Nghiên cứu bệnh viêm ruột do parvovirus trên chó tại một số tỉnh thành đồng bằng sông cửu long tt tiếng anh

MINISTRY OF EDUCATION AND TRAINING
CAN THO UNIVERSITY

SUMMARY OF DOCTORAL DISSERTATION
Major: PATHOLOGY AND TREATMENT OF
ANIMALS
Major code: 62 64 01 02

NGUYEN THI YEN MAI

RESEARCH ON CANINE PARVOVIRUS
INFECTION IN DOGS IN SOME PROVINCES OF
THE MEKONG DELTA

Can Tho, 2020
i


THE DISSERTATION WAS COMPLETED
AT CAN THO UNIVERSITY

Supervisor: Assoc. Prof. Dr. Tran Ngoc Bich

The dissertation is defendded in front of the University
Examination
Place: ...................................................., Can Tho University
Time::....................................................................................

Reviewer 1:
Reviewer 2:
Reviewer 3:

Futher information of the dissertation could be found at:
Learning Resource Center of Can Tho University
National Library of Vietnam
ii


THE LIST OF PUBLISHED WORKS RELATED TO
THIS DISSERTATION
1. Nguyen Thi Yen Mai, Tran Ngoc Bich, Pham The Lam, Nguyen
Phuc Khanh, 2016. Parvovirus disease in dogs at Can Tho.
Animal Health Clinic Office Journal of Agriculture and Rural
Development No. 11/2016, pages 151-155.
2. Nguyen Thi Yen Mai, Tran Ngoc Bich, Tran Van Thanh, 2018.
Situation of Parvovirus enteritis in dogs at Veterinary Offices of
Tien Giang, Dong Thap and Can Tho City. Science Journal of Can
Tho University. 54 (Agricultural Union No.), pages 136-142.
3. Nguyen Thi Yen Mai, Tran Ngoc Bich, Tran Van Thanh, 2018.
Situation of Parvovirus enteritis in dogs at Can Tho University
Veterinary Clinic. Journal of Veterinary Science and Technology,
volume XXV. Number 4, pages 36-41.
4. Tran Van Thanh, Tran Ngoc Bich, Thai Quoc Hieu, Nguyen Thi Yen
Mai, 2018. Situation of Parvovirus enteritis in dogs at Tien Giang
Department of Livestock and Veterinary Medicine. Journal of
Agriculture and Rural Development No. 24/2018, pages 102-107.
5. Keovongphet Phuthavong, Tran Ngoc Bich, Nguyen Thi Yen Mai,
Tran Van Thanh, 2018. Survey of Parvovirus enteritis in dogs at
Can Tho University Veterinary Clinic. Science Journal of Can Tho
University. 54 (Agricultural Union No.), pages 51-55.

iii


Chapter I: INTRODUCTION
1.1 Necessity of thesis
Inflammatory bowel disease caused by Canine Parvovirus type 2
(CPV-2) in dogs is an acute infectious disease (Kelly 1978; Appel et al.,
1979; Hoelzer et al., 2008). CPV-2 is considered the most threatening to
puppies from weaning to six months of age. However, outbreaks of
inflammatory bowel disease and dog adult mortality from CPV-2 have
been reported (Decaro et al., 2008). The disease occurs in all breeds of
dogs, all of ages and regardless of gender (Castro et al., 2007; Gombac et
al., 2008). The disease progresses rapidly, making it difficult for dog
owners to detect early, when it was detected that the dog was seriously ill
and difficult to treat. The main symptoms of the diseases include fever,
vomiting, diarrhea, loose stools mixed with fresh blood with an
unpleasant fishy odor that leads to dogs losing a lot of blood, water and
electrolytes, metabolic disorders, cardiovascular collapse, kissing
delirious and dead (Sherding, 2000; Joshi et al., 2001; Ramprabhu et al.,
2002; Miranda et al., 2016).
According to the Sub-Department of Livestock and Veterinary,
Can Tho city, Dong Thap, Tien Giang and Ben Tre, at December 2018,
in the Mekong Delta, the demand for raising pets of people is increasing;
in which dog is one of the most favourite animal as pet. For the elderly,
lonely singles, children,... as dog is friendly and consider pet as a
companion every day, even dog is consider a part of the family.
Although the incidence of inflammatory bowel disease in dogs due to
CPV-2 were 43.30% (Dung Nguyen Van et al., 2017) and the cure rate
for symptoms were quite high at 86.30% (Le Minh Thanh, 2009). But
when the pet was sicked, the days of treatment have not been cured or

not cured, will be the unavoidable days that owners were worry,
depression,... Therefore, the study of this disease in the Mekong Delta is
necessary to make the disease prevention and intervention more active in
order to reduce economic losses and especially limit the emotional
damage of dog owners. Because of all the reasons above, thesis:

1


“Research on inflammatory bowel disease caused by CPV-2 in the
Mekong Delta region” was carried out.
1.2 Thesis Objectives
Genotype sequencing, establishment of phylogenetic trees,
identification of infection rates, assessment of homologeneity of
molecular genetic characteristics of disease-causing CPV-2 genotypes
with genotypes of CPV-2 in vaccines on gene banks; evaluating the
effect of treatment and protection ability of CPV-2 preventive vaccine, in
order to help veterinary medicine that was oriented intervention to
overcome disease symptoms, producing and using vaccines to prevent
and fight against enteritis due to CPV-2 in dogs in the Mekong Delta that
were effective treatment.
1.3 The new distribution of thesis
This is the first research project in Vietnam that basically and
systematically studied a wide range of CPV-2 inflammatory bowel
disease in dogs that is being widely adopted in some provinces and cities
in the Mekong Delta. Therefore, the result of the thesis was to establish a
phylogenetic tree, determine the infection rate and the circulation of
pathogenic genotypes of CPV-2. Assessing the homogeneity of
molecular genetic characteristics of field CPV-2 with genotypes of CPV2 in vaccines on the gene banks and assessing protection ability of CPV2 vaccine in dogs currently circulating in some provinces in the Mekong
Delta.

1.4 Practical significance and applied capacity of thesis
This thesis is the first basic information source of scientific
foundation in Vietnam for researching about inflammatory bowel disease
caused by CPV-2 in dogs in some provinces and cities in the Mekong
Delta. From there, this result actively supported local veterinarians to
produced vaccines to prevent and against CPV-2, diagnosis, prognosis,
effective treatment and scientific data for the following studies.

2


Chapter III: MATERIALS AND METHODS
The thesis was carried out with 4 contents:
3.1 Content 1: Determining the incidence of CPV-2
inflammatory bowel disease in dogs treated in some clinics of some
provinces of the Mekong Delta
3.1.1 Experimental subjects: 380 dogs with symptoms of
vomiting, diarrhoea, bloody diarrhea. Making medical record sheets to
monitor age, gender, breed and vaccination.
3.1.2 Number of survey samples: 380 samples from the
Veterinary Offices of Can Tho city, Tien Giang, Dong Thap and Ben Tre
(each province selected two clinics), collecting samples according to the
formula of Thrusfield (1995).
P (1-P)
0.433 (1- 0.433)
N = 1.962
= 377.262
n = (z)2 x
2
0.05

d2
In which: n= number of sample properties
Z= value of thenormal distribution for confidence level of 95%
P= expected prevalence
d= absolute error of 5%

The samples was presented in Table 3.1.
Table 3.1: Number of samples
Numerical order

Place

Numerical of samples

1

Can Tho city

101

2

Dong Thap

90

3

Tien Giang

97

4

Ben Tre

92

Total

380

3


3.1.3 Research method
Descriptive cross-sectional research, firstly study and analysis.
Collecting fecal samples of sick dogs with symptoms of vomiting,
diarrhoea at Veterinary Clinics of some provinces above to identify dogs
positive for CPV-2 inflammatory bowel disease by Antigen Rapid CPV
test kit of American Bionote company monitors. Monitoring and
recording a chronic disease to prevention and treatment. Every day,
before the medication was gave, surveyed and recorded body
temperature, diarrhea, dehydration, blood loss, fecal,... recording the
disease progression and make a medical record sheet for each infected
dog. After that, providing supportive drugs, the course of treatment for 3
to 7 days.
3.1.4 Monitoring targets
The rate of dogs with inflammatory bowel disease by CPV-2 on
age, sex, breed and vaccination; frequency of the typical clinical

manifestations and evaluation of treatment effect of this disease in dogs
at the Veterinary Clinical Offices of the four provinces mentioned above
were recorded.
3.2 Content 2: Identification of genotypes of CPV-2
inflammatory bowel disease in dogs treated in some clinics of some
provinces of the Mekong Delta
3.2.1 Test subjects: Selecting 80 samples with positive results for
CPV-2 in content 1.
3.2.2 Research methodology: Collecting 80 fecal samples from
80 individual dogs with clear positive results for CPV-2 in content 1.
Then, selecting each province into 20 samples (with time to appeared red
line at position C that was earlier than 5 minutes after the sample
solution was put into the test and there were 2 clear pink lines),
conducted DNA extraction, performed PCR reaction with primers VP2
GMCOM (Gurpreet Kaur et al., 2015) were presented in Table 3.2.

4


Table 3.2: Primer set VP2 GMCOM (Gurpreet Kaur et al., 2015)
Primer
Primer sequencing
Accession Nucleotide
name
(5’ – 3’)
No.
position
VP2
2816GGTCAACCTGCTGT
GMCOMF

CAGAAA
2835
M19296.1
VP2
AGGTGCTAGTTGA
4525GMCOMR
GATTTTTCAT
4503
F: forward; R: reverse

Product
length
1710bp

Reaction conditions: 950C for 5 minutes, followed by 950C for 30
seconds, 580C for 30 seconds, 720C for 1 minute, this cycle was repeated
35 times, finally 720C in 5 minutes. After the PCR reaction was finished,
the PCR product was tested on 1.5% agarose gel by electrophoresis
method. The purified DNA product was sent to Phu Sa Biochem Co.,
Ltd. to decoded the VP2 gene sequence (by Sanger method on ABI
system 3130 (USA) and check the order received by BioEdit software).
When results of sequence analysis of CPV-2 genome were available,
collating the genome sequence of CPV-2 of the sequenced samples in the
above primers and compared with the nucleotide positions of the control
samples which was determined CVP-2, CPV-2a and CPV-2b.
3.2.3 Monitoring criteria: Determination of CPV-2 type and
genotype of CPV-2: CPV-2a, CPV-2b, CPV-2c, new CPV-2a and new
CPV-2b in dogs with inflammatory bowel disease treated at Veterinary
clinics of Can Tho city, Dong Thap, Tien Giang and Ben Tre.
3.3 Content 3: Determining and evaluating the homogeneity of

molecular genetic characteristics of the genotypes of field CPV-2
(DBSCL) infections in dogs with CPV-2 genotypes on gene banks
(Genbank); with CPV-2 genotypes in vaccines on gene banks of
NCBI
3.3.1 Test subjects: Selecting 32 representative samples with
clear and beautiful results in content 2 for 4 provinces and cities of the
Mekong Delta (8 samples for each province).
3.3.2 Research methodology: After the research samples were
sequenced, the complete sequences will be read and edit in Bioedit

5


7.0.5.3, this file contained at least 1 sequence of each type and included
all genotypes of CPV-2 (CPV-2a, CPV-2b, CPV-2c), original CPV-2
genotypes and recent CPV-2 genotypes were collected from NCBI's gene
bank. The sequences were fully aligned (Global alignment) by Clustal-W
2.0.11 software. The CPV-2 segment of all sequences, after being sorted,
it will be exported as a fasta file and used to draw a phylogenetic tree
with MEGA 6 software. The phylogenetic tree was drew by the
Neighbor Joining method with the nucleotide model (substitution
model), the Maximum Composite Likelihood and 1000 samples of
Boostrap relication. The percentage of the number of occurrences of a
group on the total number of sampled returns was an indicator of a
group's Bootstrap support value. The percentage difference between the
nucleotide sequences of the cloned CPV-2 from the sample and the
reference sequence was calculated by the MEGA 6 software, thereby,
determining the generation distance of the phylogenetic tree.
3.3.3 Monitoring criteria: Determining and evaluating the
homogeneity of molecular genetic characteristics of type CPV-2 causing

inflammatory bowel disease in dogs in some provinces of Mekong Delta
with CPV-2 genotypes on gene banks (Genbank), also with genotypes of
CPV-2 in vaccines gene banks of NCBI.
3.4 Content 4: Assessing the protective capacity of Parvovirus
vaccine in circulating dogs in some provinces in the Mekong Delta
3.4.1 Experimental subjects: Foreign and domestic dogs breed at
6 weeks of age, healthy, not vaccinated with Parvovirus. These dogs
were the offspring of mother dogs that have been vaccinated against
CPV-2 enteritis vaccine.
3.4.2 Experimental design: 15 foreign dogs breeds and 15
domestic dogs breeds were injected with 3 selected vaccines (symbolized
as sequencing areV1, V2 và V3).
3.4.3 Research methodology: All of domestic and foreign dogs
breeds above were cared out, nourished, prevented and treated during the

6


same study period (only different in 3 types of preventive vaccines). The
first vaccination was at 6 weeks of age, and repeated twice after the first
3 weeks (according to manufacturer's recommendations). Collecting
blood samples (each dog from 1-2 ml blood) at the time before
vaccination; after repeated second vaccination: 1 month, 3 months, 6
months, 9 months and 12 months, centrifuging (3000 rpm) for 5 minutes
and extracting serum (stored at minus 20oC). Then, the antibody content
was determined by ELISA kit (Asan Easy Test ® CPV Ab) of Korea.
3.4.4 Recorded parameters: Comparing with the protective
ability of 3 types of CPV-2 preventive vaccines in circulating dogs in
some provinces of the Mekong Delta and compared with domestic and
foreign dogs breeds in this experiment.

3.5 Statistical analysis: The first data was analyzed by Microsoft
Excel 2010 software (to calculate data: total, percentage,...) the summary
data of the report was analyzed by Chi square (χ2) and ANOVA twoway anlysis of variance of Minitab Statistical Software version 16.0.
Chapter IV: RESULT AND DISCUSSION
4.1 The prevalence of CPV-2 inflammatory bowel disease in
dogs treated in some clinics of some provinces of the Mekong Delta
4.1.1 The prevalence of CPV-2 infection in dogs
The prevalence of CPV-2 infection in dogs was presented in Table
4.1.
Table 4.1: The rate of dogs with inflammatory bowel disease caused by CPV-2
via CPV Ag test kit
Number of dogs vomiting
and diarrhea with bloody
stool (n=380)

Number of samples
infected with CPV2 (n=123)

Can Tho city

101

33

32.67

Dong Thap

90

30

33.33

Clinic

7

Rate (%)


Tien Giang

97

31

31.96

Ben Tre

92

29

31.52

Average

32.37

(P>0.05)

Table 4.1 showed that the rate of CPV-2 infection in dogs with
symptoms of vomiting, diarrhea mixed with blood in four provinces in
the Mekong Delta were no difference (P>0.05). The overall rate of CPV2 infection in dogs with diarrhea with bloody stool in the Mekong Delta
was 32.37%.
4.1.2 The prevalence of CPV-2 infection in dogs by age
The prevalence of CPV-2 infection in dogs by age was presented
in Table 4.2.
Table 4.2: The prevalence of CPV-2 infection in dogs by age
Number of samples
Ages (months)
Number of samples
positive for CPV-2
observed (n=380)
(n=123)
1 to 3
143
71
>3 to 6
141
42
>6
96
10

Rate (%)
49.65
29.79
10.42

(P<0.001)

Table 4.2 showed that the ration of dogs infected by CPV-2 in the
Mekong Delta from 1 to 3 months was the highest at 49.65%. Dogs at
the age of 3 to 6 months was 29.79%. For dogs older than 6 months, the
lowest infection rate was 10.42%. There were significantly different
(P<0.001).
4.1.3 The prevalence of CPV-2 infection in dogs by sex
The prevalence of CPV-2 infection in dogs by sex was presented
in Table 4.3.

8


Table 4.3: The prevalence of CPV-2 infection in dogs by sex
Number of samples
Number of samples
Sex
positive for CPV-2
observed (n=380)
(n=123)
Male
199
66
Female
181
57

Rate (%)
33.17

31.49
(P>0.05)

As shown in the Table 4.3, the difference of CPV-2 infection rate
among dogs with inflammatory bowel disease by sex in some provinces
in the Mekong Delta was not statistically significant (P>0.05).
4.1.4 The prevalence of CPV-2 infection in dogs by breed
The prevalence of CPV-2 infection in dogs by breed was
presented in Table 4.4.
Table 4.4: The prevalence of CPV-2 infection in dogs by breed
Number of samples
Number of samples
Breed
positive for CPV-2
observed (n=380)
(n=123)
Domestic
191
63
Foreign

189

59

Rate (%)
32.98
31.22
(P>0.05)

The results of Table 4.4 showed that the rate of CPV-2 infection in
dogs in some provinces of the Mekong Delta by domestic dog breed was
32.98%, foreign dog was 31.22%. The difference was not statistically
significant (P>0.05).
4.1.5 The prevalence of CPV-2 infection in dogs by vaccination
The prevalence of CPV-2 infection in dogs by vaccination was
presented in Table 4.5.

9


Table 4.5: The prevalence of CPV-2 infection in dogs by vaccination
Number of samples
Number of samples
Vaccination
positive for CPV-2
Rate (%)
observed (n=380)
(n=123)
Unvaccinated
281
109
38.79
Vaccinated
99
14
14.14
(P<0.001)

The results of Table 4.5 showed that 281 dogs with vomiting,

diarrhea mixed with bloody stool, unvaccinated for CPV-2 as well as
positive results for CPV-2 was 38.79%; CPV-2 vaccination was 14.14%.
The difference in the rate of CPV-2 infection in unvaccinated dogs and
CPV-2 vaccination were statistically significant (P<0.001).
4.1.6 Frequency of typical clinical symptoms in dogs with
inflammatory bowel disease caused by CPV-2
Frequency of typical clinical symptoms in dogs with inflammatory
bowel disease caused by CPV-2 was presented in Table 4.6.
Table 4.6: Frequency of typical clinical symptoms in dogs with CPV-2
inflammatory bowel disease caused by CPV-2 (n=123)
Number of samples
Clinical symptom
Frequency (%)
observed (dog)
Anorexia, lethargy
123
100
Anorexia
123
100
Diarrhea
123
100
Diarrhea with mucus
123
100
Diarrhea with bloody stool
123
100
Stinky stool

123
100
Vomiting
112
91.06
Vomiting with viscous mucus
112
91.06
Fever
33
26.83
Vomiting blood
6
4.88

The results from Table 4.6 showed that the common symptoms of
this disease such as anorexia, lethargy were 100%. The disease was not
only common symptoms but also typical symptoms such as diarrhea,

10


diarrhea mixed with blood, diarrhea with mucus, stinking stool were
100% and vomiting was 91.06%.
4.1.7 The effect of treatments in dogs with inflammatory bowel
disease caused by CPV-2
The effect of treatments in dogs with inflammatory bowel disease
caused by CPV-2 was presented in Table 4.7.
Table 4.7: The effect of
caused by CPV-2

Number
Clinic
of dog
treated
(dog)
Can Tho city
31
Dong Thap
33
Tien Giang
29
Ben Tre
30
Total
123

treatments in dogs with inflammatory bowel disease
Number of days of cure
5 to 7 dasy
8 to 10 days
Recovery Rate Recovery dog Rate Recovery Rate
dog (dog) (%)
(dog)
(%) dog (dog) (%)
27
87.10
25
91.67
2
8.33

27
81.82
26
96.00
1
4.00
25
86.21
23
92,31
2
7.69
26
86.67
24
92.31
2
7.69
105
85.37
98
93.07
7
6.67
(P>0.05)
Results of cure

The results of Table 4.7 showed that the rate of treatments in dogs
with inflammatory bowel disease caused by CPV-2 was not statistically
significant (P>0.05). The average of CPV-2 symptomatically treatments

in dogs with inflammatory bowel disease in these provinces were
85.37%.
4.2. Results of genotype determination of CPV-2 infection in
dogs
The rate of genotypes of CPV-2 was presented in Table 4.8.

11


Table 4.8: Results of genotype determination of CPV-2 infection in dogs (CPV 2a, CPV-2b, CPV-2c, New CPV-2a and New CPV-2b)
Genotype CPV
Total number of
Total number of
Rate (%)
samples tested
positive samples
CPV -2a
80
01
1.25
CPV -2b
80
0
0
CPV -2c
80
79
98.75
New CPV-2a
80

0
0
New CPV-2b
80
0
0
(P<0.01)

Through Table 4.8, genotype CPV-2c was 98.75%, CPV-2a was
1.25% and genotypes CPV-2b, new CPV-2a and new CPV-2b were not
detected. There were significantly different (P<0.01). The results also
showed that the main genotype circulating and causing intestinal disease
caused by CPV-2 in dogs in some provinces of the Mekong Delta that
was genotype CPV-2c.
4.3 Determining and evaluating the homogeneity of molecular
genetic characteristics of the genotypes of field CPV-2 (DBSCL),
infections in dogs with CPV-2 genotypes on gene banks (Genbank);
with CPV-2 genotypes in vaccines on gene banks of NCBI
The result comparison between 32 genotypes of CPV-2 collected
in some provinces of the Mekong Delta in this study has a very high
uniformity in both nucleotide and amino acid composition (98.50-100%;
98.10-100%). When compared with the reference genotypes on NCBI's
gene bank, it was found that the studied genotypes had a high
homogeneity of nucleotide composition (98.30-99.90%), amino acid
(97.40-99.20%) with 2 genotypes of CPV./dog/HCM/8/2013 and
CPV/dog/HCM/7/2013 of Ho Chi Minh City, Vietnam (Genbank
numbers was LC216907 and LC216969, respectively); and these
genotypes also showed homologeneity with other genotypes of CPV-2
in the world, in which the highest ratio of homologeneity with 2
genotypes: CPV-SD-14-12 (KR611522) from Korea (nucleotide 98.4099.90%, amino acid 96.80-99.20%) and YANJI-1 (KP749854) isolated

12


in China
99.20%).

(nucleotide

98.40-99.90%,

amino

acid

97.40-

CT5
TG39
CT21
DT01
KP749854.1 YANJI-1 (China)
BT10
MF467242.1 CPV-GX1581 (China)
CT12
CT69
KR611522.1 CPV-SD-14-12 (Korea)
CT102
TG179
LC216907.1 CPV/dog/HCM/8/2013 (Viet Nam)

BT08
TG172
CT100
LC214969.1 CPV/dog/HCM/7/2013 (Viet Nam)
CT45
CT60
DT09
TG45
BT16
BT11
DT02
DT04
TG62
BT02
BT09
DT08
DT11
BT64
BT20
DT03
DT05
TG32
TG37
TG40
KM457143.1 UY364 (Uruguay)
KM457102.1 UY243 (Uruguay)
KR002794.1 CPV/CN/HB3/2013 (China)
KR002795.1 CPV/CN/JL1/2013 (China)
EF599096.1 DH426 (Korea)
U72698.1 T37 (Taiwan)

AB054214.1 LCPV T1 (Taiwan)
AB054213.1 Taiwan 9 (Taiwan)
GU212791.1 VAC P vanguard Vaccine (Thailan)
FJ011098.1 Intervet vaccine (Taiwan)
FJ011097.1 Merial vaccine (Taiwan)

Figure 4.1: Phylogenetic tree of 32 genotypes of CPV-2 isolated in some
provinces of the Mekong Delta

The results of Figure 4.1 showed that the genotypes isolated in this
study have clear branched. In which all of genotypes CPV-2c stands
alone on the branch of phylogenetic tree. Particularly, the genotype of
CPV-2a stands apart in a new branch and almost no CPV-2 genotypes
stands in the same branch as genotype of CPV-2 in vaccines on gene
banks. The homogeneity between genotypes of CPV-2 isolated in the

13


study and genotypes of CPV-2 published in the gene bank (Genbank) of
NCBI was evaluated by compared the nucleotide and amino acid
components of this study. 8 genotypes of CPV-2 were collected in each
study site with the genotypes of CPV-2 published in the world,
specifically as follows:
4.3.1 In Can Tho city
The results showed that between 8 genotypes of CPV-2 studied
together, the nucleotide homogeneity index was 99.20-99.90%; with
genotypes of CPV-2 on gene banks (Genbank) and genotypes of CPV-2
in vaccines on gene banks, respectively 98.30-99.90% and 97.5098.40%. The homogeneity of amino acid composition between 8
genotypes CPV-2c studied together reached the homogeneity rate of

98.40-100%; with genotypes of CPV-2 on gene banks (Genbank) and
genotypes of CPV-2 in vaccines on gene banks have homogeneity ratios
of 97.40-100% and 95.50-97.40%.
Through a phylogenetic trees, analyzing branched genetic diagram
found that 8 research genotypes were divided into 2 small groups (the
first group consisted of 2 genotypes CT60 and CT100 with boot trap
index of 62.00%; the other group was on the same branch (CT5, CT12,
CT21, CT45, CT69 and CT102) with 2 genotypes SD-14-12
(KR611522.1) and YANJI-1 (KP749854.1) have been published in the
world with bootstrap values 97.00% (Figure 4.2).

14


CT5
LC214969.1 CPV/dog/HCM/7/2013 (Viet Nam)
CT21
CT12
KR611522.1 CPV-SD-14-12 (Korea)
CT100
CT102
KP749854.1 YANJI-1 (China)
LC216907.1 CPV/dog/HCM/8/2013 (Viet Nam)
MF467242.1 CPV-GX1581 (China)
CT45
CT60
CT103
KM457143.1 UY364 (Uruguay)
KM457102.1 UY243 (Uruguay)
KR002794.1 CPV/CN/HB3/2013 (China)

KR002795.1 CPV/CN/JL1/2013 (China)
EF599096.1 DH426 (Korea)
U72698.1 T37 (Taiwan)
AB054214.1 LCPV T1 (Taiwan)
AB054213.1 Taiwan 9 (Taiwan)
GU212791.1 VAC P vanguard Vaccine (Thailan)
FJ011098.1 Intervet vaccine (Taiwan)
FJ011097.1 Merial vaccine (Taiwan)

Figure 4.2: Phylogenetic tree of 8 genotype CPV-2 isolated in Can Tho city

4.3.2 In Dong Thap
The results showed that between 8 genotypes of CPV-2 in Dong
Thap, there were 99.00-99.90% nucleotide homogous levels; with
genotypes of CPV-2 on gene banks and genotypes of CPV-2 in vaccines
on gene banks, respectively, are 97.40-98.80% and 96.80-97.40%. The
amino acid homogeneity between the 8 genotypes in the study together
was almost homogeneous with the rate of 98.90-100%; with the
genotypes of CPV-2 available on the gene bank and the genotypes of
CPV-2 in vaccines on gene banks, the homologeneity were 97.5098.90% and 95.80-96.40%, respectively.

15


8 genotypes of CPV-2 in Dong Thap, when participated in the
branching scheme, the research genotypes were separated into a separate
subgroup with the second subtype including genotype SD-14-12
(KR611522 .1) and YANJI-1 (KP749854.1) with a bootstrap value of
100% (Figure 4.3) of the same main subgroup.
EF599096.1 DH426 (Korea)

U72698.1 T37 (Taiwan)
AB054214.1 LCPV T1 (Taiwan)
AB054213.1 Taiwan 9 (Taiwan)
KR002794.1 CPV/CN/HB3/2013 (China)
KR002795.1 CPV/CN/JL1/2013 (China)
KM457143.1 UY364 (Uruguay)
KM457102.1 UY243 (Uruguay)
MF467242.1 CPV-GX1581 (China)
LC216907.1 CPV/dog/HCM/8/2013 (Viet Nam)
LC214969.1 CPV/dog/HCM/7/2013 (Viet Nam)
KR611522.1 CPV-SD-14-12 (Korea)
KP749854.1 YANJI-1 (China)
DT01
DT02
DT09
DT03
DT11
DT05
DT04
DT08
GU212791.1 VAC P vanguard Vaccine (Thailan)
FJ011098.1 Intervet vaccine (Taiwan)
FJ011097.1 Merial vaccine (Taiwan)

Figure 4.3: Phylogenetic tree of 8 genotype CPV-2 isolated in Dong Thap
province

4.3.3 In Tien Giang
Results of analyzed the nucleotide homogeneity between 8
genotypes of CPV-2 in the study with each other that have the

homogeneity ratio from 98.70-100%; with genotype of CPV-2 in gene
bank was 98.40-100% and genotypes of CPV-2 in vaccine on gene banks

16


were 97.10-98.00%. The amino acid composition of these 8 genotypes
was also very high (98.10-100%) and when compared with the genotype
of CPV-2 in vaccines on gene banks, the homogeneity rate was 96.3097.70%. In addition, when considered the amino acid composition of
genotype CPV-2a (TG40) with the genotypes CPV-2a and CPV-2c on
gene banks, the difference was not significant (with CPV-2c was 98.1099.80%; with CPV-2a was 99.20-99.80%). The genotypes CPV-2a and
CPV-2c have certain differences in nucleotide composition, however, it
was not affected the amino acid modification much, similar results were
verified by study of Hoang et al. (2019).
Through a phylogenetic trees, it was shown that among the
genotypes CPV-2c was isolated in Tien Giang. Genotype TG179 forms a
major subgroup (Bootstrap 99.00%). The remaining 6 genotypes: TG45,
TG62, TG32, TG172, TG37, TG39 together with genotypes SD-14-12
(Korea), YANJI-1 and CPV-GX1581 (China) and 2 genotypes CPV/dog/
HCM/8/2013 (Vietnam) formed the remaining branch (bootstrap
60.00%). Notably, the genotype TG40 was identified as CPV-2a in the
same subgroup with the genotype CPV-2a from countries such as
Uruguay UY243 (KM457102.1), UY364 (KM457143.1) with a
bootstrap value of 93.00%; and belongs to the same sub-group with
Chinese
genotypes:
CPV/CN/HB3/2013
(KR002794.1)
and
CPV/CN/JL1/2013 (KR002795.1); genotype Taiwan: T37 (U72698.1),

LCPV T1 (AB054214.1) and Taiwan 9 (AB054214.1) and Korean
genotype: DH426 (EF599096.1) with a bootstrap value of 93.00%
(Figure 4.4).

17


TG39
TG172
LC216907.1 CPV/dog/HCM/8/2013 (Viet Nam)
LC214969.1 CPV/dog/HCM/7/2013 (Viet Nam)
MF467242.1 CPV-GX1581 (China)
TG32
TG37
TG45
TG62
TG179
TG40
KM457143.1 UY364 (Uruguay)
KM457102.1 UY243 (Uruguay)
KR002794.1 CPV/CN/HB3/2013 (China)
KR002795.1 CPV/CN/JL1/2013 (China)
U72698.1 T37 (Taiwan)
EF599096.1 DH426 (Korea)
AB054214.1 LCPV T1 (Taiwan)
AB054213.1 Taiwan 9 (Taiwan)
KR611522.1 CPV-SD-14-12 (Korea)
KP749854.1 YANJI-1 (China)
GU212791.1 VAC P vanguard Vaccine (Thailan)
FJ011098.1 Intervet vaccine (Taiwan)

FJ011097.1 Merial vaccine (Taiwan)

Figure 4.4: Phylogenetic tree of 8 genotype CPV-2 isolates in Tien Giang
province

4.3.4 In Ben Tre
The results of analyzing the nucleotide homogeneity between 8
genotypes of CPV-2 in the study with each other have the homogeneity
of 98.80-100%; with the genotypes of CPV-2 on the gene bank and with
the genotypes of CPV-2 in vaccines on gene banks have a high
proportion of homogeneity (respectively 97.20-99.80%; 97.70-98.40%).
The amino acid homogous ratio between 8 genotypes of CPV-2 was
97.90-100%; With the genotypes of CPV-2 on gene banks and in

18


vaccines on gene banks, the correlation rates were quite high (97.5099.60% and 95.20-96.40%, respectively).
Through a phylogenetic trees, it showed that all of 8 genotypes
studied in Ben Tre were grouped in the same main subgroup of
phylogenetic trees. However, in these group, it was divided into 2 subgroups (genotype BT10 forms a separate subgroup with 100% bootstrap
index; the remaining 7 genotypes include BT64, BT08, BT09, BT11,
BT16, BT20 and BT10 formed a subgroup with a bootstrap index of
69.00%) with genotype SD-14-12 (KR611522.1) from China and
YANJI-1 (KP749854.1) of South Korea published on previous world
(Figure 4.5).
BT02
BT64
BT09
BT20

BT11
BT16
BT08
BT10
KR611522.1 CPV-SD-14-12 (Korea)
KP749854.1 YANJI-1 (China)
GU212791.1 VAC P vanguard Vaccine (Thailan)
FJ011098.1 Intervet vaccine (Taiwan)
FJ011097.1 Merial vaccine (Taiwan)
LC216907.1 CPV/dog/HCM/8/2013 (Viet Nam)
LC214969.1 CPV/dog/HCM/7/2013 (Viet Nam)
MF467242.1 CPV-GX1581 (China)
KM457143.1 UY364 (Uruguay)
KM457102.1 UY243 (Uruguay)
KR002794.1 CPV/CN/HB3/2013 (China)
KR002795.1 CPV/CN/JL1/2013 (China)
U72698.1 T37 (Taiwan)
EF599096.1 DH426 (Korea)
AB054214.1 LCPV T1 (Taiwan)
AB054213.1 Taiwan 9 (Taiwan)

Figure 4.5: Phylogenetic tree of 8 genotypes of CPV-2 isolated in Ben
Tre province

19


In summary, most of the genotypes of CPV-2 isolated in some
provinces of the Mekong Delta belong to the genotype CPV-2c with the
characteristic Glutamic acid (E) at amino acid position 426. In the CPV2c group, there were also additional genotype CPV-SD-14-12 of Korea

(KR611522), 2 Chinese genotypes including JANJI (KP749854) and
GX1581 (MF467242) and 2 genotypes isolated in Vietnam including
LC216907 and LC214696. While the only CPV-2a genotype in the study
(TG40) expressed the amino acid Asparagine (N) at the amino acid
position of 426 instead of Glutamic acid like the CPV-2c genotype. This
was also found in Uruguay genotypes UY243 and UY364 Genbank
numbers KM457102 and KM457143, respectively. The above genotypes
have very high and almost homogous nucleotide and amino acid
homogeneity between the 32 genotypes of CPV-2 in the study; with
genotypes of CPV-2 in gene banks and genotypes of CPV-2 in field
vaccines. This proved that the genotypes of CPV-2 in the study, the gene
bank and the genotypes in vaccines on gene banks were closely related.
4.4 Assessing of protection of Parvovirus vaccine in circulating
dogs in some provinces of the Mekong Delta
4.4.1 Comparison of antibody content (immune response) after
vaccination against CPV-2 enteritis vaccine in dogs in some
provinces of the Mekong Delta

20


Table 4.9: CPV-2 antibody content (AC) in dogs before and after getting a
vaccination
V1
No. of
samples
with
PAC

Rate

(%)

10

8

1

10

3

V2
AC
(HI
titer)

No. of
samples
with
PAC

Rate
(%)

80

86.5

6

10

100

275

10

10

100

6

10

10

9

10

12

10

Time

Before

VC

After VC (months)

No. of
sampl
es
tested

V3
AC
(HI
titer)

No. of
samples
with
PAC

Rate
(%)

AC
(HI
titer)

60

85.0

6

60

84.0

10

100

275.5

10

100

274

516

10

100

511.5

10

100

514

100

395

10

100

400.5

10

100

388.5

10

100

308.5

10

100

314.5

10

100

314.5

10

100

224

10

100

225

10

100

227.5

P<0.01
Note: PAC: Protective antiboties sufficient; VC: Vaccination; AC: CPV-2 antibody
content

Table 4.9 showed that antibody contented generated after 01, 03,
06, 09 and 12 months after vaccination of 3 vaccines were not

statistically significant (P>0.05). The results also showed that all of three
vaccines after 12 months of vaccination have sufficient antibody
contented with a protection rate of 100%.
4.4.2 Comparison of antibody content (immune response) after
vaccination against CPV-2 enteritis vaccine in dogs by breed in some
provinces of the Mekong Delta

21

P>
0.05


Table 4.10: CPV-2 antibody contented (AC) in dogs before and after getting a
vaccination by breed
V1

Time

After VC (months)

6

No. of
sampl
es
PAC

Rate
(%)

AC
(HI
titer)

5

4

80

87.0

3

60

84.0

3

60

83.0

5

5

100

273

5

100

278

5

100

272

5

5

100

514

5

100

511

3

100

515

5

5

100

394

5

100

398

5

100

385

5

5

100

307

5

100

315

5

100

316

5

5

100

221

5

100

224

5

100

226

5

4

80

86.0

3

60

86.0

3

60

85.0

5

5

100

277

5

100

273

5

100

276

5

5

100

518

5

100

512

5

100

513

5

5

100

396

5

100

403

5

100

392

5

5

100

310

5

100

314

5

100

313

5
5
100
227
5
100
226
5
100
P>0.05
Note: PAC: Protective antiboties sufficient; VC: Vaccination; AC: CPV-2 antibody
content

229

Dome
stic

9

After VC (months)

12
Before
VC
1
3
6
9
12

V3

No. of
samples
tested

Breed

Before
VC
1
3

V2

No.
of
sampl
es
with
PAC

Foreig
n

Rate
(%)

AC
(HI
titer)

No.
of
sampl
es
with
PAC

Rate
(%)

AC
(HI
titer)

Table 4.10 showed that antibody contented generated by 01, 03,
06, 09 and 12 months after vaccination of 3 vaccines according to
domestic dog breed and foreign dog breed group were not statistically
significant (P>0.05). The results also showed that the immune response
to CPV-2 vaccine were not depend on the dog breed.
Chapter 5: CONCLUSIONS AND SUGGESTION
5.1 Conclusion
From the results of the survey, analysed and applicatially drewn
the following conclusions:

22

P>
0.05