Functional feeds. bad bug busters to reduce the impact from disease in farmed fish and shrimp. Peter Coutteau, PhD Nutriad International, Belgium

Transcription

1 Functional feeds bad bug busters to reduce the impact from disease in farmed fish and shrimp Peter Coutteau, PhD Nutriad International, Belgium

2 Antibiotics to combat shrimp disease : economic risks Zero tolerance policy by Food and Drug Administration (FDA) and European Commission (EC) on unauthorized antibiotics FDA refusals of shrimp entry for banned drug residues peak since 2014

3 Vibrio sp. are experts in acquiring antibiotic resistance

4 Antibiotic resistance plasmid mediated (Han et al., 2015) tetracycline resistance in Vibrio parahaemolyticus, HPND strain Mexico

5 Functional feed additives to improve productivity in shrimp Objective: Promoting healthy gut microflora Benefits: Healthy Gut = functional gut for digestion (growth, FCR) Healthy Gut = barrier for entry of infections (survival) Modes of action: 1. Modulating healthy gut flora : Inhibiting growth of pathogenic bacteria (parasites) Promoting beneficial bacteria (probiotics) 2. Reducing pathogenicity (Quorum Sensing Inhibition)

6 SURVIVAL % Effect of functional feed additives with combined antibacterial/qs inhibition action on survival of P. vannamei Field and lab experience in EMS and non-ems countries FIELD trials Nutriad Customer trials +44% +25% LAB challenge EMS/AHPND (Loc et al., 2015) % +18% +16% +21% +16% +107% +62% 10 0 PANAMA 2009 PANAMA 2010 HONDURAS 2014 ECUADOR 2015 VIETNAM 2013 EMS MEXICO 2014 EMS MEXICO 2014 EMS LAB EMS immerion challenge LAB EMS oral challenge CONTROL Synergistic phytobiotics (antibacterial/qs inhibition)

7 Bacterial growth (folds) Demo of bactericidal activity against Vibrio parahaemolyticus (AHPND strain) LB 0.05% 0.10% 0.30% 0 0 h 2 h 4 h Phytobiotic product (SGM, directly added to liquid culture medium) exhibited bactericidal activity against the causative agent of AHPND, Vibrio parahaemolyticus, in culture medium. Prof. Dr. Chu-Fang Lo & Dr. C.T. Lee (2014) National Cheng Kung University, Tainan, Taiwan

8 Verifying effect of heating effects on bactericidal activity against Vibrio parahaemolyticus (AHPND strain) product was pre-heated for 15 min at C. Heating does not influence the bactericidal activity of the phytobiotic product Prof. Dr. Chu-Fang Lo & Dr. C.T. Lee (2014) National Cheng Kung University, Tainan, Taiwan

9 MOA 2: Reducing pathogenicity Quorum Sensing

10 Prof. Bonnie L. Bassler DEPARTMENT OF MOLECULAR BIOLOGY University of Princeton, USA bonnie_bassler_on_how_bacteria_communicate?language=en

11 QS systems of different aquatic pathogens

12

13

14 Evaluation of the ability of a gut modulating phytobiotic in interfering on quorum sensing of AHPND Vibrio parahaemolyticus Collaboration between Nutriad and NCKU Leading researchers: Prof. Dr. Chu-Fang Lo & Dr. C.T. Lee Institute of Bioinformatics and Biosignal Transduction National Cheng Kung University, Tainan, Taiwan

15 PNAS August 25, 2015 vol. 112 no. 34 Leading researchers: Prof. Dr. Chu-Fang Lo & Dr. C.T. Lee Institute of Bioinformatics and Biosignal Transduction National Cheng Kung University, Tainan, Taiwan

16 Source: Prof. Dr. Chu-Fang Lo & Dr. C.T. Lee (2014) National Cheng Kung University, Tainan, Taiwan

17 Source: Prof. Dr. Chu-Fang Lo & Dr. C.T. Lee (2014) National Cheng Kung University, Tainan, Taiwan

18 Wild type pva1 cured + pirab AHPND tissue damage is caused by a secreted toxin AHPND HP Wild type Healthy HP pva1 cured + pirab pva1 cured Control

19

20

21 genes that may be regulated by QS in V. parahaemolyticus: (QS upregulated) (QS downregulated) opar, a luxr homologue (master regulator in V. harveyi QS) VPA1026, encoding for T6SS2 apha, a gene regulator VP1386, encoding for T6SS1 Both toxin genes, pira and pirb will be also analyzed.

22 phytobiotic

23 Mortality (%) In vivo challenge AHPND Time (h) ToP T1 T2 T3 T4 T5 ToN T1 seems to be the most effective feed (21% improvement), however, after statistical analysis, there is no significant difference between the T1 group and the positive control group.

24 NCKU study - conclusions The data suggest that phytobiotic, gut modulating feed additives may influence QS signaling and the expression of toxin genes (pir A and pir B) at RNA level. The results of in vivo trial suggest that these feed additives may contribute 21% improvement of survival after challenged with Vibrio parahaemolyticus (AHPND strain). Further research is required to fully understand the QS pathway of V. parahaemolyticus

25 The new hidden killer? 25

26 Hepatopancreatic microsporidiosis (caused by Enterocytozoon hepatopenaeid, EHP) Photo from Associated Professor Dr. Chalor Limsuwan, Thailand

27

28 EHP infection was first reported in 2004, characterized in 2009

29 Enterocytozoon hepatopenaei (EHP) obligate intracellular parasitic fungi with complex life cycle Microsporidian >1200 species belonging to 143 genera, and about 100 genera infect crustaceans and fish Oral route transmission from shrimp to shrimp Spread throughout China, SE Asia and possibly Mexico Low mortality but associated with retarded growth Very difficult to inactivate due to thick wall of spore

30 Life cycle of Enterocytozoon spp. HP cell

31 Efficacy of natural anti-parasitics: protozoan endoparasites Gregarines Penaeus vannamei Sparus aurata (Gilthead seabream) Mxyosporidae

32 Take-home message Novel functional feed additives designed to have multiple modes of action, which are relevant in the light of EMS and EHP: 1. Selective bacteriostatic action MIC concentration) against a broad range of pathogenic bacteria, including Vibrio parahaemolyticus 2. QS inhibiting activity (@ concencentrations below MIC) against Vibrio and other sp. 3. Anti-parasitic action against gut parasites, e.g. gregarines (EHP?) Results : - reduce mortality due to bacterial (co-)infections - Control growth of bacterial pathogens & gut parasites - Reduce/delay onset of EMS toxin production - Work synergistically with probiotics reduce the impact from diseases and parasites on productivity

33