A COI4PRATIVE STUDY of the Anadrc,MO•TJS 2\ND FRESHWATER

$A COI4PRATIVE STUDY of the Anadrc,MO•TJS 2\ND FRESHWATER$

-9 ,

A COI4PRATIVE STUDY OF THE ANADRc,MO•TJS 2\ND FRESHWATER

FOPthiS CF ThIE 3R0LN TROUT (SALMO ThUTTA L ) IN

THE RIVER TWEED

JAMES SCOTT CAMPBELL B,, Sc,,, N. Sc.

In partial fu]filment of the requirrrer:ts

for the degree of

Doctor of Philosophy

UNIVERSITY OF EDINBURGH

OF :1?o1d:smy AND NATI7RtL RiSQURCES 1972 a

I hereby declare that this thesis has been composed by myself and the research presented in it is my own.

.4 /

ABSTRACT

In an attempt to find differences between brown trout and sea trout (SaInt trurta Li.) of the River Tweed,

Scotland, the population biology of the two types was studied from 1969 to 1972. The sea trout, spawning in the small tributaries of the Tweed, were :cred:dnantly age 2.14 and 3.1+ maiden spawners with aslight preponderance of females. Upstream migration in these small burns appeared influsnced by rate. of flow and/or water level. Spawning brown trout were rc:st1y age 2± and 3± with a male to tamale ratio of 6 to 1, Male brown trout spent an average of 145 days on L1n apasming grounds while females remained only 2-S days. Tagging studies, of brown trout indicated summer residence in the L.aad with limited movcment until migration upstream entering the burns to spawn in the autumn. The dcnsLreac movement of sea trout fry immediately after emergence appears influenced by rate of flow, Difficulties were encountered in the estimation of age, growth, population size and mortality of trout under 3 veers of age because of the amount of immigration into and emigration out of areas of the study streams. Over 2000 downstream migrant trout were captured at stationary traps in the spring Or- 1971 and 1972. Much of the movement occurred in Liarch and April and appeared to he influenced by bo th flow and water temperature. Those trout, that migrated at the beginning of ' the season were older and larger than those that moved down later; not all fish that were taker, in the traps had a silvery appear ance. Collections of

silvered sea trout sirolts £ñ!OWGCL ?. predominance of females suggesting that there may be a greater tendency for two and three year female trout to migrate to the sea. Since male brown trout appear to participate in the spawning of sea trout, the si.igqastion is made that the freshwater- resident trout of the Tweed may actually be progeny of sea trout. Artificially produced broods of sea trout and brown trout matings, after being held in a hatchery

for one year 0 were stocked macmall streamo tributary to the Tweed. Seventeen per cent of the trout that were large enough to be tagged 0 were subsequently recaptured

Lt a stationary trap some distance downstream of the planting site. No consistent differences were found between the electrophoretic patterns of brown trout and sea trout blood serum proteins on substrates of cellulose acetate or polvacrylarnide gel. Comparison of the molecular weights of ribosomal RNA of brown trout and sja trout could conceivably become a useful technique, but attempts in this study to utilize it indicate that refinements of the method are necessary before it can be of use as a taxonomic tool at the specific level. Meristic series of wild brown trout and sea trout, as well as artificially produced progeny of the two 8 were compared but no consistent differences were found. This may be due to the absence of reproductive isolation in the natural stocks. It is concluded that the freshwater-resident and anadromous forms of the brown trout Ore taxonomically very similar and more soohisticated techniques than used in this study are roqutred to separate them.

/ ACxJowLEDGE.ENTs

I should like to thank ray supervisor, Dr DH Mill. for allowing me to undertake this study and for his cp,iidcinc.e and interest throughout the investigation. I gratefully acknowledge the assistance of my two collea ;ue.s,

Dr. M,M., Hailiday and Dc, T.Y. Aldcori WflO cave oJE their time on many occasions,

I wish to express my gratitude to the Tweed jorruissioners. for permission to work on brown trout and sea trout in

Tweed and, in. particular, I thank Lt. -Cd. R 0 M. Ryan 1 Superintendent to the Commissioners f or his assistance anc helpful suggestions. I era also grateful to the Forestry Commission and the Foresters--In--Charge: of Glentress and

Cardrona for allowing me to work in those forests. I especially thank Mr. R. Gibson of the Forestrr Co.mnai.ssicrt Cardrona for his help.

I acJcnowlc-dge the assistance of Mr 0 I. Bruce of the Berwick Salmon Fisheries Co. Ltd., who collected sea trout scales from the commercial catch and Cr, J. Stansfeld and Mr. N, Smart of Joseph Johnston and Sons. Ltd,, Montrose

Who held trout for me at the Kinriaher Tuout Fan

Nv appreciation is also due to the X-ray Department

Small Animal Clinic of the Royal (Dick) School of \Tateri.narv

Studies. I also wish to thank Mr. H. Urquart of the

Department of Zoology, University ci Edinburgh for the determination of the molecular weicfhts of ribosomal.9NA,

This study 'was financed through the award of research grants (fl3/110/1) from the Forestry Commission for 1970-71 and-1.9 - -. 72 and a University of Edinburgh post- graduate Studentship, LIST OF FIGURES

Figure Page 1 Species of North American Sa1moic3.ae 13 classified by degree of anadromy (from Rounsefell, 1958) 2 The Tweed basin and location of study area 20

3 The study area 23

4 - View of Kirk Burn as it flows northward 28 through pasture land to the Tweed. 5 Portable "back pack" electric fishing 36 apparatus 6 Box trap situated in Kirk Burn to capture 37 upstream migrant sea trout and brown trout in the autumn of 1972 7 Fyke net situated inc4lensax Burn to capture 37 downstream migrant trout in the spring of 1971 8 Plastic disc tag and applicator (scale in 41 inches) 9 Ripe female sea trout 54.4 cm and 1210 g, 45 captured in Kirk Burn on 16 November 1971 10 Ripe male sea trout, 53.3 cm and 1212 g 45 captured in Oxnam Stater on 15 December 1959 11 Movement of ripe and spent sea trout in 46 Kirk Burn, 1970 and 1971 12 Scale of a 49.5 cm female sea trout, age 3.1+, 48 captured 11 November 1970 in Kirk Burn (x 25)

13 Scale of a 49.0 cm female sea trout, age 2.1± 49 captured 27 November 1970 in Kirk.Burn (x 25) Figure Page

14 upstream movement of brown trout ralazed to - water level and water temperature in Kirk Burn1970 and 1971

15 Ripe male brown trout, 26.,2 cm and 216 g 64 captured in Kirk Burn on 19 December 1971

16 Length frequency and sex composition of 68 spawning brown trout captured moving upstream in Kirk Burn, autumn 1970 and 1971

17 Scale of a 205 cm male brown trout., age 3+, 69 captured 26 November 197! in Kirk Burn (x 30)

18 Age frequency distribution of male and female 70 brown trout captured moving upstream in Kirk

Burn, 1970 and 1971

19 Scale of a 19.1 cm female brown trout, age 7+, - 79 captured 30 October 1967 in G]entress Burn

(x 30) 20 Length frequency distributions of trout 81 captured in Glentress Burn

21 Fry trap situated in Kirk Burn to capture 84 emerging sea trout fry, spring 1972.

22 Number of sea trout fry captured in the fry 87 trap situated in Kirk Burn, spring 1972.

23 Monthly length frequency distribution of trout 9 captured in Kirk Burn, 1970-1971

24 Length frequency distribution of trout captured s€ in three sections of Kirk Burn, February 1972

25 Sea trout ernoit, 24.5-cm, seine-netted from - 103 the Tweed at Not-barn Bridge, 6 May 1971 Figure Page

26 Dotstreern rlliqrr3nt trout captured in 103

Giensax Burn by tyke net on 18 April 1971

(scale in inches),

top: silver appearance, 15.9 cm, 41 g

bottom: brown appearance, 151 cm, 36 g

27 Downstream movement and appearance of trout 105

captured in Glensax and Dead burns, 1971

28 Downstream movement and appearance of Lrcut 106

captured in Kirk Burn, 1972

29 Change in age composition of downstream migrant 112

trout in Kirk Burn, 1972

30 Change in length fraqtency and appearance of 113

downstream migrant trout in Kirk Burn, 1972

31 Length frequency distribution and appearance 114

of trout captured moving downstream in three

burns trihutaryth the Tweed, 1971

32 Calculated growth rate of trout captured 118

moving downstream in three burns tributary to

the Tweed, 1971

33 Length frequency, appearance and sex composition 119

of supposed sea trout smolts, captured at -

various locations on the Tweed

34 LateraL view of the cleared and alizarine- 128

stained head of trout showing:

deformed lower jaw (x 25)

normal lower law (x 25)

35 "Hybrid" trout : 131 cm and 24 cm, captured 133

- at the box trap in Kirk Burn, 15 April 1972 Figure P.ge 36 ScaLe of a 53.0 cm male sea trout, age 137

3. 1+ captured 16 July 3.971 in the corrnerc±ai nets at Berwi.ck-upon.-Tweed (x 25) 37 Scale of a 47.0 cm male "estuarne" brown 139 trout, age 5±, captured. 16 July 1971 in the

commercial nets at Berwick- uoon-Tweed. (a 25)

• 38 Ti-ends of the River Tweed corrnercjal sea trout 141 catches disclosed by 5-year rolling averauas 1952.-1970 1859-1895 39 River Tweed sea trout catches. Annual average produce of three periods of 15 years (the pro•iucc- of the week in which the greatest number of fish was caught is represented by 100) 40 . Electrophcretic pattern on cellulose acetate of 161 blood serum protein . from three types of trout (S.trutta) and dog (the fractions are numbered 1 to 12 with increasing mobility)

41 Electrophoretic pattern in polyacrylamide gel 164 of blood serum proteins from three types of trout (Strutta)

42 Actual polyacrylamide gels after electrcn5horesis 165 and staining for blood serum proteins of trout (S.trutta). A,& - brown trout; C.D - sea trout; E,F - brown trout x sea trout

43 Polyacrylamide gel electrophoresis of brown 177 trout ribosomal RNA and its separation from the ribosomal RA of Ec01j. Figure Page

44 Cleared and alizarin-stained body parts of 187 trout (Strutta) for meristic comparison

45 Variation of the dorsal fin, anal fin, pectoral 191 fin and brarichiostegal ray counts of trout

(S.trutta) illustrated by the Hubhs and Hubbs (1953) graphic method.

46 Variation of the precaudal, caudal and total 192 vertebrae of trout (Strutta) illustrated by

the Hubbs and Hubbs (1953) graphic method 47 Radiograph of adult trout (S.trutta) bcdy parts 195 for rneristic comparison (actual size)

43 Radiograph of juvenile trout (S€trutta) for 199 the counting of total number of vertebrae (actual size) 49 Variation of the total number of vertebrae 200 of juvenile trout (Strutta) from various

locations in the Tweed, illustrated by the

Hubbs and Hubhs (1953) graphic method. LIST CF TAJBLES

Table

2. Some phys 4 cal. characteristics of the study 26 streams 2 Stream drainage characteristics of three study 29 streams tributary to the Teed (Mills, unpublished data)

3 Chemical characteristics of Kitk ! GLentress and .30 Glensax burns, tributary to the Tweed

4 Age, growth and sex ratio of sea trout c.cotured 50 in Kirk Burn in the autumns of 1970 and 1971

5 Age, growth and sex ratios of sea trout: and 51 J.arqe brown trout captured at various locations on the Tweed in the autumns of 1970 and 1971

6 Proportion by weight and volume of bottom, 58 material of different sizes taken from the proximity of sea trout redda in Kirk Burn

7 Spawnin'g aggreqations of brown trout and ssa 59 trout electrofished from Kirk Burn 8 Number of days sea trout spent above the traps 61 in Kirk Burn in 1970 and 1971 9 Number of fish entering the box trap in Kirk 66 Burn on 23-25 November 1970

10 Age and growth of brown trout captured at the , 71

box trap in Kirk Burn in the autumns of 1.970 and 1971 ii Sex ratio of brown trout captured in Kirk Burn 73 in the autumns of 1970 and 1971 Table Page 76 12 Drown trout which were tagged in Kirk Eurfl r migrated to the Tweed and were recaptured in Kirk Burn 13 Relationship of male and female sexual maturity 78 to "length of brown trout from Glentress Burn, 1967 14: Mean lengths and weights of newly emerged trout 88 fry in Kirk Burn, 1972 15 Number, mean length and mean wet weight of age 97 group 0 trout captured each month in Kirk Burn at the study section, 1970-71 16 Number of downstream migrant, trout captured by 102 thx trap and floe net in 1971 and 1972 17 Downstream movement of trout in Glensax Burn 106 during a 24 hour period 18 Age composition by percent of downstream migrant •110 trout captured by trap in three tributaries of the Tweed, 1971 and 1972 19 Calculated lengths for each age group of trout 117 moving downstream in the three study streams, 1971 20 Summary of tag returns froth brown trout originally 125 tagged in Kirk Burn 21 Mean fork length and percent deformed of two 130

brown trout x sea trout crosses placed in Kirk Burn on 3.4 March 1972

22 Summary of tag returns of "hybrid" trout captured 132

at thl trap downstream of the planting site, 1972 Table Page 23 Calculated lengths of a sample of sea. trout 136 from the commercial catch of the Bej.wick Salmon Fisheries Co. Ltd,, 1971 24 Average weights, lengths and sex ratio of trout 133 by age groups in the commercial catch of the Bexwick Salmon Fisheries Co. Ltd., 1971 25 The mean percentage and percentage of occurrence 3.62 of each fraction of serum proteins seDarated by cellulose acetate electrophoresis

26 Mean number and standard error of some meristic 190

characters of artificially produced. progeny 27 Mean number of some meristic characters oL adult 396 Lxo.n trout and sea trout coli.ected C0 tte

Tweed. CONTENTS

P age Section I. INTRODUCTION

Historical Review of Classification 3 Degrees of Anadromy in Salmonidae 8 Naturalised Populations of Salmo 15 trutta Approach and Objectives of this 18 Study

Section Ii. THE ECOLOGICAL RELATIONSHIP OF BROWN TROUT AND SEA TROUT

A. Description of the Study Area 19 (1) The Tweed basin 19 a) Location and extent 19 V Vegetation 19 c) Geology and physiography 21 a) Climate 22 (ii) The study streams 22 a) Methods 24 b) Characteristics of the 25 streams B. Terminology and Methods 32 (1) Terms applied to various life 32 history stages (ii) Scale reading 32 (iii) Designation and recording of age 33 (iv) Methods of capture 34 (v) Treatment of material 39 Page

C Characteristics of the Spawning. 44

Populations

(1) Sea trout in Kirk Burn 44 Time, duration and size of 44 the spawning migration

Age and size of the fish 47

C) Sex composition 53 a) Factors influencing tha 54 rnigrat ion

• e) Spawning and spawning 56 aggregations

• f) Post-spawning behaviour and 50 an

(ii) Brown trout in Kirk Burn 62

• a) Time, duration and size of 62 the spawning migration

b) Age and size of the fish 67

C) Sex composition 72

d) Post-spawning behaviour 74 e) Return to the spawning grounds 75 of previously tagged trout

(iii) The (resident) brown trout in 75 Glentress Burn

a) Sex composition and maturity 75

• • b) Growth 77 Page D. Incubation and Early Developr.ient 82 of Trotft Fry

(I) Construct4.on of the traps 82. Period of Incubation and 83 emergence Size and movement after 85 emergence E. Trout in Kirk Burn Prior to 91 Downstream Migration (I) Characteristics of the nopulation 91 Age and growth calculations 93 Population estimates 95 AflflU&L survival 98 r. • Downstream Movement of Juvenile Trout 100 (I) Introduction and methods 100 (ii) Appearance of dcwnstream migrants 101 Time and duration of the migration 104

Age and size composition . 109 Sex composition 116 Downstream movement of "resident' 120 trout in Gientress Burn G. Movement of Tacrged Adults 122

(1) Sea trout kelts . 122 (ii) Recapture of adult brown trout . 123 H. The Release of Progeny of Brown 126 Trout .x Sea Trout and Their Subsequent Recapture (j) Introduction and methods 126 Page Mortality of progeny 127 Size of the progeny 129 Downstream movement of the 131 "hybrids" and their appearance; I. The Commercial Catch of Trout at 135 Berwick-upon-Tweed (1) Introduction 135 Calculated growth and age 135 The commercial catch 140 J. Discussion 143

Section IIfl ELECTROPHORETIC STUDIES OF BLOOD SERUM

PROTEINS

A. Introduction 143

(1) General principles of zone 149

- electrophoresis

(ii) Electrophoresis and the serum 150 and tissue protein of fish (iii) Factors which may affect the 151 electrophoretic pattern (iv) The experimental approach 153 B. Materials and Methods 154 (1) Trout 154 (ii) Blood collection 155 (iii) Electrophoretic methods 156 a) Cellulose acetate electro- 156 phoresis P acre b) Poiyacrylamide gel disc 158 electrophoresis Results of Electrophoresis 160 Discussion 166

Section 1'T. MOLECULAR WEIGHTS OF RIEOSOMAL RNA IN BROWN TROUT AND SEA TROUT A. Introduction 171 B. Material and Methods 173 (1) Trout 173 (ii) Preparation of RNA 173

(iii) Electropi-ioretic methods 174 C. Results 176 D. Discussion 178

Section V. MERISTIC COMPARISON OF BROWN TROUT AND SEA TROUT A. Introduction 179 B. Comparison of Artificially Produced 132 Progeny (1) Methods and problems enc3untered 182 Staining technique 185 Methods of counting rays and . 186 vertebrae Results - 189 C. Comparison of Adult Brown Trout and 194 Sea Trout

(1) Methods and techniques 194 (ii) Results and their interpretation 194 Pace Numbers of Vertetre of Juvenile 197

Trout from Different Locations (1) Methodz and the results 197 Discussion 201

Section VI, DISCUSSION ZJgD CONCLUSIONS 206

Section VI SUGGESTIONS FOR FUTURE RESEARCH 210

REFERENCES 212

APPENDICES £

I. INTRODUCTION

Brown trout and the anadrornous form, sea trout, are at the present time commonly regarded as constituting

a single species, Salmo trutta Linnaeus (]758) . That is, certain individuals of the species in the course of their life history migrate to and from the sea while others remain permanently in freshwater. The brown trout or freshwater form is indigenous to Europe! North Africa and western Asia as far as the Ural Mountains of the U0 S.S.R in the north and the Amu Dara drainage of the Aral Sea south eastward through Afghanistan and into Pakistan (Macorimrnon, Marshall and Gots, 1970). The distribution of the sea trout is not so extensive and is found in Iceland and Scandinavia, the White Sea and

Cheshkaya Gulf in the north to the Baltic, North Sea and Atlantic Ocean as far south as the Bay of Biscay (Frost and Brown, 1967) Throughout the range of this latter form of S.trutta, the two types appear to inhabit the numerous watersheds in varying proportions. In any stream or river management plan it is essential to know whether one is dealing with one homogeneous population or perhaps several. When a commercial or sport fishery depends wholly on one stock of fish, the fishery will be affected by the quantities caught in any one locality. If, on the contrary, there is more than one stock or type present then each must be treated as a separate unit. The question as to whether the migratory and non-

migratory habits of sea trout and brown trout respectivaly are the result of individual variation, environmental n pressures or genetic variation is of ±cIporLo.nce to the. proper manaqernent and conservation of the species. This controversy concerning the relationship of hrcwn trout and sea trout has been the subject of speculation for more than a century. 3

A. Historical Review of Classification.

Linnaeu, when naming the trout of Sweden Sallno

trutta 9 regarded the sea trout (Sariox) and the brook trout (S.fario) as distinct species. On describing the fishes of the Firth of Forth, Parnell (1839) felt that

the genus Salmo- contained five spec2ies of trout which included eight varieties of the bull trout (S.eriox) the salmon--trout or sea trout (S,trutta), the parr (3. sa.muius). the common trout (S.fario) and Lochlz-ven trout (S.coecifer), Marrell (1841) resorted to fin ray and vertebral counts to distinguish the trout species of Britain.. Sc included the grey trout, bull trout or roundtajl (better known from the Tweed) (S0ariox), the migrating species or salmon-trout (Strutta) the parr (S.samulus), the common trout (S,fario), the great lake trout (S. ferox) and the Lochieven trout (S. levenensis or S.coecifer), - In the Tweed, according to Scrape (1843), there

existed three species of trout; S.trutta or the salmon-- trout (white trout, phinock, sea trout, whitling or hirling), S.eriox or the grey trout, bull trout or roundtail and Sfario the common trout, Stoddart (1847) in his 11,%nglers Compztnion to the Rivers and Lochs of Scotland" described four varieties " of Salmo faria which were the gillarco (S.stornachicus), S.Ievenensis, S.f erox, and the swallow-smelt of the Tweed as well as three other species of trout, S.eriox (bull trout), S.trutta (salmon- trout) , and S.albus (finnock or herling) Lx

The taxonomic position of the various trouts were made still more complicated by Couch (18o5) who ncluded the peal (S.trutta) from the Twecd the sewen (s.cambricus) the samlet or parr (Ssamulus) as well as six other species of trout. Gunther (1866) believed that there were ten species of trout in the British Isles; namely the sea or salmon trout (S.trutta) the sewin or western sea trout (S.cambricus) the phinock or eastern sea trout (S.brach ypoma) the Galway

sea trout (sgllivensis), the Orkney sea trout (Sorcadens±s) 9 the river trout (S,fario the great lake trout (Sferox) 41 the gillaroo (S. stornachj-cus) the Welsh blacicfinned trout

(S.niqriDinnis) and the Loch Leven trout IS. ieven.ensis) He went further to separate S.ferio into a northern and southern form and distinguished 12 other species of trout from continental Europe. Although in 1880 GUnther still held that there were ten species of trout in the British Isles, he did concede that 'hybrids" might occur naturally from crossings of sewen (S.cambricus) and river trout (Sfario) and that there was evidence of 'hybrids" being formed from S.fario and S.trutta matings. Buckland (1881) concluded that the bull trout (S.eriox) of the Tweed was a much inferior trout when compared to the sea trout (s.trutta). He even went as far as to suggest that it was the cause of the salmon disease and that the netting season should be extended to rid the Tweed and other rivers of this spcci'est In this connection 9 between the years 1868 and 1871, bull trout were destroyed in the Coquet during the spawning season (Maxwell,, 1904) Ct

Both }3oughtori (138]) and Seeley (1336) agreed with

Gunther l880) as to the number of spocies of trout in the British Isles but Seeley (1886) even went, further to list 29 principle European species of trout based on the numbers of fin rays and ploric appendages

Day (187) combined all of the supposed species of trout under Salmo t rut ta o stating the sea trout and brown trout were local races of one species and that the migratory or resident habits were the result of environmental differences and the markings dependent on immediate

surroundings. Although Maxwell (1904) stated that he

adopted the view of Day (1887) and disagreed with Glintlier (isoo) he retained the bull or sea trout (S.eriox) the salmon-trout or white trout (S.trutta) and the common or

brown trout (S.fario). lie conceded that perhaos freshwater

trout and bull, trout were interchangeable and that an

attempt to destroy the latter must include measures for

the suppression of the former.

Mallocl'i (1910) grouped sea trout and bull trout

together and concluded that the "sea trout (Eh.trutta) is

the same fish as that which is called 'white trout 9 bull

trout grey trout, peel s sewin o brith-dail : salmon-trout

and many other names." Further, he stated that there

was only one species of trout (Sf aria) in Great Britain

and that in the different varieties 0 the differences were

caused by- the nature of the water in which they were found

and the food they ate.

In 19l1 Pecan published his book 'Freshwater Fishes

of the British isles" and stated that although the sea trout 6 and non-migratory brown trout differed so much in habits and appearance, there were no - structural differences and the young were indistinguishable Also he made the points that: 1. Sea trout if prevented from going to the sea will live and breed in freshwater,

2 Trout export d to New Za1an have found their way to the sea and have given rise to an anadromous race. Estuarine trout are often intermediate in appearance and have habits between the migratory and non- migratory fish. Some sea trout smolts nrobahly áo not leave the estuaries for the sea except for a few hours at a time and so become estuarine or tidal trout.

There is good reason to believe that in nature the ranks of the sea trout are teiriforceci by the offspring of the river trout and vice versa. Lamond (1916) arrived at the same conclusions as Pagan, while studying the trout of the Clyde area but at the same time he suggested that there might be other populations of sea trout winch ranged farther out to sea and were distinct from the brown trout, Nall (1930) Calderwood (1930) and Menzies A19361 agreed that sea trout and brown trout were the same species even though there was such a variation in appearance and size between non-migratory trout in various, environments and the sea trout. They further added that the hull trout (roundtail) of the Tweed arid the Tweed sea trout showed no real difference in race, as most large sea trout had rounded tails. Meanwhile, Heri}ting (1g29) and Aflnbdcn (1934)

working in Germany on the relationship of salmon sea S trout and brown trout felt that one should separate the two types of trout at the specific level because of difference in structure and rate c -f growth. Schnakcnbeck (1940) studying the three forms of Salmo trutta; the sea trout (S.trutta trutta), the river trout (S.trutta f aria) and the lake trout (S.trutta lacustris), was of the opinion that there were discrete but definite inherited differences present. Otterstràm (1936) on the other hand felt that the forms were not yet completely separated but that the sea trout and the river trout were more closely related to each other than they were to the "landlocked" form or lake trout. Berg (1932) listed six subspecies c S.t.utta distributed in Europe and Asia which were distinguishable from one another. Bnárescu (1964) described three subspecies of S,trutta from Romania and they were S.trutta labrax, Strutta fari.o and S,trutta lacustris0 Modern ichthyologists generally accept the concept of Regan (1911), Jordan (1926), Hubbs (1930) and Nall (1930) that there is but one species, Salmo trutta, and that ti-out with distinctive features should be recognised at only the subspecific level : if at all (Alms 1948; Fabricius, 1953; Trewavas 1953; Frot and Brown e 1967) 8

r Degrees of Anadromy in salrnonidae

As the term "anadromous' has been used in the title of this dissertation, it would seem prudent that one should define the word. Usually anadromous means " up running" and refers to fishes that leave seas and ascend

rivers or streams to spawn (Rourisefell and Everhart, -1 95 3D) Occasionally the term has been applied to freshwater lacustrine species which "anadromously' run up streams flowing into the lake habitat (Myers, 1949j. In this thesis, only those fish which have scent a portion of their life in the sea will be considered truly anadromous or migratory. This type of Salmo trutta will be called sea trout while those which remain in freshwater will be termed non-migratory, freshwater or more simply brown trout whether they are progeny of sea trout or not. In the family Salmonidas, there is a wide degree of

- divergence in anadromy. The life historic-s of the eastern Pacific salmon of the genus Oncorhychus include species which spend from less than one year to four years at seas The anadrornous members of the genus Salmo namely the

Atlantic salmon (Salmo salar L0) stedlhead rainbow trout (saimo aa 4 rdn eri Richardson), cutthroat trout 1 ..- (aia2 cJ2st Richardson) and sea trout (Salmo trutta L.) - spend about two years at sea before returning to freshwater to spawn.

The third group of salmonids which exhibit some anadromy 1 although in a very limited form, are the true chars of the genus Salvelinus. These fish usually spend only a short time in the see and include Salvelinus font thalis 9

(Mitchill) known on the east coast of North America

as sea-run brook trout or "coasters" (Maccrimmon and Cdmphefl., 1969) ; Salvelirus malma (Wa1baum) the Doily

Varden char; Salvelinus salvelinus (Linnaeus), the. Alpine char; Salvelinus Na,inus (Linnaeus), the Arctic char,,

In the genus Salmo the sea going habit is firmly 'I established but at the same time all of these species have forms which are resident in freshwater. Further, the presumably anadromous forms of Salmo are capable of maturing and reproducing in freshwater. "Land-locked" Atlantic salmon occur in a number of countries throughout

the world even though their access to the sea is not blocked.

Populations of this type exist in eastern North ?anerica, Norway (Dahl, 1928), Sweden Russia and New Zealand (Mills. 1971). These freshwater forms normally use a

lake as a miniature sea ascending its tributaries to spawn. Conversely: large numbers of sea-run Atlantic salmon have been planted in freshwater lakes of eastern North nerica and were found to develop and grow as well

as the landlocked form (Rounsefel].,1958). Wilder (1947) undertook a comparative study hetwedn the Atlantic salmon and what he called the lake salmon, faTh Eo sir (Girard). He found that the adults of the two generally differed only in colouration., spotting and flesh colour and so concluded that any differences were only due to' differences in the environment and diet. No consistent differences could be found in 26 body measurements or counts. 10

The rainbow trout of western North America has an anadromous form which spends a portion of its life in the sea and is known as the "steelhead' rainbow trbut.

Resident freshwater individuals which may appear as small, dark and heavily spotted inhabit snail streams and are nearly always found in streams which contain anadromous stock (Rounsefell, 1958; McAfee, 1966).

Whether there is any inheritable differences between those fish which mature in the sea and tncse remaining in freshwater has long been controversial. Neave (1944) found a significant difference between the averace scale counts of anadromous and freshwater rainbow trout of the Cowichan

River system in British Columbia and separated them into two indigenous races. Briggs (1953) agrees that the two types occur in the same streams and uses the concept of "spatial isolation" since the small rainbow trout tend to spawn in smaller trabutares. According to Snapovalo'; and Taft (1954), the situation is more coniplicated as juvenile steelbead in Waddell Creek, Cal, ifornia e may remain for a whole season in the lower portion of the stream ; some. moving out to sea, while others make an upstream migration and then a downstream migration. Also, they state that some steelhead of both sexes spawn before going to the sea, while others remain in freshwater without going to sea at ski.

The question as to whether the sea-going or resident habits of certain salmonids are the result of environmental or genetic influences has been partially answered in the case of the sockeye salmon, 2.n.corlynchus nerka (WsilDaum). There ii. are really not two but three forms of this salmon; namely the anadromous form, the dwarfed or landlocked. form (kokanee) and thirdly the 'residual" sockeye discovered by Ricker (1938) in Cultus Lake, British Columbia. He concluded that these 'residuaj' were progeny of anadromous parents because the fish differed markedly from normal nonmioratory kokanee in breeding co1our, time of spawning and other characters.. Foerster (1947) in attempting to answer the question of the relationship of kokance and sockeye salmon, reared young kokanee and liberated them in a creek below the Cultus Lake counting fence r In their fifth year, a number of these fish were recaptured in the Fraser River. They were fully grown and of a size comparable to that of five year old Fraser River sockeye salmon.

The North American brook trout (char) is usually found only in freshwater but does exhibit the anadromous habit if there is an outflow into the sea. While in the sea, they remain in the influence of the parent river (White, 1940). The tendency of a portion of the population to visit the sea appears to take the form of a feeding migration (White, 1940) and is temperature directed

(White, 1941; Smith and Saunders, 1958) Wilder (1952) found that the sea-run trout and the freshwater trout showed environmental differences such as colouration and flesh colour but he could find no significant difference in the meristic characters counted. He concluded that -L the two types constituted a single taxonomic. unit Rounse.tell (1958) established six criteria by which the anadromy of the various Ncrth American salmonid species could he classified. They were: Extent of migration in the sea, Duration of stay in the sea. State of maturity attained in the sea. Spawning habits and habitat.

S. Mortality after spawning. 6. Occurrence of freshwater forms. Each criterion was subdivided into as many categories as appeared feasible from the knowledge at the time. The category signifying the most anadromous condition was given a score of 10; the least significant, a score of 0. When the species of Salmonidee were plotted according to the six criteria, each of the four genera (Oncrhynchus. Salmo, Selvelinus and Cristivomer) occunied a distinct area in the degree of anadromy exhibited (Figure 1) It perhaps should be pointed out here that most ichthyologists now place the lake trout (char) Cristivomer namaycush in the genus Salve.li us to emphasise its close relationship with other chars. The details of the argument are presented in papers by Morton and Miller (1954) and Vladykcv (1963). Rounsefetl (1958) classified Salmo trutta as being optionally anadromous and placed the species between the rainbow trout and the cutthroat trout. At the seine time, Rounsefell (1958) found that generally the decree of anadromy within a species was Obsatcry Madt(flcMS!

o Adapt 1 -i'My & < Arvciro S :;; uj -rrcius' d 60 OriticnaUy FA Ioi ! An 50 HHC

rc 3hWit1' ' 40 27 efltS

2SC 4. I S 1.) O?CORHYt'CHUS 0 - ' £ •.• 2C I , tO 1.4 1.1 - 'N z i — r16 d ,4 . 10 1 I • ---'U RIST;VOMEP Figure 1 Species of North American Saimnide classified by degree of anadromy (from Rbunsefell, 1958). 14 greater at the higher latitudes of the species range. He also suggested that the temperature tolerance of young sal.monids showed a significant re].atior.. toth to the degree of anadrorny exhibited by the various species and to certain adaptations in their life histories. In conclusion, not only is there considerable variation in anadromy between each genus of the family Salrnonidae. but also between species within each genus. Also the proportion of anadromous individuals within a particular species varies with latitude rmis only indicates how plastic and divergent the family Salmonidee

is and how difficult it has been to quantify and designate stocks of trout or salmon as strictly anadrornous or

freshwater populations.

/, 15

C. Naturalised Populations of Salmo trutt

During the past century the endemic range of the brown trout has 'been extended : through introduction : to include waters on all continents except Antarctica (MacCrirriinon and Marshall, 1968; MacCrirr'mon, Marshall and Gots 1970) There has been a number of reports of one type of S.trutta being naturalised and leading to the development of the other variety. In most cases the type introduced has been the freshwater form, either the Loch Leven (Scotland) or the von Behr (Germany) strain and this has led to the establishment of an anadromous population. As early as 1884, eggs from Loch Leven trout were sent to

-Newfoundland. From four stocking. sites on the Avalon

Peninsula e progeny of these brown trout spread through salt water to other streams up to 60 miles away and populations of anadromous trout were established (Catt,

1950; Frost and Brown, 1967). 02 interest is the record of a sea trout angled by fly at Witless Bay in 1960 which weighed over 12.5 kg. In Nova Scotia and New Brunswick brown trout of the German vari2 etyc were imported from the United States in 1921. Plantings of this type of trout in the Guvsrough River in Nova Scotia led to the establishment of sea-run populations in both the Guysborough

itself and the Salmon River, the first stream southwards, prior to its planting with brown trout (Catt, - 1950) According to MacCrirmon and Marshall (1968) sea trout which developed from earlier plantings of brown trout -16 have been cauqht. off the Little and Mispec Rivers of

New Brunswick. On the west coast of Canada 3 plantings of brown trout were made in the Cowichari and Little Qua'licum River system of Vancouver Island during the period 1932-1935 (MacCriminon and Marshall Q 1968). Frost and Brown (1967) report that in a number of rivers on the Island, sea trout are now found which are considered. descendants of the introduced brown trout The situation is somewhat confused in New Zealand because of the large numbers of importations of various forms of trout. According to Day (1887) the initial introduction of brown trout led to some descendants resembling sea trout: in habit and appearance. Some of the progeny of the British non-migratory trout planted in the Waiou River, South Island, became aridromous while others retained the appearance related to their back-' round (Frost and Brown s 1967). However 0 Scott (1964) states that the ah,ve information is not based on substantial evidencee as there were also introductions of sea trout made at the sane time. In 1947 the Falkland Islands received as a gift from the Government of Chile,, 30000 brown trout ova listed as Sairno farto which were planted as fry in the rivers of East

Falkland (Arrowsmith and Pentelow 9 1965). This stocking resulted in the establishment of a sea-run population of trout (Frost and Brown 0 1967; MacCrirnmon and Marshall : 1968). As Salrno trutta has been so widely disseminated 17 throughout the world there are undobtediy other populations of the anadromous variety which have result--d from the introduction of the freshwater form. There is also the possibility that freshwater-resident popuit.tons have developed from the plantings of the anadrornous type of trout ORM

D. Approach and Objectives of this Study.

The study of the relationship of brown trout and - sea trout in the Tweed River system was approached from two directions. Firstiy, emphasis was piced on the ecology of the two types by actually studying the population biology of trout in a number of streams tributary to the Tweed. Attempts were made to observe the effect the migratory sea trout had on the resident brown trout. In this way it was anticipated that differences and similarities in the life history of the two forms might be quantified. Secondly, a more basic approach was taken in which the two forms of Strutta were compared using both cellulose acetate and polyacrylamido gel electrophoresis of blood. serum proteins. Also the molecular weights of rLtbosomai RNA (ribonucleic acid) of brown trout and sea trout were calculated in an attempt to find any genetic differences. Finally, artificially produced progeny of crosses of sea trout and brown trout in all possible combinations were compared meristically to find differences in form and structure. The biology of sea trout and its relationship with brown trout has received little attention; Most studies have been based on scale collections from which indirect information has been accumulated. Also the possibility that -brown trout and sea trout may reinforce the numbers of one another appeared to he of importance for the proper management of trout in the Tweed. 19

II. ThE ECOLOGICAL RELATIONSHIP OF BROWN TROUT AND SEA TROUT,

A. Description of the Study Area.

(I) The Tweed basin.

Location and extent The River Tweed rises among the upland moors of the Lowther Hills at an altitude of 520 m. approximately S km.

north of Moffati Dumfriesshire (Figure 2). The river

drains a catchment area of 3385 kin 2 . It flows in a

northerly direction from Moffat for 32 km. then swings eastward near Peebles Peehieshire and makes its way along the Tweed Valley between the Moorfoot and Larmnermuir Hills in the north and the Cheviot Hills in the south flowing for a total distance of 160 km. before entering.

the North Sea at Berwick-upon-Tweed. Both Calderwood

(1921) and Nail (11 .30) give a more detailed accoUnt of the Tweed and its longer tributaries such as the )eithen

Gala, . Ettrick1 Leader, Teviot j Jed, Till and Whit.eadder, Vegetation The upper Tweed region is made up Of Tweeddale ; the Yarrow Water the higher reaches of the Ettrick and Teviot waters' and the uplands between them. The area- is mostly unimproved hill country from' 300 to 600 nt. above

sea level with a subarctic-like climate. The uplands

are essentially open tundras ; largely made up of fescue (Fetuca ovina) and agrostis Moorland (Aqroztis canine) along with heather Callnna vuiqaris) and bracken (Pteridium

gjji1num) . The moorlands are composed of nardus grass Figure 2

The. Tweed basin and location of study, area.,

H The T%tt2d DJr, and, bctionof_StuthjArn 2.W

FiftH OF FO-iTH -t a

C 20 It3ttz , t Lv.zt4 ,

(Nardus stricta), cotton grass (Eriophorum sp,,) and sphagnum muss (S L m. sp.) 3urnett 1964). The valleys of the upper Tweed are marked by improved grasses on the lower bill slopes stretching down from about 180 m and by rotation grasses, hay and fodder crops. The Forestry Commission has acquired large estates along the upper Tweed including the plantationsof Clentress Cardrona, Elibank, Traquair and Yair Hill,

The middle Tweed lies between the upland divide to the west and the fertile lowlands of the east Rainfall is only two-thirds that of the upper Tweed, thus the moorlands are not so wet and improved grasses climb to 300 ra. Rough pasture of Agrostis and Nerdus are used for sheep and cattle grazing. The lower Tweed extends from Kelso to Berwick; the fertile plains taking over from the hills of the upper regions. Intensive acjrituiture is practised in this region, including sheep and cattle grazing. W Geology and physiography The Tweed area is made up of high-folded rocks which have been weathered to masses of broad-topped but steep- sided hills (Craig, 1965). Most of the rocks consist of qrevwackes and mudstones which have been heavily eroded and weathered to give gentle rounded uplands interspersed with steeply entrenched valleys. When weathered out, the greackes and mudstones give siliceous soils which in the uplands are acid and infertile. On lower ground a heavy glacial till is left which is slow to drain. There are broad bands of Ordovician 2

and Silurian rocks that undérlv most of the region.

Towards the east, the valle y is covered with a red or reddish brown till usually sandy with free drainage or sometimes a clay loam which originated from Old Red Sandstone times, d) Climate The weather of the'upper Tweed is relatively cloudy and wet but the lowlands, lying in the east are driEr and more sunny. As altitude increases, temperature drops and snow appears in winter. The average annual temperature at West Linton is 7.1 C while that at Kel.so as 8.3 0C. The length of the growing season (number of days/year when the daily mean temperature exceeds 5.6 °C) at Kelso is 219 days and for the upland areas (300-430 m) to the west is only 175 days. At Kelso there is low annual rainfall which amounts to 63.5 cm. but this increases to 83 cm. in the middle Tweed and as high as 178 cm. in the utter Tweed. Peebles lies in a comparatively sheltered basin and so averages abut 90 cm. of rainfall per year. Most stations record a maximum rainfall in January and a second maximum in August to October.

(ii) The study streams.

The four streams (Kirk, Glentress 9 Grensax and Dead burns) selected for study were situated within 40 km. of Edinburgh and were quite representative of many of the small tributaries of the upper Tweed (Figure 3) All were reasonably inaccessible to the general public, two of the Figure 3

The stikiy area.

•1

mE s'rtai ARFt

( N A Gi2ntrets I Ot5t

Ell FEC BLFS t)

(

N 4c C2rdrcna '1 foT(st

forc,t A - FRY TRAP UA- -RAPS C ° SYWY SZCTKY4 dy 1) —VcIFTRAP / ri —rEktT c7 -

0 1 2 3 S NJ fAj r.tCMiTt '2 L.

streams being situated in the Forestry Comm..ts.sion forests of Cardrona and Glentress. Although at times I was unable to contend with the volume of water in these stre&ms while trapping, for the most part one man could maintain the capturing facilities. These streams were selected because they could be experimentally fished at reasonable cost and they were large enough to possess a full biota. Also they permitted complete counts of all upstream and downstream migrants and thus one avoided sampling errors,

a) Methods Water level was measured by staff Qauges positioned on a stake, so that records were based on daily changes of readings with reference to a fixed level. Altho'20h stream water levels are not directly proportional tci stream discharge, they do indicate periods of high and low discharoe, In Kirk Burn., a more sophisticated water level recorder was

installed which gave a cornDlete continuous record of water level chances. This apparatus was a Munro Level Recorder with vertical drum (type 1H109) which used a Tensator sprina-balance system.

For the purpose of measuring rate of flow in Kin: Burn ? a 900 triangulation (V-notch) weir was installed near the continuous water level recorder. The formula

Q= 2.52 h 2.47 where 0 = flow in gallons per minute and h = height of water above the crest was used to calculate flow for any particular instant. Brannan maximum-minimum thermometers were used to monitor temperature fluctuations. A continuous -water temperature record was c'btaine in Kirk Burn by using a Bacharach Tempscribe supplied with charts which were renewed weekly. The method of Herrington and Dunham (1967) was used to descfibe the habitat characteristics of three of the streams. This is a line transect technique in 'which stream length, width, surface area, pool area, riffle area, depth and stream-bed composition as well as the stability and vegetative cover of the stream banks are described. b) Characteristics of the streams Kirk Burn enters the Tweed from the south about 6.0 km. east aria downstream of Peebles Peebleshire (297389). Glentress Burn flows through Gientress Forest on the north side of the Tweed Valley and joins the Eshiels Burn 1.6 kin, upstream from the point where the latter flows into. the Tweed, about 39 kin,, downstream of Peebles. The Glensax (Haystoun) Burn is the longest stream of the four studied and flows into the Tweed from the south near Scots Mill, 2.4 km. east of Peebles. The Dead Burn is located approximately 11.3 km. north west of Peebles--and flows into the Lyric Water near Romariobridge which in turn, flows into the Tweed 4.8 km. west and upstream of Peebles. Some of the physical characteristics of the four streams are shown in Table 1. The surroundings of the headwaters of Kirk Glentress and Giensax are heather moorland but both Kirk and Glentress 26

Table 1. Some physical characteristics of the study stream.

Catchment Greatest Mean Altitude Stream area length annual flow max - mm 2., (km / (km) (1/sec) (nt)

Kirk 8.1 4.9 68.0 1750--500

Glentress 2.6 2.8 56.7 1 400- 500 Glenthax 22.8 8.5 2150-500 Dead 8.2 3.8 800--700 27 burns then enter plantations of coniferous trees. Cardrona forest which contains Kirk Burn t is composed of predominantly Norway spruce (Piceaabies) ? Sitka spruce

(P 0 sitchensis) Scots pine (Pinus ylvestris•) and European larch (Larix decidua). Before entering the Tweeie the stream flows through agricultural land including sheep and cattle pasture at the edge of the forest (Figure 4) Where the burn flows thrcuqh mature stands of forest trees : the water is shallow and fast flowing with relatively few pools and undercut banks when compared to the section flowing through agricultural land. Glentress Burn flows through Glentress Forest s part of which is made up of young plantations of Douglas fir (Peud9suca taxifolia), Norway spruce and Sitka. spruce- planted in 1958. There is abundant undergrowth in this section. The lower section of the stream flows through mature forest, which was planted prior to 1921 and is composed of Douglas fir, Norway and Sitka spruce d Scots pine, and both European and Japanese larch (Larix 2&2pJRa3 Glensax and Dead burns flow through 'rough pasture land and are for the most part devoid of any forest cover. The physical and chemical characteristics of the streams are summarised in Table 2 and Table 3 respectively. The most consistent predator in the area of the study streams appeared to be the heron (Ardea cinerc.a), especially during the spring of 1972: when three birds were frequently seen at the side of the stream in the pasture section. On two occasions, an individual was observed just upstream of the bcx trap. One must assume that they were predating Figure 4

View of Kirk Burn as it flows northward through pasture land to the Tweed, e•

eg 01

' w.., 1.T• 29

Table 2. Stream drainage characteristics of three suäv

streams tributary to the Tweed (i1Is unDublisl

GJ.entress Glensx Kirk

No, of transects 100 150 40 Stream length (km) 2.82 8.49 4.90 2W. width (m) 1.09 ..,70 2.30 Surface area (ha) 0...31 3i] 1..14 Riffle area (%) 73.8 60.' 50.4 Pool area (%) 26.2 39.6 49.6

Proportion of bottom area by material cas fled rock - - Boulder 3,0 12, 227 Rubble 16.1 46.2 Gravel 64.4 36, 7.9 Silt/sand 16.3 3,0 11.8 Other 0.2

Proportion of stream banks by vegetative types Forest 56.1 16,7 50,0 Brush 3.8 8.7 12,5 Open 40.1 74' 6 37.5

Proportion of tab1e 40.5 86.1 50,0 banks Averace depth (cm) 8.8 22.7 3.8,5 Average gradient: (%) 6.5 2.1 3.9

Table 3. Chemical characteristics of Kirk, Gientress c-md Glensax burns tributary to the Tw(--ed.

Location pH Alkalinity Conductivity C 4g K NO 3-N Org.N P N

mg/i (CaCO 3 ) ( wnhos) mg/i mg/i mg/i mg/i mg/I total mg/i

K I PJ<

8,4.70 7.1 28,0 113 4.3 4.8 1.2 0.60 - 0.015 -

11.6.70 7.6 40.0 122 68 4.4 1,0 0,35 - 0.011 -

23.8.70 7.3 34.4 106 5.6 3.8 0.6 0,,23 .264 0.013 5.0

12.11.70 7.0 32.0

22.9.71 7.7 38.4 120

Total 7.3 34.6 115 5.6 4.3 0.9 0..43 .264 0.013 5.0

GLENTRESS

8.4.70 7.4 32.4 150 5.3 4.8 0.9 1.19 - 0.021 -

11.6.70 7.9 63.0 214 11.8 7..2 2.4 0.56 0.014 -•

23.8.70 7.6 48.0 172 9.0 60 0.8 0.98 .264 0.012 7,4

- -• 22.9.71 7.8 58.4 182 - 0.59 - 0.011 -

Total 7.7 50.6 180 8,7 6,0 1.4 0.84 - .264 0,015 7.4

GLEN SAX 229.7l 7.6 37.2 108 - -. 0.42 - 0040 - C on downstream migrants.. Trout which had been captured in the fyke net placed in Dead Burn in the sprinq of 1971, were found dead on three occasions. These fish had slash and stab marks, which I assumed were due to attempted predation by heron. One morning while walking toward the trap site, a heron was flushed fl- om beside the fyke net and on 1ns)ection, I found seven dead trout with--;n the net. -Herons were found to predate on salmon smolt end p&i r in the area of t:he Burrishoole Fishey, Ireland (Piggins,

1959). In the outurart of 1971, a feral mink (Mustela vision) was scent at ti mouth of Kirk Burn searching both in the water and along the stream banks for prey. A local resident one morning saw a feral mink with a trout of

about 20 cm. in its mouth. Trout and salmon have been reported as important items in the diet of feral mink

in Scotland (Akande 1972). The otter (Lutra 1itra) is also found in the area

and probably credates on trout, While collecting sea trout ova from Quair Water, a tributary of the Tweed

situated 4.8 km. from Kirk Burn, a female sea trout was

found dead on the stream bank. She had bean disemboweled

by a single bite in the region of the vent. Poaching did occur in the Peebles area especially at

the time of the autumn spawning runs. I attempted to

impress on the local residents the affect of poaching and how it would interfere with meaningful results in Kirk Burn.

On one occasion eggs were seen on the bank of the stream

and :i: expect a female sea trout was removed from the burn., 32

B. 11arminologyand. lmethods.

Terms applied to various life history stages.

Young trout which have just batched and still retain a yolk sac are considered aievios Following emercenca from the gravel after four or five weeks and absorption of the yolY sac s the small trcut are desia-rated. fry. At the end or their first year of LLEeD the fry are known

as parr. If the parr are progeny of sea trout parcnts e they will usually turn silver and become smoits at the time of their migration to the sea. Most fish remain at sea for- a year at least, but some return the autumn following their migration. Migratory trout of this 0-winter age croup are termed whitl±nc. In the second summer following migration, the fish become adult sea trout.

Adults, which return from the sea to spawn for the first time are considered maiden spawners whether they be male or female. On the way to the spawning grounds, the sexual products ripen and the fish mature, Ago at maturity is the age at which they first spawn. Just before spawning the fish are ripe but after spawning they are considered spent and called kelts until they have recovered from the effects of spawning and begin new growth.

Scale reading. The developing scale of higher bony fishes shows ridges appearing as nearly concentric rings, which are termed circuiL Generally, the scales start to register growth of the fish immediately after their formation, the circuli surrounding a central spot or focus. They are 33 widely spaced dunn' rapid growth andd cr-s narrowly spaced during slow growth rrhe circuli represerr the icargins of the successive thin plates of which the scale is constructed A prolonged cessation brief interrupion or disturhnce of arowth causes a closer spacing and irregularities of the circuli and is termed a check One that forms ]ween two growing eascns is designated. an annulus. The an:tuius is not completely formed until rapid growth is manifested in the scale itse!f by the wide spacIng or clrcuii. Freshwater growt:h will he used to denote that part of the scale which had formed during residence in freshwater, and sea or saltwater growth to designate the part formed at sea. The anterior field of the scale is usually the more useful area of the scale for identification of annual marks. Spawning is reflected in the scale by a more or less marked erosion or absorption at the scale margin. This

can cause problems in age determination and back-calculation of length as all (Crichton 1935) or a portion of the yearns

growth may be lost on the scale (Backiel and Sych, 1964)

(iii) Designation and recording of age.

At times there has been some ambiguity in the designation

of ages and the recording of scale formulas in salmon (Koo. 1962).

In this study the recommendations made at the meeting of

salmon and trout biologists in Poland in 1933 will he

followed (Anonymous, 1934).

Fish in their first year of life from the time they

hatch until the beginning of the formation of new growth 3 . following the complctio cf to first aani1us are considered O+ From the time the new groh begins, following the completion of the first annulus to the time of completion of the second annulus, the fish is in its second year and its age is recorded by the n'meral

1+. Thus a trout or smolt bavincr laid down threc annuli would be indicated by 3 and 3-:- if any additional growth was in evidence on the scale margin. The alx-'ve procedure is adeaua.te to desianate total age but when one is concerned with separating freshwater end sea life it is desirable to modify the matliod A period is used to distinguish lifo in freshwater (river or stream life) from that in salt water (sea life) The number of winters the adult sea trout spends in salt water is written after the period and the number of presrnolt winters is placed before the period Thus 2.1± is a maiden adult sea trout, with two Dresmolt years followed. by one winter spent in the sea and the scale showing new growth in the particular year of capture.. The letters SM are used to indicate a spawning mark on the scales and so represent a spawning.

(iv) Methods of capture. Trout were collected by a number of different methods including sL- ationarv traps electric Eishing apparatus and a river seine.. The electric fishing apparatus was of the type manufactured by Marine fiectrics of Donegal, Ireland.

Intai]y, the machine consisted of a 12-volt car battery and power converter which were placed on the stream bdnk 35

The electrodes were connected to ti1e power cor!verter by

100 m. of flex, the apparatus produced a direct current at about 500volts so that either the fish mced toward the anode or were stunned in a relaxed state and they could he picked up by using a wooden--handled dip net.

La-Let in the study a more portable "backpack' electric fishing apparatus was used which was made up of the same components as the other but was more suited for Le capture of fish in the small burns (Fiaure 5) This macnine was manufactured by the same c:c.moany but u.sd. two 6-volt motor cycle batteries for power and the complete apax'at:us was placed on a pack frame. The total weight of the pack was

13 kg,,

EJ.ectric fishing apparatus although perhaps the least selective of all methods of fishing, does have a number of limitations which one muse consider. There is seiact:ivi1y for size, fish of small sizes being caught least efficicndy.

The capture of individual fish was difficult at high rates of flow in the small streams, because of the increased conductivity, thus contracting the effective electric field, and, secondly, turbidity obscured the fish from the netter s view,

Boxtraps were constructed to capture upstream and

downstream migrant trout in Kirk Burn. The traps consisted of knotted nylon netting 18 mm. stretched mesh, drawn over

a square wooden frame with dimensions O z 1.2 x 1,2 .z 1.2 m.

(Figure 6). Each trap had a V-shaped entrance and an

inclined ramp so that fish which entered the trap would

find it difficult to escapee The complete width of hc Figure 5

Portable 'aback pack" electric fishing apparatus 36

'•: ;;•

•? .. : I..

- I :--- F.. .

do. Air p, T. Figute 6

Box trap situated in Kirk Burn to capture upstream migrant sea trout: aria bro:n trout in the aueIunn

of. 1972.

Figure 7 Fyke net situated in Giensax Burn to capture downstream migrant trout in the spring of 1971. 37

-4 r, VI

4• 4 - 2 • •i1.. . .- , .*•

-.. -•.d41

r .':

------i1T 1[L

-- 36

stream was blocked by attaching the trap to the banks by means of galvanized wire leaders, reinforced by metal

posts and wooden stabs. Wire mesh of two sizes was used, namely 15 x 20 mm. end 32 x 50 mm. When using the smaller sized wire mesh fish as sm•ail as 9 cm in length could he captured

t times problems were eerienced due to debris such as saw logs, leaves 9 needles and silt, which interfered with the function of the trap. Trap attendance was imperative at this time to clear this material away from the nylon mesh, On a Humber Of occasions individuals escaped over and around the leads of the traps. During the spring of 1971, fyke nets were placed in Glensax And Dead burns in an attempt to capture trout

moving downstream towards the Tweed. These nets consisted of webbing 16 mm, stretched mesh tied to seven hoops.

The diameter of the first hoop was 57 cm. The nets were positioned near the bank of the streams and the single leader 4 m. in length and consisting of 'mesh 20 inn, stretched mesh was secured to wooden stabs running diagonally to the

opposite bank (Figure 7). These nets had to be cleaned and cleared of fish each day. The advantage with using this type of net was that if washed out it could quite easily and quickly be replaced,

A Wolf-type trap (Wolf, 1951) was' used in Glentress Burn in the spring of 1970 for the capture of brown trout

moving downstream Plastic netting, 7 x 7 mm. was placed on a horizontal framework. On the downstream side of the trap, a small trough was constructed which directed trapped fish into a wooden holding box. This trap proved only marginally successful because of the large numbers of leaves and needles which were washed into the trough and holding box. When collections of sea trout smolts were required from the Tweed, a river seine 61 in, in length and 2.4 m

in depth and containing a bag or pocket 0 was used. The stretched mesh size of the nylon netting was 25 mm. in the wings and 10 mm. in the bag.

(v) Treatment of material. The fork length of each captured fish was taken to the nearest millimetre by using a measuring trough. Weights of the smaller trout were recorded from Fesola spring balances. For the larger sea trout, a metre-long measuring board and a Salter 10 kg. :c 50 : •g. spring balance were used.

Prior to examination 1 fish were usually anaesthetised with MS-222 (tricaine methanesulphonate). During the spawning season it was not difficult to distinguish the sex of the adults. Scale samples were taken from the left side of the fish just below the insertion of the dorsal fin above the lateral line. At times it was difficult to take a scale sample from mature male sea trout. The sexual condition of adult sea trout and brown trout was recorded as mature d ripe, partly spent or spent. The appearance of the downstream migrants in the 40 spring was recorded as brown or silver (B or 5) depending on the colour of the fins, type of spotting and the overall hue of the fish. In the early stages of the work, plastic disc tags of different colours were attached through the dorsal musculature of brown trout using stain1ess-steei wire. A hypodermic needle was used to form a wire loop for attachment. An internal anchor tag was employed on larger brown trout and sea trout. The dart tag itself was 5 cm, long and consisted of plastic tubing attached to a perpendicular bar which acted as a barb and was positioned between the intraneural bones (Thorson, 1967; Dell, 1968) These tags could be quickly applied even when hands were cold. A plastic disc tag was also used which was attached to the dorsai. musculature using polyethylene monofilarnent

(Figure 8) The tag was similar to those used by the Freshwater Fisheries Laboratory in Pitiochry and which are manufactured by Charles Neal and Son Ltd., London. Trout were marked by clipping certain fins at each capture. The adipose fin clip was most successful as there was no regeneration whereas with pelvic fin clips some regeneration occurred after only one years Both Stuart (1958) and Saunders and Allen (1967) found this to occur. When interpreting the results obtained from tagging and marking studies, one must he aware of the possible sources of bias that may alter the result. Not only is

S Figure 8 Plastic disc tag and applicator (scaiein inches), 41

1) 2 a j). 7 8:.:*: 42

there 'a possibility that the tag may affcct the growth

of the fish (Shetter a 1967; Canine and Brynlidson, 1972) but also different types of tags,, as used in this study, may give differing values of mortality (KennecJv 1970). and

condition and dispersal (Koshineky, 1972) . Tag loss may be another factor which must be considered. Finc±ippinq

does not appear to alter growth rate ifl raintcw trout (Shetter, 1957) but it may cause increased mortiitv rates (Stott, 1968) For the purpose of calculating growth and determining age of brown trout and sea trout, scales from individual fish were impressed on cellulose acetate strips using

metal rollers (Butler and Smith, 1953) The scale impression was then placed on the stage of a Proj ectna industrial microprojector and the image was projected on a built-in screen. Annuli were marked off on cardard

strips and the length of the fish at each annulus calculated using graph paper. The microprojector was fitted with a

Polaroid 4 x S camera attachment for photocraphic purposes, Scales to be photographed were first allowed to soak for 24 hours in 5% potassium hydroxide and then the mucus coating was removed with a soft camel hair brush. The scales were then dry-mounted between two glass slides, When calculating growth from back-calculated lcngths there frequently is a tendency for computed lengths at a given age to he smaller, the older the fish from which they are computed (Tesch, 1968). This apparent change in growth rate was first described from studies of scales of 43 herring haddock and brown trout by Lee (1912) Not all species or populations exhibit "Lee's phenomenon. Certain workers have found unequal growth (allometric) between the length and scales of trout (Lee, 1912; Sigler, 1952; Cooper, 1953) while others considered growth of the two in direct proportion (isometric) (Jensen 19587 Nicholls, 19587 Kip1ing 1962; Harris, 1970). Growth will be considered isometric in this study. Age and growth determinations were also calculated from length frequency distribution of brown trout and sea trout- •soits found in Kirk Burn. Actual age determinations were made from the scales of those fish whose lengths did not correspond to a particular length group. 44.

C. Characteristics of the Spawning Population

Ci) Sea trout in Kirk Burn.

a) Time, duration and size of the spawning migration,

Adult sea trout (Figure 9 and Figure 10) were first observed in a large pool at the mouth of Kirk Burn, where

the rate of water flow was relatively low. in i970 sea trout, as well as a number of "diseased" salmon, were seen for the first time on 29 October, and the initial sea

trout capture for that autumn occurred at the trar. on 1 November (Figure 11). Sea trout appeared in the mouth of Kirk Burn on 2 November 1971 but none reached the trap, located approximately 900 m, upstream, until

16 November. Alm (1950) also found that 67 par cent of the sea trout ascending the Ava Stream in Sweden to spawn, did so in November, while 25 per cent moved up through the trap in October. Jensen (1968) however found that sea trout ascended the River Istra in Norway in greatest numbers during August and September ; One must keep in mind that the two Scandinavian rive are located further to the north and drain somewhat larger areas,

In particular, the River Istra has a watershed of 70.5 kin 2 whereas that of Kirk Burn is only 8,1 km. 2 . Sea trout, like Atlantic salmon, may move into the larger rivers some eight months before actually arriving at the spawning area. River Tweed sea trout catches by the Berwick Salmon Fisheries Company are greatest in July and early August, but sea trout are not usually observed in the upper Tweed until late September. • 6

Figure 9 Ripe female sea trout, 54.4 cm and 1210 g captured in Kirk Burn on 16 November 1971

Figure 10

Ripe male sea trout, 53.3 cm and 1212 g 0 captured in Omani Water on 15 December 1969. 45

Ar FS11

- 'st 4, - .• •' •

I Figure 11

bveneit of ri.pe and spent sea trout in Kirk Burn 9 1970 and 1971.

C-

!C)

40 1971 M.OVINS UPSTREAM 4 30 WA EU 1.E.VEL 2 20

U 1971 f4CMNG OCMNSTRtM -J D

SEA ThOUT w 70 - 10 MALE 1970 €4) M3AM*3 UPSfltAM 8 FEMALE 50

40 WATER LEVEL 4 30

1970 MYth1NG )OWNR2A'A 44

rnrtnrrr n-rrrrrrrrrrn -i--i--r-r--r-rr-rrr -'- --1 i 10 15 20 NOVE MSER DECEMSER 47'

See. trout in various stages of sexual development were electrofished from burns in the Peebles area (Meldon, Quair, Lyre) from the first week in November to the last week in December for the purpose of egg collection. The greatest proportion of spawning fish in Kirk Burn were captured in the trap on one occasion each year, namely

24-26 November 1970, and 25, 26 November, 1971 Although the traps remained in Kirk Bunt until 18 December 1970, and 30 December 1971, no upstream migrant sea trout were captured after 10 December 1970 and

26 November 1971. Ripe adults were taken over a pertod of 26 days in 1970 and 35 days in 1971. The size of the spawning runs in Kirk Burn were somewhat disappointing as it was anticipated that one could expect at least 100 pairs. The run in 1970 consisted of 35 fish, while that of 1971 was made up of only 16 adult sea trout. b) Age and size of the fish. For the most part adult sea trout captured in Kirk Burn had spent one winter at sea after migration and so were in their second post-migration summer. In both 1970 and 1971 the predominant age class of the fish was 3.1± (Figure 12), 62.9 per cent of the catch in 1970, and 50.0 per cent or the catch in 1971 being made up of this age group. Fish, which had spent only two years in freshwater (Figure 131 made up 34.3 per cent and 37.5 per cent in 1970 and 1971 respectively (Table 4). Table 5 shows the different age groups and mean lengths of sea trout electrofished from other tributaries of the Figure 12

Scale of a 49.6 cm female sea tECUte age 3. l+.,

captured 11 November 1970 in Kirk Burn (x 25). - a - 1st freshwater annulus b - 2nd freshwater annulus c - 3rd freshwater annulus d - 1st sea annulus 48

C b a I Figure 1 3 .'..

Scale of a 49J0 cm female sea trout, age 21+ E captured 27 November 1970 in Kirk Burn Cx 2).

a- 1st freshwater annulus -

'b- - 2nd freshwater einuJ.us c- 1st sea annujus 49

'Wy / Table 4. Age, growth and sex ratio of sea trout captured in Kirk Burn in the autumns of 1970 and 1971.

1970 Age at Mean length Calculated length at end of winter No. of Pet.centage of Sex ratio caDture at capture River life - Sea life - ftsh total M/F 1 2 3 1 2 3 4

2.1± 49.9 7.5 17.3 - 36.4 - - - 12 343 4/8 3.1+ 50.6 4.1 9.8 17.8 34.6 22 62,9 12/10 3. i-SM± 49.5 4.5 12.8 20.0 29.8 1 2.8 0/1

1971

2.1+ 54.9 7.1 18.0 - 36.8 6 37.5 5/1 3.1+ .57.6 5.0 11.1 18.4 33.3 8 50.0 1/6

2.21- 66.3 8.0 22.0 - 37.0 58.5 1 6.3 0/1

3,2+ 53.6 3.5 7.5 18.0 31,0 42.0 J- 6,3 0/1

Ut 0 Table 5, Age, growth and sex ratio of sea trout and large brown trout captured at various

locations on the Tweed in the autumns of 1970 and 1971.

SEA TROUT Calculated length at the end of each winter- No. of Percentage of Sex ratio Age at: Mean tenata life Sea life capture at capture River fish total M/F 1 2 3 4 1 2

- - 27 42.4 14/13 2.1+ 52.0 7.0 17.3 - 35.6 6/12 3,1+ 50.5 5.4 11.8 18.1 - 34.9 - 18 28.3

4,1+ 47.3 4.0 8.0 13.0 17.0 31.0 - 1 1.5 0/1 3/5 2-4 55 0 9• 7.2 17.4 - 34.3 45.8 8 12.1

3.2+ 61.6 4.5 11,1 18.1 -. 33.0 50.8 7 10.6 1/6 0/1 - - 1.5 2.145M± 55,0 8,5 12.5 - 26,0 1

1. - 1.5 1/0 21+SM,l+ 54.0 6.0 175 - - 1.5 M+ 45,8 4.0 10.5 170 -- - -. 1 1/0

}3ROT6P TROUT Calculated length at the end of each Sex ratio Age at Mean length - . do. of winter fish M/F capture at capture 1 2 3 4 6 7

(fl 74. 55.4 6.8 15.4 21,3 29.2 35.1 42.7 50.0 3 0/3 0 52

Tweed (Lyne, Oxnam Eden 8 Kale), it can be seen that rather than the age 3.1+ group. predominating, the 2.1+ class is most common (42,4 per cent). Since these particular fish were taken from streams which are relatively larger and slower flowing than Kirk Burn, the fish exhibited better growth before migration than those of Kirk Burn.

Thus since smoltification is size dependent (Eisen ; 1957), they would migrate earlier. Nall (1930) found that the percentage of smolts that migrated at age 2 was 79.1 per cent and that 3-year smots made up 19.6 per cent, but he does state that since the Tweed is so large one should expect a wide range in growth of parr. The parr of the upper Tweed probably grow more slowly and have a more - prolonged river life. Alm (1950) and Jensen (1966) found that the sea trout which returned to spawn hadDspent on an average 3 years at sea. Sea trout of the 0wergowla River, Ireland, usually return the sane year as migration (Went 1949) whereas the sea trout of the Foyle System in Ireland are generally in their second post--migration summer on returning (Want, 1967) The Black Sea kumzha which are actually S.trutta and not S0salar (Dorofeeva, 1965) are relatively fast-growing and so attain sexual maturity and spawn, for the most part, in their third year (age 2±) after one (frequently incomplete) year of life an the sea (Bsrac.li g 1957) Sea trout, captured in Kirk Burn, had a mean fork length of 52.4 cm. and a mean weight of 1277 g, although one must remember that these fish were nearing spawning 53 condition. The sea trout captured in other streams were on the whole 1arger, having a mean length of 54.5 cm. and a mean weight of 1618 g. These values are smaller than those obtained by Hynd. (1964) for the large sea trout which he obtained from the Tweed. c) Sex composition. The sex ratio of adult sea trout caught in the trap in Kirk Burn was: 1970 1.19 : 1 (female : male) 1971 1.50 1 (female male) and that of sea trout caught by electrofishing in other localities was:

1.46 : 1 (female : male)

The preponderance of females is definitely Indicated. This was also shown to be the case in other studies,

(A1m0 1936, 1950; Jensen 1968; Harbis, 1970 as well as Hessle9 1935; svgrason and Anheden o 1963 as cited in Jensen 1968) Dahl (1910) found when he examined sea trout parr, that females and males were in equal proportion in the youngest age groups. However, among the older fish which had never been to sea,, the males dominated. He concluded that the young females had descended to the sea while some of the males never left the river. He also found that females were more numerous than males in samples of post-migration sea trout. Piggins (1968) concluded from tagging studies, that male kelted sea trout had a lower survival rate than female kelts.

It has been found that other salmonids, which have both a freshwater and anadromous form, exhibit this phenomenon. Wilder (1952) found an abundance of females in "sea-run' Salvelinus fontinalis as did Sumner (1953)

in cutthroat trout, Salmo clarki 9 and Foerster (1968) in adult sockeye salmon, 0n1hvnus nerka. Of interest is the change in sex composition with

different age groups for all Tweed sea trout captured during the autumn of 1970 and 1971.

Age group Number Sex ratio (female male) 21+ 35 1.05 1 38 1.47 1 2.2± 9 2.00 1 3.2+ 8 7.00 1

This would perhaps indicate a higher mortality rate of males when compared to female sea trout. d) Factors influencing the migration. As Kirk Burn is a small tributary of the upper Tweed with relatively low discharges, it appears that a substantial increase in flow is necessary before sea trout will move

upstream for spawning purposes. Although water levels give only approximate values of discharge, sea trout were first observed at the mouth of Kirk Burn in 1970 when the

water level approximated 40 cm. within 3 days and even - though the level had dropped to 35 cm 0 fish were captured

at the trap. In 3.971 the level was low (20 cm) and the- first sea trout was not captured until the first spate

raised the level to 35 cm. When water levels reached 60 cm in 1970 and 44 cm in 197]. t:he Qrcatest-proportaon of trout were captured (see Figure 11) Flows in Kirk Burn were especially low in the autumn of 1-971 so that even though there were fish available in the vicinity of the mouth of Kirk Burn t the number and magnitude of spates were insufficient to stimulate them to move upstream in the burn. The sources of Kirk Burn are situated on upland ircor from which surface water drains quickly after a rainfall. The flow of the burn can rise from 50 1/sec to 400 1/sec and back to 100 i/sec in less than 24 hours. Therefore individual runs were of short duration relative to those of the larger tributaries of the Tweed. On a number of occasions fish moved into Kirk Burn when water levels were high, hut because the level dropped so quickly, the upstream migration of these fish halted and they either returned to the Tweed or remained in a pool until there was the necessary increase in flow. A number of these fish spawned near the pool or were later caught upstream at the trap.. Munro and Balmain (1956) also found that an increase in water level stimulated adult brown trout to move upstream out of Loch Leven, Scotland. Huntsman (1945) experimented with the effects of artificial freshets upon Atlantic salmon and found evidence that all stages of this fish responded to freshets by ascent of streams. Davidson et al (1943) observed that pink salmon (Oncorhynchus grbuha) mill not enter a stream until a spate occurs and that changes in temperature alone seemed to have no effect in promoting 55 migration.

Arrival of adult sea trout at the trap nearly always occurred after, the stream had reached its highest level and greatest flow and was beginning to subside. This phenomenon was most pronounced with the brown trout. Huntsman (1945) and Lamond (1916) also found that ascents occurred as high water was falling. With each substantial increase in water level during a spate, there always followed a corresponding rise in water temperature. This too may have had an effect in Stimulating the upstream movement of sea trout. Both water level and rising water temperature were necessary for the upstream movement of rainbow trout, Salmo gairdneri (Jlartman, Northcote, Lindsey, 1962) and brook trout (Smith and Saunders, 1958) in Canada.

Others have suggested the importance of temperature in the stimulation of upstream migration of salmonids (Stuart,

1957; Gustafson, 191; Mottley0 1938). water temperature appears to he an external Priming" factor which prepares the fish endocrinologically, (Hoar, 1953; Baggerman 0 1960) Sea trout were captured at the trap for the first time when the mean water temperature of the burn was 7.9 °C in 1970 and 6.0°C in 1971. However the peaks of upstream movement occurred when the mean temperature of the water was 6.2 0 C in 1970 but only 3.6°C in 1971. e) Spawning and spawning aggregationä. Two definite spawning areas were located in Kirk Burn that were used each year by adult sea trout. The upper 57 spawning bed was situated approximately 3.. 2 km above the confluence of Kirk Burn with the Twee.d a while the second area was found below the traps 275 m from the mouth of

Kirk Burn. The lower spawning beds extended for 450 rn and were composed of material of different sizes and proportions. Table 6 indicates the proportion by weight and size of particles taken from the vicinity of four sea trout redds. Sea trout were observed moving upstream bath singly and in pairs. Females were usually accompanied by a number of brown trout males, whether a male sea trout was present or not. The male brown trout were all sexually mature and would actively dart in and out while the femUe sea trout was nest-building. Usually a male sea trout was present and at times it chased the accessory male brown trout. The usual number of these male hrotjn trout attending a pair of sea trout was from 3 to 7 (Table 7) On the night, of 25 November 1970 ? I observed, a female sea trout "cutting" a redd below the trap in Kirk Burn. Also present was a male sea trout and three male brown trout (see Table 7). While the female was digging the male sea trout maintained a position slightly downstream and to one side of her, while the 3 male brown trout were also arranged in an arc downstream approximately I at from the male sea trout. On a number of occasions, the 'male sea trout dropped further downstream in an attempt to drive off a male brown trout. While trying this either of the other brown trout moved alongside the female sea se

Table 6. Proportion by weight and volume of bottom material of different sizes taken from the proximity of sea trout reads in Kirk Burn.

Particle size Dry weight Percent of Volume Percent of (mm) (g) total dry (cc) total weight volume-

76+ 3996 25.6 1214 2l.,2

25-76 5406 34.6 1973 34.4 13-25 2876 18.4 810 14.2 6-13 866 5,5 330 5.8

3-6 518 3.3 200 3,5

2-3 476 3.0 190 3.3

0-2 1487 9.5 1005 17.6 .59

Table 7. Spawning aggregations of brown trout and -sea trout electrofisbed from Kirk Burn.

Date Type of trout Sex Length Weight - Age art g

25.117O sea trout F R 57.0 1600 3.1+ sea trout M R 51.0 1150 2I± brown trout. M R 26.9 212 4±

of It N R 22,3 135 2+

it N R 18.8 - 86 3±

2.12.71 sea trout F S 57,3 1800 31+ brown trout N R 49.0 1000 7+

U M R 28.0 228 4±

'I U M R 21,3 108 2+

-. U N R 19.0 79 2-f

11.12.71 sea trout F R 44.6 670 3.1± sea trout N R 42.1 525 2.1± brown trout N R 17.2 56 2+

- MR 24,1 154 3±

11 N R 21,3 130 2+

U M R 28.5 236 4+ N R 214 122 2+

R = ripe S = spent F. = female M = male 60

trout. They were observed to quiver. Although this reflex does not indicate the emission of sexual products (Jones and Bali, 1954) and at no time did I actually see eggs of the sea trout and milt of any male brown trout being exuded, it appears quite possible that the female sea trout could have spawned with a male brown trout. On one occasion, a complete aggregation was captured which appeared to be composed of a male and female sea trout and 3 brown trout. However when the scales of the supposed male sea trout were read this fsn was found to be a large 7-year old brown trout.

Shapovalov and Taft (1954) found that sexual?y mature male stream rainbow trout often accompanied spawning female sea-run steelhead trout. They state that the primary purpose of these male rainbow trout is to participate in the spawning activities. f) Post-spawning hehavicur and appearance. Table B gives some indication of the number of days

spawning sea trout spent above the trapping site. In 1970 51 per cent of those tagged were recaptured while in 1971, 81 per cent of the sea trout tagged at the trap were recaptured as kelts. It is possible that a few sea trout remained upstream after removal of the trap on 18 December 1970 and 30 December 1971. The difference in duration of stay above the trapping site between 1970 and 1971 may be attributed to the lower flows and water level which occurred in the latter autumn. Following spawning 9 sea trout quickly moved downstream to p

Table 8. Number of days sea trout spent above the traps in Kirk Burn in 1970 aid 1971.

Year Sex Number Number Average Range Date tagged recaptured nd. of (days) trap - days out

1970 Male 16 10 11.9 1 to 25 Female 19 8 12.1 2 to 24 18.1270

Total 35 iS 119 -

1971 Male 6 5 6.0 1 to 26 Female 10 8 4.0 1 to 10 30,12.71

Total 16 13 4,8 - 62

deeper 000ls or the Teed itself. In nearly all cases keits were in a weakened condition.

This appeared especially the case with female sea trout Fish which had spent the greatest length of time upstream were usually the most heavily infected with fungus thought to be Saprolegnia sp.

(ii) Brown trout in Kirk Burn a) Time, duration and size of the -spawning migration. The bx trap for capturing upstream migrating fish was placed in Kirk Burn on 16 October 1970 and 28 Seoten her 1971. The first substantial run of brown trout did not arrive until 27 October 1970 but occurred 11 days earlier in 1971 on 16 October (Figure 14). A total of 143 brown trout entered the trap in 1970 while 140 were captured in 1971. Ripe male brown trout (Figure 151 were captured until the trap was removed but by this time relatively few fish were being caught. Upstream movement of fish occurred at 1eat for a period of 61 days in 1970 and 58 days in 1971. Stuart (1957) found that the migration of brown trout out of a small loch near Pitlochry, Perthshire, began as early as 18 October and extended until 7 December in the four years he studied this particular population. Spawning runs in Loch Leven began in early October but extended to the end of January (Munro and Balmain 1 1956) The upstream spawning run of brown, trout in Lake storsj6n, Sweden in 1950 began on 24 June and extended to the third week in September ,Gustafson, 1951) while trout moved out of Llyn Tegid to

• •,.• •--L - ..

E H

-- 4 -- LI

S -• .-.

Figui-e14 '

A mn3mt of bpn trout related to water

1ve1 and water temperature in Kirk Burn,,

1970 and 1971, .

• • C-:- -

• -.--

•Ylt

2 • • - :-- .

I - • .• p•.f- :r - - - .-.. - - - V. •

1 ...... - -.--- - - • ••

4, - • - - 1~. .- .4 . P.. . KIRK BURN 50 N:140 . 4 40 (9 30 L fN jj Aj (9

C 20 £ 24

20 80t Maximum 0 16 60 f? 12 fr1k7C. murn 40 3 lrap in trap out 2.0 3: Ir L_n f_ij nfl r C) GOE 1970 U LI 1 K• BURN 50 ':9 ± 32 J N:43 ...... Waterkvel p C- 2E; -r J 3__) - 24 Ifl 20 20 K BO ' — eO Maxium 60 Mnimurn 40

2.0 'rap out [ LJj 3: 1 a' J nil (-A) 4 rrr_rry_r_1-1_rri- r-i-_i-H_rvnt' _n_j4_rri}.+.4 t-r-rt-y-r-y-rn-i-Pr rr irrrrr$-rnJ ZU 25 30 5 IC) 15 20 25 30 5 10 15 C?9ccmbzr Figure 15 Ripe male brown trout, 26.2 cm and 216 g captured in Kirk Burn on 19 December 1971. 64

r• • . 65

spawn in October and November (Ball arid dories,, 1962). The initial capture of trout occurred when water temperatures in Kirk Burn approximated 6.4°C in 1970 and 7,8°C in 1971. Although Stuart (1957) found that the final upstream run which immediately preceded spawning may depend on temperature there appeared to be a definite relationship between water levels and the number of fish which moved into the trap. Berry (1932) stated that although the most important factor in governing the upstream movement of fish is a rise in water level, temperature and acidity exert an influence. Munro and Balmain (1956) did not find that pH inhibited the movement of fish : but they did state that an increase in water level or some change in the nature of the water associated with such arise, is the most important stimulus to induce the upstream migration of trout to the spawning beds. As found with the sea trout, the greatest proportion of any particular run was captured in the trap after water levels had reached their peak and were beginning to slacken. This observation agrees with that of Munro and Balmain (1956). It was thought that brown trout: might be more inclined to move upstream in times of darkness. Table 9 shows the number of fish caught at various times on 23-25 November 1970, Although the rate of flow had reached a peak at 2230 hr on 23 November, the greatest number of fish caught per hour occurred from 0900 hr to 1400 hr on 24 November. It appears that on this particular occasion, daylight did not inhibit the movement of trout. 66

Table 9, Eumber of fish entering the box trap in Kirk Burn on 23-25 November 1970.

Date Time Discharge No, of Time interval Av, no, of (bra) (1/sec) fish between nan fishings cauiit/hour (h.rs)

23.11.70 1600 181 - - - 2411,70 0015 481 7 6,25 1,12 0500 323 4 4.75 0.84 0900 260 4 4,00 1.00 1400 243 11 5.00 2.20 2000 204 7 6.'00 1.17 25.11.70 1000 181 2 16.00 0.13 1200 175 3 2.00 1.50 2100 .102 0 7.00 0 67

b) Age and size of the fish, The length frequency compositions of brown trout caught at the trap in 1970 and 1971 are indicated in -

Figure 16. On the whole, the trout taken in 1970 were slightly larger than those captured in 1971. Females in both years were older and larger than males.

In 1970 age 3 males (Figure 17) were the most abundant while in the females the age 4 group were the hdst represented. However in 1971 males of age 2 and females of age 3 were the most prominent. (Figure 18) It is difficult to explain why a difference should appear in size and age of migrant trout in 1970 and 1971. There is the possibility thatp since flows and water levels

in Kirk Burn were lower in the autumn of 1971 than in 1970 3 only the smaller trout in the Tweed were stimulated to run-up into Kirk Burn in 1971. Larger fish perhaps eiher waited for higher water levels or moved on to larger streams with greater flows in the vicinity. The estimated mean flow in Kirk Burn for the autumn of 1970 was 176 1/sec while that for the same period in 1971 was only 45 1/sec. One male in 1970 and three females in 1971 were electro-fished from Kirk Burn and they had the appearance and size of sea trout. However when their scales were read it was found that they were 7 year old brown trout and may have been examples of the estuarine ("slob' or tidal) trout of the Tweed (Nail, 1930; Mills, 1971) The average calculated lengths at the end of each previous winter for each age group (Table 10) are quite flgure 16

Length frequency and sex composition of spawning u brown trout captured moving upstream in Kirk Burn autuim 1970 and 1971.: 15

¶0

LL ft

C' -u rn

I 1

C, CO 20 25 30 35 40 45 50 FOR( LENGIH (cm.)

' .nL .-. Figure 17 7 .. Scale of a 24.5 cm male J.rown'tn'it; age

captured 26 November 1971 in iCir} tan (. 30)

a -- 1st annul-us .-• .

b - 2nd annulus

c - 3rd annüliil; A

L - /

4 .. 39

10-1

It a Figure 18

Age frequency distributor of male and female

• • brown trout captured moving upstream in Kirk Burn,

1970 and 1971,, 70

1971 N 14 IMM/ifURE 60 - MALE

FEMALE

40 I I 20 I I in 1. U- 0 C': JELIL_'ItJksJli_ LI C) 4- 21 • Z 60 - igx • P4:143

40 -I If 20 urn • LLLitht t1_J 1 2 3 4 5 6 7 AGE (YEARS.) Table 10. Age and growth of brown trout captured at the lx"x trap in Kirk Burn in the autumns of 1970 and 1971,

1970 Age at Mean length Average calculated length at end of winter Mean weight No. of Percentage capture at capture 2 3 4 5 6 7 at capture fish of total

.1. 2 19.4 5,1 12,1 84.9 39 27.3

3 21.3. 4.2 9.1 15,4 1 18.0 60 42.0

.1 25.9 4,5 9.3 14.8 21.3 200,8 36 25,2 5 38.6 .3.6 8.9 16.7 26.0 32.5 670 0 6 4. 2 6 40.7 5.4 10.1 17.8 25.0 31.2 35.7 746. 5 2 1.4

1971 10.

1 12.9 5,8 25.0 9 6.4 2 19.3 4.5 10.5 91,4 84 60.0

3 22.9 4.6 10.5 16.7 150.1 30 LS, Ar 4 27,4 4.9 10.4 16.5 21.9 234.7 9 6.4 5 30.0 4.1 10.5 17.5 23.4 27.5 301.0 7 5.0

6 - - - - - - - 7 49.0 4.2 11.6 23.8 30.5 37.6 43.0 46.2 1000.0 1 0.7

-4 72

similar for the trout captured in each year, evon though

fish taken in 1.970 were slightly larger overall. The

rate of growth of the brown trout is substantially greater

than that of trout which were captured. in Kirk Burn through

the year at the study section. This would indicate that

the brown trout from the Tweed either hatched in the Tweed

or moved out of the small tributaries such as Kirk Burn in

their first or second summer. I was unable to capture

trout at the trap smaller than 90 cm so could not answer

this question.

C) Sex composition,.

The ratio of male to female brown trout in 1970 was

4.52 : 1 and nearly. double that in 1971 (Table 11)

Although in each year the number of males captured was

quite smilar 0 only 13 ripe female brown trout were taken

in 1971 whereas 23 were trapped in 1970. Again this may

have been due to the lower flows of 1971 hindering the

upstream migration of the larger females.

These findings of an abundance of males do not agree

With those of Runnstrrn (1957) who found a sex ratio close

to 1 1 for trout descending to spawn in Lake Rensj3n, Sweden ;

nor do ray observations agree with Alm (1950) Gustafson (1951)

or Hobbs (1937) all of whom found a preponderance of females.

Munro and Balmain (1956) state that a predominance of females

(as they also found) in the total run appears to be

characteristic of ITIanV saintonid spawning runs. Stuart

(1953) working on the spawning streams of three lochs in

the Tunimel--Garry catchment area found that males were always 73

Table 11. Sex ratio of brown trout captured in Kirk. Burn, in the autumrsot 1970 and 1971.

1970 1971 Sex Sex Age group male female immature male female immature

1 - - - - - 9 2 25 4 10 68 3 13

3 - 51 3 C. 26 4 --

4 26 10 - 6 3 -

5 1. 5 - 4 3 - 6

Total 104 •23 16 105 13 22

Sex ratio (14/F) 4.52 1 8.08 1 74.

more numerous than females in the total run. However,

he did not find ratios as great as those 'found in this study, r--- It appears when one is considering the se c ratios of a freshwater salmonid population which at some time in the life history is in contact with its anadromous

counterpart, there is an abundance of males. Wilder (1952) studying brook trout (S.fontinalis) found that of the fish left in the Mill Brook system of the Moser River., Nova Scotia, after the seaward migration of trout, 71 per cent of those remaining were males. As mentioned earlier, of those that ran to the sea, 72 per cent were female. Barach (1957), found that of the non-anadromous. Chernafa River population of kumzha (Salmo f aria labrax Pallas) 74 per cent were males and 26 per cent were 'females. Ricker (1938) discovered a "residual" form of the sockeye salmon 'which are supposedly wholly the progeny of anadromous parents.

In 193E he found only 63 females among 952 fish caught in Cultus Lake s British Columbia, d) Post-. spawning behaviour. In the two years that spawning brown trout were captured in Kirk Burn, 25 fish NO per cent) were recaptured moving back down in spring having spent an average at 145 days upstream. Gustafson (1951) recaptured two trout which had remained on the spawning grounds for 57 and 81 days. In Kirk Burn, the greater, number of trout were recaptured in April when the box trap was in place to capture sea trout smolts. In 1972 the trap was out from 1 January to

21 February and I expect- that many of the brown trout moved 75 out during this period, when water levels were high. The few females that were tagged as they passed upstream, for the most part returned within 2 or 3 days to the Tweed while the males remained longer.

e) Return to the spawning grounds of previously tagged trout. Four brown trout were recaptured at the traps in

Kirk Burn after having spent a period of time in the Tweed

(Table 124 One male brown trout (BTlll) was first tagged on 24 September 1970 at the study section and recaptured

on its way downstream on 23 April 1971 In the spawning migration of the autumn, 1971. it was captured on 16 November and once more taken in the traps for downstream

migrants on 30 April 1972 Finally this fish was angled on 15 May 1972 at Horsbrugh Ford in the Tweed, 1.3 km below

the location of the traps in Kirk Burn. Although the homing instinct has been demonstrated for other brown trout populations (Stuart,1957) sea trout (Zarnecki, 1966; Shearer, 1959), Atlantic salmon (white, 1936; Went, 1964), steelhead rainbow trout (Shapovaiov, 1941) and cutthroat trout (McCleave, 1967) this phenomenon does not appear to te in evidence in Tweed brown -trout0

- (iii) The (resident) brown trout in Glentress Burn a) Sex composition and maturity. A resident population of brown trout was found by Mills (1967) to inhabit a small tributary of the Tweed situated in Glentress Foit, Peebleshire. Ha found that Table 12, Brown trout which: were tagged in Kirk Burn, migrated to the Tweed and were recaptured in Kirk Burn.

Tag io, Type Date of Place of Age Length Weight Date of Place of Length Weight Sex No. of tagging tagging cm g recapture recapture cm g o.ays from ía c apt un

B]) 174 brown trout 24,1170 Kirk trap(up) 4+ 28..5 235 29.10.71 Kirk trap(up) 29,5 288 339 ST 003 It il 8.4.70 Kirk section A 3+ 20.2 75 6.11.70 'I II 23.7 142 212 PT 111 11 It 24.9.70 1, " 3+ 22,8 154 30.4,72 '(down) 25.6 184 583 J JSL' 6.4.71 Kirk trap(down) 3+ 18.2 62 2,4.72 22.9 116 363

-1 0' 77 the annual production of fish approximated 50 per cent, although values ranged from 9,0 to 3.0kg/ha depending on certain stream characteristics. Ripe male and female brown trout were electrofished from the stream in early October and were present in samples taken in the middle of November. Males in spawning conditiOn were still evident in the stream at the end of February, but all the females taken at this time were spent. A few male fish were found ready for spawning at a length of just over 9.0 cm and a number of female fish ready for spawning at a length of 11,5 cm, Age 2 and 3 trout were the most common in the samples taken in the autumn of 1967. A small percentage of the males that were in spawning condition had only one annulus on their scales while two males had spent as many as •6 winters in the stream (Table 13) . One trout was seven years of age (Figure 19). McFadden (1961), while studying the brook trout population in Lawrence Creek, Wisconsin, found that both age and rate of growth appeared to influence the attainment of sexual maturity. In Kirk Eurn, of the males, age 2 fish predominated whereas age 3 fish were the most abundant in the females. The overall sex ratio showed a slight bias towards the males (1.09 :1), although this ratio does not approach the ratio found in Kirk Burn. b) Growth. The growth rate of brown trout in Glentress was estimated by differences in length frequency distribution at various 78

Table 13 Relationship of male and female sexual maturity to length of brown trout from Glentress Burn 0 1967.

Age No of Percent Mean No: of Percent Mean males length(cm) females length (cm)

0 - - - -

1 12 14.6 8.9 - 35•5 2 34 41.5 12.3 27 13.4

31 23 28.0 16.4 44 57,9 15.8 4 10 12.1 18.6 5 6,6 18.7

5 1 1.2 19.0 - -

6 2 2,4 22.6 - -

Total 82 76

I' Figure 19 r .

• feraalebrcwh trout, ace 7+, c4$Cred'30MOobef 1967 in Glentress Burn (x 30) a:istiannu1us e 5th annulus

– - f 6th annulus

c - 3rd arnalus g - 7th annulus

£ • • H. - 4th annuilus , • ç\ Y

Sb •

• q :.' •

--••-- r 79 ,

4: 4. 4/ • N• \\ > 4 V 1%* I' • /'t >- C

41 K- StVfl ,i ft kij ; F t \

-r':4z, SOC U

capture dates (Figure 20) Although only rough approximations of rate of growth can be calculated, it anpears that the trout in Glentress Burn grow at the same rate as those from Kirk Burn. - 4

IL .

r -

I, L

;

• Figure 20

• Length frequency distributions' oftrout daptüred inG1cntccssBur.

• I t.r4 I,

••.. . ".

1- .4

,...-••-- - •._-...._ . . -•.•,-.•

. . -

• -. . .

+ - - .•

...... -...... V C' 1 OS

Ii! ul

ui J12 J L tD

U) U It ME-

2 6 10 54 18 22 2S .FO*( LENGTH (cm.) 22

D. Incubation and Early Development of Trout Fry.

Two experiments were conducted to gain information on the early development and movement after emergence of trout fry in Kirk Burn. In 1971 an attempt was made to calculate the duration of the incubation period of ova in a sea trout redd, and if possible, to follow the developnent of the alevins until emergence. The rate of development of a trout embryo depends on the water temperature (Grayç 1928). Embody (1934) found that hatchinci success was substantialy lower at ternoeratures less than 5°C and greater than 13 °C. Larval salmonids, depending on temperature may spend two or three weeks

(Frost and Brown, 1967) or several months in the gravel of stream beds (Dill ; 1969).

In 1972, a trap was placed in Kirk Burn to capture a proportion of the fry which had just emerged from the gravel.

It was hoped to find the clate of the peak of fry emergence.

(i) Construction of the Traps A screened pen was constructed which measured 210 x 90 x 80 cm, the bottom of which was open so that when placed in position, the sides extended to a depth of approximately 20 cm in the gravel. The frame was constructed of "Dexion meta]. strips bolted in place and this was wrapped with Tyaan gauze, 7 meshes to the centimetre. When placed in the stream,, the emergent trap was fixed and wired to angle-iron stakes.

On the night of 25 November 1970 a pair of sea trout S3

were observed in Kirk Burn, 70 m from its Confluence with the Tweed. The female excavated 3 redds in an area of about 1 m 2 . The constructed pen was placed over these redds so that the emerging sea trout fry might he captured the following spring.

The fry trap tFigure 21) was placed in Kirk Burn in 1972 just below a section of the stream which contained a total of 16 sea trout redds. These redds appeared in this area between 17 NovemberZ and 8 December 1971. The greatest proportion cf the redds were formed on or about 8 December.

The downstream fry trap was constructed of wire gauze (3 meshes to 1 cm) set on a'Dexion" frame with dimensions

of 100 x 60 x 46 cm. The trap was fitted with a funnel and placed facing upstream near one of the stream banks. A rock leader, faced with flat rocks was constructed so that it extended across and up the stream slightly. Preliminary observations indicated that this diversion was quite efficient in directing fry into the trap.

(ii) Period of incubation and emergence. The fry emergent trap was only marginally successful, as it was found that the gauze used was unnecessarily fine, which caused a reduced water velocity over the gravel within the pen. Any measurement of survival became impossible as only 19 sea trout fry were captured. It is felt that the decreased rate of flow and subsequent deposition of silt lead to the high mortality. These same problems were experienced by Shapovalov and Berrian (1940), and Phillips and Figure 21 Fry trap situated in Kirk Burn to capture emerging see trout fry0 spring 1972.

-a

.. 1 S.- -

.... Titr

Ale j__1 •. Y' '' • . •... i

• •- - ... - .1' • -

.•Ir •

to • - • • - I- •.' - . L Lth.47 .4

• /.- • P. )'. .••'•••---'- -

AV 4&1 85

Koski (1969)

On 25 March 1971, when water temperatures approxirated 5.4°C, 7 alive and 4 dead sea trout aievir:s were found about 12 cm below the gravel surface. The actual hatching of the ova probably occurred very close to this date as the yolk sacs of the alevins were quite 1airge Emerance of the first sea trout took place on 9 May when the midafternoon

water temperature at the pen was- about 9,2°C. If hatching actually occurred on 25 March, then the

period of incubation was 121 ) days.. As the mean ter temperature for this period was approximately 4, OPC : the results agree quite closely With those of Embody 1.9 34)

who found that eggs held at 3.6°C had an ancuration tame

of 118 days. As free-swimming fry were first captured on 9 May, the

alevins must have fed on their yolk sacs for a period of

45 days. Stuart (1953) made a detailed investigation of the effects of silt on ova and alevins of brown trout and the emergence and post larval behaviour of trout fry.

(iii) Size and movement after emergence.

In 1972., when the downstream fry. trap was used, the magnitude and extent of upstream movement of fry were considered to be limited, although no direct attempts at

measurement were made Observations at night with a torch showed fry moving downstream. The first sea trout redd was made on or aboutl7 November

1971, but the peak of spawning activity above the position of the fry trap occurred on 8 December. The fry trap was 86 placed in Kirk Burn on13 April. 1972 and the first fry was captured on 18 koril (Figure 22) when the water

temperature approximated 5 °C. The peak of emergence took place 29-30 April when over 55 per cent of the

total catch was taken.. If I the first fry were from ova deposited on .17 November, then the time interval until emergence was about 153 days. -

Observations made in 1972 suggested that fry movement took place chiefly at night. Dill (1967) found that almost all sockeye salmon fry emerged at night while Elliot (1966) observing movement of brown trout fry, and Mills (19.59b) studying the movement of planted Atlantic salmon fry, noted that the greatest movement downstream occurred during the hours of darkness. Table 14 indicates the numbers, mean lengths and weights of fry captured at the trap. It appears that only newly emerged fry were being captured and were passively moving downstream to areas where they could maintain their position more easily. Since the trap was located so close to the mouth of Kirk Burn, it is possible that a proportion of the fry may actually be swept or move into the Tweed The lower 100 in of Kirk Burn consists of undercut banks, low current, a sand/silt bottom and ample cover for the small fry. Large aggregations of fry -were observed in this section in June and July 1971, so perhaps the fry cured at the trap were moving to a more suitable habitat.

All of the captured fry appeared to be newly emerged, except for those taken after May, when the sample collected 2 E

I .• 1 3 5 S .1

Fture 22

Numler of sea trout Cry captured in the fry trap

Butfl-spring 1072; Si CO tr Sr * -C

r; I -

- - r

r 13'i

U 250

IVATER LEVEL 200 ID 150

100

M. IvtAXMUM - 4.fl

3 I--

C C MINIMUM 4 40 Cr 2 Li. t 0 i-

'A a:

20 trap out z trap Ifl /\

15 20 25 30 5, 10 15 20 25 APRIL MAY 38

Table 14. Mean lengths and weights of newly emerged

trout fry in lUrk Burn s 1972.

Date No. of fry • Mean length Mean wet weight CIII. g.

19.4.72 9 2.73 0.155

21 10 2,77 Q..146

23 26 2.52 0.103

26 73 2,65 0.116

28 38 2,66 0,118

30 154 2,72 0.130

2,5.72 18 267 0.125

4 36 2,70 0.140

6 53 - -

8 23 2.56 0.120

13 37 - -

16 74 2.82 0,153

24 62 2.86 0.157 69 contained individuals v'hich were slightly longer and heavier than those of any previous cci Lecion. While studying the movements of. young Atlantic salmon, Saunders and Gee (1964) found that fry remained within small areas during the SUIrUTLer 1 suggesting the establishment of territories. The fry in Kirk Burn may have actually been searching for more suitable habitat ; i.e. better cover, slower current where they could more easily maintain their. position; or they may have been dispersing because of the limited area available to them (Backiel and Lecren, 1967) or as Kalleberg (1953) suggested fry disperse when they begin to feed and the feeding behaviour exhibited an increasing specialization by being directed towards open water." He also sugàested that the velocity of current caused young salmon to leave exposed stations., In Kirk Burn, the area where the sea trout redds were situated containd few pools when compared with sections of the burn further upstream. Fry would find relatively few areas where they could maintain their position without difficulty. There appears to be some relationship between the numbers of fry captured in the trap and water level or discharge. The results are not conclusive as during the increased flows, the efficiency of the trap was diminished due to silt and detritus which interfered with function of the trap. Perhaps greater numbers of fry were swept downstream but away from the trap in high flows. Mills (1969b) captured the greatest number of planted salmon fry when there was a rise in water level and increased flow. Acara and Smith go

Dund that for the most part greatest fry -movement e, Oncorhmchus nerka, was associated with

dis char cTe. ci

E. Trout in Kirk Burn Prior to Downstream Migration

(1) Characteristics of the population. An area was selected in Kirk Burn which was sampled each month from April 1970 to March 1971, excluding the month. of December 1970. The study section had an area of 201.8 ra and was situated 2,6 km ahive the confluence of Kirk Burn and the Tweed (see Figure 31) . The section was located in Cardrona Forest, where cover consisted of Sitka and Norway spruce with little or no, streamside vegetation.

- The fish were sampled each month using an electrofishing machine. Some of the problems associated with electrofishing were mentioned earlier. Since it appears that the population of trout studied consisted primarily of progeny of sea trout, a strong downstream movement of trout between certain sampling periods occurred. This was especially evident in the months of March, April and May when individuals that were marked and tagged in the study section were captured further downstream at the trap. Needham and Cramer (1943) also found this, although they suggested that the trout moved less than two miles. Downstream movement will affect population parameters such as production, survival, population size and mortality - (Mills, 1959a)., although movement into the sections sampled may well counterbalance emigration from them (Nedham, Moffett and Slater, 1945) Age determinations were made from both monthly length frequency 4.istrthutiois and scale samples. The length range 92

for each age as calculated from both methods was similar. There was a slight overlap between certain age groups as determined by length frequencies so that a number of trout were aged by microscopic examination of their scales. The birth date of trout was considered to be 1 May. Population estimates were made using the formula of Seber and LeCren (1967) where two successive catches

C 1 and C 2 are taken with the same effort from a population. An estimate of the size of the populatton 0 N is given by

C - C 2

with a variance

C + c 7 ) varN= 1 2 ___ In - C '1 2'

An attempt was made to use the population estimate method of flicker (1958) after Bailey (1951) in which the

number of marked animals are treated as a random variate 1 but it was found to be unsatisfactory. The number of fish present at any one time is affected not only by survival but also immigration and emigration and it usually is difficult to separate these three parameters (Mann, 1971). The total survival rate (3) has been calculated from the formula

s=Nl/N

where N 0 = number of fish present at the beginning of the season and N 1 = number of fish present at the end of the season. 93

Survival rate was also calculated from a marking experiment in which trout, electrofished from the section, were differentially clipped in three successive months, namely March April and May. The survival rate was again calculated using the above formula but since each fish was marked after capture the annual survival rate could not include immigration into the study section.

• (ii) Age and growth calculations The length frequency distributions of trout captured each month in the study section are shown in Figure 23.

The modal values of fork length in April 1970 were 5.0 cm for the 0+ group, 8.5 cm for the 1± group and ILOSL of those fish with a fork length grnater than 10.5 cm belonged to the 2± group. For the most part, the only instances that trout showing more than three annuli were electrofished from the section, occurred from the months of September, through April when male brown trout from the Tweed were present in ripe or spent condition. The growth rate of trout in Kirk Burn is substantially less than that recorded for brown trout from other areas of the world (Schuck , 1945; Hobbs, 1949 ; Lane, 1964; Chapman, 1966; Marshall and Maccrinurion, 1970), where the populations studied inhabited larger bodies of water and food was abundant and varied. On tine River Teify and its tributaries, aobes (1970) found that the larger the water, the larger the average weight of trout caught. The growth of brown trout in some streams and rivers of the British L

-- 4'..s

-,.,...--.t-w_...-.._..-., ..J...... .. .,...... ,- . ii

4. . . •1

r 1 4

. • Figure. 23 .. . r • 1 trout

apt trc&in Kirk Burn, 1970-71.

.ti4.44 .1 - . •

r . .

......

• t-t...' - 1 1 t -

--'c

rr-

"' -k- - 94

- AVRk. ;0

8 i1L1[i w:lt3 4- I

- 12 I. MAY 197C 8- :J 4--1 jr t4 :89 I Lj

- JUNE. 1970.

- N.269 6-

jwx 197t- B- r fl 4- IJ N = 124 F--- 16 - Iii AUGUST 912- we- 1 -, - - -J Uw ----H o 12 J - ;c. N 161 th

12- OC10ER 1970

p:i29

lu ?4DVEMBER (t' 12 Lu I N129

[I, JANUARY 1971 B 4 L-L~~D-n 20 16 FEBRUARY 1971 12 W;10i S 4 -- /

16 MARCH 1971 12 - H B 4

2 6 10 14 IFI 22 25 3-0 FOR( LEJGTH (cm.)

V 95

Isles has been recorded by Frost (1945), Horton (1961),

Frost and. Brown (1967), Egishaw ( 1970 ). The calculated growth of trout caught by angling and electroflshinghas been shown for the trout of the Tweed and its tributaries (Mills, Clelland and Osborn and Watt, 1972) The length frequency distributtons of trout electrofished in February 1972 from three diffetent sections of

Kirk Burn are shown in Figure 24. In the upper or highland moor section, age group 1 trout are the most abundant and very few age 0± individuals are present. This may be due to the downstream movement of this age group after emergence because of the lack of suitable habitat or food. There appears to be a diffetent growth rate for trout from each of the three sections. This was also found by Allen (1951) The average length and weight of young-of-the-year trout is shown in Table 15. The growth in length proceeds until October when it decreases probably due to emigration downstream of the larger individuals of this age group during the winter months (Egglishaw, 1970). At the time the first annulus was laid down., the fish averaged 5 cm in length. By October of the first year the average weight of the fry had doubled but then decreased until early March when an increase occurred.

(iii) Population estimates It was found that when attempting to obtain an estimate of the population size in the study section, the method of

1 1 Bailey (1951) on the three occasions it was used, consistently I

Ficure 24

Length.fçequency distribution of trout captueft J

• in the 'Sections of Kirk Burn, February 1972

- 2 C;-o

41 w U 2 14j rX cc

8 15 U- 0 23 FEBRUARY 1972 >10 I

M LL 11 & 15 bl 0- 1972 10

2 6 10 14 18 22 26 FORK LENGTH (cm.) I 97

Table 15. Number, mean length and mean wet weight of age group 0 trout captured each month in•

Kirk Burn at the study section, 1970-71.

Date of Total Average length Average wet capture number (cm) weight Lg)

15 May, approximate date of first emergence

26 June 150 3.5 -

29 July 208 3.8 0.56 28 August 146 4.2 1.21

24 September 91 4.3 - 29 October 46 4,8 1.24 20 November 66 4.7 i18

22 January 31 4.2 1.16 11 February 48 4.5 1.00 30 March 35 5.0 1.37 98 overestimated the numbers of trout present Nhen usinci the method of Seber and LeCren (1967) estimates agreed more closely with the numbers of trout actually captured. The amount of immigration into and emigration out c-f the study section in March, April and May made it more difficult to quantify population size and survival.

(iv) Annual survival As mentioned earlier it is difficult to assess the effect of immigration and emigration on survival and mortality

(Northcote 1967). Annual survival rates calculated from the formula Nlto are shown in the followingu table:

Age group Annual survival rate Date - o - 1 0.15 July 1970 - June 1971

1 - 2 - - 0.36 April 1970 - March 1971 2 - 3 0.47 April 1970 March 1971 3 - 4 0.04 - April 1970 - March 1971

However the annual survival rate of marked fish using the same formula but not allowing for any immigration is shown in the table below:

Age group Annual survival rate Date o -1 - 1 - 2 0.16 April 1970 March 1971

2 - 3 0.09 April 1970 - March 1971

3 - 4 0.04 April 1970 - March 1971

- As suspected, an influx of second and third year fish occurred from further upstream. If this is the case, then 99

since immigration supposedly equals ernicrationa large

number of fish on their way to the Tweed will have left

the study section. Such a movement has been observed

by Horton (19611 and Needham. Moffett and Slater (1945)

The survival of fry from July 1970 to June 1971

appears somewhat high (15.4 per cent).. Pefnaps this

value would be lower if survival was considered from the

time of emergence, when fry mortality for the early months

would be greater. McFadden and Cooper (1964) divided

the post-emergence life span of brown trout into two

distinct periods on the bests of differences in survival

rates. The first few months after emergence survjval

is relatively low, perhaps from 2 to 10 per cent. The

remainder of life annual survival rates have been reported

to range from 14 to 87 per cent for various age croups under a wide range of environmental conditions (McFadden and Cooper, 1962).

The total survival rate of trout between the ages of 0 and 3 in July population estimates using the method of

Ricker (1958) was 0.34 or 34 percent. This value agrees

with that reported by Schuck (1945); Cooper (1953) D McFadden and Cooper (1962) and Marshall and MacCrirrLmon (1970). 100

F. Downstream Movement of Juvenile Trout

(i) Introduction and methods There have been very few studies on the ecology of young sea trout before they migra€e to the - sea as smolts

and the actual dynamics of this migration. However much information has been gained indirectly by the examination of scales taken from Adult sea trout (Nail, 1930;

Menzies, 1936; Went, 1962). More recently, Harris (1970) studied the sea trout smolts of the Afbnau Dyfi. and Dysvnni in Wales and Jensen (1958) captured sea trout smoits in the

River Istra of western Norway. By relying on stationary traps and nets in tributaries

of the TwcedD it was thought that more information could be obtained on the actual duration of the smolt migration and the factors which caused the day to day fluctuation

in numbers captured. In 1971, a number of smolts were tagged and although no records of recapture have been

reported to date : it is hoped that information of this nature will be accumulated over the next few years. In the spring of 1971, three trapping devices were placed in three tributaries of the Tweed in the Peebles area. Fyke nets were situated in Dead Burn and Glensax Burn while a box trap was placed in Kirk Burn. In 1972, a box trap was again stationed in Kiry Burn but at an earlier date and also the trap was slightly modified to assist in the capture

of trout. A removable grid was placed in one of the leads which facilitated easier cleaning of the trap in times of

high water levels. A greater number of migrants were ]:O1 captured in .1972 in Kirk Burn because each time the trap was cleared all trout located in the pool upstream of the trap were electrofished and included in the catch for that day. Over the two years D a total of 2018 dowñstrpam migrant trout were captured in the three streams (Table 16)

In 1971, 1 attempted to visit the traps daily but due to the time and distance involved, it was not always possible. The fyke nets of Glensax and Dead Burns in 1971 were fished more consistently day to day than was the trap of Kirk Burn in 1972. For the most part,, fish found in the box trap were caught over a two day period.

(ii) Appearance of downstream migrants Captured trout were recorded as having a brown or silver appearance. This classification was somewhat qualitative and rather arbitrary since varying degrees of brown and silver occurred. A migrant was considered 'brown' if no silvering or other sea trout smolt characteristics could be distinguished. Those trout which showed any of the following characteristics (Figure 25 1 were considered sea trout srnoits:

lack of parr marks lack of rusty red or orange spots with surrounding halos the pectoral and pelvic fins are yellowish a black line appears a short distance inside: the margin of the caudal fin the adipose fin is yellowish-brown rather than reddish a silver appearance on the sides with black spots white undersides. 102

Table 16. Number of downstream migrant trout captured

by box trap and fyke net in 1971 and'1972.

Location Duration Method of Number capture

Glensax 30 March 4 May 1971 fyke net 595

U Dead 25 March - 4 May 1971 159 Kirk 6 April 9 June 1971 box trap 343

U II 0• Kirk 17 March - 28 May 1972 -I C.

Total 2018 Figure 25. II Sea trout smo}t•, 24,5 cm, seinnetted from the Tweed at. Norham Bridge, 6 May 1971.

Figure 26

Downstream migrant trout captured in GIensax Burn by fyke net on 18 April 1.971 (Scale in inches).

top: silver appearance, 15.9 cm: 41 g, bottom: brown appearance, 15.1 cm ! 36 g. 103

21 31 45-J64L7 8 1.04.

Figure 26 shows two migrants captured in the f4ce net of Glensax Burn, the bottom fish was recorded as brown in appearance.

(iii) Time and duration of the migration In .19716. trout moving downstream were not captured in any numbers by the fyke nets in Dead or Giensax burns until 10 April (Figure 27). In both streams, a substantial increase in numbers taken occurred when the maximum daily temperature-surpassed 10°C. Trout: were not capturEd in Kirk Burn until 19 April when water temperatures at the trap exceeded 8 °C. . In 1972 the first increase in the number of trout taken occurred more than a month earlier on 15

March when water temperatures reached a maximum of 6 °C (Figure 28). The downstream movement, of trout extended in 1971: over the period 3 April in Glensax Burn and 27 March in Dead Burn until the nets were removed on 4 May, In 1972, the trap was placed in Kirk Burn at an earlier date and remained until 30 May. Fish were trapped in greatest numbers an 22-24 April 1971 in Glensax and Dead burns and on 2-3 April 1972 in Kirk Burn when, in all cases, there was a substantial increase in water level. Once fish haye reached the correct physiological

condition for migration (Bagerman. 1960 1 , there are a number of physical and climactic factors which have been suggested as influencing the. timing and intensity of the downstream migration of saimonids. There is considerable evidence that supports the hypothesis that water temperature ; a

t . ..

¼ -

L. .

- .-..--.-- V .4 • - - 1 . • . .. F .- .. -V..- ' ••• .• - •

:;.-•- ; -

Figure 27 , .. . - Downstrea-mimovemdnt an5 appearance of trout

It captured in Glensax , and - Dead burns, 197f.

• -. ,.

,,•.• .'. -, ,..

4- ,

F- &M8ZR OF F1514 8 I I

i-I C I I.. '1

'II p 1 hi /

NI p 1 > I / I F" / ID I

?4.

1' \ I

U U L r S WATER tF.MrrATu,RE (C) WATER LEVr.. (cm.) wAtEQ 1EMr•aI/nr;'a ,'C) wtiEn L(:vEI (cm ) • .• .,

4 ••. • - •--..-••.- • - -. • .-•- - .- •-- ••

) • S

4 .•'•. -

I - - -. -•'.-.

F3gLe 28

Downstream movement eCiad appearance of trout

9,' - • 'no

2!'C)

ao4 200

Lb trio

ISO

140 - 0

4AXIMUM

'20 - I-:

I' Iii Lb o 100 C- Zr La a) N z 3.- Lb 2 ui 3-

4!)

20 TRAP I 'ThP CUT / C)

MAFCO APRiL MAY 107

is of major importance in inatiati ;ng smolt migration (White 3940; Allen, 1944; Northcote, 3962) Both Mills (1964) and Osterdahi (1969) found that the main part of salmon smolt runs occurred when water

- 1 temperatures had risen aoove j O0 C. Stream discharge may also have an affect on the intensity of migration of young trout (Berry, 1932; Gustafson, 19517 Mills, 1964) Other factors such as photoperiodisrn (White, 1940 Hoar, 19537 Northcote, 1958; Eaggermarn, 19601 or perhaps a combination of temperature.. pH and content of dissolved gases of the stream (Krogius, 1954) may be necessary to cause migration. In this study the two factors which appear to 'trigger" migration response are water temperature and water level or discharge. A water temperature of 600 in Kirk Burn initiates the downstream migration while a rise in water temperature intensifies the response. Over one 24-hour period the net in Giensax Burn was cleared every four hours. , Approximately 90,.5 per cent of the migrants entering the net in that period were captured between 2000 hr and 0400 hr (Table 17). In the course of this experiment, the water temperature decreased from 11,6 to 8.-2°C but the water level remained relatively constant. The tendency for juvenile salmonids to migrate in hours of darkness has been recorded by Northcote (1962) and Alexander (1970). This did periodicity may alter depending on whether the observations are made early or late in the run (Munro, 1965; Osterdahl, 1969) Table 17. Downstream movement of trout in Glensax Burn during a 24-hour period.

Date Time Number Ternerature Water (hrs) Salmon Trout ( °C) level

22 April. 1200-4600 5 8 11.6 37 1971 1600-2000 3 1 10.4 43 2000-2400 13 56 8.2 58 23 April 2400-0400 3 87 75 52 1971 0400-0800 0 2 7.1 56 0800-1200 1 4 8,2 45

fA- I flC .1- 1 -,

(±2) Age and size composition Of the trout which were captured moving downstream in 1971, approximately 80 per cent of them from Kirk, Giensax and Dead burns had spent 2 winters in the stream before migration (Table 18). in Kirk Burn in 1972, a greater percentage of fish of ages 1 ; 3 and 4 were taken giving a lower percentage of age 2 fish. This was probably due to more complete trapping records, as the trap remained in Kirk Burn for a greater length of time and functioned more efficiently in 1972.

Jensen (1968) found that in the Istra of Norway, over 95 per cent of the srnolts were three or four years old

C at migration. In the Am Stream of Sweden, most of t:he smolts descend at an age of 2 years (Alm s 1950). The age of smelts in the Northern Baltic area, as reported by AIm (1950), is considerably higher. In Ireland, Went (1949) found that 2 year smolts comprised about three-quarters of the total catch in the Owengowla and about the same in the Inny (Went, 1948)

In the Tweed itself, Nail (1930) showed that out of 3197 fish, 79 per cent migrated to the sea as snD1ts after 2 years of freshwater growth but stated that, for the most part, sea trout from other Scottish rivers remain in freshwater for three years. Of the sea trout smoits in Wales, 58 per cent of them in the Dyfi and 83 per cent of them in. the Dysynni spent two winters in freshwater before migrating to the sea (Harris, 1970).

The older and larger migrants moved downstream earliest I 10

Table 18. Age composition by percent of downstream migrant - trout captured by trap in three tributaries of the Tweed, 1971 and 1972.

Location Age (winters) Total 1 2 3 -4 5 number

C-lensax - 79.7 19,2 ibO - 291 2 April - 4 May 1971

Dead - 80,7 17.1 2.1 - 140 25 March-4 May I

1971 I

Kirk .. - 80.6 16.0 3.4 - 294 6 April-9 June 1971

Kirk 4.9 64.4 24.9 4.9 - 205 17 March-28 May 1972 Ili

in the season (Figure -29, Figure 30) This phenomenon was also observed by Menzies (1931) Pentelow, Southgate and Bassindale (1933) and Harris (1970) . Alien (1944) considered the relationship between size and .micration of salmon smolts to be extremely complex, hut he did suagest that there is a relationshio between size and cmolt development which causes a salmon to migrate in response

to certain external stimuli

Trout which were captured in the fyke net in Dead Bern had a modal length of 14.0 cm and were somewhat larger than those of Glensax Burn, which had a modal fork length of 12.5 cm (Figure 31). The fish of Kirk Burn, taken at the box trap in 1971, were smallest (nodal lcngth of 95 cm)

but this may he biased because of the escape of the larger migrants before the box trap was placed in the stream A greater proportion of trout captured in Dead Burn. had a silver appearance which would indicate that there is

a relationship between size and silvering in sea trout Sif101ts.

A very small percentage of the migrants trapped in 'Kirk Burn' in 1971 showed any silvering characteristics. In 1972 the downstream migrants trapped in Kirk Sum were on the whole, larger than those captured in 1971. The modal length of the trout in 1972 was 12.5 cm and a greater percentage of actual smolts (csilverh) entered

the Lox trap. Both Jensen (1968) and Alm (1950) collected sea trout srrD1ts which were longer, but these fish were taken near the sea after they had left their nursery streams.

Nail (1930) calculated that sea trout smolts having spent a

7T jWl

.Fiqdxe 29

CTIänàe in age composition of downtrectrn migrant

inKirk Burn, 1972

-4

I -• C.

100

(I) LL LGO 0 0:

40 D

II IC tar 1 I Ti IlL. TZ' A Ut . - fl G CLASS H C

4 I

Figure-SO

Change in ienth frequency and appearance of H downstream migrant trout Ira Kirk Burn, 1972

•1

kilts-

4.-

4- :

- 13.3

• ..• .•• to • 4

. 5 H 46

40 • . r1fl- AP1Lji

APflARANCE • . • . - --••. 30 • - r-t.v I SILVER 0 i BR3WN - -C t •. .• 2C

a I- ru 2 - •> jm-- E I J I z

30 • . MAY •

20 • -

6 b ;o 12 14 ¶6 le 20 Fork Length (cm) I •1 I

••J- . t 1t44 I . 1

Z I t -$

11 -S

figure 2131

Length frequehcy ,4istribuQion and appearance of

trout captured moying downstream in three burns

tributary to Sppvmlweed, J971.

-• ...... r'--'

4 .

.. .. 1.14

20 15.

30 I 'I) LL o 20 1

10 z

4 6 B 10 12 14 is j3 20 22 24 FOFIK LENGTH (cm.) 115

2 years in freshwater had an average length of 18. 3 cm.

Two types of smolts have been distinguished on the basis of growth rings at the cuter margin of their scale

Went (1938), looking at scales of salmon, classified these types of smoits as:

Type A: those showing little or no freshwater growth before migration, the outer margin of the scales consisting of narrow winter rings or less than 2 summer growth rings. Type B: those showing a fairly substantial amount of

freshwater growth before migration s the outer scale margins showing more than 2 summer growth rings.

Sea trout populations in Ireland studied by Went (1962) were found to have varying proportions of the two types

but the proportion of Type A smolts never exceeded 27.2 Der cent. That is, most fish made substantial growth in freshwater before descending to the sea. However, over 88 per cent of sea trout smolts migrating to the sca in the Dyfi and Dysynni were considered by Harris (1970) to be Type A smolts

All scales taken from downstream migrants captured in the box traps in Kirk Burn and the fyke nets in Glensax and

Dead Burns showed no new summer growth rings. On 22 March 1971 a sample of sea trout smolts was electrofished from Eddleston Burn, All of them were Type A smolts showing no freshwater growth, as it was at least a month too early for the scales to show + growth in any case. Smolts seined from the Tweed at Norham Bridge on 6 May 1971 had 116 the following percentage of types:

Type A 38.8 ?4 Type B 61.7 %

It is probable that smelts from the upper Tweed add substantial growth as they migrate towards the sea; thus if captured near Ber-wick-upon Tweed at the mouth of the Tweedg. these fish would be considered Type B smolts. Pente].ow, Southgate and Bassindale (1933) stated that it was the smaller smoits which added extra growth0 The calculated lengths at times of annul! formation for migrants captured in Kirk, Glensax and Dead burns are indicated in Table 19. Figure 32 graphically depicts the calculated growth rate of trout in each of the three streams. The growth rate of fish in Dead Burn is greatest, as it experiences higher water temperature, a more gradual profile and flows through agricultural land.

( v) Sex composition The length frequency, sex composition and appearance of trout captured in the Tweedand a number of its tributaries are depicted in Figure 33. A preponderance of females appeared in each sample except that taken from Kirk Burn on 9 May 1971 117

Table 19. Calculated lengths for each age group of trout moving downstream in the three study streams, 1971.

KIRK Smolt age Length (cm) of smc'lt, No. of fish (winters) age group (winters) 1 2 3 4

2 5.9 11.8 34 3 4.5 10.1 14.4 29 4 3.9 9.0 14.0 18.3 13

Total 5.0 10.7 14.3 18.3 7

GLENSPX

2 5.8 14.0 43 3 5.2 11.7 16.3 34 4 4,3 9.8 14.1 19.6 3

Total 5.5 12.9 16,1 19.6 80

nat

2 6,2 13.5 13 5,0 11.2 17.4 13

4 3. 6 11.3 17.2 22.0 3

Total 5.4 12,2 17.4 22.0 29 N t;tflL;Et H' C

Figure 32 ,

Calculated gro€h. rac of trout captured movihg

• downstream in threburiis tributary to the 1971. • • 4- •-t -. - •

- :-. . C

DEAD BUFN

20 GLENSAX LIUWN

KRK SURN

16 E U F 0

2 z

Lii

1 2 3 4 AGE (YEARS) t

4 .

4 'Cf ,••.t• t3.

.4 C

_1 I

Figure 33

Length fyequenqyj apperanceandsex composition

of suppbsed se trout smolts 1 captured at'

various r locations on the Tweed s

SO 1 10

La- La- 0 at 1] 0 10 z .5

i--i F-'

(3 3 10 12 14 10 F- 2022 2.4 25 5 0 1.2 14 151320222426 LENGTH (cn) LENGTH kin)

120

Location Sax ratio female / male Eddleston 1.93 / 1 Dead H i.24./ 1 Norham (Tweed) Kirk 0.71 / 1

The deviation from the 1 1 ratio with an abundance of females has been shown by Meek (1925) Pentel'Gwe Southgate and Bassin-dale (1933) and Harris (1970) in Britain. Dahl (1910) found when he- examined sea trout parr that females and males were fairly evenly distributed in the youngest age groups. Among the elder fish, which had not yet been to the sea, the males dominated. He. concluded that the females had descended to the sea, while some of the males were left in the river. In accordance with this, he found that females were more numerous than males in samples of post-migration sea trout. The sex distribution of descending sea trout smolts of the River Istra in western Norway was approximately 1. 26 females to every male while Aim (1950) captured 2.84 female. sea trout smoits for every male smolt.

(vi) Downstream movement of the "resident" trout

in G1entress Burn s The trapping grid was placed in the Wclf-type trap in Glentress Burn on 4 March 1970 and the stream was visited at least at weekly intervals until 18 October. On a number of occasions when stream discharge was at a maximum, the trap and holding box became choked with debris. I

Any- fish which were moving downstream at this i:±m.s would have escaped. A total of only 14 trout were captured between 16 May and 26 June. No fish were found in the holding. box at any other time, The average fork length of the fish was lots cm. All of them were age 2 except two which were age 3. In the H0L'okjwj Stream of New Zealand, brown- trout showed no evidence of any movement (Allen, 1951) Mills (1967) studying the production of trout in Glè-ntress Burn found that a year after completely clearing -a section of the stream of trout, trout were again present in the

section giving standing crops very near the original value s thus indicating downstream movement from immediately upstream of the experimental area. The number of trout captured in the trap indicates that movement in Glentress Burn is minimal or at the Very least certainly does not approach the amount of downstream movement exhibited by trout of Kirk, Glensax or Dead burns. ] 22

G. Movement of Tagged Adults

(i) Sea trout kelts Following capture and tagging in the .Peebles area,

only 3 sea trout were recaptured by fishermen. Two fish were angled downstream and appeared to be on their way to the seat One was tagged in Kirk Burn migrating upstream on 2 December 1971 and was angled as a kelt approximately 84 km downstream in the Tweed at Norham Bridge on 14 March 1972 The second sea trout was electrofished from the MeiQon - Burn on s- December 1971. and recaptured as a kelt in the Tweed 8 km west of Berwick only 1$ days later on 19 December 1971, The sea trout showing the greatest movement was one tagged moving upstream in Kirk Burn on 11 November 1970 and recaptured 252 days later off 'Whitby, Yorkshire by a drift netter. A number of research workers have tagged sea trout from the Tweed and found that the fish do not move far away from the coast, although they may move some distance along it (NaIl, 1955) . Swain and Hartley (1955) tagged sea trout off East Anglia and a few were recaptured later off the Northumberland and Yorkshire coasts and in the

Northumbrjan rivers. Also sea trout smoits were tagged in the Tweed and the Coquet. In the few months following tagging recaptures were made off the Norfolk and Suffolk coasts and a few were reported from the coasts of Belgium, Holland, Denmark and Norway, while others were taker by trawlers in the southern North Sea,

Balmain and Shearer (1956) list the records of the 123

sea trout caught at sea off. the British Isles over the period 1927-1954. In summarisarig their results for

those sea trout tagged as kCitS Q three were recaptured off Holland, two off Denmark and one off the :Norfolk

coast Of those sea trout.taqged in the Tweed as smolts 0 37 of 46 were recaptured to the south along the English coast, Twenty seven of these were taken off the Norfolk coast, The other nine fish tagged as smolts in the Tweed were taken by drift netters or trawlers off the Dutch Danish or French coasts. - Went (1962) reviewed the information available on the movements of Irish sea trout and found that most of the recaptures of tagged sea trout were within the area

of. the river system in which they were tagged. Sea trout, tagged in the Vistula River of Poland, were mostly recaptured in the Gulf of Danzig and Vistula Bay,, although a number were caught in the Baltic Sea (Naumov, 1959; SkrochowskaD 1953; Zarnecki, 1966), Aim (1950) states that the migrations of the va Stream sea trout keep within extremely restricted areas and only very rarely extend beyond these limits. He found- that only isolated specimens had travelled up to 100 km. Most of the keits tagged in the River Istra of western Norway were recaptured within 10-15 km of the river mouth (Jensen, 1968).

(ii) Recapture of adult brown trout A total of 283 adult brown trout were tagged in Kirk Burn in 1970 and 1971 as they migrated upstream as well as 96 trout electrofished from the study section of the stream 124

between Apr11 1970 and March 1971,. Approximately 150 brown trout we-re tagged in Soonhope and Giensax burns in the autumn of 1971.

Up to 20 July 1.972.. 12 fish had been recaptured by angling at various stations on the Tweed (Table 20). This would indicate that there is substantial movement of brown trout into the small tributaries of the Tweed followed by movement out of these streams during the summer. Electrofishing surveys in tributaries situated

in the Peebles area have revealed an abundance of brown trout in the autumn in greater numbers than these small streams could hold at other times of the year (Mills,

Clelland, Osborn and Watt s 1972). All recaptured trout were taken downstream in the Tweed of the mouths of the streams in which they were first tagged; four fish being caught some distance from the tagging site, As mentioned earlier, four fish were recaptured in Kirk Burn after spending at least a growing season in the Tweed, Nall. (1955) tagged 160 brown trout in the lower Tweed and 14 per cent were recaptured. In most cases, the distance travelled was short amounting to 3 to 5 km or less but one trout tagged at Norham was reacaptured 32 km upstream.. Table 20, Summary of tag returns of brown trout originally tagged in Kirk Burn.

rn...... r n1___ -r - - ., . - - - - i1O DateLate or .saee or Age Length weigntWeight Date of Place of Distance No, of days from tagging tagging (cm) (g) recapture recapture (km) first capture

83) 52 30.10.70 Kirk trap(up) 4+ 25.9 180 10.7,71 Howford 8,1 253 RD IBE 26.11.70 4± 25.3 155 10.5,71 St.BoswellCs 46.7 165 C 5149 5.11.71 0 Cl - 4+ 27.5 236 6.5.72 Innerlejthen 11,3 183

C 5184 0 24.11.7! - 32.3 366 7.7.72 St.Bosweil°s 46.7 225 C 5211 25.11.71 II II It 2+ 24.6 178 20.4.72 Traquair 9.7 147 C 5213a 25.11.71 II ii 2+ 20.5 102 17.4.72 Ho.rsbrugh Ford 3.2 144

C 5222 26.11.71 11 11 2+ 22.1 114 1.4.72 Melrose 40.3 127

0 II U C 5350 11.12.71 2+ 21.3 130 1.6.72 Innerleithen 11.3 175

C 5356 0 .14.12,71 4± 26.2 . 216 16.4.72 Horsbrugh Ford 3.2 119 BT llib 24. 9.70 Kirk (study sect) — 22.8 164 15.5.72 Horsbrugh Ford 3.2 600

5324 912.71 Soonhope - -- - 15.5.72 Walkerhurn 13,3 158

5277 31171 Glensax - - 25.2.72 Coidstream -73,0 114

a Recaptured at the box trap moving downstream towards the Tweed before finally angled. b Recaptured 4 times migrating up and down in Kirk Burn before being finally angled, K' 126

H The Release of Progeny of Eron Trout and Sea flout arid their Subsequent Recapture.

(1) Introduction and methods

Although it has been known for many years that hybrids between Atlantic salmon and sea trout can he produced under hatchery conditions (Day 1887; Jones 1947; Alms 1955; Piggins 1965). little work has been done on the relationship' of brown trout and sea trout by artificially crossing them and observing the appearance and behaviour of theogenv. In the autumn of 1970 eggs were collected frdm 2 female sea trout arid crossed with 2 male brown trout.

C All of the parents were electrofished from the Quair Water on 3 December; below is a table indicating their age and size.

Specimen no 0 type sex age length (cm), C ST F 3.2+ 60,2 H BT N 2+ 22,.9

I ST. F 2.2± 475 J BT M 2+ 19.5

The resulting fertilised ova were brought to the Department of Forestry and Natural Resources and placed in separate trays of a vertical tray hatchery (Menzies and

Curtis, 1966). Egg mortality during incubation amounted to 5 per cent. The eggs were held in running tap water at an average temperature of 5'5°C, After an incubation period of 69 days hatching began. 'The aleiris were held in the hatchery for 6 weeks and then they were transported 127 to Kinnaber Trout Farm near Montrose, Angus on 16 March

1971. The two groups of trout were maintained in separate tanks the water for which being extracted from the River North Esk. The fry remained at the trout farm for approximately one year and over this period they were fed both by hend:and automatic feeders., On 13 March 1972, the yearlings were removed and transported to Edinburgh in galvanized iron tanks with an additional supply of oxygen. Only 2 fish were lost in transit. At Edinburgh the adipose fin of each trout was removed and all those over 9.5 cm were tagged. A total of 694 'hybrids" were handled; 116 of these were tagged. On 14 March 1972, 658 trout were placed in Kirk Burn, 3.9 km above the location of the box trap; 114 of them were tagged.. The temperature of the water in which the t'out were transported was approximately 5.3 °C while the water temperature in Kirk Burn at the time and location of planting was 45 °C.

(ii) Mortality of progeny Mortality of the trout from the time of hatching to that when the fish were placed in Kirk Burn was extremely high and amounted to 87.9 and 80.9 per cent. Mortality was greatest during the months of May and August and was attributed to an unidentified pathogen. On looking at the fish more closely, many were found to have a deformed (shortened) lower jaw (Figure 34). Day (1887) includes Figure 34 Lateral view of the cleared and alizarin-stained head of trout showing: deformed lower jaw (x 25)

normal lower jaw (x 25) 128

;;6,-

Al L2.Y

the short lower jaw in his list of monstrosities of

hatchery fish. Accordina to Ceinmill (1912) this abnormality may he caused by an "autogenetic developmental aberration or some intcrference wiun norma]: 'develonment

by an external factor." As similar, crosses were performed in 1969 and at that time there was no evidence of abnormality i.n the progeny, it seems likely that in this situation some external factor interfered with the natural development of the embryo.

While the embryo was dave1opin the water teniperat:itre fluctuated from 4.5 to 8.5°C. Also on two occasions, the flow of water ceased due to an air lock. This may have caused an increase in water temperature at the time when the lower jaw was developing. Fry of four other lots of eggs incubating at the same time in the hatchery showed no deformities but since fertilisation in these lots occurred 2 days previous to those of the "hybrids', the development of the "hybrids" was not so far advanced. Escaue of the possible affect the deformity may have on the growth and behaviour of the "hybrids" any results should be treated with reservation.

(iii) Size of the progeny

The mean fork length at the time of planting of cross ED was 10.1 cm while that of the other cross was 9.3 cm

(Table 21). The hybrids ranged in length from 5.8 to 17.9 cm.

Table 21. Mean fork length and percent deformed of two brown trout x sea trout crosses placed. in Kirk Burn on 14 March 1972

Lot -cm LI Fish tagged and clipped Fish clipped only Combined Normal Deformed Total Normal Deformed Total Normal Deformed Total Number 55 31 84 186 176 362 241 207 448 Percent 64.1 3649 100.0 - 51.4 48.6 100,0 53,8 46.2 100.0 Mean length 12.3 12.1 12.2 8.2 8. 12 8.2 10.4 9.7 10.1

Lot -EC Fish tagged and clipped Fish clipped only Combined Normal Deformed Total Normal Deformed Total Normal Deformed Total Number 32 - 32 168 46 214 200 46 246 Percent 100,0 - 100.0 78.5 21.5 100.0 81.3 1•97 100.0 Mean length 12.9 - 12.9 8.4 8.0 8.3 9.5 8.0 9,3

I-' 0 131

(iv) Downstream movement of the "hybrids' and their appearance.

A total of 20 tagged and 2 untagged but adipose- clipped trout were recaptured at the bDx trap,, 3.9 km

downstream of the release point (Table 22). Although the fish were placed in the burn on 14 March, no fish

were taken at the trap until 15 April. The daily number of the "hybrids' taken followed the same fluctuations and trends as the "wild" downstream migrants of Kirk Burn, The trap was removed from Kirk Duni on 1 June however one fish was angled from the Tweed on 9 June 180 in downstream of the mouth of Kirk Burn. Although the mesh on the trap was small enough to capture trout down to 9.0 cm in length, the mean length of the "hybrids" captured at the trap was 3-44 cm perhaps indicating that only the larger trout attained the

development necessary for migration. Wagner, Wallace and Campbell (1963) found that the mean size of hatchery- reared steelbead rainbow trout captured while moving downstream was larger than the mean size of the steelbead. stocked, indicating that the smaller individuals within

the various size groups were not moving seaward. "Hybrids" taken at the trap • after having spent at least one month in the burn, were similar in -appearance to the sea trout smolts. If anything the 'hybrids' were more silvery, on the sides; slightly paler. on the back and the pectoral 0 pelvic and anal fins were white rather than yellow-brown (Figure 35).

It would seem that the "hybrids" captured at the box 132

Table 22. Summary of tag return of 'hybrid" trout captured at the trap, downstream of the

planting site s 1972.

Date Tag no. Length Weight Clip Remarks Lot cm g

15.4.72 5362 13.1 24 AC D

17 5450 15.0 37 AC C

21 5380 13.1 23 AC D

30 5366 14.3 29 AC 1) 2.5,72.. 5359 15.6 38 AC deformed jaw 3)

2 5457 14.5 32. AC C 2 5384 149 37 AC 3)

2 5414 13,6 28 AC 3) 2 5358 14.9 37 AC 3)

4.5.72 5374 15.6 39 AC . 3) 4 5363 15,0 30 AC 3) 6 5376 14.5 36 AC 3)

8 5365 14.2 33. AC D

13 5466 13.7 27 AC C 13 5383 13.0 21 . AC 3)

13 - 103 - AC ".•

16 - I, 9.4 - AC

20 5459 15.0 37 AC C

24 5474 17.0 56 AC ô 24 5412 9.8 - AC " dead 3) 28 . 5460 14.1 29 AC C 9.6.72 5464 17.9 . AC . C angled at Cardrona rlwy. bridge Figure 35

"Hybrid" trout 0 13.1 cm and 24 cm, captured at the box trap , in Kirk Burn 15 April 1972. 133

J• A

! .41

jc -4

C 134 trap were actually migrating downstream. All of the fish taken were in good physical condition aid were not emaciated, which might be the case if they were forced downstream due to the lack of available territories. In experiments conducted by Skrochowska (1952) "hybrids" produced from crosses between brown trout and sea trout retained the migratory habit, but the generation produced by breeding "hybrids' among themselves showed a segregation into some migratory and some non-migratory forms. She also found that sea trout kept for three generations in ponds lost completely their instinct to migrate to the sea. I. The COITITTLCrCIa1 Catch. of Trout at Berwick-upon--Tweed.

Introduction

The Berwick Salmon Fishery Company Ltd 0 is situated at the mouth of the Tweed and maintains a commercial salmon and sea trout fishery both inside and outside the estuary of the Tweed. In the summer of 1971, 55 sets of sea trout scales were obtained from the commercial catch between 14 July and 11 August.

Calculated growth and age Most of the sea trout were found to be age 2a.1+ although almost an equal number were 3.j.+ (Table 23. Figure 36) From scale readings of nearly 3,000 sea trout, from the Tweed s Nail (1930) classed 23 percent as age group 2.1+ and 6.9 per cent as age group 3.1±. No explanation can he made for the difference because of the difference in numbers of scales read. The average lengths and weights of each age group are indicated in Table 24. The sex ratio is quite close to 1:1. . ,. Of interest is the inclusion of 10 brown trout which ranged from 4 to 7 years of age. These could y well be examples of estuarine trout which according to Mills (1971) adopt the intermediate habit of frequenting the brackish waters of estuaries (Figure 37). 13 6

Table 23. Calculated lengths of a sample of sea trout from the commercial catch of the Berwick Salmon Fisheries Co. Ltd. e 1971,

Calculated length at the end of each winter

Age No. of Percent of River life Sea life group fish total 1 2 3 4 1 2 3

21+ 20 46.5 7.9 15.7 - - 350 - -

3.1+ 18 41.9 5.7 114 17.2 - 33.5 - -

4.1± 1 2,3 45 9.0 15.0 25.5 AO.O - -

2.2+ 2 4.7 5.7 14.5 - - 33.3 46.7 - 3.2+ 2 4.7 5.3 10.0 15.8 - 35.O 44.8 -

el l' I • • .. • -.:

- - Figure '06, ,-.. - -' fl Scale of a'53 .O cm male' 5eattoute age' 3.l+ \

I I •a captured 16 July 171 - in, th, e cormnc:cj aJ flets

C - atBervJ1cY-L1por1-TJed (X. 25)

A. : • - L a - 1st. freshwater annulus

5 fieshwat&r annulus - - - p - -

c - 3rdfiesln.ater arnvus ia

L • • - - d -ss -Lsea -aI ulus:..7

I -

,_•'_'< C -.

- : 1

. •. •*;. - • •

• '.1•t ' *#i 4 .. ,. • • C-

• • C- 137

.,'•t . ,i ,sb 138

Table 24. Average weights and lengths and sex ratio

of trout by age groups in the commercial catch of Berwick Salmon Fisheries Company 0 1971.

SEA TROUT winters since age Number Average Average Sex migration group length weight ratio F

1 winter 2,1+ 20 48.4 1366 10 10 3.1+ 18 49.2 1465 10 : 8

4.1+ 1 57.5 2268 0:1

2 winters 2.2+ 2 58.8 2821 1 1 3.2+ 2 54.0 2183 0 2 2,1+511+ 1 57.5 2240 1 0

3 winters 3.1+2SM+.1 55,0 1871 0 : 1

BROWN TROUT

4 2 47.5 1162 0 : 2

5+ 2 45.5 1177 0 : 2

6+ 3 46.6 1257 1 2 7+ 3 51.9 1522 3 0

Figure 37

I , e rcale of a 47 - 0 ca male t Lar S ne 0 brown trout,

zqe 5-r 8 captured 16 July 197l,x the commercial c ... i.-..•

nets at Bcrwt3c- on-Twetd ç X 25) a - 1st 3rntlluc d - 4th annulus h - 2nd 'arnulus e ,5th annul uz and- c - 3rd annuwr1r ft spa.'nang mark. 4 SC

- * w C

C I

: 2

--C -4 --: . •.-.-,-. -' -

......

-- .- . •C: - - 139

- C

all 140

(iii) The commercial catch

Trends of the River Tweed sea trout catches disclosed by 5 year rolling averages for the periods 1859 to 1895 and 1952 to 1970 are indicated in Figure 38 A arid B respectively. The greatest number of sea trout entering the Tweed

seems to occur in the months of June and July. Figure 39 shows the annual average weekly produce of thred peribds of 15 years; namely 1842 to 1856, 1871 to 1885 and 1951 to 1971. In the earliest 15 year period, the netting extended from 15 February to 15 October whereas in the latter two periods netting was terminated on 14 September.

•-

- ? ' t

I, s rt

Ilk i-l-Z Figure 38 Trends of the River Tweed commercial sea trout catches disclosed by 5--.year rolling averages. A 1952-1970 B. 1859-1895

4.1

I -- 45

-----4---- •1

1952-55 1956-60 S60-54 195-63

C t 1' e

0 0

- - 0 0 - - 0 • ' S 0 C - • 0 0 0

0 0.

1860-64 1S70-74 ltS3-64 189O-94 -

Figure 39 . River Tweed sea trout catcte.J Annual ayrage-- produce of three periods or i5y,prs (tne jroduce

of the week in qhich the-ge test number of fish was c3ut is, rresented by 100) • -!.

- / -

1 '-- ----nr - -, -. -

- . r-

.4 . - IM

ScasonrFeb4 Sept,

/ Season 15 Rb.-14 S'zpt. 80- 1842-1856 / season 15 Feb 15 Oct

lkv LU .3 . z U U

/ .

March April May junq July August Scptmbcr October, 143

J. Discussion.

The sea trout is distinguished by the fact that the young migrate to the sea after two or three years and return to freshwater after spending one or more growing seasons in the sea. The true broxii trout spends its whole life in freshwater and is usually non-migratory. In the course of this study, the possibility of the existence of an intermediate variety has appeared which complements the true brown trout and the anadromous sea trout. This third type of trout spends much of its life in the Tweed but migrates upstream into the small tributaries to spawn. Some of the evidence gathered in this study suggests that this "river" trout is at least in a large part, the progeny of anadromous parents. The disproportionate sex ratios found among the adult sea trout of the Tweed are not new to the biology of sea trout. The dominance of females among sea.. trout which have spent one or more years in the sea has been found by many authors (A1m 0 1950; Jensen 1968 and others). Alm (1950) stated that although no direct explanation was possible for the predominance of females t he suggested that there might be a higher mortality among the male sea trout. Previously Dahl (1910) went as far at that time to suggest that young female sea trout descended to the sea while some of the males never left the river. The sex composition of the freshwater trout from the Tweed captured in Kirk Burn approximated 4 to I in 1970, and 8 to 1 in 1971. This is most unusual for trout as 144 most studies on brown trout have shown that the sex ratio approaches 1 to 1 or there isa slightly hicther percentage of females (Hcbbs 1937; Gustafeon, 1951; Munro and

Ealmain 9 1956) On the spawning grounds. every female sea trout observed was surrounded by at least two male Tweed trout whether a male sea trout was in attendance or not. The possibility that the male Tweed trout are present as insurance that the sea trout eggs will be fertilised cannot be overlooked. Jones (1959) described the presence of ripe male Atlantic salmon parr on the spawning grounds in November and December. He concluded that the presence of the parr ensured fertilisation of the eggs in case of .inadequate fertilisation or lack of fertilisation by the adult male salmon. According to Shapavolov and Taft (1954), spawning sea-run rainbow trout are very often accompanied by stream trout. Most of these are sexually mature males which, dart in and out during the nest building and courting act. They go on to say that 'the primary purpose in being present is probably to participate in the spawning activities. Ricker (1938) captured ripe non-migratory specimens of Oncorhynchus nerka from the same area as the redds. of the anadromous sockeye salmon, although he was not sure whether frey actually bred with the larger fish. More recent1y, Hanson and Smith (1967) found that male kokänce, from their positions as satellites, also participated in most of the observed spawning of the anadromous sbckeye salmon. In Oncorflypchus masou, usually only one ripe male spawns 145

with a female but frequently freshwater males participate

in the spawning act together with the large salmon

(Nikolsky, 1963)

The abnormal ratios among the Tweed trout and the

sea trout are not directly complementary or the result of

A single act of segregation as they are in the Jcumzha

trout of the Black Sea area (Baracli. 1957) As mentioned

earlier ? Barach found the anadrornous kumzha sampled over a period of years had a sex ratio of 85 per cent females and

15 per cent males on the spawning grounds, while the reverse

occurred with the non--anadromous population; 74 per cent

males and 26 per cent females. In populations of

Salvelinus rnalma of Kumchatka, Savvaitova (1960) found that

in the migratory trout, females made up 64 per cent and

males 36 per cent. However, in the non-migratory river

and lake forms, the relationship of the two sexes was 1:1.

The abundance of females in the samples of see trout

smoits captured in the Tweed and its tributaries is

interesting, as one, would expect a sex ratio ohich

approaches '1:1. The ratio of females to males in the

sample of silvered sea trout smelts seined at Norham on

the Tweed was 2.13 to 1 and at Dead Burn it was 1.89 to 1.

None of the fish in the sample trapped in Kirk Burn were

silvery in appearance and a sex ratio of 0.71 females to

each male migrant was found. From these meagre data

obtained on the sex composition of downstream migrants,

it would seem that those trout, which have the appearance of sea trout smelts, are predominantly female. There may

be a segregation of sexes of sorts at the time the progeny 146

of the anadrornous stock goes to the sea s

Dahl (1910) 4p when examining stocks of trout which had never been to the sea, found that in many places males

and females occurred in equal proportibri in the youngeät year classes whereas the older year classes consisted

mainly of males. He regarded this as definite 'proof that the stream in question contained fish that will

become sea trout. He goes on to say that "in many places (rivers) this is not so obvious, and we may even find that the proportion between males and females is only slightly affected by age and that spawning females

also occur which have itever migrated. 11 Ricker (1938) suggested that in the anadromous sockeye salmon (O.nerka) of Cultus Lake, British Columbia, the forces which initiate migration operate more strongly on females smolts than on males. Barach (1957) also postulated that the migratory instinct in kumzha is, to a considerable degree, associated with sex, the majority of males remaining in the river where they quickly attain sexual maturity. Following Barach's work, PanOT (1958) artificially produced progeny of pure anadromous kumzha, pure river kumzha and the intermediate. He found that of those fish which became silver in appearance, 90 per cent were female and 10 per cent were male.

The differences in sex composition of adult sea trout and adult brown trout found in Kirk Burn during the spawning period, as well as the preponderance of female smolts 3.47

captured at various locations, would suggest that the so- called brown trout are at least. partly Droganv of sea trout However they must he labelled brown trout becau:se they are riot anadrornous. . . 148

III. ELECTROi0RETIC STUDIES OF BLOOD SERUM PROTEINS.

A. Introduction

The designation and. separe.tion of iish species and populations by means of serological and biochemical methods, to a large extent, originated from problems in marine fisheries biology. In attempting to identify and measure the causes of fluctuation in the abundance and distribution of a species of fish it was found to be essential to establish the number and identity of suhpopulations

(Marr, 1957) Each subpopulation may have its own characteristic distribution s fecundity, growth rate etc. Since proteins seem to be but a few steps removed from the gene (Ingram, 1960) and do not seem to be as susceptible as are meristic characters to environmental ndification (EarlowQ 1961) they have become of value to the fisheries biologist. Cushing (1964) presented a survey of the results of studies on the blood groups of marine animals and included the basic principles and methods used. Although in the majority of investigations during the period, only blood grouping techniques were used, the review also mentions the first developments of work on proteins under genetic control. An evaluatioh of techniques, as related to the identification of fish suhpopulations by biochemical and serological methods, has been given by Parrish (1964), Another paper outlining the imnunogenetical and biochemical approach' to the study of fish populations is that of Marr -49

and Sprague (1962) Further blood group studies, but in particular the rapid developient of Biochemical

techniques for the study of genetic characters in fish populations, have resulted in the appearance of numerous

publications since 1964 (de Ligny 1 lG9)

(i) General principles of zone electrophoresis Zone electrophoresis involves the migration of charged

particles or ions 0 which are supported on a relatively inert and homogeneous substrate. The net charge of the particle determines the direction and relative rate of its migration

in an electrical field. This is particularly important with proteins because these materials are araphoteric and

can exhibit positive ; zero or negative net charues at- different pHs.

The mixture of substances to be separated is placed as a narrow band or zone at a suitable distance from two electrodes such that as migration occurs, the different

components, which move at different rates; Slowly draw, away from each other to produce a separation in the direction of migration.. A short migration generally separates the components into strongly different groups

and yields fewer fractions, while a longer migration time tends to further separate the groups.

Following migration the substrate or stabilising medium is immediately placed in a fixative which precipitates the substances being examined with the result that the separated substances remain in their correct positions of migration, 1 5O

Subsequently, staining reveals the substances as a number of discrete zones

Zone electrophoresis has been us'ed mainly as a

clinical technique and to some extent, for small scale preparative separation. A wide variety of materials can be separated by various zone electrophoretic techniques, hut the procedure is especially useful for the study of protein mixtures.

The use of zone electrophcresis has only been widely adopted since about 1950, although now there is considerable literature relating to the experimental techniques and

applications (Whipple;, 1964; Smith, 1968; Shaw ' 1969) Much of the earlier work involved the use of filter paper as the supporting medium In recent years, however, filter paper has been superseded by other supporting media

such as cellulose acetate membrane, starch gel, silica gel, agar gel, and wlvacrylamicje gel, which permit sharper separations.,

• (ii) Electrophoresis and the serum and tissue proteins of fish

The electrophoretic study of body tissues and fluids in the higher vertebrates have demonstrated the existence

of genetic variation of many proteins and enzymes (de Ligny, 1969), particularly after the method of Smithies (1955) using starch gel as a substrate. - In particular, there are a number of protein and enzyme groups found in the blood serum which have been used to differentiate fish populations Those that have been a- 4.

most successful are serum transferrin (Drilhon and 1960; Barrett and Tsuyuki, 1957; N5iier, 1970) serum

esterase (Nyman 1967, 1970, Koehn and Rasmussen 1967) haemoglobin (Sick,, 1961; Tsuyu.ki and Gadd, 1963; Wilkins, 1968) and unidentified proteins (Tsuyuki and Roberts, 1966; Mulcahy, 1967; Wright and Jias1er 1967). Tissue proteins and enzymes have also been used extensively because of the ease of collection and storage.. Some of the more important tissue proteins are muscle myagen

(Tsuyuki and Roberts, 1965; Nyman, 1967), eye lens proteins (Rabaey, 1964; Smith and Goldstein, 1967; Pollard and Pichot, 1971; Eckroat, 1971) and lrtate dehydrogenase (LDH) .(Bouck and Ball, 19687 Clayton and Gee, 1969; Northcdte, Williscroft and Tsuyuki 1970)

The species specific nature of the electrophoretic pattern of fish proteins have been subs tanti ated for many species of fish. Some of the studies are as follows:

Irisawa and Irisawa (1952) sharks; I)rilhon (1953) eels and carp; Starr and Fosherg (1 -957) sharks; Gunter Sulya and Box (1961) elasmohranchs and teleosts; Tsuyulci and Roberts (1966) Oflnchus sp.; Barrett and !rsuyuki (1967) scombroid fishes; Koehn and Rasmussen (1967) Catostomidee; Fujino and Rang (1968) tuna; Then and Tsuyuki (1970) Tilapia sp. and Nyman (1971) marine and anadromous teleosts.

(iii) Factors which may affect the electrophoretic pattern Both physiological and environmental factors may have an influence on the electrophoretic pattern of fish serum 152 proteins.. One, therefore 9 must.: be especially cautious

when interpreting results and, making conclusions (Eooke 9 1964; Thurston, 1967). Changes in serum protein patterns of fish have been attributed to differences in sex or sexual maturation (Ho and Varxstone 9 1961; Robertson et al 1961; Lysak and }3iexrz 9 1965) and life history differences

(Dri1hon 1954; ooke, 1964; Thomas and M-acCrimmon, 1964). The appearance of. a fraction associated with sexual maturation in females may Only involve an increase in concentration of a previously weak component (Thuston, 1967; Wright and Hasler, 1967) . Tsuyuki and Roberts (1966) found that fractions, supposedly, associated with sex were present in some, but not in other. species of the genus

2Q22.flyn6hus. Age and development may also alter the electrophoretic pattern (Sano 1960; Vanstone and 'Ho, 1961; Koch Bergstr6m and Evans, 1966). Intraspecific variation has been attributed to a factors such as changes in diet :am and Koch, 1969), temperature

id Hickman, 1962; Poston, 1966; Q variations (Saito 9 1957; Flemming,

smctic pressure Wordier and Barnoud a and waters with various concentrations jiya 1 1961; Neuhold and Sigler, 1960; md disease (sindermann and Mairs, 1958; y, 1972). Finally the method of ited stress may alter the electro- 3. et al, 1943; Bouck and Ball, 1965, 3.53

(iv) The experimental approach In the present study, the electrophorotic patterns

of blood serum proteins were determined for 42 sea trout 9 57 brown trout and 27 'hybrid" trout using a substrate of cellulose acetate. Electrophoresis of blood serum proteins was also carried out in polyacrylamide gel using serum samples from 37 sea trout, 46 brown trout and 19 "hybrid' trout. - The objectives were to determine if it was possibJ.e to distinguish the serum protein patterns of brown trout- and sea trout so that I could, firstly, determine the taxonomic relationship of the two and secondly, if possible, find a method to distinguish juvenile sea trout ±rorm young brown trout to assist in the management of the trout populations in small streams tributary to the Tweeds 154

B. Materials and Methods

(1) Trout

Brown trout and sea trout were captured by electro-. fishing in a number of streams tributary to the Tweed in the Peebles area, The blood samples were taken from the trout in the months of October. November and December 1970 and 1971, when the fish were mature and approaching spawning condition. Nearly all of those fish classed, as brown trout were males, while in the sea trout the ratio of males to females was approximately I to 1, The lengths of the brown trout from which samples were taken ranged from 17.0 cm to 36.7 cm while the largest sea trout sampled had a fork length of 64.2 cm. Brown trout of age group 3 were the most corrnon and age groups 21+ and 3.1+ in the sea trout predominated. The progeny of the brown trout and sea trout matings were artificially produced from two female sea trout and two male brown trout electrofished from Quair Water near Traquair (see Figure 3). The resulting two batches of ova were kept in a vertical tray hatchery in the Department of Forestry and Natural Resources until hatching was complete.

The fry were transported, at one month of age s to a trout farm and maintained in tanks. for 12 months. When they had reached a mean length of 10.2 cm,, blood samples were taken. 155

(ii) Blood collection

After removal from the stream, trout were anaesthetised

with tricaine methanasuiphonate (Ms 222) at a concentration

of about 0.25 g per litre. Blood samples were taken by cardiac puncture using a hypodermic disposable syringe of

2 ml capacity. Blood was extracted by inserting the needle into the centre of the isthmus on a line drawn

between the origins of the pectoral fins. When blood

was being taken from brown trout, a 23G x 25 rum needle

was used, but for the larger sea trout a 21G x 40 nut

needle was required, The quantity of blood renoved

varied from 0.1 ml to 1,0 ml. Following its removal from

the trout blood was immediately transferred to plastic centrifuge tubes, which were then sealed with disposable

stoppers, Samples were allowed to clot and on return

to the-laboratory, they were centrifuged at 5000 rpm for

10 minutes, Serum samples were discarded if more than

slight heemolysis had occurred, The serum was then removed using Drumrnond disposable Microcaps and stored

in Beckman Polyethylene micro test tubes at -15 0C for

periods up to 16 months before electrophoresis. Although Goswarni and Barua (1959) found that cold storage does not

affect the paper electro* phoretic pattern of serum proteins,

Haider (1969) reported slight denaturation or changes in the

electrophoretic mobility which may be minimised by sudden

freezing to -40 °C for extended storage periods.

Following withdrawal Of the blood sample, each specimen was weighed, measured and usually tagged and then 156

returned to the stream after a 10 minute recovery period. A number of trout, from which blood samples had been taken, were kept for up to six months an the laboratory. Also six brown trout were recaptured in Glensax Burr, three months after a blood sample had been withdrawn from them. Blood was taken from the smaller 'hybrids' by severing the caudal peduncle (Thurston, 1967) and using disposable pipettes to collect the sample before clotting occurred.

(iii) Electrophoretic methods a) Cellulose acetate electrophoresis !Cohn (1963) gives a detailed description of the procedure of zone electrophoresis cn cellulose acetate. The technique of separating fish proteins on cellulosa acetate has been used by Dufour and Barrette (1967) Smith and Goldstein (1967) and Mulcahy (1970) In this particular study, the following solutions were made up: veronal buffer (pH 8..6)

barbitone (diethyl-barhituric acid) 1 1.84 g barbitone sodium 10.30 g calcium acetate 0.38 g made up to 1 litre with distilled water (ionic strength 0.06) staining solution

antido black lOB 550 mg 96% methanol 250 ml conc, acetic acid 50 ml distilled water 250 ml 3-57

decolouriser

96% methanol .250 ml conc. acetic acid 50 ml distilled water 250 ml

Electrophoresis was carried out on Oxoid cellulose acetate strips (20 x 5 cm) in a Shandon 'electrophoresis

apparatus (model U77) . Power was, supplied through a - Shandon Vokam constant current/constant voltage D.C.. power unit e (type SAE_ 2761)

Following marking the strips to be used were floated on the surface of the buffer and wetted from below by

capillary action. They were then positioned in the apparatus and the current was switched on and a voltage

of 30 v was maintained for ½- hour to allow the system to equilibrate. After this period, the current was switched

off and 2 ul of sample per strip was applied. Electro- phoresis was allowed to proceed for 5 hours at a constant voltage of 120 v (3-4 mA).

After electrophoresis, the strips were placed in 96 per cent methanol for 2 minutes for fixing. The strips were then immersed in the amido black staining solution (Zweig and Whitaicer, 1967) and, in turn., three solutions of decolouriser.

The strips were allowed to dry between heavy glass plates and the resulting electropherograms were scanned using a Joyce Loeble Chromoscan integrating densitometer,

A red filter s 1010 aperture and gear ratio of 1 t 1 were required to construct a recording from which the percentages of total protein in the various bands could be calculated, 158

b) Polyacrylainide gel disc electrophoresis

This type of electrophoresis originally described

by Ornstein (1964) and Davis (1964), has the advantage cf

being more versatile, since the sample moves through the

supporting medium or gel rather than on top of it as occurs in cellulose acetate electrophoresis. This facility enables the acrylarnide gel to be used not only as a routine analytical tool based on fixed concentrations, but as a molecular seive in which certain large molecuies can be screened out. Both Latner (1967) and Smith (1968) give extensive-reviews of the techniques and the buffer systems which may be used.

In this study the following solutions were used:

Ornstein and Devises 7% separating gel

acrylamide 6.820 g

methylenebisacrylamide (BIS) 0.184 g Tris HC1 buffer (pH 8.9)

Tr is 4.575 g

NUC1 6.0 ml

distilled water 100.0 ml

TEMED 30.0 ul when required, polymerization is initiated by adding

1.0 ml of 7% ammonium persuiphate solution. Jc. buffer (pH 8.3)

Tris 0.60 g

glycine 288 g

distilled water 1000.0 ml 159

Staining solution

amido black 10 B 1 .g

7% acetic acid 100 ml

The technique of preparing the acrylamide gel was quite similar to that of Mackie (1969) . The electrophoresis apparatus had a 6-tube capacity and each tube was 7.5 cm in length with an internal diameter of 6 mm, To obtain a flat surface on the top of the gels, a volume of -- ateL' was carefully layered on top of the acrylar1iide and then the gel was allowed to polymerize, at room temperature. After one hour the water was decanted off and the tubes were placed in the electrophoresis apparatus and the tanks were filled with the Tris-glycine buffer (pH S. 3).

Ten ul sample solutions composed of equal exrnunts of 2001 sucrose solution and serum sample were then micropipetted onto the top of the gels. For the first 10-15 minutes, a constant current of 1 ma per tube was maintained but this was then increased to 4 ma per tube and continued for approximately 90 minutes. Electrophoresis was carried out in a refrigerated chamber at 4°C to contend with overheating of the gels.

After electrophoresis, the acrylamide gels were carefully removed from the tubes and placed in the staining solution for 15 minutes. They were then washed in tap water, destained in 2% acetic acid for 2-3 days end eventually stored in 7% acetic acid.

The acrylamide gels were scanned using a Joyce-Loeble U.V. Chromoscan. 1.60

P1 C. Results of Electrophoresis

Cellulose acetate electroohoresic of the serum

proteins showed as many as 12 bands or fractions, hut more commonly, a fewer number of com -ponents appeared after staining. Figure 40 indicates actual electropheroararms of serum proteins from the three types of trout as compared

to that obtained from a. sample of dog serum. No attempt: was made to idenLify the protein components and the electrophoretic areas were merely assigned numbers in order of increasing mobility.

Fraction 3 was located at the point of application of the sample; fractions 1 and 2 migrated towards the cathode while fractions 4 to 12 moved towards the anode Within the serum samples taken from each type of trout, considerable variation was apparent; however fractions l 3, 5, 8 and Ii were always present.

When qualitatively comparing the electrooherograras of brown trout, sea trout and their hybrids", (Figure 40) there is very little difference in the three. Fraction 5 with its two adjacent components 4 and 6 and the other major fractions B and 11 were common to nearly all of the patterns scanned. When the serum protein patterns of each type of trout were compared a äigrsificant difference (5 per cent level) was indicated between fraction 1 of the brown trout and that of the brown trout x sea trout Also a significant difference was found between fraction 3 of sea trout and the "hybrids'

(Table 25). Although a difference in the percentage of total serum protein in these two fractions has been shown T ., 4 "-'b • p., -

I -

Figure 40!

Iiectrophoretic pattern 9Th-cellulose, acetate of blood', seruxn 'proteins from three types 'of' trout

(S.trutta) and clog (the fractions are numbered

1 to 12 with increasing mohilty)

- r 1 ' •I --Ui-

CELLULOSE ACETATE EL.ECTROP;iR-Esis

Brown •Irout

1 12 Serum

Sea Trout - .-- Serum

---4

Sea x Brown Trout Serum

Doc - crurn

4 PONT OF APPLCATON Table 25. The mean percentage and percentage of occurrence of each fraction of serum proteins separated by cellulose acetate electrophoresis.

Fractions (slowest to fastest) - percentage of total serum protein

1 2 3 4 5 6 7 8 9 10 ii 12 Brown trout

mean percentage of each fraction 11.2* 5.9 9.5 6.7 15.8 13.4 6.3 19.7 6.8 6.8 16.1 4.5 standard deviation 3.4 2.2 3.8 2.7 5.8 3.7 3.3 4.8 2.7 2.0 4.1 1.5 percentage of occurrence of each 100.0 29.7 100.0 70.3 100.0 97.3 54.1 100.0 37.8 13.5 100.0 32.4 fraction

Sea trout

rnei percentage of each fraction 9.5 6.8 12.1* 7,5 18.1 14.4 8.6 17,5 8.4 7.3 15.3 3.6 standard deviation 5.0 .1.6 4.8 3.2 6,9 4.7 3.5 4.2 3.0 2.9 3.5 1.5 percentage of occurrence of eachloo.0 20.7 100.0 37.,9 100.0 96.6 27.6 100,0 55.2 27.6:100.0 27.6 f r act iO).

Brown trout x Sea trout

mean percentage of each fraction 9.2 5,2 8.0 8.4 15.6 10,8 5,8 23,6 8.9 - 16.8 7.0 standard deviation 3.1 - 2.5 3.0 4.6 3,1 2.1 6.0 - 2.3 - 2.5 2,4 percentage of occurrence of each 100.0 10.0 100.,0 70,0 100,0 90.0 fraction 30.0 100.0 70,0 0 100.0 40.0

* denotes significance at the 5 percent level. it-,

I feel that one should treat the results '4t- h some reservation.. Fraction 3 occurs at the point of application where non-migratable material would remain during electrophoresis. This material after stain-ing, could then give erroneous values for the amount of protein present in the

fraction. No explanation can be given for the difference in the amount of protein in fraction 1 of brown trout and

brown trout x sea trout Differences in percentage of - occurrence for a number of fractions for the three types of trout should only be attributed to the variability of this type of electrophoresis, until more serum samples have been run and/or the system refined. The technique of polyacrylamide gel electrophoresis indicated more promise as a means of separating the two

types of trout and their "hybrid". AS many as 13 bands could be distinguished, but usually 10-12 d--finite

components were separated (Figure 41). No attempt was made to quantify the amount of the various fractions because of the variations in the patterns even within each type of trout.

Figure 42 indicates the actual separation of the serum of brown trout (A and B), sea trout (C and I)) and brown trout x sea trout (E and F). There appears to be some differences in the patterns but more samples are required before any definite conclusions can be reached. :

• Figure 41 1 • Electrophoretic pattern in polyacrylarnide gel • of b1ooa:seum proteins from three types of trout (S.tr.utta) )

j . • c

• •p •. a • • c

•. I 64

)Jt Jt)Ti

POWIi IHQUT X SEA TROUT SERUM

---4 Figure 42 Actual polyacrylamide gels after electrophoresis and staining for blood serum proteins of trout

(s. trutta) A, B - brown trout'

C 1 D - sea trout E, F - brown trout x sea trout 165

A B C 166

D. Discussion

The eledtropherograrns of brown trout serum proteins compare favourably with those obtained by other research.

workers. Deutsch and McShan (1949) were one of the first contributors to the study of the interspecific differences in the family Salrnonidae. These investigators attained electrophoretic patterns for the blood serum of rainbow trout, lake trout (Saivelinun namavcusb) and brown trout and found that closely related species showed recognisable variations, They found eight components in rainbow and

lake trout and 11 fractions in brown trout serum. The serum patterns obtained for brown trout in the present study show a slight resemblance to those obtained by Deutsch and McShan (1949), both indicating three or four main fractions.

Sanders (1964) compared the electrophoretid components of sera of three species of trout and their hybrids as to mobility, amount and number of prctein components using

paper electrophoresis. He analysed 14 samples of brown trout sore and found seven fractions present 9 three of which he considered major protein fractions. Haen and OVRourke (1968) separated the serum proteins of Atlantic salmon, sea trout and their F 2 hybrid using polyacrylamide gel. In the electropherograms of the sea trout serum, they were able to distinguish 10 components. Mulcahy

(1971) also using polyacrylamide gel as a substrate 9 and looking at serum protein changes associated with ulcerative dermal necrosis (UDN) in brown trout, was able to decipher 167 three minor and five mjor components. Studying the same disease. Carberry (1970) found seven fractions on cellulose acetate. Nvrnan (1955), employing starch cci as a substrate and a discontinuous buffer system, was able to show up to nine hands in samples of sea trout serum. When eJ.ectrcphoreticaliy comparing the serum protein patterns of a number of trout species Novikov, Savvaitova and Maksimov (1970) distinguished nine compcnens in brown trout sera. Their patterns obtained for hrovn-x trout closely resembled those obtained in this study in that they found two components which migrated towards the cathode and seven components which moved in the direction of the anode Also three components were especially prominent and were located in approximately the same position, relative to the other bands, as found in the electropherograms of sera from Tweed brown trout;. When searching for natural hybrids between European trout and Atlantic salmon, Payne, Child and Forrest (1972) analysed general serum protein patterns using polyacrylamide gal electrophoresis. Although they only found up to five components in sera from brown/sea trout, they were able to distinguish natural hybrids between Atlantic salmon and European trout. Nyman (1965) found that serum protein patterns of the Atlantic salmon were easily distinguishable from those of the sea trout and the brown trout. He was also able to find hereditary differences between "brook locked" brown trout upstream of a series of impassable falls in a 168

tributary of the River Indalslveri in .northern Sweden and

sea-run trout of the main river (Nyman perscornmJ These differences were located i.n the blood plasma proteins. He did state however that since he was 'unable to cross- breed specimens of the to populations ; he has no definitive proof that the differences are purely genetic.

Schumann (1959) investigated the haemoglobin portion of fish blood proteins using agar electrophoresis and found that he could separate sea-run brown trout (Salmo trutta trutta) which consistently showed two distinct bands 0 from the lake form (Salmo trutta lacustras) which indicated three distinct haemoglobin fractions. The haemoglobin of the brook form of brown trout (Saln'n trutta farjoY migrated as two fractions resembling that of the sea-run trout, There have been a number of other electrophoretic. studies in which the patterns of certain protein groups of freshwater-resident salmonids and their anadromous counterpart have been compared. When looking at the muscle myogen patterns on starch gel of some members of the family Salmonidae, Tsuyuki and Roberts (1963) could find no apparent differences between the rainbow and steelhead (sea-run rainbow) trout SaJ.mo cairdneri Vanstone Roberts and Tsuyuki (1964) found that haemoglobin from "land-locked" and anadromous sockeye salmon (Cncoflynchus nerka) yielded identical patterns on starch gel. Finally,, Gray and McKenzie (1970) found muscle protein patterns of anadromous Atlantic salmon from three river systems in New Brunswick and from Greenland, similar to those of 169

landlocked salmon from Newfoundland and New Brunswick Sera taken from the three forms of trout in this particular study all show nearly identical patterns on cellulose acetate. Although this type of electrophoresis is suited for derisitometric scanning of the patterns the separation is not as comp1±e as it is in gel media such as polyacrylarnide with a molecular sieving effect.. On polyacrylamide gel,, there is a suggestion of a. difference in the serum protein patterns of the various trout forms. Differences in the pattern obtained from "hybrid" trout and the other two forms could be due to age and maturity as the "hybrids" were only one year of age while the brown trout were predominately 34. and the sea trout 2.1+ and 3.1±. Gradual alterations in protein patterns with age and development do occur (Sanders and Wright, 1967; Koch, Bergstro"m and Evans, 1966; Uthe and Tsuyuki, 1967).

When looking at the arrangement of the serum protein bands in oolyacrylamide gel of the brown trout (A and B) and sea trout (C and D) in Figure 42, there appears to be a difference of the furthest migrating fractions between 40 and 65 rrm 0 Whether this difference can be considered an actual genotypic effect, is unknown until more samples are processed.

If representatives of the three forms of trout had been kept in a hatchery under identical environmental conditions, much of the variability experienced in the present study of blood serum proteins might have been ruled out. Also the converse may occur and that is in 170 view of the suggestion made in the previous section that those fish considered brown trout are actually progeny of sea trout, the differences observed in the serum protein patterns on polyacrylamide gel, are more likely due to physiological and environmental influences rather than genetic, factors. 171

IV. MOLECULAR WEIGHTS OF RIBOSOMAL RNA IN BROWN TROUT

AND SEA TROUT..

A. Introduction

The molecular weights of ribosomal RNA (rRNA) can be compared using electrophoresis on polyacryla'inide gels, which give more extensive and precise separations than are possible by density-gradient centrigugation (Loc-nina

1967) Ribosomal PITh is made up of a number of ribz'soiriai subunits or components two of which in marPnials have bew labelled 28s" for the larger and lBs for the smaller subunit.

ifl his study of the molecular weights of r.Ri'A in relation to evolution, Loaning (1968) found that bacteria,

Acinomycetes, blue-green algae and higher plant chloroplasts all had rRNA ("23s" and "16s") with molecular weights of

1.1 x 10 daltonE and 0,56 x 10 daltons The weights or rRNA ("25s" and "18s") of the higher plants 0 ferns algae and fungi and of some prctozoa were approximately

1.30 x 106 d and 0,7 x 10 d0 The 0.7 x 106 d component was found to be common to all the animals studied, but the larger rRNA component ( " 28s") appeared to have evolved with each major step in animal evolution from 1.40 x 106 d in sea urchins to 1075 x 10 d in mammals.

According to Loening (1963) Dr. 0 HL. Bishop of the

Department of Microbiology, University of Illinois D using double labelling, found differences not related to 172 composition, of about 1 per cent between sp?c5es of hacte. More recently, Loening (in press) has actually been able to distinguish three species of Amoeba by differences in the molecular weights of the ribosomal components. With this in mind, it was thought that the species specific molecular weights of rRNA of the various forms of Saimo trutta might give some indication as to, the similarity or distinctions of the two types of trout at a basic level. 173

B. Materials and Methods

.(j) Trout

Brown trout were collected by electrofishing in

Glensax Burn in the early months of 1972. The fish were all spent males ? between 21 and 25 cm in length, and had moved into this tributary of the Tweed during the autumn of 1971. Sea trout smolts which had a silver appearance, we-re captured in May 1972 at the box trap situated in Kirk Burn.

Sea trout x brown trout progeny ("hybrids") were artificially produced and kept in a hatchery near Montrose, Perthshire until they were one year of age.

(ii) Preparation of RNA

• Whole livers were removed from the trout immediately after death and stored at -20 0C until samples were required. RNA was isolated by homogenization of the tissue in a medium which contained (Parish and Kirby, 1966):

tri-isopropylnaphthaiene sulphonate (TNS) 1% (w/v) 4-amino sal.icyiate (4-AS) 5% (w/v)

NaCi 1% (w/v) phenol--cresol mixture 6% (w/v)

Homogenization was performed by a motor-driven Teflon-in- glass homogenizer, using at- least S volumes of the medium, at 0 to 5°C.

The homogenates were shaken with phenol-cresol-water (Parish and Kirby, 1966) and centrifuged at 2000 g at 5oc 174

for 10 minutes. The supernatant solutions from the phenol extraction were brought to 3 per cent in NaCl and the phenol extraction was repeated once or twice, RNA and DNA were precipitated from the final supernatant

solution with 2 volumes of ethanol at 0 °C. To purify the RNA further, and to remove the detergents and phenol, it was dissolved in a solution containing 0.15 M-sodiuxn acetate and Oe 5 per cent sodium dodecyl sulphate (SS) at pH 6. The RNA was precipitated with 25 volumes of ethanol. The method of preparation is explained more fully in Loening (1967).

RNA was prepared from Escher.i.chia coil by the sodium dodecyl sulphate-phenol method (Penman, 1965).

(iii) Electrophoretic methods

Polyacrylamide gels were prepared as described by Loaning (1967) and Loening and Ingle (1967) Briefly, dilute gels containing 2.2 to 3.0 per cent of acrylamide were made up, and the concentration of the bisacrylamide was 5 per cent of that of the acryltimide throughout. The buffer contained: Tris 30 mM sodium dihydrogen phosphate 36 mN sodium EDTA (disodium salt) 1 IT1M to give apR between 7.Q and 7,8e The use of EDTA9 which has a high electrophoretic mobility, precludes the use of the discontinuous buffer system described by Ornstein (1964) and used in the separation of blood serum proteins in the 17.5

earlier section. Thfrrimning buffer in the buffer compartments also contained sodium dodecyl sulphate

(c). 2%) - The gels were pre-run, at 5 irA/gel for

approximately 30 minutes. The RNA samples were dissolved in the electrophoresis buffer containing 14 per cent

sucrose and 0,2 per cent sodium dodecyl sulphate, The

concentration of RNA was approximately 1 mg/ml and 5 to 50 ul were layered over the gels, Mixtures containing RNA from 2 or 3 types of trout were prepared beforehand arid layered as a single solution, Electrophoresis was continued at room temperature (20 to 25 °C) at 7v/cm length of gel and 5mA/gel for from 2,5 to 5.5 hours. The gels were then scanned in a Joyce-Loebi Chromoscan using a medium pressure mercury lamp and an interference filter

at 265 mu. In most cases,. trout--type RNA was run with E.coli RNA as the marker. In order to identify the peaks in a mixture, the individual samples were also run separately in adjacent gels in the same electrophoretic tank.

The molecular weights of E.coli RNA were assumed to be 1.07 x 106 daltons and 0.56 x 10 daltons (Loening 19.68)

-Th The mobility of an RNA molecule in polyacrylamide gels is inversely related to the log of the molecular weight. Loaning (1968) found that the reproducibility of the molecular weights waswas always within - 5 per cent and usually within 2 per cent. 176

C. Results

Electt'c'phoresis of a mixture of E.coli and "hybrid" trout RNA showed four separate peaks,, two peaks of

E 0 coli P.NA and two peaks of trout RNA (Figure 43).

Considering the larger ribosomal component of E.CO1± as having a molecular weight of 1.07 x 10 d and the smaller component as 0. 156 x 10 dv then the weight of "28s" RNA in the trout is 1.45 x 106 d and that of the "iBs" is 0.72 x 10 d. Comparison of each of the other types of trout, namely sea trout and brotn trout, with E O coi± RNA as a marker showed that the mobility and therefore the molecular weight of the two subunits of rRNA of each type were similar.

Mixtures of E.coli RNA and two or three types of trout also indicated that there was no distinguishable difference between the molecular weights of rRNA of brown trout, sea trout or brown trout x sea trout. Any variations which did occur were not reproducible and so were attributed to experimental error. 4 Fgure43

-- Pblyacrylarnide gel- electrophoresis of brown trout ribosomal •RNA and its separation from soma] RNA of E.co1i (SOuiof trout RNA and 20 ul of Eco1i RNA were applied to a 24% gel. Electrbphoresis was for 4 hr at 50 v; scanned. at an T5ticaLdqnsity of 5.0 T.— frout subunits of RNA E ---. E.coli subunits of RNA

DNA - deoxyribonucleic acid ...... -'

t 9.0

(1) Lii C) -J C U 3-

t-J Distance rnqratcd (cm) --.1 1.73

D. Discussion

As it is probable that the molecular weight of 18s RNA of all animals approximates 0.70 x 10 6 a (Loening, 1969) and this has been found to be the

case for the trout in this experiment, this particular subunit is of little use in distinguishing animals species. The value obtained for the molecular weight of

28s RNA for Salmo trutta fits quite well within tha evolutionary scale described by Loening (1968). The molecular weight appears similar to that of Xeipus (tadpole) and chick (liver) but is not as great as rabbit (reticulocytes) or mouse (liver).

Perry et al (1970) in assessing the extent of variation in the size of the rRNA transcription unit among different organisms and to infer its possible evolution, determined the molecular weight of r RN , A in several organisms. From cultures of rainbow trout gonad, they calculated the molecular weight of the 23s 6 subunit to be 1.5 x 10 d and the lBs subunit as 0.65 X 6 10 d. It is not known at this point whether tne1 disimilarity of these molecular weights of rainbow trout with those of brown trout-sea trout are actually specific, due to experimental error, or due to differences in experimental technique.

It would seem that the comparison of molecular weights of rRNA in different species of fish may be a useful technique to employ in fish taxonomy and systematics. However, before meaningful comparisons of this nature cart be made, refinements of the method are necessary to reduce the experimental error. 179

V. MERISTIC COMPARISON OF EROV2N TROUT AND SEA TROUT

A Introduction

The term 'meristic' has at least two interpretations

in the ichthyological literature. A general usage is synonymous with "numerical' or "capable of being counted" (Quast, 1964) A restricted usage applies to those countable characters which appear to be anatomically associated with the segmentation of the body or body

somites (Rouns3feli and Everhart, 1953) . Gillraker counts are 'meristic in the general sense but not in

the restrictive. Here, the more general usage will be employed in attempting to find differences in the anatomical counts of brown trout and sea. trout.

In systematic ichthyology the numbers of vertebrae9 fin rays and other numerical characters have always played an important part an the cescrxptaon and definition of

species and subspecies (Tning 9 1944) This use of meristic characters became an accepted tool in the taxonomic description of fishes after the early works of Gunther (1866); Jordan and Evermann (1896) and Heincke (1898) More recently, counting of meristic series in fish sampled over the range of the species has been found to be a. convenient technique in looking for evidence of population segregation (Marr 1 1957) Meristic characters are useful because, in contrast to mcrphometric measurements or body proportions, they are supposedly constant throughout the life of an individual and subject to discrete and objective measurement (Lindsey, 1961) Morphological characters

are almost always related to hody size, thus to use these criteria0 the fish taxonomist must eliminate the effect of body size. This is usually accomplished through the use of ratios. Difficulty has been experienced when ratios have been compared, because fish parts are seldom related to body size in a simple ratio which is constant throughout the life of the fish (Royce 1957) Demonstration of consistent meristic difference between samples of fish from two areas or two groups of fish from one area may usually be taken as evidence for some degree of racial segregation, even though hereditary and environmental effects cannot be separated. More useful and reliable conclusions may be drawn, however, if the results of environmental variation can be separated from genetic differences.

The rearing of fish in the laboratory under controlled conditions has demonstrated that several environmental factors are capable of altering meristic counts, Further, the fixation of counts is usually influenced by environmental factors during critical stages in embryonic and larval development (Tning, 1952; Orska,1956) A variety of factors have been found to be capable of modifying meristic counts, including temperature (Schmidt, 1919; Tning 1952;

Marckmann, 1954; Orska, 1956) oxygen concentration (Tning, 1952; Garside, 1966), salinity (Hubbs, 1925; Heuts, 1947), light intensity (Dannevig, 1932; McHugh, 1954; MacCrimmon and Kwain, 1969) light duration (Lindsey, 1958) and osmotic 181

stress (Tay. and 3arside, 1972) The influence of temperature on the numbers of certain meristic structures in fishes has probably been studied to the greatest extent bOth in the 1abDratory and in wild populations Under- natural cond.itions there is a tendency for closely related fishes to have more parts (particularly vertebrae) towards the polar end of their range (Lindsey, 1961). within species latitudinal dines, which may be genetically fixed, are probably still the result of selection operating in relation to gradients in environmental factors. Temperature and its effect on rate of development is an- obvious factor to examine when considering meristic dines (Hubbs, 1922; Campbell and NacCrimmon, 1970) but light intensity and light duration, both of which vary regularly with latitude 9 are capable of altering mer-istic counts (McHugh, 1954; Lindey, 1958), With the above problems and factors in rnind an attempt was initiated to find if one could distinguish brown trout and sea trout by counting various meristic series of firstly, artificially produced progeny and secondly, adults and juveniles of the two types taken from natural waters. 1.82

13. Comparison of Artificially Produced Progeny

(1) Methods and problems encountered Experimental F 1 progeny were obtained from artificial

fertilization of Tweed sea trout and brown trout. Wild, adult, sea trout and brown trout were electrofished from a number of tributaries of the Tweed in the autumn of 1969

and 1970 when they were nearing spawning condition. Because of the difference in size and appearance of the

adult fish 0 no difficulty was experienced in distinguishing the freshwater-resident trout from sea trout.

In 1969 five lots of eggs were obtained, each lot from one set of parents. Two batches were "pure' sea trout, two lots were "hybrids', one lot the reciprocal of the other (female sea trout x male brown trout and female brown trout x male sea trout) , and one lot was produced from the artificial spawning of two brown trout. In 1970, only four broods resulted; namely two lots of sea trout, one lot of brown trout and an intermediate lot of brown trout x sea trout

Following fertilization in 1969 the five batches of eggs were placed in -a hatchery near Galashiels, Se1kirkshire where the water temperature during the incubation period approximated 50C. The eggs were held in running water for 93 days until hatching began. The resulting alevins were transported to fry-rearing tanks supplied with running tap water from the mains at the Department of Forestry and Natural Resources. 183

In 1970/1971,, the fertilized eggs wera immediately placed in vertical hatching trays in the Department where

the water temperature approximated 6e5 0CG HchJ.ng was initiated about 71 days after fertilization.

Originally it was anticipated that the resulting fry of the various crosses would he held for at least a year

and observation could be made on their appearance ô but problems were experienced with the holding facilities.

In1970, five days after the alevins were placed in the water supply in the Department,, losses occurred. This mortality continued until the middle of June and a total of 5500 trout had been lost. All the fry were preserved in S per cent formaldehyde. The cause of the mortality in 1970 has been attributed to concentrations of between 0.25 and 0.40 mg/l of copper (cu) in the water supply which was introduced into the water as it passed through the copper pipes of the building. Mortality of the individual trout did not occur suddenly,, but the copper appeared to interfere with the alevins ability or desire to feed. Although a few of the fry were feeding, a far greater number did not initiate . a feeding response, On close examination,, the trout were usually emaciated and in poor condition even though ground liver and Ewos No,1 trout food was available to them. When the concentration of copper in the water supply was first suspected of causing the observed mortality, correspondence with Mr, V.M. Brown of the Water Pollution Research Laboratory at Stevenage, Herts. indicated that the 184

48-hour median lethal boncerit rat j.on. of this metal for

adult trout is only about 0.5 mg/i in hard water (300 mg/1'

as CaCO 3 ) and in very soft water (20 mg/i) is as low as 0,05 mg/i. Further, he found that, although rainw trout fry took food (cladocera, copepoda) at 0i mg/i of copper, at concentrations of 0.3 mg/j, of copper and higher they showed no interest in feeding in the first 18 hours of exposure.

McKim and Benoit (1971) in studying the effect of long-term exposures of copper (Cu) on the survival, growth and reproduction of brook trout Salvelinus fontina1±s concluded that approximate concentrations of .03 and .02 mg/l. (water hardness 1 45 mg/1 CaCO 3 ) had marked adverse effects on survival and growth of alevins and juvenile trout. During the summer of 1970, a new water transport system was installed in the Department which consisted of PVC (plastic) piping which brought water direct from the mains to hatchery trays and holding tanks. Water did not come in contact with copper pipes. In 1971, the four broods of eggs hatched in the second week of February and actual feeding began on about 15 March. Fry development proceeded until approximately

14 April when a mortality of about 2 per cent of the total number of fry per day was noticed. This continued to the first week of June and a total of 4500 trout fry had been lost. Although there is some controversy in the Department as to the reason for the failure of my second attempt to maintain broods of trout fry in the laboratory ; the mortality M

may be attributed to the minute amounts of Chlorine

(supposedly an amount to give a. concentration of 0.1 mq/l but pernaps as much as 0.2 mg/].) added to the City of

Edinburgh water SUPp1V to maintain the water in -a potable condition. Sprague and Drury (1969) found that the 12-day lethal level for rainbow trout was 0.01 mg/l while in

brook trout, survival was Only 1-2 days in concentrations between 0.08 and 0.04 mg/i (Dandy8 1972). •Merkens (1958) suggested that the safe threshold for rainbow trout was perhaps as low as 0.004 mg/1.

(ii) Staining technique (after Hollister (1934) and Orska (1956)

All specimens ranged from 20 to 30 mm in length and were first preserved in a 5 per cent solution of formaldehyde. This caused the specimens to harden and so assisted handling. After remaining in the formaldehyde solution for periods up to 16 months, specimens for staining were removed and

rinsed in running water for 24 hours. Following this, the specimens were placed in a solution of 4 per cent potassium hydroxide. The fish were allowed to remain in this solution for at least 24 hours or until the vertebrae could be dimly seen for the last half to two-thirds of the length of the body. The potassium hydroxide solution was then drawn off

and hydrogen peroxide (20 vold was added and the specimen remained in this solution for at least one hour or until all pigment was lost due to bleaching.. A 1 per cent solution of alizarin red S (alizarin sulphonate sodium) in 4 per cent potassDura hydroxide was 166

made up and the specimens to be stained were placed in

this solution for a period of - 5 to 10 minutes. Then - the stain was drawn off and fresh 4 per cent potassium hydroxide was added which leached most of the stain out of the flesh in 2 to 3 days, - Finally0 the specimens were allowed to remain in a solution of 4 per cent potassium hydroxide in the following proportions of glycerine for varying periods of time depending on the size of the specimens and the amount of stain still retained.

30 per cent glycerine 50 per cent glycerine 70 per cent glycerine

The specimens were then held in 100 per cent glycerine until fin ray, branchiostegal ray and vertebral counts were made

(iii) Methods of counting rays and vertebrae

Eight meristic characters were examined for possible use in separating Tweed brown trout and sea trout (Figure 44) They were adapted from Hubbs and Lgler (1958) and were as follows:

The number of dorsal fin rays included all of the rudimentary as well as principal rays and the last two rays were counted as one. The number of anal fin rays included all of the rudimentary as well as principal rays and the last two rays were counted as one, - Figure 44

Cleared and alizarin-stained body parts of trout (S.trutta) for meristic comparison. Anal fin rays (x j. 1) Dorsal fin rays (x 15) Pectoral fin rays (x 18) Branchiostegal rays (x 10) Whole specimen (x 4.4)

a 187

. \\\

A/ B. \ \__ •\ \__ \__\ \ _\ C I

1'5

huh

E 198

3 The number of PeCtOral fin rays were - counted on the left fin and included the smallest ray at the lower or inner end of the fin base. The number of pe]vic fin rays were also counted on the left fin and included, the smallest nv at the lower

or inner end of the fin base. A snail ray preceded the first well developed ray but was bound very closely to t. This particular ray was not counted - The number of hranchstegal rays included all the rays situated on the outer side of the hyoid arch and those inserted more anteriorly and ventrally on the inner face of the arch.

The vertebral ccunt did not include the basioccipiral and terminated at the base of the hypurai plate,, so that the last three upturned ossified centre were not counted, The number of precaudal vertebrae did not include the basioccipital and extended to the last vertebra without a definite hemal spine.

The number of caudal vertebrae included the first vertebra bearing a definite hemal spine and the last vertebra before the hypural plate.

* IM

(iv). Results

Data on each of the eight meristic characters (Table 26) were graphed according to a modification of the method proposed by Huhbs and Hubbs (1953) (Figure 45u and Figure 46). The horizontal line depicts the range, the triangle the mean, the hollow rectangle one standa'd deviation on either side of the mean, and the solid black rectangle two standard errors on either side of the mean. Broad overlap of the black rectangles indicates low probability that the observed differences between- any pair of samples is significant or conversely when the solid bars just meet, the means are just about significantly different. The solid bar also indicates approximately the 95 per cent fiducial limits of the mean, Thus by this method measures of reliability and the measures of dispersion are both depicted, The number of specimens was arbitrarily held to 30 individuals from each of the eleven samples, although it was recognised at the outset that a larger number of specimens and samples would have considerably broadened the scope and effectiveness of the analysis. It was originally anticipated that a more critical

discriminant function proposed by Fisher (1936) would be used in an attempt to distinguish brown L:rout, sea trout: and their hybrids but after graphing the data according

-, to Hubbs and Huhbs ( 1953) it was decided that thisLflsb mLLIIO of analysis was not warranted because of the irregular nature of the results. Table 26. Mean number and standard error of some meristic characters of artificially produced progeny.

1970 Lot Type No. Mean counts female male dorsal anal pectoral pelvic branchio- precaudal caudal vertebrae stegal SE SE SE SE SE SE SE SE

1 ST xST 30 13.80:0.10 11.47:0.10 12.73:0.11 8,63:0,06 8.83:0.11 28.37:0.11 27.37:0.17 55,73:0.13 2 ST x ST 30 13.43:0.11 12.13:0.09 13.00:0.05 8.90:0.02 8.70:0.14 28.77:0.12 27.40:0.12 56.17:0.10 3 ST xBT 30 13.50:0.09 11.87:0.12 12.83:0,08 9.00:0 9.07:0.13 28.23:0.11 28.40:0,11 5E,63:0.13 4 BT xST 30 14.10:0.06 11.93:0.11 12.93:0.05 8.87:0.02 8.47:0.14 27.97:0.17 28.40:0.12 56,37:0.15 5 .BT XBT 30 13,83:0.08 12.17:0.10 13.07:0.10 9.00:0 8.87:0.11 27.07:0.14 28.03:0.13 55,10:0.11

ST x ST 30 14.07:0.12 11.47:0,10 13.33:0.09 9,00:0 9.20:0,11 26.43:0,09 27.83:0,14 54.26:0,13 ST xST 30 14,23:0.13 11.53:0.11 13,20:0.08 9.00:0 9.50:0.13 26.80:0.11 28.97:0.16 55,770.18

ST XBT 30 14.17:0.13 11.20:0.10 13,10:0.11. 9.00:0 9.33:0.15 27.00:0,14 .28.770.14 55.71:0.12 BT x ET 30 14,17:0.12 11.63:0,10 1340;0,06 903:0 10,00:0,11 27.07:0.13 28.43:0,19 55,500,20

- 15 14.40:0,18 12.33:0,12 13.80:0.10 9.00:0 11,00:0.20 26,93:0,18 29.20:0,20 56.13:014

- 30 13.90:0.09 11,63:0.10 13,27:0,08 8.97:0,01 9.63:0.10 27.03:0,12 28,83:0,15 55,870.11

I-,

C ç. .. C __-,_'-- -.a------

Figure 45 ,._.Lt' -'r:--,_ .-.-.. Variation ofof the dorsal fin, anal fin, pectoral fin and branchiostegal ray counts of trout (S.trutta)

. illustrated by the, Hubbs and Huibs (953) graphic

• method. (The base line indicates the range the

• small triangle, the mean; the solid bar, two

standard errors on either side of the mean and

• the ..hollow bar, one standard deviation; on either

• ..... side of the mcan). -

. ___•*• • - - - . •- r;-3,-a

• t.* ....-, •• . -

• •. •. .

•&. -• -.• • - I'C', -L J

Id V l•3 13 14

0OSA(. FIN RAYS . AML FIN RMS:

r_Z I 37 ST C 1969.

I rflTL__, 37 x ST CI99i

5' xIn I99)

-. ST. ST 1963 JLt2tk1iL -

. e:r. x (U (S4,9)

-, ST 51 I 1390) çnntticnt._ --

Si Si i !-'701

ST (1970)

Six 91 04X)

KIRK (1970) .

hRK (I:)7l) -

II 12 13 14 - 7 6 9 13 II 12

PEC10R.L FIN 1AYS CFA:-(CHiOSTCGLt RAVS

'ss

STK ST (1969)

ST 4 (ST (1969)

ST (1969)

.91 ,c St (969)

SixST (1970)

STIcSTIIS/0)

r__C_30Sk5iCflflt3. -, CIT Ic 51 0973; .

r1r ST (197))

CJ KiRK (1070)

(197 fl

-J •

- - vie ...-- .

Figure 46 4•••• -

a - • r..Vtiation- '. of the precüda1, caudal,- and toal vertebrae of trout (rLt%a) illustrated by .Tthè Hubbs and Hubbs (i93Y'graphic method. (The....base Line indicatësthe range, the small trianqle the mean;- the solid bar e two standard be 0 .t ,-- errors on either..sideaoftheftme.arvtnd th3 hollov I ...... bar, one standard deviation on either side of

. ,.;...... the mean)

.a4.CtvC- L;4C,flt..

..'r .-

...... -- ...... --.

. -r . 3.92

tC 27 20 29 33 2 27 23 29

P?ECA'JOAL VEFflEIflAE cgurk. V1EJt

ST M ST (T3C9)

z UT (1969)

9TxS

ETx ST t969)

ST 11970)

SIX ST't"70)

STx ST cem - -

Blx UT (197O) - .

KIRK (1970) -

KIRK (1371)

5 1 53 34 t5 € 57 58

TOTAL VEt-JC'RAE

Six 5? ( 1369)

ztsi (1969)

ST nBT (1C)

F3TxST (1969)

FsTxBT(1%3)

St x ST ( 19 N)

tI m ST 09701

81 x ST (¶370)

ST St (1970)

KIRK (1970)

URK (IS 711 C. 3.93

The amount of variation in a number of the meristic series,, even though within each year the various batches of developing embryos were held under identical condition5 9 would indicate the effect of some intrinsic or genetic

factor passed to the offspring from the parents. This factor, which causes one meristic series to be significantly different from another, does not appear to be common to the sea trout Q the brown trout or the "hybrid" broods. Examples of this phenomenon appear in the anal fin ray

counts of the two STxST 11969) crosses and in the dorsal fin ray counts of the STxBT (1969) and the BTxST (1969) crosses where graphical comparisons indicate differences

that are statistically significant (5% level) counts of total vertebrae of the two STxST (1970) crosses also show a significant difference (5% level). Little can be said when attempting to compare the broods of 1970 and those of 1971 as development after

fertilization occurred at different temperatures. The same is perhaps true for the fry taken from Kirk Burn in

3970 and 1971. The mean number of dorsal anal, pectoral and branchiostegal rays are significantly different (5% level) for the two years; but this is understandable as this difference is most probably caused by the influence of some environmental factor such as temperature. 194

C. Comparison of Adult Brown Trout and Sea Trout

Methods and techniques

Adult brown trout and sea trout were electrofished from a number of streams tcibutary to the upper Tweed in the autumns of 1969, 1970 and 1971. Parents of the artificially produced progeny were also used. In tota1 23 sea trout and 30 brown trout were retained in a frozen state until they were required 1- or the counts. Ray and vertebral counts were performed in the same mariner as those of the stained trout fry but enumeration was taken from radiographs which were made using a standard X-ray diagnostic machine 0 Satisfactory exposures

ranged from 0.2 to 05 seconds using 50-100 kv and 25 amp depending on the size of the specimen. The processed plates were exainined.on a light table (Figure 47) Gillraker counts were taken from the first gill arch of the left side of the fish. All rudimentary rakers were included in the counts and if a gillraker straddled the angle of the arch, it was included in the count of the lower or ventral limb . In counting the pyloric caeca all. tips were enumerated.

Results and their interpretation

Table 27 indicates the mean values for the counts performed. The "U' test showed. that for .the numbers of precaudal caudal and total vertebrae; dorsal gill rakers; pyloric caeca; pectoral and anal fin rays there was no Figure 47 Radiograph of adult trout (S.trutta) body parts for meristic comparison. Pectoral fin Pelvic fin Branchiostegal rays I).. Anal fin F. Dorsal fin 195

1/ 9

.r,.,-_

E Table 27. Mean number of some meristic charactrsof adult brown trout and 4ea trout collected from the Tweed.

Type 'No. of vertebrae gill takers pyloric branchiostegal fin rays sped- precaudal caudal total dorsal ventral total cacca rays mci-is - pectoral dorsal pelvic anal

rout 23 28.42 29.58 58.00 6.75 12.00 18.75 53.00 10,75 13.50 13.58 10,00 11.33 troift 30 28.73 29.40 2.13 6.47 11.33 17-.80 52.38 10.21 13.07 12.57 9.36 1L57

Significance * * * * *

denotes sigMficint difference at the 5% level.

Cl' 197

significant difference at S per cent level between brown trout and sea trout,.

A significant difference did appear9 however, between the ventral and total gillrakers; branchiostegal and

dorsal and pelvic fin rays of brown trout and sea trout, Although a significant difference was shown for the above five meristic series,, these results should be

ixfterpreted most conservatively. Counts of gilirakers have proved useful in distinguishing races of kokenee, Oncorhynchus nerka Q (Vernon, 1957) and whitefish,

Corecionus 5P.t (Lindroth, 1957). More recently, giliraker numbers have been found to increase with age in the grayling, Thymailus thymailus, (Peterson, 1968), in the whitefish (Svrdson, 1966) and in the Pacific thread herring, ()pisthonema libertate, (Berry and Barrett, 1963). Concerning the significant difference found between the branchiostegal, dorsal and pelvic rays, difficulty was experienced in enumerating the smallest rays at either end of the individual series of the smaller soecimens

Since the brown trout were, on the whole 9 smaller than the sea trout, some of the rays on the radiographs perhaps were not indicated.

D. Numbers of Vertebrae of Juvenile Trout from Different Locations

(I.) Methods and the results

Samples of juvenile trout were collected by electra- fishing tributaries of the upper Tweed and by seine netting 198

the Tweed itself near Norham Bridge. Two collections were made in Kirk Burn (A 0 B) while onecollect ion in Eddleston Burn could be divided by appearance into trout showing a silver appearance (A) and those having no sea trout characteristics W. Vertebral counts were made from radiographs of the whole fish (Figure 48).

The number of vertebrae of 20 fish from each location were distinguished as previously.

The mean number of total vertebrae for each sample of 20 trout is depicted in Figure 49 following the method of Hubbs and Hubbs (1953). As there is a broad overlap of the black rectangles, the observed differences between the mean vertebral counts of the samples are probably very slight and not significant (50' level). Figure 48 Radiograph of juvenile trout (S.trutta) for the counting of total number of vertebrae (actual size).

I r .'

* *-

Figure 49 Variation of the total number of vertebrae of juvenile trout (s.trutta) from various locations in the Tweed, illustrated by the Hubbs and Hubbs (1953) grà1fic inethod. '(The base line indicates the. range, the small triangle 8 the mean; the solid bar, two standard errors oh either'-side-d' the mean and the hollow bar,, one standard deviation on either

side of the - ,mean) ,;

• - -. _.t I , ._. f . .ft -? . • "C.

-I 200

57 58 59 •60 61

VERTEBRAE

NORHAM fl•__iflrca

GLENTRESS

DEAD :

KIRK tid 9rT: i. 1: •:;.:.:t .. L

- K!RKupstçenJ..

SOONK 0 PE -

EDDLESTON A silver cErJ

CDDI..ESTCN B brown 201

E. Discussion

Meristic characters exhibit plasticity under the

influence of environmental factors, - especially temperature during the incubation period and early larval life

(Schmidt., 19.20; Tning 1944; Lindsey, 19541. However numerous studies have shown a genetic as well as phenotypic basis for rneristic character in various fish species (Barlow,, 1961). In the family Salmonidae 0 meristic and/or morphometric characters have proved useful as a method to

distinguish subpopuiation.s of salinorsids (Vernon ? 1957; McPhail, 1961; Amos, Anas and Pearson, 1963) where the cause of the dbserved effect is not important.

Coupled with additional biological and biochemical information, the technique of counting meris€ic series can provide useful evidence in delimiting populations. Tester (1949) when considering populations of Pacific herring Clupea pailasfi, found that the nature of mixtures of stocks was such that the study of mean vertebral counts had to be supplemented by tagging studies,, The number of studies of meristj.c characters of ariádromous and freshwater-resident populations of fish

species is somewhat limited, wilder (1947) studied populations of the Atlantic salmon, Sall -no salar L. and the lake salmon Salrno salar sebaso (Girard) and found that when the two types of approximately the same size were compared, no consistent differences in the meristic

characters could be demonstrated. He concluded that any di.fferenoes that were observed were not inherent but the 202

result of differences in the environment and diet. Wilder (1952) also made a comparative study of anadromous and freshwater populations of brook trout Salvelinus fontina1is in the Moser River system of Nova Scotia, He found that the so-called sea trout and freshwater trout did not differ significantly in any of the meristic - structures counted and so he concluded that they constituted one taxonomic unit. Savvaitova (1960) compared dwarf or "subsidiary" males of the Dolly Varden, Saivelim !L2kE, which spend their whole lives in freshwater, with their anadromous counterpart. She did find some morphological and meristic differences but •attributed then to dissimilarity in size and so concluded that both male forms were members of one population.. While studying the biology of a landlocked form of the normally catadromous salinoniform fish 9 Galaxias culatns : Pollard (1971) concluded, after studying the two types mor6hologicaily9 that the landlocked form should be regarded as only an isolated 'ecological rac&'. On the other hand, there have been a number of studies of meristic characters of fish populations which have shown distinguishable differences between types of the same species. McCart and Craig (1971) compared the gill-raker and pyloric caeca counts of populations of anadromous and freshwater- resident arctic char, Salvelinus alpinus, in a river system in Alaska and found that the anadrbmous type had lower mean giliraker and pyloric caeca counts. However, the two types appear to be at least partially, if not wholly reproductively 203

isolated as the freshwater-resident char are lake-dwelling

with no anaciromous component and entry of arsadtcmous stock

into the lakes is blocked because of nearly impassable fails.

Meristic differences have been found between spring

and autumn spawning. AtLantic herring, Cltpea harencius off

southwestern Newfoundland which has been attributed to

differences in water temperature during larval develooment

and differences in developmental rate (Parsons aftd Hodder,

1971). Smith (1969) has been able to separate summer

and winter spawning races of steelhead trout, Saint crairdneri.

He found that some taxonomic characteristics were heritable

to the extent that meristic series ; such as numbers of

vertebrae 0 gillrakers and parr marks in F 1 yearling fishy

could be used. However detailed examination of migratory

behaviour and taxonomic and physiological characters of

steelbead in one river indicated that the two races do not

interbreed and so they exhibit reproductive isolation.

The concept of reproductive or spatial isolation

appears necessary before racial differences can be

distinguished between two populations. Thompson (1930)

as cited in Gordon (1957),suggested that isolation is a more effective cause of racial differences within a fish

species than differences in environments to which fish may

respond. Further, he stated that as long as populations

are isolated, heritable differences from the original condition will accumulate. If however 9 interbreeding is occasionally possible between oartially isolated populations, these differences will tend to be less pronounced. 204

Comparison of various méristic series of adult

brown trout and sea trout from tributaries of the Tweed have shown some differences but because of the small sample sizes, these must he viewed with reservation As it has been found that intermingling of the two types of trout do occur on the spawning grounds the possibility of separating the two using meristic analysis of adult trout appears rather remote., - The influence of heredity in maintaining the identity of two races of the brown trout, reared in a similar

environment was studied by Alm (1948) The large silvery form lacustris or ferox was obtained from large Swedish lakes, while the small brownish form -farto was taken

from small brooks in forests and mountains. After maintaining the two races for two generations he found differences in the colouration of certain 'fins and the

time of reaching soxue]. maturity. He concluded that the racial divergence between the two forms was clue to genetic' factors.

Differences which appear in the meristic counts of the ,artificially produced progeny in this study, though in a number of instances significant do not appear to

follow a consistent pattern. Even though the various broods each year were kept under identical conditions from the time of fertilization to that of hatching, the amount of variation between batches with similar types of parents make meaningful analysis impossible. This variation can only be due to the effect of the parents 1 that is high 205 parental vertebral numbers are associated with high numbers in the offspring and lpw parental numbers are. associated with low counts in the offspring,. The rnerjstic comparison of Tweed brown trout and sea trout has not been altogether successful but it perhaps has given some indication that t:ne two types of trout are so similar morphologically that this method of analysis is not so refined as to effect a. separation. fl C-'- U

V1 9 DISCUSSION AND CONCLUSIONS

While studying the trout population of the Dabs

River in Norway 0 Dahl (1910 1933) was unable to find

in the non-migratory stock 0 females older than four winters or spawning females that had not been to seas. He states that similar conditions are met-fl in many of the Norwegian streams to which sea trout have access. In examining the stock which had never been to the sea r he found that in many places males and females occurred in equal proportions in the' youngest year classes, whereas the older year classes consisted mainly, and at the end exclusively, of males. In some rivers, he continues, the proportion of males and females may be only slightly affected by age so that spawning females may also occur which have never migrated -

The trout population of the River Tweed exhibits many of the characteristics outlined by Dahl (1910) in his

studies. The observed sex ratios of the freshwater resident trout, the sea trout smolts and to a lesser extent the anadronious stock indicate that the relationship of Tweed sea trout and brown trout may be very complex. This fact is borne out by the spawning behaviour of these two trout types on the spawning grounds where male brown trout may actually participate in the spawning of sea trout. This phenomenon has been found to occur in a number of saimonid species which are composed of both anadromous and freshwater forms. Perhaps the best documented example of this is the sockeye salmon (Oncorhynchus nerka) of 207

Cultus Lake, British Columbia (Ricker, 1938) where "residuals" from sea-run fish, occur in addition to a self-maintaining stock of freshwater ]cokance. In

attempting to explain the rather complex segregation 9

by sex and size, of the stocks of sockeye salmon s Ricker (1938) had to make the following assumptions: the forces, which bring about migration, operate more strongly as the young fish increase in size and for fish of a given size 1 more strongly on females than on males. that after a certain size is reached, something connected with approaching maturity inhibits the effects of (l) v this latter influence being much stronger among males than among females. The assumption that factors influencing migration act more strongly on larger fish has been given support by the present study as well as by Fessier and Wagner (1969) for steelhead trout (Salmo jaeri) and by Johnson and Eales (1970) for Atlantic salmon. That such factors act more strongly in female fish is suggested by disproportionate sex ratios of adult fish and the preponderance of females in samples of silvered sea trout srnolts taken from Dead Burn and Norham Bridge, Although more samples of actual smolts are required before a definite conclusion can be reached, there appears to be a distinct possibility that the brown trout which move into the tributaries of the Tweed during the spawning season are at least part1y, progeny of sea trout, 208

Any attempt to delimit two types of trout which actually intermix and reinforce one another's ranks to

the degree that appears to occur, in the Teed makes it almost impossible to find a criterion for phenotypic separation. At least some measure of reproductive

isolation is required before differences in meristic characters can be utilized to distinguish two stocks.

The rearing of artificially produced progeny of supposedly known parentage will limit any differences in rr.eristic

characters to the effect of genetic factors. However when this was attempted with brown trout and sea trout 9 no consistent differences or trends were found

There have been a number of experiments desiqrieci to find physiological differences between brown trout and sea trout, Spaas (1960) compared the temperature resistance at different ages and in relation to the individual size and found that the differences between sea trout and brown trout were not statistically significant. Gordon (1959) studying the osmotic and ionic regulation in Scottish brown trout and sea trout, concluded that the patterns of regulation were very nearly the same in both forms. Landgrebe (1941), on the other hand, suggested that the anadromous and non-seagoing types of brown trout are genetically distinct as he was able to cause "silvering" in brown trout after injecting them for one month with thyroid extract s He considered that the difference between the two types was "associated with functional activity of the thyroid gland,' 239

The electrcphoretic comparison of blood serum protein has proved to he a useful technique in fish taxonomy. However the amount of variability experienced in the electropherograms and polyacrylamide gels would indicate that to separate brown trout and sea trout from

the Tweed. more refined techniques are required. The sare may, perhaps, he said of the comparison of molecular weights of ribosomal RNA, but, I feel, this techhique may eventually be of use in fish systematics and taxonomy. .The behavioural, genetical and morphological evidende accumulated in this study, strongly indicates that the freshwater-resident trout and anadromous sea trout of the Tweed are taxonomically similar or nearly so To separate these types of trout in the Tweed more sophisticated techniques, that perhaps at present are just being developed must be utilised,, 210

VII. SUGGESTIONS TOR FUTURE RESEARCH

An attempt has been made to ecologically and

physiologically distinguish brown trout and sea trout in the Tweed, and to discover the factors which influence

the population biology of the two types. If any management plan for the Tweed is to he meaningful, it is essential that the number of and identity of subpopulations within

the brown trout/sea trout complex be established. The following areas should be profitable for future research:

continuation of the tagging progranuna of both sea trout kelts and smolts as well as the brown trout which move from the Tweed into the tributaries to spawn. compare the biology and vital statistics of a non-migrant population of brown trout isolated above a waterfall or barrier with that of a more migrant population. investigate the sex composition of sea trout smolts captured at various locations and times on the Tweed and relate the results to the sex ratio of adult brown trout and sea trout,

examine the environmental and physiological factors which influence the upstream migration of soawning sea trout and then compare these with those factors which cause the downstream movement of smolts,

further studies on the reproductive biology of sea trout and the importance of the freshwater-resident brown trout during spawning.

intensive investigation into growth, mortality and 21]. movement of the young stages of sea trout and notation

of possible differences from brown trout. - electrophoretic studies of the two types of trout but utilise brown trout of known ancestry; rely on definite tissue or serum Drotein (enzymes) groups using a polyacrylamide or starch gel substrate, refine the technique of determining the molecular weights of ribosomal RNA so that, in the first instance, fish species can be separated and then, if possible,, attempt to distinguish brown trout and sea trout. 212

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Appendix 1. Daily mean temperature (mean of means of maximum and means of minimum) and total rainfall at Glentress D Peebleshire, 1970-71. Monthly Weather Reports, H.M. Meteorolodical Office, Met, 0. 837,

Temperature ( °c) Total rainfall (mm) 1970 January 1.9 78 February 0.7 76 March 2 7 50 April 5.0 62 May 11. 3 17 June 14.5 43 July 12.9 112 August 14.3 60 September 12.3 72 October 8.9 90 November 5.0 83 December - 62 1971 January 3.3 53 February 4.1 62 March - -- April 6.5 47 May 9.5 55 June 10.7 43 July 14.7 97 August 13.5 94 Septeither 12.7 24 October 9 0 9 61 Appendix 2. Record averages of River Tweed flow station at Peebl3s 9 Peebieshire

- (period of record averages October 1959 - September 1966) The surface water year book of Great Britain (1971). H.M. Stationery Office! London 0 186p

Oct Nov Dec jan Feb March Apr May June July Aug Sept Year

Gauged flow m 3/s 1 16.31 Run-off (mm) 66 105 108 92 73 71 58 40 29 26 58 71 797 Rainfall (mm) 114 151 115 113 78 76 88 74 83 91 149 134 1264 1916/50 av rainfall mm 132 122 122 137 91 79 79 84 76 107 114 109 1252

Appendix 3. Species other than trout captured in the 7 study streams 1971-72.

Date Species Number Remarks KIRK

Spring 1972 brook lamprey. B captured in fry trap near mouth sprinc: S;2 :el 3 smolt trap autu17l-72 salmon 27 diseased adults at the mouth

- r '1 1 IS ripe male pa r moving upstream

T c a'7 '," .. rn

autumn 1971 salmon 4 ripe adults spring 1971 126+ sinolts captured in fyke net 1971 eel .. 7 captured in fyke net 1971 river lamprey 5 II

DEAD

134.71 salmon 2 smolts captured in fyke net spring 1971 brook lamprey numerous vicinity of fyke net 20.4.71 grayling I captured in fyke net

spring 1971 eel 1 91 11 11

GLENTRESS

None Reproduced from Report on Forest Research, 1971

The Ecological Relationship of Brown Trout and Sea Trout in Forest Streams

By J. S. CAMPBELL

Depaitment of Forestry and Natural Resources, University of Edinburgh

Printed in England by Her Majesty's Stationery Office at St. Stephen's Parliamentary Press

(886823) (9) Dd. 558311 200 11171 THE ECOLOGICAL RELATIONSHIP OF BROWN TROUT AND SEA TROUT IN FOREST STREAMS By J. S. CAMPBELL

Department of'Forestry and Natural Resources. University of Edinburgh

Duringthe last year trout poulations in both Glentress Burn, (lentress Forest, and Kirk Burn, Cardrona Forest (Peeblesshire), have bedn investi- gaSd....ese forest streaths were chosen because they were reasonably accessible, large enough to possess a full biota, and small enough to be experimentally fished at reasonable cost. Also, they permit complete counts of all upstream and downstream migrants and, thus, avoid sampling errors. Glentress Burn has a resident population of slow-growing brown trout. This was sampled by means of a fish trap to capture downstream migrants, and by electrofishing gear. Kirk Burn, on the other hand, contains both brown trout and sea trout. Over the year emphasis has been placed on a study of the sea trout. Sections of the burn were electrofished monthly to calculate changes in production, distribution and vital statistics . of young fish. During the autumn, box traps were erected to capture upstream and downstream migrant fish. A total of 41 adult sea trout and 112 brown trout were captured during their ascent and tagged and released upstream. Environmental factors such as stream flow, discharge, temperature fluctuations, and water chemistry of Kirk Burn have been monitored. The effect of these and other physical characteristics on the biomass of fish present is being prepared for publication.

140 Reprint from Tweed Commissioners Annual Report for 1971 Dated 6th March 1972 -

A PROGRAMME FOR THE STUDY OF THE RELATIONSHIP OF BROWN TROUT AND SEA TROUT

J. SCOTT CAMPBELL Department of Forestry and Natural Resources. University of Edinburgh

Printed by the Tweeddale Press Ltd., Hawick A PROGRAMME FOR THE STUDY OF THE RELATIONSHIP OF BROWN TROUT AND SEA TROUT by J. Scott Campbell, Department of Forestry and Natural Resources. University of Edinburgh

- Introduction Brown trout and sea trout are, at the present time, commonly regarded by fishery biologists as constituting a single species, Sabno trutta Linnaeus. As is well known, certain individuals in the course of their life history migrate to and from the sea, while others remain permanently in freshwater. Other species of salmonids which have access to the sea also exhibit this phenomenon. The rainbow trout, Sairno gairdneri, of western North America has a "steelhead" form whose life history is quite similar to that of sea trout. Also the North American brook trout, Sa/velinus fontina/is, the cutthroat trout, Sa/mo clarki, and the Arctic char, Salvelinus alpinus, have forms which spend at least a portion of their life in estuaries or the sea itself. Whether fishes 'which behave so differently constitute separate populations with genetic differences, or whether the differences in behaviour are merely due to environmental pressures, has been a subject of speculation among ichthyologists for more than 'a century. As early as 1866, Gunther catagorised many populations of trout into separate species and gave diagnostic characters for each species. Day (1887) considered that sea trout and brown 'trout were local races of one species, and that the migratory or resident habits were the result of environmental differences and the markings dependent on immediate surroundings. Began '(1911) also treats the two types as one species since he could find no structural difference between the two forms. In 1916 Lamond arrived at the same conclusion while studying the trout of the Clyde area but at the same time he suggested that there might be other populations of sea trout that ranged farther out to sea and were distinct from the brown trout. Henking (1929) and Altnoder (1934) felt that one should separate brown trout and sea trout at the specific level, but more recently Alm (1949), Fabricius (1953), Trewavas (1953) and Frost and Brown (1967) have concluded that the two are only forms of Sabno trutta. To further substantiate the premise that one is dealing with two types of trout, 'there have been reports of progeny of introduced brown trout resembling sea trout in habit and appearance. Accord- ing to Day (1887) this was the case in New Zealand where non-migratory brown trout assumed the habits and appearance of sea trout. However, Scott (1964) states that this is not based on substantial evidence as there were also introductions of sea trout made at the same time. Frost and Brown (1967) report that sea trout are now found on Vancouver Island in Canada where they are considered to be descendents of introduced brown trout. The Falkland Islands have a sea run population of trout which are descended from brown trout obtained from the Chilean government in 1947 (Arrowsmith and Pentelow, 1965). At a more local level, there has been some controversy over the origin, nature and terminology of many forms of trout in Tweed. As early as 1864, the River Tweed Commissioners appointed a Com- mittee to collect information on the different kinds of "salmon" in Tweed. The results obtained from a questionnaire they sent out on this subject was published as a report in 1867. There are still differing local opinions on this subject. In any stream management plan it is necessary to understand the ecological relationship existing between the various fish species present. For this reason a thorough understanding of the relationships existing among salmon and the two types of trout is needed. Unfortunately it is not possible to distinguish young "brown" from young "sea" trout, until the latter become silver when moving to the sea as smolts, and yet it is most necessary that one should, as while young sea trout are frequenting the stream for a limited period, young brown trout are present for a greater length of time. For this reason a study of the biology and ecological relationships of brown trout and sea trout of the River Tweed was initiated in 1970 under the supervision of Dr. Derek Mills. The study was approached in two ways: (a) ecologically and (b) taxonomically. The ecological approach. Actual populations of trout were sampled using both electrofishing apparatus and - box traps to see if one could distinguish the two forms of trout. Such parameters as age structure, growth, size and movement were recorded because little is known of the effect of young and adult sea trout on resident brown trout. The trout populations in Kirk, Glensax and Glentress burns, situated east of Peebles, were investigated. These streams were chosen because they were reasonably accessible, small enough to be experimentally fished at reasonable cost and they permitted complete counts of all upstream and downstream migrants. - During the autumn of 1970 and 1971 the complete spawning migration of brown trout and sea trout in 'Kirk Burn was trapped, tagged and released. Also the seaward movements of sea trout smolts was monitored so that these fish could be tagged, the age and size frequencies calculated and the appearance of these migrants recorded. Approximately 500 trout have been tagged in an attempt to follow their subsequent movements. Preliminary observations of the spawning habits of brown trout and sea trout suggest that the two may actually "hybridize" in nature as female sea trout and male brown trout have been observed spawning together in Kirk Burn. The taxonomic approach. Sea trout and brown trout were artificially crossed in all possible combinations to observe differences in form of the alevins. Numbers of vertebrae, fin rays and branchiostegal rays are being counted in each cross and compared one with the other as well as with the parents. Also the "hybrid" progeny of a brown trout/sea trout cross 'have been kept in a hatchery near Montrose, Angus. These fish will be tagged and released in K-irk Burn to observe if they have inherited the migratory habit of the sea trout or the nonmigratory tendency of the brown trout. Brown trout and sea trout are also being compared using the technique known as electrophoretic analysis of blood serum proteins. When a sample of trout serum is placed on a substrate and subjected to an electric current the various protein groups in the serum migrate at different rates. After staining, these groups exhibit a pattern which is specific for different species of fish. This technique is now so refined that individuals from different populations can be distinguished, in this particular work, it will be especially interesting to observe the serum protein pattern of the brown trout/sea trout "hybrids". This study will be completed in the summer of 1972 and results published shortly afterwards

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