Large scale mortality in cultured Nile tilapia (Oreochromis niloticus): natural co-infection with Aeromonas hydrophila and Streptococcus iniae

Document Type : Full paper (Original article)

Authors

1 Ph.D. Student in Fisheries Microbiology, Department of Aquatic Animal Health Management, College of Fisheries, Karnataka Veterinary, Animal and Fisheries Sciences University, Mangalore-575 002, India

2 Department of Aquatic Animal Health Management, College of Fisheries, Karnataka Veterinary, Animal and Fisheries Sciences University, Mangalore-575 002, India

Abstract

Background: Nile tilapia is a highly valuable fish in the aquaculture sector. A culture farm has reported heavy mortalities of tilapia. Aims: The present study aimed to identify the etiological agent responsible for the heavy mortality in cage cultured tilapia. Methods: The moribund and freshly dead fishes were analyzed for clinical signs. Biochemical and molecular characterizations were performed to identify the etiological agents of the disease. Also, polymerase chain reaction (PCR) assay was used to detect the presence of the virulence genes. The susceptibility of the isolates to various antibiotics was tested by the disk diffusion method. Results: The results of the biochemical tests and PCR assay confirmed that co-infection with Aeromonas hydrophila, and Streptococcus iniae was responsible for the disease severity. Phylogenetic analysis of the 16S rRNA gene showed that A. hydrophila and S. iniae isolates shared 99% and 98% sequence homology with A. hydrophila and S. iniae previously deposited in the Genbank database. The multiple antibiotic resistance (MAR) index of A. hydrophila was 0.16 and that of S. iniae was 0.71. The PCR test revealed that both pathogens harbored numerous virulence factors. The experimental infection study confirmed that the synergistic action of A. hydrophila and S. iniae led to increased mortality in tilapia. Histopathological changes were observed in the liver and spleen tissues of the co-infected fishes. Conclusion: These findings indicate that the disease outbreak in the tilapia culture farm occurred as a result of co-infection by A. hydrophila and S. iniae.

Keywords

References

Abdel-Latif, H; Dawood, M; Menanteau-Ledouble, S and El-Matbouli, M (2020). The nature and consequences of co-infections in tilapia: A review. J. Fish Dis., 43: 651-664. https://doi.org/10.1111/jfd.13164.
Abdel-Latif, H and Khafaga, AF (2020). Natural co-infection of cultured Nile tilapia Oreochromis niloticus with Aeromonas hydrophila and Gyrodactylus cichlidarum experiencing high mortality during summer. Aquac. Res., 51: 1880-1892.
Aboyadak, IM; Ali, NGM; Goda, AMAS; Aboelgalagel, WH and Salam, AME (2015). Molecular detection of Aeromonas hydrophila as the main cause of outbreak in tilapia farms in Egypt. J. Aquac. Mar. Biol., 2: 2-5. doi: 10.15406/jamb.2015.02.00045.
Aisyhah, MAS; Amal, MNA; Zamri-Saad, M; Siti-Zahrah, A and Shaqinah, NN (2015). Streptococcus agalactiae isolates from cultured fishes in Malaysia manifesting low resistance pattern towards selected antibiotics. J. Fish Dis., 38: 1093-1098.
Asao, T; Kinoshita, Y; Kozaki, S; Uemura, T and Sakaguchi, G (1984). Purification and some properties of Aeromonas hydrophila hemolysin. Infect. Immun., 46: 122-127.
Assis, G; Tavares, G; Pereira, F; Figueiredo, H and Leal, C (2017). Natural coinfection by Streptococcus agalactiae and Francisella noatunensis subsp. orientalis in farmed Nile tilapia (Oreochromis niloticus L.). J. Fish Dis., 40: 51-63.
Ausuhel, FM; Brent, R; Kington, RE; Moore. DD; Seidman. JG; Smith, JA and Struhl, K (1990). Current Protocols in Molecular Biology. 1: Greene Publishing Associates, Inc. and John Wiley and Sons, Inc., NY.
Baiano, JC and Barnes, AC (2009). Towards control of Streptococcus iniae. Emerg. Infect Dis., 15: 1891-1896. doi: 10.3201/eid1512.090232. PMID: 19961667; PMCID: PMC3044519.
Barrow, GI and Feltham, RKA (2004). Cowan and Steel’s manual for the identification of medical bacteria. 3rd Edn., Cambridge University Press. P: 329.
Basri, L; Roslindawani, MN; Salleh, A; Yasin, ISM; Saad, MZ; Rahaman, NYA; Barkham, T and Amal, MNA (2020). Co-infections of tilapia lake virus, Aeromonas hydrophila and Streptococcus agalactiae in farmed red hybrid tilapia. Animals. 10: 2141. doi: 10.3390/ ani10112141.
Baums, CG; Hermeyer, K; Leimbach, S; Adamek, M; Czerny, CP; Schwark, GH; Weigand, VP; Baumg€artner, W and Steinhagen, D (2013). Establishment of a model of Streptococcus iniae Meningoencephalitis in Nile Tilapia (Oreochromis niloticus). J. Comp. Pathol., 149: 94-102.
Bigarre, L; Cabon, J; Baud, M; Heimann, M; Body, A; Lieffrig, F and Castric, J (2009). Outbreak of betanodavirus infection in tilapia, Oreochromis niloticus (L.), in fresh water. J. Fish Dis., 32: 667-673.
Clinical and Laboratory Standards Institute (CLSI) (2006). Performance standards for antimicrobial disk susceptibility tests. Approved Standard, 9th Edn., IN Clinical and Laboratory Standards Institute document M2-A9 (ISBN 1-56238-586-0). Clinical and Laboratory Standards Institute, 940 West Valley Road, Suite 1400, Wayne, Pennsylvania, 19087-1898, USA.
Clinical and Laboratory Standards Institute (CLSI) (2016). Methods for antimicrobial dilution and disk susceptibility testing of infrequently isolated or fastidious bacteria. 3rd Edn., CLSI guideline M45. Wayne, PA: Clinical and Laboratory Standards Institute.
Clinical and Laboratory Standards Institute (CLSI) (2017). Performance standards for antimicrobial susceptibility testing. 27th Edn., CLSI supplement M100. Wayne, PA: Clinical and Laboratory Standards Institute.
Cutuli, MT; Gibello, A; Rodriguez-Bertos, A; Blanco, MM; Villarroel, M; Giraldo, A and Guarro, J (2015). Skin and subcutaneous mycoses in tilapia (Oreochromis niloticus) caused by Fusarium oxysporum in coinfection with Aeromonas hydrophila. Med. Mycol. Case Rep., 9: 7-11.
Deng, ML; Yu, ZH; Geng, Y; Wang, KY; Chen, DF; Huang, XL; Ou, Y; Chen, ZL; Zhong, ZJ and Lai, WM (2015). Outbreaks of Streptococcosis associated with Streptococcus iniae in Siberian sturgeon (Acipenser baerii) in China. Aquac. Res., 48: 1-11. doi: 10.1111/are.12934.
Dong, HT; Nguyen, VV; Le, HD; Sangsuriya, P; Jitrakorn, S; Saksmerprome, V; Senapin, S and Rodkhum, C (2015). Naturally concurrent infections of bacterial and viral pathogens in disease outbreaks in cultured Nile tilapia (Oreochromis niloticus) farms. Aquaculture. 448: 427-435. doi: 10.1016/j.aquaculture.2015.06.027.
Dong, HT; Techatanakitarnan, C; Jindakittikul, P; Thaiprayoon, A; Taengphu, S; Charoensapsri, W; Khunrae, P; Rattanarojpong, T and Senapin, S (2017). Aeromonas jandaei and Aeromonas veronii caused disease and mortality in Nile tilapia. Oreochromis niloticus (L.). J. Fish Dis., doi: 10.1111/jfd.12617.
El-Sayed, AFM (2019). Tilapia culture. 2nd Edn., Cambridge, MA: Academic Press. PP: 1-275.
Eyngor, M; Zamostiano, R; Tsofack, JEK; Berkowitz, A; Bercovier, H; Tinman, S and Eldar, A (2014). Identification of a novel RNA virus lethal to tilapia. J. Clin. Microbiol., 52: 4137-4146.
FAO (2017). Statistics and Information Branch, Fisheries and Aquaculture Department/FAO. Fishery and Aquaculture Statistics. Global Production by Production Source 1950-2015 (FishstatJ). FAO Fisheries and Aquaculture Department (online), Rome.
FAO (2020). The state of world fisheries and aquaculture, 2020. Sustainability in Action. Food and Agriculture Organization of the United Nations, Rome. https://doi.org/10.4060/ca9229en.
Girisha, SK; Puneeth, TG; Nithin, MS; Kumar, BN; Ajay, SK; Vinay, TN; Suresh, T; Venugopal, MN and Ramesh, KS (2020). Red sea bream iridovirus disease (RSIVD) outbreak in Asian seabass (Lates calcarifer) cultured in open estuarine cages along the west coast of India: First report. Aquaculture. 520: 734712. https://doi.org/10.1016/j.aquaculture.2019.734712.
Grim, CJ; Kozlova, EV; Sha, J; Fitts, EC; van Lier, CJ; Kirtley, ML and Shak, JR (2013). Characterization of Aeromonas hydrophila wound pathotypes by comparative genomic and functional analyses of virulence genes. mBio., 4: e00064-e13. https://doi.org/10.1128/mBio.00064-13e00064-00013.
Julia, WP and Phillip, HK (2011). Molecular identification and virulence of three Aeromonas hydrophila isolates cultured from infected channel catfish during a disease outbreak in west Alabama (USA) in 2009. Dis. Aquat. Org., 94: 249-253.
Krumperman, PH (1983). Multiple antibiotic resistance indexing of Escherichia coli to identify high-risk sources of fecal contamination of foods. Appl. Environ. Microbiol., 46: 165-170.
Kumar, S; Stecher, G; Li, M; Knyaz, C and Tamura, K (2018). MEGA X: Molecular evolutionary genetics analysis across computing platforms. Mol. Biol. Evol., 35: 1547-1549.
Lane, DJ (1991). 16S/23S rRNA sequencing. In: Stackebrandt, E and Goodfellow, M (Eds.), Nucleic acid techniques in bacterial systematic. New York, John Wiley and Sons. PP: 115-175.
Ma, J; Bruce, TJ; Oliver, L and Cain, KD (2019). Co-infection of rainbow trout (Oncorhynchus mykiss) with infectious haematopoietic necrosis virus and Flavobacterium psychrophilum. J. Fish Dis., 42: 1065-1076. https://doIorg/10.1111/jfd.13012.
Menaga, M and Fitzsimmons, K (2017). Growth of tilapia industry in India. J. World Aquac. Soc., 48: 49-52.
Monir, MS; Yuso, SM; Mohamad, A and Ina-Salwany, MY (2020). Vaccination of tilapia against Motile Aeromonas Septicemia: A review. J. Aquat. Anim. Health. 32: 65-76.
Nguyen, HT; Kanai, K and Yoshikoshi, K (2001). Experimental Streptococcus iniae infection in Japanese flounder Paralichthys olivaceus. Fish Pathol., 36: 40-41.
Nithin, MS; Girisha, SK; Kushala, KB; Chandan, DV; Puneeth, TG; Naveen Kumar, BT; Vinay, TN; Suresh, T; Sahoo, L and Ramesha, KS (2021). Novel lytic bacteriophages (AhFM4 & AhFM5) as bio-control measures against multidrug resistant biofilm producing Aeromonas hydrophila (AhZ1K). Aquaculture. 544: 737106. https://doi.org/10.1016/j.aquaculture.2021.737106.
Niu, G; Khattiya, R; Zhang, T; Boonyayatra, S and Wongsathein, D (2020). Phenotypic and genotypic characterization of Streptococcus spp. isolated from tilapia (Oreochromis spp.) cultured in river-based cage and earthen ponds in Northern Thailand. J. Fish Dis., 43: 391-398. https://doi.org/10.1111/jfd.13137.
Ortega, C; Garcıa, I; Irgang, R; Fajardo, R; Tapia-Cammas, D; Acosta, J and Avendãno-Herrera, R (2018). First identification and characterization of Streptococcus iniae obtained from tilapia (Oreochromis aureus) farmed in Mexico. J. Fish Dis., 41: 773-782. https://doi.org/10.1111/jfd.12775.
Pakingking, RJ; Takano, R; Nishizawa, T; Mori, KI; Iida, Y; Arimoto, M and Muroga, K (2003). Experimental coinfection with aquabirnavirus and viral hemorrhagic septicemia virus (VHSV), Edwardsiella tarda or Streptococcus iniae in Japanese flounder Paralichthys olivaceus. Fish Pathol., 38: 15-21.
Palang, I; Hirono, I; Senapin, S; Sirimanapong, W; Withyachumnarnkul, B and Vanichviriyakit, R (2020). Cytotoxicity of Streptococcus agalactiae secretory protein on tilapia cultured cells. J. Fish Dis., 43: 1229-1236. https://doi.org/10.1111/jfd.13230.
Pauzi, NA; Mohamad, N; Azzam-Sayuti, M; Ina-Salwany, MD; Zamri-Saad, M; Nasruddin, NS and Amal, MNA (2020). Antibiotic susceptibility and pathogenicity of Aeromonas hydrophila isolated from red hybrid tilapia (Oreochromis niloticus - Oreochromis mossambicus) in Malaysia. Vet. World. 13: 2166-2171.
Perretta, A; Antúnez, K and Zunino, P (2018). Phenotypic, molecular and pathological characterization of motile aeromonads isolated from diseased fishes cultured in Uruguay. J. Fish Dis., 41: 1559-1569.
Pulkkinen, K; Suomalainen, LR; Read, AF; Ebert, D; Rintamäki, P and Valtonen, ET (2010). Intensive fish farming and the evolution of pathogen virulence: the case of columnaris disease in Finland. Proc. Biol. Sci., 277: 593-600. https://doi.org/10.1098/rspb.2009.1659.
Rahmatullah, M; Ariff, M; Kahieshesfandiari, M; Daud, HM; Zamri-Saad, M; Sabri, MY and Ina-Salwany, MY (2017). Isolation and pathogenicity of Streptococcus iniae in cultured red hybrid Tilapia in Malaysia. J. Aquat. Anim. Health. 29: 208-213. https://doi.org/10.1080/08997659. 2017.1360411.
Raj, NS; Swaminathan, TR; Dharmaratnam, A; Raja, SA; Ramraj, D and Lal, KK (2019). Aeromonas veronii caused bilateral exophthalmia and mass mortality in cultured Nile tilapia, Oreochromis niloticus (L.) in India. Aquaculture. 512: 734278. doi: 10.1016/j.aquaculture.2019. 734278.
Ray, SD and Homechaudhuri, S (2014). Morphological and functional characterization of hepatic cells in Indian major carp, Cirrhinus mrigala against Aeromonas hydrophila infection. J. Environ. Biol., 35: 253-258.
Salah, MA; Wael, GN; Mounir, M and Salem-Bekhi, MM (2012). Bacteriological and histopathological studies on Enterobacteriacea in Nile tilapia Oreochromis Niloticus. J. Pharm. Biomed., 2: 94-104.
Sen, K and Rodgers, M (2004). Distribution of six virulence factors in Aeromonas species isolated from US drinking water utilities: a PCR identification. J. Appl. Microbiol., 97: 1077-1086. https://doi.org/10.1111/j.1365-2672.2004. 02398.x.
Shoemaker, CA; Evans, JJ and Klesius, PH (2000). Density and dose: Factors affecting mortality of Streptococcus iniae infected tilapia (Oreochromis niloticus). Aquaculture. 188: 229-235.
Smith, P; Hiney, MP and Samuelesen, OB (1994). Bacterial
resistance to antimicrobial agent used in fish farming: a critical evaluation of method and meaning. Annu. Rev. Fish Dis., 4: 273-313.
Stehr, F; Kretschmar, M; Kroger, C; Hube, B and Schafer, W (2003). Microbial lipases as virulence factors. J. Mol. Catal. BEnzym., 22: 347-355. doi: 10.1016/S1381-1177(03)00049-3.
Swaminathan, TR; Ravi, C; Kumar, R; Dharmaratnam, A; Basheer, VS; Pradhan, PK and Sood, N (2018). Derivation of two tilapia (Oreochromis niloticus) cell lines for efficient propagation of Tilapia Lake Virus (TiLV). Aquaculture. 492: 206-214.
Swift, S; Lynch, JM; Fish, L; Kirke, DF; Tomas, JM; Stewart, GS and Williams, P (1999). Quarum sensing-dependent regulation and blockade of exoprotease production in Aeromonas hydrophila. Infect. Immun., 67: 5192-5199.
Wang, GH; Clark, CG; Liu, CY; Pucknell, C; Munro, CK and Kruk, TM (2003). Detection and characterization of the hemolysin genes in Aeromonas hydrophila and Aeromonas sobria by multiplex PCR. J. Clin. Microbiol., 41: 1048-1054. doi: 10.1128/JCM.41.3.1048-1054.2003.
Zhou, SM; Fan, Y; Zhu, XQ; Xie, MQ and Li, AX (2011). Rapid identification of Streptococcus iniae by specific PCR assay utilizing genetic markers in ITS rDNA. J. Fish Dis., 34: 265-271. https://doi.org/10.1111/j.1365-2761.2010. 01233.x.

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