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STUDIES ON COOPERIA CURTICEI (RANSOM 1 907) A NEMATODE
PARAS ITE OF SHEEP
A thesis
presented in partial ful filment of the requirements for the degree of
Doc t or of Phi l os ophy at Massey University
JAGJIT S INGH AHLUWALIA 1 970
i.
ABSTRACT
This the sis records in part I studies on th e e c ology of the fre e living stage s of Cooperia curt icei , b oth under c ont rolled and natural c onditi ons .
At constant t empe ratures free living stages develop ed throughout the temperature range of 1 0-37 0 c. At all t emperatures each larval stage occ upied th e s ame prop ortion of the to tal developmental time t o reach the infec tive stage . The relati onship be tween the rate of deve l opment in l og days and t emperature wa s found to be linear . Under natural c onditions the rate of development was most strongly c orrelated with mean maximum air t emperat ure and was not significantly d ifferent to that
obs erved under controlle d conditions . When faecal cultures were kept at 10° C, 27°C and 37°C a higher prop ortion of eggs c ompleted d evel opment t o the infective stage at 27°C than at the o ther t e mperatures . Unde r natural conditions t h e percent age rec overy was influenc e d b y weather c onditions particularly rainfall .
Submergenc e of the free living stages in wate r inhibited their further developme nt . First and second stage larvae survive d longest at t emperatures b e tween 5°C and 1 5°C but for a much-shorter time than in
fectiv e l arvae . Between the extreme s of -6 and 52°C, the longest s ur
vival of infective larvae was 3 1 2 days at 1 0°C.
Techniques are described for th e rec overy of Cooperia c urticei larvae from sample units of pasture, soil and faec al p ellet s . Unde r natural conditions the maximum survival of larvae from monthly experi
ments range d from 9 - 2 6 weeks . Naximum survival was particularly influenced by t emperature . Infe c tive larvae survived through th e winter . There was an exp onential r elationship b e t ween the perc entage survival and percentage of larvae recovered from the herbage . Vertical migration of larvae appear ed to b e primarily affe cted by rainfall and evaporation .
It is c onclude d that infec tive larvae of Cooperia cur t ic ei are available to grazing sheep throughout the year . The oret ic al ly the nemato d e can comp le t e from 9 - 1 1 generations in each year .
iio Part II of this the sis r e c ords experiments on the relationshi p b etween Cooperia c urt i c e i and the h os t sheep .
Experiments carri ed out in
�
and in vitro d emonstrated that infe c t iv e larvae o f C . cur t i c e i exshe ath under c onditions provided by the rumen . The proce ss of exshe athment was similar to that d escribed f or H . cont ortus.A seri e s of experimental obs ervati ons were made on the effe c t of Cooper ia cur t i c ei infection in sheep using animals of differing age s , on d ifferent diets and with vari ous s izes of inf e c ti on . The prepatent
period of infe c ti on was 1 4- 1 6 days . Peak egg c ounts were re c orded 5-7 days af ter inf e ction became pat ent . There aft er th ey d e cl ined gradually in sh e ep given 1 0 , 000 larvae but in she ep given 50 , 000 to 1 00,000 lar
vae the decl ine was more abrupt. The .egg output per female worm was found to range up to 1 , 958 e ggs per day.
No c linic al sign of inf e c tion was obs erved from any experiment
al animal . Body weights , w o ol growth and blood anal yse s sh owed no signific ant changes and no gro s s les ions or significant hist opathologi
cal changes were obs erve d . Th e results ind i cate a well balanc ed relati onship b et w e en C . curtic e i and the she e p .
The distribut ion o f the C . curt icei i n the small int e stine was skewed , and most of the worms were re c overed from 5-1 0 feet from th e gastric pylorus . A pred ominanc e of f emal e worms was obs erved at all levels of the small int e s t ine. Maximum per c entage r e c overy of C.curti
££i was observed in sheep given 1 0 , 000 larvae. Experimental animals with h igher dos e s besides giving a lower rat e of recovery showed inhibi
tion of d evelopme nt and stunte d growth of w orms .
Serum and int e s tinal mucus sample s from inf e c t e d animals wer e t e s t ed for pre cipitating antibodies by gel diffusion against five antigens . Ant igens were prepared from first stage, s e cond stage , ensheathed third s tage , exsheathed third stage larvae and exsh eathing fluid . Variabl e numbers of pre cipitin lines were obtained with s erum and mucus from infec t e d sh e ep more than 6-7 months old . She ep 2-3 m onths old showed no s uch resp onse but did show evidence of an ac quire d resistance to inf e c t i o n .
ACKNOVJLEDGEMENT
I am pleased t o acknowledge and thank my Chief Sup ervisor, Dr W . A.G . Charl e s t on , f or the stimulation , guidance and sincere con
s truc tive criticism that he has given me during this period . I am espe cially ind ebt e d t o Mr P�L� Car ter for as sisting me with many of the exp eriments in this the sis�
iii ..
The constructive c riticism of D r B.W. Mankte l ow , Pro f e s s or of Animal Health and Dr L . S . Forbes is much appreciated . I am als o grate•
ful t o Mr S . Hus sain (D epartment of Mathematics) for his advi c e on
st atistical analysis ; t o Dr K. M . Moriarty (Department of Animal Health) f or his guidanc e in the immunological s tudies ; and t o Dr R . M. Gre enway (Department of Bio ch emistry) f or h e lping me in th e protBin e s t imations on antigens . Mis s Y. Gray, D.S.I. R. , has been most helpful in suppl y
ing the mete orologi cal data .
I am als o thankful for th e as sistance of th e s taff of the his t o
pathology section of the Animal Health Department and of the Library, Mas s ey Universit y .
I would like t o thank Miss Ann Waugh and Mrs Rose Couling for t yping the thesis . Mr P . Herbert kindly print e d the figures.,
I would like t o expr ess m y gratitude to the Governments of India and New Zealand , and parti cul arly the Bihar Governm ent and the University Grants Committ e e, New Zealand , for granting me the opp or
t unity and financial assist ance to undertake thi s work .
Finall y , I wish to expre s s my appreciati on t o m y vufe , Lakhinde r , f or her enc ouragement and support through out the whole p eriod of this stud y .
CHAPTER I.
CONTENTS
ABSTRACT
ACKNOWLEDGEMENT INDEX TO FIGURES INTRODUCTION
PART I . ECOLOGY OF COOPERIA CURTICEI REVIEW OF LITERATURE
1 . CLASS IFICAT ION AND GENERAL
DESCRIPTION OF THE GENUS COOPERIA 2 . MORPHOLOGY AND DEVELOPMENT OF THE PRE-PARAS IT IC STAGES AT DIFFERENT TEMPERATURES
i ) The egg
i i ) First and s e c ond stage larvae iii ) Third stage infective larvae 3 . S URVIVAL OF PRE-PARASITIC STAGES
UNDER LABORATORY CONDITIONS
Page No .
i ..
iii.
iVa 1 .
4 ..
4 . 6 ..
8.
i ) The egg 1 0 .
i i ) First and sec ond stage larvae 1 0 . i i i ) Third s tage inf e c t ive larvae 1 0 . 4 . E COLOGY OF FREE LIVING STAGES
i ) General c onsiderat i ons i i ) Ec ol ogy and d evel opm ent of
free living stages iii ) Ec ology and survival of
infec tive larvae
iv ) Higrati on of inf e c tive larvae 5• SEASONAL VARIATION OF WORH BURDENS OF
COOPERIA CURTICEI IN SHEEP
1 1 . 1 1 .
1 2 .
1 4 . 1 7 .
CHAPTER II.
CHAPTER III .
GENERAL MATERIALS AND METHODS
1 o GEOGRAPHICAL DEFINITION OF t�NAWATU 23.
2 . THE ESTABLISHMENT AND t-'lA INTENANCE OF
A PURE CULTURE OF COOPERIA CURTICEI 23 . 3 . TECHNIQUE USED FOR EGG COUNTS 24 .
4. ROUT INE CULTURE OF FAECES 24 . 5• TECHNIQUE USED FOR LARVAL COUNTS 24 .
6 . PARASITE-FREE SHEEP 2 5 .
S TUDY ON DEVELOPMENT AND SURVIVAL OF THE FREE LIVING STAGES OF COOPERIA CURTICEI UNDER LABORATORY CONDIT IONS .
INTRODUCTION
1 . DEVELOPMENTAL STUDY OF COOPERIA CURT ICEI AT DIFFERENT TEHPERATURES i )
ii )
Materials and methods Results
2 . EFFECT OF EXCESS MO ISTURE ON DEVELOP•
MENT OF THE EGGS AND PRE- INFECT IVE
2 6 .
2 6.
26 . 28o
LARVAE OF COOPERIA CURT ICEI AT 27°C 3 5·
i ) i i )
Materials and methods Results
3 . EFFECT OF MOISTURE ON SURVIVAL OF THE EGGS OF COOPERIA CURT ICEI AT D IFFERENT TEHFERATURES
i ) i i )
Materials and me thods Results
4. SURVIVAL OF THE FREE LIVING STAGES OF COOPERIA CURTICEI AT D IFFERENT TEMPERm ATURES
i ) i i )
Mat erials and me thods Results
3 5 ..
3 6 .
3 9 ..
3 9 · 40 .
CHAPTER IV:.,
CHAPTER V .
5, SU:tv'Jlv!ARY O F EXPERIHENTAL FINDINGS 6 . D ISCUSS ION
THE ECOLOGY OF THE FREE LIVING STAGES OF COOPERIA CURTICEI
1 . MATERIALS AND METHODS
2 . TEST OF A TECHNIQUE FOR THE RECOVERY OF INFECTIVE LARVAE OF COOPERIA CURTICEI FROH SMALL SAMPLE UNITS i)
i i)
Descript ion of the techni que a) Separation of infec tive
larvae from herbage b ) Separation of infect ive
larvae fr om t op s oil surface of flower pots
c ) Separation of infective larvae from faecal pellets Test s of the technique
3 . RESULTS
i ) Development of the free living stages
so.
52s
62 .
i i ) Survival of the infec tive larvae 91 . iii ) Vertical migration of the infect -
ive larvae
4 .. SUMMARY OF EXPERIMENTAL FINDINGS 5· DISCUSS ION
PART II . HOST PARAS ITE RELATIONSHIP OF COOPERIA CURTICEI AND THE SHEEP.
REVIEW OF LITERATURE 1 .. EXSHEATHivlENT
2 . DEVELOPMENT OF THE RARAS ITIC STAGES OF COOPERIA CURTICEI, MORPHOLOGY OF
1 1 4 ..
1 1 9 ..
1 22 .
1 32 .
CHAPTER VI .
CHAPTER VII .
THE ADULT NE1'-1ATODE , LOCATION IN THE
Sl'-1ALL INTESTINE AND EGG PRODUCTION 1 3 5"
i )
i i )
Development o f the parasitic stages
Morph ology of the adult stage iii ) Location in the small int e s t ine iv ) Egg produc tion
3 . PATHOGENES IS , PATHOLOGY AND IMNUNE RESPONSE OF COOPERIA INFECT ION i) Sympt oms
i i ) Feed intake , body weight and
1 3 5·
1 3 5 ..
1 36 . 1 37 ..
1 37 . 1 38 .
wool produc t ion 1 39 .
i i i ) Eff e c t s on c onstituents of the
bl ood 141 .
iv ) Gross lesions and histopathology 1 42.
v ) Resistance and immune respons e 1 43 .
EXSHEATHMENT OF THE INFECTIVE LARVAE OF COOPERIA CURT ICEI BOTH IN VITRO AND IN VIVO
1.. MATERIALS AND METHODS 2 . RESULTS
3 . SUHHARY 4 . D ISCUSS ION
EFFECT OF COOPERIA CURTICEI IN SHEEP 1 . EFFECT OF COOPERIA CURT ICEI IN SHEEP
147 ..
1 48 . 1 50 . 1 51 .
6 - 7 MONTHS OLD 1 53•
i ) Materials and me thods a ) Experimental design
b ) Parasitological t echnique c ) Fae c al e gg c ounts
d ) Body weight and wool growth
1 53 . 1 53 . 1 54 ..
1 54 ..
1 54 .
CHAPTER VII I .
i i )
e ) Blo od analysis f ) Necropsy procedure g ) Histopathology h) Immunol ogi cal study
S ource o f antibodies Preparation of antigen Ser ological method Results
1 55·
1 55·
1 56 ..
1 56 . 1 57 . 1 57 . 1 58 . 1 58 ..
a ) Clinic al observations 1 58 . b ) Faecal egg c ount s 1 58 . c ) Body weight and wo ol growth 1 58 . d ) Blo od anal yses 1 63 . e ) Gros s lesions, frequency
d is tribution in small i ntes tine , sex rati o and percentage established in the host
f ) His topatho logy
g ) Immunologi cal resp onse 2 . EFFECT OF COOPERIA CURT ICEI IN SHEEP
1 74 . 1 74 .
2 - 3 MONTHS OLD 1 8 1 .
i ) i i )
Materials and methods Res ults
3. EFFECT OF D IET ON COOPERIA CURTICEI
INFECTION IN SHEEP 9 - 1 0 MONTHS OLD 1 99 . i )
i i )
Materials and wethods Results
4 . SUMMARY OF EXPERIMENTAL FINDINGS 5 • DISCUSS ION
GENERAL DISCUSS ION AND CONCLUS IONS 1 . GENERAL DISCUSS ION
2 . GENERAL SUMNARY AND CONCLUS IONS BIBLIOGRAPHY
APPENDICES
1 99 ..
200 . 209 . 2 1 2 .
2 1 7 ..
222 .
Figures
1.
2.
4.
INDEX TO FIGURES
Egg of Cooperia cur ticei
Days d evelopment t o inf ec tive stage (T90) at c ontr olled t emperatures
Regr e s s ion of d evelopment time ( log d ays) on temp eratur e
Percentage survival of infe c t ive larvae o f C . c urticei in wat er at c ontrolled t emperatures Maximum survival of firs t , s e c ond and third
stage larvae of C . c urtice i in water at controlled t emperature s
Days development t o inf e c t ive stage ( > 90%) and mean maximum air temp erature during period of d evelopment
Regr e s s ion of development time (log days ) on mean maximum air t emperature
8. Larval survival and migration with mete or ological data .
10�
11.
12 ..
Experiment 1. ( March 1968)
Larval survival and mi gration with me t e orologi cal dat a .
Exp eriment 2. (April 1968)
Larval survival and migrati on with met e or ol ogical data .
Expe riment 3 ( May 1968)
Larval survival and migration with m e t e orological dat a .
Exp er iment 7 ( S eptember 1968)
Larval survival and migrat i on with met e or ological data .
Exp eriment 8 ( Oc t ober 1968)
Larval survival and migrat ion with met e or ological data .
Experiment 9 ( November 1968)
iv .
Page No .'
30 ..
30.
48.
90.
90.
104.·
106�
107.
108.
109 ..
Figures Page No . 14. Larval survival and migration with m e t e orologi cal
datao
Experiment 10 ( D e c ember 1968) 1 10.
15. Larval survival and m igration wi th met e or ological d ata .
Exp er iment 11 ( January 1969) 111.
16. Larval survival and migration with mete orological dat a .
Experiment 12 ( February 1969) 112.
17. Arcsin of p ercentage of larvae r e c ov ered from h erbage ( migrat ed ) plotted agains t p erc entage of larvae sur-
\iving , expressed as mid point s 1 18.
18. Gel-d iffusion reactions of s erum from she ep inf e ct e d
f o r 34 d ays against var ious antigens 176.
19. Gel d i f fusion reac t i ons of muc us extract from small i nt est ine of sheep inf e c t ed for 34 days against
various antigens 177.
20. G el diffusion r eact ions of s erum from she ep inf e ct e d
f o r 56 d ays against var ious ant igens 178.
21. Gel diffus ion react ions of mucus extract from small int est ine of sh eep inf e cted for 56 d ays against
var ious ant igens 179.
