Evaluation of the Antibacterial Effect of Three Carps Species

Published: 2021-06-17 06:31:52
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In present study, all selected carps (H. nobilis, C. idella and C. carpio) secreted huge amount of mucus. Also mucus secretion varied among selected fish species. Blackstock and Pickering (1982); Negus (1983) and Nigam, Kumari, Mittal and Mittal (2012) also reported the similar results. This variation in mucus secretion might be due to different ecological and physiological conditions and also different mucus producing cells situated in epidermis and epidermal layers of different fishes. Jung et al. (2012) also revealed the variation in mucus secretion and its component during the summer and winter, which also supports our finding as mucus secretion by fishes was higher in summers than winters. Present study also revealed more dense and increased amount of mucus in all fish species, after challenge with A. hydrophila. Increased amount of mucus might be the indicator of increased stress level and activation of innate immune system against bacterial attack. Many previous reports by number of authors such as, Subramanian et al.(2008) and Nigam et al.(2012) demonstrated that environmental perturbations such as, pH, temperature, dissolved oxygen etc.; different ecological niches, bacterial stress, different developmental ages influence the amount of mucus secretion in fishes. Our results are in substantial agreement with above studies. Jones (2001) also reported that Gyrodactylus infected fish produced high amount of mucus than normal which also supports our finding that increased bacterial stress enhanced mucus secretion in challenged group. Madetuja, Dalsgaard and Wiklund (2002); Subramanian et al. (2008) and Holm et al. (2015) also observed similar results.
It is now well established that fish skin mucus provides mechanical barrier to fishes by lying at interface between them and surrounding pathogens. In addition to trapping and sloughing of infectious pathogens, it is also the reservoir of bioactive components which acts in different way and gifted innate antibacterial properties to mucus. Present results on antibacterial activity shown by selected exotic carps also confirmed that fish mucus is a source of antimicrobial products. Earlier studies by many authors viz. Austin and Mcintosh (1988); Oren and Shai (1996); Cole, Weis and Diamond (1997); Ellis (2001); Nagashima et al. (2003); Sarmasik (2002); Balasubramanian, Baby Rani, Arul and Prakash (2012); Prakash, Loganathan, Arul, Senthilraja and Gunaesekaran (2013); Nurtamin, Nurman and Hafizah (2016) and many others also demonstrated strong antibacterial activity in several fishes. Ebran et al. (1999) and Ebran, Julien, Orange and Molle (2000) demonstrated that only the hydrophobic components of crude epidermal mucus of fresh water and sea water fish exhibited strong pore-forming properties, which were well correlated with antibacterial activity. Manivasagan, Annamali, Ashokkumar and Sampathkumar (2009) also reported the antibacterial activity in skin mucus soluble and insoluble friction of A. maculatus and A. thallasinus against E. coli and P. aeruginosa. However, ZOIs values against same microbes shown by mucus extracts in our study were higher. crude mucus extract of C. straitus showed ZOI of 8 mm against A. hydrophila and no antibacterial effect against E. coli and K. pneumonia studied by Wei et al. (2010).Bragadeeswaran and Thangraj (2011) revealed the presence of strong antimicrobial effect of crude mucus extract of eel fish against E. coli, P. aeruginosa and S. aureus and no activity was seen against K. pneumonia. Furthermore, they did not notice bactericidal effect in aqueous mucus extract against P. aeruginosa. However, in the present study, crude mucus, as well as aqueous mucus (HFM and CFM) extracts of all selected fishes, exhibited antibacterial activity against all microbial strains. The test species of bacteria were similar to the studies of Wei et al. (2010) and Bragadeeswaran and Thangraj (2011).
Variation in antibacterial activity might be due to different fish species, different habitats, different stress levels etc. Also different fishes having different susceptibility against different or same pathogen. Expression of different immune components in different fish species against different pathogen could be one of the reasons responsible for variation in bactericidal effect. Subramanian, Mackinnon and Ross (2007) also reported antimicrobial activity of aqueous mucus extract of several fishes. But in their further studies, they didn’t find any bactericidal effect in aqueous mucus extract of wider range of fish species such as S. alpines, S. fontinalis, C. carpio, M. saxatalis, M. aeglefinus and M. glutinosa. Low level of enzymatic activities due to incubation temperature or pH conditions used in the antimicrobial assay might be one of the reasons behind the absence of antibacterial activities. But in present study, aqueous mucus extracts of all selected fishes exhibited bactericidal effect against all pathogenic bacteria taken under study. Anbuchezhian, Gobinath and Ravichandaran (2011) also noticed the similar results which also support our results. Aqueous mucus extracts R. rita and C. punctatus exhibited ZOIs of 9.75 ± 1.70 mm and 8.00 ± 0.47 mm against S. arueus. In the same studies, aqueous mucus of both fishes was not effective against E. coli and P. aeruginosa reported by Kumari et al. (2011). However, their report on low ZOIs values against S. aureus was similar to our results. Balasubramanian et al. (2012) demonstrated the antibacterial effect shown by C. idella against E. coli (17mm), K. pneumonia (7mm) and P. aeruginosa (15mm). Our findings on aqueous HFM and CFM also similar to above said study. It is evident from this discussion that different fish species exhibit variation in their antibacterial activity against similar or different microbial strains which might be due to difference in quality and quantity of proteins/ enzymes in secreted mucus.
Contradictory to our results Hellio et al. (2002) observed no antibacterial activity in aqueous mucus extracts of 13 fish species. However, they did not study crude mucus extract. Similarly, strong inhibitory effect of acidic extracts of Tilapia and bigrid cat fish against human pathogenic bacteria was reported by Rao et al. (2015). Whereas, no bactericidal effect was noticed in crude and aqueous extracts.
Current investigation revealed the change in antibacterial activity (either increased or decreased) after bacterial challenge. Subramanian et al. (2008) also reported that extruded slime produce by bacterial stress showed higher bactericidal effect than normal mucus in M. glutinosa L. Raj et al. (2011) observed increased antibacterial activity in mucus of C. carpio after virus infection. Our studies are in agreement with these studies. Futhermore, this increased or decreased inhibitory effect in fish species might be due to different enzymes level or different protein secretion in same/ different fish, during adverse conditions. Antibacterial activities shown by HFM and CFM of exotic carps, suggesting the presence of one or more antibacterial components in fish skin mucus. Oren and Shai (1996); Cole et al. (1997) and Ebran et al. (1999) isolated antibacterial peptides such as Paradaxin and pleurocidin and many others from epidermal mucus of different fishes. Andreu and Rivas (1999) reported that antibacterial components acts non-specifically, they form a pores in cell membrane of pathogenic bacteria; release their cell content and finally leads to cell lyses. Thus, we may assume that strong antimicrobial activity of epidermal mucus extracts of exotic carps against microbial strains may be due to pore formation ability of their antibacterial peptides in target cell membrane. Many authors such as Wei et al. (2010); Vennila et al. (2011) and Rao et al. (2015) revealed the MICs of different epidermal mucus extracts of many fishes viz. C. statius, Desyatis sephen and Himantura gerradi, tilapia, C. nigrodigitatus against several infectious microbes. Ebran et al. (1999) reported that the epidermal mucus of O. mykiss exhibited MIC at 50 µgml-1 against S. aureus, T. ticna at 60 µgml-1 against A. hydrophila and C. carpio at 50 µgml-1 against S. aureus, supporting the present results. Also Hellio et al. (2002) observed the MICs value of 13 fish species against different pathogen ranged from 25μgml-1 to 48 μgml-1. Our results are in agreement with above study. Contradictory to our findings, Oren and Shai (1996) and Cole et al. (1997) reported that the mucus extracts of winter flounderand moses fish have been found to inhibit the growth ofP. aeruginosa, E. coli and A. hydrophila and the MIC values obtained were in range of 40mgl-1 to 200 mgl-1. Lemaitre et al. (1996) reported that glycosylated proteins of MW 27kDa and 31kDa isolated from C. carpio strongly inhibits the growth of all tested microbial strains at approximately 5 µgml-1. Similarily, Ebran et al. (2000) noticed that three proteins (45kDa, 49kDa and 65 kDa) from hydrophobic supernatant of skin mucus of eel, tench and trout showed MICs values ranging from 1 µgml-1 to 5 µgml-1. However, same fish or different fishes exhibited different MIC against different or same bacterial strains. Different age, habits and habitats of different fishes could be the reason behind these variations. Thus, more studies are required to unhide the role of skin mucus of these economically important exogenous carps species as a good antimicrobial component against diseases causing fish and human bacteria.
Conclusion
In the present study, the increase in amount of mucus secretion after exposure to bacterial challenges indicates its involvement in protection against pathogenic attacks. High mucus secretion could be utilized as an indicator of increased stress level in fishes. Both crude and aqueous mucus extracts of healthy and challenged fishes showed broad spectrum of antibacterial activity against tested human and fish pathogenic bacteria. Therefore, fish skin mucus might possess bioactive components and could be utilized as a novel alternative of antibiotic used in aquaculture and for humans. Being a natural product, it could be helpful in reducing the antibiotic resistance and would be a cost effective.

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