- Er det sant at medisin mot lakselus

tar knekken på rekene?

Almennur fundur um hóttanir móti vistskipanunum á føroysku firðunum Føroya Fróðskaparfelag saman við Umhvørvisstovuni – Tórshavn, Færøyene, 9. juni 2015

IRIS Lab-team: Renée K. Bechmann, Emily Lyng, Stig Westerlund, Shaw

Bamber, Sree Ramanand, Mark Berry, Marianne Nilsen

Elisa Ravagnan - Modelling

NIVA

Katherine Langford – Analysis of DFB Jannicke Moe & Dag Ø. Hjermann - Modelling

Université du Québec à Rimouski Piero Calosi – Scientific advisor

University of Leicester Paul Seear – Genomics

FLUCLIM (Effects of diflubenzuron on Northern shrimps (Pandalus borealis) at ambient and future climate conditions) is funded by Forskningsrådet – Havet og Kysten (2014 – 2016)

B a c k g ro u n d

Foto: John Øystein Berg, http://www.tronderbladet.no/debatt/article10050639.ece

• Ikke bra for laksens helse • Ikke bra for økonomien til oppdrettsnæringen • Ikke bra for ville laksefisk

Rensefisk

Mekanisk fjerning av lus

Beskyttelsesskjørt

Kjemikalier (= Medisiner)

o Badebehandling Laksen svømmer i et bad med medisin noen minutter (hydrogen-peroksid, azametifos, cypermetrin, deltametrin)

o Spesialfôr Godkjent legemiddel tilsettes fiskefôret (Diflubenzuron, teflubenzuron, emamektin, maks 14 dager)

B e h a n d l i n g a v l a k s e l u s

0

1000

2000

3000

4000

5000

2005 2006 2007 2008 2009 2010 2011 2012 2013 2014

Fôrmiddelet diflubenzuron (kg aktiv substans)

= 8000 tonn spesialfôr med diflubenzuron (Releeze) brukt i 2014

L a k s e l u s er en parasitt som plager laksen

Shrimps Pandalus borealis

F u t u r e c l i m a t e

Ocean acidification (pH 7.6) and increased temperature (10C)

D i f l u b e n z u r o n from medicated fish feed

Anti-parasitic drug against salmon lice

A m b i e n t

c l i m a t e pH 8.1

7C

Benzoylurea pesticide

Log Kow 3.8

A chitin synthesis inhibitor

- Can salmon lice medicine be a problem for shrimps?

Experiments with shrimps exposed to medicated salmon feed containing the chitin syntesis inhibitor diflubenzuron (DFB) as the active substance

Feed waste

Medicated fish feed

A d u l t s h r i m p s : • Can eat feed waste and contaminated sediment • Suspended particles of medicated feed and fish feces • Dissolved DFB from feed and fish feces

Positive effect for salmon: Salmon lice die during

moulting

- H o w c a n s h r i m p s b e ex p o s e d ?

L a r v a e : • Suspended particles of

medicated feed and fish feces • Dissolved DFB from feed and

fish feces

14 days treatment of salmon to get rid of

salmon lice

Yes! Free lunch!

Yummi fish feed!

Stock solutions of sediment slurry with medicated or clean fish feed

10°C 7°C

+ CO2

Continuos flow exposure system

C

Pressure regulator

Solenoid valve

Connection plug

Aqua Medic

system

CO2

Temperature controlled seawater

Heat exchangers

The larvae

2 g 3.5 mm pellets

Main DFB exposure:

Flow: 200 ml/min

A little extra DFB:

8% fish feed in sediment:

0.5 mg/L

Shrimp larvae were exposed to the medicated fish feed for 2 weeks from hatching

The larvae

Hatching

Two weeks exposure to medicated pellets with DFB

Climate acclimation of shrimps (females with embryos)

DFB

Ambient Climate

Future Climate

DFB

Age of larvae (days)

10°C, pH 7.6

10°C, pH 7.6

7°C, pH 8.1

7°C, pH 8.1

-14 -7 0 7 14 21 28

Tr e a t m e n t s & T i m e :

DFB at Ambient Climate

DFB at Future Climate

x 6

x 6

x 6

x 6

The larvae

200

X 6 batches

200

X 6 batches

200

X 6 batches

200

X 6 batches

Ambient climate: 7°C and pH 8.1 Future climate: 10°C and pH 7.6

Control DFB OA/T OA/T + DFB

X 6 X 6 X 6 X 6

Exposure to DFB

Exposure to DFB

The larvae

Analysis of DFB by Katherine Langford, NIVA

0.0

0.5

1.0

1.5

2.0

2.5

3.0

Total Concentration of DFB in the aquaria (mean + SE)

i.e. DFB attached to feed particles and dissolved in the water

Ambient Climate + DFB Future Climate + DFB

4 h

24 h

4 h

24 h

Controls samples: <0.01 – 0.02 µg/L

µg

/L D

FB

n = 3 n = 3 n = 3 n = 3

DFB was analysed at NIVA by liquid chromatography coupled to tandem mass spectrometry (LC/MS/MS)

Concentrat ion of d i f lubenzuron

1 mm mesh

200 ml/min After 4 hours: 2 µg/L After 24 hours: 1 µg/L

Conc. of DFB

The pellets after 24 h in the cylinder:

3.5 mm pellets

More DFB remained in the flow-through aquaria than we expected

Analysis of DFB by Katherine Langford, NIVA

The mesh in the cylinder

SURVIVAL for shr imp lar vae

0

20

40

60

80

100

0 5 10 15 20 25 30

Pe

rce

nt

surv

ival

of

shri

mp

larv

ae

Days post-hatch

- 25 %

- 56 %

- 82 %

E x p o s u r e t o D F B o r c l e a n p e l l e t s

Mean percent survival for 6 replicate batches of shrimp larvae exposed to DFB at two climate scenarios

Ambient Climate Control

Future Climate Control

Ambient Climate + DFB

Future Climate + DFB

Time

Development time (zoeal progression) for shrimp larvae

Photo: Tandberg & Arnberg

Stage I zoea Stage II zoea Stage III zoea Stage IV zoea

The tail of the shrimp larvae change when they develop from stage II – III - IV

The larvae

Shrimp larvae go through five pelagic zoea stages and one megalopa stage before they settle as post-larva We followed them until stage 4

0

20

40

60

80

100

0

20

40

60

80

100

% stage II % stage III % stage IV

Day no = age of shrimp larvae Day 6 Day 17 Day 19 Day 9 Day 13

Co

nt

ro

l

DF

B

DFB

Co

nt

ro

l

L A R VA L D E V E L O P M E N T T I M E f o r t h e s u r v i v o r s

Future

Climate

Ambient

Climate D

FB

Co

nt

ro

l

DFB

Co

nt

ro

l

DFB

Co

nt

ro

l

DFB

Co

nt

ro

l

DFB

Co

nt

ro

l

DFB

Co

nt

ro

l

No stage II No stage III No stage IV

Day 27

Exposure to medicated (DFB) or clean (control) pellets

5 shrimp larvae and

150 Artemia nauplii

in 1 litre seawater.

Wait 5 hours.

Count remaining Artemia.

Feeding rate

for stage II and IV larvae

Pandalus borealis larvae

Artemia nauplii

1 litre

Nu

mb

er

of A

rtem

ia e

ate

n p

er

shrim

p larv

ae

pe

r ho

ur

0

1

2

3

4

5

6

1 2 3 4 5 6 7 8

code B

Num

ber

of A

rtem

ia e

aten

per

s

hrim

p la

rvae

per

hou

r Stage II zoea Stage IV zoea

Ambient Climate Ambient Climate Future Climate Future Climate

DFB

DFB

DFB DFB

Control Control

Control

Control

No stage 4, these are stage 3!

Feeding rate for the survivors

B S T

« B a m b e r ’ s S w i m m i n g T e s t »

L i g h t

I R l i g h t Detector

Shrimp larvae are photo-positive

(they swim towards the light)

- Will DFB affect swimming activity at

ambient or future climate conditions?

Shaw Bamber, IRIS

Future Climate

• STAGE I: Similar swimming activity for control and DFB exposed larvae.

• STAGE II: Lower swimming acitivity for DFB exposed larvae than control larvae.

STAGE I: Before first moult

STAGE II: Those that survived first moult

Ambient Climate

Similar swimming activity for control and DFB

exposed STAGE I and II larvae.

Example - Raw data

Control (future climate)

DFB (future climate)

Shaw Bamber, IRIS

S w i m m i n g a c t i v i t y

Day 0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30 32 34

Day 0 2 4 6 8 10 12 14 16 18 20 22 24 26 28

Determine survival and stage of larvae

2 weeks exposure

2 weeks exposure

1 day exposure

Ambient Climate, 7C

Ambient Climate, 10C

Ambient Climate, 7C

Ambient Climate, 10C

Determine survival

0

20

40

60

80

100

Day 11 Day 22 Day 33

Ambient Climate Control

Ambient Climate + DFB

Future Climate Control

Future Climate + DFB

7.2C, pH 8.06

9.6 C, pH 7.63

* * *

% s

urv

ival

of

shri

mp

larv

ae

O n e d ay ex p o s u r e : S u r v i va l o f s h r i m p l a r va e

Mean (st.dev.), n = 4 x 140 shrimp larvae

At 10C one day exposure caused high

mortality, but not at 7C

Effect of DFB exposure on adult shrimps

Control shrimps moulted successfully in the lab a few weeks after all eggs had hatched

Shrimp exit

Pilot experiment with adult shrimps 2014

Flubenzurones are chitin synthesis inhibitors: they stop the organism from casting its exuvium during the molting process resulting in death.

Pilot experiment with adult shrimps 2014

A d u l t s h r i m p s d i e d d u r i n g m o u l t i n g

w h e n e x p o s e d t o m e d i c i n e p e l l e t s

c o n t a i n i n g d i f l u b e n z u r o n

0

20

40

60

80

100

120

140

160

19

- m

ars

21

- m

ars

23

- m

ars

25

- m

ars

27

- m

ars

29

- m

ars

31

- m

ars

2-

apri

l

4-

apri

l

6-

apri

l

8-

apri

l

10

- ap

ril

12

- ap

ril

14

- ap

ril

16

- ap

ril

18

- ap

ril

20

- ap

ril

22

- ap

ril

24

- ap

ril

26

- ap

ril

28

- ap

ril

30

- ap

ril

2-

mai

4-

mai

6-

mai

8-

mai

Acc

um

ula

ted

nu

mb

er o

f m

ou

lts

0

10

20

30

40

50

60

70

80

90

Acc

um

ula

ted

nu

mb

er o

f

de

ad s

hri

mp

s

Moult related mortality

Medicated fish feed (0.6 g Diflubenzuron/kg)

Controls

Successful m o u lt ing

Controls

Medicated fish feed

Control A

Control B

DFB A

DFB B

172 162 179 151

Mixed size shrimps 9.4C

pH 8.0, 400 L tanks

no moulting for previously exposed shrimps

Expo Depuration

Expo Depuration

Number of shrimps at start

Dose: Ca 1 pellet (3.5 mm) per

shrimp per day for 8 days

Feed waste

There is no such thing as a free

lunch

1-2 weeks exposure (with or without sediment) 1 month depuration Shrimps eat salmon feed (normal or medicine pellets) High mortality at both temperatures and for all sizes of adult shrimps during the 1 month depuration after ending 1-2 week exposure

Effects on adult shrimps

Rekelarver og andre krepsdyr i planktonet kan få i seg dette…

Rekelarver er veldig følsomme for diflubenzuron fra medisin-fôr!

Feces med flubenzuroner

Fôr - spill

Medisinfôr med flubenzuroner

Lite metaboliseres Alt skilles ut raskt

Flubenzuroner har «lang holdbarhet» i det marine miljø

Voksne reker kan spise dette og få problemer under skallskiftet

Oppsummering!

Strømmen tar med seg rester av medisin-fôr, feces og forurenset

sediment

Både rekelarver og voksne reker dør under skallskifte når de utsettes for medisin-fôr med diflubenzuron. Resultatene tyder på mer negative effekter ved 10C enn ved 7C

Konklusjon!

Så svaret er:

Ja! Lakselusmedisin kan ta knekken på rekene hvis for mye

slippes ut i fjordene våre.

Paul Seear is working with the shrimp genomics

R e m a i n i n g p r o j e c t w o r k :

Genomics

The aim is to develop a genetic marker for exposure to chitin synthesis inhibitors in shrimps

Modelling

…. analyse the multiple stressors impacts of increased temperature and OA together with

DFB exposure on Northern shrimp populations

F u t u r e ( 2 0 1 6 ) p r o j e c t w o r k :

Jannicke Moe & Dag Ø. Hjermann

Elisa Ravagnan

Kathy Langford will analyse DFB in the shrimps from the experiments

Chemistry

- How much diflubenzuron have the shrimps accumulated?

H A V K Y S T

M o r e i n f o :

r k b @ i r i s . n o

Stig Westerlund Emily Lyng

Shaw Bamber Mark Berry

Elisa Ravagnan Marianne Nilsen

Sreerekha S. Ramanand Renée K. Bechmann

Ph

oto

: Elisabeth

Tøn

nessen

Katherine Langford, Jannicke Moe

Dag Ø. Hjermann

Université du Québec à Rimouski

Piero Calosi

University of Leicester

Paul Seear

Partners

T h a n k y o u f o r y o u r a t t e n t i o n !

NFR

The local crowd

Thank you for the money:

Thak you for the science: