Sara Persson Rodrigo Esparza-Salas Mariam Khammari Jannike Ring - - PowerPoint PPT Presentation

Determination of the nematode parasites Pseudoterranova sp and Contracaecum sp in the stomach of grey seals, ringed seals and harbor seals from Sweden

Sara Persson

Rodrigo Esparza-Salas Mariam Khammari Jannike Ring Elina Thorsson

Photo: Lars-Arne Söderberg

• Baltic cod

– Fewer – Smaller – Low body condition – Parasites

• Causes?
• Fishing
• Increasing population of seal

– increasing amount of parasites in cod?

• Knowledge about how the

parasites vary in the seal is lacking

Background

• Pseudoterranova decipiens in

cod muscle

– ”Cod worm” or ”seal worm”

• Contracaecum osculatum in cod

liver

– ”Liver worm”

• Seals are main host
• Life cycle with many

intermediate fish hosts

Cod/seal parasites

• 41 ringed seals 2015-2017
• 60 grey seals 2017-2018
• 55 harbour seals 2017-2018

Materials and methods

• Parasites were weighed (if all nematodes could be

collected) and stored in 75% alcohol

• Stomach fluid from necropsied seals was preserved in

buffer for DNA analysis

• Parasite lesions were noted

• Cod worm and liver worm look identical macroscopically
• Ringed seal: 4.7%

– only 1-2 worms

• Grey seals: 100%

– Mean 20.5 gram – 4-95 gram

• Harbour seals: 98%

– Mean 1.8 gram – 0.2-13 gram

Results: prevalence, weight

Stomach lesions in the seals

Ringed seal 0% Grey seal 65% Harbour seal 64%

• Adult parasites (L5) reproduce and produce eggs
• Large variation in larvae size within the seal
• Proportion of nematodes of the larger type varied a lot in

both GS and HS (0 - 100 %)

• Weighing parasites does not say much about capacity to

spread eggs (and can be very time consuming)

Variation in larvae stages

• Snap shot
• Differences in defense of the seal host
• Differences in fitness of the parasites (in earlier life stages)
• P.decipiens fecundity (egg production) will be affected by the seal

– immune system, – age, – diet – how many parasites that the seal is continuously infected by (McClelland, 2002, Johansen et al., 2010).

Why the large variation in the amount of adult nematodes?

• A new set of DNA barcode markers was designed to

target a short fragment of the mitochondrial LSU gene of parasitic worms of the family Anisakidae.

• The LSU fragment was amplified in DNA extracts from

114 seal stomach samples

• LSU was amplified from DNA abstracts from the ethanol

used to preserve the nematodes (N=12)

• The sequence output data was de-multiplexed, filtered

and identified to species by comparing to a custom- made reference sequence database of LSU genes of Anisakidae

• 59 of the studied samples amplified the LSU fragment

successfully

– Inhibiting substances, degraded DNA

DNA analysis of stomach fluid

Proportion of nematode species in stomachs from seals collected from different areas (mean and range) Bothnian bay N Contracaecum

• sculatum
• P. decipiens

sensu lato Unidentified Anisakidae, Ascaridida och Chromadorea Ringed seal 1 99% 0.002% 0.002% Grey seal 3 100% (99.99-100) 0.003% (0-0.008) 0.004% (0-0.01) Bothnian sea Grey seal 11 99.6 % (97.5-100) 0.2% (0.001-2.5) 0.1% (0.001-0.05) Baltic proper Grey seal 22 97.1% (38.6-100) 2.9% (0-61.4) 0.003% (0-0.013) Harbour seal 2 0.07% (0.005-0.02) 93.9% (87.9-99.9) 6.1% (0-12.1) Skagerack+ Kattegatt Harbour seal 20 7.7 % (0-94.4) 88.0% (1.9-100) 2.5% (0-23.3)

Results

• Parasite lesions in the gastric wall are common but most
• ften mild (GS and HS)
• Pattern:

– grey seals are most often infected by Contracaecum spp only – harbour seals are most often infected by Pseudoterranova spp

• nly

The DNA barcoding provide a time-efficient method for the identification of nematode parasites that would be otherwise impractical to carry out in large-scale monitoring

Conclusions

• Score/classification of parasite load
• Score/classification of number of

large nematodes (~fecundity)

• DNA analysis of species and species

proportion within each seal

• Adding more data
• Geographical study (ideas?)
• Harbour seals from the Baltic proper

interesting

• Variation with, age, sex. Health.

Further studies