Differences in AChE and BChE Enzyme Activation Levels in Liver and Brain Tissues in Rainbow Trouts Exposed to Different Bacterial Diseases

Yıl 2021, , 356 - 364, 30.06.2021

Şükrü Önalan , Aslı Çilingir Yeltekin

https://doi.org/10.29133/yyutbd.853386

Cited By: 3

Öz

In this study, symptomatic fish samples were taken from rainbow trout farms. Isolation and identification of agents isolated from fish samples were made. DNA isolations from different purified colonies were carried out with the mericon bacterial DNA kit. Real-Time PCR procedure was performed by using universal bacterial primers. Molecular identifications were performed by blasting the nucleotides obtained by sequence analysis of PCR amplicons. Spectrophotometric measurements were performed at 412 nm wavelengths for AChE activity and 412 nm for BChE activity from liver and brain tissues of fish samples. The activity differences of different disease factors among themselves and according to the control group were examined. As a result of the study, isolation and identification of Bacillus subtilis, Lactococcus garvieae and Staphylococcus epidermidis from 5 different farms were performed. Over 98% similarity was observed as a result of sequencing analysis of the isolates. In this study, it was observed that three different bacteria isolated from trout farms suppressed AChE and BChE enzyme activities in both tissues of trout.

Anahtar Kelimeler

AChE, Bacteria, BChE, Rainbow trout, Real-Time PCR, Sequencing

Farklı Bakteriyel Hastalıklara Maruz Kalmış Gökkuşağı Alabalıklarında Karaciğer ve Beyin Dokularında AChE ve BChE Enzim Aktivite Seviyesi Farklılıkları

Öz

Bu çalışmada, gökkuşağı alabalığı işlemelerinden semptomlu balık örnekleri alınmıştır. Balık örneklerinden etkenlerinin izolasyonu ve identifikasyonları yapılmıştır. Saflaştırılan farklı kolonilerden DNA izolasyonları mericon bacterial DNA kiti ile gerçekleştirilmiştir. Real-Time PCR prosedürü evrensel bakteriyel primerler kullanılarak gerçekleştirilmiştir. Moleküler identifikasyonlar PCR amplikonlarının dizi analizi ile elde edilen nükleotidlerin blastlaması ile gerçekleştirilmiştir. Balık örneklerinin karaciğer ve beyin dokularından AChE aktivitesi için 412 nm ve BChE aktivitesi için 412 nm dalga boylarında spektrofotometrik ölçümler yapılmıştır. Farklı hastalık etkenlerinin kendi aralarında ve kontrol grubuna göre aktivite farklılıkları incelenmiştir. Çalışma sonucunda, 5 farklı işletmelerin 3’ünden Bacillus subtilis, Lactococcus garvieae ve Staphylococcus epidermidis etkenlerinin izolasyon ve identifikasyonları gerçekleştirilmiştir. İzolatların dizileme analizi sonucunda %98'in üzerinde benzerlik gözlenmiştir. Çalışmada, alabalık çiftliklerinden izole edilen üç farklı bakterinin her iki dokusunda da AChE ve BChE enzim aktivitelerini baskıladığı görülmüştür.

Anahtar Kelimeler

AChE,, Bakteri, BChE, Rainbow trout, Real-Time PCR

Kaynakça

• Al-Hebshi, N. N., Nasher, A. T., Idris, A. M., Chen, T. (2015). Robust species taxonomy assignment algorithm for 16S rRNA NGS reads: application to oral carcinoma samples. Journal of oral microbiology, 7(1), 28934.
• Austin, B., & Austin, D. A. (2016). Bacterial fish pathogens. 6. th edition. Springer International Publishing, Switzerland, 82-721.
• Baldissera, M. D., Souza, C. F., Doleski, P. H., Júnior, G. B., Vargas, A. C., & Baldisserotto, B. (2016). Involvement of cholinergic and purinergic systems during the inflammatory response caused by Aeromonas hydrophila in Rhamdia quelen. Microbial Pathogenesis, 99, 78-82.
• Baldisseraa, M. D., Souza, C. F., Santos, R. C. V., & Baldisserotto, B. (2018). Streptococcus agalactiae alters cerebral enzymes of phosphoryl transfer network in experimentally infected silver catfish: Impairment on brain energy homeostasis. Aquaculture, 489, 105–109.
• Chatonnet, A., & Lockridge, O. (1989). Comparision of butyrylcholinesterase and acetylcholin esterase. Biochem J, 260, 625-634.
• Chuiko, G. M. (2000). Comparative study of acetylcholinesterase and butyrylcholinesterase in brain and serum of several freshwater fish: specific activities in vitro inhibition by DDVP, an organophosphorus pesticide. Comp Biochem Physiology Part C, 127, 233-242.
• Das, U. N. (2007). Acetylcholinesterase and butyrylcholinesterase as possible markers of low-grade systemic inflammation. Med Sci Monit, 13, 214-221.
• Das, P., Pandey, P., Harishankar, A., Chandy, M., & Bhattacharya, S. (2016). A high yield DNA extraction method for medically important Candida species: A comparison of manual versus QIAcube based automated system. Indian journal of medical microbiology, 34(4), 533.
• Ekholm, M. (2001). Predicting relative binding free energies as substrate and inhibitors of acetyl- and butyrylcholinesterase. Theo chem, 572, 25-34.
• Ellman, G. L., Courtney, K. D., & Andres, V. J. (1961). A new and rapid colorimetric determination of acetylcholinesterase activity. Biochem Pharmacol, 7, 88-95.
• Inoue, D., Wada, K., Sei, K., Ike, M., & Fujita, M. (2005). Comparative evaluation of quantitative polymerase chain reaction methods for routine enumeration of specific bacterial DNA in aquatic samples. World Journal of Microbiology and Biotechnology, 21, 1029-1035.
• Jaguezeski, A. M., Perin, G., Rhoden, L. A., Da-Silva, T. M. A., Mendes, R. E., & Bottari, N. B. (2018). Changes on the activity of cholinesterase's in an immunomodulatory response of cattle infected by Listeria monocytogenes. Microbial Pathogenesis, 114, 36–40.
• Massouliè, J., Pezzementi, L., Bon, S., Krejci, E., & Valette, F. M. (1992). Molecular and cellular biology of cholinesterases. Neurobiology, 41, 31-91.
• Metin, S., Kubilay, A., Onuk, E. E., Didinen, B. I., & Yildirim, P. (2014). First isolation of Staphylococcus warneri from cultured rainbow trout (Oncorhynchus mykiss) broodstock in Turkey. Bulletin of the European Association of Fish Pathologists, 34(5), 165-174.
• Nadkarni, M. A., Martin, F. E., Jacques, N. A., & Hunter, N. (2002). Determination of bacterial load by Real-Time PCR using a broad-range (universal) probe and primers set. Microbiology, 148(1), 257-266.
• Pinchi, V., Focardi, M., Martinelli, D., Norelli, G. A., Carboni, I., Gozzini, A., & Ricci, U. (2013). DNA extraction method from teeth using QIAcube. Forensic Science International: Genetics Supplement Series, 4(1), 276-277.
• Ryhänen, R. J. J. (1983). Pseudocholinesterase activity in some human body fluids. Gen Pharmacol, 14, 459-460.
• Schwertz, C. I., Do-Carmo, G. M., Bottari, N. B., Da-Silva, E. S., Gabriel, M. E., & Da-Silva, A. S. (2016). Relationship between pathological findings and cholinesterase activity and nitric oxide levels in cattle infected naturally by Eurytrema coelomaticum. J Comp Pathology, 154, 150–156.
• Touraki, M., Karamanlidou, G., Karavida, P., & Chrysi, K. (2012). Evaluation of the probiotics Bacillus subtilis and Lactobacillus plantarum bioencapsulated in Artemia nauplii against vibriosis in European sea bass larvae (Dicentrarchus labrax, L.). World Journal of Microbiology and Biotechnology, 28(6), 2425-2433.
• Yeltekin, A. Ç. (2018). Comparison of toxic metal, trace element and macro element levels in trout cultivated in Latvia and Turkey. Fresenius Environmental Bulletin, 27(10), 7039-7044.
• Yeltekin, A. Ç., & Oğuz, A. R. (2018). Antioxidant responses and DNA damage in primary hepatocytes of Van fish (Alburnus tarichi, Güldenstadt 1814) exposed to nonylphenol or octylphenol. Drug and Chemical Toxicology. 41, 415.
• Zhu, L., Shen, D. X. Zhou, Q., Liu, C. J. Z., Li, X., Fang, Q., & Li, Z. (2014). Universal Probe Library based real-time PCR for rapid detection of bacterial pathogens from positive blood culture bottles. World Journal of Microbiology and Biotechnology, 30(3), 967-975.