UNIVERSITI PUTRA MALAYSIA GASTROINTESTINAL PARASITISM IN GOATS - BIONOMICS OF THE SUPRAPOPULATION AND INCIDENCE IN YOUNG GOATS

UNIVERSITI PUTRA MALAYSIA GASTROINTESTINAL PARASITISM IN GOATS - BIONOMICS OF THE SUPRAPOPULATION AND INCIDENCE IN YOUNG GOATS DAUD AHMAD BIN ISRAF ALI FPV 1991 3

GASTROINTESTINAL PARASITISM IN GOATS - BIONOMICS OF THE SUPRAPOPULATION AND INCIDENCE IN YOUNG GOATS By DAUD AHMAD BIN ISRAF ALI A Thesis Submitted in Fulfilment of the Requirements for the Degree of Master of Science in thefaculty of Veterinary Medicine and Animal Science, Universiti Pertanian Malaysia. November 1991

ACKNOWLEDGEMENT The constant encouragement and comments given by my supervisor, Dr. Rehana Abdullah Sani is sincerely appreciated. A warm thanks to Dr. Ridzwan Abd. Halim who not only helped a lot with the statistical analysis of data but also provided the experimental plots. A special thanks to Dr. Zamri Saad for diagnosing the causes of death. Throughout the course of the work Dr. Ali Raj ion, En. Sharifuddin and the staff of the university's Small Ruminant Unit have been very helpful in providing the goats for the study. En.lslahuddin has worked very hard to maintain the experimental plots. A special thanks to all of you. I would also like to thank Mr.Lee Chu Chong, Bohari Yaacob, Nuzolazuan Ishak and Rosli Malek from the faculty 's parasitology laboratory and Dr. Frank Jackson from Moredun Research Institute, U.K for their kind assistance and comments. My sincere gratitude to the Dean of the faculty, Prof. Latif Ibrahim for providing research facilities and financial support. Last but not least a special thanks to my parents, my wife, Imah and son, Faisal who have been a source of inspiration throughout the course of study. ii

PREFACE This thesis comprises two parts. Part 1 describes the testing and further modification of the Modified Parfitt's Technique (MPT) which is used to recover and enumerate infective larvae from herbage. An assessment of the concentration of infective nematode larvae on pasture herbage gives an indication of the infection to which grazing animals are exposed. The technique was then tested under field conditions to study the survival of infective trichostrongyle larvae. An experiment aimed at determining the rates of development of the various preparasitic stages is also included. Part 2 is a study on the incidence and significance of gastrointestinal parasites in goats from birth to nineteen months old. iii

PERPUST AI<AAN UNIVERSITI PERTAN1AN MALAYSIA TABLE OF CONTENTS Page ACKNOWLEDGMENT PREFACE ii iii LIST OF TABLES iv LIST OF FIGURES ABSTRACT ABSTRAK vi vii x CHAPTER PART 1. BIONOMICS OF THE FREE-LIVING STAGES OF GOAT TRICHOSTRONGYLES I II INTRODUCTION LITERATURE REVIEW Technical aspects in the Recovery of Infective Nematode Larvae from Herbage General Herbage Sampling 1 3 3 3 5 Separation of Larvae from Herbage.......................... 9 Separation of Larvae from Debris........................... 11 Microscopical Examination of Larvae............................. 15 Conditions in Microhabitats 16 Faeces 16 Soil, Mat and Herbage 20

The Effect of Climate upon Development and Survival of Free-living Stages of Trichostrongyles........................... 22 III MATERIALS AND METHODS 31 The Modified Parfitt's Te hnique 31 Spiking of Grasses 33 Infective Larvae 33 Experimental Procedures 34 Validation of the Modified Parfitt's Technique 34 Improvement of the Modified Parfitt's Technique.................. 37 Survival of Larvae on Grass Plots........................... 38 Development of Preparasitic Stages................................. 40 IV RESULTS 41 Validation of the Technique 41 Field Trial 41 Larval Loss Laboratory Trials 41 43 Improvement of the Technique 43 Initial Modifications 43 Further Modifications 47 Survival on Grass Plots 47 Larval Development 49 V DISCUSSION 51 The Modified Parfitt's Technique and its Application............................ 51

Preparasitic Development of Haemonchus contortus............ 59 PART 2. INCIDENCE OF GASTROINTESTINAL PARASITES IN KIDS I II III IV INTRODUCTION LITERATURE REVIEW The Malaysian Goat Industry Effects of Gastrointestinal Parasitism upon Productivity Helminthoses in Goats Coccidioses in Goats Interpretation of Faecal Egg Counts MATERIALS AND METHODS Animals Samples and Laboratory Techniques Data Analysis RESULTS Incidence of Parasites Prevalence of Parasites Liveweight Gain Causes of Death and Worm Counts Relationship between Faecal Egg Counts and Total Worm Counts 62 64 64 66 68 72 77 84 84 85 86 89 89 94 97 99 102 Changes in Worm Populations and Fecundity.................................. 102 Relationship between Faecal Egg/Oocyst Counts and Age............................ 107

v DISCUSSION 109 Gastrointestinal Tract Parasites 109 Significance of Gastrointestinal Parasitism................................. 116 Relationship between Worm and Faecal Egg Counts................................. 119 BIBLIOGRAPHY 126 APPENDIX 139 A Means and Standard Deviations of Variables from all Goats... 139 VITA B C Means and Standard Deviations of Variables from the Goats that Survived throughout the Study Period.............................. 141 Means and Standard Deviations of Variables from the Goats that Died throughout the Study Period.............................. 143 144

LIST OF TABLES Table No. 1 2 3 Assessment of Mean Larval Loss by the MPT using Panicum maximum Spiked with 300 L3.... Validation of the MPT on Various Grasses Spiked with 200 L3.... Validation of the MPT on Various Grasses Spiked with 300 L3..... Page 42 44 45 4 Initial Modifications to the MPT and Recovery Rates using Digi taria setivalva Spiked with 200 L3............ 46 5 Further Modifications to the MPT and Recovery Rates from Panicum maximum, Setaria splendida and Digi taria setivalva Spiked with 200 L3........................... 48 6 7 Recovery Rates of the MPT using Lukewarm water from Panicum maximum, Setaria splendida and Digitaria setivalva Spiked with 200 L3.... Mean Interval for Development of Preparasitic Stages of H.con tortus under Three Different Grass Species.... 48 50 8 Percent Prevalence of Gastrointestinal Parasites in Goats.... 9 Distribution of Causes of Death and Mean Worm Counts 98 101 10 Correlation Analysis between Egg and Worm Counts, Haemonchus and Trichostrongylus Counts, and Egg Counts and TPI.... iv 103

11 Correlation Analysis between Egg/ Oocyst Counts and Age of Animals at Various Age Groups after Logarithmic Transformation of Counts [Log 10 (x+l)].... 108 v

LIST OF FIGURES Figure No. Page 1 2 3 4 5 6 7 8 Incidence of Strongyles among Kids.... Comparison of Strongyle Egg Counts between Kids that Survived and those that Died.... Incidence of Coccidia among Kids.... Comparison of Oocyst Counts between Kids that Survived and those that Died Incidence of Moneizia among Kids.... Comparison of LWG Between Animals that Survived and those that Died.... Comparison of Worm Burdens at Various Age Intervals.... Comparison of Mean Egg Production per Female Worm at Various Age Intervals.... 90 92 93 95 96 100 105 106 vi

Abstract of thesis submitted to the Senate of Universiti Pertanian Malaysia in fulfilment of the requirements for the degree of Master of Science. GASTROINTESTINAL PARASITISM IN GOATS - BIONOMICS OF THE SUPRAPOPULATION AND INCIDENCE IN YOUNG GOATS By DAUD AHMAD ISRAF ALI NOVEMBER 1991 Supervisor Faculty Dr. Rehana Abdullah Sani Veterinary Medicine and Animal Science The Modified Parfitt's Technique (MPT) is used to assess the concentration of infective nematode larvae on pasture which gives an indication of the infection to which grazing animals are exposed. Validation of this technique showed that it was not successful when applied in the field and variation in results occurred among different grass species. Using Panicum maximum (Guinea), Setaria sphacelata var splendida (Setaria) and Digitaria setivalva (Mardi Digit) grasses, modifications were introduced to improve the larval recovery rate. The soaking of herbage in lukewarm water ( 38 o C) instead of tap water increased the recovery rate significantly without variation between the three grasses tested. vii

Using this improved MPT, plot trials were conducted to assess the longevity of third-stage larvae (L3) of goat trichostrongyles under the forementioned grass leys. However the technique failed to recover any L3. Reasons for this were low number of larvae, rainfall, and larval- grass/debris adherence. There is a need for further investigation into this technique. Studies were also conducted to establish the moulting times of the pre-parasitic stages of H.contortus under the three grass leys. Development to Ll and L2 took the same time under the various grasses. Development time to L3, however, was longest under Mardi Digit followed by Setaria and Guinea. The lower temperatures under Guinea aided a more rapid development compared to the other grass species. In order to establish the incidence and significance of gastrointestinal parasitism among kids, faecal samples and liveweights were obtained weekly for seventy-nine weeks from forty-six newly born kids kept under a semi-intensive management. Strongyle infections were detected from two months of age and coccidia from one month. The strongyles were identified Infection as with Haemonchus and Trichos trongylus species. strongyles and coccidia was continuous throughout the study period. Patent infections with Moniezia viii

occurred at eleven weeks of age and continued until thirty eight weeks of age. Comparisons of egg/oocyst output and livewe ight gain (LWG) of animals that survived and those that eventually died showed a significant difference in egg output and LWG but not in oocyst output. The prevalence of parasites was moderate in kids below three months of age and high among those above three months. The possibility of reduced average daily gain (ADG) due to parasites was evident and the combined effects of H.contortus and T.colubriformis significantly increased mortality rate among the animals. Egg and worm counts were positively correlated except for T.colubriformis counts among animals above six months of age. The use of the "total pathogenic index" (TPI) as a diagnostic measure is of valuable significance. Correlation analys is showed suppression of eggs and oocyst production after twelve and six months of age respectively, suggesting acquisition of immunity. ix

Abstrak tesis yang dikemukakan kepada Senat Universiti Pertanian Malaysia bagi memenuhi syarat-syarat ij azah Master Sains. PARASITISME GAS TROUSUS PADA KAH ING - BIONOMIK SUPRAPOPULASI DAN INSIDENS PADA KAHBING MUDA Oleh DAUD AHMAD ISRAF ALI NOVEMBER 1991 Penyelia Fakulti Dr. Rehana Abdullah Sani Kedoktoran Veterinar dan Sains Peternakan Teknik Modified Parfitt's (TMP) didapati tidak berj aya mencapai matlamatnya apabila digunakan di ladang dan terdapat perbezaan dalam keputusan apabila spesies rumput yang berlainan digunakan. Pengubahsuaian ke atas teknik ini dengan menggunakan spesies- spesies rumput Pan icum maximum (Guinea), Setaria sphacelata var splendida (Setaria) dan Digitaria setivalva (Mardi Digit) menunj ukkan peningkatan peratus perolehan yang bermakna tanpa perbezaan di antara ketiga-tiga spesies rumput telah tercapai dengan merendam rumput di dalam air suam (38 C). Ujikaji-ujikaj i plot dengan menggunakan TMP yang diubahsuai, gagal mengesan larva daripada rumput. Di antara x

sebab-sebab kegagalan ini adalah bilangan larva pada rumput yang terlalu sedikit, huj an yang lebat dan kesukaran memisahkan larva daripada rumput. Kaj ian untuk menentukan jangkamasa perkembangan peringkatperingkat pra-parasitik cacing H.contortus di bawah rumputrumput pastura menunj ukkan peringkat Ll dan L2 mengambil masa yang sarna di bawah rumput- rumput tersebut. Perkembangan kepada L3 mengambil masa paling lama di bawah Mardi Digit diikuti dengan Setaria dan Guinea. Suhu persekitaran yang lebih rendah di bawah rumput Guinea membantu mempercepatkan perkembangan. Dalam kaj ian insidens dan kepentingan parasitisma gastrousus pada anak-anak kambing, infeksi-infeks i strongil dikesan sej ak umur dua bulan dan koksidia sej ak satu bulan. Strongil-strongil dikenalpasti sebagai spesies Haemonchus dan Trichostrongylus. Infeksi dengan strongil dan koks idia berlarutan sehingga tamat tempoh uj ikaj i. Infeks i-infeksi paten dengan Honiezia dikesan pada umur sebelas minggu dan berterusan sehingga umur tiga puluh lapan minggu. Perbandingan hasilan telur cacing dan peningkatan berat badan antara haiwan-haiwan yang hidup dengan yang telah mati dalam tempoh uj ikaj i menunjukkan perbezaan yang bermakna. Tidak terdapat perbezaan di antara kedua - dua kumpulan haiwan ini dari segi has ilan oosista koksidia. xi

Prevalens parasit sederhana pada anak- anak karnbing di bawah urnur tiga bulan dan tinggi selepas urnur ini. Kernungkinan berkurangnya purata pertambahan berat harian disebabkan kesan parasit telah ditunjukkan dan kornbinasi kesan-kesan H.contortus dan T.colubriformis rnengakibatkan kadar kernatian yang berrnakna. Kiraan telur pada tinj a dan bilangan cacing pada saluran gastrousus telah rnenunjukkan korelasi positif. Bagairnanapun bagi T.colubriformis, tidak terdapat korelasi di antara keduadua pernbolehubah ini bagi haiwan-haiwan yang berumur lebih daripada enam bulan. Penggunaan "indeks jurnlah patogenik" (IJP) sebagai ukuran diagnosis didapati arnat berkesan. Analisa korelasi juga telah rnenunjukkan pengurangan penghasilan telur cacing dan oosista koksidia dalarn tinj a karnbing-karnbing rnasingrnasing selepas rnencapai umur dua belas dan enarn bulan. lni rnenandakan kehadiran pelalian perolehan. xii

PART 1 BIONOMICS OF THE FREE-LIVING STAGES OF GOAT TRICHOSTRONGYLES

CHAPTER I INTRODUCTION The Malaysian small ruminant industry is suffering losses due to trichostrongylid infections, mainly haemonchosis. One of the main options of controlling trichostrongylosis is grazing management since pastures on which animals graze are considered as a centre for the dispersal and exchange between sheep and parasite populations (Crofton, 1971). For this, the bionomics, that is factors affecting the development and survival of the suprapopulation (free-living population) of trichostrongyles has to be known. There is virtually no study reported on the bionomics of trichostrongyles in the humid tropics. The findings in other climatic regions cannot be applied to the humid tropics. Hence this part of the study was devoted to the bionomics of freeliving stages of trichostrongyles. The first obj ective was to assess a widely acceptable technique, the Modified Parfitt's Technique, used for the recovery of third stage trichostrongyle larvae from herbage. Modifications, where found necessary, were applied to the initial technique. The second obj ective was to apply this technique to field conditions to estimate the survival time of goat trichostrongyle larvae under three different grass 1

leys. The third obj ective was to determine the hatching and 2 moulting times of Haemonchus contortus eggs under these grasses.

CHAPTER II LITERATURE REVIEW Technical Aspects in the Recovery of Infective Nematode Larvae from Herbage General The estimation of larval populations of trichstrongyles on pasture has become a common means of monitoring the risk of infection in Europe (Cabaret et a1., 1986). Various methods for the recovery, differentiation and enumeration of infective larval stages found on herbage have been described (Taylor, 1939 ; Crofton, 1954 ; Donald, 1967; Heath and Major, 1968 ; Bawden, 1969 ; Heath et a1., 1970; Lancaster, 1970; Smeal and Hendy, 1972; Armour, 1982 ; Lee and Ikeme, 1985 ; Gettinby et a1., 1985; Cabaret et al., 1986). According to Gettinby et a1. (1985), there are three currently used methods to assess the concentration of infective larvae of parasitic nematodes on pasture. The first methdod describes a technique by which recovery, enumeration and identification of infective larvae could be achieved by manual sampling of herbage (Taylor, 1939). Another method is the grazing method using either animals free of parasites (Tetley, 1959) or rendered free by the use of anthelmintics (Anderson, 1973; Armour, 1982). These animals, usually referred to as "tracers" are slaughtered soon after completion of grazing and 3

4 their worm burdens analysed. The third method, described by Heath et a1. (1970) uses oesophageal fistulated animals to collect herbage samples. Bawden (1969) was able to obtain a mean recovery of 90.4% with his apparatus. However as stated by Smeal and Hendy (1972), the cost of the sieve and filtering apparatus would prohibit its constant use. Parfitt (1955) reported that when a known number of larvae was placed on grass, his technique yielded an average of 43%. Durie (1959) tested his technique by adding known numbers of larvae to sediment obtained by washing samples of pasture which were free of infective larvae. Recoveries ranged from 61% to 94% with an average of 74%. Gettinby et a1. (1985) observed a three to nine fold increase in recovery rates from oesophageal fistulated animals compared to manual sampling. However Cabaret et a1. (1986) observed that ewes ingested on average less infected herbage than would be predicted from a random sample collected on pasture. It is acceptable that throughout the labourious, time consuming process of separating larvae and subsequent enumeration, there will be a considerable loss of larvae. However these losses can be minimized by modification of the current techniques. In general the techniques employed in the

5 recovery of infective nematode larvae comb ine four major aspects : * Sampling of the herbage * Separating the larvae from the herbage * Separating the larvae from the debris * Microscopical examination of the larvae Herbage Sampling The first and most important aspect in recovering infective nematode larvae is the sampling of the herbage. The method employed should be able to pick up as many infective larvae as possible. Various methods of sampling herbage have been described. Taylor (1939) preferred a double zigzag ( "W" ) fashion and collections made at about one hundred different places by pinching two or three pinches of grass from four places at each spot. Plucking should be as close to the ground as possible without uprooting the grass. Lee and Ikeme (1985) only used a single zigzag and collection made at every fifth step. They preferred cutting the top clusters of herbage in front and on both sides so as to simulate the grazing behaviour of the animals. Smeal and Hendy (1972) preferred to follow Taylor 's method. Lancaster (1970) also preferred Taylor's method but stated that "N " shaped transects are more convenient for a single collector. He stated

6 that there was no apparent difference between results obtained by either method of sampling. According to Crofton (1954), Taylor 's method, in theory, was inefficient due to the low number of traverses. He claimed that only if larvae were numerous and randomly distributed would Taylor 's method be sufficient. However, larvae are neither numerous nor randomly distributed in relation to the total number of pinches. Therefore, it seems obvious that the samples must be either greatly increased in number or size, wh ich in either case increases bulk and makes the recovery more difficult (Crofton, 1954). The various methods for the recovery of infective larvae have been subj ected to various tests in order to validate the technique. However, most of the workers (Donald, 1967 ; Heath and Major, 1968; Bawden, 1969; Lancaster, 1970; Smeal and Hendy, 1972; Lee, 1985) based their results on "spiked" herbage (herbage to which larvae have been added after sampling) and not with natural populations. They did not take into account the effect of grass characteristics upon recovery rates. Bawden (1969) was able to obtain a high recovery rate with his apparatus but again, his results were based on "spiked" herbage and he failed to mention an appropriate herbage sampling method. Therefore, it is expected that his technique

7 would give a low recovery rate if the herbage contained natural populations of larvae. This is due to the fact that there is a closer association between natural larval population and debris particles (Lancaster, 1970; Lee, 1985}. Bryan and Kerr (1988) concluded that manual pasture sampling based on observed grazing behaviour seems the only method which meets all criteria. Lee and Ikeme (1985) also noticed the importance of manual sampling and tried to simulate animal grazing habits. Generally it has been accepted that from the single plant, sheep and cattle eat leaf in preference to stem (Arnold and Dudzinski, 1978) and goats tend to nibble atthe shoots and leaves of growing plants and reject the stems (Devendra and McLeroy, 1982). Since ruminants are selective in their grazing habits, the importance of contamination and availability of infective larvae has been recently highlighted. Gettinby et a1. (1985) claimed that sampling herbage using oesophageal fistulated sheep gave a three to nine fold increase in infective larvae recovery compared to manual collection through a zig-zag route. However, Morley and Donald (1980) stated that the contamination of the pasture with infective larvae may simulate availability after a single shower of rain. According to Gettinby et a1. (1985), manual collection of herbage leads to substantial underestimation. Therefore the