INDUKSI KEKEBALAN SPESIFIK PADA IKAN MAS, Cyprinus carpio Linn. TERHADAP INFEKSI KOI HERPESVIRUS (KHV) MELALUI TEKNIK KOHABITASI TERKONTROL

Penelitian dengan tujuan untuk mengetahui masa induksi kekebalan spesifik hingga level protektif terhadap KHV pada populasi ikan mas yang diinfeksi secara buatan melalui teknik pemaparan terkontrol telah dilakukan pada skala laboratorium. Pemaparan terkontrol terhadap ikan positif KHV dilakukan selama 3 hari pada suhu 24oC-26oC, selanjutnya ikan dipindahkan ke wadah volume 300 liter yang diisi ikan uji sebanyak 200 ekor/wadah dan suhu air berkisar antara 31oC-34oC. Perlakuan yang diterapkan adalah periode induksi: (A) periode induksi selama 1 minggu dari akhir masa pemaparan terhadap KHV, (B) periode induksi selama 2 minggu dari akhir masa pemaparan terhadap KHV, (C) periode induksi selama 3 minggu dari akhir masa pemaparan terhadap KHV, (D) periode induksi selama 4 minggu dari akhir masa pemaparan terhadap KHV, dan (E) tanpa periode induksi setelah dilakukan pemaparan terhadap KHV. Pada hari ke-21, seluruh kelompok perlakuan diinfeksi KHV secara buatan dengan teknik kohabitasi. Pengamatan dilakukan terhadap tingkah laku, gejala klinis, dan mortalitas ikan uji yang dilakukan setiap hari hingga akhir percobaan. Rataan persentase sintasan ikan uji tertinggi pada akhir riset diperoleh pada periode induksi selama 3 minggu sebesar 53,75%; diikuti oleh periode induksi selama 2 minggu sebesar 33,75%; selama 1 minggu sebesar 18,75%; selama 4 minggu sebesar 12,5%; dan kelompok kontrol sebesar 7,5%.The research with the aim to study an effective period to develop specific immunity against Koi Herpesvirus on common carp population which was obtained by cohabitation technique had been done in laboratory scale. Cohabitation of KHV infected fish at 24oC-26oC was for 3 days, and then the fish was moved to fiber glass tank at 31oC-34oC to develop specific immunity. The treatments applied in the research were: (A) a week period induction, (B) two weeks period induction, (C) three weeks period induction, (D) four weeks period induction, and (E) without period induction as a control group. Fish test were challenge to KHV infection at the end of each defined period induction by cohabitation method for 2 weeks lasting. Examination on behavior, clinical signs, and mortality of fish test were taken place daily. The results showed that the highest survival rate was found on three weeks period induction (53.75%), and followed by two weeks period induction (33.75%), one week period induction (18.75%), four weeks period induction (12.5%), and the lowest was found on the control (7.5%).

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The Effectiveness of Onion Extract Allium sativum to Prevent Koi Herpesvirus (KHV) Infection on Common Carp Cyprinus carpio

Jurnal Akuakultur Indonesia ◽

10.19027/jai.7.139-150 ◽

2008 ◽

Vol 7 (2) ◽

pp. 139

Author(s):

Sri Nuryati ◽

P. Giri ◽

Y. Hadiroseyani

Keyword(s):

Immune System ◽

Survival Rate ◽

Common Carp ◽

Cyprinus Carpio ◽

Allium Sativum ◽

Garlic Extract ◽

Herpesvirus Infection ◽

Onion Extract ◽

Koi Herpesvirus ◽

Herpès Virus

Common carp is one of consumption fish that has delicious meat, high pritein level, and easy in farming. The serious problem in common carp farming is koi herpesvirus infection. Onion extract potency to improve immune system was estimated to prevent disease infection. The testing of the garlic extract through food could be used as efforts to increase endurance of common carp fish Cyprinus carpio to koi herpesvirus infection that was considered from blood parameter. Fish that was used was measuring 9-11 cm with the treatment of food containing 30, 50, and 70 gr/100 ml onion extract. Fish was acclimated for seven days in 60×30×30 cm3 aquarium before used. Garlic extract diet in food gave increasing of fish immune system that was infected by koi herpesvirus. The increased of leucocytes of blood fish with onion extract diet was faster than possitive control. The dose of B treatment (50 gr/100 ml) was the best dose gave short incubation periode comparing other treatment. Survival rate (SR) of this B treatment was highest, i.e. 91.7%, while survival rate of negative control was 50%. Key word: common carp, Cyprinus carpio, onion, Allium sativum, koi herpesvirus ABSTRAK Salah satu jenis ikan konsumsi air tawar yang banyak digemari oleh masyarakat adalah ikan mas Cyprinus carpio karena rasa dagingnya gurih, memiliki kadar protein tinggi dan cukup mudah dalam pemeliharaannya. Permasalahan yang muncul saat ini adalah wabah Koi Herpes Virus (KHV). Potensi ekstrak bawang putih sebagai anti mikroba spektrum luas, diduga dapat mengobati dan mencegah penyakit ikan. Pengujian bawang putih secara in vivo melalui pakan dapat digunakan sebagai upaya untuk meningkatkan ketahanan tubuh ikan mas Cyprinus carpio terhadap infeksi penyakit KHV yang ditinjau dari gambaran darahnya. Ikan uji yang digunakan adalah ikan mas berukuran 9-11 cm dengan perlakuan pakan yang mengandung bawang putih sebanyak 30, 50, dan 70 gr/100 ml. Sebelum dilakukan penelitian ikan diadaptasikan selama 7 hari pada akuarium berukuran 60×30×30 cm3. Hasil penelitian menunjukan bahwa ekstrak bawang putih efektif untuk meningkatkan ketahanan tubuh ikan mas Cyprinus carpio yang diinfeksi oleh Koi Herpes Virus (KHV). Pengamatan gambaran darah ikan yang terinfeksi KHV setelah pemberian ekstrak bawang putih selama 30 hari dapat meningkatkan jumlah leukosit lebih cepat dibandingkan dengan perlakuan kontrol positif. Peningkatan jumlah limfosit dalam darah mampu meningkatkan pertahanan tubuh. Peningkatan jumlah leukosit ditunjukkan dengan meningkatnya jumlah limfosit dan penurunan jumlah monosit. Perlakuan B (50 gr/100 ml) merupakan dosis yang paling baik karena masa inkubasi KHV terlewati lebih cepat dibanding perlakuan lain yang ditandai dengan penurunan jumlah leukositnya. Hal ini didukung oleh gejala klinis yang ringan dibandingkan kontrol positif dan tingkat kelangsungan hidup yang tinggi sebesar 91,7%. Sedangkan pada kontrol negatif kelangsungan hidupnya sebesar 50%. Kata kunci: ikan mas, Cyprinus carpio, bawang putih, Allium sativum, KHV dan Herpes

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Description of an as Yet Unclassified DNA Virus from Diseased Cyprinus carpio Species

Journal of Virology ◽

10.1128/jvi.79.4.1983-1991.2005 ◽

2005 ◽

Vol 79 (4) ◽

pp. 1983-1991 ◽

Cited By ~ 46

Author(s):

Marina Hutoran ◽

Ariel Ronen ◽

Ayana Perelberg ◽

Maya Ilouze ◽

Arnon Dishon ◽

...

Keyword(s):

Common Carp ◽

Cyprinus Carpio ◽

Secondary Infection ◽

Clinical Signs ◽

Viral Disease ◽

Infected Fish ◽

Etiologic Agent ◽

Amino Acid Sequences ◽

Virus Species ◽

Enveloped Virus

ABSTRACT Numerous deaths of koi and common carp (Cyprinus carpio) were observed on many farms throughout Israel, resulting in severe financial losses. The lethal viral disease observed is highly contagious and extremely virulent, but morbidity and mortality are restricted to koi and common carp populations. Diseased fish exhibit fatigue and gasping movements in shallow water. Infected fish had interstitial nephritis and gill necrosis as well as petechial hemorrhages in the liver and other symptoms that were not consistent with viral disease, suggesting a secondary infection. Here we report the isolation of carp nephritis and gill necrosis virus (CNGV), which is the etiologic agent of this disease. The virus propagates and induces severe cytopathic effects by 5 days postinfection in fresh koi or carp fin cell cultures (KFC and CFC, respectively), but not in epithelioma papillosum cyprini cells. The virus harvested from KFC cultures induced the same clinical signs, with a mortality of 75 to 95%, upon inoculation into naive koi and common carp. Using PCR, we provide final proof that the isolated virus is indeed the etiologic agent of food and ornamental carp mortalities in fish husbandry. Electron microscopy revealed viral cores with icosahedral morphology of 100 to 110 nm that resembled herpesviruses. Electron micrographs of purified pelleted CNGV sections, together with viral sensitivities to ether and Triton X-100, suggested that it is an enveloped virus. However, the genome of the isolated virus is a double-stranded DNA (dsDNA) molecule of 270 to 290 kbp, which is larger than known herpesviruses. The viral DNA seems highly divergent and bears only small fragments (16 to 45 bp) that are similar to the genomes of several DNA viruses. Nevertheless, amino acid sequences encoded by CNGV DNA fragments bear similarities primarily to members of the Poxviridae and Herpesviridae and to other large dsDNA viruses. We suggest, therefore, that the etiologic agent of this disease may represent an as yet unclassified virus species that is endemic in C. carpio (carp).

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Isolation, molecular identification, and pathological lesions of Saprolegnia spp. isolated from common carp, Cyprinus carpio in floating cages in Mosul, Iraq

Veterinary World ◽

10.14202/vetworld.2020.2759-2764 ◽

2020 ◽

Vol 13 (12) ◽

pp. 2759-2764

Author(s):

Fawwaz Fadhil Ali ◽

Shahbaa Khalil AL-Taee ◽

Zahraa Mustafa AL-Jumaa

Keyword(s):

Common Carp ◽

Cyprinus Carpio ◽

Clinical Signs ◽

Inflammatory Cells ◽

Infected Fish ◽

Universal Primers ◽

Tissue Samples ◽

Common Carp Cyprinus Carpio ◽

Carp Cyprinus Carpio ◽

Gill Filaments

Background and Aim: Saprolegniasis is a fungal disease that infects freshwater fish. The condition is characterized by a cotton-like appearance in the gills and body. This study aimed to isolate Saprolegnia from common carp, Cyprinus carpio, raised in a floating cage in Wana district, Mosul, Iraq. Materials and Methods: Samples were collected from 15 infected fish and examined microbiologically, molecularly, and histopathologically. Saprolegnia DNA was extracted which was amplified using universal primers give a 540 bp DNA fragment, and gill and muscle tissue were also examined for histopathological changes. Results: Isolated colonies of Saprolegnia were characterized by a circular, white cottony appearance with long hair. Lactophenol staining demonstrated hyphae as branched non-septate, transparent masses. The genomic DNA of isolates was consistent with Saprolegnia spp. The infected tissue samples showed variable pathology in gills. Severe hemorrhage and edema were observed in primary gill filaments with hyperplasia in epithelial cells and infusion in secondary gill filaments. Hyphae of Saprolegnia were seen between necrotic and edematous myofiber with inflammatory cells infiltration. Conclusion: Saprolegnia can cause economic impacts through lethal infection of fish. Clinical signs of Saprolegnia infection were confirmed molecularly and microscopically, and these findings were supported by histopathological lesions in gill and muscle tissues.

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Hematology of common carp following DNA vaccination and koi herpesvirus challenge test

Jurnal Akuakultur Indonesia ◽

10.19027/jai.9.9-15 ◽

2010 ◽

Vol 9 (1) ◽

pp. 9 ◽

Cited By ~ 2

Author(s):

Sri Nuryati ◽

N.A. Maswan ◽

. Alimuddin ◽

. Sukenda ◽

K. Sumantadinata ◽

...

Keyword(s):

Survival Rate ◽

Dna Vaccine ◽

Common Carp ◽

Cyprinus Carpio ◽

Lethal Dose ◽

Challenge Test ◽

Phagocytic Index ◽

Viral Glycoprotein ◽

Post Vaccination ◽

Koi Herpesvirus

The study was aimed to determine the effectiveness of DNA vaccine doses on hematological aspect which represent immune response and its influence on common carp survival rate. DNA vaccines encoding the viral glycoprotein of koi herpesvirus (KHV) have been proved to highly protect the fish under laboratory condition. A dose of 12.5 µg/100 µl vaccine had resulted in a survival rate of 96.67 % during 30 days after challenge test with a lethal dose of KHV. Fish vaccinated using lower doses, i.e. 2.5 and 7.5 µg/100µl showed 100% mortality after 15 and 19 days challenge test respectively, whereas non vaccinated fish as a control showed 100% mortality after 17 days challenge test. Total leucocytes of the vaccinated fish were higher than control until 42 days post vaccination, but declined afterward. Phagocytic index of the vaccinated fish using 12.5 µg/100 µl was declined after 49 days post vaccination or 7 days post challenge test. Key words: DNA vaccine, Koi herpesvirus (KHV), leucocyte, phagocytic index, Cyprinus carpio ABSTRAK Penelitian ini bertujuan untuk menguji pengaruh vaksinasi menggunakan vaksin DNA dengan dosis berbeda terhadap gambaran darah ikan sebagai respresentasi tanggap kebal ikan mas serta pengaruhnya terhadap tingkat kelangsungan hidup ikan mas. Vaksin DNA penyandi glikoprotein koi herpesvirus (KHV) dapat memberikan proteksi yang tinggi pada percobaan skala laboratorium. Vaksinasi dengan dosis 12,5 µg/100µl dapat mempertahankan kelangsungan hidup sebesar 96,67% selama satu bulan setelah uji tantang dengan virus KHV menggunakan dosis letal. Ikan yang divaksin dengan dosis yang lebih rendah yaitu 2,5 dan 7,5 µg/100µl mengalami kematian total berturut-turut setelah 15 dan 19 hari uji tantang, sedangkan ikan kontrol yang tidak divaksin mengalami kematian total setelah 17 hari uji tantang. Jumlah leukosit total ikan yang divaksinasi lebih tinggi dibanding dengan kontrol sampai hari ke-42, setelah itu mengalami penurunan. Indeks fagositosis ikan yang divaksin dengan dosis 12,5 µg/100µl mengalami penurunan setelah hari ke-49 atau 7 hari setelah uji tantang. Kata kunci: Vaksin DNA, Koi herpesvirus (KHV), leukosit, indeks fagositosis, Cyprinus carpio

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Enrichment of common carp (Cyprinus carpio) diet with Malic acid: Effects on skin mucosal immunity, antioxidant defecne and growth performance

Annals of Animal Science ◽

10.2478/aoas-2020-0092 ◽

2020 ◽

Vol 0 (0) ◽

Author(s):

Roghieh Safari ◽

Seyed Hossein Hoseinifar ◽

Maryam Dadar ◽

Hien Van Doan

Keyword(s):

Common Carp ◽

Cyprinus Carpio ◽

Malic Acid ◽

Control Group ◽

Skin Mucus ◽

Antioxidant Genes ◽

Treatment Comparison ◽

Genes Expression ◽

Significant Difference ◽

And Control

AbstractThe present study investigated possible effects of dietary malic acid on the expression of immunity, antioxidant and growth related genes expression as well as skin mucus immune parameters in common carp. Common carp (Cyprinus carpio) fingerlings were fed diets supplemented with different levels (0 [control], 0.5%, 1%, 2%) of malic acid (MA) for 60 days. The results revealed highest expression levels of immune-related genes (tnf-alpha, il1b, il8 and lyz) in skin of common carp fed 2% MA (P < 0.05). Regarding 1% MA treatment comparison with control group, significant difference was noticed just in case of lyz (P < 0.05). Evaluation of growth related genes expression revealed no significant difference between treatments (P > 0.05). The study of antioxidant related genes (gsta and gpx) in common carp skin fed with MA, showed significant difference between treated groups and control (P < 0.05). Carps fed with 2% MA had highest alkaline phosphatase activity in skin mucus compared other treated groups and control (P < 0.05). There were no significant difference between 0.5% and 1% and control (P > 0.05). The study of total protein and total immunoglobulin (Ig) in common carp skin musus revealed no alteration following MA treatment (P > 0.05). The present data demonstrated that feeding with MA altered immune and antioxidant genes expression in skin mucus of common carp.

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Effects of a Trans-Galactooligosaccharide on Minerals Content of Common Carp (Cyprinus carpio L.) Tissues

Biological Trace Element Research ◽

10.1007/s12011-021-02600-w ◽

2021 ◽

Author(s):

Ewa Ziółkowska ◽

Joanna Bogucka ◽

Jan Mazurkiewicz ◽

Mateusz Rawski ◽

Szymon Różański ◽

...

Keyword(s):

Common Carp ◽

Cyprinus Carpio ◽

Control Diet ◽

Colorimetric Method ◽

Bifidobacterium Bifidum ◽

Feed Additives ◽

Control Treatment ◽

Control Group ◽

Whole Cells ◽

Freeze Dried

AbstractCommon carp (Cyprinus carpio L.) is a dominant fish species in aquaculture, and as it is a stomachless species, absorption and digestion of nutrients take place in the intestine. The aim of the study was to evaluate the effects of a prebiotic on the content of selected minerals found in the meat, gills, and skeleton of common carp. The research applied trans-galactooligosaccharide (GOS) prebiotic produced by enzymatic transgalactosylation of milk lactose by whole cells of Bifidobacterium bifidum. The following diets have been applied: control diet without feed additives (C), diet 2 (B1) with 1% of GOS, and diet 3 (B2) with 2% of GOS. In the freeze-dried samples, concentrations of the analyzed metals were determined using atomic absorption spectroscopy (AAS). The content of phosphorus was determined using colorimetric method. The analyses confirmed that the highest level of Mg was detected in the skeleton of fish fed with 1% GOS (2.51 g kg−1) and was significantly higher compared the control treatment (2.11 g kg−1) (P < 0.05). Zn content in fish meat fed with 1% GOS (35.41 mg kg−1) was significantly higher (P < 0.05) than in the control group (24.59 mg kg−1). The tissue that accumulated the greatest amount of Zn was the gills. GOS had a positive effect on Fe accumulation in the meat, gills, and skeleton. It has been concluded that supplementation of feed with 2% GOS significantly influenced the positive correlations between Mg and P in the meat and skeleton, Fe–Ca correlation in gills, and Fe–Zn correlation in the skeleton.

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Potency of Garlic Extract Against Koi Herpesvirus (KHV) in Common Carp

Jurnal Akuakultur Indonesia ◽

10.19027/jai.6.147-154 ◽

2007 ◽

Vol 6 (2) ◽

pp. 147

Author(s):

Sri Nuryati ◽

D. Puspitaningtyas ◽

D. Wahjuningrum

Keyword(s):

Survival Rate ◽

Common Carp ◽

Clinical Symptoms ◽

Hemoglobin Concentration ◽

Blood Parameters ◽

Garlic Extract ◽

Virus Suspension ◽

Cell Numbers ◽

Koi Herpesvirus ◽

Leukocyte Number

Prevention of koi herpesvirus (KHV) infection using chemicals or medicines was ineffective way. In this study garlic extract was used to prevent KHV infection. Virus suspension of 0.1 ml and garlic extract of 0.1 ml in different dosage, i.e., 100, 200 and 300 ppt, was injected into common carp body. Hemoglobin concentration, red and white blood cell numbers, and leukocyte number were counted. The results of study showed that administration of 300 ppt of garlic extract could produce higher survival rate (67.5%), good blood parameters and clinical symptoms compared to other treatments. Keywords: garlic, KHV, common carp ABSTRAK Upaya penanggulangan wabah Koi Herpesvirus (KHV) menggunakan bahan-bahan kimia atau obat-obatan adalah tidak efektif. Pada penelitian ini dilakukan pemberian ekstrak bawang putih untuk menanggulangi infeksi KHV. Suspensi virus sebanyak 0,1 ml ditambahkan dengan 0,1 ml ekstrak bawang putih dengan berbagai konsentrasi, yaitu 100, 200 dan 300 ppt, disuntikkan ke dalam tubuh ikan mas. Kadar hemoglobin, jumlah sel darah merah dan sel darah putih jenis dan jumlah leukosit diamati. Hasil penelitian menunjukkan bahwa pemberian ekstrak bawang putih sebanyak 300 ppt menghasilkan kelangsungan hidup (67,5%) yang lebih baik dibandingkan dengan perlakuan lainnya, dan begitu pula dengan gambaran darah serta gejala klinisnya. Kata kunci: bawang putih, KHV, ikan mas

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Effect of β-glucan on behavioral, biochemical and hematological parameters against toxicity of copper sulfate in common carp Cyprinus carpio L.

The Iraqi Journal of Veterinary Medicine ◽

10.30539/iraqijvm.v38i2.234 ◽

2014 ◽

Vol 38 (2) ◽

pp. 128-137

Author(s):

Jamal K. Al-faragi,

Keyword(s):

Common Carp ◽

Copper Sulfate ◽

Cyprinus Carpio ◽

Hematological Parameters ◽

Control Group ◽

Enzymes Activity ◽

Common Carp Cyprinus Carpio ◽

Carp Cyprinus Carpio ◽

Wbc Count ◽

Hb Concentration

The aim of the present study was to assess the effect of a commercial ß-glucan against acute toxic effect of copper sulfate on common carp Cyprinus carpio L. behavioral, hematological parameters and biochemical tests. A total of 250 Fish (150 ± 2 g) were fed a basal control diet or the basal diet plus a ß-glucan (1 g kg-1 diet) for 60 days. LC50 was calculated for 72 h which was 10.83 mg L-1. At the end of the trial, CuSO4.5H2O was added to all treatments at a dose of 5 mgL-1 for T1 and T2, 7mg L-1 for T3 and T4, 9 mgL-1 for T5 and T6 for 96h. Fish behavioral changes were recorded during 96h of exposure to different Cu concentrations with and without β-glucan, which showed abnormalities such as increase swimming activity, jerky movement, loss of equilibrium, respiratory stress, increase operculum movement, frequent jumping, erratic swimming and swimming near the water surface. On the other hand, ß-glucan supplemented groups showed significant increase (P≤0.05) in Hb concentration, PCV%, RBC and WBC count in comparison with groups without ß-glucan. The results of dietary groups without ß-glucan showed a significant decrease (P≤ 0.05) in Hb concentration, hematocrit value, RBC count, but there were significant increase (P≤ 0.05) in WBC count with an increase of Cu concentration compared to control groups. Results showed significant increase (P≤ 0.05) in dietary groups without ß-glucan in values of ALT, AST and ALP enzymes activity with elevation of Cu levels compared to control group. But, ß-glucan supplemented groups showed significant decrease (P ≤ 0.05) in values of ALT, AST and ALP enzymes activity in comparison with dietary groups without ß-glucan. These results suggested that β-glucan has protective effect against CuSO4 toxicity and future studies should be pursued particularly immune response and molecular studies

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Efficacy Testing of Orally Administered Praziquantel to Common Carp Naturally Infected by Caryophyllidean Tapeworms (Platyhelminthes: Eucestoda)

Acta Veterinaria Brno ◽

10.2754/avb201079s9s073 ◽

2010 ◽

Vol 79 (9) ◽

pp. S73-S78 ◽

Cited By ~ 3

Author(s):

Eliška Sudová ◽

Veronika Piačková ◽

Josef Velíšek ◽

Martin Pijáček ◽

Zdeňka Svobodová

Keyword(s):

Body Weight ◽

Oral Administration ◽

Common Carp ◽

Cyprinus Carpio ◽

Treatment Efficacy ◽

Infected Fish ◽

Individual Species ◽

Clean Water ◽

Single Oral Administration ◽

Heat Treated

The aim of the study was to assess the effects of orally applied praziquantel on the treatment efficacy against most abundant tapeworms (Atractolytocestus huronensis Anthony, 1958 and Khawia sinensis Hsü, 1935) of common carp (Cyprinus carpio Linnaeus). Naturally infected fish (1.5-year-old; K1-2) were divided into 6 groups. Three trial groups were administered praziquantel mixed in heat-treated amyloid vehicle (at doses of 50 mg kg-1 body weight). In 3 groups of control, the fish were administered amyloid vehicle only. Fish were examined using gut dissection 2, 4 and 6 days after the administration. The location, intensity and prevalence of individual species of tapeworms were recorded. A. huronensis and K. sinensis were found in the gut of carp together. A. huronensis was situated in the oesophageal part of foregut, and K. sinensis came after – immediately before the first intestine loop, of the intestines of carp. The invasion of A. huronensis was fully eliminated in all trial groups after 4 days whereas some specimens of K. sinensis still persisted in the gut of few fish. The full elimination of both parasites was observed 6 days after administration. Our results showed that elimination of K. sinensis takes longer than elimination of A. huronensis. The dose of 50 mg kg-1 of orally administered praziquantel is effectual to control A. huronensis and K. sinensis infections in common carp successfully, however, a significant decrease of the number of parasites was also caused by a single oral administration of big bolus vehicle and leaving unfed fish in the clean water.

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