Study on epidemiological characteristics of fasiolosis in buffaloes and bovines in thai nguyen, bac kan, tuyen quang province and preventive and treatment methods (2010 2013)

MINISTRY OF EDUCATION AND TRAINING
THAI NGUYEN UNIVERSITY


PHAM DIEU THUY


STUDY ON EPIDEMIOLOGICAL CHARACTERISTICS OF
FASCIOLOSIS IN BUFFALOES AND BOVINES IN THAI
NGUYEN, BAC KAN, TUYEN QUANG PROVINCE AND
PREVENTIVE AND TREATMENT METHODS
(2010 - 2013)






SUMMARY OF PH.D. DISSERTATION IN
VETERINARY MEDICINE



Thai Nguyen, 2014
MINISTRY OF EDUCATION AND TRAINING
THAI NGUYEN UNIVERSIY


PHAM DIEU THUY

STUDY ON EPIDEMIOLOGICAL CHARACTERISTICS OF

FASCIOLOSIS IN BUFFALOES AND BOVINES IN THAI
NGUYEN, BAC KAN, TUYEN QUANG PROVINCE AND
PREVENTIVE AND TREATMENT METHODS
(2010 - 2013)



Speciality: Parasites and Veterinary Microbiology
Code: 62.64.01.04

SUMMARY OF PH.D. DISSERTATION IN
VETERINARY MEDICINE
ADVISOR: Prof. Nguyen Thi Kim Lan, Ph.D.




Thai Nguyen, 2014



3

LIST OF WORKS RELATED TO THE DISSERTATION

1. Nguyen Thi Kim Lan, Pham Dieu Thuy, Pham Thi Trang
(2012), “Infection rate and intensity of Fasciola gigantica in
buffaloes and bovines in Thai Nguyen”, Animal husbandry magazine
July, pages 19 - 23.
2. Pham Dieu Thuy, Nguyen Thi Kim Lan, Hoang Thi Ngan

(2012), “Status of Fasciola infection in buffaloes and bovines in Bac
Kan province”, Animal husbandry magazine, August, pages, 26 - 31
3. Pham Dieu Thuy, Nguyen Thi Kim Lan, Tran Nhat Thang,
Tran Thi Phuong Thao (2014), “Study on egg viability and
development time of Fasciola gigantica larvae in the external
environment and in the intermediate host”, Journal of Agriculture
and Rural Development, June, pages 122 - 126.
4. Pham Dieu Thuy, Nguyen Thi Kim Lan, Tran Nhat Thang,
Tran Thi Phuong Thao (2014), “Study on the contamination of
Fasciola gigantica eggs and larvae in the environment”, Veterinary
sciences and Techniques, Vol. XXI, No6, pages 76 - 81.




1

INTRODUCTION

The Fascioliasis in buffaloes and bovines (Fassciolosis) caused
by two liver fluke species namely Fasciola hepatica and Fasciola
gigantica, is considered as a common parasitic disease which
causes great losses in the economy of buffalo and bovine
production in the world (Soulsby E. J. L., 1987). In recent years,
the Fascioliasis in buffaloes and bovines has occurred widely and
increased in the number due to the climate change and the
migration of animals between countries and/or regions (Kasib
Muhammad Khan et al., 2013).
There are some studies on the epidemiological characteristics of
fascioliasis in buffaloes and bovines and the effectiveness of the

treatment. But, only few works has been paid which considers the
issue in the mountainous area such as Thai Nguyen, Bac Kan and
Tuyen Quang province. Therefore, an effective measures of
prevention and the control of this diease in the mountainous area are
obviously needed.
CHAPTER 1
OVERVIEW

CHAPTER 2
MATERIALS, CONTENTS AND METHODS OF STUDY

2.1. Objects of study
- Buffaloes and bovines in three provinces: Thai Nguyen, Bac
Kan and Tuyen Quang
- Fasciolosis in buffaloes and bovines.
2.1.2. Study period: From 2010 - 2013
2.1.3. Places of study
2.1.3.1. Places of implementation: Thai Nguyen, Bac Kan and Tuyen
Quang province.
2.1.3.2. Places of samples



2

- Laboratory of Veterinary and Animal Science faculty - College
of Agriculture and Forestry, Thai Nguyen University
- Institute of Ecology and Biological Resources.
- Sperstructure Laboratory - National Institute of Hygiene and
Epidemiology.

2.2. Materials of study
2.2.1. Materials and samples of study
* Animals of study: buffaloes and bovines at diffenrent age, and
freshwater snails
* Samples of study: samples of fresh feces from Buffaloes and
bovines; samples of soil from animal house (sediment) floors, aquatic
grass samples, samples of snails without covers of the mouth,
Fasciola samples collected from necropsied buffaloes and bovines,
and bile samples, etc.
2.2.2. Instruments and chemicals: Light microscopy, counting
chamber Mc Master, cattle bedding, jars for specimen, surgical
instrument set, glass sinks, glass pots, plastic trays, enamel tray,
anthelmintic drugs against liver fluke, etc.
2.3. Contents of study
2.3.1. Determining parasitic liver flukes in buffaloes and bovines in
three Northern mountainous provinces
2.3.2. Epidemiological characteristics of fasciolosis caused by F.
gigantica in buffaloes and bovines
2.3.2.1. Investigating current status of animal husbandry and the
prevention of buffaloes and bovines from parasiolosis in three
studied provinces
2.3.2.2. Situation of F. gigantica infection in buffaloes and bovines
2.3.2.3. Study on eggs and larvae of F. gigantica in the external
environment and the intermediate host
2.3.3. Studying correlation between the number of F. gigantica
eggs in 1 gram of feces with the number of parasitic flukes
buffaloes and bovines

3

2.3.4. Studying prevention and control measures of fasioliasis
caused by F. gigantica in buffaloes and bovines
2.4 Methods of study
2.4.1. Method of necropsy and examination are used to collect and
identify F. gigantica species in buffaloes and bovines in Thai
Nguyen, Bac Kan and Tuyen Quang province
2.4.2. Method of investigating veterinary hygiene and prevention of
fascioliasis caused by F. gigantica in cattle and bufaloes
2.4.3. Methodologies for studying epedemic charateristics of
Fascipla spp. in buffaloes and bovines: use of Benedek sedimentation
method (1943) for examination of Fasciola eggs.
2.4.4. Studying eggs and larvae of F. gigantica in the external
environment and the intermediate host
2.4.5. Studying lifetime of F. gigantica eggs in the external
environment (when not released into water)
2.4.6. Study of time of miracidium released from eggs and survival in water
2.4.7. Study on the duration of F. gigantica larval in Lymnae viridis
snails - intermediate hosts
2.4.8. Method of determining correlation between the number of F.
gigantica eggs in 1 gram of feces and the number of parasitic liver
flukes in buffaloes and bovines.
2.4.9. Method of determining the effect of medicines against F.
gigantica
2.4.10. Trials on preventive measures of fascioliasis caused by F.
gigantica in buffaloes and bovines.
- Place of implementation: Dong Hy district - Thai Nguyen
Nguyen province.
- Trial object: buffalo at 2 - 4 years old.

Content:
* Arranging the test



4

* Determining the rate and intensity of infection F. gigantica of
bufalo in 2 months, 4 months after the trial
* Sugegesting the prevention and control procedures of
fascioliasis caused by F. gigantica in buffaloes and bovines.
2.5. Methods of data processing
The data acquired were analyzed and processed using the method
of Biostatistics (according to Nguyen Van Thien, 2008) based on
Excel 2003 and Minitab 14.0 software.
CHAPTER 3
RESULTS AND DISCUSSION

3.1. Results of the species determination of parasitic Fasciola spp.
in buffaloes and bovines in three provinces (Thai Nguyen, Bac
Kan and Tuyen Quang)
Table 3.1. Results of the necropsy examination of buffaloes and
the collection of liver flukes
Place
(province)
Number of
necropsied
buffaloes
(bufalo)
Number of

infected
buffaloes
(bufalo)
Percentage
(%)
Infection intensity

(liver
fluke/bufalo)

Thai Nguyen 150 89 59.33 5 - 89
Bac Kan 150 78 52.00 3 - 72
Tuyen Quang 150 97 64.67 6 - 78
Total
450 264 58.67 3 - 89
Table 3.2. Results of the necropsy examination of bovines and the
collection of liver flukes
Place
(province)
Number of
necropsied
bovines
(bovine)
Number of
infected
bovines
(bovine)
Percentage
(%)
Infec

tion intensity
(liver
fluke/(bovine)

Thai Nguyen 20 8 40.00 7 - 52
Bac Kan 20 6 30.00 1 - 45
Tuyen Quang 20 8 40.00 4 - 64
Total
60 22 36.67 1 - 64



5

The results in table 3.1 and 3.2 show that at post - mortem
examination of 450 buffaloes and 60 bovines in three provinces, the
infection rate in buffaloes and bovines was 58.67% and 36.67%,
respectively.
Table 3.3. Indentification of parasitic liver fluke species in
buffaloes and bovines
Results of species determination
Fasciola
gigantica species
Fasciola
hepatica
species
Intermidiate
forms between
the 2 species
Place

(province)
Number of
liver flukes
for
determinat
ion
(fluke)
Number
of
flukes
%
Number
of
flukes
%
Number
of
flukes
%
Thai Nguyen 262 258 98.47 0 0 4 1.19
Bac Kan 356 347 97.47 0 0 9 2.29
Tuyen Quang 538 517 96.10 0 0 21 4.06
Total
1156 1122 97.06 0 0 34 2.94
Table 3.3 shows that in the total of 1156 liver fluckes
classified, 97.06% of them was Fasciola gigantica, this was
from 96.10% - 98.47% between the provinces, and there was no
fluke belonging to Fasciola hepatica species. However, there
were 34 flukes (2.94%) in the intermediate forms between the
two species (F. gigantica and F. hepatica) (these flukes had

"shoulder" but not obvious). So we had redefined these samples
by biological molecular methods. The result of DNA sequencing
of three reprenstative samples showed that 99% of the samples
were homologous with the gen bank of F. gigantica. Thus, the
flukes in the intermediate forms were also F. gigantica species.
Comparing nucleotide sequence and amino acid sequence, it is
obvious that 2 samples of F. gigantica having the same sequences.
The other one had 5 nucleotides and 3 amino acids different



6

compared with the 2 samples mentioned above. So the intermediate
forms were also F. gigantica species.
3.2. Epidemiological characteristics of fascioliasis caused by
Fasciola in buffaloes and bovines
3.2.2. Situation of F. gigantica infection in three Northern
mountainous provinces
3.2.2.1. Infection rate and intensity of F. gigantica in buffaloes and
bovines in Thai Nguyen, Bac Kan and Tuyen Quang province
Table 3.6. Infection rate and intensity of F. gigantica in buffaloes
in the places studied
Infection intensity
(eggs/g feces)
≤
200
> 200 - 500 > 500
Place
(province,

district)
Number
of
buffaloes
examined
(buffalo)
Number
of
infected
buffaloes
(buffalo)
Percentage
(%)
n % n % n %
* Thai Nguyen
1800 851 47.28
b
447 52.52 262 30.79 142 16.69
Dong Hy
600 335 55.83
c
162 48.36 111 33.13 62 18.51
Vo Nhai
600 238 39.67
a
151 63.45 58 2437 29 12.18
Dai Tu
600 278 46.33
b
134 48.20 93 33.45 51 18.35

χ
2
= 31.779; P = 0.000


* Bac Kan
1800 733 40.72
a
461 62.89 191 26.06 81 11.05
Cho Moi
600 320 53.33
b
186 58.13 102 31.87 32 10.00
Bach Thong
600 190 31.67
a
141 74.21 30 15.79 19 10.00
Ngan Son
600 223 37.17
a
134 60.09 59 26.46 30 13.45
χ
2
= 63.055; P= 0.000

* Tuyen Quang
1800 934 51.89
c
568 60.81 289 30.94 77 8.25
Yen Son

600 336 56.00
b
228 67,86 89 26.49 19 5.65
Ham Yen
600 275 45.83
a
166 60.36 82 29.82 27 9.82
Tuyen Quang
city
600 323 3.83
b
174 53.87 118 36.53 31 9.60
χ
2
= 13.784; P = 0.001

5400 2518 46.63 1.476 58.62 742 29.47 300 11.91
Total
χ
2
= 45.551; P = 0.000
* Notes: In vertical line, the numbers with superscriptions are
different remarkably in stastistics.



7

Table 3.6 shows that in three provinces the infection rate of F.
gigantica in buffaloes was 46.63%, ranging from 40.72% to 51.89%.

The difference between the three provinces was significant (P <
0.001). Particularly, in Tuyen Quang, the infection rate in buffaloes
was highest (51.89%), followed by Thai Nguyen (47.28%), the
lowest was in Bac Kan proince (40.72%).
In general, the infection intensity in buffaloes was mild. However,
buffaloes infected intensity was accounted for 58.62%; the average intensity
made up 29.47%; while the severe intensity accounted for 11.91%.
Table 3.7. Infection rate and intensity of F. gigantica in bovines in
the places investigated
Infection intensity
eggs/g feces)
≤
200
> 200 - 500 > 500
P
laces
(province,
district)
Number of
bovines
examined
(bovine)
Number of
infected
bovines
(bovine)
Percentage

(%)
n % n % n %

* Thai Nguyen

630 243 38.57
b
144 59.26 75 30.86 24 9.88
Dong Hy
210 110 52.38
b
47 42.73 44 40.00 19 17.27
Vo Nhai
210 63 30.00
a
45 71.43 15 23.81 3 4.76
Dai Tu
210 70 33.33
a
52 74.29 16 22.86 2 2.85
χ
2
= 25.846; P = 0.000

* Bac Kan
630 188 29.84
a
123 65.43 51 27.13 14 7.44
ChoMoi
210 80 38.10
b
48 60.00 27 33.75 5 6.25
Bach Thong

210 69 32.86
b
52 75.36 12 17.39 5 7.25
Ngan Son
210 39 18.57
a
23 58.97 12 30.77 4 10.26
χ
2
= 20.485; P= 0.000

Tuyen
Quang

630 225 35.71
b
136 60.45 77 34.22 12 5.33
Yen Son
210 67 31.90 39 58.21 25 37.31 3 4.48
Ham Yen
210 75 35.71 46 61.33 23 30.67 6 8.00
Tuyen Quang
city
210 83 39.52 51 61.45 29 34.94 3 3.61
χ
2
= 2.655; P = 0.265

1.890 656 34,71 403 61.43 203 30.95 50 7.62
Total

χ
2
= 11.015; P = 0.004
* Notes:I vertical line, the numbers with superscription are different
remarkably in statistics.
Table 3.7 shows that the infection rate of F. gigantica in bovines was
34.71% (much lower than buffaloes), the highest rate found in Thai Nguyen



8

(38.57%), and the lowest rate was in Bac Kan province (29.84%). However,
the differences in prevalence of bovines Fasciola in Thai Nguyen and Tuyen
Quang were not significant (P > 0.05). In different districts the prevalence was
apparent (P < 0.001), but in Tuyen Quang province, the differences in
prevalence between districts were not clear (P > 0.05). In term of infection
intensity, there was lighter and more moderate intensity in bovines compared
with buffaloes, but the severe infection intensity was less.
3.2.2.3. Infection rate and intensity of F. gigantica in buffaloes and
bovines acording to seasons
Table 3.10. Infection rate and intensity of F. gigantica in
buffaloes according to seasons
Infection intensity
(eggs/g feces)
≤
200
> 200 - 500 > 500
Places

(province)

Season
Number of
buffaloes
examined
(buffalo
Number
of
infected
buffaloes

(buffalo)

Percentage
(%)
n % n % n %
Sping 450 173 38.44
a
78 45.09

63 36.42

32 18.49

Summer

486 291 59.88
c
165

56.70

80 27.49

46 15.81

Autumn

414 218 52.66
b
117

53.67

69 31.65

32 14.68

Winter 450 169 37.56
a
87 51.48

50 29.59

32 18.93

Thai
Nguyen
χ

2
= 66.906; P= 0.000

Sping 378 115 30.42
a
80 69.57

31 26.96

4 3.47
Summer

468 245 52.35
c
146

59.59

66 26.94

33 13.47

Autumn

450 202 44.89
b
119

58.91

54 26.73

29 14.36

Winter 504 171 33.93
a
116

67.84

40 23.39

15 8.77
Bac Kan
χ
2
= 55 697; P = 0.000

Sping 503 254 50.50
b
136

53.54

93 36.61

25 9.85
Summer

453 271 59.82

c
187

69.00

63 23.25

21 7.75
Autumn

412 220 53.40
bc
138

62.73

67 30.45

15 6.82
Winter 432 189 43.75
a
107

56.61

66 34.92

16 8.47
Tuyen
Quang

χ
2
= 23.653; P = 0.000

Sping
1.331 542 40.72
a
294

54.24

187

34.5 61 11.26

Summer

1.407 807 57.36
c
498

61.71

209

25.9 100

12.39

Autumn

1.276 640 50.16
b
374

58.44

190

29.69

74 11.87

Winter
1.386 529 38.17
a
310

58.60

156

29.49

65 11.91

Total
χ
2
= 129.022; P = 0.000

* Notes: in vertical line, the numbers that carry different letters are
different in stastistic signinficance.



9

The results in table 3.10 shows that regarding infection rate:
overall, the infection rates of liver flukes in buffaloes in three
provinces were different with seasons. The highest rate was in
Summer (57.36%), followed by Autumn (50.16%); Spring (40.72%)
and the lowest was in Winter (38.17%). However the infection rate of
liver flukes in buffaloes between Winter and Spring was not
markedly different (P > 0.05).
* Regarding the intensity of infection:
Buffaloes infected mainly in mild and moderate intensity in all four
seasons of the year. In severe intensity the infection rates of liver flukes in
buffaloes and bovines in summer and were the lowest in Winter.
3.2.2.4. Infection rate and intensity of F. gigantica in buffaloes and
bovines with different sex.
Table 3.13. Infection rate and intensity of F. gigantica in
buffaloes with sex of cattle
Infection intensity
(eggs/g feces)
≤
200
> 200
- 500 > 500
Places

(province)
Sex
Number of
bovine
examined
(cattle)
Number of
infected
cattle(cattle)
Percentage
(%)
n % n n % n
Bull
239 85 35.56 51 60.00

26 30.59

8 9.41
Thai Nguyen

Cow
391 158 40.41 93 58.86

49 31.01

16 10.13

Bull
284 86 30.28 53 61.63

28 32.56

5 5.81
Bac Kan
Cow
346 102 29.48 70 68.63

23 22.55

9 8.82
Bull
164 58 35.37 34 58.62

21 36.21

4 5.17
Tuyen
Quang
Cow
466 167 35.84 102

61.08

56 33.53

8 5.39
Bull
687 229 33.33 138

60.26

75 32.75

17 6.99
Cow

1203 427 35.49 265

62.06

128

29.98

33 7.96
Total
χ
2
= 0.901; P = 0.342
Table 3.13 shows that the infection rates of F. gigantica in bulls
and cows were 33.33% and 35.49% respectively. Statistical



10

comparing shows that the prevalence between bulls and cows was not
markedly different (P > 0.05). Statistically there were also no
significant differences of infection intensity between bulls and cows.
This result was consistent with study findings on cattle by Pham

Ngoc Vinh, Nguyen Trong Kim (1997). The study results on goats by
Nguyen Thi Kim Lan et al. (1999). Khan M. K. et al. (2009).
3.2.3. Study on eggs and larvae of F. gigantica in the external
environment and in intermediate hosts
3.2.3.2. Contamination of F. gigantica eggs on grazing land
Table 3.16. Contamination of F. gigantica eggs in grazing land of
buffaloes and bovines
Top soil from grazing land Pool
Place
(province
, district
)
Number of
samples
examined

Number of
positive
samples (+)

Percentage
(%)
Number of
samples
examined

Number of
positive
samples (+)

percentage

(%)
* Thai Nguyen 450 33 7.33 450 61 13.56
Dong Hỷ 150 11 7.33 150 20 13.33
Vo Nhai 150 10 6.67 150 16 10.67
Dai Tu 150 12 8.00 150 25 16.67
* Bac Kan 450 38 8.44 450 63 14.00
Cho Moi 150 14 9.06 150 23 15.33
Bach Thong 150 13 8.84 150 25 16.67
Ngan Son 150 11 7.56 150 15 10.00
* Tuy
enQuang 450 42 9.33 450 78 17.33
Yen Son 150 16 10.67 150 32 21.33
Ham Yen 150 14 9.33 150 24 16.00
Tuyen
Quang city
150 12 8.00 150 22 14.67
1.350 113 8.37 1.350 202 14.96
Total
χ
2
= 1.178; P = 0.555 χ
2
= 3.016; P = 0.221

Table 3.16 shows that overall 8.37% of the samples of grazing
land surface and 14.96% of water sampes from the hollows on
grazing land in the places of the three provinces under study were
contaminated with liver fluke germs.

11

3.2.3.3. The distribution of freshwater snails - intermediate hosts of
F. gigantica
Table 3.17. Results of freshwater snail classification
Lymnaea viridis
species
Lymnaea
swinhoei
species
Other species*
Places
(province, district)

Number of
snails
classified
(snail)
n (%) n (%) n (%)
* Th
ai Nguyen 2160 768 35.56 621 28.75 771 35.69
Dong Hỷ 720 224 31.11 195 27.08 301 41.81
Vo Nhai 720 293 40.69 235 32.64 192 26.67
Dai Tu 720 251 34.86 191 26.53 278 38.61
* B
ac Kan 2160 663 30.70 437 20.23 1.060 49.07
Cho Moi 720 221 30.69 174 24.17 325 45.14

Bach Thong 720 230 31.94 147 20.42 343 47.64
Ngan Son 720 212 29.45 116 16.11 392 54.44
*Tuy
en Quang 2160 520 24.07 978 45.28 662 30.65
Yen Son 720 234 32.50 286 39.72 200 27.78
Ham Yen 720 120 16.67 365 50.69 235 32.64
Tuyen Quang city 720 166 23.05 327 45.42 227 31.53
T
otal
6480 1951 30.11 2036 31.42 2493 38.47

Table 3.17 shows that in 6480 snails collected 30.11% belonged
to the L. viridis species; 31.42% belonged to L. swinhoei species
38.47% were the other snail species (not the intermediate host of
liver fluke). Thus more than 60% of the snails collected belonged to
two species L. viridis and L. swinhoei. According to Dang Ngoc
Thanh et al. (1980), Nguyen Trong Kim and Pham Ngoc Vinh
(1997), intermediate hosts of Fasciola in Northern part of Viet Nam
are two snails species: L. viridis and L. swinhoei. Results of snail
classification in Thai Nguyen, Bac Kan and Tuyen Quang showed
that two snail species accounted for a high proportion of snails
collected. Common distribution with large numbers of two snail
species creates favorable conditions for F. gigantica complete their
life cycle.



12

3.2.3.4. The prevalence of liver fluke larvae in freshwater snails -

intermediate hosts
Table 3.19. The prevalence of F. gigantica larvae in freshwater
snails
Place (province)
Description

Thai Nguyen

Bac Kan

Tuyen Quang

Snail species

Lymnaea
viridis
Lymnaea
swinhoei
Lymnaea
viridis
Lymnaea
swinhoei
Lymnaea
viridis
Lymnaea
swinhoei
Number of snails
examined (snail)
768 621 663 437 520 978
Number of snails

infected with
F. gigantica
larvae(snail)
108 40 173 41 208 318
Percentage (%)
14.06 6.44 26.09 9.38 40.00 32.52
n (snail)
55 13 95 18 76 124
Sporocyst
(%)
50.93 32.50 54.91 43.90 36.54 38.99
n (snail)
63 19 97 25 96 131
Redia
%
58.33 47.50 56.07 60.98 46.15 41.19
n (con)
66 17 104 22 103 141
Cercaria
(%)
61.11 42.50 60.12 53.66 49.52 44.34
Table 3:19 shows that on the prevalence of larvae in snails there were
14.06% of L. viridis and 6.44% of L. swinhoei in Thai Nguyen; 26.09% of
L. viridis 9.38% L. swinhoei in Bac Kan; 40% of L. viridis and 32.52% of
L. swinhoei in Tuyen Quang being infected with F. gigantica larvae. Thus,
both species of snails were infected with liver fluke larvae ranging 20% -
70%. In which L. viridis was infected with liver flukes with significantly
higher rate compared with L. swinhoei (this shows in all three provinces
studied). It is believed that L. viridis species was more susceptible to F.
gigantica compared with L. swinhoei.

- Regarding the prevalence of larval forms of F. gigantica in snail:
L. viridis was infected with sporocyst larvae from 36.54% to 54.91%; Redia
larvae from 46.15% to 58.33% and Cercaria larvae 49.52% to 61.11%. L.
swinhoei infected with Sporocyst larvae from 32.50% to 43.90%; Redia larvae
from 41.19% to 47.50% and Cercaria larvae from 42.50% to 53.66%.



13

3.2.3.8. Study on the time of Miracidium hatched released and
surviving in water
Table 3.24. Time of Miracidium hatched and released into
water (from releasing eggs in water)
Initial time of
Miracidium
releaseed from
eggs
Time for
all Miracidium
released from eggs
Season

E
xperiment
Number
of
samples

Temper

ature
and pH
water
(
)
X
X
m
±

(day)
total
(day)

(
)
X
X
m
±

(day)
total
(day)

I 10
15.40 ± 0.45

14.40 ± 0.36

II 10
14.60 ± 0.41

16.80 ± 0.25

Spring

III 10
22 - 23
o
C,
6 - 7
15.20 ± 0.25

14
15.20 ± 0.62

16
I 10
8.80 ± 0.33

9.70 ± 1.58
II 10
8.90 ± 0.31

8.30 ± 1.78
Summer
III 10
26 - 27
o

C,
6 - 7

9.50 ± 0.23

8
10.10 ± 2.18

10
I 10
12.00 ± 0.58

14.10 ± 2.79

II 10
13.70 ± 0.30

12.20 ± 2.81

Autumn

III 10
24 - 25
o
C,
6 - 7

11.40 ± 0.21

11

15.40 ± 2.92

15
I 10
20.00 ± 0.47

19.70 ± 0.63

II 10
19.30 ± 0.74

21.10 ± 0.79

Winter

III 10
18 - 19
o
C,
6 - 7
19.90 ± 0.42

19
20.30 ± 1.57

21

Table 3.24 shows that in Spring Miracidium hached and released
from egg earliest was 14 days, no later than 16 days; In Summer the
earliest was 8 days and no later than 10 days; In Autumn, the earliest

time was 11 days and no later than 15 days; In Winter the earliest
was 19 days and no later than 21 days. Thus, the time required for
development from the egg into the water to hatch into miracidium
varied from 8 to 21 days. In winter, due to cold weather the
development of eggs in water lasted longer. Whereas, in Summer the
weather was hot, the development of eggs was markedly shorter.



14

3.2.3.9. Study of time for development of F. gigantica larvae in snail
- intermediate hosts
Table 3.26. Time to complete the larval stages of F. gigantica
(from the time the eggs fall into the water)

Season
Number of
samples
Stage of larvae
Time to
complete (day)
Egg à Miracidium 14 - 16
Miracidium à Sporocyst 3 - 4
Sporocyst à Redia 6 - 8
Redia à Cercaria 18 - 20
Cercaria à Adolescaria 2 - 4
*
Spring 5
* Egg à Adolescaria 41 - 48

Egg à Miracidium 8 - 10
Miracidium à Sporocyst 2 - 3
Sporocyst à Redia 4 -7
Redia à Cercaria 15 - 17
Cercaria à Adolescaria 2 – 4
*
Summer 5
* Egg à Adolescaria 29 - 37
Egg à Miracidium 11 - 15
Miracidium à Sporocyst 3 – 4
Sporocyst à Redia 5 – 8
Redia à Cercaria 17 - 18
Cercaria à Adolescaria 2 - 4
*
Atumn 5
* Egg à Adolescaria 36 - 45
egg à Miracidium 19 - 21
Miracidium à Sporocyst 4 - 5
Sporocyst à Redia 8 - 9
Redia à Cercaria 20 - 21
Cercaria à Adolescaria 3 - 5
*
Winter 5
* Egg à Adolescaria 51 - 56
The results in table 3.26 shows that in winter, the time of
developing egg and larval forms in the snail were the longest. The
rule of the development time of larvae in intermediate hosts with
seasons reveals that the temperature is an important factor and a
major influence on the development of eggs and larvae of F.
gigantica in the external environment and in intermediate hosts.

15

x
y
9080706050403020100
800
700
600
500
400
300
200
100
0
3.3. Studies of correlation between number of parasitic F. gigantica in
buffaloes and bovines with number of eggs in 1 g of feces.
* The correlation between the number of parasitic Fasciola
/buffalo with number of eggs in 1 gram of feces was determined on
Minitab 14.0 software. The results were as follows:
Linear regression equation: y = a + bx
(y: number of eggs / g of feces. x: number of parasitic Fasciola / buffalo)
Among them: a = 0.194
b = 8.101
→ linear regression equation: y = 0.194 + 8.101 x
Correlation coefficient: r = 0.96
The correlation between the number of parasitic Fasciola / buffalo
and number of eggs /g feces is illustrated in Figure 3.10

Figure 3.10. The graph illustrating the equation y = a + bx of the
relationship between the number of parasitic Fasciola /buffalo
with number of eggs /g faeces.
The graph 3.10 shows that number of corresponding points
between the parasite count through buffaloes at postmortern
examination with number of eggs in 1 gram of feces were mostly
located around the curvature of the linear regression equation y = a +
bx, away from lower left to upper right, there were no points lying far
from this curvature. That means that the correlation between the



16

x
y
706050403020100
500
400
300
200

100
0
number of parasitic Fasciola / buffalo and number of eggs /g feces
was correlated.
The correlation coefficient of r = 0.96 indicates a very close
correlation.
* Correlation between number of eggs /g faeces and number of
parasitic Fasciola / bovine was as follows:
The linear regression equation y = a + bx
(y: number of eggs/g faeces. x: number of parasitic Fasciola /bovine)
Among them: a = 4.145
b = 8.094
→ the linear regression equation: y = 4.145 + 8.094x
The correlation coefficient: r = 0.969
Correlation between number of F. gigantica eggs /g faeces and
number of parasitic Fasciola /bovine is illustrated in figure 3.11.











Figure 3.11. The graph illustrating the equation y = a +
bx of the relationship between number of parasitic Fasciola
/bovine with number of eggs /g faeces.

The graph 3.11 shows that number of corresponding points
between the parasite count through bovines at necropsy with number
of eggs in 1 gram of feces were mostly located around the curvature
of the linear regression equation y = a + bx, from lower left to upper
right, there was only one point lying a bit far from this curvature.



17

That means that the correlation between the number of parasitic
fasciola /bovines and number of eggs /g of feces was direct
correlated.
The correlation coefficient r = 0.969 shows that this is closely
corelated.
3.4. Study of prevention and treatment measures of fascioliasis
caused by F. gigantica in buffaloes and bovines
3.4.1. Identifying drug for prevention and treatment of fascioliasis
caused by F. gigantica that is highly effective and safe
3.4.1.2. Trial on efficacy and safety of some drugs against F. gigantica in
buffaloes and bovines
* Testing on a small scale
Table 3.30. Efficacy of three drugs against F. gigantica in
experimental bovine
Day after
treatment
(eggs / g feces)
Necropsy after treatment

Drugs and dosage

Order
number

of bovine
Prior to
treatment

(eggs / g
feces)
5 10 15
Day
at necropsy
after
treatment

Comparison

sluke/cattle
(fluke)
1 285 30 0 0 -
2 315 30 0 0 -
3 375 45 0 0 -
4 405 60 0 0 35 0
Albendazol
(12mg/kg B.W)
5 255 15 0 0 -
1 450 55 0 0 -
2 410 60 0 0 25 0
3 380 40 0 0 -
4 295 30 0 0 -

Triclabendazole
(15 mg/kg kg

B.W)
5 320 35 0 0 -
1 260 25 0 0 -
2 350 35 0 0 -
3 300 30 0 0 30 0
4 430 45 0 0 -
Nitroxinil - 25
(12mg/kg kg B.W)

5
330 40 0 0
-




18

Table 3.30 shows that three drugs used for treatment of cattle
infected with F. gigantica at dose above, efficacy achieved similar to
the test on buffaloes (15/15 removed parasite eggs in feces after 10
days of dosing, 3/15 of cattle that were necropsied to examine liver
flukes in bile ducts after 25-35 days using dewormers). At the same
time, all 15 cows were no signs of any unusual after dosing. Thus,
doses of three drugs used for cattle were safe.
* Trial on a large scale
We tested the efficacy of triclabendazole treatment at dose 15 mg

/kg B.W albendazole and nitroxinil - 25 at dose 12mg /kg B.W in 450
buffaloes and 270 bovines in the studied places. The results are
illustrated in table 3.31 and 3.32.
Table 3.31. Efficacy of three dewormers angainst F. gigantica
in cattle on a large scale
Before dewworming 15 days after deworming Dewworming eficacy
Time

Type of drug
Dose (mg/

kg B.W)
Number of
infected
buffaloes
(buffaloes)

Infection intensty

(
x
mX ±
)
(eggs/ feces g )
Number of
infected
buffaloes
(buffaloes
Intensity
(

x
mX ±
)
(eggs/ feces g )
Number of
buffaloes
cleared from
eggs
(buffalo)
Efficacyy
(%)
Albendazol

12 50 465.00 ± 56.00

3 45.60 ±10.30

47 94.00
Triclabendazole

15 50 418.30 ± 48.65

0 0 50 100
I
Nitroxinil
- 25
12 50 445.28 ± 30.50

3 35.20 ±8.70 47 94.00
A

lbendazol
12 50 520.80 ± 53.10

5 60.80 ±14.40

45 90.00
Triclabendazole

15 50 560.00 ± 46.50

0 0 50 100
II
Nitroxinil
- 25
12 50 490.35 ± 28.50

4 50.00 ±13.40 46 92.00
Albendazol

12 50 420.30 ± 46.00

5 25.80 ±6.72 45 90.00
Triclabendazole

15 50 380.50 ± 60.78

0 0 50 100
III
Nitroxinil
- 25

12 50 415.27 ± 23.60

5 40.50 ± 9.86 45 90.00
Effectiveness of albendazole was quite high, reaching 90 - 94%.
the infection intensity decreased from 420-465 eggs /g feces to 25-
60 eggs /g faeces. Nitroxinil - 25 also had althelmintic effectiveness
reaching 90 - 94%, reducing infection intensity from 415 - 490 eggs



19

The results in table 3:31 shows that in all three treatments.
triclabendazole has the highest althelmintic efficacy (100%). 15 days
after dosing, examining feces, eggs were not detected. The
althelmintic /g of feces to 35-50 eggs /g of feces 15 days after
deworming. Observing buffaloes before and after dosing, we found
that the activities of the whole buffaloes were normal, eating and
ruminating normally, no buffaloes had any side effects after receiving
the drug. So we evaluated: dose levels of three drugs used were
100% safe for cattle.
3.4.2. Trial on therapeutic measures in treating fascioliasis
caused by F. gigantica in buffaloes
Infection rate and intensity in buffaloes of the experimental group
and the control prior to the experiment are illustrated in table 3.33.
Table 3.33. Infection rate and intensity of F. gigantica in
buffaloes prior to the experiment
Group

Description

Experime
ntal group

Control
Statistical
significance
(P)
Number of buffaloes examined
(buffalo)
80 80 -
Number of infected buffaloes
((buffalo)
31 30 -
Percentage (%)
38.75 37.50 > 0.05
n
18 17 -
≤ 200
%
58.06 56.67 > 0.05
n
11 11 - >200
- 500
%
35.48 36.67 > 0.05
n
2 2 -
Infection
intensity
(number of eggs/

gram of feces)
>
500
%
6.45 6.67 > 0.05
Table 3.33 shows that infection rate and intensity of F. gigantica
in the experimental group and control group prior the experiment
were equivalent
3.4.2.2. Infection rate and intensity of F. gigantica after 4 months
of the experiment



20

Table 3.35. Infection rate and intensity of F. gigantica after 4
months of the experiment
Group
Description
Experimental
group
Control
Statistical
significance (P)
Number of buffaloes examined
(buffalo)
80 80 -
Number of infected buffaloes
(buffalo)
14 36 -

Percentage (%)
17.50 45.00 < 0.001
n 13 24 -
≤ 200
% 92.86 66.67 < 0.001
n 1 10 -
>200 -500
% 7.14 27.78 < 0.001
n 0 2 -
Infection
intensity
(number of
eggs/ gram
of feces)
> 500
% 0.00 5.56 < 0.001
Table 3.35 shows that the infection rate after 4 months of
experiment, the prevalence of F. gigantica in buffaloes in the
experimental group was 17.50%, while the prevalence in the control
group was 45.00%. This difference was very clear (P <0.001).
The intensity of infection:
+ Mild infection intensity: 92.86% of buffaloes were infected with
liver fluke in experimental groups in light intensity, whereas in the
control group, mild prevalence was 66.67%, this difference was very
clear (P <0.001).
+ Average intensity of infection: In experimental group mean intensity of
infection was 7.14%, while that in the control group was 27.78%.
+ Heavy infection intensity: There were no buffaloes in
experimental group being infected in severe intensity, while the
proportion of severe infections in the control group was 5.56%. The

difference had statistical significance (P <0.001).



21

3.4.3. Recommendation of combining preventive and control
procedures to prevent fascioliasis caused by F. gigantica in
buffaloes.
From the results of the study we recommend combining
preventive and control procedures against fascioliasis in buffaloes
and bovines follows:
1 Deworming F. gigantica for buffaloes and bovines: Deworm
periodically liver fluke twice/ year (phase 1 in April- May; phase 2 in
September - October) for the whole herd of buffaloes and bovine
once examining feces shows that 20-30% of these animals are
infected with liver fluke.
Three drugs at dose levels having been tested (triclabendazole, at
dose 15 mg /kg B.W, albendazol and nitroxinil - 25, at dose of 12
mg /kg) all were effective in removing liver fluke. Depending on the
locale and on the specific conditions, one of those drugs to remove
liver fluke from buffaloes can be selected. However, the use of
triclabendazole is the best effective.
2 Treatment of cattle manure to kill eggs of F. gigantica: daily
collect manure from stable and put in pitches of manure composting,
or gather them in one place as a pile and cover with thick mud
contained 5 -10 cm
3 Cleaning cattle stables every 2 months, using disinfectants to
spray in order to kill F. gigantica eggs in the floor of cattle houses
and their surrounding areas.

4. Improving pastures, grazing land for buffaloes and bovines:
Removing stagnant water on pastures, collecting manure from
grazing land for composting are needed in order to limit the spread
and development of eggs and larvae of trematodes in the
environment.



22

5. Killing intermediate hosts of Fasciola, drainage of water,
drying muddy, wet pastures and grazing land are effective measures
to kill Lymnaea spp Fresh snails can be killed by using lime poder,
copper sulffate or rearing waterfowls (ducks, muscovy ducks, geese)
and black carps (mylopharyngodon piceus). However it depends on
place to apply appropriate measures in order to kill intermediate hosts
of Fasciola effectively.
6. Strengthening the care and feeding animals: For buffaloes and
bovines at different ages (especially calving cows), it is necessary to
ensure adequate rations in terms of quantity and quality. Cattle over 8
years old must be culled so that they will not shed F. gigantica eggs
into the environment.
CONCLUSIONS AND RECOMMENDATIONS
1. Conclusion
1- 450 buffaloes and 60 cattle in 3 provinces were necropsied, the
infection rate in buffaloes was 58.67% and in cattle was 36.67%.
F. gigantica is the only species of trematode parasitising liver and
bile ducts of buffaloes and bovines the places invstigated, common
prevalence is 100%
2 - Epidemiological characteristics:

+ Status of animal husbandry and prevention of parasitosis in
buffaloes and bovines in 3 provinces is not good, especially
preventive measures of parasitosis in buffaloes and cattle.
+ The prevalence of F. gigantica in buffaloes and bovinein three
provinces is 46.63%; in cattle is 34.71%. The rate and intensity of
infection increases with aging.
Buffaloes and bovines are infected with F. gigantica in larger
number and more severe in summer. The rate and intensity of
infection in the other seasons are lower and lighter.



23

Variation in the rate and intensity of infection of cows and bulls
are markedly irregular.
+ The areas around stables and barns of buffaloes and bovines are
contaminated with F. gigantica eggs at the rate of 20 - 43%.
8.37% of the soil samples from the surface of grazing land and
14.96% of water samples from the hollows on grazing land were
contaminated with F. gigantica eggs.
+ L. viridis and L. swinhoei - intermediate hosts of F. gigantica
are commonly distributed in all investigated places.
The infection rate of liver fluke larvae in snails varies 20 - 70%.
Infection rate of L. viridis is 61%.
+ Samples of aquatic grasses in Thai Nguyen. Bac Kan and Tuyen
Quang are contaminated with Adolescaria at the rate of 14 - 23%.
+ In Spring, survival of F. gigantica eggs lasts 12 - 48 days (in
dry feces), 43 - 125 days (in wet feces). Survival of eggs is shorter
than that in Summer, longer in Autumn and the longest in winter (19

- 80 days in dry feces, 75 - 160 days in wet feces).
The survival time of eggs is short in dry soil, longer in wet soil. In
Summer, the survival time of eggs in the ground is shorter than in
other seasons.
+ In Spring, Miracidium hatches and is releases from the egg
earliest in 14 days, no later than 16 days; in Summer time this is 8
and 10 days; in Autumn 11 and 15 days; in Winter 19 and 21 days.
After hatching from the egg, Miracidium in the water is not more
than 10 hours (in Spring), 11 hours (in Summer), 14 hours (in
autumn) and 9 hours (in Winter).
The time from releasing F. gigantica eggs in water until forming
Adolescaria is the longest in winter (51 - 56 days), followed by
Spring (41 - 48 days). Autumn (36 - 45 days) and the shortest is in
Summer (29 - 37 days).



24

3- Correlation between the number of eggs /g of feces with
number of parasitic liver flukes / animal is correlated quite closely, in
accordance with the linear regression equation y = a + bx.
4- At the recommeded doses, efficacy of han - dertin B and
fasciolid against F. gigantica is not high (78 - 86%).
Efficacy of triclabendazole against F. gigantica at dose of 15 mg
/kg B.W is 100%; Efficacy of albendazole and nitroxinil - 25 at the
same dose (12mg /kg B.W) reaches 90 - 96%. All of 3 drugs are safe
for cattle and buffaloes.
5- Prevention and control of F. gigantica in buffaloes and cattle
should be performed by using five synchronized key measures

(deworming liver flukes with triclabendazole, composting manure.
killing snails, intermediate hosts, sanitation of feed, water, stables
and grazing land), culling the animals older than 8 years old.
2. Recommendation
In the Northern mountainous provinces, cattle are raised under
grazing methods, utilizing the existence of natural conditions.
Therefore, it is needed to implement the combined prevention and
control of Fascioliasis gigantica for buffaloes and bovines as the six
contents of the above conclusion. This is believed to help reducing
the economic losses caused by the parasiolois disease, improving
livestock productivity, and promoting the industry of animal
husbandry.



25