INTERNSHIP FINAL REPORT

WITH TOPIC OF INTEREST

SEROPREVALENCE AND ASSOCIATED RISK FACTOR OF

TRANSMISSIBLE GASTROENTERITIS IN SWINE OF BHAKTAPUR
AND KAVRE DISTRICT

SUBMITTED BY:
NAME: SUSHIL ACHARYA
ROLL NO: 37
LEVEL: B.V.SC AND AH (INTERNSHIP)
IAAS, PAKLIHAWA, TU

SUBMITTED TO:
TRIBHUVAN UNIVERSITY
INSTITUTE OF AGRICULTURE AND ANIMAL SCIENCE
PAKLIHAWA CAMPUS
RUPENDEHI, NEPAL

IN PARTIAL FULFILLMENT FOR THE REQUIREMENT

FOR THE DEGREE OF
BACHELORS OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY B.V.SC
AND A.H
OCTOBER 2018
Declaration by student

I, Sushil Acharya, hereby declare that the work presented herein is original
and the work done by me and has not been published or submitted elsewhere
for the requirement of a bachelor’s degree program. Any literature date or
work done by other and cited within this thesis has given due
acknowledgement and listed in the reference section.

Name: Sushil Acharya

Institution: IAAS, Paklihawa Campus
Tribhuvan University
Certificate

It is tocertified that the thesis entitled “STUDY ON SEROPREVALENCE AND

ASSOCIATED RISK FACTOR OF TRANSMISSIBLE GASTROENTERITIS IN PIG
AT BHAKTAPUR AND KAVREPLANCHOK DISTRICT” prepared and submitted by
Mr. Sushil Acharya in partial fulfillment of the requirement for the degree of Bachelor in
Veterinary Science and Animal Husbandry (B.V.Sc and A.H), Tribhuvan university,
Institute of Agriculture and Animal Science, Paklihawa campus, is hereby accepted.

Dr. Awadesh Jha (M.V.Sc)

Site Advisor
Senior veterinary officer
Central Veterinary hospital
Date:

Assoc.Prof.Shambhu Shah (Ph.D) Prof. Keshav Raj Adhikari (Ph.D)

Internship coordinator Chairperson
Tribhuvan University Internship Advisory Committee
IAAS, Paklihawa Dean, IAAS,
Paklihawa, Rupendehi TU
Date: Date:
ACKNOWLEDGEMENT

I would like to express my sincere gratitude and appreciation to Dr. Awadesh

Jha. A senior veterinary officer at central veterinary hospital, Tripureshwor,
Kathmandu as my internship advisor for his valuable guidance, continuous
encouragement, advice support, meticulous supervision, constructive criticism
and detailed editing till the completion and final preparation of the
manuscript.
I am grateful to Asso. Prof. Dr. Shambhu shah, internship coordinator and
Prof. Dr. Dinesh Kumar Singh for their valuable suggestions, continuous
encouragement, incredible aid and detail editing to refine the manuscript to
this degree.
I respectfully acknowledge and express my profound thanks to Dr. Swyam
Prakash Shrestha, Dr. Dhojraj Khanal and Dr. Riddhi Shrestha for their
incredible help and guidance during research period. I express my sincere
thanks to Miss Eliza Ranjit and all the staffs of AHRD, NARC who
facilitated, critiqued and assisted me during this research.
Regards are with my batch mates Anil Subedi, Sonu Adhikari, Sangam
Acharya, Saraswati and all the helping hands for their affection and
cooperation during the study period.
Last but not least, I feel my proud privilege to mention the feeling of
gratefulness to my parents, brother, sister who persistently encouraged me
throughout my B.V.Sc and A.H course and research job. Words of thanks are
insufficient for their ultimate understanding patience and sacrifice.

September 2018 Sushil Acharya

 TABLE OF CONTENT
TITLE PAGE
ACKNOWLEDGEMENT i
TABLE OF CONTENT ii
LIST OF TABLE iii
LIST OF FIGURES iv
LIST OF APPENDICES v
ACRONYM AND ABBREVIATION vi

PART 1
1. INTRODUCTION TO INTERNSHIP PROGRAM 1
1.1 BACKGROUND INFORMATION
1.2 OBJECTIVES
1.3 DURATION OF INTERNSHIP
1.4 NATURE OF WORK
2. PROFILE OF INSTITUTE OF AGRICULTURE AND ANIMAL SCIENCES 2
2.1INTRODUCTION TO IAAS
2.2VETERINARY PROGRAM AT IAAS
3. PROFILE OF THE SITE 3
3.1 COUNTRY PROFILE
3.2 PROFILE OF THE LOCAL SITE
4. INTRODUCTION TO ACTIVITIES AT DIFFERENT SITES 5
4.1 CENTRAL VETERINARY HOSPITAL
4.2 NEPAL AGRICULTURE RESEARCH COUNCIL
4.3KATHMANDU ANIMAL TREATMENT (KAT) CENTRE
4.4 NEPAL CAVALRY
5 OTHER ACTIVITIES DURING INTERNSHIP PERIOD 7
6 CONCLUSION

PART 2
CASE STUDIES
CASE1: CANINE DISTEMPER 10
CASE2: INFECTIOUS BURSAL DISEASE IN POULTRY 16
CASE3: PARVOVIRAL ENTERITIS IN A DOG 20
 TABLE OF CONTENT
PART 3

ABSTRACT 27
1. INTRODUCTION
BACKGROUND AND SIGNIFICANCE OF STUDY 29
OBJECTIVE OF THE STUDY
General objective
Specific objective
2. TIME AND PLACE OF STUDY 29
3. LITERATURE REVIEW 29
4. METHODOLOGY 30
4.1 STUDY AREA, DISTRICT PROFILE AND POPULATION
4.2 MATERIALS AND METHOD
4.3 SAMPLE SIZE AND SAMPLING PROCEDURE
4.4 DATA COLLECTION AND ANALYSIS
4.5 VALIDITY OF TEST
5. RESULTS 33
6. DISCUISSION 36
7. CONCLUSION AND RECOMMENDATION 36
8. REFERENCES 38
9. APPENDICES 40
10. GLIMPSE OF INTERNSHIP ACTIVITIES 42
LIST OF TABLE page
I. Haematological value of disease dog 21
II. Differential diagnosis of canine parvovirus enteritis 23
III. Sexwise prevalence of TGE 34
IV. Agewise prevalence of TGE
34
V. District wise prevalence of TGE
35
VI. Overall prevalence of TGE
35
LIST OF FIGURES page
1. Canine distemper infected dog 10
2. Pathogenesis of canine distemper 11
3. Clinical signs of CDV infected dog 11
4. Diagnosis of CDV by rapid test kit 12
5. IBD lesions 16
6. IBD kit 16
7. Parvovirus infected dog 20
8. CPV rapid test kit 20
9. Map of Nepal showing study area and district profile 30
10. A value obtained in ELISA analyser 33
11. Sex wise prevalence of TGE 33
12. Age wise prevalence of TGE 34
13. District wise prevalence of TGE 35
14. Overall prevalence of TGE 36
15. Glimse of research activities 42
LIST OF APPENDICES

Appendix page
1. Questionnaire survey 40
2. Elisa protocol 41
3. Glimpse of research activities 42
ACRONYMS AND ABBREVIATIONS

B.V.Sc&A.H: Bachelor in Veterinary Science and Animal Husbandry

IAAS : Institute of Agriculture and Animal science
CVH : Central Veterinary Hospital
& : And
TGE : Transmissible Gastroenteritis
ELISA : Enzyme linked immunosorbent assay
% : percentage
S : significant
N/S : non-significant
+ve : positive
-ve : Negative
CDV : Canine Distemper virus
IBD : Infectious Bursal Disease
CPV : Canine parvovirus
AHRD : Animal Health Research Division
NARC : Nepal Agriculture Research council
OD : Optical Density
DLSO : District livestock service office
SPSS : Statistical Package for the Social Sciences
MS EXCEL : Microsoft excel
℃ : Degree Celsius
Ml : Milliliter
µl : Microliter
PART 1
INTRODUCTION TO INTERNSHIP PROGRAM
1.1. Background information
Internship program is a pre-service fieldwork training organized for students of Bachelor
In Veterinary Science and Animal Husbandry (B.V.Sc. & A.H.) as an integral part of
Degree program at the Institute of Agriculture and Animal Science (IAAS), Tribhuvan
University. B.V.Sc. & A.H. is an undergraduate course of five years which includes ten
Semesters with nine academic semesters and the last semester as an Internship program.
The program starts just after completion of ninth semester. The duration of internship
Program is of six months and the student spends this period at veterinary hospitals, Disease
diagnostic laboratories and other relevant institutions and organizations of Veterinary
importance where they undergo skill development training. This helps to perform research,
understand the field problems solve those problems and to carry out the Field work.
The internship sites were selected on the basis of the facilities available like veterinary
Hospital, disease diagnostic laboratories, and capable manpower to guide the intern. The
Internship advisory committee decided the sites in consultation with government
Organization, private organizations and other concerned authorities. Number of students
was allocated to different sites according to the facilities available and the availability of
Veterinary officer.
1.2. Objectives
General Objective
• To offer pre-service field training to undergraduate students of B. V. Sc. & A.H.In the
field of veterinary science and animal husbandry in veterinary hospitals, disease diagnostic
laboratories and livestock farms.
Specific objectives
• To offer pre-service field training to undergraduate students of B. V. Sc & A.H.
• To understand the situation of veterinary practices at different levels.
• To provide opportunity for practical training to students through variety of work
experience
• To develop confidence among the students to utilize their knowledge and skill to
Solve clinical problems.
• To familiarize students with the administrative procedure at the allocated site.

DURATION OF INTERNSHIP
Duration of internship program was six months, and it started after completion of 9th
semester examination at IAAS. The students were assigned to start the work from 29th
Chaitra, 2074 with an orientation of internship program at IAAS organized by internship
advisory committee on the 31 of Chaitra. The students were assigned to start their work
from 7th Ashwin at their respective sites.

NATURE OF WORK
This research is carried out on animal health division, NARC khumaltar under the
guidance of Dr.Awadesh Jha, a senior veterinary officer at central veterinary hospital,
tripureshwor, topic kathmandu. The research was carried out in the topic
“SEROPREVALENCE OF TRANSMISSIBLE GASTROENTERITIS IN PIG AT
BHAKTAPUR AND KABHREPLANCHOK DISTRICT”. During that time participation
in various activities organized by the department of livestock services was also done.
Besides regularly attending activities at internship site, visit to other internship sites within
and outside the kathmandu valley was also done. Last one month of work was allocated for
report writing and submission. The report contains three parts: part i- Includes work done
during the internship period; part ii- includes case studies and part iii- includes research
work done on the specific topic of interest.

2. PROFILE OF THE INSTITUTE OF AGRICULTURE AND ANIMAL

SCIENCE
2.1. Introduction to IAAS
Institute of Agriculture and Animal Science (IAAS) is the only agricultural institute under
Tribhuvan University in Nepal for providing higher-level education and training in
agriculture and animal sciences. The IAAS traces its origin to a school of agriculture,
established in 1957 in Kathmandu to train Junior Technical Assistants (JTAs). In 1968, the
school was upgraded with I.Sc. Agriculture program to college of agriculture. In 1972, the
college was recognized as the Institute of Agriculture and Animal Science under Tribhuvan
University and moved to the present location at Rampur, Chitwan in 1974. Presently, the
institute has three campuses Rampur Campus at Rampur, Chitwan, Lamjung Campus at
Sundarbazar, Lamjung, and Paklihawa Campus at Paklihawa, Rupendehi. IAAS offers a
B.V.Sc. & A.H. (Bachelor of Veterinary Science and Animal Husbandry), B.Sc.
Agriculture (Bachelor of Science in Agriculture), M.V.Sc., M.Sc. Agriculture, M.Sc.
Animal Science and Ph. D programs at Rampur Campus.
2.2. Veterinary program at IAAS
In the beginning five and half years Bachelor of Veterinary Science andAnimal Husbandry
(B.V.Sc. & A.H.) program was started at IAAS in 1992. From 1992 to 1994, Condensed
and Conversion program of B.V.Sc. & A.H. was offered to the students of B.Sc. Animal
science at Kathmandu as well as at Rampur. The veterinary science program at IAAS has
made significant progress since its inception in 1995, including the graduation of its first
regular batch on 2000, the development of a 10 semester based B.V.Sc. & A.H. curriculum
and the acquisition of 21 faculty members. The institute has its own veterinary teaching
hospital, livestock farm and laboratories for the academic purpose of veterinary science.
The regular course of five and half years was in annual system but now the curriculum has
been updated and the course of veterinary science has been converted into semester system
and the duration of course has been reduced to five years from 2006. The curriculum has
been designed to integrate basic, production and management, practical and clinical
subjects to convey both extensive and intensive knowledge in veterinary clinical sciences,
animal breeding, livestock production and management.

3. PROFILE OF THE SITE

3.1 Country Profile
Nepal is one of the biodiversity richest countries in the world due to its unique
geographical and altitudinal variation. The elevation ranges from 60m above sea level to
the highest point on the earth, Mount Everest at 8,848m, all within a distance of 150km
resulting into climatic condition from subtropical to arctic. Nepal occupy only 0.03% of
the earth, is home to 2% of the flowering plants in the world, 8% population of birds of the
world that includes more than 848 species, 4% of the mammals on earth, 11 of the world's
15 families of butterflies (more than 500 species), 319 species of exotic orchids. Nepal is a
landlocked country situated between 26022' N to 30027' N latitudes and 8004' E to 88012'
E longitudes that covers a total land area of 147,181 sq. km. Livestock is crucial part of
agricultural production system of Nepal that contributes 16.6 % of the total GDP in the
form of milk, ghee, meat, skin, hair, wool, manures and draft power and make a substantial
contribution to household livelihoods of rural poor people. Poverty level is extremely high
and more than 80% of the population relies on the agriculture sector for employment and
income generation. However, livestock resource management is the basic problem in the
development of this crucial sector. As Nepal is endemic for different diseases of livestock,
they are the major problem in livestock, which cause big economic losses every year.
Besides this the high prevalence of zoonotic and infectious diseases poses constant threats
to human beings and other healthy herds too.

3.2 Profile of the Local Site

My internship site was Katmandu district, the capital city of Nepal and is the eldest
Metropolitan city of the country. It is located in the mid-hill region of the country at an
altitude of 1,300m above sea level and has a fertile alluvial soil covering an area of 600 sq.
km. Kathmandu valley, which includes three districts Kathmandu, Bhaktapur and Lalitpur,
experiences temperate type of climate with an average minimum temperature 9.7 0 C during
winters and average maximum temperature 20.30 C during summer. The average annual
rainfall is recorded to be 1200 mm. Mostly the monsoon sets in June and ends in
September. The relative humidity varies from a low of 42.9% in March to a high of
75.06% in July.

More than three million people inhabit the Kathmandu valley. The population of
Kathmandu is made up of people of different castes and creeds following different
religions and observing variety of cultural practices. Thus they all make up a composite
cultural society and enjoy wide range of cultural and festive ceremonies throughout the
year. Kathmandu has well facilities of transport, communication, electricity and other
infrastructures. The economy is also sound with flourishing trade, industry, business and
service activities along with a very strong agricultural base. Central veterinary and
livestock related offices, NGOs and INGOs working for the welfare of animals and
development of livestock sector and wildlife conservation offices are located at
Kathmandu. Beside this many commercial livestock and poultry farms as well as small-
scale livestock business are run at Kathmandu valley at different level.

4. INTRODUCTION TO ACTIVITIES AT DIFFERENT SITES

I visited different places that are associated with providence of veterinary services during
the period of internship program. Overall I got chances for wider exposure on veterinary
field in the capital city.

4.1. Central Veterinary Hospital (CVH)

The Central Veterinary Hospital, Tripureshwor was inaugurated on June 1970. Since then
it is strongly committed to providing quality veterinary services as a central level hospital
in the country. Currently there are 3 doctors working in the hospital. The hospital is well
facilitated with ultrasonography and x-ray machines. Also it has effective coordination
with Central Veterinary Laboratory for laboratory examinations and diagnosis. CVH
mainly provides following services to the livestock and pet owners.
 `

· Treatment of sick animals and birds.

· Surgical correction and gynecological interventions.

· Immunization of dogs and cats against Rabies and infectious diseases.

· Fecal Examination.

· Expert advice on livestock production and health management.

· Artificial Insemination of cattle and buffaloes.

I was benefitted by maximum clinical exposure and skill enhancement on handling surgical
and medical cases of canine, poultry, feline, swine, caprine and bovine species at CVH.
Canine cases hold the major share among the animal species brought at the hospital
followed by poultry, caprine and other large animals

4.2. Nepal Agriculture Research Council (NARC)

Nepal Agriculture Research Council, Khumaltar is an autonomous body at the national

level to conduct agriculture research activities for optimizing agricultural production and
productivity. It was established in 1991 AD under the authority of "Article No.9 of the
Nepal Agriculture Research Council Act 1991" with main objectives to develop
appropriate agro-technologies suitable to various agro-ecological zones. Its diversified
field of research includes crops as cereals, grains, legumes, oilseed, cash crops,
horticulture, and livestock research on swine, bovine, rabbit, fish, poultry and animal
health. The main objective of working at NARC was to get acquainted with new research
in health, management, feeding practices of farm animals and birds. The five main
divisions of Nepal Animal Science Research Institute under NARC are:
· Animal Health Division, Khumaltar
· Animal Breeding Division, Khumaltar
· Animal Nutrition Division, Khumaltar
· Fodder and Pasture Management Division, Khumaltar
· Fisheries Division, Godawari
4.3. Kathmandu Animal Treatment (KAT) Centre
The Kathmandu Animal Treatment Centre (KAT centre), Chapaligaun is a registered
non-profit charitable organization dedicated to the humane management of street dogs for
community benefit in Kathmandu district. Currently there are two veterinary doctors and
other staffs working in the organization. Activities of KAT centre for dog sterilization is
concentrated on one area of the city at a time. Trained staffs pick up a quota of female
dogs, which are examined by veterinary professionals. After a day of rest bitches are
vaccinated against rabies, spayed, dewormed and treated for any other problems. Each
dog is identified by an ear notch and is tattooed with an individual identification number.
The dogs also receive a red collar to indicate treatment. After recovery dogs are released
in their same location. Besides this KAT centre also catches the dogs that are severely
injured, infected or affected with diseases and treats them by keeping them in isolation
chamber. After recovery they are released to the location from where they were brought.
This organization also facilitates the adoption of healthy dogs. KAT has also been
running public education program regarding dogs and diseases of dogs. At KAT center I
learned the method of spaying by flank method, general medical intervention on diseased
stray dogs and general management of the organization.

4.4. Nepal Cavalry

The stud farm of Nepalese army at Singh durbar is the wing of Directorate of Livestock
Development and Animal Health, Nepal Army for the treatment and training of the
military horses. Currently it holds nearly 200 horses mainly of three different types, i.e.
heavy, used for coach; light and pony for riding. We were guided by Lieutenant Dr.
Nirvik Neupane. Horses are examined with the staff of the farm by horse-to-horse
approach. Major disease conditions noted in the farm-included lameness due to sand
crack, capped elbow, laminitis, chronic wounds, bruises, tumors, seasonal dermatitis and
colic. I was acquainted with disease prevalence and their correction approach in this
farm. Besides these management practices as the process of shoeing, method of casting
and restraining by lifting the limb, application of twitch and casting with rope by breast
knot or crisscross method, application of head belt and saddle, identification of horses by
branding, body color and markings were also learnt.

5. OTHER ACTIVITIES DURING INTERNSHIP PERIOD

Besides working at the Central Veterinary Laboratory and Central Veterinary Hospital I
was also involved in the following activities during my internship period:
 Attended rally on world veterinary day in pokhara organized by DLSO,
 kaski
 Attended World Rabies Day and World Animal Day Programs organized by
Directorate of Animal Health.
 Attended talk program on brucellosis in Tripureshwor Vet Complex,
Kathmandu
Site of veterinary importance visited during internship
• Central Veterinary Hospital (CVH), Tripureswor,kathmandu
• Central veterinary Laboratory (CVL), tripureswor kathmandu
• Animal medicle centre,kathmandu
• Regional Veterinary Lab, Pokhara
• DLSO (bhairahawa,pokhara,chitwan
• Nepal Army cavalary, singh durbar, kathmandu
• Dlso bhaktapur,kathmandu
• Avian lab ,chitwan
• Nepal Agriculture Research Council, Khumaltar

6. CONCLUSION
The internship period was fruitful in the sense that I got accustomed to the different
laboratory techniques involved in the disease diagnosis. I also got opportunities to be
exposed to different cases that were brought in the animal hospital. I also got many
opportunities to handle and treat animals in the hospital. I learned so many things and met
many people of different faculty and broaden my knowledge. I got the chance to broaden
my knowledge and practically implement it. However, I have felt the need of new and
sophisticated diagnostic approaches to strengthen the veterinary practices in coming
generations. During the short duration of internship, what I realized is that there is a vast
scope of veterinary sector in the country.
PART 2
Case study: Canine Distemper
INTRODUCTION
It is highly contagious disease of dog. Characterized by diphasic fever leukopenia and
gastrointestinal catarrhal and usually pneumonic and neurological complication. The virus
can also be found in wildlife such as foxes, wolves, coyotes, raccoons, skunks, mink and
ferrets and has been reported in lions, tigers, leopards and other wild cats as well as seals.
The virus may also cause the footpads to thicken and harden, leading to its nickname “hard
pad disease.”
Transmission
Puppies and dogs most often become infected through airborne exposure (through sneezing
or coughing) to the virus from an infected dog or wild animal. The virus can also be
transmitted by shared food and water bowls and equipment. Infected dogs can shed the
virus for months, and mother dogs can pass the virus through the placenta to their puppies.
Canine distemper also impacts wildlife populations, contact between wild animals and
domestic dogs can facilitate the spread of the virus.

HISTORY

Name of owner: Hemnath adhikari

Address: Teku, Kathmandu
Breed of dog: Labrador
Name: lucky
Age: 2.5 month
Sex: male
Vaccination: not vaccinated Fig1: CDV Infected dog

PATHOGENESIS

The virus initially replicates in the lymphatic tissue of the respiratory tract. A cell-
associated viremia results in infection of all lymphatic tissues, which is followed by
infection of respiratory, GI, and urogenital epithelium, as well as the CNS and optic
nerves. Disease follows virus replication in these tissues. The degree of viremia and extent
of viral spread to various tissues is moderated by the level of specific humoral immunity in
the host during the viremic period.
Fig 2: Pathogenesis of canine distemper
CLINICAL SIGNS

 Reduced appetite and watery discharge from eyes since 1 week

 Fever
 nasal discharge, coughing, lethargy, and vomiting was noticed.
 circling behavior was revealed.
 head tilt, salivation, seizures, and partial paralysis was noticed.
 Footpads thickened and hardened.

Lacrimation Lethargy Head tilt

Salivation Depression loss of appetite

Fig3: Clinical signs of CDV infected dog

DIAGNOSIS

Diagnosis was done by

I. Rapid test kit

Fig4: Diagnosis by rapid test kit

II. Clinical signs and symptons

 Head tilt
 Fever
 Loss of appetite
 Lethargy
 Salivation
 Lacrimation
 Hard pad
 Circling behavior
DIFFERENTIAL DIAGNOSIS

• Kennel cough
• Rabies
• listeriosis

TREATMENT
Initially for symptomatic treatment is recommended

• Neurobion tab.

Sig. ½ tab. B.i.d for 7 days

• Vit. C b.i.d for 15 days

• clavam

sig.3.5 ml b.i.d for 7days

• Ciplox eye drop

Sig. 2 drops t.i.d for 7 days

Since, prognosis was worse, it was then euthanized.

References:

chakrabarti, A 2011. A Textbook of preventive medicine. Kalyani publishers, India. 4th

edition . Pp232-238
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Krakowka S, Axthelm M K, Gorham J R 1987 Effects of induced thrombocytopenia on
viral invasion of the central nervous system in canine distemper virus infection. Journal of
Comparative Pathology 97: 441–450
Haines D M, Martin C L, Chelack B J, Sargent R A, Outerbridge C A, Clark E G 1999
Immunohistochemical detection of canine distemper virus in haired skin, nasal mucosa,
and footpad epithelium: a method for antimortem diagnosis of infection. Journal of
Veterinary Diagnostic Investigation 11: 396–399
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canine pathogens among free-ranging jackals in Kenya. Journal of Wildlife Diseases 30: 4
486–491
Alldinger S, Wunschmann A, Baumgartner W, Voss C, Kremmer E 1996 Up-regulation of
major histocompatibility complex class II antigen expression in the central nervous system
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(Berlin) 92: 3 273–280
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1275 .

Case2: Infectious bursal disease in poultry

 Case history and description:
Sample of cob 500 broiler of age 4 weeks from the flock size of 2500 was presented on the Nation
avian laboratory, chitwan, Nepal. History of decreased feed intake for last 3 days, depression, diarrhe
morbidity rate 7 and mortality rate was 3 birds/day. Birds was reported with soiled (pasty) vent a
ruffled feathers. Lesion such as enlarged bursa and hemorrhagic bursa , cheesy mass in bursa, urates in
ureters and enlarged kidney was seen Haemorhage on thigh muscle

Fig5: IBD lesions

Tentative diagnosis
 Infectious bursal disease (IBD) or gumboro disease.
 Bursal swab was taken to check with rapid test kit.

Fig6: Diagnosis of IBD by rapid test kit

Confirmation
 Based on history, clinical findings, postmoretem lesions and result from IBDV rapid test kit it w
confirmed as infectious bursal disease (IBD).
Treatment
1. Gumbovit pv.
Sig: 1 gm/2lit of DW for 3 days
2.Brotone liq.
Sig:7ml/100 birds for 3-4 days
3.immunocare liq
Sig: 15ml/100 birs od for 3-4 days
4.Neodox forte pv.
Sig: 1 gm/2 lit of DW for 3 days

DISCUSSION
Infectious bursal disease( also known as IBD, Gumboro Disease, infectious
Bursitis and infectious Avian Nephrosis) is a highly contagious disease of young chickens caused
infectious bursal disease virus (IBDV), characterized by immunosuppression and mortality generally a
to 6 weeks of age (caston et al.,2008).it is a widely contagious disease occurring in brooding and growi
stage of chickens causing significant morbidity and low mortality rate . it is characterized by init
enlargement of bursa of fabricus followed by atrophy ( chakrabati,1993).

Clinical signs
Disease may appear suddenly and morbidity typically reaches 100 %. In the acute form birds
prostrated, debilated and dehydrated. They produce a watery diarrhea and may have swollen faec
stained vent. Recumbent and ruffled feathers are also the typical signs.
Lesions
Necropsy examination will usually show changes in the bursa of fabricus such
swelling,oedema,haemorrhage may also be seen in the skeletal muscle, intestines, kidneys and spleen.
Diagnosis
Based on flock history, clinical signs and postmortem ( necropsy) examinations. Definitive diagnosis c
only be achieved by the specific detection and /or isolation and characterization of IBD
Immunofluorescence or immunohistochemistry tests, based on anti-IBDV labeled antibodies are use
for the specific detection of IBDV in infected tissues . Real time PCR, ELISA can also be used
diagnosis.
Treatment and control:
Breeder flocks must be immunized against IBD so that they would transfer protective antibodies to th
progenies, such as broiler and pullet chicks. Biosecurity with adequate restriction to farm visit a
distancing from others flocks is suggested

Refrrences :
 Caston et al.2008.”Infectious Bursal Disease Virus (IBDV)” Segmented Double
stranded RNA virus : structure and molecular Biology. Caister Academic press.
ISBN 9798-1-904455-21-9. Accesed on 5 jun,2018 from

http://en.wikipedia.org/wiki/infectious_bursal-disease
 Chakrabrati,A.1993.A Textbook of preventive veterinary medicine. Pp 735-739
 Chauhan,H.V.S. and S.Roy.2008. poultry disease diagnosis and treatment. Third
edition, new age international publishers.
 Dhakal,I.P.2002.commom poultry disease and their management in Nepal. The
Bluecross. Annual Bulletin. NVSA 5: 3-9.
 Jordan,F.T.W. and M. Pattison. 1998.poultry diseases.fourth edition. W.B. saunders
company ltd.

Case3:
PARVO-VIRAL ENTERITIS IN A DOG

Case history and observation

A puppy name lucky of 2 month was presented in the Animal medical centre ,Kathmandu.
On taking history it was found that dog was not vaccinated and clinical sign such as
frequent vomition ( foamy gel like), whitish loose diarrhea, and lethargic was reported.
On clinical examination mucous membrane was pale, capillary refill time was 2 second,
respiration rate and heart rate was normal, slight fever with temperature 103°f ,and
intestinal motility was increased.
 Figure7: Diseased dog
Laboratory examination
Test with parvovirus antigen kit

Fig8: parvo virus rapid test kit

1. Rectal swab was taken and mixed in a specimen tube containing 1 ml assay diluents
buffer
2. 4 drops of sample from specimen tube were added into the sample well of the kit
3. Purple colour moved across the result window in the centre of thest6 kit
4. 4. The presence of two lines on the cpv Ag test area indicates that the case was
positive.
Haematological values:
Table1 : Haematological val;ues of the diseased dog
Parameters Obtained value* Refrence value**
Haemoglobin 9.3g/dl↓ 11.9-18.9g/dl
PCV 32%↓ 35-57%
Total leucocytes count 4.4*103/µl ↓ 5.0-14.1* 103 µl
Neutrophil 55%↓ 58-85%
Lymphocytes 41.6%↑ 8-21%
Monocytes 2.8% 2-10%
Basophils 0.00% 0-1%
Albumin 2.6 g/dl 2.3-3.1 g/dl
Bun 34.2 mg/dl ↑ 8-28 mg/dl
 Creatinine 1.4 mg/dl 0.5-1.7 mg/dl
*Animal medical centre ( AMC)
** merck veterinary manual, 2018 ( Accesed online )

Diagnosis :
Canine parvo virus infection based on history , clinical signs and symptoms, test kit and
laboratory tests.
Treatment :
 Ringer’ lactate –
Sig: 100 ml slow I/v BID for 7 days
 Inj.Aciloc ( antacid)
Sig: 0.3 ml s/c BID for 7 days
 Inj.onden (Anti- emetic)
Sig: 0.5 mg/kg bw I/V BID for 7 days
 Inj. Veticef- 0.5 gm ( cephalosporonin)
Sig : 1 ml slow I/V BID for 7 days
Prognosis: Recovered

DISCUSSION
Introduction
 canine parvovirus ( CPV) is highly contagious and relatively common
cause of acute, infectious GI illeness in young dogs.
 Non-enveloped, single-stranded DNA virus
 The disease is hoghly contagious and is spread from dog to dog by direct
orindirect contact with their feces.
 Young (6 week to 6 month) ,unvaccinated or incompletely vaccinated dogs
are most susceptible.
 Puppies born to a dam with CPV antibodies are protected from infection
for the first weeks of life sufficient colostrum is ingested.

.Pathogenesis:
Clinical findings:
Enteric form : ( chakrabarti,2007)
 Rise of temperature but decreases in advanced
 stages of diarrhea and vomition.
 Inappetance, refusal to food, vomition and diarrhea.
 Diarrheic stool may contain blood.
 Vomitus may contain yellow froathy material.
Cardiac form(chakrabrati,2007)
 Heart muscles are damaged
 Pulmonary edema, cyanosis
 Myocardial scaring, progressive cardiac insufficiency( sellen,2007)
Diagnosis:
 Signs and symptoms
  Virus isolation
 HA/HI
 ELISA(CITE test) ( poster& smith, 2011)
 Hematology
 Abdominal radiographic ( foster & smith, 2011)

Differential diagnosis :
Table 2 : differential diagnosis of canine parvovirus enteritis
Enteritis : Differentiating characters:
1.CD Diphasic fever , Neurologic signs)
2.ICH Corneal edema, cough, jaundice
3.corona virus Vomition not seen
4.salmonella infection mucus in stool
5.campylobacteteriosis Bacterium isolation
6.HGE No fever and no low WBC count
7.poisonings History
8.Hookworm infection fecal test shows no evidence

Myocarditis
1.CD
2.ICH
3.canine herpes virus infection yellow/ green feces, seizures, blindness
4.streptococcus infection Bacterial isolation
5.congenital heart anomalies

Treatment :
 Only symptomatic and supportive treatment is suggested.
 Fluid therapy
 Systemic antibiotics- Amyglycoside and beta- lactam antibiotics, fluoroquinolones
( contraindicated in young puppies)
ADVICE: Nothing but oral until diarrhea and vomition persist ( sellen, 2001)
Control and prophylaxis:
 Attenuated live CPV 2 vaccine
 Active immunity: vaccination ( primary at 6-9 weeks of age: secondary at 12-14
weeks of age and then booster dose annually) ( sellen,2001)
 Once recovered lifelong immunity.
References :
 Foster and smith. 2011. Parvovirus: serious diarrhea in puppies & dogs. Veterinary &
aquatic services department, foster & smith, inc- 2253 Air park road,
Rhinelander, Wisconsin, 54501.
 Goddard, A. and A.l. leisewitz. 2010. Canine parvovirus. Journal of veterinary clinical
small animal 40 : 1041-1053.
 Prittie, j. 2004. Canine parvoviral enteritis: A review of diagnosis, management, and
prevention. Journal of veterinary Emergency and critical care 14 ( 3) : 167-
176 .
 Parthiban, s.p. pothiappan and H.k. Mukhopadhya. 2016 . hematological and
Therapeutic Aspects of canine parvovirus infection in Non- descript pups.
Indian veterinary journal 93 ( 21):35-37
 The merck veterinary manual, 2012. Canine parvovirus
http:/www.merckvetmanual.com/mvm/index.jsp?cfile=htm/bc/23301.htm.
(accesed on 1st june 2018).
 PART 3

SEROPREVALENCE AND ASSOCIATED RISK FACTORS OF

TRANSMISSIBLE GASTROENTERITIS IN SWINE OF BHAKTAPUR AND
KAVRE DISTRICT OF NEPAL
Acharya. S1 and Jha. A.2
1
Paklihawa Campus, Institute of Agriculture and Animal Science, Tribhuvan University
2
Central Veterinary Hospital, Tripureshwor, Kathmandu

ABSTRACT
Objective: To find the Seroprevalence and associated risk factors of transmissible
gastroenteritis in swine of Bhaktapur and Kavre district of Nepal.
Study Design: Cross-sectional study
Methodology: 92 samples (50 samples from Bhaktapur and 42 sample from Kavre district)
of different farm, were selected randomly. A volume of 5 ml of blood was collected from
each selected swine, serum was separated using centrifuge and stored at -20 0 Celsius. The
ELISA test was conducted to detect the anti- TGE antibody using kit manufactured by
NOVATEINBIO, as per manufacturer’s PROTOCOLS. The test titre was determined
using serial dilution technique.
Result: Out of the 92 serum sample tested for the presence of antibody against TGE,
90samples (98.9%) were found to be positive. 43males(100%) were positive out of 43
male swine and 47 females (97.91%) were positive out of 48 female swine, 1 (0.0108%)
well was kept blank as manufacturer’s instruction . This variation between male and
female was not found to be significant at 95 percent confidence interval. Among the total
swine samples 34 were (< 6 months) in which 33 were positive and 57 were (> 6 months)
in which 57 were positive. But this difference between age and sex was not significant at 5
percent level of significance (p>0.05).
Conclusion and Recommendations: The Seroprevalence of TGE was (98.90) % in the
Bhaktapur and Kavre district of Nepal. Presence of antibodies in serum against TGE in the
swine population of the country clearly indicates that swine surveyed in this districts have
already been exposed and the disease should be given adequate attention in Nepal. Further
researches are essential to determine the economic losses due to TGE and to develop
control strategies against this disease in the country.
Key Words: Seroprevalence, Transmissible gastroenteritis, Risk factor, swine, Nepal.

Introduction:

Porcine transmissible gastroenteritis (TGE) is a disease in pigs caused by a coronavirus

belonging into the Coronaviridae family of the order Nidovirales. Coronaviruses are
enveloped and the genetic information is coded by a copy of a single stranded RNA
genome of positive polarity, so far known as the largest such stable RNA genome of
animal viruses, ranging in size between 26.4 – 31.7 kilobases (kb.it spreads rapidly to pigs
of all ages and causes vomiting, watery diarrhoea, dehydration, and death, particularly in
pigs under 2 weeks of age. The hemagglutinating activity of transmissible gastroenteritis
virus (TGEV), an enteric porcine coronavirus, was analyzed and found to be dependent on
the presence of alpha-2,3-linked sialic acid on the erythrocyte surface(Saif LJ, Wesley RD)
,but it differs from Porcine respiratory coronavirus (PRCV) in the fact that it is
enteropathogenic,  (Rasschaert D, Duarte M, Laude H). TGEV multiplies in and damages
the enterocytes lining the small intestine, producing villous atrophy and enteritis. The virus
is mostly isolated from the intestinal tract and from faeces. By contrast, PRCV is mostly
isolated from the upper respiratory tract, trachea, tonsils or the lungs. (D. L. Hank Harris,
1995)

The genus coronavirus comprises three groups, or most recently referred to as subgenera,
namely Alpha-, Beta and Gamma-coronavirus. Coronaviruses have some rather prominent
members that cause important diseases, both of human and veterinary interest such as,
Severe Acute Respiratory Syndrome Virus (SARS-CoV), Infectious Bronchitis Virus
(IBV), Feline Infectious Peritonitis Virus (FCoV), Porcine Hemagglutinating
Encephalomyelitis (PHEV) and Transmissible Gastroenteritis (TGEV), just to mention a
few of them. All in all the complete genome of some 26 coronaviruses have been described
to this date. But there are probably more out there and this number is in no doubt going to
change in 3 a not too distant future. After the SARS outbreak in 2002-2003 (Peiris et al.,
2004), research into coronaviruses has really picked up the pace and new interesting things
were found out, among them the zoonotic potential of some members of the genus (Shi and
Hu, 2008; Hon et al., 2008). The ability to break the species barrier is also evident in case
of the newly emerging enteric bovine coronaviruses that share a 98 % RNA sequence
homology with the HECoV-4408 human enteric coronavirus, frequently isolated in
diarrhea cases of children (Han et al., 2006). Also noteworthy in this respect is the fact that
the recently emerged canine respiratory coronavirus (CRCoV), was originally a bovine
coronavirus (Decaro et al., 2007). Coronaviruses are well known for their flexibility in
adapting to new species or new environmental conditions in the host. Their plasticity has
mainly been attributed to three factors, the high mutation rate due to the lack or limited
proof reading activity of the RNA dependent RNA polymerase (Jenkins et al., 2002; Duffy
et al., 2008); the polymerase may move to a different template during replication (Lai,
1992); and also there seems to be a negative correlation between genome size and
mutation-rate. If the genome grows beyond a certain size the amount of accumulated
deleterious mutations will prove fatal to the virus (Eigen, 1987). All these together make
for a potentially fast evolving virus, which is able to adapt to new circumstances rather
fast. 

2. Significance of study:
 This study try to explore the seroprevalence of the transmissible gastroenteritis
among the herds as well as it gives idea about the epidemiology of the disease
occurrence pattern and play tremendous role in disease surveillances and reporting.
 Since, transmissible gastroenteritis in pig causes loss in swine industry. Hence,
finding seroprevalence of TGE will obviously help to implement control strategies
against swine disease in those districts.
3.Objective of the study:
 General objective:
To know the Seroprevalence of procaine transmissible gastroenteritis.
 Specific objective:
To check the presence of TGE antibody in the serum of the swine.
To find the age-wise Seroprevalence of TGE
To find the sex-wise Seroprevalence of TGE
To access the practices among swine owners regarding risk factors for TGE in swine
Risk factors potentially associated with the seropositivity of TGEv in Bhaktapur district

Time and place of study:

This study will be conducted in between July 2018-september2018 in Bhaktapur and Kavre
district and seroprevalence will be tested at NARC, Khumaltar (Lalitpur). Under the
supervision of Dr. Awdesh jha
Review of literature
Several research have been carried out in order to investigate the seroprevalence of
transmissible gastroenteritis in Spain and Korea and the prevalence ranges 5%-65%. Close
study of 71 herds revealed 90% incidence between late December and early April
(Haelterman, 1962-1963). But in Nepal there is no any single research carried out to
investigate this disease. (Cubero MJ1, 1993 Mar 6) In south east Spain found that the
prevalence of seropositive pigs ranged from 5 to 60 per cent. In the United States,
outbreaks that were likely TGE were seen as early as 1943 but the first documented
outbreaks were reported in 1946. , the disease was identified in many other countries.
Serologic surveys indicate that TGE is widespread throughout the US. Although initial
outbreaks are acute, a less severe, endemic form in some herds. TGE can cause up to 100%
neonatal mortality during the initial stages of an outbreak and is therefore unlikely to be
misdiagnosed in a previously naïve herd. (Iowa State University College of Veterinary
Medicine, 2018)
4.METHODOLOGY:
4.1 Study area, district profile and population:

The study will be conducted in Bhaktapur district located in the eastern part of Kathmandu
valley, is the smallest among the seventy-seven districts of Nepal. It is part of Province No.
3. The district, with Bhaktapur as its district headquarters, covers an area of 119 km2 (46
sq. mi) and in 2011 had a population of 304,651 of whom 9,701 people were absent
(mostly working abroad). The district is divided into four municipalities: Bhaktapur,
Changunarayan, Madhyapur Thimi and Suryabinayak Municipality. Next district that the
research will include is kavreplanchok district which is part of Province No. 3, is one of the
seventy-seven districts of Nepal, a landlocked country The of South Asia. The district,
(htt1) population (2011) of 381,937. Following picture illustrate the basic concept about
study area and district profile of this research

bhaktapur
Fig9: map .of Nepal showing study area and district profile

4.2Materials and methods:

 materials
Pigs:
92 pigs from 2district (Bhaktapur and Kavre) will be used for the study of seroprevalence
of gastroenteritis in swine

Others:
 Pig catcher
 Syringe
 Syringe needle
 Ice box
 Blood collecting tubes
 Zipper plastic bag
 EDTA vile
 Gloves
 Methods:

 Study design and Time Frame: Cross sectional study will be conducted from June
2018 to September 2018 in Bhaktapur district of Nepal.
 Sampling Frame: Administratively, the district has four municipality: Bhaktapur,
Changunarayan, Madhyapur Thimi, and Suryabinayak Municipality.
  Sample Size Determination: The sample size necessary for detection of TGE
antibodies will be calculated from EpiTools epidemiological calculators by Ausvet,
sample size is calculated by formula
n=Z P (1-P) (Naing et al, 2006)
2

d2
Where, n= sample size,
Z= Z static for a level of confidence,
P= Expected prevalence or proportion
d=Precision (in proportion of one, if 5% d=0.05)
 Sampling procedure:
. Cross sectional purposive sampling technique is used to farmers.
4.3Blood sampling and data procedure
Blood sampling:
Pig catcher is used to restrain the selected animal and about 5 ml blood was collected from
jugular vein by using sterilized syringe and needle of 18 gauge from pig and immediately
subjected to blood collecting tube and preserve it in ice box to prevent hemolysis. The
sample was taken to NARC, Khumaltar for seroprevalence study through ELISA kit.
Serum samples was stored in a deep freezer at -20°C. Blood sampling:

Data collection:
A structured questionnaire containing associated with TGE was inquired using both closed
and open-ended questions. Questions pertaining to individual swine included age, breed
(indigenous/ cross), sex, BCS, history of abortion and deworming. Additional data were
gathered on general farm and management data such as geographical location of farm,
farm size, husbandry system,
Laboratory Analysis:
Serum sample was tested in porcine transmissible gastroenteritis virus antibody, TGEV
ELISA kit having 96 wells. Which was stored at 2-8℃, manufactured by NOVATEINBIO
Data Analysis:
Data collected in the field using individual data sheet was entered in the MS Excel
spreadsheet. Data from laboratory analysis was coded in Excel spreadsheet. The data from
Excel spreadsheet will be imported into SPSS to complete proportions of seropositive
animals. All analyses wil be based on c-ELISA serological test results.
Prevalence rate will be calculated by dividing no. of seropositive samples by ELISA by
total no. of samples tested.
Association between categorial variables and the outcome variable (seropositive) will be
assessed by using Pearson chi-square test. The multiple effect between predicted variable
and outcome variable was assessed in 95% CI values in a logistic regression model.
Validity of the Test
For optical density (OD 450) value of sample is (A value)
If the A≥0.38, the sample is classified as positive for TGEV antibodies
If the 0.2≤A≤0.38, the sample is classified as suspicious for TGEV antibodies
If the A≤0.2, the sample is classified as negative
Protocols that was demonstrated by Transmissible gastroenteritis virus antibody, TGEV
ELISA Kit, was followed

Results:

Fig10: A values obtained in ELISA analyser

 SEX wise prevalence

Fig11: sexwise prevalence of TGE

Table3: sexwise prevalence of TGE

Agewise prevalence

Agewise prevalence
 Fig12: Agewise prevalence of TGE

Table 4: Agewise prevalence of TGE

District wise prevalence

Fig13: district wise prevalence of TGE

Table5: district wise prevalence of TGE

Overall seroprevalence of TGE in swine of Bhaktapur district based on spss

Test assay Classification prevalence CL
ELISA Positive 90 95%
Negative 1
Total 91
Table6: Overall prevalence of TGE

98.9%

Fig14: Overall prevalence of TGE

DISCUSSION
The first and only outbreak of transmissible gastroenteritis in pigs in Ireland occurred in
1984 in a 650-sow (J Cubero, M Luis 1993)After that Uganda has reported TGEV in
1999(Muley Thorsten 2012)With the appearance and spread of prcov the incidence of TGE
gradually decreased and from an OIE A list disease it became an almost forgotten disease
throughout the world. Occasional reports (Elvander et al., 2000, Brendtsson et al., 2006) of
TGEV-specific seropositivity (“singleton reactors”) indicated that the virus is still present
in pig herds but without clinical manifestation.Morbidity in some groups was estimated at
about 50%, mortality stayed low, at about 4-6%.(Cubero MJ1, 1993 Mar 6) In south east
Spain found that the prevalence of seropositive pigs ranged up to 60 per cent.
Pigs were exposed to transmissible gastroenteritis (TGE) virus when three days old or
when 21 days old. Diarrhea was earliest in onset, most frequent, most profuse and most
prolonged in the youngest group( Norman JO, Lambert G, Moon HW, Stark SL.)
TGE can cause up to 100% neonatal mortality during the initial stages of an outbreak
(Iowa State University College of Veterinary Medicine, 2018)
In the Murcia region of south east Spain, epidemics of transmissible gastroenteritis disease
have occurred in pigs every three years since 1980.(J Cubero, Leon L1993)

Conclusion and Recommendation

This study conclude that the overall prevalence was 98.90% This result closely resembles
with the (Haelterman, 1962-1963) study where 71 herds revealed 90% incidence between
late December and early April.From above statistical analysis there was no significant
difference in Seroprevalence with respect to Age, sex, abortion, housing system and
geography. Implementation of the concept of biosecurity in swine farms of Nepal is
important to strengthen the swine industry.Further researches are essential to determine the
economic losses due to TGE
REFERENCES:
Norman JO, Lambert G, Moon HW, Stark SL. Age dependent resistance to transmissible
gastroenteritis of swine (TGE). II. Coronavirus titer in tissues of pigs after exposure. Can J
Comp Med. 1973 Apr;37(2):167–170. [PMC free article] [PubMed] Saif LJ, Wesley RD.
Transmissible gastroenteritis and porcine respiratory coronavirus. In: Straw BE, D'Allaire
S, Mengeling WL, Taylor DJ, editors. Diseases of Swine. 8th ed. Ames, Iowa USA: Iowa
State University Press; 1999. p. 295-325.
Rasschaert D, Duarte M, Laude H. Porcine respiratory coronavirus differs from
transmissible gastroenteritis virus by a few genomic deletions. J Gen Virol 1990;71: 2599-
607.
http://www.janatapostdaily.com/news-details/1482/2018-02-16.
Cubero MJ1, L. L. (1993 Mar 6). Transmissible gastroenteritis in pigs in south east Spain:
prevalence and factors associated with infection. US National Library of Medicine
National Institutes of Health, 132.
Stock Sanitary Association, Haelterman, E. O. (1962-1963). Epidemiolpgical studies of
transmissible gastroenteritis of swine. Proceedings. United States Live 1962 1963 pp.305-
315 pp.
Haelterman, E. O. (1962-1963). Epidemiolpgical studies of transmissible gastroenteritis of
swine. Proceedings. United States Live Stock Sanitary Association, 1962 1963 pp.305-315
pp.
lowa State University College of Veterinary Medicine. (2018). history of ransmissible
gastroenteritis. Veterinary Diagnostic and Production Animal Medicine.
transmissible gastroenteritis. (1995). Merck & Co., Inc., Kenilworth, NJ, USA.
Gelberg HB: 2007.  Alimentary system - Transmissible gastroenteritis.  IN: Pathologic
Basis of Veterinary Disease.  4th ed., McGavin MD, Zachary JF eds.  Mosby-Elsevier, St.
Louis, MO.  p. 3
 
Saif LJ, Sestak K: 2006.  Transmissible gastroenteritis and porcine respiratory
coronavirus.  In: Diseases of Swine, 9th ed. Straw BE, Zimmerman JJ, D'Allaire S, Taylor
DJ eds. Blackwell Publishing, Ames, IA.  Pp 489-516.75.

Cox, E., Pensaert, M.B. and Callebaut, P. 1993. Intestinal protection against challenge with
transmissible gastroenteritis virus of pigs immune after infection with the porcine
respiratory coronavirus. Vaccine 11: 267-272.
. Bernard, S., Bottreau, E., Aynaud, J.M., Have, P. and Szymansky, J. 1989. Natural
infection with the porcine respiratory coronavirus induces protective lactogenic immunity
against transmissible gastroenteritis. Vet. Microbiol. 21: 1-8.
Carman, S., Josephson, G., McEwen, B., Maxie, G., Antochi, M., Eernisse, K., Nayar, G.,
Halbur, P., Erickson, G. and Nilsson, E. 2002. Field validation of a commercial blocking
ELISA to differentiate antibody to transmissible gastroenteritis virus (TGEV) and porcine
respiratory coronavirus and to identify TGEV-infected swine herds. J. Vet. Diagn. Invest.
14: 97-105.

Cox, E., Pensaert, M.B. and Callebaut, P. 1993. Intestinal protection against challenge
with transmissible gastroenteritis virus of pigs immune after infection with the porcine
respiratory coronavirus. Vaccine 11: 267-272.

Food Safety and Consumer Bureau, Ministry of Agriculture, Forestry and Fisheries. 2009.
Morbidity of Infectious Diseases (2.24 Swine transmissible gastroenteritis). Statistics
Anim. Hyg. 2005: 38

Furuuchi, S., Shimizu, Y. and Kumagai, T. 1975. Comparison of properties between

virulent and attenuated strains of transmissible gastroenteritis virus. Nat. Inst. Anim. Hlth.
Quart. 15: 159-164.

Hao Zou, Dante S. Zarlenga, Karol Sestak, et al. Transmissible gastroenteritis virus:
Identification of M protein-binding peptide ligands with antiviral and diagnostic potential.
Antiviral Research. 2013, Vol.99, No.3, p.383.

Márta Lőrincz, Imre Biksi, Simon Andersson, et al. Sporadic re-emergence of enzootic
porcine transmissible gastroenteritis in Hungary. Acta Veterinaria Hungarica. 2014,
Vol.62, No.1, p.125.
APPENDICES
Appendix:1 questionnaire
Questionnaire for transmissible gastroenteritis
1. Name of farm:
2. District :
3. Municipality :
4. Name of owner:
5. contact no:
6. No of swine :
7. Type of housing system
a) Intensive b) semi intensive c) free range.
8. Feed
a) Hotel waste b) feed ration
9. Sex of swine
a) Male b) female
10. Id no:
Appendix2: protocol of TGE ELISA test
1. All reagents must be allowed to come to room temperature(18-26℃) before use
2. Take out the required coated plates. Set 1 well for blank control, 2 wells for
negative control and positive control each other wells for samples. The Unused
micro-well strip should be sealed as soon as possible and stored at 2-8℃.
3. Add 100 µl sample diluent solution to the blank control well. Add 100µl negative
serum and positive serum to the appropriate wells. Add 100µl of the diluent serum
(6.2) to the remainder of the plate
4. Mix gently, incubate at 37℃ for 30 min.
5. Remove adhesive foil. Pour the liquid out of the wells, add 1×washing solution into
each well, 300µl well, stand for 30 sec. Repeat 5 times, at last time pat to dry on
absorbent paper.
6. Add 100µl Enzyme conjugate into each well(except the blank well). Cover plate
with new adhesive foil. Incubate at 37℃for 30 min.
7. Repeat step 5 (washing)
8. Add 50µl substrate A and 50µl substrate B into each well, mix properly, incubate
for 10 min at 37℃ in the dark with new adhesive foil.
9. Add 50µl stop buffer into each well, mix gently and determine the result within 10
min.
10. Measure the optical density(OD) with a photometer at 450 nm (reference
wavelength: 630nm). Set 0 for the blank well, and read the OD450 value of sample
(A value) on the microplate reader.
11. For the assay to be valid, the negative control mean OD450 should be less than or
equal to 0.1, the positive control mean OD450 should be grater than or equal to 0.6
SOME GLIMPSE OF INTERNSHIP ACTVITIES