara acak selama 10 detik setiap. Swimming activity in each aquarium was Aktivitas berenang di dalam akuarium masing-masing subjectively classified as high, medium, or low, and was given a score of 3, 2, or 1, subyektif diklasifikasikan sebagai tinggi, sedang, atau rendah, dan diberi skor 3 2,, atau 1, Page 13 Page 13 88 respectively. masing. Reproductive behavior (subjectively defined here as a male actively Reproduksi perilaku (subjektif didefinisikan di sini sebagai laki-laki aktif chasing other fish out of his territory) was also classified in each aquarium as high, mengejar keluar ikan lainnya dari wilayahnya) juga diklasifikasikan dalam setiap akuarium sebagai tinggi, medium, or low, corresponding to scores of 3, 2, or 1, and the number of males exhibiting menengah, atau rendah, sesuai dengan nilai dari 3 2,, atau 1, dan jumlah laki-laki menunjukkan chasing behavior was recorded. perilaku mengejar tercatat. The reproductive activity score was calculated by adding Nilai kegiatan reproduktif telah dihitung dengan menambahkan the reproductive behavior score to the number of mature males (those with reddened fins, skor perilaku reproduksi dengan jumlah laki-laki dewasa (yang memerah sirip, which were chasing others out of their territory). yang mengejar orang lain di luar wilayah mereka). Experiment T-2. In this 28-day experiment, 8-month old Topeka shiners Percobaan T-2 .Dalam percobaan 28 hari, 8-bulan tua Topeka shiners infected with Asian tapeworms were used, although the tapeworms were not discovered terinfeksi cacing pita Asia digunakan, meskipun tidak ditemukan cacing pita until after the test was completed. sampai setelah ujian selesai. The fish were randomly distributed among five ikan secara acak didistribusikan di antara lima treatments with two replicates (Table 1). perawatan dengan dua kali ulangan (Tabel 1). Each replicate contained 10 fish, for a total of Setiap mereplikasi berisi 10 ikan, untuk total 10 aquaria and 100 fish. 10 akuarium dan 100 ikan. All aquaria began at the acclimation temperature of the fish (23 akuarium Semua mulai pada suhu aklimatisasi ikan (23 C), and temperatures were changed daily over 8 days to reach desired treatment levels. C), dan suhu yang berubah setiap hari selama 8 hari untuk mencapai tingkat perawatan yang diinginkan. Fish were given either one or two fin clips at the time of initial measuring to identify Ikan diberi satu atau dua klip sirip pada saat pengukuran awal untuk mengidentifikasi

individuals. individu. Initial and final weights and lengths from each fish were used to calculate dan terakhir bobot awal dan panjang dari masing-masing ikan yang digunakan untuk menghitung specific growth rate for individual fish. laju pertumbuhan spesifik ikan individu. At the conclusion of the experiment, all fish from Di akhir percobaan, semua ikan dari the two highest temperature treatments were used in experiment CTM-2. dua perlakuan suhu tertinggi percobaan yang digunakan adalah CTM-2. Experiment T-3. In this 28-day experiment, 12-month old Topeka shiners from Percobaan T-3 .- Dalam percobaan 28 hari, 12-bulan tua Topeka shiners dari the same cohort as fish used in experiment T-2 were randomly distributed among five kohort yang sama seperti ikan yang digunakan dalam percobaan T-2 didistribusikan secara acak di antara lima treatments with two replicates (Table 1). perawatan dengan dua kali ulangan (Tabel 1). Each replicate contained 14 fish, for a total of Setiap mereplikasi berisi 14 ikan, untuk total 10 aquaria and 140 fish. 10 akuarium dan 140 ikan. In each replicate, seven fish had been treated with Praziquantel Dalam setiap kali ulangan, tujuh ikan telah diperlakukan dengan Praziquantel 2 weeks prior to the start of the experiment to rid them of tapeworms. 2 minggu sebelum memulai percobaan untuk menyingkirkan mereka dari cacing pita. The other seven Yang lainnya tujuh fish were not treated and were assumed to be 100% infected. ikan tidak diperlakukan dan diasumsikan 100% terinfeksi. All aquaria began at the akuarium Semua dimulai di fish's holding temperature (20 C), and water temperatures were changed daily over 13 suhu memegang ikan (20 C), dan suhu air berubah setiap hari lebih dari 13 Page 14 Page 14 99 days to reach desired treatment temperatures. hari untuk mencapai suhu pengobatan yang diinginkan. Fish were fin-clipped to identify Sirip ikan-dipotong untuk mengidentifikasi individuals, so that growth of infected fish could be compared with that of non-infected individu, sehingga pertumbuhan ikan bisa terinfeksi dibandingkan dengan yang tidak terinfeksi fish. ikan. Specific growth rates for individual fish were determined as in T-1 and T-2. tingkat pertumbuhan yang khusus untuk ikan individu ditentukan seperti pada T-1 dan T-2. At the Pada conclusion of the experiment, all fish from the 30 C temperature treatment were used in Kesimpulan dari hasil penelitian, semua ikan dari suhu perlakuan C 30 telah digunakan dalam experiment CTM-3. percobaan CTM-3. Critical Thermal Maximum Kritis Thermal Maksimum The goal of this group of experiments (CTM-1, -2, and -3) was to determine upper Tujuan dari kelompok eksperimen (CTM-1, -2, dan -3) adalah untuk menentukan atas lethal temperatures of Topeka shiners acclimated to high temperatures. suhu mematikan shiners Topeka menyesuaikan suhu yang tinggi. The fish used in Ikan yang digunakan dalam these experiments had been acclimated to high temperatures as a result of undergoing percobaan ini telah menyesuaikan diri suhu tinggi sebagai akibat dari menjalani high temperature treatments in growth tests T-1, T-2, or T-3. perlakuan suhu tinggi dalam tes pertumbuhan T-1, T-2, atau T-3. The apparatus used to alat yang digunakan untuk determine CTM was an aerated 10-liter static-water aquarium. menentukan CTM adalah liter soda 10-statis-air akuarium. An MGW Lauda MS MS Lauda MGW

heater/ chiller/ mixer, with precise temperature control, was placed in one end. heater / chiller / mixer, dengan kontrol suhu yang tepat, ditempatkan di salah satu ujungnya. A small Sebuah kecil mesh enclosure was placed in the other end that allowed the fish access to the surface but kandang mesh ditempatkan di ujung lainnya yang memungkinkan akses ke permukaan ikan tetapi kept the fish away from the heater's circulating motor. terus ikan dari pemanas yang beredar motor. The initial aquarium temperature Akuarium suhu awal was set at the fish's acclimation temperature. ditetapkan's aklimatisasi suhu ikan. One, two, three, or four fish along with a Satu, dua, tiga, atau empat ikan bersama dengan digital thermometer probe were placed in the mesh container and fish were given one probe termometer digital ditempatkan dalam wadah mesh dan ikan diberi satu minute to acclimate before temperature was increased. menit untuk menyesuaikan diri kpd suatu iklim sebelum suhu meningkat. Water temperature was raised 0.3 Temperatur air dibesarkan 0,3 C per minute, and the CTM endpoint was reached when the fish could no longer regain C per menit, dan titik akhir tercapai CTM saat ikan tidak bisa lagi kembali equilibrium (Becker and Genoway 1979). kesetimbangan (Becker dan Genoway 1979). CTM values were recorded for individual fish. nilai CTM direkam untuk ikan individu. Experiment CTM-1. Twenty-eight Topeka shiners from the 28 C treatment and Percobaan CTM-1 .- Dua puluh delapan Topeka shiners dari perlakuan C 28 dan 23 fish from the 33 C treatment from experiment T-1 were used in this experiment, 12 23 ikan dari perlakuan C 33 dari percobaan T-1 yang digunakan dalam percobaan ini, 12 Page 15 Page 15 10 10 days after the conclusion of the growth experiment. hari setelah akhir percobaan pertumbuhan. During the holding period between Selama periode memegang antara T-1 and CTM-1 fish were held at 28 and 31 C, respectively. T-1 dan CTM-1 ikan diadakan pada 28 dan 31 C, masing-masing. Experiment CTM-2. Sixteen Topeka shiners from the 27 C treatment and 14 Percobaan CTM2 .- Enam belas Topeka shiners dari perlakuan C 27 dan 14 fish from the 32 C treatment from experiment T-2 were used in this CTM determination, ikan dari perlakuan C 32 dari percobaan T-2 digunakan dalam penentuan CTM, conducted within 1 week after conclusion of the growth experiment. dilakukan dalam waktu 1 minggu setelah kesimpulan dari percobaan pertumbuhan. Experiment CTM-3. Nineteen fish without Asian tapeworms and six fish with Percobaan CTM-3 .- Sembilan belas ikan tanpa cacing pita Asia dan enam ikan dengan Asian tapeworms from the 30 C treatment in T-3 were used within 1 week after cacing pita Asia dari perlakuan C 30 di T-3 digunakan dalam waktu 1 minggu setelah conclusion of the growth experiment (presence or absence of Asian tapeworm was kesimpulan dari percobaan pertumbuhan (ada atau tidak adanya cacing pita Asian verified through dissection after completing the experiment). diverifikasi melalui pembedahan setelah menyelesaikan percobaan). Fish from the highest Ikan dari yang tertinggi treatment temperature (35 C) in T-3 were not used because of high mortality at that perlakuan suhu (35 C) pada T-3 tidak digunakan karena kematian yang tinggi pada temperature. suhu.

Effect of Dissolved Oxygen on Growth Pengaruh Oksigen terlarut terhadap Pertumbuhan The goals of experiments O-1 and O-2 were to determine the effect of reduced Tujuan percobaan O-1 dan O-2 untuk mengetahui pengaruh penurunan dissolved oxygen concentration on growth for Topeka shiners at relatively high konsentrasi oksigen terlarut pada pertumbuhan untuk shiners Topeka di relatif tinggi temperatures. suhu. Dissolved oxygen concentration was controlled using an oxygen ladder konsentrasi oksigen terlarut dikontrol menggunakan tangga oksigen modified from Brungs (1971). dimodifikasi dari Brungs (1971). Briefly, cold well water was heated in a head tank and Singkatnya, air sumur dingin dipanaskan di kepala dan tangki vigorously aerated to bring dissolved gases to atmospheric equilibrium. penuh semangat soda untuk membawa gas terlarut untuk keseimbangan atmosfer. Water at the Air di desired test temperature then flowed by gravity to a stripping column in which nitrogen suhu uji yang diinginkan oleh gravitasi dialirkan ke kolom stripping di mana nitrogen was bubbled through the water in a counter current direction to reduce the dissolved itu menggelegak melalui air di arah arus berlawanan dengan mengurangi terlarut oxygen concentration to near zero. oksigen konsentrasi untuk mendekati nol. From the stripping column, water flowed to the top of Dari kolom pengupasan, air mengalir ke atas the oxygen ladder, then flowed over baffles separating individual compartments, and tangga oksigen, kemudian mengalir ke baffle memisahkan kompartemen individu, dan finally to a drain. akhirnya ke saluran pembuangan. As water moved over the baffles it gradually absorbed oxygen from the Ketika air bergerak selama baffle secara bertahap menyerap oksigen dari Page 16 Page 16 11 11 atmosphere. atmosfer. Airstones were placed in some of the compartments of the ladder to increase Airstones ditempatkan di beberapa kompartemen tangga untuk meningkatkan the rate of oxygen absorption. tingkat penyerapan oksigen. Water was withdrawn from the compartments of the Air ditarik dari kompartemen dari ladder where the desired treatment concentrations had been attained. tangga di mana konsentrasi perlakuan yang diinginkan telah tercapai. For all three experiments, the desired treatment concentrations were saturation, 5, 4, 3, Untuk semua tiga percobaan, perlakuan konsentrasi yang diinginkan adalah saturasi, 5, 4, 3, and 2 mg L dan 2 mg L -1 -1 of dissolved oxygen. oksigen terlarut. However, temperature fluctuations as well as calcium Namun, fluktuasi suhu serta kalsium deposits and algal growth in the tubing caused some variability in the actual dissolved deposito dan pertumbuhan alga dalam tabung menyebabkan variabilitas beberapa di terlarut aktual oxygen concentrations (Table 2). oksigen konsentrasi (Tabel 2). Experimental chambers were 40-liter aquaria with ruang eksperimen telah akuarium 40 liter dengan water inflow positioned diagonally opposite water outflow. masukan air diposisikan secara diagonal keluar air yang berlawanan. Fish were individually Ikan secara individu identified with unique fin clips. diidentifikasi dengan klip sirip unik. Specific growth rate was measured as described in the laju pertumbuhan khusus telah diukur seperti yang dijelaskan dalam

temperature experiments above. suhu percobaan di atas. After conclusion of the experiments, fish from the two Setelah kesimpulan dari percobaan, ikan dari dua lowest dissolved oxygen levels were used in the LC terendah tingkat oksigen terlarut digunakan di LC 50 50 determinations. penentuan. TABLE 2. TABEL 2 .Mean dissolved oxygen concentrations (mg L Berarti konsentrasi oksigen terlarut (mg L -1 -1 ) for the five treatments in ) Selama lima perawatan di experiments O-1, and O-2, with standard error in parenthesis. percobaan O-1, dan O-2, dengan kesalahan standar dalam tanda kurung. Treatment Pengobatan Experiment Fish age (months) Ikan Percobaan umur (bulan) 11 22 33 44 55 O-1 O-1 19 19 2.02 2,02 (0.0396) (0,0396) 2.91 2,91 (0.0593) (0,0593) 3.87 3,87 (0.0419) (0,0419) 4.78 4,78 (0.0560) (0,0560) 6.14 6,14 (0.0865) (0,0865) O-1 O-1 77 2.42 2,42 (0.0474) (0,0474) 3.12 3,12 (0.0615) (0,0615) 4.31 4,31 (0.0530) (0,0530) 5.23 5,23 (0.0581) (0,0581) 6.27 6,27 (0.0861) (0,0861) O-2 O-2 18 18 2.42 2,42

(0.0527) (0,0527) 3.46 3,46 (0.0445) (0,0445) 4.31 4,31 (0.0552) (0,0552) 5.23 5,23 (0.0833) (0,0833) 6.27 6,27 (0.0426) (0,0426) Experiment O-1. This 28-day experiment was conducted at a temperature of Percobaan O-1 .ini percobaan 28 hari dilakukan pada suhu 24.8 C ( 0.8 C) based on the findings of experiment T-1. 24,8 C ( 0,8 C) berdasarkan temuan dari percobaan T-1. This temperature was suhu ini considered at the time of O-1 to be optimum for growth. dipertimbangkan pada saat O-1 akan optimal untuk pertumbuhan. Both 19 month old and 7- Kedua berusia 19-bulan dan 7 Page 17 Page 17 12 12 month old Topeka shiners were used in each of the five treatments. bulan tua Topeka shiners digunakan dalam masing-masing dari lima perlakuan. Plastic divider Plastik pembagi screens placed parallel to the shorter sides of the aquarium separated older and younger layar ditempatkan sejajar dengan sisi yang lebih pendek dari akuarium terpisah lebih tua dan lebih muda fish. ikan. The position of older and younger fish in relation to inflow and outflow was Posisi ikan yang lebih tua dan muda dalam kaitannya dengan masuk dan keluar adalah randomly selected in one replicate and was the opposite arrangement in the other dipilih secara acak dalam satu replikasi, dan sebaliknya adalah pengaturan yang lain replicate. ulangan. Dissolved oxygen and temperature were measured in each compartment. Oksigen terlarut dan suhu diukur di setiap kompartemen. The Itu compartments varied slightly in both temperature and oxygen, so the actual dissolved kompartemen bervariasi sedikit di kedua suhu dan oksigen, sehingga sebenarnya terlarut oxygen levels varied slightly between groups of fish. kadar oksigen bervariasi sedikit di antara kelompok ikan. Each replicate contained 12 fish in Setiap mereplikasi berisi 12 ikan di each age group, for a total of 24 fish per aquarium. setiap kelompok umur, untuk total 24 ikan per akuarium. Water flow rate was 270 ml min laju alir Air 270 ml min -1 -1 .. During the 11-day acclimation period, the initial oxygen concentrations of between 3.3 Selama periode aklimatisasi hari-11, konsentrasi oksigen awal antara 3,3 and 7.6 mg L dan 7,6 mg L -1 -1 were gradually lowered to the desired treatment concentrations while berangsur-angsur diturunkan dengan konsentrasi perlakuan yang diinginkan sementara temperature was increased from 21 C to 25 C. Dissolved oxygen started in this range of suhu meningkat dari 21 C sampai 25 C. oksigen terlarut dimulai dalam rentang

concentrations because the oxygen ladder's design did not allow for multiple water karena konsentrasi oksigen yang tangga desain tidak memungkinkan untuk beberapa air withdrawals from the same compartment in the oxygen ladder. penarikan dari kompartemen yang sama dalam tangga oksigen. The 7-month-old fish had The-month-old ikan 7 telah Asian tapeworm infection but this was not discovered until midway through the test. infeksi cacing pita Asia tetapi ini tidak ditemukan sampai pertengahan melalui tes. The 19-month-old fish did not have tapeworm infection. Bulan ini berusia 19 ikan tidak punya infeksi cacing pita. Fish from both age groups were Ikan dari kedua kelompok usia tersebut randomly chosen for dissection at the end of the experiment to verify the level of dipilih secara acak untuk pembedahan pada akhir percobaan untuk memastikan tingkat tapeworm infection. infeksi cacing pita. Experiment O-2. This 28-day experiment was designed to examine the Percobaan O-2 .- hari ini percobaan 28 dirancang untuk menguji relationship between dissolved oxygen (Table 2) and Topeka shiner growth at a hubungan antara oksigen terlarut (Tabel 2) dan bintang Topeka pertumbuhan di temperature higher than the optimum temperature for growth. pertumbuhan yang lebih tinggi daripada suhu suhu optimum untuk. The temperature during Suhu selama this experiment was 28.2 C ( 0.6 C). penelitian ini adalah 28,2 C ( 0,6 C). Eighteen-month old fish were used in each of the bulan tua ikan-Delapan belas digunakan dalam masing-masing five treatment levels, and the aquaria were not divided because only the one age group pengobatan tingkat lima, dan akuarium itu tidak dibagi karena hanya satu kelompok umur was used. digunakan. These fish were previously infected with Asian tapeworm, but treatment of all Ikan ini sebelumnya terinfeksi dengan cacing pita Asia, tetapi perlakuan terhadap semua Page 18 Page 18 13 13 fish with Praziquantel had eradicated the infection. ikan dengan Praziquantel telah diberantas infeksi. Each replicate contained 20 fish, and Setiap mereplikasi berisi 20 ikan, dan flow rate to each aquarium was 300 ml min laju alir masing-masing untuk akuarium 300 ml min -1 -1 . . During the 7-day acclimation period, Selama periode aklimatisasi hari-7, temperatures were slowly increased from 26 C, and oxygen was adjusted to the final suhu yang perlahan-lahan meningkat dari 26 C, dan oksigen disesuaikan dengan akhir treatment concentrations starting from a range between 3.1 and 7.1 mg L perlakuan konsentrasi mulai dari kisaran antara 3,1 dan 7,1 mg L -1 -1 as in seperti dalam experiment O-1. percobaan O-1. Four separate behavior trials were conducted in which activity was measured by Empat uji coba perilaku terpisah dilakukan dalam aktivitas yang diukur dengan counting line-crossings. menghitung garis-penyeberangan. A narrow side of each aquarium was divided into quarters using Sebuah sisi sempit setiap akuarium dibagi menjadi empat dengan menggunakan colored tape, and each time the fish in question crossed a tape line, it was counted as a pita berwarna, dan setiap kali ikan di pertanyaan menyeberangi garis tape, itu dihitung sebagai

line-crossing. line-persimpangan. Three different chaser fish and three non-chaser fish were observed Tiga berbeda "pemburu" ikan dan tiga "non-pemburu" ikan diamati for 30 seconds in each aquarium during each trial. selama 30 detik di setiap akuarium selama persidangan masing-masing. Chaser fish were selected as being "Chaser" ikan dipilih sebagai aggressive male fish that were most actively chasing other fish. ikan jantan agresif yang paling aktif mengejar ikan lainnya. Non-chaser fish were "Non-pemburu" ikan fish that were judged subjectively to be among the least active fish in the aquarium. ikan yang dinilai secara subjektif menjadi salah satu ikan paling aktif di dalam akuarium. Lower Lethal Dissolved Oxygen Concentrations Konsentrasi Lethal rendah Oksigen terlarut Two 96-hour LC Dua 96-jam LC 50 50 determinations were conducted to determine lower lethal penentuan dilakukan untuk menentukan lebih rendah mematikan oxygen concentrations. oksigen konsentrasi. Dissolved oxygen was measured twice per day using the Winkler Oksigen terlarut diukur dua kali per hari dengan menggunakan Winkler method (APHA 1998) and averaged for calculation of LC metode (APHA 1998) dan rata-rata untuk perhitungan LC 50 50 values (Table 3). nilai (Tabel 3). Experiment LC Percobaan LC 50 50 -1. This test began 11 days after the conclusion of O-1, and -1 .- Tes ini dimulai 11 hari setelah kesimpulan O-1, dan used fish from the two lowest dissolved oxygen treatments. digunakan ikan dari dua oksigen terlarut terendah perawatan. Fish remained at lowered Ikan tetap di menurunkan dissolved oxygen levels between the growth test and LC tingkat oksigen terlarut antara uji pertumbuhan dan LC 50 50 -1. -1. The arrangement of older Penyusunan lebih tua and younger fish in relation to inflow and outflow was randomized as in experiment O-1, dan ikan muda dalam kaitannya dengan masuk dan keluar secara acak seperti pada percobaan O-1, and age groups were separated with plastic screens. dan kelompok umur dipisahkan dengan layar plastik. Fish were exposed to three dissolved Ikan yang terkena tiga terlarut oxygen concentrations with two replicates each (Table 3). oksigen konsentrasi dengan dua kali ulangan masing-masing (Tabel 3). Each aquarium contained Setiap akuarium berisi Page 19 Page 19 14 14 eight fish from each age group. delapan ikan dari masing-masing kelompok usia. Flow rate to each aquarium was 390 ml min Laju Alir untuk akuarium masing-masing 390 ml min -1 -1 , and , Dan temperature was between 24.5 C ( 0.5 C). suhu adalah antara 24,5 C ( 0,5 C). Plastic covers were placed on the surface of Plastik mencakup ditempatkan pada permukaan the water but not sufficiently tight to prevent fish from having slight access to the water air tapi tidak cukup ketat untuk mencegah ikan dari sedikit memiliki akses ke air

surface. permukaan. Fish were not fed during the 96 hours, and deaths were recorded every 24 hours. Ikan tidak diberi makan selama 96 jam, dan kematian dicatat setiap 24 jam. Experiment LC Percobaan LC 50 50 -2. This test began 2 days after the conclusion of O-2 and used -2 .- Tes ini dimulai 2 hari setelah kesimpulan O-2 dan digunakan fish from the lowest two dissolved oxygen treatments. ikan dari terendah dua perawatan oksigen terlarut. Four treatments with two Empat perawatan dengan dua replicates were used (Table 3), for a total of eight 40-liter aquaria. ulangan digunakan (Tabel 3), untuk total delapan akuarium 40 liter. Each aquarium Setiap akuarium contained either 8 or 9 fish. berisi baik 8 atau 9 ikan. Flow rate to each aquarium was 300 ml min Laju Alir untuk akuarium masing-masing 300 ml min -1 -1 , and , Dan temperature was 27.8 C ( 0.4 C). suhu 27,8 C ( 0,4 C). Plastic covers with foam insulation around the edges Plastik tutup dengan isolasi busa di tepinya prevented fish from gaining access to the surface more effectively than in LC mencegah ikan dari mendapatkan akses ke permukaan yang lebih efektif daripada di LC 50 50 -1. -1. Fish Ikan were not fed during the 96 hours, and deaths were recorded every 24 hours. tidak diberi makan selama 96 jam, dan kematian dicatat setiap 24 jam. TABLE 3. TABEL 3 .Mean and standard error (in parenthesis) of dissolved oxygen Mean dan standard error (dalam kurung) oksigen terlarut concentrations (mg L konsentrasi (mg L -1 -1 ) for treatments in experiments LC ) Untuk perawatan dalam percobaan LC 50 50 -1, and LC -1, Dan LC 50 50 -2. -2. Treatment Pengobatan Experiment Percobaan Fish Age Umur Ikan (months) (Bulan) 11 22 33 44 LC LC 50 50 -1 -1 20 20 0.62 0,62 (0.0478) (0,0478)

0.87 0,87 (0.0436) (0,0436) 2.16 2,16 (0.0450) (0,0450) -LC LC 50 50 -1 -1 88 0.64 0,64 (0.0592) (0,0592) 0.88 0,88 (0.0417) (0,0417) 2.17 2,17 (0.0460) (0,0460) -LC LC 50 50 -2 -2 19 19 0.46 0,46 (0.0499) (0,0499) 0.80 0,80 (0.0395) (0,0395) 1.15 1,15 (0.0174) (0,0174) 1.98 1,98 (0.0547) (0,0547) Page 20 Page 20 15 15 Data Analysis Analisis Data Optimum temperature. Optimum temperatures for growth were calculated by Suhu optimum .suhu optimum untuk pertumbuhan dihitung oleh plotting mean SGR values for each replicate vs. mean aquarium temperature for merencanakan berarti nilai SGR untuk setiap mereplikasi vs berarti suhu akuarium untuk experiment T-1, and by plotting the median SGR value for each replicate vs. mean percobaan T1, dan dengan memetakan nilai SGR rata-rata untuk setiap replikasi vs berarti aquarium temperature for experiments T-2 and T-3. akuarium temperatur untuk percobaan T-2 dan T-3. Median values were used because nilai rata-rata digunakan karena they eliminated the decision of whether or not to remove outliers, while adequately mereka dieliminasi keputusan apakah atau tidak untuk menghilangkan pencilan, sementara cukup representing the central tendency of fish in the sample; however, median values could not mewakili kecenderungan pusat ikan dalam sampel, namun nilai rata-rata tidak bisa be used in analysis of experiment T-1 because individual fish were not identified. digunakan dalam analisis percobaan T-1 karena ikan individu tidak diidentifikasi. Mean Berarti

values weighted for number of fish per replicate were used for experiment T-1. nilai tertimbang untuk jumlah ikan per mereplikasi digunakan untuk percobaan T-1. Third Ketiga order polynomial regression was performed in Arc, a statistical regression program (Cook regresi polinomial order dilakukan di Arc, program regresi statistik (Cook and Weisberg, 1999) using the Delta method (Weisberg, 2005) to calculate estimates and dan Weisberg, 1999) dengan menggunakan metode Delta (Weisberg, 2005) untuk menghitung perkiraan dan standard errors for each experiment's optimum temperature for growth. standar kesalahan untuk itu setiap percobaan temperatur optimum untuk pertumbuhan. Third order Urutan ketiga regression was used because it gave the best fit based on R-squared values. regresi digunakan karena memberikan sesuai terbaik berdasarkan nilai-nilai R-squared. A separate Sebuah terpisah linear regression was performed in Arc on data from experiment T-3 to examine regresi linier dilakukan di Arc pada data dari percobaan T-3 untuk memeriksa significant effects of tapeworm infection on fish growth. signifikan dampak infeksi cacing pita pada pertumbuhan ikan. Chi-square tests were Uji Chi-square adalah performed for T-1, T-2, and T-3 to compare survival rate between treatments for each dilakukan untuk T-1, T-2, dan T-3 untuk membandingkan tingkat kelangsungan hidup antara perawatan untuk masing-masing experiment. percobaan. Also, two t-tests were used to compare survival rates between fish with and Juga, dua t-tes digunakan untuk membandingkan tingkat ketahanan hidup antara ikan dengan dan without tapeworm infection for experiment T-3; one t-test included all five treatments, tanpa infeksi cacing pita untuk percobaan T-3; satu t-test termasuk kelima perawatan, and the other excluded the highest temperature treatment (35 C) because of low survival. dan yang lainnya dikecualikan dengan suhu tertinggi (35 C) karena kelangsungan hidup rendah. Activity scores recorded during T-1 were first examined visually for time trends Aktivitas nilai tercatat selama T-1 pertama kali diperiksa secara visual untuk tren waktu (eg a time trend exists if one aquarium shows consistent increase in activity over time) (Misalnya waktu ada kecenderungan jika salah satu akuarium konsisten menunjukkan peningkatan aktivitas dari waktu ke waktu) and then replicates were averaged and examined visually for any patterns occurring dan kemudian ulangan adalah rata-rata dan diperiksa secara visual untuk setiap pola yang terjadi between treatments. antara perlakuan. Each aquarium's score was then averaged over all observations, Masing-masing akuarium skornya kemudian rata-rata seluruh pengamatan, Page 21 Page 21 16 16 resulting in 10 data points total ( n =2 for each of five treatments). menghasilkan total 10 titik data (n = 2 untuk setiap lima perawatan). One-way ANOVAs Satu-cara ANOVAs were then performed in Statistica (StatSoft, Inc., 1997) to determine if temperature had a kemudian dilakukan di Statistica (StatSoft, Inc, 1997) untuk menentukan apakah suhu punya significant effect on either swimming activity or reproductive behavior. signifikan berpengaruh terhadap aktivitas baik berenang atau perilaku reproduktif. Critical Thermal Maximum. Estimates of mean, standard deviation, and Kritis Thermal Maksimum Perkiraan .-, standar deviasi mean, dan

standard error of the mean were calculated for each experiment. standard error dari mean dihitung untuk setiap percobaan. Different acclimation Berbeda aklimatisasi temperature groups were separated for experiments CTM-1 and CTM-2, and fish treated kelompok suhu dipisahkan untuk eksperimen CTM-1 dan CTM-2, dan ikan perlakuan and not treated for tapeworm were separated for CTM-3. dan tidak diperlakukan untuk cacing pita dipisahkan untuk CTM-3. To test for significant differences in CTM between groups, t-tests were performed in Microsoft Excel. Dissolved Oxygen Effect on Growth and Behavior. Specific growth rates for each fish were calculated in experiments O-1 and O-2. For experiment O-1, a split-plot design ANOVA was used for analysis in the following procedure. First, a model including oxygen level, fish age group, and interactions between aquarium and oxygen level as well as age and oxygen level was fitted in Arc to examine within-treatment differences and effect of age on growth. Next, separate one-way ANOVAs and post-hoc Tukey tests were performed in Statistica for each age group to examine only effect of dissolved oxygen level on growth. For experiment O-2, a one-way ANOVA and posthoc Tukey test was performed in Statistica to test for effect of dissolved oxygen on growth. pertumbuhan. The behavior observations (line crossings) that were recorded during O-2 were analyzed with a 2-way ANOVA and post-hoc Tukey test in Statistica to determine significant differences in activity between oxygen treatment levels and between 'chaser' and 'non-chaser' fish. Finally, Chi-square tests were performed for both O-1 and O-2, comparing survival rates between treatments for each experiment. Page 22 Page 22 17 17 Lethal Oxygen Concentrations . For both LC 50 50 -1 and LC 50 50 -2, the number of deaths at 96 hours was used to calculate LC 50 50 values using the binary method (Newman 1995). 1995). Results Hasil Optimum Temperature and Effect of Asian Tapeworm on Growth Experiment T-1. The optimum temperature for growth as SGR in weight (ie mass) was 18.0 C (SE 0.348), and for length 18.4 C (SE 0.604) (Table 4, Figure 1). All Semua treatments had 100% survival except for the highest treatment temperature (33 C) in which 28 of 29 fish survived. Temperature had no significant effect on fish survival ( P= 0.417). Dissections revealed no tapeworms in any of the fish. Reproductive behavior Reproduksi perilaku was significantly higher at the 23 C treatment and above ( P <0.05), and activity was significantly higher at two highest temperature treatments compared to the lowest temperature treatment ( P <0.05) (Figure 2). No time trends were found for any of the behavior observations.

Experiment T-2. The optimum temperature for growth in weight was 32.1 C (SE 2.32), and 29.5 C (SE 1.36) for length. However, the highest mean treatment temperature was 31.2 C and therefore the estimate for weight is an extrapolation. The Itu highest growth rate for weight occurred at the highest temperature tested (Figure 3, Table 4). 4). Survival was 90%, 75%, 90%, 85%, and 70% for the five treatments in order of increasing temperature, and no significant effect of temperature was found on fish survival rate ( P= 0.346). Post-experiment dissections revealed that Asian tapeworms Page 23 Page 23 18 18 were present in fish in all treatments, with 94%, 89%, 100%, 94%, and 43% infection in the five treatments, in order of increasing temperature. TABLE 4 TABEL 4 . Optimum temperature for growth (SE in parenthesis), treatment temperature where highest specific growth rate (SGR) occurred, and that treatment's mean SGR (%/day) in experiments T-1, T-2, and T-3. Experiment T-3. Fish that had been treated with Praziquantel, and therefore were free of tapeworms during the experiment, had an optimum temperature for growth in weight of 27.4 C (SE 0.811) and 27.1 C (SE 0.757) for growth in length (Figure 4, Table 4). Tabel 4). For fish that were infected with tapeworms (those that had not received the Praziquantel treatment), the optimum temperature for growth in weight was 28.1 C (SE 0.429), and for length it was 26.3 C (SE 1.63) (Figure 4, Table 4). In weight, growth rate Experiment Percobaan Measure Mengukur Fish age (months) (Bulan) Optimum Optimal Temperature Suhu (C) (C) Temperature Suhu with highest dengan tertinggi mean SGR (C) Highest Paling tinggi Mean Berarti SGR SGR T-1 T-1 Weight Berat 13 13 18.0 18,0 (0.348) (0,348) 18.1 18,1 0.804 0,804 T-1 T-1 Length Panjangnya 13 13 18.4 18,4

(0.604) 18.1 18,1 0.137 0,137 T-2 T-2 Weight Berat 8.5 8,5 32.0 32,0 (2.32) (2,32) 31.2 31,2 2.04 2,04 T-2 T-2 Length Panjangnya 8.5 8,5 29.5 29,5 (1.35) (1,35) 31.2 31,2 0.507 0,507 T-3 T-3 Weight, Berat, no worms 12 12 27.4 27,4 (0.811) 24.4 24,4 2.21 2,21 T-3 T-3 Length, Panjang, no worms 12 12 27.1 27,1 (0.757) (0,757) 24.4 24,4 0.606 0,606 T-3 T-3 Weight, Berat, with dengan worms 12 12 28.1 28,1 (0.429) (0,429) 29.6 29,6 1.42 1,42 T-3 T-3 Length, Panjang, with dengan worms

12 12 26.3 26,3 (1.63) 24.4 24,4 0.342 0,342 Page 24 Page 24 19 19 FIGURE 1 GAMBAR 1 .Relationship of specific growth rate (SGR) to temperature for weight and length in experiment T-1. Regression equation for weight is y = 0.0007x 33 - 0.0567x 22 ++ 1.1331x - 9.0468, R 22 =0.952. Regression equation for length is y = 0.0002x 33 - 0.0137x 22 ++ 0.321x - 2.2413, R 22 =0.815. Figure 2. Observations of behavior during experiment T-1. Reproductive-related activity and general swimming activity were scored as high (3), medium (2) or low (1). Number of reproductive males was added to reproductive activity score. Results shown are mean scores for each of the five treatment temperatures. Letters indicate significantly different growth rates, as determined by post-hoc Tukey tests. 00 11 22 33 44 55 66 77 12.7 12,7 18.1 18,1 23.4 23,4 27.8 27,8 32.9 32,9 Temperature (C) Suhu (C) MM ee

a sebuah nn a sebuah cc tiv TIV ity dasarkan ss cc oo rr ee Swimming Renang Spawning Hal ikan bertelur AA a sebuah AA a sebuah AB AB BB bb BB bb bb -0.8 -0,8 -0.6 -0,6 -0.4 -0,4 -0.2 -0,2 0.0 0,0 0.2 0,2 0.4 0,4 0.6 0,6 0.8 0,8 1.0 1,0 10 10 15 15 20 20 25 25 30 30 35 35 Temperature (C) Suhu (C) Weight Berat Length Panjangnya SGR (% growth per day) Page 25 Page 25 20 20

FIGURE 3. Relationship of specific growth rate (SGR) to temperature for weight and length in experiment T-2. Regression equation for weight is y = -0.0012x 33 + 0.0832x 22 1.7942x + 11.862, RR 22 =0.937. Regression equation for length is y = -0.0004x 33 + 0.0276x 22 0.5698x + 3.6862, R 22 =0.864. FIGURE 4. Relationship of specific growth rate (SGR) to temperature for weight and length in experiment T-3. Regression equation for weight, without tapeworms, is y = -0.0017x 33 + 0.1039x 22 - 1.8766 x + 10.521, R 22 =0.838; for weight, with tapeworms, y = -0.0025x 33 + 0.1677x 22 - 3.564x + 24.17, R 22 =0.883; for length, without tapeworms, y = -0.0004x 33 + 0.0251x 22 - 0.4356x + 2.3458, R 22 =0.869; and for length, with tapeworms, y = -0.000202x 33 + 0.0112x 22 0.188x + 0.9954, R 22 =0.657. -1 -1 -0.5 -0,5

00 0.5 0,5 11 1.5 1,5 22 2.5 2,5 33 10 10 15 15 20 20 25 25 30 30 35 35 40 40 Temperature (C) Suhu (C) Weight, no worms Weight, worms Length, no worms Length, worms -1 -1 -0.5 -0,5 00 0.5 0,5 11 1.5 1,5 22 2.5 2,5 10 10 15 15 20 20 25 25 30 30 35 35 Temperature (C) Suhu (C) Weight Berat Length Panjangnya Page 26 Page 26 21 21 was reduced by 42.2% at 20 C, 38.9% at 25 C, and 26.5% at 30 C; in length, growth rate was reduced by 56.0% at 20 C, 45.1% at 25 C, and 61.2% at 30 C. These values were calculated by comparing median values at each treatment temperature. Growth rate was not reduced at 15 C, and too few fish survived the 35 C treatment for a valid growth rate estimate to be calculated for percent growth rate reduction. Post-experiment dissections revealed no tapeworms in fish that had undergone Praziquantel treatment. Asian Asia tapeworms were present in fish that had not received Praziquantel, with 77%, 89%, 83%,

and 54% infection in the lowest four temperature treatments, in order of increasing temperature. suhu. For the 35 C treatment, no Asian tapeworms were found upon dissection at the conclusion of the experiment. Survival rate for the four lowest treatment temperatures of fish without tapeworms was 100%, and was 21% for the 35 C treatment. Survival rate for fish with tapeworms was 93%, 64%, 86%, 79%, and 21% in the five treatments in order of increasing temperature. Temperature had a significant effect on fish survival of fish both with ( P <0.0001) and without ( P= 0.0005) tapeworm infection. There was no significant difference in survival between fish with and without tapeworms when all treatments were considered ( P= 0.239). However, when the highest temperature (35 C) was excluded from the analysis, fish without tapeworms had a significantly higher survival rate ( P= 0.005). Growth rate was reduced in fish with tapeworm infection. Critical Thermal Maximum In experiment CTM-1, fish acclimated to 28 C had a mean CTM of 38.9 C ( n= 27) (Table 5). (Tabel 5). Fish acclimated to 31 C had a mean CTM of 39.6 C ( n= 23). The CTM values for these two acclimation groups were significantly different ( P <0.0001). Fish in Page 27 Page 27 22 22 experiment CTM-2 acclimated to 27 C had a mean CTM of 38.7 C ( n= 16) and fish acclimated to 31.5 C had a mean CTM of 39.8 C ( n= 14). These two groups were also significantly different ( P <0.0001). In experiment CTM-3, fish with tapeworms ( n= 19) as well as fish without tapeworms ( n= 6) both had mean CTM values of 39.5 C. No significant difference in CTM was found for fish with and without tapeworm infection ( P =0.467). Effect of Dissolved Oxygen on Growth Experiment O-1. Age was a significant factor for growth in length ( P <0.01) but there was no significant difference in growth in weight. For the 19-month-old shiners, dissolved oxygen had a significant effect on growth in weight ( P= 0.0003) and weight was significantly higher at 6 mg L -1 -1 and 5 mg L -1 -1 than at 2 mg L -1 -1 ( P <0.05) (Figure 5a). Growth in length for the 19-month-old shiners was not significantly affected by oxygen level. tingkat. For the 7-month-old shiners, variability was high and no significant differences were observed in growth rates (Figure 5b). Survival rates for the 19-month-old fish were 100%, 96%, 100%, 96%, and 96% for the five treatments in order of increasing dissolved oxygen concentration with no significant effect of dissolved oxygen on survival ( P= 0.73). Survival rates for the 7month-old fish were 96%, 88%, 92%, 92%, and 88% in order of increasing dissolved oxygen treatments with no significant effect of dissolved oxygen on survival ( P= 0.84). Post-experiment dissections revealed no tapeworms present in 19-month-old fish and 89% infection in 7-month-old fish (ie 16 of 18 randomly sampled 7-month-old fish over

the three highest dissolved oxygen treatments of were infected with tapeworms). Page 28 Page 28 23 23 TABLE 5 TABEL 5 .Critical thermal maxima from experiments CTM-1, CTM-2, and CTM-3. P values from one-tailed t-tests indicate comparisons between acclimation groups in CTM1 and CTM-2, and between groups with and without tapeworms in CTM-3. Experiment O-2. Dissolved oxygen concentration had a significant effect on growth in weight ( P= 0.0001) for 18-month-old fish (Fig. 6). Growth in weight was significantly lower for the 2.4 and 3.5 mg L -1 -1 dissolved oxygen treatments than the 4.3 mg L mg L -1 -1 dissolved oxygen treatment and higher ( P <0.05) (Figure 6). The effect of Pengaruh dissolved oxygen on growth in length was not significant. Survival rates were significantly different among treatments ( P= 0.01) with 93%, 98%, 95%, 78%, and 95% surviving in order of increasing dissolved oxygen concentration. Tapeworm infection was not determined by dissection, but was assumed to be 0% for the duration of the experiment because of the efficacy of the Praziquantel treatment observed in other experiments. percobaan. Experiment Percobaan nn Acclimation Aklimatisasi Temp. Temp. (C) (C) CTM CTM Mean Berarti CTM CTM SD SD CTM CTM Minimum Minimum CTM CTM Maximum Maksimum PP 23 23 31 31 39.6 0.273 38.5 38,5 39.9 39,9 CTM-1 27 27 28 28 38.9 0.203 38.5 38,5 39.3 39,3

} <0.0001 14 14 31.5 31,5 39.8 0.181 39.4 39,4 40.1 40,1 CTM-2 16 16 27 27 38.7 0.203 38.4 38,4 39.1 39,1 } <0.0001 CTM-3 (worms) (no worms) 19 19 66 30 30 30 30 39.5 39,5 39.5 39,5 0.340 0,340 0.082 0,082 38.3 38,3 39.4 39,4 39.9 39,9 39.6 39,6 } 0.467 Page 29 Page 29 24 24 Dissolved Oxygen (mg L -1 -1 )) A) 19-month-old Topeka shiners 00 0.2 0,2 0.4 0,4 0.6 0,6 0.8 0,8 11 1.2 1,2 1.4 1,4 1.6 1,6 2.0 2,0

2.9 2,9 3.9 3,9 4.8 4,8 6.1 6,1 Weight Berat Length Panjangnya B) 7-month-old Topeka shiners 00 0.2 0,2 0.4 0,4 0.6 0,6 0.8 0,8 11 1.2 1,2 1.4 1,4 1.6 1,6 1.8 1,8 22 2323 3131 4242 5151 6565 Weight Berat Length Panjangnya AA AA AA AA AA a sebuah a sebuah a sebuah a sebuah a sebuah Dissolved Oxygen (mg L -1 -1 )) FIGURE 5 GAMBAR 5 .Mean specific growth rates (SGR) (+/- SE) of 19-month old fish (A) and 7month old fish (B) in relation to dissolved oxygen concentration in experiment O-1. Letters indicate significantly different SGR between treatments as determined by posthoc Tukey tests. AA AB AB AB AB

BB BB a sebuah a sebuah a sebuah a sebuah a sebuah Page 30 Page 30 25 25 00 55 10 10 15 15 20 20 25 25 2.4 2,4 3.5 3,5 4.3 4,3 5.2 5,2 6.3 6,3 Chasers Chaser Non-Chasers AA AB AB BC SM ABC ABC CC a sebuah a sebuah a sebuah a sebuah a sebuah -0.2 -0,2 -0.1 -0,1 00 0.1 0,1 0.2 0,2 0.3 0,3 0.4 0,4 0.5 0,5 2.4 2,4 3.5 3,5 4.3 4,3 5.2 5,2 6.3 6,3

Weight Berat Length Panjangnya AA AA BB AB AB AB AB a sebuah a sebuah a sebuah a sebuah a sebuah FIGURE 6 GAMBAR 6 .Mean specific growth rates (SGR) (+/- SE) for weight and length in experiment O-2 for 18-month old fish. Letters indicate significantly different SGR between treatments as determined by post-hoc Tukey tests FIGURE 7 GAMBAR 7 . Summary of behavior observations in experiment O-2. Mean number of line crossings are shown (+/- SE) for four different observational periods for chaser and non-chaser fish. Letters indicate significant differences. Dissolved Oxygen (mg L -1 -1 )) Dissolved Oxygen (mg L -1 -1 )) Page 31 Page 31 26 26 In the analysis of behavior, there were significant effects of oxygen levels on line crossings for 'chaser' fish ( P= 0.001) (Fig. 6). Significant differences occurred in line crossings between 2 and 4 mg L -1 -1 , between 2 and 6 mg L -1 -1 , and between 3 and 6 mg L -1 -1 treatments (Figure 7). Dissolved oxygen level did not affect line crossings by 'nonchaser' fish ( P= 0.31). Lower Lethal Dissolved Oxygen Concentrations In experiment LC 50 50 -1, the 96-hour LC 50 50 value for the 19-month old fish acclimated to oxygen concentrations between 2.0 and 2.9 mg L

-1 -1 was 1.16 mg L -1 -1 , with a , Dengan 95% confidence interval of 0.62 - 2.16 mg L -1 -1 . . No 96-hour LC 50 50 value was calculated for the 7-month old fish because the binary method of calculation requires at least one treatment level with 100% survival and one with 100% mortality, and at no treatment was there 100% mortality. In experiment LC 50 50 -2, the 96-hour LC 50 50 value for fish acclimated to oxygen concentrations between 2.4 and 3.5 mg L -1 -1 was 1.26 mg L -1 -1 , with a 95% confidence interval of 0.80 - 1.98 mg L -1 -1 .. Discussion Diskusi High temperature and low dissolved oxygen alone are probably not factors in Topeka shiner population declines. Topeka shiners were able to survive and grow in conditions of both high temperature and low dissolved oxygen. Published historical information describing Topeka shiner habitat as being cool prairie streams does not seem to adequately describe their possible range of habitats. A combination of factors other Page 32 Page 32 27 27 than high temperature and low oxygen are probably responsible for local Topeka shiner extirpations. Although little information was previously available about temperature and oxygen tolerance of the Topeka shiner, information on other Notropis species suggests considerable tolerance of high temperatures. In Ohio, Mundahl (1990) reported observing Notropis chrysocephalus (striped shiners) and N. stramineus (sand shiners) surviving in habitats with water temperatures up to 39.5 C. However, shade may have provided lower temperatures in some areas of those habitats. CTM for Notropis species indicates values are as high as 39.65 C for N. lutrensis (red shiner) after fish were acclimated to 30 C (Rutledge & Beitinger 1989) (Table 6, adapted from Beitinger et al. 2000). 2000). Additionally, as summarized in Coutant (1977) a few studies have described preferred temperatures and optimum temperature for growth for species in the genus

Notropis . Cherry et al. Cherry et al. (1975) acclimated N. rubellus (rosyface shiner) and N. spilopterus (spotfin shiner) to a range of temperatures and reported a preferred temperature for each level of acclimation. When acclimated to 27 C, on average N. rubellus preferred 26.8 C and N. dan N. spilopterus preferred 28.1 C. Similarly, Barans and Tubb (1973) tested both young and adult N. atherinoides (emerald shiners) for preferred temperature after being acclimated to water between 20 and 25 C. Both age groups preferred temperatures between 21 and 23 C. Kellogg and Gift (1983) demonstrated that preferred temperature was related to optimum temperature for growth in the four species they tested. One of Salah satu these species was N. atherinoides . They report N. atherinoides ' preferred temperature as being 29 C after acclimation to 25 C, and they report an optimum temperature for growth Page 33 Page 33 28 28 for N. atherinoides of 27.3 C. Previously, McCormick and Kleiner (1976) studied optimum temperature for growth for N. atherinoides , and reported a value of 28.9 C. These past studies show that the genus Notropis has other members with high temperature tolerance, which supports our findings for N. topeka. Little information is available about low dissolved oxygen tolerance in Notropis spp. spp. Smale and Rabeni (1995) reported critical dissolved oxygen tolerances (dissolved oxygen concentration at which opercular movement and ventilation ceases, conducted at 26 C over a 4 to 6 hour period) for N. rubellus, N. nubilus, N. dorsalis, and N. stramineus (rosyface shiner, Ozark minnow, bigmouth shiner , and sand shiner) as 1.49, 1.45, 1.02, and 0.93 mg L -1 -1 , respectively. , Masing-masing. Ostrand and Wilde (2001) reported dissolved oxygen concentrations at which loss of equilibrium occurred over about 15 minutes for N. buccula and N. oxyrhynchus as 2.11 and 2.66 mg L -1 -1 , respectively. , Masing-masing. Results indicate that Hasil menunjukkan bahwa Topeka shiners have similar high-temperature, low-oxygen tolerances as other species of Notropis . Growth rates differ quite a bit between the three experiments in which I determined optimum temperature for growth. Although statistically it is not possible to draw a comparison between experiments, it is important to note the difference. One Satu possible explanation for this difference is that the three groups of fish had different thermal and photoperiod histories, which could affect their circadian or circannual rhythms. ritme. Seasons, because of differing photoperiod, affect fish growth, and that factor could have affected the results. Another factor to consider is fish age, which is discussed in more detail below in regards to experiment T-2. Page 34 Page 34 29 29 TABLE 6 TABEL 6 .Critical thermal maxima for Notropis spp. adapted from Beitinger et al. (2000). (2000). Entries include pretest acclimation temperature (C), rate of temperature increase

during CTM trial (T Cmin -1 -1 ), reported test endpoint (OS=onset of muscular spasms, LOE=loss of equilibrium), mean CTM and standard deviation of all CTMs tested, n= number of fish tested, and reference as given by Beitinger et al. (2000). (2000). Species Jenis Acclimation Aklimatisasi temperature suhu T T End Akhir point titik Mean Berarti CTM CTM SD SD NN Reference Referensi N. atherinoides 25 25 1.0 1,0 OS OS 37.6 37,6 0.40 0,40 10 10 Matthews & Maness (1979) 10 10 1.0 1,0 OS OS 34.1 34,1 0.65 0,65 88 Lutterschmidt & Hutchinson (1997) N. buccula 25 25 0.5 0,5 LOE 39.65 0.23 0,23 15 15 Ostrand & Wilde (2001) (2001) 30 30 0.5 0,5 LOE 36.5-

37.9 37,9 -15 15 Ostrand & Wilde (2001) (2001) N. N. chrysocephalus 24(field) 0.50.8 0,8 LOE 36.2 36,2 1.0 1,0 66 Mundahl (1990) 11(field) 1.0 1,0 LOE 30.8 30,8 1.8 1,8 88 Hockett & Mundahl (1988) (1988) N. cornutus 15 15 1.0 1,0 OS OS 31.9 31,9 0.48 0,48 88 Schubauer et al. (1980) (1980) 15(Dec) 1.0 1,0 OS OS 30.6 30,6 0.97 0,97 10 10 Kowalski et al. (1978) 15(Mar) 1.0 1,0 OS OS 32.0 32,0 0.54 0,54 16 16 Kowalski et al. (1978) N. cummingsae

88 1.0 1,0 LOE 29.0 29,0 1.3 1,3 -McFarlane et al. (1976) (1976) 88 0.1 0,1 LOE 28.0 28,0 1.1 1,1 -McFarlane et al. (1976) (1976) N. dorsalis 26 26 0.017 0,017 LOE 36.6 36,6 0.49 0,49 99 Smale & Rabeni (1995) N. girardi 25 25 1.0 1,0 OS OS 38.6 38,6 0.32 0,32 10 10 Matthews & Maness (1979) N. lutipinnis 13 13 1.0 1,0 LOE 30.0 30,0 1.2 1,2 -McFarlane et al. (1976) (1976) 13 13 0.1 0,1 LOE 29.0 29,0 0.7 0,7 -McFarlane et al. (1976) (1976)

N. lutrensis 30 30 0.3 0,3 LOE 39.65 0.23 0,23 10 10 Rutledge & Beitinger (1989) (1989) N. nubilus 26 26 0.017 0,017 LOE 36.2 36,2 0.62 0,62 99 Smale & Rabeni (1995) N. oxyrhynchus 25 25 0.5 0,5 LOE 36.537.9 37,9 -15 15 Ostrand & Wilde (2001) (2001) 30 30 0.5 0,5 LOE 39.2 39,2 0.2 0,2 15 15 Ostrand & Wilde (2001) (2001) N. rubellus 15 15 1.0 1,0 OS OS 31.8 31,8 0.51 0,51 55 Kowalski et al. (1978) 26 26 0.017 0,017 LOW RENDAH

35.3 35,3 0.23 0,23 77 Smale & Rabeni (1995) N. spilopterus 11(field) 1.0 1,0 LOE 31.8 31,8 1.8 1,8 26 26 Hockett & Mundahl (1988) (1988) N. stramineus 15(4 seasons) 1.0 1,0 OS OS 32.333.0 33,0 0.45- 0,450.68 0,68 10- 10 18 18 Kowalski et al. (1978) 26 26 0.017 0,017 LOE 37.0 37,0 0.27 0,27 66 Smale & Rabeni (1995) The estimate of 18 C in T-1 was probably not an accurate estimate for the Topeka shiner's optimum temperature (Figure 2). At higher temperatures and long daylengths, mature fish became sexually active, with considerable chasing of subordinate fish by dominant males. High energy expenditures resulting from this intense reproductive Page 35 Page 35 30 30 activity at or above 23 C probably reduced energy available for growth, resulting in slower growth of sexually active fish and a lower estimate for temperature of optimum growth. pertumbuhan. Subsequent experiments using a shorter daylength reduced the reproductive behavior considerably, although all chasing behavior was not eliminated. In both of the experiments with shorter daylength, the optimum temperature for growth was at least 9 C higher than with longer daylength (and more reproductive activity), which supports the conclusion that the optimum of 18 C was an underestimate.

It appears that an accurate estimate of optimum temperature for growth lies between 27 and 31 C, regardless of whether growth is measured as length or weight. However, the 31 C estimate may be too high because of the presence of tapeworms. It is Hal ini clear from the experiment comparing infected and non-infected fish that tapeworms reduce growth rate. However, lower parasite loads at higher temperatures suggest that tapeworms could not tolerate the highest temperature, as demonstrated in experiment T-2 where only 43% of the fish at 31 C had tapeworms, compared to between 89 and 100% infection at all lower temperatures. Similarly, tapeworm infection in experiment T-3 for fish that had not been treated with Praziquantel was 54% at 30 C and 0% at 35 C, compared with 77%, 89 %, and 83% infection at 15, 20, and 25 C, respectively. While I Sementara saya could not document exactly when fish shed tapeworms during the test, it is likely that the reduction in parasite load increased their subsequent growth rate. Thus, the high growth rate of infected fish at 31 C in experiment T-2 was relatively higher than fish at 27 C, which had a 94% tapeworm infection at the end of the experiment. This relatively higher growth rate at 31 C than at 27 C would cause the estimate of optimum temperature for growth to be high. Also, younger fish were used in experiment T-2 (in which all fish Page 36 Page 36 31 31 were initially infected with tapeworm). Age of fish is an important factor to consider for at least two reasons. First, younger fish do not exhibit reproductive behavior, although individuals still exhibit some chasing behavior; second, younger fish also could have different growth potential than adults, resulting in a higher optimum temperature for growth. pertumbuhan. However, there is no way to separate effect of age from effect of tapeworm infection in the experiment, as no experiment was performed on young fish without tapeworm infection. The best estimate, therefore, is that optimum temperature for growth is probably closer to 27 than 31 C for Topeka shiners. The estimate of 27 C is similar to other species in the genus Notropis . Overall, the presence of tapeworms reduced fish growth rate and survival. A A reduction in growth has been reported for several other fish species parasitized by tapeworm (Brouder 1999; Pulkkinen and Valtonen 1999; Sirois and Dodson 2000; Saksvik et al. 2001). 2001). Hoffman (1998) suggests that mortality due to tapeworms is not often a major concern, but this study indicated reduced growth and higher mortality and suggests that infected fish should not be used for population rehabilitation. Furthermore, Selanjutnya, because the Asian tapeworm is an exotic parasite, regulations prohibit the release of infected fish in waters where the parasite is not already present. CTM values were similar to the highest values reported for other Notropis species (Table 6). (Tabel 6). Although I did not test for the Topeka shiner's ultimate upper incipient lethal temperature (UULT; the maximum tolerable temperature regardless of acclimation temperature) it is probably between 33 and 35 C. This is suggested by the results, which showed that 97% of fish survived temperatures of 33 C, compared with 80% mortality at 35 C over 30 days. Additionally, a high CTM value supports the estimate of a high

Page 37 Page 37 32 32 optimum temperature for growth, because for many fish, the optimum temperature for growth is relatively close to the CTM value. Overall, survival at temperatures as high as 33 C with a CTM of 39 C is further evidence that Topeka shiners can tolerate high temperatures. suhu. Topeka shiners are also tolerant of low dissolved oxygen levels. Although a high Meskipun tinggi level of variability was present not only in the oxygen levels themselves (Table 2) but in growth of individual fish, results still showed that growth occurred at dissolved oxygen levels as low as 2 mg L -1 -1 . . Growth in length and weight was observed for both younger and older fish at 2 mg L -1 -1 . . Although the results for effect of oxygen on growth were not exactly the same between the two oxygen-growth experiments, the general downward trend of growth rate with reduced oxygen suggests that Topeka shiner growth is dependent on oxygen concentration. At oxygen levels below about 4 mg L -1 -1 , reduction in growth may be substantial. Results also show some evidence that activity level, including reproductive behavior, decreased as dissolved oxygen level decreased (Figure 7). 7). Inhibition of reproductive behavior could impact isolated populations. Results from the lower lethal dissolved oxygen tests support oxygen-growth test results, showing that Topeka shiners are quite tolerant of low oxygen conditions. However, three aspects of experimental conditions should be noted for the lower lethal dissolved oxygen tests. One aspect is that the first experiment was disrupted twice over the 96-hour period because the compressed nitrogen gas tank used for stripping oxygen from the water ran out, allowing oxygen levels in the experimental tanks to rise for a maximum of about 4 hours. Another aspect from the first test is that although tanks were equipped with plastic covers floating on the surface of the water, fish still had access to Page 38 Page 38 33 33 the surface via small gaps between the covers and the sides of the tank, and most fish in the treatments below 2 mg L -1 -1 congregated there during the experiment, taking advantage of the slightly more oxygenated surface layer of water. Finally, the second experiment was at a slightly higher temperature than the first (about 28 C vs. about 25 C). In spite of Meskipun these three differences, the estimates of 1.26 and 1.16 mg L -1 -1 for 96-hour LC

50 50 values nilai obtained from the two experiments are similar, and the estimate of 1.2 mg L -1 -1 for a 96hour LC 50 50 value is probably valid. Tolerance of high temperatures and low oxygen may result from adaptation to the demanding conditions of naturally occurring seasonal drought associated with prairie streams; however, other Notropis species show similar tolerances and some are not prairie headwater species, so the hypothesis about adaptation to prairie streams may be more complicated. lebih rumit. Topeka shiner tolerance of high-temperature and low-oxygen conditions supports the idea that off-channel habitats may be population sources, rather than population sinks. Certain microhabitats may allow populations to persist by supplying organisms to the areas in the habitat from which organisms are frequently lost (Pulliam 1988). The off- Offchannel habitats may provide refuges for Topeka shiners in times of drought. When Ketika water again becomes abundant the off-channel habitats reconnect, at least temporarily, to the main stream, allowing Topeka shiners to repopulate the stream. This study gives new information about physiological limits of the Topeka shiner that should be applied in the protection of known Topeka shiner habitat, including off-channel habitats, and could help to ensure success of future population restoration efforts. Page 39 Page 39 34 34 REFERENCES DAFTAR PUSTAKA APHA (American Public Health Association), American Water Works Association, and Water Environment Federation. 1998. 1998. Azide modification. Pages 4-131 and 4132 in Clesceri, LS, AE Greenberg, AD Eaton, and MAH Franson, editors. Standard Methods for the Examination of Water and Wastewater. Metode standar untuk Ujian Air dan Air Limbah. APHA, APHA, Washington. Washington. Barans, CA and RA Tubb. 1973. 1973. Temperatures selected seasonally by four fishes from Western Lake Erie. Journal of the Fisheries Research Board of Canada 30:1697-1703. Bayless, MA, M.. G. McManus, and JF Fairchild. 2003. 2003. Geomorphic, water quality and fish community patterns associated with the distribution of Notropis topeka in a central Missouri watershed. The American Midland Naturalist 150:58-72. Becker, CD and RG Genoway. 1979. 1979. Evaluation of critical thermal maximum for determining thermal tolerance of freshwater fish. Environmental Biology of Fishes 4:245-256. Beitinger, TL, WA Bennett, and RW McCauley. 2000. 2000. Temperature tolerances of North American freshwater fishes exposed to dynamic changes in temperature. Environmental Biology of Fishes 58:237-275. Berg, JA, TA Petersen, Y. Anderson, and R. Baker. 2004. 2004. Hydrogeology of the

Rock River watershed, Minnesota, and associated off-channel habitats of the Topeka shiner. Minnesota Department of Natural Resources, Divisions of Waters and Ecological Services. Brouder, MJ 1999. Relationship between length of Roundtail Chub and infection Page 40 Page 40 35 35 intensity of Asian fish tapeworm Bothriocephalus acheilognathi . Journal of Jurnal Aquatic Animal Health 11:302-304. Brungs, WA 1971. Chronic effects of low dissolved oxygen concentrations on the fathead minnow ( Pimephales promelas ). Journal Fisheries Research Board of Canada 28:1119-1123. Cherry, DS, KL Dickson, and J. Cairns, Jr. 1975. Temperatures selected and avoided by fish at various acclimation temperatures. Journal of the Fisheries Research Board of Canada 32:485-491. Cook, RD and S. Weisberg. 1999. 1999. Chapter 7: Introduction to Multiple Linear Regression in Applied Regression Including Computing and Graphics. Barnett, V., N. Cressie, N. Fisher, I. Jonstone, J. Kadane, D. Kendall, D. Scott, B. Silverman, A. Smith, J. Teugels, R. Bradley, and JS Hunter, editors. John John Wiley & Sons, Inc., New York, NY. Coutant, CC 1977. Coutant, 1977 CC. Compilation of temperature preference data. Journal of the Jurnal Fisheries Research Board of Canada 34:739-746. Dahle, SP 2001. Studies of the Topeka shiner ( Notropis topeka ) life history and distribution in Minnesota. MS Thesis, University of Minnesota, St. Paul. Hatch, JT 2001. What we know about Minnesota's first endangered fish species: the Topeka shiner. Journal of the Minnesota Academy of Science 65:39-46. Hatch, JT and S. Besaw. 2001. 2001. Food use in Minnesota populations of the Topeka shiner ( Notropis topeka ). Journal of Freshwater Ecology 16:229-233. Hoffman, GL 1998. Parasites of North American Freshwater Fishes, Second Edition. Cornell University Press, Ithaca, New York. Page 41 Page 41 36 36 Hrabik, RA 1996. A new distributional record of Notropis topeka (Teleostei: Cypriniformes) from the Mississippi River drainage in Missouri. Transactions of the Missouri Academy of Science 30:1-5. Kellogg, RL and JJ Gift. 1983. 1983. Relationship between optimum temperatures for growth and preferred temperatures for the young of four fish species. Transactions of the American Fisheries Society 112:424-430. Kerns, HA, and JL Bonneau. 2002. 2002. Aspects of the life history and feeding habits of the Topeka shiner ( Notropis topeka ) in Kansas. Transactions of the Kansas Academy of Science 105:125-142. Kuitunen, A. 2001. Microhabitat and instream flow needs of the Topeka shiner in the Rock River watershed, MN. Minnesota Department of Natural Resources, Division of Ecological Services, Stream Habitat Program, 1221 East Fir Avenue,

Fergus Falls, MN 56537. McCormick, JH and CF Kleiner. 1976. 1976. Growth and survival of young-of-the-year emerald shiners ( Notropis atherinoides ) at different temperatures. Journal of the Fisheries Research Board of Canada 33:839-842. Minckley, WL and FB Cross. 1959. 1959. Distribution, habitat, and abundance of the Topeka shiner Notropis topeka (Gilbert) in Kansas. The American Midland Naturalist 61:210-217. Mitchell, AJ 2004. Effectiveness of Praziquantel bath treatments against Bothriocephalus acheilognathi in grass carp. Journal of Aquatic Animal Health 16:130-136. Mundahl, ND 1990. Heat death of fish in shrinking stream pools. The American American Page 42 Page 42 37 37 Midland Naturalist 123:40-46. Newman, MC 1995. The LC LC 50 50 : Binomial Method. Page 134 in Quantitative Methods in Aquatic Ecotoxicology. CRC Press, Inc., Boca Raton, Florida. Ostrand, KG and GR Wilde. 2001. 2001. Temperature, dissolved oxygen, and salinity tolerances of five prairie stream fishes and their role in explaining fish assemblage patterns. pola. Transactions of the American Fisheries Society 130:742-749. Pulliam, HR 1988. Sources, sinks, and population regulation. The American Midland Naturalist 132:652-661. Pulkkinen, K. and ET Valtonen. 1999. 1999. Accumulation of plerocercoids of Triaenophorus crassus in the second intermediate host Coregonus lavaretus and their effect on growth of the host. Journal of Fish Biology 55:115-126. Rutledge, CJ and TL Beitinger. 1989. 1989. The effects of dissolved oxygen and aquatic surface respiration on the critical thermal maxima of three intermittent-stream fishes. Environmental Biology of Fishes 24:137-143. Saksvik, M., F. Nilsen, A. Nylund, and B. Berland. 2001. 2001. Effect of marine Eubothrium sp (Cestoda: Pseudophyllidea) on the growth of Atlantic salmon, Salmo salar L. Journal of Fish Diseases 24:111-119. Schrank, SJ, CS Guy, MR Whiles, and BL Brock. 2001. 2001. Influence of instream and landscape-level factors on the distribution of Topeka shiners and Notropis topeka in Kansas Streams. Copeia 2001: 413-421. Sirois, P. and JJ Dodson. 2000. 2000. Influence of turbidity, food density and parasites on the ingestion and growth of larval rainbow smelt Osmerus mordax in an estuarine turbidity maximum. Marine Ecology-Progress Series 193:167-179. Page 43 Page 43 38 38 Smale, MA and CF Rabeni. 1995. 1995. Hypoxia and hyperthermia tolerances of headwater stream fishes. Transactions of the American Fisheries Society Transaksi dari American Society Perikanan 124:698-710.

Stark, WJ, JS Luginbill, and ME Eberle. 2002. 2002. Natural history of a relict population of the Topeka shiner ( Notropis topeka) in Northwestern Kansas. Transactions of the Kansas Academy of Science 105:143-152. StatSoft, Inc. 1997. STATISTICA for Windows [Computer program manual]. StatSoft, Inc., 2300 East 14th Street, Tulsa, OK 74104. Tabor, VM 1998. Final rule to list the Topeka shiner as endangered. Federal Register Federal Daftar 63:69008-69021. Weisberg, S. 2005. Applied Linear Regression, Third edition. John Wiley & Sons, New John Wiley & Sons, New York, NY. York, NY