Immunomodulatory potential of different herbal plant extracts on striped catfish (Pangasianodon hypophthalmus)

: in vitro and in vivo approaches

Abstract

Striped catfish (Pangasianodon hypophthalmus) is one of the economically important fish species, which is widely developed in several Asian countries. Like other fish species, striped catfish also suffer from many kinds of diseases including bacteria, fungi and parasites under stress conditions due to over-expansion farming areas and increase intensification. To control the diseases, regular use of antibiotics and other chemotherapeutics has been causing many negative impacts on the environment and food safety. Therefore, there is an increased demand to look for eco-friendly preventive measures to promote sustainable aquaculture. In recent years, the application of medicinal plants has attracted a lot of attention and become the active subject of scientific investigation. The present study aimed to elucidate the efficiency of different plant extracts in striped catfish in order to find an alternative replacement and reduce the use of antibiotics.
Many medicinal plants have been documented to possess positive effects on the immune responses of aquatic cultivated animals. However, the use of natural products has not been yet popularly applied in striped catfish farms. It may spend a lot of time and the number of experimental animals when all plant products were examined in case of an in vivo application. Hence, based on bibliography review data and on a survey in fish farms of Mekong Delta, 20 plants possessing potential immunostimulatory activities were selected for in vitro study using the striped catfish peripheral blood mononuclear cells -PBMCs and head kidney leukocytes- HKLs models. Five plant extracts were then validated for their effects on the blood indices, humoral immune responses and disease resistance in striped catfish via oral administration. Moreover, the effects of single supply diets versus mixture diets with Phyllanthus amarus and Psidium guajava extracts on striped catfish health were continuously submitted. In addition, the protein expression profile was also investigated to provide a better understanding of the metabolic pathways related to immune response, antioxidation and lipid metabolism in fish liver after oral administrated plant extracts. The different polarities of the extract solvents may cause a wide variation in the level of bioactive compounds in the extract, which may lead to different effective degrees on aquatic animal immune responses. The immunomodulatory effects of crude ethanol extract of P. amarus and P. guajava as well as their fractions including n-hexane, dichloromethane, ethyl acetate, aqueous and non-tannin were therefore compared via in vitro experiment. The effects of different pure compounds isolated from P. amarus and P. guajava were also investigated in this study. Furthermore, biological activities plant
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products related to inflammation are largely unknown in fish. The molecular mechanism behind the effects of P. guajava extracts on fish immune responses were also highlighted and discussed.
The results demonstrated the ability of several plant extracts in activating of humoral immune responses (lysozyme, complement, and total immunoglobulin) in a dose dependent manner in striped catfish PBMCs and HKLs after 24 h. Several extracts also induced a strong upregulation of 4 cytokines (il1β, ifnγ 2a and 2b, and a2 mhc class II) according to the concentration, time points and kind of leukocytes. Of the ethanol extracts, Phyllanthus amarus, Psidium guajava, Mimosa pudica, Azadirachta indica and Euphorbia hirta could stimulate the striped catfish innate immune response (lysozyme and complement) and adaptive immune response (total Immunoglobilin) as well as protecting better resistant capacity in striped catfish against Edwardsiella ictaluri pathogen. The study was further revealed how single or mixture supply of P. amarus and P. guajava extract-based diets acted in the regulation of immune responses, significantly reducing fish mortality. Moreover, oral administration of P. amarus and P. guajava extracts displayed an upregulation of several proteins involved in immune response in striped catfish liver, the positive synergistic effects of liver proteome profile related to immune system processes were observed under mixture of administration. The crude ethanol extracts of P. guajava and P. amarus, their fractions and the pure compounds at certain concentration can potentially act as immunomodulators in HKLs of striped catfish at 24h. P. guajava dichloromethane fraction beneficially enhanced TLRs-MyD88-NF-κB signaling pathway, following induce the levels of inflammatory and apoptosis cytokines as well as lysozyme, iNOS and RBA productions, providing the better understanding about the mechanism behind the effects of P. guajava extracts on fish immune response at molecular level. Taken together, our studies suggested that boosting the immune response by ecologically friendly products, especially Phyllanthus amarus and Psidium guajava derived products is an effective strategy to promote sustainable aquaculture via improvement the health status as well as resistance to pathogens in striped catfish.

Date of Award	19 Dec 2019
Original language	English
Awarding Institution	University of Namur
Sponsors	ARES-CCD
Supervisor	Patrick Kestemont (Supervisor), Bui Thi Bich Hang (Co-Supervisor), Frederik De Laender (President), Joelle Quetin-Leclercq (Jury), M. L. SCIPPO (Jury), Robert Mandiki (Jury) & Silvia Torrecillas (Jury)