OCCURRENCE AND RISK FACTORS OF ENDOPARASITES AND ASSOCIATED

LESIONS IN DONKEYS IN SELECTED ABATTOIRS IN KENYA

DR. NANCY NDINDA MULWA (BVM, UoN)

A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR

THE DEGREE OF MASTER OF SCIENCE IN VETERINARY PATHOLOGY AND

DIAGNOSTICS OF THE UNIVERSITY OF NAIROBI.

DEPARTMENT OF VETERINARY PATHOLOGY, MICROBIOLOGY AND

PARASITOLOGY,

FACULTY OF VETERINARY MEDICINE,

UNIVERSITY OF NAIROBI

2019
 DECLARATION

This thesis is my original work and has not been presented for a degree in any other

University

Dr. Nancy Ndinda Mulwa (BVM, UoN)

Signature: …………………… Date: ……………………….

This thesis has been submitted with our approval as supervisors:

Signature: …………………. Date: …………………………

Prof. Samuel Maina Githigia, BVM, MSc., PhD

Department of Veterinary Pathology, Microbiology and Parasitology

University of Nairobi

Signature: …………………. Date: ………………………….

Dr. Davis Njuguna Karanja, BVM, MSc., PhD

Department of Veterinary Pathology, Microbiology and Parasitology

University of Nairobi

Signature: ………………. Date: …………………………

Dr. Cecilia Kathure Mbae, BSc, MSc., PhD

Centre of Microbiology Research

Kenya Medical Research Institute

ii
 DEDICATION

I dedicate this work to my family members for their unending love and support throughout

my study period.

iii
 ACKNOWLEDGEMENTS

I offer my tribute and gratitude to God for bestowing me health and endurance to complete

this major task. I also thank the University of Nairobi for the award of a scholarship. I have

grown immensely in terms of skills and expertise in this area of study. My heartfelt gratitude

to Eberhard Zeyhle and Erastus Mulinge (KEMRI) for the support offered during the

proposal development and sample collection in the field. Special thanks go to my supervisors

Prof. Samuel Maina Githigia, Dr. Davis Njuguna Karanja and Dr. Cecilia Kathure Mbae for

their unending support, motivation and guidance during my research project.

I cannot forget to mention Mr. R.O. Otieno, J. Mugendi and Ms. Edith Keya of Parasitology

Laboratory, Mr. John Mukiri, Mr. David Muriithi and Ms. Grace Mwangi of Histopathology

Laboratory and Ms. Virginia Mumbi of Biochemistry Laboratory. Indeed the entire staff

department of Veterinary Pathology, Microbiology and Parasitology fraternity for

passionately guiding me throughout the various aspects of my research project and an

opportunity to use the laboratory and other facilities. I must say that my research skills have

been enhanced. Your commitment and expertise is highly appreciated.

Special gratitude goes to Dr.Gerald Muchemi for his statistical advice on the various aspects

of data handling and analysis.

Finally, I give plenty of thanks to my beloved family members who have stood by me

through thick and thin to make this course a success.

iv
 Table of Contents

DECLARATION .......................................................................................................................ii

DEDICATION ......................................................................................................................... iii

ACKNOWLEDGEMENTS ...................................................................................................... iv

Table of Contents ....................................................................................................................... v

LIST OF TABLES ................................................................................................................. viii

LIST OF FIGURES .................................................................................................................. ix

LIST OF APPENDICES ........................................................................................................... xi

LIST OF ABBREVIATIONS ..................................................................................................xii

ABSTRACT ........................................................................................................................... xiii

1.0 CHAPTER ONE: INTRODUCTION .................................................................................. 1

1.1 OBJECTIVES OF THE STUDY ..................................................................................... 3

1.1.1 General Objective ...................................................................................................... 3

1.1.2 Specific Objectives .................................................................................................... 3

1.2 JUSTIFICATION ............................................................................................................. 3

2.0 CHAPTER TWO: LITERATURE REVIEW ...................................................................... 5

2.1: OVERVIEW OF DONKEY ENDOPARASITES AND ASSOCIATED LESIONS ..... 5

2.1.1: Occurrence of Various Species of Donkey Endoparasites ....................................... 5

2.1.2: Lesions Associated with Nematodes ........................................................................ 8

2.1.3: Lesions Associated with Gasterophilus species ..................................................... 10

2.1.4: Lesions Associated with Cestodes.......................................................................... 11

2.1.5: Lesions Associated with Trematodes ..................................................................... 11

2.2: Animal Risk Factors for Occurrence of Gastrointestinal Parasites .............................. 12

2.3: Occurrence of Donkey Hemoparasites and Associated Lesions ................................... 12

3.0 CHAPTER THREE: MATERIALS AND METHODS .................................................... 16

3.1: Study Areas ................................................................................................................... 16

v
 3.2: Study design .................................................................................................................. 17

3.3: Sample Size Determination ........................................................................................... 18

3.4: Determination of Prevalence and Intensity of Gastrointestinal Parasites and Risk

Factors for Helminth Infestation in Donkeys ....................................................................... 19

3.4.1: Prevalence of Gastrointestinal Parasites ................................................................. 19

3.4.2: Intensity of Gastrointestinal Helminths Infection .................................................. 19

3.4.3: Risk Factors for Helminth Infestation in Donkeys ................................................. 20

3.5: Determination of Occurrence of Hemoparasites in Donkeys ....................................... 20

3.6 :Characterization of Lesions Associated with Gastrointestinal Parasites in Donkeys in

Selected Abattoirs in Kenya ................................................................................................. 21

3.7: Data Analysis ................................................................................................................ 21

4.0 CHAPTER FOUR: RESULTS .......................................................................................... 23

4.1: GENERAL INFORMATION ....................................................................................... 23

4.2: Prevalence and Intensity of Gastrointestinal Parasites and Risk factors for Helminth
Infestation in Donkeys in Three Abattoirs in Kenya............................................................ 24

4.2.1: Intensity: Fecal Egg Count ..................................................................................... 24

4.2.2: Animal Risk Factors and Helminth Infestation ..................................................... 25

4.2.3 Prevalence of the Helminth Eggs in the Slaughtered Donkeys ............................... 26

4.2.4: Prevalence of the Cysts .......................................................................................... 29

4.3: Occurrence of the Various helminths in Donkeys and Associated Lesions.................. 29

4.3.1 Stomach ................................................................................................................... 29

4.3.2: Intestines ................................................................................................................. 33

4.3.3: Liver ....................................................................................................................... 42

4.4: Occurrence of Hemoparasites in the slaughtered donkeys ........................................... 45

4.4.1: Buffy Coat Smears ................................................................................................. 46

4.4.2: Thin Blood Smears..................................................................................................... 48

5.0: CHAPTER FIVE: DISCUSSION, CONCLUSIONS AND RECOMMENDATIONS ... 53

5.1: DISCUSSION ............................................................................................................... 53

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 5.1.1: Proportion and Intensity of Endoparasites Infestation in Donkeys ............................ 53

5.1.2: Risk Factors to Occurrence of Helminths and Strongyle Egg Count ........................ 55

5.1.3: Occurrence of Hemoparasites .................................................................................... 56

5.1.4: Characterization of Lesions Associated with Helminths in donkeys ......................... 57

5.1.4.1: Stomach ............................................................................................................... 57

5.1.4.2: Intestines .............................................................................................................. 58

5.1.4.3: Liver .................................................................................................................... 60

5.2: CONCLUSION AND RECOMMENDATIONS .......................................................... 61

5.2.1 CONCLUSION ....................................................................................................... 61

5.2.2 RECOMMENDATIONS......................................................................................... 61

6.0: REFERENCES ................................................................................................................. 62

7.0: APPENDICES .................................................................................................................. 73

vii
 LIST OF TABLES

Table 1: Proportion of Animals Examined per Slaughterhouse .............................................. 23

Table 2: Mean Egg Count between the Gravid and Non-gravid Female Donkeys ................. 24

Table 3: Table showing the intensity of the Strongyle egg infection rates for slaughtered
donkeys .................................................................................................................................... 25

Table 4: Showing infestation intensity ranges and means ±standard error of the mean of
helminth eggs, cysts and oocysts recovered from the fecal samples ....................................... 28

Table 5: Table showing the number of positive cases for thin blood smears and the buffy coat
smears from slaughtered donkeys from the three slaughter houses ......................................... 45

Table 6:Occurrence of Hemoparasites as Examined on the Thin Blood Smears from

slaughtered donkeys ................................................................................................................. 49

Table 7: Cellular Morphological Alterations due to Hemoparasites on slaughtered donkey

blood smears in Kenya ............................................................................................................. 52

viii
 LIST OF FIGURES

Figure 1: Map of Kenya showing the three Sites of Study; Turkana (A), Baringo (B) and Nakuru (C)
County (Source: d-maps.com accessed at https://d-maps.com/pays.php?num_pay=30&lang=en)...... 17

Figure 2: Prevalence of the Helminth Eggs in Fecal Samples of Slaughtered Donkeys ...................... 26

Figure 3: Photomicrograph showing a typical strongyle type egg(S), thin shelled and ovoid in shape
from donkey (ID: 22 KNB)................................................................................................................... 27

Figure 4: Photomicrograph showing the spherical brownish egg of Parascaris equorum (PE) from
donkey (ID: 51 KNB) and an air bubble (A). ....................................................................................... 27

Figure 5: Photomicrograph Showing the Egg of Oxyuris equi from donkey (ID: 75 MGT) the mucoid
plug (MP) and the flattened side (FS) are illustrated. ........................................................................... 28

Figure 6: Proportion of the various species of the Gastrophilus larvae in the stomach of slaughtered
donkeys ................................................................................................................................................. 29

Figure 7: An ulcer (shown by the black arrow) on the non-glandular area of the donkey stomach with
some areas of the mucosa tinted yellow from donkey (ID: 20 KNB); several Gasterophilus species are
present (white block arrow). ................................................................................................................. 30

Figure 8: Photomicrograph of a stomach section of a slaughtered donkey (20 KNB) illustrating the
disruption of lamina propria (white block arrow) and discontinuity of the stratified squamous
epithelium (X10; H&E) ........................................................................................................................ 31

Figure 9: Photomicrograph of the stomach of a slaughtered donkey (20 KNB) showing engorged
blood vessels at the submucosa as illustrated by the white block arrows (X40; H&E) ........................ 32

Figure 10: Photomicrograph of the stomach mucosa of a slaughtered donkey (ID: 20KNB) showing
vacuolar degeneration (illustrated by the white arrows) in the submucosa of the stomach due to
Gasterophilosis (X40; H&E)................................................................................................................. 33

Figure 11 A and B: A photomicrograph of Anaplocephala perfoliata recovered from the ileocecal

junction of slaughtered donkey (ID: 10 MGT), illustrating the lappets (L) and the scolex (SC) which is
distinct and smaller than the body. On the right side is the same cestode after staining with acetoalum
carmine stain illustrating the scolex (SC), lappets (LP) and the proglottids (P). The white block arrow
shows an artifact (air bubble). ............................................................................................................... 34

Figure 12: Prevalence of the various helminth species in the intestines of slaughtered donkeys in
Kenya .................................................................................................................................................... 35

Figure 13: A picture of the cecal wall from slaughtered donkey (ID: 13MGT) showing focal nodular
lesions, about 2-5 mm (white block arrow) and a reddish nematode attached onto the mucosa;
Strongylus vulgaris (thin black arrow).................................................................................................. 37

Figure 14: Photomicrograph of the ceca of donkey (13 MGT) showing four regular structures
identified as nematode larvae suspected to be larval stages of Strongylus vulgaris enclosed in a fibrous

ix
capsule(White block arrow) A portion of the cecal mucosa has been disrupted also(Black arrow) (X4).
.............................................................................................................................................................. 37

Figure 15: Photomicrograph of slaughtered donkey (ID: 13 MGT) illustrating the fibrous capsule zone
(Black arrow) that’s encapsulating the nematode larva (white block arrows). The fibrous connective
tissue has mixed inflammatory infiltrate (black arrow) (X10; H&E). .................................................. 38

Figure 16: Picture showing attached nematodes on the cecal mucosa (black arrow), Strongylus
vulgaris (SV). The mucosa is edematous, hyperemic and eroded (EHM) (ID: 3 MGT) ............... 39

Figure 17: photomicrograph of donkey (ID: 3MGT) illustrating the larvae encapsulated within a
lumen (L) with red blood cells adjacent (RBC) and a thin fibrous capsule layer (FC). The mucosa can
be seen with a few glands (M), the blood vessels are also engorged with blood (EBV) (X4; H&E) ... 39

Figure 18: Photomicrograph of donkey (ID: 3MGT) illustrating the mixed inflammatory infiltrate as
shown by the arrow and the fibrous capsule (FC) (X40; H&E) ........................................................... 40

Figure 19: Photomicrograph of donkey ceca showing the submucosa of a donkey (9 MGT) illustrating
the mixed inflammatory infiltrate with the eosinophils predominating (black arrow) (X40; H&E) .... 41

Figure 20: Liver of a slaughtered donkey (ID: 60 MGT) showing the cyst-like swelling (white arrow)
protruding above the capsule. ............................................................................................................... 43

Figure 21: Photomicrograph of the liver of a slaughtered donkey (60 MGT) showing the well
demarcated zone (ZC) with the central vein engorged with blood (CV) , the enclosed material(CD) is
where the retrieved helminth (Strongylus edentatus) was residing and is composed of necrotic
material and no hepatic tissue can be appreciated(X10; H&E) ............................................................ 43

Figure 22: Picture of a liver of a slaughtered donkey (ID: 18 LDW) showing multifocal greyish areas
on the liver surface. ............................................................................................................................... 44

Figure 23: Photomicrograph of a donkey liver (ID: 18 LDW) showing a focal area with necrotic
material with interspersed neutrophils (N) and an accompanying fibrous capsule (FC)-Hepatic
abscess. The central vein is also illustrated (CV) (X10; H&E). ........................................................... 44

Figure 24:Occurrence of Hemoparasites as Examined on the Buffy coat smears of slaughtered

donkeys ................................................................................................................................................. 46

Figure 25: Photomicrograph of a buffy coat smear of animal (ID: 2LDW) showing trypanosome spp
(white block arrows) in a donkey, (X100;Giemsa Stain) ...................................................................... 47

Figure 26: Photomicrograph of a buffy coat smear from donkey (ID: 4LDW) showing a lymphocyte
with purple staining inclusions at the margin (Anaplasma spp-Black arrow). (X100;Giemsa Stain) .. 48

Figure 27: Photomicrograph of a buffy coat smear of a slaughtered donkey(ID: 80KNB) showing
Setaria equina microfilariae(MF), a band neutrophil(N) (X100;Giemsa Stain) ................................... 48

Figure 28: Photomicrograph of a thin blood smear from donkey (ID: 4LDW) showing red blood cells
with two pyriform shaped inclusions black arrow( Babesia caballi). Note the red blood cells that are
polychromatophilic with varying shapes and sizes.(X 100;Giemsa Stain) ........................................... 49

x
Figure 29: Photomicrograph of a buffy coat smear from donkey (ID: 78 KNB) showing a red
blood cell with Babesia caballi (X100 Giemsa stain) ........................................................................ 50

Figure 30: Photomicrograph of a buffy coat smear from a donkey (ID: 78KNB) trypanosome spp can
be seen (White arrow head) (X100; Giemsa Stain). ............................................................................. 51

Figure 31: Photomicrograph if a thin blood smear from donkey( ID: 69 MGT) illustrating a
lymphocyte with vacuolated cytoplasm(white arrow head) and the variation in shape and size of the
red blood cells can also be appreciated(X100; Giemsa Stain). ............................................................. 52

LIST OF APPENDICES

Appendix 1: Field Data Collection Sheet ............................................................................................. 73

Appendix 2: Body Condition Scoring System for Donkeys ................................................................. 74

Appendix 3: Analyzed Results .............................................................................................................. 75

xi
 LIST OF ABBREVIATIONS

< Less than

> Greater than

Cm centimeters

EDTA Ethylenediaminetetraacetic acid

EPG Eggs per Gram

FEC Fecal Egg Count

g Grams

H&E- Hematoxylin and Eosin

ID Identity

KEMRI Kenya Medical Research Institute

KNB Kinamba

LDW Lodwar

MGT Mogotio

Mm millimeters

MAFF Ministry of Agriculture Fisheries and Food

MALF Ministry of Agriculture Livestock and Fisheries

mls millilitres

OIE World Organization for Animal Health

PCR Polymerase Chain Reaction

r.p.m Revolutions per Minute

SPSS Statistical Package for the Social Sciences

Spp Species

UoN University of Nairobi

xii
 ABSTRACT

The donkey is a significant animal species in the arid and semi-arid areas, it serves as an

important means of transport and provision of draught power. Gastrointestinal parasites and

hemoparasites have been reported to cause the most prominent diseases in donkey as

compared to other diseases. Helminth infestation has been reported to cause mortality in

Ethiopia besides retarding growth, decreasing work output in addition to distress and pain.

Recent studies to show the diverse nature of gastrointestinal parasites and their pathological

effects have not been done in Kenya. A survey was done in order to determine the diversity

and intensity of donkey endoparasites and to characterize associated lesions in selected

abattoirs in Kenya. Three abattoirs (Kinamba, Mogotio and Lodwar) were visited between

July-September, 2017. A total of 282 donkeys presented for slaughter were systematically

sampled for gastrointestinal parasites and associated lesions. Blood, fecal, parasite and

gastrointestinal organ samples were collected and screened for hemoparasites, quantification

of the fecal eggs, identification and characterization of pathological lesions respectively. The

diverse nature of gastrointestinal parasites and hemoparasites were determined by virtue of

their morphological characteristics whereas the intensity was assessed by quantifying the

fecal egg count using the McMaster technique. The pathological lesions were characterized

grossly and microscopically.

Adults helminths parasites identified were; Anaplocephala magna 2.5%, Anaplocephala

perfoliata 10.3%, Cylicocyclus auriculatus 2.1%, Cylicocyclus leptostomus 0.4%

Cyathostome species 2.1%, Parascaris equorum 20.2% , Strongylus edentatus 12.1%,

Strongylus equinus 0.4%, Strongylus vulgaris 52.8%, Setaria equina 3.5% and

Triodontophorus serratus 0.4%. Fifty one point eight percent were positive for Gasterophilus

species larvae with an occurrence of 38.3%, 5.7% and 7.8% for G. intestinalis, G. nasalis and

G.pecorum respectively. Forty four point seven percent of the donkeys were positive for

xiii
strongyle eggs; Parascaris equorum at 5.3%, Oxyuris equi at 1.1%, Triodontophorus

tenuicolis and Habronema species each at 0.7% and cestodes eggs at 0.4%. For intensity

determination, 55.3% had no eggs present, 39% had a low infection (up to 500 EPG), 5% had

a medium infection (501-1000EPG) with 0.7% having a high infection (>1000 EPG). For the

hemoparasites, Anaplasma phagocytophilum occurred at 2.5%, trypanosomes at 5.6%,

Babesia caballi at 2.5% and microfilaria at 0.60% of the examined donkeys. Pathological

changes observed in the stomach mucosa included diffuse areas of erosion after dislodging

the various Gasterophilus species. Microscopic observations made included disruption of the

keratinized layer of the non-glandular portion of the stomach, yellowish discoloration of the

epithelium, loss of keratin and thickening in some areas. In the liver, hepatomegally was

observed in 6% of the donkeys and a cyst-like focal swelling measuring 2-3 cm in diameter

protruded above the capsule in 3% of the donkeys. The other 3% had pinpoint necrotic foci

on the hepatic parenchyma. Microscopically, there was a well demarcated zone comprising

fibrous connective tissue, hemosiderosis and cellular infiltration. In the intestines, focal

nodules were present in 3% of the donkeys. Microscopic examination revealed disruption of

the mucosal lining and nematodes larvae encapsulated in the sub mucosa. Intestinal

ulcerations were observed in 2.5% of the donkeys and were mainly characterized by

circulatory disturbances and cellular infiltrations.

The survey revealed that up to 86% of the donkeys in Kenya are infested by a variety of

gastrointestinal helminths and 11.3% by hemoparasites. Gastrointestinal parasites may cause

severe pathological lesions in the gastrointestinal tract and these interfere with digestion and

assimilation of nutrients thereby leading to poor work performance of the donkeys. There is

need to control these parasites in donkeys in Kenya.

xiv
 1.0 CHAPTER ONE: INTRODUCTION

Donkeys (Equus asinus) are amongst the first tamed equines that have been used as draft

animals for a long time (Saul et al., 1997). They are frequently depicted as the

underprivileged man’s horse, a state best revealed in the animal-energy agriculture in many

of third world countries (Getachew et al., 2012). The global donkey population is at

approximately 44 million (Starkey and Starkey, 2000). In Africa, the population of donkeys is

estimated to be 13 million (Starkey and Starkey, 2001). Kenya has approximately 1,832,519

donkeys with majority of the donkeys in arid and semiarid areas (Kenya National Bureau of

Statistics, 2010). Over half of this population is being used for works in transport and tillage

operations (Kenya National Bureau of Statistics, 2010). The donkey has been known to play

an important function in packing, riding, carting and ploughing. The main reason is due to

cheapness and availability of donkeys. They are also an alternative transport means in places

where the road network is inadequately developed (Pearson et al., 1999 and Negasa et al.,

2017). Donkey power is an environmentally friendly means of transport and vital in areas of

inadequate roads. Donkeys can easily pull more load than it can carry on condition that the

harness is appropriate (Saul et al., 1997). Equines, mainly the donkeys have been entirely

neglected regardless of their important role in both rural and urban societies (Etana et al.,

2011)

Donkey meat is considered a delicacy in some parts of North West Kenya and Southern

Ethiopia and its milk is also believed to be medicinal and has been used to treat whooping

cough (Fred and Pascal, 2006). The government of Kenya recognized donkeys as a food

animal in Kenya in 2012 (The Meat Control Act, 2012). A study done by Mugachia and

Muthusi (2015), shows that donkey owners wish to use their donkeys for work other than for

sales and that donkey owners do not uphold the consumption of donkey meat.

1
Currently, there are four donkey export slaughter houses in Kenya; Goldox limited, in

Mogotio, Baringo County; Star-Brilliant Export slaughter House in Naivasha, Nakuru

County, Silzha Limited in Lodwar, Turkana County and Fuhai Machakos Trading Company

Limited. However, this venture has faced lots of resistance from local communities who do

not accept or uphold consumption of donkey meat and also due to environmental degradation.

In addition to this there has been a lot of donkey theft, illegal sale of the skin and meat. Due

to lack of proper breeding practices, the population has been alarmingly declining. Recently

there has been a report by the animal welfare lobbies that the donkeys in Kenya could be

extinct in four years unless measures are put in place to guarantee the welfare of the animals.

The trade in donkey meat and skin should be halted until measures are put in place to protect

the species (Okech and Nyoike, 2019). The Chinese believe that donkey skin derived

medicine called ejiao treats anemia, delays aging, increases libido, reverses infertility,

prevents miscarriage and menstrual irregularity and treats side effects of chemotherapy

(Daily Nation, 25th July 2019)

Gastrointestinal parasites and hemoparasites have been reported to be most prominent

diseases in the donkey as compared to other diseases (Ahmed et al., 2008). Saul et al. (1997)

reported helminth infestation as the most common cause of death besides retarding growth,

decreasing work output in addition to distress and pain (Svendsen, 1997). More recently, a

study in Ethiopia reported a high prevalence of gastrointestinal helminths interfering with

donkey health and welfare (Mohamee et al., 2017). Similar studies (Lewa et al., 2000 and

Karanja et al., 1994) have shown gastrointestinal parasites as prevalent in donkeys. However,

there are no recent studies in donkeys in Kenya to demonstrate that parasites are still a threat

to donkey welfare as well as to public health. Additionally, there is need to document the

impact of these parasites and pathological lesions caused by gastrointestinal parasites in

donkeys.

2
This research study hence aims to determine the diversity, intensity and the lesions associated

with endoparasitism. This will help in planning and implementation of parasite control

measures in donkeys and contribute to the improvement of donkey health and productivity in

Kenya. Given that the donkey is now considered food animal, information on these parasites

and subsequent control strategies will ensure farmers earn a better income for improved

livelihoods.

1.1 OBJECTIVES OF THE STUDY

1.1.1 General Objective

Determine the occurrence and risk factors of gastrointestinal infestation and haemoparasitic

infection and associated lesions in donkeys in selected abattoirs in Kenya

1.1.2 Specific Objectives

1. To determine the prevalence, intensity and risk factors for gastrointestinal infestation in

donkeys in three abattoirs in Kenya

2. To determine the occurrence of hemoparasites in donkeys in three abattoirs in Kenya

3. To characterize lesions associated with gastrointestinal parasites in donkeys in three

abattoirs in Kenya

1.2 JUSTIFICATION

Agriculture accounts for 27% of Gross Domestic Product and employs about 75% of the

Kenyan population, majority of these are small holder farmers with limited finances and

access to farm machinery (MALF, 2015). Consequently, agriculture is viewed as

unrewarding hard job and youth are not ready for it. In addition, there is a yawning gap in

food production to feed the rapidly growing human population using agricultural technology.

There is need to explore draught power from donkeys and other farm animals to increase

3
efficiency and productivity. In Kenya's vision 2030, and Jubilee's government 2018-2022

strategic plan, food security is a priority agenda. The government is undertaking bold

measures aimed at increasing acreage under cultivation in order to enhance grain supplies

from arid and semi-arid lands (ASAL) through irrigation. However, there is need to match

these efforts with mechanization

Donkeys are important working animals in Kenya especially the arid and semi-arid areas

where people live below the poverty line. The health care of these animals is neglected as

well as a vast number of the owners live in the remote areas. When in good health and well

harnessed, the donkey can till possibly 100 times more land than the hand-held hoe.

However, donkey power is hampered by myriad problems including malnutrition, infectious

diseases and more recently booming donkey trade for slaughter that is decimating population

at alarming rate. Many ignorant people believe donkeys are disease free, but incidentally,

they suffer from clinical and subclinical diseases. A survey of the parasites affecting the

donkeys has been done previously but not extensively as the animals involved were few. This

study will also add to current information on parasites present in Kenyan donkeys.

Knowledge of these parasites is important as control measures can be instituted to improve

the general productivity for better use of these animals. So as to come up with effective

treatment and control programs, it is crucial to have data on the occurrence of the parasites as

well as the lesions they cause. Control measures include regular deworming of the animals

and proper manure management.

4
 2.0 CHAPTER TWO: LITERATURE REVIEW

2.1: OVERVIEW OF DONKEY ENDOPARASITES AND ASSOCIATED LESIONS

2.1.1: Occurrence of Various Species of Donkey Endoparasites

A parasite can be defined as a smaller organism that lives on or in a host and at the expense

of the host. Endoparasites on the other hand can be defined as parasites within the body of the

host (Bowman, 2014).The major categories include Gasterophilus species instars,

Habronema, Draschia, Trichostrongylus, Parascaris equorum, Strongyloides westeri,

Anaplocephala species, Oxyuris equi, Onchocerca cervicalis, Setaria equina, Thelazia

lachrimalis, Strongylus species and small strongyles (Powell and Russell, 2012).

Donkeys are described as sturdy though they are vulnerable to a number of disease

conditions. Among these, parasitic infections are major cause of illness in donkeys (Negasa et

al., 2017). The deficiencies of appropriate management and by virtue of their wandering

behavior, the donkeys are exposed to a wide range of parasitic infections especially

gastrointestinal nematodes because they frequently come into contact with contaminated

pasture (Raman et al., 2014). Parasites are a major problem of donkeys amid other problems

such as wounds, other infectious and noninfectious diseases (Abebew et al., 2011).

In a research carried out in Ethiopia by Ahmed et al (2011), the percentage of helminth

infection in donkeys was 98.45%. Twenty two (22) helminth species including Fasciola

hepatica, Anaplocephala perfoliata, Parascaris equorum, large strongylid, small strongylids,

Habronema muscae, Habronema megastoma, Habronema microstoma, Setaria equina and

Oxyuris equi were identified. Hydatid cysts were also encountered. Fecal egg examination

showed 99% strongyle spp, 80% Fasciola spp, 51% Parascaris spp, 30% Gastrodiscus spp,

11% Strongyloides westeri, 8% Anaplocephala perfoliata cestodes and 2% Oxyuris equi

5
infection prevalence. More than 55% of donkeys had higher than 1000 eggs per gram of

faeces (EPG) (Ahmed et al., 2011).

In Ethiopia, forty two diverse species of parasites that include 33 nematodes, 3 trematodes, 3

cestodes and 3 arthropod larvae were identified from donkeys at postmortem. This study

showed that working donkeys in Ethiopia harbor a variety of helminths and arthropod larvae

which represent most of the significant pathogenic parasites found in equines globally

(Getachew et al., 2010).

Thirty adult donkeys necropsied from Burkina Faso indicated that strongylids were the most

common species with 100% prevalence linked to Strongylus vulgaris. Four species of

Strongylus, two of Triodontophorus and six of the Cyathostominae were also identified. All

donkeys were also infected with Habronematid nematodes with the oxyurid and ascaridid

nematodes being less. Coprological exam of 131 samples from donkeys indicated an egg

count of 100-9,200 for strongylid eggs (Vercruysse et al., 1986). In a study carried out by

Ibrahim et al (2011) in Southern Ethiopia the prevalence of helminth eggs and helminth

species was 96.9%. The prevalence of gastrointestinal parasite in different age group

indicated that there was a significant difference (p<0.05) between the young and the adult

(Wako et al., 2016). In a current research done in Ethiopia from December 2015 to May

2016, the gastrointestinal helminths recorded by Abdulahi et al. (2017) were strongyles (79.7

%), Parascaris equorum (44.8%), Oxyuris equi (37.5%), Fasciola spp (1.6%) and 46.9%

mixed infections. Death in equines has been commonly reported due to strongyles,

tapeworms, ascarids, trypanosomes and Babesia species (Soulsby, 1982)

In six donkeys examined at necropsy in Kenya, the variety of species of internal parasites

identified included Dictyocaulus arnfieldi, Gastrophilus intestinalis, Strongylus vulgaris and

Strongylus edentatus (Lewa et al., 2001). In another study done in Kenya by Karanja et al.

(1994), the common parasites found in eight donkeys included Gasterophilus nasalis,

6
Cyathostomum tetracanthum, Habronema muscae, Strongylus spp, Setaria equina and

Oxyuris equi. The fecal egg count indicated a low to moderate strongyle infestation (Karanja

et al., 1994). A reduction in worm burden has also been shown to result to a 14%

improvement of body condition (Svendsen, 1986).

Parasites affecting the equine stomach include Gasterophilus spp. larvae and gastric

nematodes. Four species of nematodes identified in the stomach include Habronema muscae,

Habronema majus, Draschia megastoma and Trichostrongylus axei (Jacobs, 1986). A

retrospective study done to identify causes of rectal prolapse in donkeys showed that majority

of rectal prolapse cases were linked to Gasterophilus nasalis. Other causes included

overworking the donkeys and diarrhea (Getachew et al., 2012).

Genus echinococcus has two important species of veterinary significance and these include

Echinococcus granulosus and Echinococcus multilocularis. Echinococcus granulosus

equinus is the sub species whose larval stages (hydatid cysts) are found in the equines. A case

of hydatid cyst (E.granulosus) in the liver of a horse was demonstrated with the horse

displaying no clinical signs despite the presence of 20-30 cysts (Bowman, 2014). Evidence of

hydatidosis was established at 17.2% of 122 donkeys necropsied in Central Jordan (Abo-

Shehada., 1988).

Equine has been reported as a less common host for Fasciola and the rate of infection is

relatively low as compared to the ruminants (Jones et al., 1977). Necropsy of 65 donkeys in

Egypt revealed that only two donkeys were infested with Fasciola hepatica in their bile ducts

indicating a low occurrence (Ahmed et al., 2011).

Microfilaria was diagnosed in 20 blood samples collected from healthy horses with a

prevalence of 30.76% (Suleiman et al., 2012). Setaria equina could affect the donkey eye

7
with severe lesions such as continuous lacrimation and ulcerative dermatitis (Suleiman et al.,

2012).

Sharma and Pachauri (1981) showed that the filarial worms meet their nutritional requirement

from the host tissues and fluids and in situ transfer of nutrients from the host tissues to the

body of microfilariae and this causes various deficiency symptoms including anemia

depending on the worm load of the host. In a study undertaken in Hungary, 9.2% of the

donkeys were infected with Setaria equina (Hornok et al., 2007).

2.1.2: Lesions Associated with Nematodes

Cyathostomes result in an inflammatory enteropathy affecting the caecum and colon (Love et

al., 1999). When the cyathostomes move to the surface from the bowel, they cause rupture of

the muscularis mucosae, local eosinophilia, edema and infiltration by neutrophils and

macrophages (Maxie et al., 2016).

Hepatic macroscopic changes associated with experimental infection with Parascaris

equorum infection included focal hemorrhages and tiny, white diffuse or nodular lesions.

Microscopically, lesions were seen mainly around the portal triads and were characterized by

cellular infiltration of eosinophils and lymphocytes as well as fibrosis (Brown and Clayton,

1979).

Trichostrongylus axei has been associated with chronic catarrhal gastritis, which may lead to

weight loss. The lesions include nodular areas of thickened mucosa bordered by a zone of

congestion and covered with an inconsistent amount of mucus (MSD Veterinary Manual,

2016). The third stage larvae of Trichostrongylus axei enters the tunnels in the epithelium of

the gastric glands in the fundic and pyloric regions leading to mucous metaplasia and

hyperplasia of the glands. Cellular infiltration by eosinophils and lymphocytes will be seen in

the lamina propria (Maxie et al., 2016)

8
In the livers of the donkeys, traumatic hepatitis, bile duct thickening and abscess formation

were seen and were linked to Strongylus species larvae. Sections of parasitic larvae were

observed to be connected with hemorrhage and cellular degeneration. The liver tissue was

also highly infiltrated with eosinophils (Lewa et al., 2000).

Both the large and small strongyles have been shown to cause enteritis with the mucosal

surface having parasitic nodules. The intestinal mucosa in the majority of the donkeys was

highly folded and covered with mucoid material. Microscopically, the mucosa and the sub

mucosa had cellular infiltrations (Lewa et al., 2000). Hemomelasma malei are subserosal

hemmorhagic lesions associated with the migration of strongyle larvae (Maxie et al., 2016).

Strongylus vulgaris has been linked to endo-arteritis and thrombi formation. Strongylus

equinus has been associated with formation of cecal and colonic nodules as they penetrate the

mucosa and hemmorhagic tracts in the liver and pancreas (Soulsby, 1982). The nodules are

surrounded by debri, polymorphonuclear cells and macrophages (Maxie et al., 2016).

Strongylus edentatus on the other hand has been associated with hepatic hemorrhagic nodules

in the liver associated with the 4th stage larvae 3-5 months post infection (Soulsby; 1982).

Parenchymal scars and tags of fibrous tissue on the liver capsule have also been associated

with Strongylus edentatus (Maxie et al., 2016). Strongylus equinus has been associated with

hemorrhagic subserosal nodules in the caecum and colon (Maxie et al., 2016)

Invasion by Habronema species triggers the secretion of great quantities of thick and

tenacious mucus in the glandular portion of the stomach adjacent to Margo plicatus, where

adult worms are implanted (Jacobs, 1986). Gastric habronemiasis is accompanied by

catarrhal gastritis, diarrhea, progressive weight loss and ulcers (Traversa et al., 2006).

Histopathological survey indicated two distinct lesions characterized by widespread

infiltration of inflammatory mononuclear cells with some eosinophils in the mucosa and

submucosa. A huge granulation mass located in the deep layer of the submucosa containing

9
granulomatous necrotic tissue and cross sections of Draschia megastoma were also seen

(Nadalian et al., 1997). Grossly, Draschia megastoma causes tumor-like nodules in the

Margo plicatus when the parasites burrow in the sub-mucosa (Maxie et al., 2016).

Triodontophorus and esophagodontus species are linked with nodules in the mucosa and sub-

mucosa of the caecum and colon (Maxie et al., 2016).

2.1.3: Lesions Associated with Gasterophilus species

In a study done by Al-Mokaddem et al (2015) in Egypt, Gasterophilus larvae infestations

were discovered in every case where the larvae were attached to the non-glandular part of the

stomach, gastric outlet and the proximal duodenum. Upon removal of the larvae, tiny

superficial erosions were seen at their sites of attachment. Microscopically, the attachment

site appeared as a deep, pitted ulcer with exposed lamina propria, granulation tissue

formation and neutrophilic infiltration. Grossly, the stomach mucosa revealed hyperemia,

edema, erosions and ulcers. Microscopical examination showed hyperkeratosis, acanthosis,

vacuolar degeneration of stratified squamous cells, gastritis, erosions, ulcerations, scarring,

hyperactivity of mucus glands, periglandular fibroplasia and parasitic granulomas with

infestation by Gasterophilus larvae (Al-Mokaddem et al., 2015). The epithelial margins and

deep layers of the stomach also develop rete pegs (Maxie et al., 2016). Additional health

concerns that may result due to high infestation of these larvae are; chronic gastritis, gastric

ulcers, esophageal paralysis, peritonitis, rupture of the stomach, squamous cell tumors and

anemia (Williams and Knapp, 1999).

10
2.1.4: Lesions Associated with Cestodes

Macroscopic and microscopic lesions linked to Anaplocephala perfoliata include mucosal

erosion, ulcerations, edema accompanied by eosinophilic and mononuclear infiltration in the

mucosa and basal membrane of the caecum and colon (Sangioni et al., 2000).

The tapeworm larvae (Hydatid cyst) form large cysts in the liver of the donkey which can

occlude the liver (Svendsen, 1986).

2.1.5: Lesions Associated with Trematodes

Hepatic fibrosing granulomas were found to be a result of chronic schistomiasis due to the

eggs that were escaping from the blood vessels containing the adults (Buergelt and Greiner.,

1995).

The liver may have an irregular outline and appear pale and firm. In chronic cases, hepatic

fibrosis and hyperplastic cholangitis is usually seen. The types of fibrosis include post

necrotic scarring, ischaemic fibrosis and fibrosis associated with flukes in the bile ducts.

Calcification of the bile ducts is more common in bovines (Boray and Murray., 1999). In

Egypt, bovine histopathological examination of tissues due to acute hepatic fascioliasis

showed hepatic necrosis and degeneration with numerous abscesses in the hepatic

parenchyma that was characterized by homogenous mass of necrotic cells bordered by heavy

aggregations of inflammatory cells chiefly neutrophils, histiocytes and lymphocytes. The

abscesses were surrounded by fibrous connective tissue capsule (Sohair and Eman, 2009).

Chronically, increased fibrous connective tissue in the portal triads was seen. The biliary

epithelium were hyperplastic characterized by formation of a large numbers of new bile

ductules and existence of mature fasciola worm inside the lumen of the main bile

ducts(Sohair and Eman, 2009).

11
2.2: Animal Risk Factors for Occurrence of Gastrointestinal Parasites

Significantly higher positivity for helminths has been observed in males (85.18 %) when

compared with female (50%) donkeys (Sathiyamoorthy et al., 2016). According to Jajere et

al (2016), age, sex and season were not statistically associated with the risk of helminth

infection. On the other hand, body condition scores, settlement, anthelmintic medication

history and management practices were significantly linked with the risk of gastrointestinal

helminthosis. A high prevalence of helminthic infections was seen in donkeys that had a poor

body condition, those from rural settlements, those with no anthelmintic treatment and those

that are raised under poor management systems (Jajere et al., 2016). In a study done in

Ethiopia by Ibrahim et al (2011) they established that body condition was an important factor

that influenced the occurrence of some helminth parasites as the parasites were more

prevalent in animals with a poor body condition than those that were well conditioned. It was

then recommended that donkey owners ought to be trained to better the management system,

particularly with regard to the level of nutrition to ensure that the donkey has a good body

condition that bestows some level of resistance against helminth infection (Ibrahim et al.,

2011)

2.3: Occurrence of Donkey Hemoparasites and Associated Lesions

African animal trypanosomiasis is one of the key hurdles to livestock development and

agricultural production (Mekuria et al., 2010, Boada-Sucre et al., 2016). In a cross sectional

study of trypanosomosis done in South Ethiopia, 10.7% of the donkeys were positive for

Trypanosoma vivax and Trypanosoma congolense as seen by dark ground and phase contrast

buffy coat method and Giemsa stained blood smears (Mekuria et al., 2010).

Three species of trypanosomes identified in donkeys by Abebe and Wolde (2010) included

Trypanosoma congolense (52.4%), Trypanosoma brucei (28.6%) and Trypanosoma vivax

12
(19.05%). The observed red blood cell alterations are as a result of mechanical and

biochemical damage due to host-parasite interaction that occurs in the bloodstream (Boada-

Sucre et al., 2016). Additionally, the morphological changes of the red blood cells in

Trypanosoma vivax infection are seen to be a contributory factor for the pathophysiology of

the disease (Boada-Sucre et al., 2016). In an experimental infection carried out in five

donkeys in Brazil, Trypanosoma evansi infection resulted to decrease in hemoglobin levels,

packed cell volume and erythrocyte count. Biochemically, the icteric index and serum

globulins was high. The serum albumin and glucose levels had also decreased. At necropsy

the donkeys exhibited splenomegally characterized by increased white pulp,

lymphadenopathy of the mediastinal lymph nodes, pulmonary and hepatic congestion as well.

Meningoencephalitis was also seen and was characterized by perivascular cuffing.

Demyelination in some aspects of the cerebellum pediculus and neutrophil vacuolization was

observed. Hemosiderosis and lymphoid hyperplasia in the lymph nodes and spleen was seen.

The kidneys also exhibited chronic nephritis (Cadioli et al., 2006). In a more recent study

undertaken by Garba et al (2016) in Nigeria, a gross and histopathological study was done on

18 donkeys, where six of them were infected with Trypanosoma spp. The gross lesions in six

donkeys included lung congestion with frothy exudate in the trachea in cases that were acute.

In chronic cases, grey hepatization, hydrothorax, hydroperitoneum, serous atrophy of fat and

mild adherence of renal capsule was observed. Microscopically, the lungs showed congestion

and mononuclear cellular infiltration. Hemosiderosis and lymphocytic depletion was seen in

the germinal centres of the spleen. The liver was congested with moderate focal necrosis and

cellular infiltration of mononuclear cells in nature (Garba et al., 2016).

Poikilocytosis is a general terminology that refers to red blood cells with an abnormal shape.

The specific cell changes can also be described and are of diagnostic significance.

Acanthocytes are defined as sphere shaped red blood cells with blunt tipped spicules that are

13
irregular in pattern at the margins. Echinocytes/crenated red blood cells are those that have

sharp blunt spicules of uniform length and are more evenly spread at the periphery. In horses,

the echinocytes may have blunt ends owing to relatively smaller sizes of red blood cells. This

morphological alteration has been seen in horses with colic and diarrhea

(http://eclinpath.com/hematology/morphologic-features/red-blood-cells/poikilocytosis/).Light

microscopic studies have demonstrated that Trypanosoma evansi produces poikilocytosis

(Silva et al., 1995).

Morphological alterations in the ovine red blood cells due to Trypanosoma evansi as

observed under scanning electron microscope in Venezuela included vesicle formation in the

red blood cell, microspherocytosis and deformation of the red blood cells (Boada-Sucre et al.,

2016). Several erythrocytes abnormalities were observed in the blood of Trypanosoma spp

infected dogs and horses and included the appearance of microspherocytes, acanthocytes,

dacrocytes, codocytes, vacuolated and sometimes bizarre shapes of red blood cells.

Polychromasia and poikilocytosis were present in both dogs and horse erythrocytes (Silva et

al., 1995).

Equine piroplasmosis is a tick-borne disease caused by hemoparasites namely; Babesia

caballi and Theileria equi (Formerly Babesia equi). It is a notifiable disease and a key

limitation to the international movement of equines (Gizachew et al., 2013). Babesia equi has

however been moved to the genus Theileria by Mehlhorn and Schein based on its

preerythrocytic developmental stages in the lymphocytes (Mehlhorn and Schein, 1998).

Infected animals may remain carriers for protracted periods and act as sources of infection for

ticks, which are the vectors (OIE, 2014). In acute clinical cases when the parasitaemia is high

it is difficult to detect the parasite and for this reason serological methods are preferred

though they may also give a false positive/negative (OIE, 2014). Necropsy findings include;

anemic and icteric organs, hydrothorax, ascites, hydropericardium, subcutaneous edema,

14
splenomegally, hepatomegally with the distension of the gall bladder, pulmonary congestion

and lymphadenopathy. In cases of Theileria equi, mucosal hemmorhages and hypertrophy

with accompanying inflammation of the lymph nodes is more pronounced (Brandt, 2009). In

a retrospective study done by Rassouli and Aghazaman (2015), a certain group of dog blood

films were subjected to examination to look for tick borne pathogens and the red blood cells

showed polychromatophilic macrocytes, hypochromic reticulocytes, poikilocytosis and

anisocytosis. Sex-related variation in the prevalence of Theileria and Babesia infections in

donkeys is absent as shown by serohematological study (Gizachew et al., 2013).

Babesia caballi and Theileria equi are most often associated as they share common vectors.

However, infections with Theileria equi are more prevalent than Babesia caballi infections

(Friedhoff et al., 1990).

Setaria equina, family: Onchocercidae, subfamily: Setariinae; is a nematode filarial parasite

usually found in the peritoneal cavity of equines in diverse geographical regions of the world

(Coleman et al., 1985). The adult; in the peritoneal cavity is usually harmless and are found

incidentally during pathological examination (Hillyer et al., 2001). The infection is

transmitted by mosquitoes. The sheathed microfilaria worms are found in the blood (Yeargan

et al., 2009). Microfilaria of equine in Baghdad was reported to have an occurrence rate of

11.11%. The technique used for screening was Knott technique (Afkar and Amall, 2014).

Another study in Hungary reported a percentage occurrence of 9.2 of microfilaria of Setaria

spp using a similar technique which is regarded as a more sensitive (Hornok et al., 2007). In

Iraq, the percentage of infection amongst horses was 30.76% by Knott technique (Suleiman

et al., 2012). The presence of adult worms in the peritoneal cavity is not always

accompanied by microfilaraemia (Hornok et al., 2007).

15
 3.0 CHAPTER THREE: MATERIALS AND METHODS

3.1: Study Areas

Three study sites were purposively selected (Figure 1) based on the presence of donkey

abattoirs. These included:

Site A: Silzha Company in Lodwar town, Turkana County. Turkana County is the second

largest in terms of land area. It is bordered by the country of Uganda to the west. Lodwar is

the largest town and is Turkanas’ capital (Wikipedia, 2019).

Site B: Goldox Kenya Limited in Mogotio, Baringo County. The economy is based on

agricultural activities with hides and skin as one of the main livestock products. Mogotio is

one of the urban centres (Wikipedia, 2019).

Site C: Star Brilliant Donkey Export Slaughter House in Naivasha, Nakuru County. Naivasha

is a major significant urban centre in Nakuru County (Wikipedia, 2019).

16
 SITE A

SITE B

SITE C

Figure 1: Map of Kenya showing the three Sites of Study; Turkana (A), Baringo (B) and
Nakuru (C) County (Source: d-maps.com accessed at https://d-
maps.com/pays.php?num_pay=30&lang=en)

3.2: Study design

A cross-sectional study was undertaken in the three donkey slaughter houses in the month of

July to September 2017 in order to examine the donkeys, collect blood, fecal, parasite and

organ samples for hemoparasites, parasitological and pathological analyses respectively.

Upon arrival at the slaughter house, the movement permits of the donkeys were verified by

the meat inspector. The animals were then subjected to antemortem inspection and put in

holding pens with water ad lib. The period the animals were put in the holding pens depended

on the total number of donkeys present for slaughter. If the donkeys present were small in

number, they weren’t slaughtered due to economic reasons. The donkeys for slaughter were

17
led into the lairage and then into the stunning box where they were humanely killed via use of

a captive bolt piston. They were then hoisted up and the jugular vein slithered to achieve

bleeding. Skinning was then done followed by evisceration of gastrointestinal tract and the

red offals. Each section of the gastrointestinal system was then inspected.

3.3: Sample Size Determination

The sample size was determined according to the number of animals presented for slaughter.

A total of 282 out of the 1,538 donkeys presented for slaughter were examined for

gastrointestinal parasites. Systematic sampling method was applied at the slaughter house

level whereby every 5th animal was examined depending on the number presented for

slaughter. While examining the thin blood and buffy coat smears simple random sampling

method was used and the random numbers for the slides to be examined was obtained via use

of a scientific calculator (FX-82 MS). A total of 160 thin blood smears and 160 buffy coat

smears were examined under light microscope. The formulae used to derive the sample size

for the smear examination was as follows:

In a finite population; N=282

n= 384 which is the sample size for an infinite population

Adjustment; where n (the new sample size);

n=1/ (1/384+1/282) = 162 (Hewson, 2004).

Sampling proportional to size:

Mogotio; 114= 114/282=0.4X 162=65

Kinamba; 148= 148/282= 0.52X162=84

Lodwar; 20= 20/282=0.07X162=11

18
0.4+0.52+0.07=0.99≈ 1

All samples=160X 2=320 thin blood and buffy coat smears.

65+84+11=160

3.4: Determination of Prevalence and Intensity of Gastrointestinal Parasites and Risk

Factors for Helminth Infestation in Donkeys

3.4.1: Prevalence of Gastrointestinal Parasites

Collection of Gastrointestinal Parasites and Identification: The gastrointestinal system

was examined from the esophagus to the rectum and parasites were picked from the

gastrointestinal tract and the liver. The gross lesions linked with the parasites were noted

down. The parasites were then rinsed gently with water and preserved in 80% alcohol for

further analysis. The parasites were then identified based on their gross/microscopic

morphological features as described by Soulsby (1982). Cestode samples collected were

processed according to a technique described by International Institute of Parasitology (1994)

and identified morphologically according to Soulsby (1982). The arthropod larvae were

obtained from the gastric mucosa by use of a forceps. The different species were identified by

virtue of their morphological features as described by Soulsby (1982). The proportion of

donkeys positive for a specific parasite was determined in terms of percentages.

3.4.2: Intensity of Gastrointestinal Helminths Infection

Fecal sample (5-10 grams) was collected directly from the rectum of each animal post

slaughter so at to determine the intensity of helminth infection. The sample was labelled and

preserved in 10-15 mls of 80% alcohol to prevent hatching of the eggs. The samples were

transported to the Parasitology laboratory at the department of Veterinary Pathology,

Microbiology and Parasitology (UoN). McMaster egg count technique was applied in the

processing of the samples according to MAFF (1986). Identification of the helminth eggs and

19
cysts was done and their counts performed. The intensity of infection for strongyle eggs was

recorded for each donkey and classified according to established infection intensity classes

into one of the four classes: none, low (up to 500EPG), medium (501-1000 EPG), high

(>1000 EPG) (Soulsby 1982).

3.4.3: Risk Factors for Helminth Infestation in Donkeys

Data regarding approximate age, sex, origin of the animal, pregnancy status, physical strength

and the body condition score estimate were noted (Appendix 1.0). The body condition scores

were given in a scale of 1 to 5(1-Poor 2-Moderate, 3-Ideal 4-Fat 5-Obese) according to

National Equine Welfare Council (2005); (see attached appendix 2). This data was compared

with the fecal strongyle eggs intensity and the prevalence of the adult helminths.

3.5: Determination of Occurrence of Hemoparasites in Donkeys

To determine the proportion of donkeys infected with hemoparasites, the number of donkeys

positive for hemoparasite was divided by the overall number of donkeys examined and

multiplied by 100.

Sample collection and processing: A 10ml venous blood sample was collected from each

animal during bleeding after the donkey was stunned. A thin blood smear was made

immediately, air dried and fixed in absolute methanol. The remaining volume of the blood

sample was immediately transferred into an EDTA blood vacutainer and the whole lot was

centrifuged at 300 r.p.m for 15 minutes to separate blood components. A thin buffy coat

smear was prepared, fixed in absolute methanol and stained using Giemsa (1:5dilution). The

smear was then screened for the presence of hemoparasites under the oil immersion objective

20
lens (×1000). Identification of hemoparasites was based on their morphological features (Brar

et al., 2002). The cellular changes on thin blood smears were also noted down.

3.6 :Characterization of Lesions Associated with Gastrointestinal Parasites in Donkeys

in Selected Abattoirs in Kenya

The postmortem inspection of gastrointestinal tract was carried out in the green area whereas

the liver was inspected in the red area. The green and red area areas are specific zones in the

slaughter where the inspection of these organs is carried out. The lesions associated with

helminths were characterized using the macroscopic and microscopic appearance. Gross

lesions were identified and characterized according to their variation in size, shape, color and

consistency as compared to normal organs. Special attention was paid to liver, stomach and

the intestines. This was because; a previous pilot study done by Mulwa et al (unpublished

data) that indicated that the helminth infestation was high in these organs. Gastrointestinal

organ samples were collected and fixed in 10% buffered formalin then processed routinely

via the paraffin wax method and stained with Hematoxylin and Eosin according to Carson

and Hladik (2009). The tissue sections obtained thereafter were examined using a light

microscope at X40, X100 and X400 magnifications. The lesions observed at each section

were then recorded as per the organ affected and type of lesion.

3.7: Data Analysis

The data was entered into Microsoft Office Excel Data sheet (2007), cleaned and coded. It

was later on transferred into statistical software (SPSS) version 22 for descriptive statistics

and various association tests. The proportion of the donkeys with the parasites was

determined using simple percentage methods. The proportion of the four classes of strongyle

egg infection was also determined. Prevalence estimates were obtained for overall infection.

Nonparametric correlation tests and independent t test were used to quantify associations

21
between independent variables (age, pregnancy status and sex) and intensity of strongyle

infection as well prevalence of helminths. The level of significance was set at p<0.05. Results

were considered significant in cases where the P value was less than 0.05. The confidence

level was held at 95%.

22
 4.0 CHAPTER FOUR: RESULTS

4.1: GENERAL INFORMATION

A total of 282 donkeys were examined; 20 from Lodwar, 114 from Mogotio and 148 from

Kinamba slaughterhouses (Table 1). Of these, 40% (114/282) were females while 60%

(168/282) were males. Ninety four percent of the sampled animals were adults whereas 6%

were juveniles. The number of animals observed to be weak prior slaughter were 1.4%

(4/282) of the animals while 98.6% (278/282) of the animals appeared physically strong.

A high proportion of the animals had a poor body condition (89%; 250/282). Moderate body

condition was recorded in 11% (32/282) of the animals. Samples examined included; 36

tissues from the gastrointestinal system, 282 fecal, 160 blood smears and 160 buffy coat

smears.

Table 1: Proportion of Animals Examined per Slaughterhouse

Lodwar Kinamba Mogotio Total

No. slaughtered 70 148 1320 1538

No. sampled 20 148 114 282

Proportion of the (20/282); (148/282); (114/282); 100%

sampled animals 7.1% 52.5% 40.4%

Gravid females donkeys presented for slaughter were 16.7% (19/114) whereas non-gravid

ones were 83.3% (95/114). Out of the 19 gravid female donkeys, 52.6% (10/19) were positive

for strongyle eggs whereas, 47.4% (9/19) females were negative for strongyle eggs. The

23
study found out that pregnant females had no statistically significant fecal egg count (178

±265.8 E.P.G) as compared to the non-pregnant females (141.1 ±228.5 E.P.G)

(P=0.522) (Table 2).

Table 2: Mean Egg Count between the Gravid and Non-gravid Female Donkeys

Mean of
No. of the
female strongyle Std. Error
Pregnancy status donkeys eggs Std. Deviation Mean

Strongyle egg Pregnant

19 178.9474 265.78803 60.97596
count

Not Pregnant
95 141.0526 228.54523 23.44825

4.2: Prevalence and Intensity of Gastrointestinal Parasites and Risk factors for

Helminth Infestation in Donkeys in Three Abattoirs in Kenya

4.2.1: Intensity: Fecal Egg Count

Fifty five point seven percent (157/282) of the fecal samples analyzed using the McMaster

technique were positive for helminth eggs while 44.3% (125/282) were negative. Fifty five

point three percent of the donkeys (156/282) of the donkeys were negative for strongyle eggs,

39% (110/282) had a low infection rate (up to 500 EPG), while 5% (14/282) had a medium

infection rate (501-1000EPG) with 0.7% (2/282) having a high infection rate (>1000 EPG).

See table 3 below.

24
Table 3: Table showing the intensity of the Strongyle egg infection rates for slaughtered

donkeys

Level of Infection No. of donkeys in each category Proportion (%)

High (>1000) 2 0.7

Medium (501-1000) 14 5

Low( up to 500) 110 39

None 156 55.3

Total 282 100

4.2.2: Animal Risk Factors and Helminth Infestation

There was no significant difference in the fecal strongyle egg count between the adults and

the juveniles (P=0.283). Additionally, there was no significant difference in strongyle eggs

shed between males and females (P=0.529). There was a significant difference in the

strongyle egg count shed between the poor and moderate body condition scores (p= 0.001). A

correlation test to determine the presence of helminth parasite in the age and sex revealed that

there was no association between age and sex of a donkey and the presence of helminths

(p=0.739; p=0.624). There was no association between body condition scores and the

prevalence of the helminths (X2(O.121a, p=0.488). (Appendix 2).

25
4.2.3 Prevalence of the Helminth Eggs in the Slaughtered Donkeys

A total of 44.7 % (126/282) of the donkeys were positive for strongyle eggs(Figure 3),

Parascaris equorum affected 5.3% (15/282) of the donkeys(Figure 4); followed by Oxyuris

equi at 1.1% (3/282); (Figure 5),Triodontophorus tenuicolis and Habronema both at 0.7%

(2/282) and cestodes eggs had the lowest prevalence at 0.4% (1/282) (Figure 2). The means

and ranges of the various eggs and cyst were demonstrated. The strongyle egg counts ranged

from 0-1900 with an average of 136.52 eggs per gram of feces (EPG) followed by Parascaris

equorum with a mean of 13.12 EPG. The least was the cestode egg with a mean of 0.35 EPG.

Coccidia and Giardia cysts had an average of 3.90 and 2.13 oocysts per gram of feces

respectively (Table 4).

Figure 2: Prevalence of the Helminth Eggs in Fecal Samples of Slaughtered Donkeys

26
Figure 3: Photomicrograph showing a typical strongyle type egg(S), thin shelled and

ovoid in shape from donkey (ID: 22 KNB)

Figure 4: Photomicrograph showing the spherical brownish egg of Parascaris equorum

(PE) from donkey (ID: 51 KNB) and an air bubble (A).

27
Figure 5: Photomicrograph Showing the Egg of Oxyuris equi from donkey (ID: 75

MGT) the mucoid plug (MP) and the flattened side (FS) are illustrated.

Table 4: Showing infestation intensity ranges and means ±standard error of the mean of
helminth eggs, cysts and oocysts recovered from the fecal samples

Identity of the eggs/cysts Number Intensity Mean± Standard error of the

sampled ranges mean

Cestode spp 282 0-100 0.35±.355

Coccidia spp 282 0-500 3.90±2.027

Giardia spp 282 0-400 2.13±1.584

Habronema spp 282 0-100 0.71±.501

Oxyuris equi 282 0-300 2.13±1.323

Parascaris equorum 282 0-500 13.12±3.660

Strongyle eggs 282 0-1900 136.52±14.153

Triondontophorus 0-100
282 0.71±.501
tenuicolis

28
4.2.4: Prevalence of the Cysts

A total of 2.1% (6/282) of the donkeys were positive for Giardia cysts while 0.7%(2/282) of

the donkeys were positive for coccidian oocysts.

4.3: Occurrence of the Various helminths in Donkeys and Associated Lesions

Out of the 282 donkeys sampled, 85.5%(241/282) were positive for various helminth

parasites whereas 14.5%(41/282) were negative. A total of 36 donkeys had severe gross

lesions. Twenty two were from Mogotio, eleven from Kinamba and three from Lodwar

slaughterhouses. Lesions were mainly found in the intestine 47.2% (17/36), followed by the

liver 41.7% (15/36) and lastly the stomach at 11.1% (4/36).

4.3.1 Stomach

In the stomach, Gasterophilus species occurred at a rate of 51.8% (146/282). A proportion of

38.3% (108/282) of the animals were positive for G. intestinalis, G.pecorum had an

occurrence of 7.8 % and 5.7% of the donkeys were positive for G. nasalis (Figure 6).

Figure 6: Proportion of the various species of the Gastrophilus larvae in the stomach of

slaughtered donkeys

29
The area that was affected by Gasterophilus species was the non-glandular region of the

stomach. Grossly multiple ulcers that were regular, circular with elevated margins and a

depressed center were observed after dislodgement of the various Gasterophilus species. The

mucosa was tinted yellow in some areas (Figure 7). In a donkey (ID: 72 MGT) the

keratinized layer of the non-glandular epithelium of the stomach had peeled off.

Microscopically, the mucosal epithelium was thickened in some areas and there was loss of

tissue architecture with accompanying dilatation of the sub-mucosal blood vessels.

Additionally there was disruption of the keratinized layer of the non-glandular stomach. The

keratin covering the surface epithelium had a yellow tinge. Empty white spaces were visible

in the lamina propria (Figure 8, 9 and 10). In another case there was a notable neutrophilic

cellular infiltration in the exposed lamina propria and accompanying formation of rete pegs.

Vacuolar degeneration was observed in one case and appeared like empty white spaces below

the mucosa (Figure 10).

Figure 7: An ulcer (shown by the black arrow) on the non-glandular area of the donkey

stomach with some areas of the mucosa tinted yellow from donkey (ID: 20 KNB);

several Gasterophilus species are present (white block arrow).

30
Figure 8: Photomicrograph of a stomach section of a slaughtered donkey (20 KNB)

illustrating the disruption of lamina propria (white block arrow) and discontinuity of

the stratified squamous epithelium (X10; H&E)

31
Figure 9: Photomicrograph of the stomach of a slaughtered donkey (20 KNB) showing

engorged blood vessels at the submucosa as illustrated by the white block arrows (X40;

H&E)

32
Figure 10: Photomicrograph of the stomach mucosa of a slaughtered donkey (ID:

20KNB) showing vacuolar degeneration (illustrated by the white arrows) in the

submucosa of the stomach due to Gasterophilosis (X40; H&E)

4.3.2: Intestines

Anaplocephala magna had an occurrence of 2.5%(7/282) whereas Anaplocephala perfoliata

(Figure 11) had an occurrence rate of 10.3%(29/282). Cylicocyclus auriculatus was found in

2.1% (6/282) of the donkeys. Cyathostome species were present in 1.4% (4/282) of the

donkeys. Thelazia equi was present in one donkey represented at (0.4%) 1/282, Parascaris

equorum was represented in 20.2% (57/282) of the donkeys, Strongylus edentatus at 12.1%

(34/282), Strongylus equinus at 0.4 % (1/282), Strongylus vulgaris at 52.8 % (149/282),

Setaria equina at 3.5 %( 10/282) and Triodontophorus serratus at 0.4% (1/282) (Figure 12).

33
Figure 11 A and B: A photomicrograph of Anaplocephala perfoliata recovered from the

ileocecal junction of slaughtered donkey (ID: 10 MGT), illustrating the lappets (L) and

the scolex (SC) which is distinct and smaller than the body. On the right side is the same

cestode after staining with acetoalum carmine stain illustrating the scolex (SC), lappets

(LP) and the proglottids (P). The white block arrow shows an artifact (air bubble).

34
Figure 12: Prevalence of the various helminth species in the intestines of slaughtered
donkeys in Kenya

In the ceca of 8 donkeys, focal elevated areas measuring about 2-3mm that were firm in

consistency were visible (Figure 13). Hyperemia and oedema were also evident on the

mucosa. On incision into the cavity a slender reddish worm was observed and identified as

Strongylus edentatus. Microscopically, there was disruption of the mucosal lining; nematode

larval stages were encapsulated within a lumen (Figure 14). The outer zone of the capsule

was surrounded by fibroblasts and neovascularization was also evident (granulation tissue).

The submucosal blood vessels were also dilated (Figure 15).There was also a case (ID:

9MGT) with parasite in early developmental stages coupled with eosinophilic infiltration. In

some instances there was cellular infiltration in the submucosa characterized by plasma cells,

macrophages and eosinophils. The eosinophils seemed to form a vast number of the

population as they were also present in the lamina propria. On incision into other elevated

35
areas on the intestinal mucosa that were firm and whitish in color, a purulent discharge was

visible. Microscopically, there was necrotic debri with degenerated neutrophils surrounded by

a capsule at the periphery.

Irregular circumscribed hyperemic areas of epithelial loss were seen on the ileocecal junction

region after dislodgement of cestodes attached at the region. Microscopically, eosinophils

were present in the submucosa with accompanying dilatation of the submucosal blood

vessels. Mucosa was disrupted with lymphocytic cellular infiltration, localized areas

surrounded by high density fibrous connective tissue and goblet cells were hyperplastic. In

another case, there was massive infiltration by macrophages, eosinophils and lymphocytes

with accompanying neovascularization in the submucosa. There were instances where the

lymph vessels were fully dilated with a cellular infiltration (eosinophils, neutrophils and

macrophages). In this case Strongylus vulgaris were retrieved from the cecal nodules. There

was cecal tissue that had pink eroded areas whose wall was hyperemic and edematous.

Numerous Strongylus vulgaris species were attached on the mucosa as well (Figure 16). On

microscopic examination, there was a parasitic larva within the tunica mucosa where the

larva was surrounded by fibrous connective tissue with a characteristic eosinophilic

infiltration (Figure 17, 18 and 19).

36
Figure 13: A picture of the cecal wall from slaughtered donkey (ID: 13MGT) showing

focal nodular lesions, about 2-5 mm (white block arrow) and a reddish nematode

attached onto the mucosa; Strongylus vulgaris (thin black arrow).

Figure 14: Photomicrograph of the ceca of donkey (13 MGT) showing four regular

structures identified as nematode larvae suspected to be larval stages of Strongylus

vulgaris enclosed in a fibrous capsule(White block arrow) A portion of the cecal mucosa

has been disrupted also(Black arrow) (X4).

37
Figure 15: Photomicrograph of slaughtered donkey (ID: 13 MGT) illustrating the

fibrous capsule zone (Black arrow) that’s encapsulating the nematode larva (white

block arrows). The fibrous connective tissue has mixed inflammatory infiltrate (black

arrow) (X10; H&E).

38
Figure 16: Picture showing attached nematodes on the cecal mucosa (black arrow),

Strongylus vulgaris (SV). The mucosa is edematous, hyperemic and eroded (EHM) (ID:

3 MGT)

Figure 17: photomicrograph of donkey (ID: 3MGT) illustrating the larvae encapsulated

within a lumen (L) with red blood cells adjacent (RBC) and a thin fibrous capsule layer

(FC). The mucosa can be seen with a few glands (M), the blood vessels are also engorged

with blood (EBV) (X4; H&E)

39
Figure 18: Photomicrograph of donkey (ID: 3MGT) illustrating the mixed

inflammatory infiltrate as shown by the arrow and the fibrous capsule (FC) (X40;

H&E)

40
Figure 19: Photomicrograph of donkey ceca showing the submucosa of a donkey (9

MGT) illustrating the mixed inflammatory infiltrate with the eosinophils predominating

(black arrow) (X40; H&E)

41
4.3.3: Liver

In the liver, Strongylus edentatus was retrieved. Hepatic enlargement was observed as the

margins were rounded in the 15 donkeys sampled and in addition, eight of the liver tissues

exhibited cyst-like focal swelling measuring 2-3 cm in diameter that protruded above the

capsule (Figure 20). On incision, blood filled cavity containing a slender nematode was

observed in the eight cases. The nematode was identified as Strongylus edentatus.

Microscopically, there was a well demarcated zone characterized of hemorrhage, loss of

hepatic architecture and brown pigment deposition. In some cases there was a well

demarcated zone between the liver and the nodule with red blood cells, necrotic debri filling

the area where the helminth resided (Figure 21). In some cases, the well demarcated zone

had deposition of fibrous connective tissue proliferated bile ducts in the portal triad, cellular

infiltration and disrupted sinusoids.

Greyish pinpoint necrotic foci were also observed in one of the donkeys which had a high

infestation of Parascaris equorum. On microscopic examination, the foci were composed of

neutrophils interspersed amongst cell debri at the centre and a fibrous capsule at the margin

which was suggestive of hepatic abscessation (Figure 22 and 23).

42
Figure 20: Liver of a slaughtered donkey (ID: 60 MGT) showing the cyst-like swelling

(white arrow) protruding above the capsule.

Figure 21: Photomicrograph of the liver of a slaughtered donkey (60 MGT) showing
the well demarcated zone (ZC) with the central vein engorged with blood (CV) , the
enclosed material(CD) is where the retrieved helminth (Strongylus edentatus) was
residing and is composed of necrotic material and no hepatic tissue can be
appreciated(X10; H&E)

43
Figure 22: Picture of a liver of a slaughtered donkey (ID: 18 LDW) showing multifocal

greyish areas on the liver surface.

Figure 23: Photomicrograph of a donkey liver (ID: 18 LDW) showing a focal area with

necrotic material with interspersed neutrophils (N) and an accompanying fibrous

capsule (FC)-Hepatic abscess. The central vein is also illustrated (CV) (X10; H&E).

44
4.4: Occurrence of Hemoparasites in the slaughtered donkeys

In the 160 buffy coat smears examined, 11.3% (18/160) were positive for the various

hemoparasites while 88.8% (142/160) were negative. As for the blood smears, 6.25%

(10/160) were positive whereas 93.75% (150/160) were negative (Table 5). Mixed infections

were represented at 1.25% (2/160) for Babesia caballi and Trypanosome spps as examined in

buffy coat smear.

Table 5: Table showing the number of positive cases for thin blood smears and the buffy

coat smears from slaughtered donkeys from the three slaughter houses

Hemoparasite No. of donkeys No. of donkeys

identified positive on thin positive on buffy

blood smears coat smears

examination examination

Anaplasma spp 0 4

Trypanosoma spp 1 9

Babesia caballi 8 4

Microfilariae 0 1

Theileria equi 1 0

Total number of 10/160 18/160

positive donkeys

Overall Proportion 6.25% 11.3%

(%)

45
4.4.1: Buffy Coat Smears

Anaplasma spp had an occurrence of 2.5%(4/160), trypanosomes at 5.6%(9/160), Babesia

caballi at 2.5%(4/160) and microfilaria at 0.60%(1/160) (Figure 24). Photomicrographs of

the various hemoparasites have been illustrated (Figure 25,26 and 27).

Figure 24:Occurrence of Hemoparasites as Examined on the Buffy coat smears of

slaughtered donkeys

46
Figure 25: Photomicrograph of a buffy coat smear of animal (ID: 2LDW) showing
trypanosome spp (white block arrows) in a donkey, (X100;Giemsa Stain)

47
Figure 26: Photomicrograph of a buffy coat smear from donkey (ID: 4LDW) showing a
lymphocyte with purple staining inclusions at the margin (Anaplasma spp-Black arrow).
(X100;Giemsa Stain)

Figure 27: Photomicrograph of a buffy coat smear of a slaughtered donkey(ID:

80KNB) showing Setaria equina microfilariae(MF), a band neutrophil(N) (X100;Giemsa
Stain)

4.4.2: Thin Blood Smears

In the 160 animals that were screened, Babesia caballi had an occurrence of 5% (8/160),

Theileria equi with an occurrence rate of 0.6% (1/160) Trypanosoma spps at an occurrence

rate of 0.6% (1/160) as illustrated in Table 6. Some of the hemoparasites identified have been

shown also (Figure 28, 29, 30 and 31).

48
Table 6:Occurrence of Hemoparasites as Examined on the Thin Blood Smears from

slaughtered donkeys

Hemoparasite Frequency Proportion (%)

Babesia caballi 8/160 5%

Theileria equi 1/160 0.60%

Trypanosoma spps 1/160 0.60%

Figure 28: Photomicrograph of a thin blood smear from donkey (ID: 4LDW) showing
red blood cells with two pyriform shaped inclusions black arrow( Babesia caballi). Note
the red blood cells that are polychromatophilic with varying shapes and sizes.(X
100;Giemsa Stain)

49
Figure 29: Photomicrograph of a buffy coat smear from donkey (ID: 78 KNB) showing

a red blood cell with Babesia caballi (X100 Giemsa stain)

50
Figure 30: Photomicrograph of a buffy coat smear from a donkey (ID: 78KNB)

trypanosome spp can be seen (White arrow head) (X100; Giemsa Stain).

The various cellular morphological alterations associated with the hemoparasites included;

poikilocytosis, anisocytosis, polychromasia, hypersegmented neutrophil and reactive

lymphocytes (Figure 31) (Table 7).

51
Table 7: Cellular Morphological Alterations due to Hemoparasites on slaughtered

donkey blood smears in Kenya

Hemoparasite(s) Associated Morphological Alterations

Trypanosoma Poikilocytosis(echinocytes) and anisocytosis

spps

Babesia caballi Poikilocytosis(echinocytes),anisocytosis,polychromasia and

hypersegmented neutrophil

Babesia caballi Poikilocytosis(echinocytes), anisocytosis and hypersegmented neutrophil

Babesia caballi Poikilocytosis(echinocytes) anisocytosis, polychromasia and reactive

lymphocyte

Theileria equi Poikilocytosis(echinocytes) anisocytosis and polychromasia

Figure 31: Photomicrograph if a thin blood smear from donkey( ID: 69 MGT)
illustrating a lymphocyte with vacuolated cytoplasm(white arrow head) and the
variation in shape and size of the red blood cells can also be appreciated(X100; Giemsa
Stain).

52
5.0: CHAPTER FIVE: DISCUSSION, CONCLUSIONS AND RECOMMENDATIONS

5.1: DISCUSSION

5.1.1: Proportion and Intensity of Endoparasites Infestation in Donkeys

According to this study, parasitism was found to be a prevalent condition (85.5%) amongst

donkeys in Kenya. This is supported by similar studies done by Karanja et al. (1994) and

Lewa et al. (2000) that showed gastrointestinal parasites are prevalent in donkeys in Kenya.

This differed a little to a study done in Ethiopia where the prevalence was 96.9% (Ibrahim et

al., 2011). This difference could be attributed to differences in geographical zones and

management systems as well. Majority of the donkeys in this study (89%) had a poor body

condition. This could be due to helminthosis in conjunction with a low plane of nutrition.

This is in agreement with (Saul et al., 1997) who reported helminth infestation as the most

common cause of death besides retarding growth or decreasing work output in addition to

distress and pain (Svendsen, 1997). Lewa et al. (1998) also noted that the unthrifty state in

donkeys was caused by heavy helminth infections, majorly the adult worms. An overall of 10

adult helminth species obtained included; Anaplocephala magna 2.5%(7/282),

Anaplocephala perfoliata 10.3%(29/282), Cylicocyclus auriculatus 2.1%(6/282),

Cylicocyclus leptostomus 0.4%(1/282), Cyathostome species 2.1%(6/282), Parascaris

equorum 20.2% (57/282), Strongylus edentatus 12.1%( 34/282), Strongylus equinus 0.4 %

(1/282), Strongylus vulgaris 52.8 %( 149/282), Setaria equina 3.5% (10/282) and

Triodontophorus serratus 0.4 %( 1/282).This finding differs with Ahmed et al. (2011) who

found the overall prevalence to be 98.5% with 22 adult helminth species and this could be

attributed to the different geographical regions and management systems. The most common

species in donkeys in our study was the Strongylus vulgaris (52.8%). Lewa et al. (2001)

found Gastrophilus intestinalis, Strongylus vulgaris and Strongylus edentatus in the six

donkeys examined at necropsy. Gasterophilus nasalis, Cyathostomum tetracanthum,

53
Habronema muscae, Strongylus spp, Setaria equina and Oxyuris equi were the common

parasites found in eight donkeys in Kenya (Karanja et al., 1994).This study has revealed

more other species occurring in donkeys in Kenya which include Triodontophorus serratus,

Cylicocyclus auriculatus, Cylicocyclus leptostomus Parascaris equorum, Anaplocephala

magna and Anaplocephala perfoliata. Habronema species eggs were also observed in two

donkeys in our study. The additional species could be due to the larger sample size used in

this study and also the donkeys were mostly from the arid and semiarid areas of Kenya

contrary to the previous study in which the donkeys were from Kiambu County only.

Getachew et al. (2010) observed 8% prevalence of Anaplocephala perfoliata in working

donkeys in Ethiopia and this is almost similar to the prevalence found in this study (10%).

Fecal examination in this study revealed that 44.7 % (126/282) of the donkeys were positive

for strongyle eggs, Parascaris equorum occurred in 5.3% (15/282) of the donkeys followed

by Oxyuris equi at 1.1 %( 3/282), Triondontophorus tenuicolis and Habronema spp occurred

each at 0.7 %( 2/282) infection rate and cestode eggs which were present in only one donkey

at 0.4 %( 1/282). According to a report by Ahmed et al (2011) in Ethiopia, fecal examination

showed 99% strongyle, 80% Fasciola, 51% Parascaris, 30% Gastrodiscus, 11%

Strongyloides westeri, 8% Anaplocephala perfoliata cestodes and 2% Oxyuris equi infection

prevalence. The infection level in this study was lower and this could be attributed to the

seasons in Kenya as the animals were sampled during the fairly dry season. Additionally,

donkeys originated mostly from the dry areas and these include Isiolo, Loitoktok, Laikipia,

Tana River, Suswa-Narok, Kajiado, Narok, Kitui and Karamoji (Uganda). Some animals also

originated from Kericho. According to Lewa et al. (1998) egg counts seemed to be relatively

low during the dry season. The egg count for the strongylid eggs ranged from 0-1900, which

is low as compared to that of Vercruysse et al. (1986) who established an egg count of 100-

9,200 for strongylid eggs. Karanja et al. (1994), in Kenya, found the infection to be low to

54
moderate for strongyle infestation, this is in agreement with the results of this study as most

donkeys had an egg count of 0-499 EPG. The number of adult helminths observed was

however high and this shows that it does not correlate to the fecal egg count. As observed in

this study, a fecal egg count of zero did not mean the donkey was parasite free. Additionally,

the animal could be harboring a majority of the immature stages of the helminths in high

numbers hence the resultant low/no egg count. This is an agreement with an observation

made by Matthews and Burden (2013), who stated that donkeys can harbor substantial levels

of parasitic infection but the parasites aren’t detectable via routine faecal egg count (FEC)

analysis.

5.1.2: Risk Factors to Occurrence of Helminths and Strongyle Egg Count

There was no significant differences in strongyle egg count between the adults and the

juveniles (P>0.05). This is in contrary to the study in Lesotho which indicated an inverse

association between age and intensity of strongyle infection (Upjohn et al., 2010, Yoseph et

al., 2005). Additionally no significant differences were observed in strongyle eggs shed

between the males and the females. This could suggest that the management of the animals at

the farm level is the same. This is in contrary to the study in Lesotho where a significant

strongyle infection for females was observed by Upjohn et al. (2010). Variations in the

husbandry practices between the age and sex were however not determined and this requires

further investigation. A significant difference was seen between the strongyle eggs shed and

body condition scores (P< 0.05). Yoseph et al. (2005) reported that animals with a poor body

condition had a high fecal egg count. Ibrahim et al. (2011) also found out that donkeys with a

poor body condition score had a high prevalence of helminth parasites as compared to well-

conditioned donkeys. This is agreement with this study that showed a high prevalence of

strongyle egg counts in poor body conditioned animals. A recent study carried out in

Morocco by Crane et al. (2011) showed that considerable increase in body condition score in

55
the equids that received anthelmintic treatment contrary to those in which a placebo was

administered. With regard to the prevalence of the helminths there was no association found

between the body condition score and the presence of the helminths in our study. This is in

agreement with a recent study done by Tesfu et al. (2014) who reported no association of

nematode infection with regard to body condition score of the donkeys. In this study, both

body condition score groups were equally affected and could mean that body condition score

of 2 or less could indicate a need for therapeutic intervention with antihelminthic drugs.

A nonparametric correlation test to determine the presence of helminth parasite in the age

and sex in this study revealed that there was no association between these two factors and the

presence of helminths. This is in agreement with Ibrahim et al. (2011) who found that all age

groups were equally affected. For sex however, the females were found to be mostly affected

as compared to males (Attia et al., 2018).This differs with this study and this could be

because both male and female donkeys are raised in the same environmental conditions.

5.1.3: Occurrence of Hemoparasites

A higher occurrence of Trypanosome spps were observed under the buffy coat smears(5.6%)

as compared to the thin blood smears((0.6%).This denotes that buffy coat smears were more

sensitive in detecting the Trypanosome spps. A proportion of 10.7 % of the donkeys in South

Ethiopia were positive for trypanosomes (Mekuria et al., 2010). This is a slightly higher

prevalence than the findings in this study as the method used in the latter was dark ground

and phase contrast buffy coat method (Mekuria et al., 2010). In this study, Theileria equi had

an occurrence of 0.6% whereas Babesia spps had an occurrence of 5%. This is in contrary to

a recent study done in Northern part of Kenya by Hawkins et al (2015), where 72% of the

56
donkeys were positive for Theileria equi whereas no animal was positive for Babesia caballi.

This variation could be explained by the technique used; PCR which is highly sensitive.

Microfilaraemia in equine in Baghdad was reported to have an occurrence rate of 11.11% via

use of Knott technique and species were identified as those of Setaria equina (Afkar and

Amall, 2014). In Iraq the percentage of infection amongst horses was 30.76% via use of

Knott technique (Suleiman et al., 2012). The occurrence rate in this study was at 0.6% which

is low and could be due to the standard technique used to screen for the hemoparasite and

also differences in occurrences could be due to the different geographical areas.

5.1.4: Characterization of Lesions Associated with Helminths in donkeys

5.1.4.1: Stomach

Perforation or rupture of the gastrointestinal tract leading to peritonitis has been documented

as consequence of Gasterophilus infestation (Van der Kolk et al., 1986). Ulcers frequently

occur in the non-glandular mucosa of the stomach which doesn’t have adequate protection

against the detrimental effect of stomach acids (Andrews et al., 2005). In our study

widespread areas of erosion and ulcers were seen after dislodging the various species of

Gasterophilus species in the non-glandular region. The bots significantly affected the non-

glandular region. This was also observed by Al-Mokaddem et al. (2015) in Egypt where they

found tiny superficial erosions upon removal of the larvae. Microscopic vacuolar

degeneration of the squamous cells, thickened keratinized layer of the stomach lining were

observed in this study. According to Al-Mokaddem et al. (2015) the microscopic examination

showed the attachment site appearing like deep, pitted ulcer with exposed lamina propria,

granulation tissue formation and neutrophilic infiltration. Additionally, they found;

hyperkeratosis, acanthosis, vacuolar degeneration of stratified squamous cells, gastritis,

57
erosions, ulcerations, scarring, hyperactivity of mucus glands, periglandular fibroplasia and

parasitic granulomes with infestation by Gasterophilus larvae.

This current study revealed a similar pathological pattern whereby the mucosal epithelium

was thickened and there was loss of tissue architecture. Additionally there was disruption of

the keratinized layer of the non-glandular stomach. The keratin covering the surface

epithelium however had a yellow tinge and this could be due to bile as the donkeys were

starved in the lairages prior slaughter. Empty white spaces were visible in the lamina propria

(vacuolar degeneration).

5.1.4.2: Intestines

The mucosa of the intestines had nodules on gross examination and was covered with mucoid

exudate. On incision, a worm was retrieved in most cases. Microscopically, there was dilation

of the submucosal blood vessels with four regular structures suspected to be larva of

Strongyle spp encapsulated by granulation tissue. This was also accompanied by a high

cellular infiltration characterized by eosinophils, neutrophils and macrophages within the

sub-mucosa. This is similar to observations by Lewa et al. (2000) who found out that both

the large and small strongyles cause enteritis with the mucosal surface having parasitic

nodules. Microscopically, the mucosa and the sub-mucosa had cellular infiltrations.

Penetration of the third stage larvae of Strongylus vulgaris causes small hemorrhages through

the intestinal wall. The small nodules could be due to Strongylus edentatus owing to its life

cycle in the liver and also the intestines. A similar study done by (Lewa, 1998), showed that

the 3rd stage larvae of Strongylus edentatus was partially the cause of the nodules observed

in the small intestines. The larvae go through the wall of the intestines and reach the liver via

portal veins where they grow to 4th stage which migrate within the liver and also contribute

to necrosis and inflammation. The nodules in this current study were majorly affecting the

ceca. Cyathostomes are obtained by horses within pasture which upon ingestion, primarily go

58
through a period of arrested development in the mucosa, submucosa, or both, of the large

intestine and cecum as well. Their emergence has been associated with ill effects including

diarrhea, emaciation and instigates an inflammatory protein-losing enteropathy and changes

in intestinal motility (Love et al., 1999). As the cyathostomes are emerging they cause

rupture of the muscularis mucosae, local eosinophilia, edema and infiltration by macrophages

and neutrophils as well (Soulsby, 1982). One of the life threatening parasites is the small

strongyles that cause larval cyathostominosis and usually encyst into the large intestine and

their larvae can initiate severe damage in the lining of the intestine (Oryan et al., 2015) .The

findings in our study could imply that there was a possibility of mixed infection by both the

small and large strongyles as both categories of helminths were identified. Oryan et al. in

(2015) established that the lesions in the caecum of a six year old working donkey were

associated with non-suppurative enteritis with characteristic infiltration of eosinophils,

plasma cells, lymphocytes and macrophages in the intestinal mucosa, submucosa and lamina

propria. The parasites identified on parasitological examination revealed two species of

cyathostomes that included Cylicocyclus elongatus and Cyathostomum pathratum.

Stress, interconnected with work and inadequate nutrition frequently results in a loss of

condition and cyathostomid nematodes may then bring about the clinical disease (Krecek and

Guthrie, 1999).

Lesions associated with Anaplocephala perfoliata observed in this study included; grossly

mucosal erosions were observed, lymphocytic cellular infiltration, localized area that was

surrounded by fibrous connective tissue and hyperplastic goblet cells. Sangioni et al. (2000)

reported macroscopic and microscopic lesions linked to Anaplocephala perfoliata which

included mucosal erosion, ulcerations, edema accompanied by eosinophilic and mononuclear

cell infiltration in the mucosa and the basal membrane of the caecum and colon. It also causes

59
hypertrophy of the intestinal wall at the site of attachment and can as well cause perforation

leading to colic (Soulsby, 1982).

5.1.4.3: Liver

In this study, tiny focal necrotic lesions and hepatomegally were seen in liver samples

grossly. On microscopic examination chronic hepatitis was observed. Hepatic abscessation

was also observed in donkeys that had an infestation with Parascaris equorum. The same

animal was positive for Strongylus vulgaris. Similar findings were found in livers of donkeys

examined by Lewa et al. (2000) who observed traumatic hepatitis, bile duct thickening and

abscess formation which were linked to Strongylus species larvae. Hepatic lesions have also

been associated with migrating larval stages of Parascaris equorum (Maxie et al., 2016).

Hepatic hemorrhagic nodules were seen in livers of eight donkeys. In our study, hemorrhagic

nodules due to Strongylus edentatus was common. Strongylus edentatus has been associated

with hepatic hemorrhagic nodules in the liver associated with the 4th stage larvae 3-5 months

post infection (Soulsby, 1982). Migration of strongylus equinus through the liver also causes

formation of nodules and formation of fibrous tissue. An acute reaction can also result in the

liver due Strongylus edentatus infection (Johnstone, 1998).

60
5.2: CONCLUSION AND RECOMMENDATIONS

5.2.1 CONCLUSION

1. Donkeys in Kenya are affected by a variety of gastrointestinal helminth parasites and

hemoparasite species.

2. The prevalence of gastrointestinal parasites is high in donkeys (85.5%) in Kenya and

interferes with their work performance as a majority of them (89%) had a poor body

condition.

3. These parasites cause significant pathological lesions in the gastrointestinal tract and liver.

4. These lesions interfere with the normal functioning of the body resulting in lowered meat

and skin quality which are a prime cut in the donkey value chain enterprise.

5. Risk factors such as poor body condition scores predispose the donkeys to gastrointestinal

parasites

5.2.2 RECOMMENDATIONS

1. Control measures for gastrointestinal parasites and hemoparasites in donkeys is

recommended so as to ensure the wellbeing of these animals.

2. Further studies on the effects of the hemoparasites on the performance of donkeys in

Kenya should be carried out in areas where the donkeys are raised. Such study should involve

larger sample sizes and sensitive diagnostic techniques should be applied.

3. Further study on zoonotic parasites in donkeys is recommended.

4. Further hematological studies in donkeys should be done.

61
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72
 7.0: APPENDICES

Appendix 1: Field Data Collection Sheet

Name of Abattoir: Source of animals:

Average no. of animals

slaughtered/day:

Animal Id Approximate Sex Comments(weakness, approximate body

Age condition score, pregnancy status, gross

lesions observed)

73
Appendix 2: Body Condition Scoring System for Donkeys

CONDITION NECK AND WITHERS RIBS BACK HINDQUA

SHOULDER AND AND R-
S BELLY LOINS TERS
1. Neck thin, all Dorsal spine Ribs can be Backbone Hip bones
POOR bones easily of withers seen from a prominent, visible and
felt. Neck Prominent distance and can feel dor- felt easily
meets shoul- and easily felt felt with ease. sal and trans- (hock and pin
der abruptly, Belly tucked verse proc- bones). Little
Shoulder up. esses easily muscle cover.
bones felt May be cavity
easily, angu- under tail.
Lar
2. Some muscle Some cover Ribs not visi- Dorsal and Poor muscle
MODERATE Development over dorsal ble but can be transverse cover on
Overlying withers, felt with ease processes felt hindquarters,
bones. Slight spinous proc- with light hip bones felt
step where esses felt but pressure. with ease.
neck meets not prominent Poor muscle
Shoulders development
either side
midline.
3. Good muscle Good cover of Ribs just cov- Cannot feel Good muscle
IDEAL development, muscle/fat ered by light individual cover in hind-
bones felt un- over dorsal layer of fat/ spinous or quarters, hip
der light cover spinous proc- muscle, ribs transverse Bones
of muscle/fat. esses withers can be felt with processes. rounded in
Neck flows flow smoothly light pressure. Muscle devel- appearance,
smoothly into into back Belly firm with opment either can be felt
shoulder, good muscle side of mid- with light
which is tone and flat- line is good Pressure
Rounded tish outline.
4. Neck thick, Withers Ribs dorsally Can only feel Hindquarters
FAT crest hard, broad, bones only felt with dorsal and rounded,
shoulder cov- felt with firm firm pressure, transverse bones felt
ered in even Pressure ventral ribs processes only with firm
fat layer may be felt with firm pressure. Fat
pres-
more easily. sure. Slight Deposits
Belly overde- crease along evenly placed
veloped. midline

5. Neck thick, Withers Large, often Back broad, Cannot feel

OBESE crest bulging broad, unable uneven fat de- unable to feel hip bones, fat
with fat and to feel bones posits covering spinous or may over-
may fall to dorsal and transverse hang either
one side. possibly ven- processes. side of tail

74
 Shoulder tral aspect of Deep crease head, fat of-
rounded and ribs. Ribs not along midline ten uneven
bulging with palpable. Belly bulging fat and bulging
fat. pendulous in either side.
depth and
width.

Copyright NEWC 2005

Appendix 3: Analyzed Results

Independent Samples t-Test( Strongyle egg count between adults and juveniles)

Levene's t-test for Equality of Means

Test for
Equality of
Variances
F Sig. T df Sig. Mean Std. Error 95%
(2- Difference Difference Confidence
tailed) Interval of the
Difference
Lower Upper
Equal
-
variances 1.644 .201 1.075 280 .283 63.907 59.449 180.930
53.117
assumed
Strongyle
Equal
eggs
variances -
1.396 19.928 .178 63.907 45.780 159.425
not 31.611
assumed
No significant difference in strongyle egg count between adults and juveniles, p>0.05

Independent Samples t-Test (Strongyle egg count between males and females)

Levene's t-test for Equality of Means

Test for
Equality
of
Variances

75
 F Sig. T Df Sig. Mean Std. Error 95% Confidence
(2- Difference Difference Interval of the
tailed) Difference
Lower Upper
Equal
- -
variances .256 .614 280 .529 -18.202 28.872 38.631
.630 75.035
assumed
Stronyle
Equal
eggs
variances - -
246.822 .527 -18.202 28.731 38.387
not .634 74.791
assumed
No significant difference in strongyle eggs shed between males and females p>0.05

Group Statistics: Body condition score versus strongyle egg count

Body condition Std. Std. Error

score N Mean Deviation Mean

Strongyle egg 1 250 154.0000 247.07932 15.62667

count
2 32 .0000 .00000 .00000

Independent Samples Test for strongyle egg counts between the Poor and moderate
body condition scores
Levene's
Test for
Equality
of
Variances t-test for Equality of Means
Sig. Std. 95% Confidence
(2- Mean Error Interval of the
Sig taile Differen Differen Difference
F . T Df d) ce ce Lower Upper

76
Strongy Equal
le egg varianc
28.81 .00 3.52 154.000 43.7458 67.8875 240.112
count es 280 .001
2 0 0 00 3 4 46
assume
d
Equal
varianc
9.85 249.0 154.000 15.6266 123.222 184.777
es not .000
5 00 00 7 70 30
assume
d
Significant difference in strongyle eggs shed between the two body condition scores

Chi-Square Tests(Association between body condition scores and prevalence of

helminths)

Asymp. Sig. Exact Sig. (2- Exact Sig.

Value df (2-sided) sided) (1-sided)
Pearson Chi-Square .121a 1 .728
Continuity Correctionb .007 1 .935
Likelihood Ratio .125 1 .723
Fisher's Exact Test 1.000 .488
Linear-by-Linear
.120 1 .729
Association
N of Valid Cases 282

77