Veterinary World

Open Access Open Access  Restricted Access Subscription Access

Molecular Identification and Histopathological Study of Natural Streptococcus agalactiae Infection in Hybrid Tilapia (Oreochromis niloticus)


Affiliations
1 School of Fisheries and Aquaculture Sciences, Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
2 Institute of Tropical Aquaculture (AQUATROP), Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
3 Department of Microbiology, Zhejiang University School of Medicine, 866 YuHuaTang Lu, Hangzhou, Zhejiang, China
 

Aim: The main objective of this study was to emphasize on histopathological examinations and molecular identification of Streptococcus agalactiae isolated from natural infections in hybrid tilapia (Oreochromis niloticus) in Temerloh Pahang, Malaysia, as well as to determine the susceptibility of the pathogen strains to various currently available antimicrobial agents.
Materials and Methods: The diseased fishes were observed for variable clinical signs including fin hemorrhages, alterations in behavior associated with erratic swimming, exophthalmia, and mortality. Tissue samples from the eyes, brain, kidney, liver, and spleen were taken for bacterial isolation. Identification of S. agalactiae was screened by biochemical methods and confirmed by VITEK 2 and 16S rRNA gene sequencing. The antibiogram profiling of the isolate was tested against 18 standard antibiotics included nitrofurantoin, flumequine, florfenicol, amoxylin, doxycycline, oleandomycin, tetracycline, ampicillin, lincomycin, colistin sulfate, oxolinic acid, novobiocin, spiramycin, erythromycin, fosfomycin, neomycin, gentamycin, and polymyxin B. The histopathological analysis of eyes, brain, liver, kidney, and spleen was observed for abnormalities related to S. agalactiae infection.
Results: The suspected colonies of S. agalactiae identified by biochemical methods was observed as Gram-positive chained cocci, β-hemolytic, and non-motile. The isolate was confirmed as S. agalactiae by VITEK 2 (99% similarity), reconfirmed by 16S rRNA gene sequencing (99% similarity) and deposited in GenBank with accession no. KT869025. The isolate was observed to be resistance to neomycin and gentamicin. The most consistent gross findings were marked hemorrhages, erosions of caudal fin, and exophthalmos. Microscopic examination confirmed the presence of marked congestion and infiltration of inflammatory cell in the eye, brain, kidney, liver, and spleen. Eye samples showed damage of the lens capsule, hyperemic an d hemorrhagic choroid tissue, and retina hyperplasia accompanied with edema. Brain samples showed perivascular and pericellular edema and hemorrhages of the meninges. Kidney samples showed hemorrhage and thrombosis in the glomeruli and tubules along with atrophy in hematopoietic tissue. Liver samples showed congestion of the sinusoids and blood vessel, thrombosis of portal blood vessel, and vacuolar (fatty) degeneration of hepatocytes. Spleen samples showed large thrombus in the splenic blood vessel, multifocal hemosiderin deposition, congestion of blood vessels, and multifocal infiltration of macrophages.
Conclusion: Therefore, it can be concluded that pathological changes in tissues and organs of fish occur proportionally to the pathogen invasion, and because of their high resistance, neomycin and gentamicin utilization in the prophylaxis or treatment of S. agalactiae infection should be avoided.

Keywords

16S rDNA, Antibiotic Resistance, Aquaculture, Histopathological Examination, Polymerase Chain Reaction, Streptococcus agalactiae.
User
Notifications
Font Size

  • Marcel, G., Sabri, M.Y., Siti-Zahrah, A. and Emikpe, O.B. (2013) Water condition and identification of potential pathogenic bacteria from red tilapia reared in cage-cultured system in two different water bodies in Malaysia. Afr. J. Microbiol. Res., 7(47): 5330-5337.

  • Ma, Y.P., Ke, H., Liang, Z.L., Liu, Z.X., Hao, L., Ma, J.Y. and Li, Y.G. (2016) Multiple evolutionary selections involved in synonymous codon usages in the Streptococcus agalactiae Genome. Int. J. Mol. Sci., 17(3): 1-12.

  • Pradeep, P.J., Suebsing, R., Sirthammajak, S., Kampeera, J., Jitrakorn, S., Saksmerprome, V., Turner, W., Palang, I., Vanichviriyakit, R., Senapin, S., Jeffs, A., Kiatpathomchai, W. and Withyachumanarnkul, B. (2016) Evidence of vertical transmission and tissue tropism of Streptococcosis from naturally infected red tilapia (Oreochromis spp.). Aquac. Rep., 3: 58-66.

  • Pereira, U.P., Mian, G.F., Oliveira, I.C.M., Benchetrit, L.C., Costa, G.M. and Figueiredo, H.C.P. (2010) Genotyping of Streptococcus agalactiae strains isolated from fish, human and cattle and their virulence potential in nile tilapia. Vet. Microbiol., 140: 186-192.

  • Austin, B. and Austin, D.A., editors. (1999) Bacterial Fish Pathogens Diseases of Farmed and Wild Fish. 4th ed. Praxis Publishing, Chichester, UK.

  • Siti-Zahrah, A., Padilah, B., Azila, A., Rimatulhana, R. and Shahidan, H. (2008) Multiple streptococcal species infection in cage-cultured red tilapia but showing similar clinical signs. In: Bondad-Reantaso, MG., Mohan, CV., Crumlish, M., Subasinghe, RP., editors. Diseases in Asian Aquaculture VI. Manila: Fish Health Section, Asian Fisheries Society; 2008. pp: 313-320.

  • Abuseliana, A., Daud, H., Aziz, S.A., Bejo, S.K. and Alsaid, M. (2010) Streptococcus agalactiae the etiological agent of mass mortality in farmed red tilapia (Oreochromis sp.). J. Anim. Vet. Adv., 9(20): 2640-2646.

  • Musa, N., Wei, L.S., Musa, N., Hamdan, R.H., Leong, L.K., Wee, W., Amal, M.N., Kutty, B.M. and Abdullah, S.Z. (2009) Streptococcosis in red hybrid tilapia (Oreochromis niloticus) commercial farms in Malaysia. Aquac. Res., 40(5): 630-632.

  • Pretto-Giordano, L.G., Muller, E.E., de Freitas, J.C. and da Silva, V.G. (2010) Evaluation on the pathogenesis of Streptococcus agalactiae in nile tilapia (Oreochromis niloticus). Braz. Arch. Biol. Technol., 53(1): 87-92.

  • Rodkhum, C., Kayansamruaj, P. and Pirarat, N. (2011) Effect of water temperature on susceptibility to Streptococcus agalactiae serotype Ia infection in nile tilapia (Oreochromis niloticus). Thai. J. Vet. Med., 41(3): 309-314.

  • Ye, X., Li, J., Lu, M., Deng, G., Jiang, X., Tian, Y., Quan, Y. and Jian, Q. (2011) Identification and molecular typing of Streptococcus agalactiae isolated from pond-cultured tilapia in China. Fish. Sci., 77(4): 623-632.

  • Zamri-Saad, M., Amal, M.N. and Siti-Zahrah, A. (2010) Pathological changes in red tilapias (Oreochromis spp.) naturally infected by Streptococcus agalactiae. J. Comp. Pathol., 2-3: 227-229.

  • Najiah, M., Aqilah, N.I., Lee, K.L., Khairulbariyyah, Z., Mithun, S., Jalal, K.C.A., Shaharom, H.F. and Nadirah, M. (2012) Massive mortality associated with Streptococcus agalactiae infection in cage-cultured red hybrid tilapia Oreochromis niloticus in Como River, Kenyir Lake, Malaysia. J. Biol. Sci., 12(8): 438-442.

  • Jolaine, M.W., Ralph, M.B. and Anthony, J.C. (2009) Evaluation of rapid cooling and tricaine methanesulfonate (MS222) as methods of euthanasia in zebrafish (Danio rerio). J. Am. Assoc. Lab. Anim. Sci., 48(6): 785-789.

  • Breed, S.R., Murray, E.G.D. and Smith, N.R. (1957) Bergey’s Manual of Determinative Bacteriology. 7th ed. The Williams & Wilkins Company, USA. p1-194.

  • Buller, N.B. (2004) Bacteria from Fish and Other Aquatic Animals: A Practical Identification Manual. CABI Publishing, Alabama, USA. p1-361.

  • Evans, J.J., Pasnik, D.J., Klesius, P.H. and Al-Ablani, S. (2006) First report of Streptococcus agalactiae and Lactococcus garvieae from a wild bottlenose folphin (Tursiops truncatus). J. Wildl. Dis. Assoc., 42(3): 561-569.

  • Yang, Y., Liu, Y., Ding, Y., Yi, L., Ma, Z., Fan, H. and Lu, C. (2013) Molecular characterization of Streptococcus agalactiae isolated from bovine mastitis in Eastern China. PLoS One, 8(7): 1-8.

  • Fay, A.B., Corrigan, J. and Murphy, R.A. (2016) Short-term effects of mechanical drainage on fungal and bacterial community structure in a managed grassland soil. Appl. Soil Ecol., 101: 93-100.

  • Bauer, A.W., Kirby, W.M., Sherris, J.C. and Turck, M. (1966) Antibiotic susceptibility testing by a standardized single disk method. Am. J. Clin. Pathol., 45(4): 493-496.

  • CLSI. (2012) Performance Standards for Antimicrobial Susceptibility Testing; Twenty-Second Informational Supplement. Vol. 32. Clinical and Laboratory Standards Institute, Wayne, PA. p1-184.

  • Al-Darwesh, A.A., Al-Shabbani, M.A.A. and Faris, B.H. (2014) Diagnostic and pathological study of Argulus japonicus in goldfish (Carassius auratus). Glob. J. BioSci. Biotechnol., 3(4): 384-387.

  • Abdullah, S., Omar, N., Yusoff, S.M., Obukwho, E.B., Nwunuji, T.P., Hanan, L. and Samad, J. (2013) Clinicopathological features and immunohistochemical detection of antigens in acute experimental Streptococcus agalactiae infection in red tilapia (Oreochromis spp.). Springerplus, 2(286): 1-7.

  • Chiang, Y.C., Pai, W.Y., Chen, C.Y. and Tsen, H.Y. (2008) Use of primers based on the heat shock protein genes hsp70, hsp40, and hsp10, for the detection of bovine mastitis pathogens Streptococcus agalactiae, Streptococcus uberis and Streptococcus bovis. Mol. Cell Probes., 22(4): 262-266.

  • Netto, L.N., Leal, C.A.G. and Figueiredo, H.C.P. (2011) Streptococcus dysgalactiae as an agent of septicaemia in nile tilapia, Oreochromis niloticus (L.). J. Fish Dis., 34(3): 251-254.

  • Lukkana, M., Jantrakajorn, S. and Wongtavatchai, J. (2015) Antimicrobial susceptibility and enrofloxacin resistance of Streptococcal bacteria from farmed nile tilapia, Oreochromis niloticus (Linnaeus 1758) in Thailand. Aquac. Res., 47(10): 3136-3144..

  • Suanyuk, N., Kanghear, H., Khongpradit, R. and Supamattaya, K. (2005) Streptococcus agalactiae infection in tilapia (Oreochromis niloticus). Songklanakarin J. Sci. Technol., 27: 307-319.

  • Suanyuk, N., Kong, F., Ko, D., Gilbert, G.L. and Supamattaya, K. (2008) Occurrence of rare genotypes of Streptococcus agalactiae in cultured red tilapia Oreochromis sp. and nile tilapia O. niloticus in Thailand-Relationship to human isolates? Aquaculture, 284(1): 35-40.

  • Yiagnisis, M. and Athanassopoulou, F. (2011) Bacteria isolated from diseased wild and farmed marine fish in Greece. Recent Adv. Fish Farms., 27(2): 61-69.

  • Mian, G.F., Godoy, D.T., Leal, C.A.G., Yuhara, T.Y., Costa, G.M. and Figueiredo, H.C.P. (2009) Aspects of the natural history and virulence of S. agalactiae infection in nile tilapia. Vet. Microbiol., 136: 180-183.

  • Russo, R., Mitchell, H. and Yanong, R.P.E. (2006) Characterization of Streptococcus iniae isolated from ornamental cyprinid fishes and development of challenge models. Aquaculture, 256(1): 105-110.

  • Filho, C.I., Muller, E.E., Pretto-Giordano, L.G. and Bracarense, F.R.L. (2009) Histological findings of experimental Streptococcus agalactiae infecion in nile tilapias (Oreochromis niloticus). Braz. J. Vet. Pathol., 2(1): 12-15.

  • Hernández, E., Figueroa, J. and Iregui, C. (2009) Streptococcosis on a red tilapia, Oreochromis sp., Farm: A case study. J. Fish Dis., 32(3): 247-252.

  • Noraini, O. and Jahwarhar, N.A. (2013) The effect of heat stress on clinicopathological changes and immunolocalization of antigens in experimental Streptococcus agalactiae infection in red hybrid tilapia (Oreochromis Spp.). Vet. World, 6: 997-1003.

  • Suwannasang, A., Dangwetngam, M., Issaro, A., Phromkunthong, W. and Suanyuk, N. (2014) Pathological manifestations and immune responses of serotypes Ia and III Streptococcus agalactiae infections in nile tilapia (Oreochromis niloticus). Songklanakarin J. Sci. Technol., 36(5): 499-506.

  • Bowater, R.O., Forbes-Faulkner, J., Anderson, I.G., Condon, K., Robinson, B., Kong, F., Gilbert, G.L., Reynolds, A., Hyland, S., McPherson, G., Brien, J.O. and Blyde, D. (2012) Natural outbreak of Streptococcus agalactiae (GBS) infection in wild giant Queensland grouper, Epinephelus lanceolatus (Bloch), and other wild fish in Northern Queensland, Australia. J. Fish Dis., 35(3): 173-186.

  • Chen, C.Y., Chao, C.B. and Bowser, P.R. (2007) Comparative histopathology of Streptococcus iniae and Streptococcus agalactiae-infected tilapia. Bull. Eur. Assoc. Fish Pathol., 27(2): 2-9.

  • Eldar, A., Bejerano, Y., Livoff, A., Horovitcz, A. and Bercovier, H. (1995) Experimental streptococcal meningoencephalitis in cultured fish. Vet. Microbiol., 43(1): 33-40.

  • Okwari, O.O., Ettarh, R.R., Akpogomeh, B.A. and Eteng, M.U. (2000) Gastric anti-secretory and anti-ulcerogenic effects of Dombeya buettneri in rats. J. Ethnopharmacol., 71: 315-319.

  • Koo, H.J., Kwak, H.S., Yoon, S.H. and Woo, G.J. (2012) Phylogenetic group distribution and prevalence of virulence genes in Escherichia coli isolates from food samples in South Korea. World J. Microbiol. Biotechnol., 28(4): 1813-1816.

  • Amyes, S.G.B. (2007) Enterococci and Streptococci. Int. J. Antimicrob. Agents, 29: S43-S52.

  • Geng, Y., Wang, K.Y., Huang, X.L., Chen, D.F., Li, C.W., Ren, S.Y., Liao, Y.T., Zhou, Z.Y., Liu, Q.F., Du, Z.J. and Lai, W.M. (2012) Streptococcus agalactiae, an emerging pathogen for cultured ya-fish, Schizothorax prenanti, in China. Transbound Emerg. Dis., 59(4): 369-375.

  • Wang, K., Chen, D., Huang, L., Lian, H., Wang, J., Xiao, D., Geng, Y., Yang, Z. and Lai, W. (2013) Isolation and characterization of Streptococcus agalactiae from nile tilapia Oreochromis niloticus in China. Afr. J. Microbiol. Res., 7(4): 317-323.


Abstract Views: 133

PDF Views: 0




  • Molecular Identification and Histopathological Study of Natural Streptococcus agalactiae Infection in Hybrid Tilapia (Oreochromis niloticus)

Abstract Views: 133  |  PDF Views: 0

Authors

A. A. Laith
School of Fisheries and Aquaculture Sciences, Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
Mohd Azmi Ambak
School of Fisheries and Aquaculture Sciences, Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
Marina Hassan
School of Fisheries and Aquaculture Sciences, Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
Shahreza Md. Sheriff
School of Fisheries and Aquaculture Sciences, Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
Musa Nadirah
School of Fisheries and Aquaculture Sciences, Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
Ahmad Shuhaimi Draman
Institute of Tropical Aquaculture (AQUATROP), Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
Wahidah Wahab
Institute of Tropical Aquaculture (AQUATROP), Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
Wan Nurhafizah Wan Ibrahim
School of Fisheries and Aquaculture Sciences, Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
Alia Syafiqah Aznan
School of Fisheries and Aquaculture Sciences, Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia
Amina Jabar
Department of Microbiology, Zhejiang University School of Medicine, 866 YuHuaTang Lu, Hangzhou, Zhejiang, China
Musa Najiah
School of Fisheries and Aquaculture Sciences, Universiti Malaysia Terengganu, Kuala-21030, Terengganu, Malaysia

Abstract


Aim: The main objective of this study was to emphasize on histopathological examinations and molecular identification of Streptococcus agalactiae isolated from natural infections in hybrid tilapia (Oreochromis niloticus) in Temerloh Pahang, Malaysia, as well as to determine the susceptibility of the pathogen strains to various currently available antimicrobial agents.
Materials and Methods: The diseased fishes were observed for variable clinical signs including fin hemorrhages, alterations in behavior associated with erratic swimming, exophthalmia, and mortality. Tissue samples from the eyes, brain, kidney, liver, and spleen were taken for bacterial isolation. Identification of S. agalactiae was screened by biochemical methods and confirmed by VITEK 2 and 16S rRNA gene sequencing. The antibiogram profiling of the isolate was tested against 18 standard antibiotics included nitrofurantoin, flumequine, florfenicol, amoxylin, doxycycline, oleandomycin, tetracycline, ampicillin, lincomycin, colistin sulfate, oxolinic acid, novobiocin, spiramycin, erythromycin, fosfomycin, neomycin, gentamycin, and polymyxin B. The histopathological analysis of eyes, brain, liver, kidney, and spleen was observed for abnormalities related to S. agalactiae infection.
Results: The suspected colonies of S. agalactiae identified by biochemical methods was observed as Gram-positive chained cocci, β-hemolytic, and non-motile. The isolate was confirmed as S. agalactiae by VITEK 2 (99% similarity), reconfirmed by 16S rRNA gene sequencing (99% similarity) and deposited in GenBank with accession no. KT869025. The isolate was observed to be resistance to neomycin and gentamicin. The most consistent gross findings were marked hemorrhages, erosions of caudal fin, and exophthalmos. Microscopic examination confirmed the presence of marked congestion and infiltration of inflammatory cell in the eye, brain, kidney, liver, and spleen. Eye samples showed damage of the lens capsule, hyperemic an d hemorrhagic choroid tissue, and retina hyperplasia accompanied with edema. Brain samples showed perivascular and pericellular edema and hemorrhages of the meninges. Kidney samples showed hemorrhage and thrombosis in the glomeruli and tubules along with atrophy in hematopoietic tissue. Liver samples showed congestion of the sinusoids and blood vessel, thrombosis of portal blood vessel, and vacuolar (fatty) degeneration of hepatocytes. Spleen samples showed large thrombus in the splenic blood vessel, multifocal hemosiderin deposition, congestion of blood vessels, and multifocal infiltration of macrophages.
Conclusion: Therefore, it can be concluded that pathological changes in tissues and organs of fish occur proportionally to the pathogen invasion, and because of their high resistance, neomycin and gentamicin utilization in the prophylaxis or treatment of S. agalactiae infection should be avoided.

Keywords


16S rDNA, Antibiotic Resistance, Aquaculture, Histopathological Examination, Polymerase Chain Reaction, Streptococcus agalactiae.

References