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Pictures of fin clipping for DNA research
Frank Magallanes, OPEFE
August 11, 2000
Pulling specimens from Aquariums
Steadying specimen for extraction of dna sample
Clip small portion of fin
Wash, wipe and clean sample
Place specimen carefully into tube
Fill tube with ethanol
Begin procedure with next specimen
Specimen swabbed with antibiotics and then returned to aquarium.
The above images are of me extracting live piranhas from the aquarium for DNA research. The fins are clipped and then sent to the Orti Laboratory (School of Biological Science, University of Nebraska-Lincoln) for testing. At no time is the fish significantly harmed during the procedure.
WARNING: Piranhas have razor sharp teeth and powerful jaws. Some images display me placing my fingers in close proximity to the fishes mouth. Do not try this at home. I have handled piranhas for over forty years and my experience has taught me how to properly handle this type of fish outside of the aquarium.
TOP ROW: Image #1; I'm extracting fish from the aquarium, Serrasalmus altuvei. Image #2; The fish bit through the net, you can imagine what I'm thinking at this time. I'm holding on to fish with my left hand. Image #3; I'm positioning the fish for the photograph (Don't try this at home--keep your fingers away from the fishes mouth! The teeth are sharp and the jaws strong). Image #4; fish has calmed down some, but I'm not ready to let it go just yet for the primary photograph.
MIDDLE ROW: Pygopristis denticulata Image #1; I'm clipping the edge of the caudal fin. The scissors are very sharp. All the instruments are sterilized before I do any cutting, this insures that no contamination is present. Also, sterile gloves are worn at all times. I change these for each fish I clip. Image #2; The cut fin is put into a small container for an ethanol bath. The fin is rewashed several times. Image #3; I'm placing clipping into left hand to position it for wiping. Image #4; I'm wiping the clipping with a dry sterile gauze. This is done several times with a clean gauze to ensure no contamination is present prior to placing it in the shipment vial.
BOTTOM ROW: Image #1; I'm inserting the clipping to the vial using sterile tweezers. Image #2; Ethanol is poured into vial then the vial is sealed. An identification number is assigned and labeled to each vial with the scientific name of the fish. Image #3; Pygopristis denticulata is positioned for having its picture taken. Image #4; Final picture before fish is returned to the aquarium. The fin grows back after a period of time. The fish is kept moist at all times using an eye dropper, before and during the procedure. The eye dropper has antibiotic (tetracycline) to help prevent infections. The fishes cut caudal fin is the last part antibiotic is added just prior to the fish going back into the aquarium. Stress Guard™ is added to the aquarium water to help the fish recover from the stress of being handled outside of the aquarium.
Additional specimens that were not live, but also sent for DNA analysis (muscle tissue):
sterile gauze pads
1-2 inch diameter plastic tub to hold liquid for wash
2 pairs of latex gloves
2-pre-sterilized vials & sterilized tops
1-encapsulated tetracycline, liquefied
1-sterile eye dropper (to hold liquid tetracycline)
Stress Guard™ formulation
(Samples provided for DNA fin clipping from OPEFE for these results are Catoprion mento, Metynnis sp., Pygocentrus nattereri, Myleus rubripinnes, and Piaractus brachypomus). These were obtained from a commercial source, 1993.
Phylogeny of the Serrasalminae (Characiformes) based on mitochondrial DNA sequences
Previous work (Ortí et al. 1995) based on DNA sequences of mitochondrial (mt) rRNA genes showed three main groups within the subfamily Serrasalminae:
(1) a basal clade of herbivores (Colossoma, Mylossoma, Piaractus);
(2) the "Myleus" clade (Myleus, Mylesinus, Tometes);
(3) the "piranha" clade (Serrasalmus, Pygocentrus, Pygopristis, Pristobrycon, Catoprion, Metynnis). The genus Acnodon was placed as the sister taxon of clade (1+2). However, poor resolution within each clade was obtained due to low levels of variation among rRNA sequences. Complete sequences of the hypervariable mtDNA D-loop are now presented for a total of 40 taxa representing all genera in the subfamily to address intragroup relationships. Phylogenetic analyses of these sequences identify the same groupings as before and provide further evidence to support the following observations:
(a) the genera Serrasalmus and Pristobrycon are paraphyletic and form a group that also includes Pygocentrus;
(b) Catoprion, Pygopristis, and Pristobrycon striolatus form a well supported clade, sister to the group described above in 'a';
(c) distinction of subgenera within Myleus (i.e., Myleus, Prosomyleus, Myloplus) is not supported;
(d) Mylesinus and Myleus are paraphyletic, since Tometes sp. is the sister taxon of Mylesinus paraschomburgkii and Mylesinus paucisquamatus is most closely related to other species of Myleus.
Dr. Paulo Petry, Bio-Amazonia International, provided the vials and instructions to collect DNA samples.
Ortí, G., P. Petry, J. A. Porto, M. Jegú, and A. Meyer. 1996. Patterns of nucleotide change in mitochondrial ribosomal RNA genes and the phylogeny of piranhas. Journal of Molecular Evolution 42 (2): 169-182.
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