NOAA Technical Memorandum NMFS 

This TM Series is used for documentation and timely cornrnunicatlon of preliminary results, imterirn reports, or 

Special purpose information, and have not received complete formal review. editorial control. or detailed editing 

NOVEMBER 198 1 

AN EVALUATION OF TAGGING, MARKING, AND 

TATTOOING TECHNIQUES FOR SMALL DELPHINIDS 

Merrill J. White, Jr. 

Hubbs/Sea World Research in Institute 

San Diego, CA 

Jacqueline G. Jennings 

Southwest Fisheries Center 

National Marine Fisheries Service 

La Jolla, CA 

Walter F. Gandy 

Southeast Fisheries Center 

National Marine Fisheries Service 

Bay St. Louis, MS 

and 

Lanny H. Cornell 

Sea World, Inc. 

San Diego, CA 

U.S. DEPARTMENT OF COMMERCE 

Malcolm Baldrige. Secretary 

National Oceanic and Atmospheric Administration 

John V. Byrne. Administrator 

National Marine Fisheries Service 

William G. Gordon, Assistant Administrator for Fisheries 
ACKNOWLEDGEMENTS 

Preparation of this report was covered under NMFS Contract 03-7-208-35282 

to the Hubbs/Sea World Research Institute. It serves to synthesize results of 

a coordinated set of studies by the NMFS, HSWRI and other NMFS contractors. 

The following individuals and institutions participated in the research effort: 

David De Blanc, Biological Technician, Mississippi Laboratories, Southeast 

Fisheries Center, National Marine Fisheries Service, Bay St. Louis, 

Mississippi. 

Dr. William E. Evans, Director, Hubbs/Sea World Research Institute, 

San Diego, California. 

Dr. Keith Farrell, Professor of Cryobiology, Washington State University, 

Pullman, Washington. 

Dr. Jeffrey Graham, Research Associate, Physiology Research Laboratory, 

Scripps Institution of Oceanography, La Jolla, California. 

Ms. Ingrid Kang and Staff, Training Department, Sea Life Park, 

Waimanalo, Hawaii. 

Romaine Maiefski, Civil Engineer, Maiefski Engineering Consultants, 

San Di ego , Calf i ronia. 

Jeffrey Norris, Research Assistant, Hubbs/Sea World Research Institute, 

San Diego, California. 

Dr. Edward Shal lenburger, Vice President, Director of Operations, Sea 

Life Park, Waimanalo, Hawaii. 

Dr. Jay P. Tullis, Civil Engineer, Utah State Water Research Laboratory, 

Logan, Utah. 

Jacqueline Jennings of the Southwest Fisheries Center served as Chief 

Official Technical Representative for the contract. Referral to products 

by name does not imply endorsement by the government. 

i 
IN MEMORIAM 

The authors wish to particularly acknowledge the invaluable contributions 

of the late Dr. R. Keith Farrell to the research project. Dr. Farrell, Professor of Cryobiology at Washington State University, was a pioneer in the 

use of cryogenic marking techniques for terrestrial and marine animals. 

“La joie de vivrell emanated from Dr. Farrell for his work and the finer 

things of life. He is sincerely missed. 

ii 
INTRODUCTION 

Marking and tagging have been recognized as important techni-ques for 

collecting data on marine mama1 s since the rnid-1920'sY when "discovery type" 

tags were developed for commercially exploited stocks of whales. 

An increasing dependence on marine resources has created social pressures 

against exploitation of marine mammals and a concern for proper management 

that has become international in scope. 

the Marine Mammal Protection Act of 1972, and international regulations via 

the International Whaling Commission, IJnited Nations (FAO) and CITES, are 

pending to provide blanket protection for all marine mammals. 

To obtain important field information for marine mammal management the 

Increased awareness has resulted in 

National Marine Fisheries Service has developed marine mammal programs in 

biology, technology, and assessment. Visual identification of individuals is 

essential for this type of research and an important aspect of this program 

involves porpoise marking, tagging, and subsequent observation using a Porpoise 

School Impoundment System (PSIS), designed by NOAA's National Fisheries 

Engineering Laboratory and Southwest Fisheries Center. 

The Porpoise School Impoundment System functions as a platform for 

marking and tagging small pelagic delphinids, primarily spotter porpoise, 

Stenella attenuata. Specialized procedures and equipment are necessary to 

process sufficient numbers of animals in an efficient and humane manner to 

insure adequate sample size. 

Under the guidance of Ms. Jacquel ine G. Jennings (Leader, Porpoi se 

Tagging Project, Southwest Fisheries Center), the Hubbs/Sea World Research 

Institute conducted a research program to evaluate marking and tagging 

techniques and equipment for use with the Porpoise School Impoundment System 

iii 
(PSIS). 

mentioned in the Acknowledgements Section. 

This work was coordinated with that of the other NMFS contractors 

Procedures and equipment for tagging were developed according to the 

following criteria derived from the Marking and Tagging Workshop convened 

by the Southwest Fisheries Center, National Marine Fisheries Service -- 

May 1977; 

1) 

2) 

3) 

Tag design that minimizes tissue trauma. 

Tag fabrication from biocompatible materials. 

Tag fabrication from materials durable to field environmental 

factors for at least one year. 

4) Tag application in less than one minute per animal. 

Procedures and equipment for marking were developed according to the 

following criteria: 

1) Equipment designed with sufficient coolant capacity and insulation, 

or pigment reservoir to allow marking large numbers of animals. 

Equipment designed for marking with multiple symbol capacity. 

Marking equipment fabricated from materials durable at cryogenic 

temperatures and in field situations. 

Marking equipment portable in field situations. 

Mark application in less than one minute per animal. 

Legible mark for at least one year. 

2) 

3) 

4) 

5) 

6) 

Experiments were conducted to provide the National Marine Fisheries 

Service with observations and recommendations based on the proposed criteria, 

although the time schedule for field operations with the Porpoise School 

Impoundment System (PSIS) did not allow all tags 

for the proposed year period. 

and marks to be evaluated 

However, tags and marks cited in the report 

iv 
as "during the study period" were eva 

October, 1978 and April, 1980. 

This report will review research 

1) Tagging equipment design and 

uated for at least a full year between 

progress in three separate sections: 

evaluation for small delphinids. 

2) Cryogenic marking equipment design and evaluation for small 

delphinids. 

3) The feasibility of tatooing small delphinids. 

For the purpose of clarity the following definitions for "tag" and 

'?nark" will be used: 

Tag -- A visible object attached to, implanted in or covering the 

epidermis for the purpose of identification. 

Mark -- A visible symbol produced by depigmentation of the epidermis 

with cryogenic temperatures or its implantation with indelible 

pigments for the purpose of identification. 

V 
TABLE OF CONTENTS 

Introduction 

Section I - Tagging Equipment Design and Evaluation 

Tag Group #1 Hampshire Pig (Sus scorfa) 

Tag Group #2 Common Dolphin (Delphinus delphis) 

Tag Group #3 Common Dolphin (Delphinus delphis) 

Tag Group #4 Common Dolphin (Delphinus delphis) 

Tag Group #5 Pacific White-sided Dolphin 

(Lagenorhynchus ob1 i qui dens ) 

Tag Group #6 Pacific White-sided Dolphin 

Tag Group #7 Hawai i an Spi nner (Stenel 1 a 

(Lagenorhynchus ob1 i qui dens ) 

longirostri s) 

Tag Tests - Utah State Water Research Laboratory 

Tag Group #8 Hawai ian Spi nner (Stenel 1 a 

longirostris) 

Sect i on I I - Cryogenic Marki ng Equipment Des i gn and Eva1 uati on 

Cryogenic Marking Evaluation - Phase I 

Cryogenic Marking Evaluation - Phase I1 

Section I11 - The Feasibility of Tatooing Small Delphinids 

I. Polystyrene Particle Spray 

11. Schvco Injector 

I I I. The Verni tron Hypodermic Inject i on Apparatus 

vi 
Section I - Tagging Equipment Design and Evaluation 

vi i 
Tag Evaluation _. - Hampshire Pig (Sus scrofa) 

Initial tag evaluations were performed with a Hampshire pig (Sus 

scrofa) maintained at the University of California's Laboratory Animal 

Facility. The pig's epidermal - fat - muscle layers are anatomically similar 

to delphinids and they can sustain multiple cutaneous implants without health 

complications. 

Prior to tag implant, the animal was manually restrained and scrubbed 

with septodyne disinfectant. After tagging, it was monitored periodically by 

Institute and Laboratory Animal Facility staff. 

Tag Group #1 Hampshire Pig (Sus scrofa) 

Tag Description and Application 

1. Floy Spaghetti Tag (Fig. 1) 

The Floy spaghetti tag had a thin, flat, pointed type 304 stainless 

steel barb (35 mm x 8 mm) attached to a flexible monofilament lead with 

f 1 uorescent polypropylene streamer (14 cm) . 

material. The tag was applied with a jab stick (Fig. 2). The length of the 

jab stick's holding pin determined depth of penetration. 

penetrated the tissue at an acute angle (from verticle plane) and anchored by 

Stai nl ess steel i s a bi ocompati bl e 

The anchor barb 

rotating at a right angle to the entry point when back pressure was applied 

to the streamer. 

Design Concept 

This tag was originally designed for fish. When applied properly, 

the tag head swiveled 90" relative to the streamer as shown in Fig. 1. 

-1- 
The barb no longer pointed mddially, which was intended to reduce tag 

shedding. The tag head was flat and thin with a slight curvature, 

allowing penetration and anchoring in body tissue. 

2. Floy Arrowhead Tag (Fig. 3) 

Also designed for use with fish, the Floy arrowhead tag had a round, 

pointed, nylon dart (16 mm) with two adjoining flukes (30 mm). 

a biocompatible material. The dart was attached to a flexible vinyl 

Nylon is 

tubing streamer (14 cm). This tag was applied adjacent to the dorsal 

fin with a jab stick. Length of the jab stick's tag holding pin determined 

depth of penetration. The arrowhead anchored when the back pressure was 

applied to the streamer while removing the jab stick spread the flukes 

outward . 

Design Concept 

The pliable twin flukes formed the anchoring mechanism for this tag 

design. 

against the streamer. Once the tag penetrated to the required tissue depth, 

back pressure from jab stick withdrawal spread the flukes. 

point remained positioned medially in the tissue. 

As the flukes penetrated into the body tissue they were pressed 

The dart's 

3. Disc-Arrowhead Tag (Fig. 4) 

The disc-arrowhead tag had a pointed nylon head (16 mm) with two 

adjoining rigid flukes (8 mm), and a rigid shaft (22 mm). The shaft united 

-2- 
the flukes and a disc 2.54 cm in diameter. Application was made by incising the tissue with a scalpel and firmly pressing the tag into position by hand. 

Design Concept 

The disc was designed to rest on the epidermis and control the tag's 

depth of penetration. 

a highly visible streamer (not tested) while reducing tissue trauma 

caused by the streamer's flexion. The shaft length was intended to position the flukes in the blubber layer only to determine anchoring ability. 

It was also intended to act as a pivot point for 

4. Disc-Conehead Tag (Fig. 5) 

Fabricated from Delrin, the disc-conehead tag had a pointed, cone 

shaped anchor head (13 mm x 10 mm). 

Application procedures were the same as the disc-arrowhead tag. 

Oelrin is a biocompatible material. 

Design Concept 

This anchor head design was intended to separate rather than cut 

the tissue during application. It was hypothesized that the tissue's 

resilient nature may hold this anchor head design more securely. 

5. Carbon-Coated Disc-Conehead Taa 

This tag was structurally identical to the Disc-Conehead design. 

Application technique was also the same. 

carbon coated a1 1 surfaces. 

A low temperature isotrophic 

-3- 
Design Concept 

A low temperature isotrophic carbon coating was used on the tag's 

surface areas. 

La Jolla, California. This type of coating has successfully promoted 

tissue adherence and augmented surgical implant healing cycles in primates 

and humans. A magnified view of the carbon coating is shown in Fig. 6. 

The tags were coated by the Gulf General Atomic Corp., 

Results 

The tag evaluations are summarized in Table la, b. 

Appl ication 

The Floy spaghetti and arrowhead tags were implanted with a jab 

stick and presented no difficulties. 

minute. However, all three of the new tag designs were difficult to 

implant without incising the skin with a scalpel. 

area and rigid nature of their anchor heads may have been respons ble. 

application tool of specialized design was needed to provide suff cient 

force for penetration of the tag. 

tion was observed, particularly with the conehead design. 

Each tag was implanted within one 

The increased surface 

An 

Hemorrhaging subsequent to tag applicaDurabi 1 i ty 

No structural failure was observed with any of the tags. The carbon 

coating used on the disc-conehead tag was absent on the disc's ventral 

surface and at its junction with the shaft. 

-4- 
Vi s i bi 1 i ty 

The Floy tag streamers (fluorescent orange, yellow) contrasted well 

against the pig's black skin. No streamers were attached to the other 

tags, but the white nylon and delrin discs also contrasted well with the 

pig's skin color. 

Retention Time 

Retention times of tag group #l were as follows: 

1) 

2) 

3) 

4) 

The Floy spaghetti tag remained implanted during the study period. 

The Floy arrowhead tag was retained for 60 days. 

The disc-arrowhead tag was retained for 34 days. 

The disc-conehead tag (uncoated) was retained for 5 days. 

5) 

Retention time of the new tag designs (disc-arrowhead, disc-conehead, 

The carbon coated disc-conehead tag was retained for 5 days. 

carbon-coated disc conehead) were low when compared to the Floy spaghetti 

and arrowhead designs. The nylon disc-arrowhead tag design may have been 

retained longer due to its 22 mm shaft length which could have anchored 

in the fibrous fascia tissue. 

Tissue Trauma 

The pig was frequently observed to rub all the tags against it's 

pen wall, which subjected the tags and tissue to considerable abrasion. 

The tags' entry hole enlarged until shedding occurred. The Floy 

-5- 
spaghetti and arrowhead tags also displayed an erosion of tissue where 

the streamers entered the epidermis. Individual tag sites appeared to 

be healed within 30 days after the tag was shed and no infections or 

other complications were noted. 

Discussion 

Use of a pig for the initial tag implants proved to be important 

since the animal's rubbing behavior subjected all the tags and surrounding 

epidermal tissue to trauma similar to the water friction expected in a 

delphinid environment. Although retention times for disc-conehead, 

than the 

bil ity 

in two 

carbon-coated disc conehead, and disc arrowhead tags were lower 

Floy types, the absence of significant structural and b 

problems influenced our decision to implant them all as 

ocompat 

a group 

common dol phi n (Del phinus del phis) specimens. 

Tag Evaluation - Common Dolphin (Del phinus del phis) 

Tag evaluation experiments were conducted with two common dolphin 

(Delphinus delphis) specimens maintained in a 10.3 m x 13 m 240,000 liter 

pool. 

and aerial behavior, similar to Stenella sp. the tags were being developed 

for. 

tuna purse seine fishing. 

This species was selected since they display frequent rapid swimming 

They are also the third most important pelagic species taken during 

The animals were handled with careful attention 

to their health and safety. The holding pool was lowered prior to tagging 

and each animal was placed in a stretcher, then lifted by crane onto a soft 

- 6- 
foam rubber pad 

by tag implant. 

in the days fol 

The skin was scrubbed with Septodyne disinfectant followed 

Blood samples were taken before tagging and periodically 

owing to monitor health status. Water quality was checked 

on a daily basis. 

Tag Group I1 Common Dolphin (Delphinus delphis) 

Tag Description and Application 

The tag designs evaluated in the pig were implanted in two Delphinus 

delphis. Since blubber thickness of the Delphinus was similar to the pig 

(>18 mm) shaft lengths of the two arrowhead tags were 22 mm to extend the 

anchor head into the fascia. A streamer was added to one of the arrowhead 

tags to observe its influence on tag retention. 

The disc-conehead tags were fabricated with a 16 mm shaft to anchor 

in the blubber. One half the tag number were carbon coated for comparison 

of tissue reaction and healing cycle times with the uncoated tags. 

the tags were implanted using a plastic headed mallet (Fig. 7). 

All 

1. Disc-Arrowhead Tag (Fig. 4) 

Delrin was used instead of nylon. A yellow polyurethane streamer 

was attached to the disc. 

2. Carbon Coated Disc-Arrowhead Tag 

Low temperature isotropic carbon coating. No streamer was attached. 

-7 - 
3. Disc-Conehead Tag (Fig. 5) 

The shaft was 16 mm instead of 20 mm and nylon was used instead 

of delrin. No streamer was attached. 

4. Carbon Coated Disc-Conehead Tag 

Low temperature isotrophic carbon coating. No streamer was attached. 

Results 

The tag evaluation results are summarized in Table lb. 

Appl ication 

Application with the mallet was intended to improve tag penetration 

into the tissue, Each tag was implanted within one minute. Hemorrhaging 

was observed during the application of all tags and continued for approximately thirty minutes subsequent to tagging. The disc-arrowhead tags 

were implanted with no difficulty. However, the conehead tags required 

greater force to implant, and several tags were fractured during application. 

Conehead and arrowhead tag discs protruded above the epidermis within 

minutes after the animals were released, leaving a visible gap between the 

disc and the skin surface (Fig. 8). 

Durabil i ty 

No structural failure was observed with any of the tags. Carbon 

coated tags displayed areas where the carbon was removed, primarily on 

the disc's ventral surface at the junction of the disc and shaft. These 

-8- 
sites probably sustained the most significant mechanical trauma from the 

animal's body flexion, and water friction. 

Visibility 

The yellow polyurethane streamer attached to the uncoated discarrowhead tag was easily observed against the black skin of the Delphinus. 

The discs seated on the epidermis were visible due to their white color 

but their small size would limit useful viewing at a distance. Carbon 

coated discs could only be viewed with considerable effort at tankside. 

Retention Time 

Retention times of tag group #2 were as follows: 

1) Disc-arrowhead tag (uncoated) remained implanted for 4 days. 

2) Carbon coated disc-arrowhead tag remained implanted for 

14 days. 

3) Disc-conehead tag remained implanted for 6 days. 

4) 

Within two hours after application, the disc arrowhead tag (uncoated) 

Carbon coated disc-conehead tag remained implanted for 5 days. 

with streamer was protruding above the epidermis. 

observed pull ing on the streamer repeatedly. Its subsequent shedding 

(4 days) may be attributed in part, to this behavior. 

the uncoated conehead tag was also observed. 

to jump frequently after tagging and usually landed on the tag sites 

A companion animal was 

Disc protrusion of 

The Delphinus were observed 

when hitting the water (Fig. 9). Although aerial behavior is common with 

-9- 
this species at sea, these Delphinus had not been observed jumping prior 

to tag appl ication for several weeks. 

The Delphinus implanted with the disc-conehead tags was observed to 

rub against the tank wall in an apparent attempt to remove them. 

behavior appeared to influence retention time of the tags. 

This 

Once the tag 

discs protruded above the epidermis (Fig. 8), water friction was able to 

accelerate tag shedding. 

Tissue Trauma 

The mechanical trauma resulting from aerial and rubbing behavior, 

and pulling by mouth, appeared to retard completion of a healing cycle. 

The tag entry holes enlarged until they were larger than the anchor heads, 

and the tags were shed. 

once the tag discs projected above the epidermal surface. No infection 

of the tag sites was observed, but some swelling was noted during the 

period tags were implanted. Once the tags were shed, a healing cycle 

Water friction was also a significant factor, 

appeared to be complete within 23 days. 

Discussion 

Tissue adjacent to the dorsal fin of the Delphinus appears to be very 

resilient. Without a scalpel incision, the disc-conehead proved difficult 

to implant. Our rationale had been that the resilient nature of this 

tissue would serve to hold the tag in place. Since retention times for 

this tag type were the lowest of the group tested, its further evaluation 

was di sconti nued. 

-1 0- 
A streamer attached to the disc promotes visibility and prevents tissue abrasion where a constantly flexing lead or streamer may pass through 

the epidermis (i .e., spaghetti tag). 

behavior exhibited by a companion animal may have had a significant effect 

on retention times of this tag (4 days). 

with spaghetti tag streamers, but was not observed; "body slapping" was 

However, the "streamer pulling" 

This behavior may have occurred 

also observed throughout the time this tag group remained implanted. 

Carbon coatings were removed from several tag areas even though tag 

retention time was brief. Milled delrin and nylon appears to be too 

smooth for low temperature isotropic carbon to adhere well. Further use 

of low temperature isotropic carbon coating was discontinued to economize 

research time and funds. If carbon ooating is used for field tagging, 

the tag's dorsal surface should not be coated to permit better observation. 

A tag design that will withstand the mechanical trauma of rubbing, 

jumping, and pulling by mouth is necessary if field tag retention for 

at least one year is to be realized. 

healing cycle with tags implanted in the Delphinus due to the brief 

We have yet to observe a complete 

retention times. 

The rapid shedding of all prototype designs from the Delphinus indicated a need for further design improvements and subsequent testing with 

emphasis on reducing the disc size of prototype tags, revision of anchor 

head design, and continuation of Floy tag evaluation. 

common dolphin (Delphinus delphis) specimens were used as test subjects. 

Efforts were initiated to design a tag for application to the dorsal fin. 

Once again, two 

-11- 
%-Group #3 Common Dolphin (Delphinus delphis) 

Tag Description and Application 

The Floy tags were implanted with a jab stick and the disc-arrowhead 

and disc-conehead tags were implanted (after a scalpel incision) with a 

plastic headed mal 1 et. 

1. Floy Spaghetti Tag (Fig. 1) 

The Floy spaghetti tag design remained unchanged. 

2. Floy Arrowhead Tag (Fig. 2) 

The flukes were rounded and their length reduced by 50 percent 

(14 mm) for improved anchoring. 

3. Disc-Arrowhead Tag (Fig. 4) 

The disc size was reduced to a 25 mm x 15 mm oval. The shaft was 

hollowed to allow drainage of body fluids away from the wound. 

4. Adjustable Disc-Arrowhead Tag (Fig. 10) 

A Floy Co. pointed nylon barb (8 mm x 5 mm) with a single fluke 

(10 mm) was attached to a 90 mm flexible nylon conduit tie band. 

polycarbonate disc (25 mm diameter) with the wire tie locking mechanism 

bonded in its center was designed to thread the wire tie band material 

and lock in place. 

A flat 

-1 2- 
The tag head and flexible wire tie penetrated the tissue via a 

5 mm diameter trocar implanted to a predetermined tissue depth. Once 

the trocar was positioned, the tag head was threaded through it until 

the fluke cleared the opposite end. After removal of the trocar, the 

wire tie was pulled to anchor the fluke. 

the locking mechanism in the disc to "seat" it on the epidermis. Excess 

The wire tie was threaded through 

wire tie material was clipped off. 

5. Flag Tag (Fig. 11) 

This was the first dorsal fin tag design tested, ith two 80 mm x 5 mm 

orthoplast flags secured by a nylon nut and bolt. 

cored manually with a #8 trocar (Fig. 12) at a position intersected by 

the middle one-third of the fin measured from the dorsal tip and the first 

one-third measured from the anterior leading edge. A nylon sleeve (23 mm 

x 5 mm) was inserted in the hole prior to securing the flags on each side 

of the dorsal fin. Nylon is a biocompatible material. Once in place, the 

nylon nut and bolt were heat welded with a hot metal spatula. 

the sleeve and the nut and bolt head allowed 360' rotation of the flags. 

The dorsal fin was 

Space between 

Results 

The tag evaluation results are summarized in Tables la, b, c. 

Application 

No problems were encountered while implanting the Roy tags. Their 

application required only several seconds. However, the rigid shaft 

-13- 
disc-arrowhead tag was still difficult to implant even with a scalpel incision, prior to seating with a plastic headed mallet. If the tag was 

not struck on the disc's center, the shaft fractured or popped the entire 

tag out of the incision, prolonging application time. 

The adjustable disc-arrowhead tag was easily positioned but the fluke 

would not anchor in the tissue without considerable maneuvering. Application time was approximately 2.5 minutes. 

Coring a hole in the dorsal fin was not easily accomplished due to 

the extreme resilience of its connective tissue. 

tag application time (2 minutes). 

This prolonged Flag 

The dorsal fin must be held rigid and 

considerable leverage must be used to core this tissue. 

was noted but insertion of the sleeve in the hole controlled it effectively 

after about five minutes. 

Hemorrhaging 

Dura bi 1 i ty 

Only the Floy arrowhead and flag tags exhibited structural failure. 

One fluke of the Floy arrowhead tag evidently fractured during implant. 

After 28 days, one of the flag tag's flags fractured near the post. 

of water friction during the animal's respiration and aerial behavior 

caused a forward rotation of the flags and subsequent flexion when the 

animal returned below the water surface. Repeated action of this type 

caused fatiguing of the flag and finally fracture. 

Absence 

-14- 
Retention Time 

Retention times of tag group #3 were as follows: 

1) 

2) 

3) 

4) 

5) 

The Floy spaghetti tag was surgically removed after 31 days. 

The Floy arrowhead tag was surgically removed after 31 days. 

The disc-arrowhead tag was shed in 6 days. 

The adjustable disc-arrowhead tag was shed in 1 day. 

The flag tag was intact for 43 days until the animal's 

death from causes not related to the tag. 

Tissue Trauma 

The Floy spaghetti and arrowhead tags were removed since raised 

areas developed around each tag and blood sample analysis indicated potential infection. 

tinued penetration into the muscle tissue, as shown by a change in position of the script on the streamer (Fig. 13 a, b). Removal of this tag 

revealed that one fluke had fractured, possibly during implant. 

abcess was present. The Floy spaghetti tag was found to have anchored well 

Photographs of the Floy arrowhead tag indicated a conAn 

in the fascia. The barb was encapsulated and no infection was present. 

Both disc-arrowhead tags were shed rapidly, again indicating that 

water friction against the disc did not allow completion of a healing 

cycle. 

have been a significant factor in the shedding of these tags. 

Rubbing and jumping behavior was frequently observed, and may also 

The flag tag was removed only because the Delphinus specimen expired 

of causes not related to the tag. Sectioning of the post hole revealed 

-15- 
I 

that a healing cycle had been progressing well, with reepithelialization 

noted adjacent to the sleeve location (Fig. 14). 

Di scussi on 

Although the Floy tag depth of penetration should be limited by the jab 

stick shaft length, the arrowhead type which fractured appeared to migrate 

medially (Fig. 13a, b). 

tag anchored accounted for continued penetration of the barb. 

It is possible that muscle layer flexion where the 

Testing of the Floy arrowhead tag was discontinued in favor of the Floy 

spaghetti tag. Reduction of tag migration after application, and abrasion at 

the entry site were important design criteria to be considered. 

The disc-arrowhead tag, despite design changes in the disc and shaft, 

was still difficult to implant and retention time was low. Further evaluation 

of this tag type was discontinued. 

Testing of the adjustable disc-arrowhead design was discontinued, 

atthough reducing the diameter of the trocar and improving the method for 

advancing the barb beyond the trocar may improve anchoring and reduce the time 

needed for tag implant. 

With the exception of the Floy spaghetti tag, the flag tag displayed 

longest retention time and also the most complete healing cycle. 

nylon sleeve appeared to minimize tissue trauma encountered with other tag 

designs. 

Use of the 

Selection of a more durable material was of primary importance for 

further testing of this tag type. 

-16- 
Tag Group #4 Common Dolphin (Delphinus delphis) 

New tag designs developed by the National Fisheries Engineering 

Laboratory, Bay St. Louis, Mississippi, were tested on two Delphinus specimens. All of the tags were fabricated from biocompatible materials and 

could be applied in less than one minute, without use of tools. Three 

of these tags were designed for application to the dorsal fin since the 

most significant retention time (43 days) to date was recorded for a 

dorsal fin tag. 

Taa DescriPtion and ADD1 ication 

1. Anterior Dorsal Fin Tag with Wrap-around Streamer (Fig. 15) 

Spring-loaded stainless steel tines were intended to hold the flag 

to the anterior edge of the dorsal fin. 

pierces the fin is approximately 2.5 cm posterior to the anterior edge. 

The flag was a continuous piece of hypalon that wrapped around the lower 

anterior aspect of the dorsal fin. It was connected to the clasp by a 

nylon fastener. 

The element of the clasp that 

Design Concept 

The thin spring-loaded stainless steel tines were intended to hold 

the flags against the dorsal fin's leading edge, aided by water flow over 

the flags. The wrap-around flags were thin and pliable to minimize 

-17- 
mechanical trauma of the dorsal fin. The spring placement and sharp tines 

allowed rapid tag application. 

2. Metal Scoop Tag (Fig. 16) 

The anchoring devices were M-shaped type 316 stainless steel wire 

tines with a 14 cm hypalon streamer fastened at the center angle with a 

nylon rivet. 

penetrate the tissue = 2 cm, just posterior to the dorsal fin. 

flow from the forward motion of the animal was intended to hold the 

anchored tines in position. 

The 3 cm tines were curved downward in a semi-circle to 

Water 

Design Concept 

This tag's tines were shaped in downward curved semicircles so that 

once implanted, water flow would augment their anchoring by forcing them 

"into" the tissue. 

3. Anterior Dorsal Fin Tag with Side-mounted Streamers (Fig. 17) 

Same clasp as tag #l. Streamers were attached to the clasp on each 

side of the leading edge without wrapping around the anterior aspect of 

the dorsal fin as in tag #l. 

Design Concept 

The flag placement was changed to observe water flow effects as 

compared to the wrap-around flags. 

-1 8- 
4. Posterior Dorsal Fin Spring Clip Tag (Fig. 18) 

Stainless steel spring-loaded tines (similar to tag #1 and 3) with 

the tines piercing the fin 3 cm anterior to the dorsal fin's trailing 

edge. No streamer was attached to this tag. 

Design Concept 

Positioning was adjacent to the dorsal fin to observe water flow 

effects as compared to tags #I and 3. 

Results 

The tag evaluation results are summarized in Table Id. 

Application 

Each of the tags were applied within thirty seconds. Considerable 

force was needed to penetrate the dorsal fin's connective tissue even 

though the thin stainless steel wire clasps were sharpened and spring 

loaded. No tools were required for application. 

Durabi 1 i ty 

The stainless steel wire anchoring mechanisms remained structurally 

intact. However, all the flags used on the dorsal fin applications were 

shed within three days, indicating a need for improvement of their attachment mechanisms. 

Visibility 

Whi7e intact the streamers attached to the dorsal fin were highly 

visible. Yellow appeared to be the color of choice for future evaluation 

of these design types. 

-19- 
Retention Times 

All tags applied to the dorsal fin were removed three to seven days 

after application. 

application. 

The scoop tag was shed within several hours after 

Tissue In jury 

Constant inward pressure produced by the spring-loaded tines of the 

dorsal fin tags combined with water friction against thin wire 

significant tissue abrasion within several days after appl ication. 

tags were removed to prevent further tissue trauma and insure the animal's 

health. 

promoted 

The 

Discussion 

The primary advantage of the tags was their relative ease of application without tools, and the visibility of dorsal fin streamers. Since 

significant reepithelialization of dorsal fin tissue was observed with a 

sleeved anchor pin in tests, this application technique should be evaluated with these tag designs. 

diameter of the stainless steel wire, which may serve to distribute water 

flow forces more evenly against the tissue. 

An alternative would be to increase the 

Insertion and Withdrawal Tests of Tag Head Designs 

Tags designed for implant in the body exhibited low retention times 

(with the exception of the spaghetti tag). Nore precise data was needed 

-20- 
to determine the relationship between tag head design, tissue morphology 

and tag retention. 

to measure the pressures (lbs/in ) required for insertion and withdrawal. 

Tests were conducted with specially designed tag heads 

2 

Methods and Materials 

Insertion and withdrawal pressures were measured for eight specially 

designed tag heads and two spaghetti tag heads (Fig. 19a). 

were inserted and withdrawn from the dorso-lateral tissue of a thawed and 

a fresh Delphinus delphis carcass. 

spring scale were used to measure insertion and withdrawal pressures in 

lbs/in2 (Fig. 79b). 

These tag heads 

A specially designed tag head chuck and 

The tag head chuck allowed penetration through muscle and fascia 

tissue, and could be adjusted for blubber penetration only. 

insertions and withdrawals were made for a tag head in each category and 

At least three 

the x of each test group was recorded. Surgical removal of some tag heads 

after insertion was also conducted to observe their positions in the tissue. 

Results 

Results for the insertion and w 

Table 2. 

2 The 1 bs/in values are higher 

thdrawa 

for tag 

tests are summarized in 

head insertion and withdrawal 

through muscle-fascia tissue when compared to blubber tissue measure values. 

The freezing and thawing process appeared to impart a change in the tissue's 

integrity since thawed tissue pressure values were also higher. 

-21- 
Tag heads with low insertion and high withdrawal pressure values had 

a fluked or barbed head design with less mass. 

examples were the spaghetti tag designs. The modified spaghetti tag design 

was more difficult to insert and neither spaghetti tag design could be 

withdrawn. 

The most significant 

Surgical removal of tag heads revealed that cone head designs often 

caused enough physical tissue trauma during insertion to allow easier 

withdrawal, especially if inserted only into the blubber tissue. 

spaghetti tags with rounded flukes usually caused less tissue trauma and 

often anchored in the fibrous fascia tissue (Figs. 20-21). 

The 

Discussion 

Additional insertion and withdrawal tests should be conducted for 

evaluation of tag head designs. Fresh carcasses should be used for an 

evaluation of this type since the freezing and thawing process (in Delphinus 

tissue) tends to produce distorted 1 bs/in2 values. 

The muscle-fascia layer appears to promote higher withdrawal pressure 

values which would contribute to longer tag retention time. 

removal of tag heads related that some involvement of muscle tissue was 

usually observed after insertion since the fascia layer is very thin (compared 

to the blubber and muscle layers). 

physical trauma will affect healing cycle time. 

However, surgical 

The extent of this involvement and the 

-22- 
Insertion and surgical removal of spaghetti tags provides a good 

example. The Standard Floy tags three-fluked barb was easier to insert 

and rotate but the sharp points cause more tissue trauma. 

design may prolong the healing cycle. 

rounded off) was recommended for further live animal testing. 

This barb 

A three-fluked barb (fluke points 

Tag Group #5 Pacific White-sided Dolphin (Lagenorhynchus ob1 iquidens) 

The dorsal fin represented the most visible area of the body to tag 

and an encouraging healing cycle and retention time had been establ ished 

with the flag tag design. 

healing cycles or retention times with the exception of the Floy spaghetti tag design. Therefore, emphasis was placed on a tag design for the 

dorsal fin (disc tag) and improvement of the Floy spaghetti tag design. 

The peduncle tag, a new design concept, was also evaluated. 

Implanting body tags had not shown significant 

Tag Description and Application 

1. Dorsal Fin Disc Tag (Fig. 22) 

This dome-shaped disc tag was designed by the National Fisheries 

Engineering Laboratory. 

holes and radial and perimeter slits. 

of a teflon multi-notched pin on one disc's concave side which inserted into a 

ridged tube on the concave side of the other disc. 

Each teflon disc was 5 cm in diameter with vent 

The self-locking anchor pin consisted 

-23- 
This disc tag was applied by cutting a hole through the dorsal fin, 

slightly smaller than the anchor pin, with a scalpel. 

hole was not cauterized. 

placed in the hole and snapped together. 

The anchor pin 

The anchor pin sections of both discs were 

Design Concept 

The disc's dome-shaped vent holes, and slits were intended to channel 

water flow between the discs and the dorsal fin. They functioned to keep 

the disc edge from abrading the tissue. 

facilitated rapid application by snapping together when placed in the dorsal 

The tlmale-female" anchor pin 

fin hole. The teflon, a biocompatible material, had a smooth surface to 

curtail fouling growth. 

2. Spaghetti Tag - Modified Lead and Anchor Head (Fig. 23) 

A 30 x 10 x 1 mm type 316 stainless steel anchor head was attached 

to a 1.5 mm diameter type 316 stainless steel wire lead. A 21 x 1.27 cm 

polyurethane streamer was attached to the wire lead by a chrome plated 

brass McMahon fishing swivel (Fig. 24). The tag was applied with a jab 

stick. 

Design Concept 

This tag was designed to minimize stress on the lead and anchor head 

by means of a pivot point for the streamer. The streamer shape was enlarged and flattened to improve visibility and to observe its hydrodynamic 

characteristics. 

- 24- 
3. Snap Fastener Peduncle Belt (Fig. 25) 

The peduncle tag was a 21 x 10 cm band of thin soft red synthetic 

rubber with a polypropylene male-female fastener at each end. The belt 

was applied around the peduncle, at the insertion of the tail flukes. 

The male-female fasteners were over1 apped and pressed together to secure 

the belt in place. 

Desi gn Concept 

The thin soft synthetic rubber belt iras intended to minimize tissue 

abrasion by flexing with the peduncle and flukes as the aniriial moved 

through the water. The polypropylene mal e-feinale fasteners could be 

secured rapidly and allowed adjustment for variance in peduncle yirth. 

Results 

The tag evaluation results are summarized in tables la, c, and e. 

Application 

Coring the dorsal fin with a scalpel for placement of the disc tag 

was accomplished with no difficulty in about 30 seconds. Caution in 

incising the connective tissue must be exercised since the scalpel blade 

is thin and rather fragile. 

Considerable force was needed to penetrate the tissue and seat the 

new spaghetti tag. The first application attempt did not penetrate the 

anchor head deep enough into the tissue or toggle it sufficiently to 

prevent withdrawal. The second attempt was successful requi ri ng several 

seconds. A sharper point and edye for the anchor head of this tay should 

be used. 

-25- 
The peduncle tag was appl ied in a matter of seconds with .no 

pro bl ems. 

Dura bi 1 i ty 

The self-locking anchor pin of the dorsal fin disc tag failed due 

to the soft nature of the virgin teflon. The spaghetti tags swivel corroded and finally fractured, shedding the streamer after twenty-three 

days. The anchor head remained implanted. The polypropylene fasteners 

used on the peduncle tag were also too soft to withstand water friction 

and rapidly pulled loose. 

Visibility 

Of the three tags evaluated, the dorsal fin disc tag was most visible above and below the water surface. The spaghetti tag could only be 

observed clearly above the water surface during respiration. 

tag was also difficult to observe since it rarely was exposed above the 

water surface. 

visi bil i ty. 

The peduncle 

The red and white color of this tag di'd not augment 

Retention Time 

Retention times of tag group #5 were as follows: 

1) The dorsal fin disc tag was shed in less than 1 day. 

2) The spaghetti tag with modified lead and anchor head 

remained implanted during the study period, a1 though the 

streamer needed replacement. 

-26- 
3) The peduncle tag was shed in less than one day. 

Tissue Trauma 

The dorsal fin disc tag and peduncle tag were shed in less than one 

day, causing no tissue injury. A blood vessel ruptured during anchor pin 

hole coring continued to hemorrhage for approximately twenty minutes 

after the animal was returned to the pool. The modified spayhetti tag 

displayed minimal tissue abrasion at the barb's entry site (Fig. 24). 

This abrasion appeared to heal within eighteen days. No additional 

tissue trauma problems have been noted with the spaghetti tag. 

Discussion 

Coring post holes for dorsal fin disc tays in the .field with scalpel 

blades is not advisable since they are fragile and dull quickly. Ulood 

vessels in the dorsal fin's arterial tree inay be ruptured by coring a 

post hole for dorsal fin tags. Prolonged hemorrhaging could attract 

sharks in a field tagging situation. Tissue constriction by using a 

slightly laryer post and chemical cantery can control this blood flow. 

Short retention times for the dorsal fin disc tag and peduncle tag 

precluded a thorouyh evaluation. More durable inaterials for locking 

mechanisms kiere def i ni tely needed to continue eval uat ion of these tag 

designs. The rigid lead eiire and swiveling pivot point of the modified 

spaghetti tag appeared to effectively reduce tissue trauma and promote a 

complete healiny cycle. However, the streamer was shed due to swivel 

corrosion and was replaced with a barrel swivel (brass) which also 

-27- 
corroded and fractured after thi rty-six days. The streamer was final ly 

attached to the lead with a type 316 stainless wire loop and has remained 

intact during the study period. 

Tag Group #6 Pacific White-sided Dolphin (Lagenorhynchus obliquidens) 

Since the spaghetti tag remained intact (with the exception of the 

streamer) and no bi ocornpat i bi 1 i ty problems were observed, cmphasi s for 

tag yroup #6 centered on further development of the dorsal fin disc tag 

and the peduncle tag. 

Tag Descri pt i on and Appl i cat i on 

1. Dorsal Fin Disc Tag (Fig. 22) 

This dorsal fin disc tag was identical to the design used in tag 

yroup #5 except for the anchor pin. The anchor pin of this tag was a G 

nim diameter smooth nylon rod, secured with a zinc coated steel retainer 

riny. Additional security was provided by heat welding the rod tip with 

a hot spatula. 

Uesi yn Concept 

Nylon is biocompatible and more rigid than teflon. It was 

hypothesized that a nylon anchor pin would maintain its structural 

inteyrity under stress froin water flow or the animal's aerial behavior. 

The new anchor pin design alloided adjustments for variance in dorsal fin 

thickness. 

- 28- 
2. Velcro Rubber Peduncle Belt (Fig. 26) 

This peduncle tag was fabricated from a 21 cm x 4 cm band of soft 

synthetic rubber. A 37 cm x 2 cm x 1.5 mm strip of Velcro was sewn to 

the rubber band with nylon thread. 

to secure the rubber band around the animal's peduncle. 

The Velcro functioned as a fastener 

Design Concept 

Velcro (trademark) is a soft non-corrosive nylon fastener material. 

Its soft flexible nature and holding ability were intended to improve the 

tag retention time. Velcro is also a manufactured item available in 

quanti ty . 

3. Ty Rap Rubber Peduncle Belt (Fig. 27) 

Thjs peduncle tag was fabricated from a 19 x 8 x 1.5 mm band of 

soft synthetic rubber. 

was sewn to the rubber section with nylon thread. 

a locking device, holding the rubber band in place on the animal's 

A Thomas and Betts polyethylene Ty Rap wire tie 

The tie functioned as 

peduncle. 

end which contains a tine that seats between the teeth. The threaded 

belt is folded over and secured under two posts giving this tag two 

locking mechanisms. 

The toothed belt is threaded through a slot on the opposite 

Design Concept 

Ty Raps (trademark) are commonly used to bundle together heavy objects such as cable and pipe. Its locking mechanism is strong, adjustable 

-29- 
and non-corrosive. It is also a manufactured item available in 

quantity. 

4. Nasco Multi-loc Cattle Leg Belt (Fig. 28) 

A 34 x 3.2 an x 3 nun polypropylene cattle leg tag. 

locking mechanism similar in design to the Ty Rap (wire tie). 

This tag had a 

Adjustments could be made on the lockiny end of the band at 6 mm 

intervals. 

Desiqn ConcePt 

Once again the emphasis was on a strong and durable locking 

mechanism for the belt. This locking mechanism is similar in design to 

the Ty Rap wire tie except the teeth are only located on one end of the 

belt, and the belt is not folded over. The tooth and tine arrangement is 

the only means of locking. This tag has been successfully used on cattle 

and can withstand considerable stress without fatigue. 

Results 

The tag evaluat 

Appl i cat i on 

on results are summarized n Tables IC, and e. 

The disc tag was appl ied in the same anchor pin hole used for the 

tag group #5 disc tag. The peduncle tags were easily applied in a few 

seconds. 

to miniinize epidermal trauma. Approximately 1-3 mn of clearance between 

the epidermis and tag was allowed for both tag types. 

There was some question about the need for a loose or tight fit 

-30- 
Durabil i ty 

After 80 days, sections of the disc tag on one side of the dorsal fin 

began to fracture and shed until only the portion around the anchor pin 

was left (Fig. 29). The zinc-coated steel retainer ring was badly corroded when removed at 100 days. Although no structural change occurred 

with the Velcro peduncle belt, continuous water flow caused the Velcro 

mesh to separate, and the tag was shed. The Ty Rap and Cattle Leg peduncle 

belts did not exhibit any structural integrity problems. 

Vi si bi 1 i ty 

Although the white teflon disc tag contrasted well with the dorsal 

fin's black skin, algae growth was beginning to establish itself on surfaces of both discs when the tag was removed. Continued growth would have 

significantly reduced visibility of the discs. The red and black colors 

of the Velcro and Ty Rap peduncle belts were not easily observed above or 

below the water surface. However, the white color of the cattle leg tag 

contrasted well with the black skin of the Lagenorhynchus. 

Retention Time 

Retention times for tag group #6 were as follows: 

1) The dorsal fin disc tag was removed from the dorsal fin 

after 100 days to allow implant of a new disc tag design. 

2) The Velcro peduncle belt was shed in one day. 

-31 - 
3) 

4) 

The Nasco cattle leg belt was removed after five days. 

The Ty Rap peduncle belt was removed after five days. 

Ti ssu e Trauma 

An indented ring of darkened tissue around the disc tag's perimeter 

was observed (Fig. 30), but no apparent tissue damage was noted and the 

tag was left in place. No tissue injury was observed with the Velcro 

peduncle belt. The cattle 1 eg and Ty Rap peduncle belts were removed 

after five days due to significant tissue abrasion at the junction of the 

peduncle and tail flukes (Fig. 31). 

Discussion 

Use of a smooth nylon post with retainer ring for the dorsal fin disc 

tag prolonged retention time and reduced tissue injury. Further study was 

needed of more durable materials for this tag design. 

A1 though the locking mechanism of bo peduncle belt designs improved 

retention times, tissue abrasion prompted removal to safeguard the test 

animal's health. 

or flat, wide, and soft (Ty Rap rubber belt). 

these designs seemed to press the belt material down against the epidermis 

and then lift it upward to create a "grating" action. A very soft, round 

material may "roll" with the body flexion, and distribute the pressure of 

the belt on the epidermis. 

The belt materials were flat and rigid (cattle leg belt) 

Water flow acting on both 

-32- 
Tag Group #7 Hawaiian Spinner (Stenella longirostris) 

Hawaiian Spinner Tag Tests - Spring 1978 

During the week of March 15-19, 1978, further evaluation of the dorsal 

fin disc tag was conducted at Sea Life Park in Hawaii using Spinner porpoise 

(Stenella longirostris). The spinning behavior of Stenella longirostris 

and availability of a large 45 m x 45 m 680,000 liter pool provided ideal 

conditions for evaluating the dorsal fin disc tag design. The initial 

tagging experiment involved six animals. Safe and humane procedures for 

handling the animals were followed. The water level was lowered to allow 

placement of an animal into a stand-supported stretcher in the pool. The 

dorsal fin was washed with Septodyne before coring was attempted. 

tags were secured in place and the animal was returned to the pool. 

The 

Tag Description and Application 

Modified Disc Tag 

The disc tag used in this experiment is described in experiment #5 

(Fig. 22). Tag application technique was slightly modified from previous 

experiments. A 7 mm hole was cored through the fin. The coring tool was 

notched corresponding to the various teflon post and sleeve lengths. 

appropriate sleeve was mounted on the post of the male tag section, the 

post inserted in the protruding coring tool, and pushed back through the 

hole. 

it served as a compress within the wound to curtail blood flow. The female 

The 

Since the sleeve was slightly larger than the hole (7 mrn vs. 6 mm) 

-33- 
side was then affixed to the post and the protruding post material 

removed with wire cutters. 

Design Concept 

In an attempt to eliminate previous disc tag application problems 

the male tag disc section was modified to accomodate different post and 

sleeve lengths and the coring tool as described. 

Results 

The tag evaluation results are suirimarized in Tables la, c. 

Tay Application 

The tacjs required much longer to apply than expected (>1 minute). 

Since the sleeve was sliyhtly laryer than the post and cored hole, some 

difficulty was encountered in its positioning. Each tag was individually 

iiiachined, causing some variability in the male and fernale sections. Soine 

holes in the fernale disc section would not fit over the post. 

The teflon post was observed to bend when pressure necessary to 

affix the fernale section was applied. After the post and sleeve were 

inserted through the hole, it was necessary to cut back the post by 

depending on dorsal fin thickness. Wire cutters used to trim the post 

flattened the soft teflon, rriaking it difficult to attach the female section. Sharper snips and a harder tag material such as polyurethane 

should eliminate these problems. 

Durabi 1 i ty 

Spinning and re1 ated behavior caused repeated tag structural fai 1 - 

ures soon after application. Problem areas were determined to be the post, 

-34- 
which may have fractured in two, and the discs which fractured in segments 

or everted entirely (Figs. 32 and 33). 

caused male and female tag sections to pull apart. 

The teflon's pliable nature also 

Retention Time 

A summary of tag retention times is presented in Table 3, None of 

the tags were retained for longer than a four-day period. 

Tissue Trauma 

The apparent mechanical trauma from water friction on the tag post 

caused erosion and softening of the surrounding tissue. 

was observed as well as necrotic tissue and lesions (Fig. 34). 

Some inflamation 

The presence 

of one or more of these conditions on all the animals necessitated removal 

of tags remaining intact after four days. 

Discussion 

The problems encountered with structural integrity, and to1 erances 

between sleeves, posts and discs, suggested that injection molding of the 

disc tag's components may improve the tag's application, and retention time 

by rendering them more durable and consistent in dimension. 

Considerable time was needed to core the dorsal fin. Since the 

dorsal fin is flexible, a small piece of wood placed on the side opposite 

the coring tool's entry should aid the coring process and sleeve positioning. 

A larger coring tool handle for more leverage, with a sharp cutting edge, 

is also desirable. 

-35- 
A harder more durable material is needed to prevent the disc fracture 

and eversion observed in these tests. Flow tank testing of materials at 

flow rates relative to the swimming behavior of Stenella (20-30 f.p.s.) 

were needed to further develop tags before continuing live animal tests. 

Tag Tests - Utah State Water Research Laboratory 

It was difficult to accurately observe tag design performance (i.e., 

structural failure) while the tag was affixed to the animal. This was 

due to the animal's agil4ty and physical factors such as light reflection 

off the water. It was necessary to test old and new tag designs in a 

"laboratory" situation before continuing live animal tests, and utilize 

the data to improve tag designs prior to a second tagging effort with the 

Stenella at Sea Life Park. 

Methods and Materials 

Flow tank tests were conducted at the Utah State Water Research 

Laboratory in Logan, Utah. Selected tag designs were observed in a 

specially constructed 35.6 x 30.4 x 35.6 cm viewing chamber (Fig. 35). 

A Stenella sp. dorsal fin was secured to a metal mount within the 

chamber. Tags representing four different categories (spaghetti tags, 

disc tags, flag tags, and cattle ear tags) were attached to the dorsal 

fin and subjected to pump-induced varied water flow rates of up to 30 ft ./ 

second at different mount angles. 

Mitchell Monitor 10 movie camera at 24 f.p.s. 

Each test was recorded on film, with a 

The contact of tag surfaces 

-36- 
with the skin and any changes in skin appearance were used as criteria 

for recording observations. 

Results 

Flow tank test data is presented in Table 4. Observations on biocompatibility factors are as follows: 

Disc Tags - Polyurethane, polypropylene, and delrin tags were fitted to 

the dorsal fin using a nylon sleeve and screw-type anchor pin. The flow 

tank tests revealed several important physical characteristics of the 

dorsal fin, and tag design factors which may influence tag biocompatibility: 

1. The dorsal fin flexed to the right or left, changing the 

position of the disc relative to the skin surface, as the 

water flow rates increased (Fig. 36). 

2. This change in dorsal fin curvature allowed the water flow 

to bend or tear a thin-walled, flexible disc tag. 

A thick-walled, non-flexing disc tag chafed the skin at the 

top and bottom of the disc as the water flow rate increased. 

3. 

The edges of all disc tags tested were thin. 

(thick-walled type) discs with equally-spaced holes were found to be the 

Since delrin and polyethylene 

most durabl e, mod 

to distribute the 

cal trauma to the 

fying the edges of tags fabricated from these materials 

discs contact surface with the epidermis may minimize mechaniskin. 

Flag Tags - 

a tendency to contact the dorsal fin during flow tank testing. This 

The dorso-anterior margin of the flag tags tested displayed 

-37- 
contact involved flexion of the dorsal fin, the individual flags, and increased as a function of water flow and dorsal fin flexion. A new flag 

design was fabricated at the test site with double sleeves and anchor 

pins. The sleeves projected an additional 3 mm beyond the dorsal fin on 

each side. 

except at the posterior edge of the dorsal fin. 

possibility of flag rotation forward when the dorsal is out of the water 

(i.e., respiration or aerial behavior). 

This design reduced flag flexion and contact with the skin, 

It also eliminated the 

Spaghetti Tags - A Floy spaghetti tag and the improved design with modified 

lead and barb, were each implanted in the dorsal fin. The streamers of 

both tags appeared to lay flat against the dorsal surface, but there was 

flexion observed. No significant differences between the tag types at 

flow rates up to 30 ft./sec were observed. 

However, there was no way of approximating the animal's respiration behavior. The alternating presence and absence of water friction on the 

streamer during respiration or aerial behavior may account for its spiralTissue damage was not evident. 

ing motion observed in live animal tests and subsequent tissue erosion 

at the skin surface. The improved spaghetti tag version now has a 360" 

pivot point where the streamer is attached, which appears to have eliminated 

tissue erosion on the Lagenorhynchus test subject. 

Cattle Ear Tags - The cattle ear tag tested, appeared to contact the skin 

unless positioned toward the anterior margin of the dorsal fin, allowing 

the flags to project beyond a point of contact. We were able to test a 

- 38- 
flexible flat type and no significant change in skin appearance at flow 

rates up to 30 ft./sec was observed. 

Flexion of the dorsal fin relative to the tag was easily observed 

and photographed during the flow tank tests (Fig. 36). 

in combination with water friction, had a direct influence on tag biocompatibility and retention. The dorsal fin used in these tests had been 

preserved in formalin, However, we feel the flexibility of this dorsal 

fin compares favorably with those of live animals previously examined. 

(Manual flexion of dorsal fins and observation of swiming Stenella 

through Sea Life Park's underwater view ports.) 

This flexion, 

High speed movie film 

of tagged Stenella through these view ports would allow a more accurate 

comparison. 

Tag Group #8 - Hawaiian Spinner (Stenella longirostris) 

During the week of June 11-18, 1978, additional tagging tests were 

conducted at Sea Life Park, Oahu, Hawaii, on the six Hawaiian spinner 

porpoise (Stenella longirostris) tagged previously. Due to infection 

problems encountered during the previous tag evaluations with these animals, 

copper sulfate was injected into the pool water and antibiotics were administered to the animals in their food. Blood analysis for each animal was 

regularly performed and water quality was monitored daily. 

Tag Description and Application 

1. Dorsal Fin Disc Tag (Fig. 37) 

-39- 
On the basis of flow tank tests conducted at Utah State University, 

three types of material were used for the 7 cm diameter disc tags: 

polyurethane (red), high density polyethylene (orange) , and delrin (blue). 

These discs differed from the original teflon disc tags. They were molded 

to different thicknesses, disc slits were eliminated, and the number of 

holes was reduced from six to four. Disc edges were smooth and rounded. 

The polyethylene and polyurethane discs were applied using a notched 

post of delrin (a biocompatible plastic) which was cored to allow for 

insertion of the applicator tool. 

enough to allow for passage of the post through the locking section of the 

- The delrin discs were not pliable 

on sleeve. The 

nut was used to 

thout sleeves to 

disc so a 316 stainless steel bolt was used with a tef 

locking section was cut away and a 316 stainless steel 

lock the disc in place. The posts were used withand d 

determine sleeve effectiveness. 

2. Spaghetti Tags (Figs. 1, 23) 

Two types of spaghetti tags were tested. The standard orange Floy 

spaghetti tag with a 304 stainless steel anchor head and a monofilament 

lead, and a modified spaghetti tag with a 316 stainless steel dart, 316 

stainless steel lead wire extended from the anchor head to a stainless 

steel swivel which was attached to a polyurethane streamer. 

head of the experimental tag was wider, thicker, shorter, and blunter than 

The anchor 

the standard Floy type. 

Earlier investigations have indicated that 316 stainless steel is 

preferable as an implant material to 304 stainless or flexible plastic 

-40- 
monofilament. 

continuous movement of the streamer, we have tested various types of leads 

and swivels. 

on a Lagenorhynchus at Hubbs/Sea World Research Institute. Since the animals 

could be easily approached and handled, a 15.24 cm hand-held jab stick was 

used for tag applications. 

In order to minimize mechanical trauma to the wound caused by 

The one used on this test animal group was successfully tested 

3. Multi-post Flag Tag (Fig. 38) 

The flags were fabricated of Kydex, measuring about 10 cm x 6 cm. The 

forward edge was secured to the fin by two sleeved nylon bolts, nuts, and 

sleeves to prevent the tag from pivoting about a single post as was observed 

in earlier flow tank and captive dolphin tests. 

pivot, it can become oriented in such a way that the flags are forward of the 

post. The oncoming water, rather than flipping the flags back, can spread 

them apart resulting in considerable pressure on the post and often breakage 

of the flags or post. 

If the tag is allowed to 

Two models of the flags were tried: a tag with the 

th the trailing edges bolted open with 

1.4 cm (Fig. 39). The rationale for 

bolting the tag open was to assure continuous water flow under the tag and to 

prevent abrasion of the trailing edge of the fin. 

to ride about 0.25 cm out from the fin on both sides. 

a fluorescent yellow color for improved visibility. 

Both models were designed 

The flags were painted 

Tag application involved placement of a flag against the dorsal fin 

as a template to mark positions of the post holes. The coring tool was 

designed with a door knob type handle for improved leverage and used in 

trai 1 

nylon 

ng edges unsecured, and one w 

bolts, nuts and sleeves about 

-41 - 
conjunction with a small block of wood placed on the opposite side of the 

dorsal fin to reduce flexibility. 

4. Suraical Tubina Peduncle Taa 

A new peduncle tag was evaluated, as shown in Fig. 40. It was 

fabricated from rubber surgical tubing (1.2 cm width) with a nylon wire 

tie inserted inside to secure the tag around the peduncle. The surgical 

tub ng's rounded surface and flexibility were considered important for 

min mizing skin abrasion. 

for visibility. 

It was coated with orange latex rubber paint 

Results 

A summary of test results for the Hawaiian spinners is presented in 

Tables la, c, and e. 

Application 

The posts for the disc tags were barely long enough to engage the 

locking section of the opposing disc. 

Utah State University revealed that the tags had to be snugly fitted to 

prevent water friction from fracturing or everting the disc. We wanted 

to determine the degree to which the dolphin's fin could accommodate 

pressure around the periphery of the disc. The dolphin's fin apparently 

flexes also causing pressure to be applied at the top and bottom of the 

tag. 

Results of the flow tank tests at 

The tags were generally not flush to the epidermis due to the curva 

ture of the fin. 

-42- 
Application of the spaghetti tags were not well executed. Lack of 

pressure behind the jab stick did not allow the anchor heads to rotate 

properly. The experimental spaghetti tags placed anterior to the dorsal fin 

on the right side made a larger than expected insertion hole because the 

animal moved during application. The standard type spaghetti tags placed 

adjacent to the fin on the left side did not penetrate well (the barbs were 

showing) and were further inserted by hand. 

Durabi 1 i ty 

A summary of tag durability is presented in Table la, c, and e. 

Durability problems with disc tags were attributed to fracture of 

delrin posts (4), and polyurethane discs (2) (Fig. 41). The mult 

tags (2) eventually fractured adjacent to the posts (Fig. 42). D 

unsl eeved 

-post flag 

sc and flag 

tags tested displayed varying degrees of fouling by algae. The flag tag's 

fouling was extensive enough to obscure its number (Fig. 43). 

No structural failures were noted with the spaghetti or peduncle tags. 

The orange latex paint on the peduncle tag was observed to peel within two 

days. The spaghetti and peduncle tags were not retained for a long enough 

period of time to evaluate durability. 

Visibility 

Of the three disc tag colors (blue, red, and orange) orange displayed 

the highest visibility above and below the water surface. 

fluorescent orange streamers of the spaghetti tags were visible, but their 

relatively small surface area 1 imits observation, especially underwater 

The yellow and 

-43- 
(Fig. 44). Although placed adjacent to the dorsal fin, these tags -rarely 

were exposed above the water surface. 

The large surface area and yellow color of the flag tags displayed the 

highest visibility of all tags tested, above and below the water surface 

(Fig. 39). 

above the water surface displayed minimal visibility. 

The peduncle tags, with reduced surface area, and rare exposure 

Retention Ti me 

Retention times for all tags tested are shown in Table la, c, and e. 

The delrin disc tag with sleeved stainless steel post and the rear posted 

Kydex flag tag were retained intact for the longest period of time (58 days). 

A modified spaghetti tag was retained for 233 days but the streamer had shed 

after 33 days. 

Tissue Trauma 

Tissue trauma still appears to be a significant problem around the 

periphery of the disc tag, particularly at the dorsal and ventral aspects 

(Figs. 41, 45). However, Fig. 45 shows reepithelialized tissue along the 

posteri or periphery i ndi cati ng some mediation of t rauma by the ani mal. Si nce 

some of the tags were removed as a healthy precautionary measure or had posts 

that fractured, it was difficult to determine if a complete healing cycle 

would have occurred. 

The Floy and modified spaghetti tag designs were not retained for 

sufficient time to fully evaluate their healing cycles. 

three spaghetti tags was without sufficient force to seat them properly. 

The application of 

The fourth (Floy type), however, lost its streamer and eventually caused 

an infection, prompting surgical removal. 

had mi grated medi a1 ly. 

It was not determined if the barb 

-44- 
Flag tagged animals sustained abraded areas along the anterior margin 

of the dorsal fin. This abrasion was not significant enough to warrant 

removal of the tags. 

The peduncle-tagged animals sustained significant tissue erosion at the 

junction of tail flukes and peduncle (Fig. 31 ). The tags were removed as a 

health precautionary measure. This tag type needs to be evaluated for a 

longer period of time to determine if the animal could regenerate enough 

tissue to mediate abrasion by the tag. 

Di scussi on 

Posts for dorsal fin disc tags are in need of further improvement to 

reduce application time, and increase retention time. 

sleeved stainless steel post combination provided best tagging results but 

A delrin disc and 

the value of the sleeve is difficult to determine. A sleeveless stainless 

steel post was not evaluated. If nylon or delrin posts are to be evaluated 

further, a comparison should be made between posts with and without sleeves. 

As shown in Fig. 46, the dorsal fin exhibits continuous curvature in 

several different planes. 

adjustment of the post to minimize disc pressure on the epidermis. 

Effective placement of a disc tag requires careful 

This is 

ished with a threaded post strengthened 

Fig. 47) resulting in flexion of the 

where contact is made with the tag 

mal to be tagged in 

may not be as 

time consuming, but can best be accomp 

by a sleeve. Typical aerial behavior 

dorsal fin may still produce abrasions 

(Figs. 41, 45). Since Stenella attenuata, the pr 

the field, is not prone to aerial behavior, these 

i mport ant. 

mary an 

factors 

-45- 
The Floy and modified spaghetti tags were not retained long enough 

for a meaningful evaluation, although the Floy type which lost its 

streamer eventually caused infection and was surgically removed. Absence 

of the streamer may have allowed inedial migration of the barb. This tag 

type has shown good retention in previous tests, and can be applied 

rapidly. We would suggest that they be retested when possible. 

Factors limiting the retention of flag tags appeared to be related 

to materials rather than biocompatibility. Use of sleeved stainless 

steel pins and smooth delrin or polyurethane flays should improve the 

durability of this tag and minimize fouling. 

rather than two, to reduce flexion of the flags. 

Four posts should be used 

Peduncle tags may still warrant further evaluation, using a wider 

and softer foam type material similar to the lining of delphinid radio 

tracking packayes. This material may reduce the tissue abrasion observed 

with the surgical tubing (Fig. 40), and improve visibility. The wire tie 

fastener appears to be an effective locking rnechanisrii. 

A1 though nurnerous i nstances of tissue trauma were observed, a1 1 the 

test animals cornpleted tag site healing cycles rapidly. Antibiotics were 

adiriinistered in their food and copper sulfate was added to the pool 

water. However, it should be emphasized that the open ocean environment 

is essentially sterile, and therefore water treatment or use of 

antibiotics should not bias this tagging data enough to limit its value. 

Recomrnendat i ons 

Field tagqing should be accomplished using dorsal fin d 

or flag tags, or spaghetti tags, with improvements in desiqn 

-46- 

sc taw 

and 
materials as discussed. At this t 

application (per unit) can be comp 

for spaghetti tags), since careful 

me, it should not be expected that tag 

eted in the 60 seconds requested (except 

post adjustments are still necessary. 

Delrin or polyurethane in bright yellow appear to be the most qualified 

materials for discs or flags. Stainless steel posts sleeved with delrin 

should be used for attachment. All these materials are biocompatible. Since 

tag design progress has not completely eliminated tissue trauma problems, we 

recommend that a medication be used during field tagging to supplement the 

animal's adjustment to the tag. 

Progress in evaluation of tags during this project has been affected by 

limited access to test animals specifically designated for tagging research. 

The success of future work will depend on establishing a group of pelagic 

delphinids designated for tagging research by permit, in a controlled 

environment situation. 

-47- 
List of Figures 

Figure 1 - Floy spaghetti tag. 

Figure 2 - Spaghetti tag applicator tool (jab stick). 

Figure 3 - Floy arrowhead tag. 

Figure 4 - Disc-arrowhead tag. 

Figure 5 - Disc-cone head tag. 

Figure 6 - Magnified view of low temperature isotropliic carbon and tissue 

7 - Tag application with a plastic-lieatled mallet - Delphinus. 

8 - Protrusion of tag disc above epidermis - Delphinus. 

interface. 

Figure 

Figure 

Figure 9 - Aerial behavior of tagged ___~ Dtblphinus. 

Figure 10 - Adjustable disc arrowhead tag. 

Figure 11 - Flag tag. 

Figure 12 - Trocaring of the dorsal fin - -_- Delphinus. 

Figure 13.3 - Floy arrowhead tag - ___--- Delphinus. 

Figure13b - Evidence of migration in the tissue - -___ Delphinus. 

Figure 14 - Section of anchor post hole in dorsal fin showing extensive re-epithFigure 15 - Anterior dorsal fin tag with wrap--around streamer. 

Figure 16 - Metal scoop tag. 

Figure 17 - Anterior dorsal fin tag with side mounted streamers. 

Figure 18 - Posterior dorsal fin spring clip tag. 

Figure 19a - Tag headq used for insertion and withdrawal tests 

Figure 1Db - A specially designed spring scale and tag head chuck for measurement 

of insertion and withdrawal forces (psi). 

Figure 20 - Sp.tglietti tag barb arxhored in the fascia. Delphinus. 

Figure 21 - Dissection showing toggled spaghetti tag barbs in fascia layer between 

elialization - Delphinus. 

blubber and muscle tissue - -- Delphinus. 

Figure 22 - Dorsal fin disc tag. 

Figure 23 - Spaghetti tag - modified lend and barb. 

Figure 24 - Spaghetti tag (modified lead and barb) showing healed entry site 

La g e no r hy n c hu s . 

Figure 25 - Snap fastener rubber peduncle belt. 

Figure 26 - Velcro-riibber peduncle belt. 

Figure 27 - Ty rap rubber peduncle belt. 

Figure 28 - Nasco multi-loc cattle leg belt. 

-48- 
Figure 29 - 

Figure 30 - 

Figure 31 - 

Figure 32 - 

Figure 33 - 

Figure 34 - 

Figure 35 - 

Figure 35 - 

Figure 37 - 

Dorsal fin disc tag showing fractured disc - Lagenorhynchus. -- __ 

Idented ring of darkened epidermis around the dorsal fin disc tag's 

perimeter - Lagenorhynchus. 

Abrasion caused by peduncle belt - Lagenorliynchus . 

Dorsal fin disc tag showing fractured disc - - Stenella. 

Disc tag everted by water friction - - Stenella. 

Inflammation and lesions on disc tagged dorsal fin - - Stenella. 

Flow tank test apparatus for tag evaluation. 

Flexion of dorsal fin during flow tank test. 

Dorsal fin disc tag. 

Figure 38 - Multi post flag tag. 

Figure 39 - Multi post flag tag on Stenella dorsal fin. 

Figure 40 - Surgical tubing peduncle belt - Stenella. 

Figure 41 - Fractured dorsal fin disc tag showing healing of abraded tissuc - 

Figure 42 - Fractured multi post flag tag on Stenella dorsal fin. 

Figure 43 - Algae covered multi post flag tag - Stenella. 

Figure 44 - Spaghetti tags - Stenella. 

Figure 45 - Dorsal fin disc tag showing primary abrasion points - Stt~r.1La. 

Figure 46 - Dorsal fin disc tag (improved design) showing fit on Jorsal fin - 

Figure 47 - Disc tagged -___ Stenella - aerial behavior. 

Stenella. 

-____ 

Stenella. 

-49- 
FIGURE 1 - FLOY SPAGHETTI TAG 

-50- 
TAG 

FIGURE 2 - SPAGHETTI TAG APPLICATION TOOL 

-51 - 
FIGURE 3 - FLOY ARROWHEAD TAG 

-52- 
FIGURE 4 - DISC - ARROWHEAD TAG 

DISC 

FIGURE 5 - DISC - CONE HEAD TAG 

-53- 
Isotrophi c 

carbon 

mater i a1 

Tissue 

Figure 6 - Magnified view of the interface where the 

tissue is growing into the low temperature 

isotrophic carbon. 

Photo is courtesy of CarboMetics, Inc. 

- 54- 
Figure 7 - Tag application with a plastic-headed mallet - Delphinus. 

Figure 8 - Protrusion of tag disc above epidermis - Delphinus. 

-55- 
Figure 9 - Aerial behavior of tagged Delphinus. 

-56- 
STREAM ER 

c SHAFT 

-FLUKE 

DISC 

BARB 

MECHAN ISM 

FIGURE 10 - ADJUSTABLE DISC ARROWHEAD TAG 
1 SLEEVE 

ANCHOR POSTFIGURE 11 - FLAG TAG 

-58- 

FLAG 
Figure 12 - Trocaring of the dorsal fin - Delphinus. 

-59- 
Figure 13a - Floy arrowhead tag - Delphinus. 

Figure 13b - Evidence of migration.in the tissue - Delphinus. 

Note "Sport" has di sappeared bel ow epidermis . 

-60- 
Figure 14 - Section of anchor post hole in dorsal fin showing extensive 

re-epitheliazation - Delphinus. 

-61 - 
1 

FIGURE 15 - ANTERIOR DORSAL FIN TAG 

WITH WRAP AROUND STREAMER 

C 

t 

TINE 

t __IIJ 

STREAMER 

FIGURE 16 - METAL SCOOP TAG 

-62- 
- STREAMER 

FIGURE 17 - ANTERIOR DORSAL FIN TAG WITH 

SIDE MOUNTED STREAMERS 

SPRING 

FIGURE 18 - POSTERIOR SPRING CLIP TAG 

-63- 
Figure 19a - Tag heads used for insertion and withdrawal tests. 

-64- 
Figure 19b - A specially designed spring scale and tag head chuck for measurement of insertion and withdrawal forces (psi). 

-65- 
Figure 20 - Spaghetti tag barb anchored in the fascia between muscle and 

blubber layers. Delphinus. 

-66- 
Figure 21 - Dissection showing toggled Spaghetti tag barbs between 

blubber and muscle tissue. Delphinus. 

-67- 
VENT HOLE 

FIGURE 22 - DORSAL FIN DISC TAG 

BARB 

FIGURE 23 - SPAGHETTI TAG - MODIFIED LEAD AND BARB 

-68- 
Figure 24 - Spaghetti tag - (modified lead and barb) - showing healed entry 

site. Lagenorhynchus. 

-69- 
I 

c I 

FEMALE FASTENER 

t 

BELT 

MALE FASTENER 

FIGURE 25 -SNAP FASTENER PEDUNCLE BELT 

VELCRO FASTENER 

FIGURE 26 -VELCRO RUBBER PEDUNCLE BELT 

TY RAP FASTCUER 

FIGURE 17 - TY RAP 9UBJE7 PE3UIICLE BELT 

BELT 

f 

FASTENER 

FIGURE 28- NASCO MULTI-TOE CATTLE 

LEG BELT 

- 70- 
Figure 29 - Dorsal fin disc tag showing fractured disc. 

Lagenorhynchus. 

F,igure 30 - Indented ring of darkened epidermis around the dorsal 

fin disc tag's perimeter. Lagenorhynchus. 

-71 - 
Figure 31 - Abrasion caused by peduncle belt. Lagenorhynchus. 

Figure 32 - Dorsal fin disc tag showing fractured disc - Stenella. 

-72- 
Figure 33 - Disc tag everted by water friction - Stenella. 

Figure 34 - Inflammation and lesions on disc tagged dorsal fin - 

Stenella. 

- 73- 
F-igure 35 - Flow tank test apparatus for tag evaluation. 

-74- 
Figure 36 - Flexion of dorsal fin during flow tank test. 

-75- 
WATER VENT HOLE 

DISC 

FIGURE 37 - DORSAL FIN DISC TAG 

FLAG 

FIGURE 38 - MULTI POST FLqG TAG 

-76- 
Figure 39 - FluIti post flag tag on Stenella dorsal fin. 

-77- 
WIRE TIE FASTENER 

FIGURE 40- SURGICAL TUBING PEDUNCLE BELT 

-78- 
Figure 41 - Fractured dorsal fin disc tag showing healing of 

abraded tissue - Stenella. 

Figure 42 - Fractured multi post flag tag on Stenella dorsal fin. 

-79- 
Figure 43 - Algae covered multi post flag tag - Stenella. 

Figure 44 - Spaghetti tags - Stenella. 

-80- 
F-igure 45 - Dorsal fin disc tag showing primary abrasion points - Stenella. 

-81 - 
Figure 46 - Dorsal fin disc tag (improved design) showing fit on dorsal fin - 

Stenella. 

-82- 
Figure 47 - Disc tagged Stenella - aerial behavior. 

-83- 
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-92- 
Section I1 - Cryogen a3 Marking Equipment Design and Evaluation 

-93- 
Cryogenic Marking Evaluation - Phase I 

A cooperative cryogenic marking experiment involving representatives 

from the Hubbs-Sea World Research Institute, Southwest Fisheries Center 

and the National Fisheries Engineering Laboratory of the National Marine 

Fisheries Service (NMFS) was conducted at Hubbs-Sea World Research Institute 

in December of 1977. 

Methods and Materials 

Heat sink and evaporative techniques were used for coolant application 

to the epidermis of a Delphinus delphis specimen. The heat sink techniques 

required placement of a marking tool cooled to cryogenic levels, on the 

epidermis. 

tool resulted in refrigeration of the tissue. 

The evaporative technique involved spray application of a compressed 

Heat drawn from the warm epidermal tissue to the cooled marking 

gas on the epidermis. 

decreased allowing heat transfer from the epidermis to the evaporating coolant. 

Heat sink marking with liquid NE was conducted using a self-contained 

As the gas was sprayed, its pressure and temperature 

vacuum flask (Foster Froster) modified by the National Fisheries Engineering 

Laboratory (Fig. 1). 

liquid N2 into a hollow brass marking head. 

A button depressed on top of the cannister released 

The marking head was fitted with 

a valve for venting 

with a polyethylene 

in 1 iquid N2 before 

N2 gas. An additional heat sink marking iron was used 

handle and 2.54 cm diameter copper head. It was immersed 

application to the epidermis (Fig. 2). 

-94- 
For marking by the evaporative technique, a freon (monochlorodifluromethane) applicator designed by Dr. R. K. Farrell of Washington State University 

was used (Fig. 3). Dr. Farrell personally made the applications in this 

experiment. The applicator had a hand activated valve to regulate coolant 

flow through a .025 inch orifice, with high pressure line connected to a 

cannister containing the coolant. Epidermal temperatures were monitored with 

a potentiometer and .005 diameter alurnel/chromel constantan thermocouple 

inserted approximately 3 mm into the epidermis. 

Our initial cryogenic marking research focused on the physical and 

biological factors that determine mark legibility. The following factors 

were evaluated during this experiment: 

1) 

2) 

3) 

4) Serial marks with both coolants; 

Heat sink and evaporative techniques; 

Liquid N2 and freon coolants; 

Specialized application equipment for each coolant; 

5) 

6) 

Application times ranging from one to twenty seconds; 

Temperature monitoring equipment to record skin temperature 

reduction as a function of coolant type and application time; 

Marking sites to include the dorsal fin and dorso-lateral areas 

adjacent to the dorsal fin. 

The length of time required for "marked" tissue to regain its original 

integrity with no visual sign of continued swelling, sluffing or repair, was 

used for healing cycle criteria. 

7) 

Results 

Results for the cryogenic marking experiment with a Delphinus delphis are 

summarized in Table I. No infection or other healing cycle complications were 

observed and the animal's social behavior appeared to be normal after marking. 

-95- 
Dorsal fin marks using liquid N? (Fig. 4) 

Very little swelling was observed after marking. Tissue sluffing was 

observed within 3 days. 

marks within 14 days. 

A complete healing cycle was observed for all 

Marks on the dorsal fin with 8, 10 and 12 second application times 

displayed only slight and diffuse depigmentation. 

displayed a pronounced central ized depigmentation, bordered by diffuse 

depigmentation and was the most distinct of the group. 

The 14 second mark 

Dorso-lateral marks using liquid N7 (Fig. 5-top row) 

Pronounced swelling of the mark sites was observed within 30 minutes 

after marking. Tissue sluffing was observed within 3 days. After 51 days, 

only two mark sites displayed a complete healing cycle (application times: 

1 and 3 seconds). 

of % 20% as compared to original size. 

complete healing cycle in 72 days. 

Both of these marks displayed a depigmented area shrinkage 

The other mark sites displayed a 

Dorso-lateral marks of 1 and 3 seconds application time also displayed 

pronounced centralized depigmentation with diffuse depigmentation occurring 

around the perifery. 

distinct "pie wedge effect" of pigmented tissue (Fig. 6). 

Marks of 4 and 5 seconds application time show a 

Dorso-lateral marks using Freon 22 (Fig. 5-bottom row) 

The most pronounced swelling and tissue sluffing was observed with 

this group of marks using Freon 22 (evaporative technique). After 39 days 

-96- 
only one mark displayed a complete hea ing cycle (application time: 10 

seconds). 

76 days respectively (application time 

The other marks displayed a complete healing cycle in 50 and 

11 and 20 seconds). 

Dorso-lateral marks with 11 and 20 second application time displayed 

a fairly uniform depigmentation over the entire mark site. 

displayed isolated areas of pigmented tissue. 

application time displayed a centralized depigmentation bordered by 

extensive diffuse depigmentation. 

Both marks also 

The mark at 10 seconds 

Discus.sion 

The dorsal fin marks were not very distinct (depigmented). The presence 

of dense connective tissue and an arterial tree, which can act as a heat 

radiator, may not have allowed for adequate tissue temperature reduction. 

also may have been ice on the marking head that acted as an insulator during 

There 

application. A larger marking head with more surface area and greater mass 

is also desirable to minimize temperature variance during application. 

Dorso-lateral iriarks using liquid N2 displayed a wide range of variability 

in terms of depigmentation, and tissue refrigeration, within an application 

time span of only five seconds (i.e., 1 second application time reaching a 

temperature of -4.2OC compared to an application time of 5 seconds reaching 

a temperature of -29OC). The latter appears to have produced excess tissue 

trauma that could obscure an arabic numeral mark applied in a field situation 

due to migration of pigmented cells (melanocytes) into the mark site during 

the healing cycle (Fig. 6). 

-97- 
Lower application times would not leave much margin for error in a 

field marking situation since the best mark in this group had an application 

time of only 3 seconds. 

resulted with freon at 11 seconds application time, reaching temperatures of 

-1OOC. However, a freon produced mark with an application time of 20 

seconds produced a minimum temperature only 2OC different than the above. 

This may have been caused by a difference in placement of the thermocouples, 

and pressure of the marking head on the epidermis. 

consistent placement of thermocouples is needed. 

The dorsal fin and dorso-lateral marks were made according to a 

The most satisfactory mark of the experiment 

A method to insure 

"predetermined application time". The Freon 22 marks were applied according 

to a "predetermined temperature level reached" before removal of the coolant 

source. Results using both techniques show variability in depigmentation 

of the mark site (Figs. 5, 6). 

nique with Freon 22 eliminates some factors affecting this variability such 

as marking tool pressure and marking tool mass and surface area. Our next 

phase of cryogenic marking research focused on eliminating the variables 

affecting the degree of mark depigmentation and obtained more data on 

temperature change during the tissue freezejthaw cycle, as a function of time. 

However, the evaporative application tech- 

-98- 
CRYOGENIC MARKING EVALUATION - PHASE I I 

Further marking research acti vi ty focused on the eval uation of 

cryogenic marking equipment developed by the National Fisheries Engineering 

Laboratory (Fig. 7). 

with the National Marine Fisheries Service's Porpoise School Impoundment 

System (PSIS). It consisted of a 30-liter durer connected by two 8-foot 

long vacuum-lined hoses to a branding tool with a "TI'-shaped branding head. 

The flow of liquid nitrogen was regulated by valves on the durer to flow 

down one hose, through the head, and the gas was vented out of the other hose. 

This equipment was designed for field marking operations 

Potentiometers with digital readouts were interfaced with thermocouples placed 

in the epidermis and branding head for monitoring temperature. 

Methods and Materials 

The initial marking experiment was conducted on a Hampshire pig 

(Sus - scrofa) maintained at the Laboratory Animal Facility, University of 

California at San Diego. 

marking system was assembled and tested at the Hubbs/Sea World Research 

Institute. A video system was also tested to record marking procedures, 

temperature changes of the animal's skin and the marking tool head. 

Prior to marking, the self-contained cryogenic 

In addition to equipment evaluation we also wanted to determine a 

temperature reduction plateau, which can indicate a state of intra- and 

inter-cellular ice crystallization in the epidermis. This crystallization 

is important in producing a well-defined mark, as is the rate of cellular 

freeze and thaw. 

The pig was sedated with Ketamine ("Vetalar" 100 mgjml). 

15 cm Teflon template with a "T"-shaped silhouette slightly larger than the 

A 15 cm x 

-99- 
brand head was secured around the pig (Fig. 8). 

to stabilize the marking iron head and allow accurate insertion of the 

thermocouple probes at prescribed depths of 1, 2, or 3 mm. The thermocouples were incorporated into 21 and 31 gauge hypodermic needles, and 

This template was designed 

interfaced with digital readout potentiometers. The temperature changes 

were recorded on video tape. 

epidermis, the thermocouples were inserted to an approximated depth of 3 mm. 

They were left in place after the marking tool was removed to record the 

rate of tissue thaw. 

Due to the firm cornified nature of the pig's 

Application tool pressures of 10 and 20 psi were 

A 

remova 

within 

used. 

Results 

data summary 

of the mark 

30 minutes. 

problems, the exper 

for the pig marking is presented in Table I. After 

ng tool, the mark site became reddened and edematous 

Due to the amount of sedative given and instrumentation 

ment was extended over a 2 day period for a total of 8 

marks. 

healing cycle within 30 days (Fig. 9). No infection occurred but the pig 

was observed rubbing marked areas against the enclosure walls on numerous 

occasions. 

Tissue sluffing was noted after 5 days and all marks completed a 

Figures 10 and 11 illustrate the difficulties of obtaining accurate 

tissue temperature data. Figure 10 shows that the temperature for mark A 

dropped at a constant rate to -2OOC in 776 seconds, before leveling off. 

Figure 11 shows a variable temperature reduction for mark B to +18"C with 

an elapsed time of 157 seconds. 

- 100- 
Discussion 

1 

Testing the experimental cryogenic marking equipment on the pig 

proved to be an informative exercise. Although self-contained, the 

cryogenic marking system was difficult to maneuver due to the weight of 

the liquid I12 in the durer and hose stiffening produced by the cryogenic 

temperatures. 

reposi ti oning of the template , thermocouples, and marking tool head 

making it nearly impossible to record accurate application times and 

temperatures. Frequently, there was interrupted contact between the marking 

iron and the epidermal tissue and thermocouple probes inserted through 

the template appeared to shift position in the tissue. 

The animal's movernent was a major problem, which caused 

Determination of temperature reduction plateaus was not practical 

under these circumstances. 

The apparent slow temperature reduction during marking tool application (Figures 10 and 11) may have been due to the firm, cornified nature 

of the pig's epidermis, placement of the thermocouples at too great a 

tissue depth, or marking tool pressure on the thermocouples. 

Cryogenic Marking of Hawaiian Spinner Porpoise (Stenella 1 ongirostris) 

Two Hawaiian spinner porpoises (Stenella longirostris) at Sea Life 

Park were cryogenically marked to provide additional data for the National 

Marine Fisheries Service field operations. 

-1 01 - 
Methods and Materials 

A Stenella longirostris specimen was captured in the holding tank, 

placed in a stretcher, and taken to a sheltered area out of the sun. Tne 

stretcher was secured to a portable metal stand where the epidermal areas 

to be marked were dried and disinfected. 

careful physical restraint of the Stenella test subjects without sedation to 

insure their health. 

Special attention was given to 

Serial marking was performed on the body and dorsal 

fin, using 1 iquid nitrogen and Freon (monochlorodifluoromethane) coolants. 

Heat sink and evaporative application techniques were used. For heat 

sink application with liquid N2, the portable cryogenic marking system 

developed for fie1 d marking by the National Fisheries Engineering Laboratory 

was tested (Figs. 7, 12). The marking tool and thermocouples were used without 

the template previously tested on the pig, For heat sink application with 

freon, a copper headed marking tool was immersed in a coolant bath. Evaporative 

application of freon was performed with a valved applicator head and high 

pressure line connected to a freon cannister (Fig. 3). 

Each cryogenic mark was made according to a predetermined application 

time, and temperature change during the freeze-thaw cycle was monitored by 

a potentiometer, with copper constantan thermocouples inserted in the 

mark site epidermis. Temperature changes indicated by the potentiometer's 

digital readouts were video taped to compare temperature change to mark 

legibility. 

marks with freon (Figs. 13, 14, 15). 

seconds. 

A total of 21 marks were made with liquid N2 coolant and 12 

Appl ication times ranged from 1-33 

-1 02- 
Results 

Results of the cryogenic marking with Stenella are summarized in 

Tables 2 and 3. All mark sites displayed some degree of swelling within 

30 minutes after marking iron removal. 

8 days. 

Tissue sluffing was observed after 

If the marking iron was applied for longer than 20 seconds, epidermal 

and dermal skin layers were prone to sluff, 

filled blubber tissue (Fig. 16). Even slight trauma would cause these mark 

sites to hemorrhage. Mark sites completed healing cycles within 8-51 days 

sometimes expo5ing capillaryMarks with application times of 12 to 16 seconds and reduced tissue 

temperatures of between -2OC and -6OC appeared to remain the most legible 

(8 months to date) with a minimum of tissue trauma. 

varying degrees of depigmentation and legibil i ty (Figs. 13, 14, 15). 

of marks for comparative data is difficult due to factors such as animal 

All marks displayed 

Use 

marking tool contact with the epidermal surface, tissue compression and thermocouple sensitivity. 

The experiment was shifted from tankside to a covered staging area 

to faci 1 i tate video taping of marking procedures. Transporting the test 

subject an extended distance combined with serial marking and careful 

placement of thermocouples, prolonged their out-of-water time. 

A review of the video tape and plots of epidermal temperature reduction 

across time revealed a mark site's temperature reduction was frequently slow 

or irregular (Fig. 17), which may have resulted from the animal's movement 

breaking contact between the marking iron and epidermis or the thermocouple 

may have changed its depth of penetration. Pressure of the marking iron 

on the epidermis may have compressed tissue and flexed the thermocouple, 

producing an irregular readout. 

-103- 
The animals position in the stretcher usually dictated that 

the marking iron was applied at a 45" to 70" angle to the epidermal 

surface, which may not have allowed even contact (Fig. 17). A comparison 

of temperature reduction and mark legibility as a function of coolant 

types was not possible with the variability observed between individual 

marks. 

Due to the out-of-water time, the animal was frequently wet down 

with a hose to facilitate thermoregulation. Water inadvertenetly covered 

several sites during marking which also slowed epidermal temperature 

reduction. 

dorsal fin due to the presence of dense connective tissue (Fig. 18). 

Epidermal temperature reduction occurred more rapidly on the 

It does not appear that application time can be used as the only 

ssue density and 

ty to compress also appears to be important. 

internal temperature and thaw times for mark #4 and #7 with 

Note the difference 

liquid N2 on Lioele (Figs. 19 and 20). Both of the marks were made with 

the same application times. The dorsal fin's dense connective tissue 

reached a colder temperature and thawed over a longer period of time. 

criteria 

its abil 

in final 

to consistently replicate a legible mark. T 

Discussion 

Comparison of each mark in terms of epidermal and marking tool 

temperatures, application times, pressure, and coolant type was biased by 

a number of factors such as: 

1. Body curvature--Depressions in the epidermal surface formed by 

muscle bundles may not allow complete marking iron contact. 

-1 04- 
2. Body movement--Immobilizing the test subject was difficult. 

Contact of the marking tool with the epidermal surface was 

often broken by sudden body moveiiients. 

Tissue compressibi 1 i ty/marki rig tool pressure--Compression of the 

tissue by the marking tool effects the rate of freeze and thaw. 

Increased tissue compression will promote freezing and slow the 

thawing process because the density increases. 

Thermocouple position in the tissue--The ability to position and 

maintain thermocouples at a constant tissue depth in each mark site 

proved to be difficult, especially with the marking tool compressing 

the tissue around the probe. 

3. 

4. 

5. The animal 's out-of-water time--Since this experiment involved serial 

marking, each mark was allowed to thaw slowly while additional 

marks were made. The test animals were out of the water in excess 

of 40 minutes each. 

be reduced to approximately 3 minutes, the rate at which this tissue 

thaws should also be evaluated in terms of mark legibility. 

Blood shunting by the animal--Tissue freeze and thaw rates may 

be influenced by increased or decreased blood flow to the mark 

site, particularly on the dorsal fin. 

temperature and emotional state may produce blood flow changes in 

a given tissue area. 

To fully evaluate these "physiological and anatomical factors", 

In a field situation this marking time could 

6. 

The animal Is overall body 

the tissue's reaction to cryogenic temperatures at the cellar level must 

-7 05- 
be histologically docuinented. If, for cxmplc, the physiological response 

of the tissue to cryogenic temperatures is too great, excess fibroblasts 

and collagen fibers may bc formed causing a "puckering" of the inark site 

(Fig. 74). iiyperpigr;ictitation can also result. 

At this stage in the research program, we have established a number 

of factors that appear to influence mark legibility: 

1. The temperature of the marking iron as it is applied to the 

animal 's epidcrmi s. 

2. The ability of the iron to function as a heat sink while in 

contact with the epi denni s . 

3. 

4. 

The amount of tinic the iron is in contact with the epidemis. 

The amount of time allowed for cryogenically marked cpiderinis 

to thaw before immersion in water. 

5. 

6. 

To fully evaluate these physical cnd anatomical factors, the 

The density and compressibility of the epidermis. 

Blood shunting by the animal in a given tissue area. 

tissw's reaction to cryogenic tembzratures at the cell ular 1 eve1 - nust be 

histologically analyzed. Cellular activity during the healing cyc:c such 

as pigment migration into the mark site and the production of collagen 

and fibroblasts (whidi in excess can I'pticker" or "shrink" the mark) should 

be carefully studied. 

Further marking should he conducted when marking irons and 

tcmpe~~ture moni tori tlg eqt:iprncnt have b,neii thoroughly tested on 1 z5oratory 

aniiual s . 

-106- 
A thorotrgti refererice search will be required for background 

information as well as consultation with professional specialists in 

histology, derr;latology and cryogenic marking to rea? ize developnient of 

productive experimental design. 

-107- 
List of Figures 

Figure 

Figure 

Figure 

Figure 

Figure 

Fi gure 

Figure 

Figure 

gure 

gure 

gu re 

1. 

2. 

3. 

4. 

5. 

6. 

7. 

8. 

9. 

0. 

1. 

Figure 12. 

Figure 13. 

Figure 14. 

Figure 15. 

Figure 16. 

Foster Froster modified for use with liquid nitrogen (heat 

sink technique). 

Farrell heat sink marking tool used with liquid nitrogen. 

Farrell freon applicator for evaporation technique spraying 

of Freon 22 (monochlorodifluroinethane). 

Cryogenic marks on the dorsal fin - Delphinus delphis. 

Cryogenic marks on the dorso-1 ateral area - Del phi nus del phis. 

Migration of pigment cells into the mark site - Delphinus 

delohis. 

Cryogenic marking equipment developed by the National Fisheries 

Engineeri ng Laboratory. 

The teflon template secured around the pig (Sus -scrofa) prior 

to cryogenic marking. 

Cryogenic marks on a pig (Sus -scrofa) after completion of 

healing cycle. 

Constant temperature reduct ion during cryogenic marki ng w 

1 iquid nitrogen (Sus -scrofa). 

Variable temperature reduction during cryogenic marking w 

1 iquid nitrogen (Sus -scrofa). 

the 

th 

th 

Cryogenic marking equipment developed by the National Fisheries 

Engi neeri ng Laboratory. 

Heat sink technique using liquid nitrogen. 

Marking a Stenel la 1 ongi rostri s. 

Example of cryogenic marks on a Stenella longirostris. 

sink technique using liquid nitrogen. 

Heat 

Close up view of cryogenic marks on a Stenella longirostris 

showing mark site puckering and depigmentation. Heat sink 

techi nque using 1 iquid nitrogen. 

Exaniples of cryogenic marks on a Stenel la longirostris. 

Heat sink technique using Freon 2~hlorodifluromethane). 

Bottom row, evaporative technique with same coolant. 

Tissue sluffing of a mark site exposing capillary filled 

blubber tissue (Stenella longirostris) after cryogenic marking. 

-108- 
Figure 17. Variable temperature reduction during cryogenic marking with 

1 iqui d nitrogen (Stenel 1 a 1 ongi rostri s ). 

Figure 18. 

Figure 19. 

Rapid epidermal temperature reduction during cryogenic marki ny 

of the dorsal fin with liquid nitrogen (Stenella longirostris). 

Temperature reduction during cryogenic marking with liquid 

nitrogen (mark #4) on Stenella longirostris. 

Figure 20. Temperature reduct ion during cryogenic marki ng with 1 iquid 

nitrogen (mark #7) on Stenella longirostris. 

-1 09- 
Figure 1 - Foster Frcster modified for use with liquid N2 

(heat sink technique). 

Figure 2 - Farrell heat sink marking tool used with liquid N2. 

-1 10- 
I 

Figure 3 - Farrell freon applicator for evaporation technique 

spraying of Freon 22 (monochlorodifluromethane). 

Figure 4 - Cryogenic marks on the dorsal fin (Delphinus delphis). 

(Branding times from left to right were 14, 12, 10, 

and 8 seconds.) 

-11’1- 
Figure 5 - Cryogenic marks on the dorso-lateral area - 

Delphinus delphis. 

Figure 6 - Migration of pigment cells into the mark site - 

Delphinus delphis. 

-112- 
Figure 7 - Cryogenic marking equipment developed by the National Fisheries 

Engineering Laboratory. 

-113- 
Figure 8 - The teflon template secured around the pig prior to 

cryogenic marking. 

Figure 9 - Cryogenic marks on a pig (Sus - scrofa) after completion 

of the healing cycle. 

-1 14- 
FIGURE 10 CONSTANT JEMPERAlURE REDUCTION DURING 

MARKING IRON TIME 176 SECONDS 

CRYOGENIC MARKING WITH LIQUID NITROGEN (SUS scmiaJ 

5 +10 

B 

Iz 

L .............. 0 .................................... 4 

* 

10 20 30 80 50 60 70 so so io0 110 120 130 140 150 160 170 is0 is0 200 210 220 

TIME (~econd~l 

FIGURE 11 VARIABLE TEMPERAJURE REDUCTION DURING 

CRYOGENIC MARKING WITH LIQUIO NIJROCEN (Sus scroial 

MARKING IRON TIME: 157 SECONDS 

-15 ................................................................................................................. * 

OFF 

-187’~ 

io 20 30 40 50 60 70 so so iw 110 120 130 WJ 150 160 170 180 190 zoo 210 220 

TIME lsecondsl 

-115- 
Figure 12 - Cryogenic marking equipment developed by the 

National Fisheries Engineerinq Laboratory. 

Marking a Stenel la longirostris. 

technique using liquid nitrogen. 

Heat sink 

Figure 13 - Example of cryogenic marks (liquid N2) on a 

Stenella longirostris. Heat sink technique 

using liquid nitrogen. 

-116- 
F-.qure - 14 - Close up view of cryogenic marks on a Stenella 

longirostris showing mark site puckering and 

uneven depiqmentation. Heat sink technique 

using 1iqui;i nitrogen. 

Figure 15 - Examples of cryogenic marks on a Stenella + lon irostris. 

Freon 22 monochlorodifluromethane). Bottom 

row, evaporative technique with same coolant. 

Heat sink technique using 

-1 17- 
Figure 16 - Tissue sluffing of a mark site exposing capillary 

filled blubber tissue (Stenella longirostris) after 

cryogen i c mark i ng . 

-118- 
- +4, 

= 

3 

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FIGURE 17 VARIABLE TEhlPERATL'RE REDUCTION DURING 

CRYOGENIC MARKING WITH LIQUID NITROGEN (Sleneila loogrrosrris) 

MARKING IRON TIME 10 SECONDS ON DORSAL FIN 

5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 85 90 95 1W 105 

TIME (seconds1 

t FIGURE 18 RAPID EPIDERHAL JEMPERAJURE REDUCTION DURING 

CRYOGENIC MARKING OF THE DORSAL FIN, WITH 

LIQUID NITROGEN (Stenella iongrroitrrsi 

MARKING IRON TIME 32 SECONDS 

10 20 30 40 50 60 70 80 90 1W 110 120 130 140 150 160 170 180 190 200 210 22 

TIME lsecondsl 

-719- 
?5 :I E 

- 20 

t 

FIGURE 20 TEMPERATURE REDUCTION DURING CRYOGENIC 

MAKKINC WITH LIQUID NIJROCEN (MARK #7J 

ON Stenella iongrrosrrrs 

MARKING IRON TIME 10 SECONDS ON DORSAL FIN 

5 10 15 20 25 30 35 40 45 Y) 56 60 65 70 75 80 85 90 95 100 105 

TIME iremndsl 

- 1 20- 
List of Tables 

Table 1. Cryogenic Marking Data Summary - Liquid nitrogen, Freon. 

Hubbs/Sea World Research Institute, Common Dolphin (Delphinus 

delphis). 

Cryogenic Marking Data Summary - Liquid N;z. 

Hawaiian Spinner Porpoise (Stenella longirostris). 

Cryogenic Marking Data Summary - Freon 22. 

Hawaiian Spinner Porpoise (Stenella longirostris). 

Table 2. Sea Life Park, 

'Fable 3. Sea Life Park, 

-1 21 - 
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Section I11 - The Feasibility of Tatooing Small Delphinids 

% 

-127- 
The Feasi bi 1 i ty of Tatooi ng Small Del phi ni ds 

Tatooing may be defined as the implantation of pigment in the 

skin's dermal tissue, where it is permanently retained and readily 

visible. It has been employed by various tribal groups as cosmetic 

adornment for thousands of years. More recently, livestock, dogs, 

seals, and even fish have been successfully tatooed as a means of 

identification. To determine the feasibility of tatooing small 

delphinids in a safe and humane manner, preliminary experiments 

were conducted at the University of California at San Dieso Medical 

School Is laboratory animal facility. The following is a summary 

report on these experiments. 

Polystyrene Particle Spray 

Methods and Materials 

Minute polystyrene particles (biologically inert) coated with 

fluorescent "day glow" pink pigment were applied to 7" x 5" x 1.5" 

section of fresh Delphinus epidermis with compressed nitrogen. The 

spray apparatus is shown in Fig. 1. Tissue spraying was conducted 

according to the following criteria: 

Distance 

Test # (nozzle to target) Pressure on Tissue Time 

1 6 " 100 psi 3 sec 

2 1 " 150 psi 3 sec 

'3 1 125 psi 3 sec 

4 1 It 100 psi 3 sec 

Use of a face mask was necessary during application to avoid breathing 

the polystyrene particles (Fig. 2). After application, the tissue 

- 128- 
was incised across the spray area and examined under ultra-violet 

light magnification to determine the depth of particle penetration. 

Results 

Results of the polystyrene particle spray application are 

summarized in Table 1. The first test displayed no particle penetration 

and no tissue erosion. In the other three tests the polystyrene 

particles appeared to abrade the outer cornified epithelial layer. 

Only test #2 displayed particle penetration into the epidermis in 

sufficient quantity to be visually detected. However, tissue damage 

was sufficient to preclude further testing at higher application 

pressures, The polystyrene particles appeared to abrade tissue 

without penetrating sufficiently for permanent retention. 

Schvco Injector 

Methods and Materials 

The Schvco Injector (Fig. 3) is designed for subcutaneous injection 

of liquids used in medicine. A spring-loaded piston is cocked and 

fired manually, forcing a predetermined amount of liquid from its 

reservoir into the tissue. No information was available on its psi 

rating. Carmine red was used for pigment. Injections were made at a 

fixed pressure into a dog cadaver and a section of fresh Delphinus 

epidermis. The dog cadaver was injected first, positioning the 

injector head on the epidermal surface and at .25, .50, and 1 inch 

above it to determine depth of pigment penetration and surface area. 

The Delphinus epidermis was injected in the same manner. 

,129- 
Results 

Results of the Schvco Injector evaluation are summarized in 

Table 1. Pigment penetration was only realized when the injector 

head was held in contact with the epidermal surface of both tissue 

types. The surface area of the mark was .02 sq. in. When the dog 

epidermis was injected, the pigment was forced completely through 

the epidermal and dermal layers into the blubber. Injection of 

pigment into the Delphinus tissue appeared to diffuse throughout 

the epidermal and dermal layers, providing the desired results. 

The Vernitron Hypodermic Injection Apparatus 

Methods and Materials 

The Vernitron Hypodermic Injection Apparatus (Fig. 4) is also 

designed for subcutaneous injection of liquids used in medicine. 

It is capable of injecting viscous liquids such as penicillin. The 

unit consists of a self-contained compressor and a piston-driven 

injection gun. The liquid is conveyed from an inverted rubber 

capped bottle (60 cc volume) via a stainless steel tube into a 

holding chamber when the gun's piston is cocked, creating a vacuum. 

A trigger is pulled and the compressed air driven piston forces 

liquid into the tissue at 1200 psi. 

The dog cadaver, a fresh section of Delphinus epidermis, and 

Delphinus epidermis with the cornified eipthelium removed, were 

injected with white Burdizzo tatoo ink paste mixed with normal saline 

(2 parts tatoo ink paste and 1 part saline by volume). Subsequent 

tests used a 1:l ratio by volume. The Schvco Injector tests indicated 

the injector head must be in contact with the epidermal surface for 

-1 30- 
pigment penetration. The Vernitron Injector's air pressure was also 

fixed, requiring the head to be placed in contact with the epidermal 

surface, or with a material covering the epidermis, for pigment 

penetration. Several types of materials used to cover the epidermis 

and regulate the depth of pigment penetration. These materials included 

single and multiple layers of cellophane tape, adhesive tape, and a 

.25 inch thick section of closed cell urethane foam. 

Results 

Test results for the Vernitron Hypodermic Injection Apparatus 

area are summarized Table 1. Injection of the dog cadaver with no 

covering allowed pigment penetration into the muscle tissue. 

Cellophane tape covering the epidermis allowed pigment penetration 

between the dermal and muscle tissue. Injection of pigment through 

adhesive tape appeared to distribute it throughout the epidermal 

and dermal tissue, achieving the desired penetration (Fig. 5.). 

Injection of normal Delphinus tissue with an adhesive tape 

covering produced no pigment penetration. When the pigment mixture 

ratio was changed to 1 part pigment and 1 part normal saline by volume 

(used for all remaining tests) the same test was repeated, producing 

a blister of pigment under the cornified epithelium, 

Removal of the cornified epithelium and injection with no covering 

allowed pigment penetration into the muscle tissue: Addition of an 

adhesive tape covering again allowed pigment penetration into the 

muscle tissue. A single layer of cellophane tape on the epidermis 

(no cornified epithelium) appeared to allow the desired penetration 

-1 31 - 
throughout the epidermal and dermal layers. Two layers of cellophane 

tape did not allow any pigment penetration. 

Discussion 

Implanation of polystyrene particles with compressed nitrogen 

using the equipment shown in Fig. 1 does not appear to be practical. 

The large quantities of airborne particles which result during 

application, may jeopardize the animals' health and the tissue trauma 

observed may produce sufficient scar tissue to obscure the tatoo. 

The equipment's bulky nature does not lend itself to field use, ' 

although the technique should be evaluated further. 

The Schvco Injector is designed to inject liquids at a fixed 

pressure; depth of pigment penetration into the tissue is difficult to 

regulate. This device must also be cocked manually and its liquid 

capacity is only 12 cc. It appears that many injections would be 

needed to mark an adequate surface area on a delphinid (i.e. dorsal 

fin) for field observations. 

Pneumatic injection of indelible pigments using the Vernitron 

Injector appears to have some potential for field tatooing of delphinids. 

It is portable, lightweight, can inject pigment rapidly, and has a 

greater capacity (60 cc). If the pigment is not to viscous, this 

injection apparatus does not appear to cause significant tissue 

trauma. It will be necessary to eliminate removal of the cornified 

epithilium from th'e animal or cover the epidermis with material to 

insure required depth of pigment penetration. Air pressure, pigment 

viscosity and injector nozzle design are all factors that appear to 

influence pigment penetration and tissue trauma. They should receive 

further eval uat i on. 

-132- 
Recommendations 

The biocompatibility and tissue resorption of indelible pigments 

should be studied with a live animal experiment. Three genera of 

delphinids (i.e., Tursiops, Delphinus and Stenella) could each be 

tatooed using the "jailhouse technique", requiring application of the 

pigment (sterile) to the epidermal surface and placement in the tissue 

with a needle (sterile), similar to a small pox vaccination. A 1" 

diameter tatoo would allow sufficient diameter for biopsy and 

hi stol ogi cal evaluation. 

Once biocompatibility and tissue resorption of the inedlible 

pigments has been evaluated, tatoo size and location (i.e. body, 

dorsal fin) should be determined. When this data is available, 

further progress can be made to develop portable equipment for rapid, 

safe and humane tatoo marking of delphinids in a field situation. 

-1 33- 
List of Figures 

Figure 1. Spray apparatus for polystyrene particles. 

Figure 2. Spray application of polystyrene particles on Delphinus tissue. 

Figure 3. The Schvco Injector. 

Figure 4. The Vernitr'on Hypodermic Injection Apparatus. 

Figure 5. Injection of dog cadaver with pigment (adhesive tape cover), 

using the Vernitron Hypodermic Injection Apparatus. Note 

pigment distribution throughout epidermal and dermal layers 

at tip of scalpel blade. 

-1 34- 
Fi 

-1 35- 
Figure 2 - Spray application of polystyrene particles on Delphinus tissue. 

-1 36- 
Figure 3 - The Schvco Injector. 

-1 37- 
Figure 4 - The Vernitron Hypodermic Injection Apparatus. 

- 138- 
Figure 5 - Injection of dog cadaver with pigment (adhesive tape cover) using 

the Vernitron Hypodermic Injection Apparatus. Note pigment 

distribution throughout epidermal and dermal layers at top of 

scalpel blade. 

-139- 
List of Tables 

Table 1 - Summary of tatoo experiment data. 

-1 40- 
m 

+J 

4 

? 

m 

2 

13 

H 

-. . . . . .- . . . . 

L 

0 

tJ 

0 

cu 

. . . . . - - 

0 

u 

a, 

G 

0 

d 

I 

c. 

M 

0 

n 

0 +J 

G 

.ri 

a, 

C 

0 

C 

I 

0 

0 

c\I 

d 

tJ 

0 

rd 

+J 

C 

0 

0 

M 

0 

u 

-141 - 
H 

a! 

c 

a, 

a 

tJ 

.,-I .r 

a++ 

I 

a! 

ri 

V 

m 

2 

E 

0 

.P 

C 

.cl 

a 

e, 

CJ 

(d 

si 

tJ 

a, 

C 

a, 

a 

CJce, e,=! 

Em 

wm 

.4 '4 

acl 

a, 

n 

m 

iJ 

a, 

3 

.d 

m 

aJ 

c 

a 

m 

I 

- 

-- -._.. 

e, 

tJ 

rd 

Fi 

I 

- 

, . . _. 

a 

ri 

ri 

v 

.. 

0 

P 

ri 

c\1 

.. 

-142-