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Development of Indirect ELISA for the Diagnosis of Bovine Hypodermosis (Hypoderma lineatum) in the Cattle of Subtropical Region of Pakistan

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Development of Indirect ELISA for the Diagnosis of Bovine Hypodermosis (Hypoderma lineatum) in the Cattle of Subtropical Region of Pakistan

Abstract and Figures

The aim of this study was to develop an indirect ELISA for the detection of Hypoderma lineatum antibodies and to determine the influence of seroepidemiological factors on the seroprevalence of bovine hypodermosis in cattle of subtropical region of Pakistan. For this purpose a total of 1000 blood samples were taken from twenty eight villages of Rawalpindi, Attock, Chakwal and Jhelum districts. First instar larvae (L-1) of the warble fly were collected from the surrounding abattoirs to obtain the antigen (HyC) used for development of indirect ELISA. The seroprevalence was 17.4% (174/1000). The highest seroprevalence was recorded in the Rawalpindi district (28.81%), followed by Attock (21.51%), Jhelum (10%) and Chakwal (6.66%). In present study the sensitivity and specificity was 97.28% and 96.44%, respectively. The epidemiological factors showed that the village having hilly locations, local breed, young animals, water bodies, extensive grazing, primary exposed and non medicated were more seropositive as compare to others (P<0.05). In conclusion, indirect ELISA developed during this study is very useful tool for early detection of bovine hypodermosis in cattle grazing in subtropical region of Pakistan.
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Kafkas Univ Vet Fak Derg
18 (Suppl-A): A215-A219, 2012
Summary
The purpose of the present study to determine the geographical distribution of Hypoderma in the Northern Punjab, Pakistan. One
thousand cows were examined by palpation method for the presence of warbles in dierent areas from northern Punjab from August
2010 to February 2011. The overall prevalence was 18.40% with significant dierences in the geographical distribution of Hypoderma,
when the grazing pattern, Topography, Water Bodies and Village were considered in dierent geographic areas. The prevalence was
higher in hilly and semi-hilly areas as compared to the plane areas.
Keywords: Hypodermosis, Geographical distribution, Cattle, Northern Punjab, Pakistan
Pakistan’ın Kuzey Pencab Bölgesinde Hypodermosis`in
(Hypoderma sp.) Coğrafi Dağılımı
Özet
Sunulan bu calışmanın amacı, Pakistan`ın Kuzey Puncab Bölgesi’nde Hypodermanın cografik dağılımını belirlemektir. Ağustos,
2010 ile şubat, 2011 arasında 1.000 sığır Kuzey Pencab`ın farklı böolgelerinde nokranın varlığını araştırmak için palpasyon yoluyla
incelenmiştir. Değişik coğrafik alanlardaki otlatma modeli, topoğrafya, sulak bölgeler ve köylerin durumuna göre ortalama prevalans
%18.4`tür. Prevalans tepelik ve yarı tepelik bölgelerde, ovalık alanlara göre daha yüksekti.
Anahtar sözcükler: Hypodermosis, Coğrafik dağılım, Sığır, Kuzey Pencab, Pakistan
Geographical Distribution of Hypodermosis (Hypoderma sp.)
in Northern Punjab, Pakistan
Haroon AHMED * Mobushir Riaz KHAN ** Rosario PANADERO-FONTAN ***
Ceferino López SANDEZ *** Muhammad Farooq IQBAL **
Syed Muhammad Saqlan NAQVI **** Mazhar QAYYUM *
*
**
***
****
Department of Zoology, PMAS Arid Agriculture University, Rawalpindi, 46300, PAKISTAN
Department of Meterology, Comsats Institute of Information Technology, Park Road, 46300, Islamabad - PAKISTAN
Departamento de Patología Animal: Sanidad Animal, Facultad de Veterinaria de Lugo, Universidad de Santiago de
Compostela, 27071 Lugo - SPAIN
Department of Biochemistery, PMAS Arid Agriculture University, Rawalpindi, 46300, PAKISTAN
Makale Kodu (Article Code): KVFD-2012-6533
Ectoparasitic infestations are one of the major
veterinary problems aecting livestock in many parts of
the world 1. Amongst these, the warble y infestation (WFI)
by Hypoderma and Przhevalskiana (Diptera: Oestridae) is a
notorious and common disease of cattle, bualoes and
goats in Pakistan 2.
Hypodermosis is a parasitic disease caused by the
development of the larval stages of insects of genus
Hypoderma. Each species of this genus is strictly a parasite
of a ruminant species. H. diana and H. actaeon are specific
of deer, H. tarandi of reindeer and H. bovis and H. lineatum
of cattle. Cattle hypodermosis is widely spread over each
continent of the northern hemisphere 3.
Hypodermosis is common in the USA, Canada and in
many parts of Africa and Europe, for example in Spain
where levels of parasitism have been estimated to be
between 26% and 42% depending upon the region
studied 4,5. Hypodermosis can be the cause of economic
INTRODUCTION
İleşim (Correspondence)
+09251-9290267
haroonahmad12@yahoo.com
RESEARCH ARTICLE
A216
Geographical Distribution of ...
losses due to meat trim at slaughter, and the eect on
hides is well established 6. The importance of the disease
has led to the formation of official campaigns to control
and/or eradicate the parasite in several countries such as
Britain 7. and Denmark 8. Financial losses prior to the start
of the eradication campaign were suggested to be £13
million in Britain 9. Similar estimates suggest losses of more
than £600 million in the USA 10,11.
Pakistan is an agricultural country with semi arid
landscape and subtropical climate. Most of the people
earn their livelihood from selling agro-livestock products
and rearing of livestock (cattle, sheep, goats and bualoes).
The prevalence of WFI has been reported in bualoes,
cattle and goats from dierent areas of Pakistan 12,13. In
dierent endemic areas, the prevalence of WFI was 20-
84% in goats and 22-24% in cattle 14. In Chakwal district
(northern Punjab) the prevalence of hypodermosis in
bualo is 5.20% 15.
Keeping in view the importance of cattle in this country
an epidemiological survey was conducted to establish the
prevalence and geographical distribution of hypodermosis
in dierent areas of Northern Punjab (Pakistan).
MATERIAL and METHODS
Location and Sample Size
The present study was carried out in the Northern
part of Punjab Province, Pakistan (30°34° N and 70°74°E).
Covering area is about 13.000 k m2 which is 2.9% of Punjab
total area. The area of Pakistan covered by this province is a
total of 205.345 km2 with 8 Divisions and 32 Districts.
A total of 1.000 animals were examined randomly from
different herds from Attock, Chawkal, Rawalpindi and
Jhelum districts, from august 2010 to February 2011.
In the present study geographical information were
taken in the form of questionnaire from dierent villages of
Northern Punjab, Pakistan. The animals were observed in
dierent herds belonging to dierent villages of northern
Punjab in dierent from august 2010 to February 2011.
Geographical Factors Considered for Risks Analysis
The geographical information’s were collected in the
form of questionnaire. It comprises of District, Village,
Topography, Water Bodies and Grazing Pattern
Statistical Analyses
The Statistical analyses was done by using the statistical
package SPSS for Windows 18.0 and SPSS answer Tree
3.1 (SPSS Inc., Chicago, IL USA) and the maps showing
the geographical distribution were prepared by using
Geographical Information System, ArcInfo.
RESULTS
In the present study, the Prevalence of the bovine
hypodermosis was 18.4% (95% CI 2.2-4.5). In the present
study a total of four districts were studied for the
geographical distribution of bovine hypodermosis. The
results show that in the district Rawalpindi and Attock the
prevalence was higher, as 29.92% and 25.34% respectively.
Fig 1. District-wise geographical
distribution of Bovine Hypodermosis
in Northern Punjab
Sekil 1. Pakistan’ın Kuzey Pencab
Bölgesi’nde sığır Hypodremosis’inin
bölgesel coğrafik dağılımı
A217
AHMED, KHAN, PANADERO-FONTAN
SANDEZ, IQBAL, NAQVI, QAYYUM
Whereas, in Jehlum and Chakwal district the prevalence
10.04% and 7.55% respectively. The results show that there
is a significant dierence in the geographical distribution
of bovine hypodermosis in all the districts of northern
Punjab (Fig. 1).
The village wise geographical distribution of bovine
hypodermosis was also determined during this study.
In the present study a total of twenty nine villages from
the dierent districts of northern Punjab, Pakistan were
examined for the geographical distribution of bovine
hypodermosis in the northern Punjab, Pakistan. Dierent
villages from the district Jehlum were examined for the
geographical distribution of bovine hypodermosis. In
village Hadali and D. Bypass the geographical distribution
of bovine hypodermosis were 0%, while in village Stad pur,
Purana metha and N. Rhotas were 26.9% (14/52), 29.6%
(8/27) and 12.6% (3/24) respectively. In the Rawalpindi
district the different villages were examined for the
geographical distribution of bovine hypodermosis. There
were no WFI were found in the village Dhoke Amban,
Pangrah, Dhoke Meera, Chappar, Chohah Chalsa, Buccha
were non-infested. While the geographical distribution
of bovine hypodermosis in the village Ghroli, Ghoran
Lohran, Mankiala, Dhoke Major, Purha, Kallar Sydian, Pidh
Bhunir, Gangrila, Missa, Guliana, Ghrabi, 37.5% (9/27),
56.9% (9/17), 23.1% (6/26), 3.3% (4/122), 47.8% (11/23),
28.9% (4/14), 37.5% (3/8), 50%(7/14), 50% (15/200), 75%
(15/20), 97.1% (33/34) and 60% (3/5) respectively. In the
district Chakwal the four villages were examined for the
geographical distribution of bovine hypodermosis. There
were no WFI were found in the village Ibrahim Abad, while
in the Hiraj, Hindol, Dhoke Chodrian the geographical
distribution of bovine hypodermosis were 5.3% (5/95), 6%
(3/50) and 23.7% (9/38) respectively. In the Attock district
the dierent villages were examined for the geographical
distribution of bovine hypodermosis. The hypodermosis
was geographical distributed in the Kharimurat and Jhund
Mirza, where it was 17.3% (14/81) and 26.4% (24/91)
respectively (Fig. 2).
The Topography of the villages is very important factor
in the geographical distribution of bovine hyoodermosis.
Three type of location were Hilly, Semihilly and plain were
existing in this area. In the plane area the distribution was
8.9% (43/484). In semihilly area distribution was 30.8%
(105/341). In the hilly area the distribution was 20.6%
(36/175) (Fig. 3). The grazing pattern is a very important
geographical factor influencing the epidemiology of
bovine hypodermosis. The geographical distribution in
Fig 2. Geographical distribution of
Bovine Hypodermosis in different
herds of Northern Punjab, Pakistan
Sekil 2. Pakistan’ın Kuzey Pencab
Bölgesi’nin farklı sürülerinde sığır
Hypodermosis’inin coğrafik dağılımı
91.11%
79.42%
69.21%
30.79%
20.58%
8.89%
0%
10%
20%
30%
40%
50%
60%
70%
80%
90%
100%
Plane Hilly Semihilly
Loca tion
Prevalance
Non Infested (%)
Infested (% )
Fig 3. Role of Location in the geographical distribution
of Bovine Hypodermosi from different districts of
Northern Punjab, Pakistan
Sekil 3. Pakistan’ın Kuzey Pencab Bölgesi’nin farklı
alanlarında sığır Hypodermosis’inin coğrafik dağılı-
mında lokalizasyonun rolü
A218
Geographical Distribution of ...
the animals having gone to field was 28.8% (179/622). The
animal kept at home having distribution of hypodermosis
was 1.3% (5/378) (Fig. 4). The presence of Water-bodies is
very important factor in the geographical distribution of
bovine hypodermosis. In the area where the water- bodies
were not present the distribution of bovine hypodermosis
were 10.8% (61/563). In the area where the water- bodies
were present the distribution of bovine hypodermosis was
28.1% (123/437) (Fig. 5).
DISCUSSION
In the present study the Prevalence of bovine hypo-
dermosis in cattle from northern Punjab was 18.4%
(184/1000). The results show that the geographical
distribution in the Rawalpindi and Attock district was
higher as compared with Jehlum and Chakwal district
was 10.04% and 7.55% respectively. Our results correlate
with the distribution of hypodermosis in dierent areas of
district Chakwal 23.50% 15. There is a wide variation in the
geographical distribution of WFI among dierent parts of
the world 4 and even within Pakistan 12. The distribution of
hypodermosis was recorded 21.62-23.8 percent in cattle 16.
Warble y infestation (WFI) in cattle was recorded to be
29% in cattle in Dera Ghazi Khan District and 26% in cattle
of Rajanpur districts 17. WFI has also been reported from
Dera Ismail Khan, Kohat, Malakand and Abotabad districts
of NWFP 16 and Dera Ghazi Khan & Barkhan districts 16,18.
Similarly, the Prevalence of hypodermosis in cattle in Kars,
Afyonkarahisar and Nigde province of Turkey were 31.9% 19,
2.96% 20, 5.08% 21 respectively. This variation in the rate
of geographical distribution of bovine hypodermosis
in dierent areas might be due to the dierences in the
environmental conditions (topography of the land, season,
humidity, temperature, climate, rain fall, wind velocity)
aecting the development of the warble 7.
The Topography of the villages is very important factor
in the geographical distribution of bovine hypodermosis.
Three type of topography were Hilly, Semihilly and plain
were existing in this area. In the plane area the distribution
was 8.9% (43/484). In semihilly area, the distribution was
30.8% (105/341). In the hilly area the distribution was
20.6% (36/175). So it is concluded from the results that the
geographical distribution of hypodermosis in the hilly and
semihilly area is higher as compared to the plane locations.
The intensity of hypodermosis in hilly and semihilly area is
higher due to suitable climatic conditions that favors the life
cycle of hypoderma. Our results correlates with the other
studies as Hypodermosis, is an important ectoparasitic
infection of cattle and goats prevalent in hilly and semi-
hilly areas of Pakistan 15. Similarily, the Hypodermosis is
98.67%
71.22%
1.37%
28.72%
0%
10%
20%
30%
40%
50%
60%
70%
80%
90%
100%
Home Field
Grazzing Pattern
Preval ance (%)
Non Infested
Infes ted
Fig 4. Role of Grazzing Pattern in the geographical
distribution of Bovine Hypodermosis from
different districts of Northern Punjab, Pakistan
Sekil 4. Pakistan’ın Kuzey Pencab Bölgesi’nin
farklı sürülerinde sığır Hypodermosis’inin coğrafik
dağılımında otlatma şeklinin rolü
28.10%
10.80%
71.9%
89.2%
0%
10%
20%
30%
40%
50%
60%
70%
80%
90%
100%
Present Absent
Wa ter Bodies
Pre va lanc e
Infes te d
Non-Infested
Fig 5. Role of Water Bodies in the geographical
distribution of Bovine Hypodermosis from
different districts of Northern Punjab, Pakistan
Sekil 5. Pakistan’ın Kuzey Pencab Bölgesi’nin
farklı sürülerinde sığır Hypodermosis’inin coğrafik
dağılımında su kaynaklarının rolü
A219
AHMED, KHAN, PANADERO-FONTAN
SANDEZ, IQBAL, NAQVI, QAYYUM
an endemic disease reported in mountainous areas of
Pakistan 16.
The grazing pattern is a very important geographical
factor influencing the epidemiology of bovine hypo-
dermosis. The geographical distribution in the animals
having gone to field was 28.8% (179/622). The animal
kept at home having distribution of hypodermosis was
1.3% (5/378). Other determinants aecting the distribution
might include host specificity, breeds, husbandry and the
use of insecticides. Many other factors can also inuence
the distribution of hypodermosis like grazing pattern 19.
Similarly, it was observed that the major risk factor for
hypodermosis herd’s positivity is the free grazing practice.
The waterbodies is an important geographical factor
inuencing the bovine hypodermosis. The presence of
Water-bodies is very important factor in the geographical
distribution of bovine hypodermosis as shown in the
results. Because the areas have water bodies having
environmental condition (Relative humidity, Wind speed)
that favors the developmental stages of Hypoderma larvae
be. So the geographical factors, Villages, Topography,
Grazing pattern and water bodies has a significant eect
on the geographical distribution of Hypodermosis in
Northern Punjab, Pakistan as Shown in figures.
In Pakistan hypodermosis is an endemic disease that
has a significant eect on the economy, because Pakistan
is a major exporter of hides, leather and their products, so
infested hides were not able to export so resulting a major
economic losses.
It is concluded from the present study that the hypo-
dermosis is a serious problem in the cattle of dierent
villages of Northern Punjab, Pakistan. So it is very important
to explore these disease in future in dierent areas of
Pakistan. The government should control this disease,
because is very important in livestock sector of Pakistan.
This study is very useful in determining the geographical
distribution of hypodermosis in the cattle of Northern
Punjab, Pakistan.
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... There are different risk factors responsible for the presence of hypodermosis in Pakistan, Iran and Turkey. The potential risk factors are categorized into the following types; (a) Biological factors such as age, sex, breed, previous exposure and medication schedule (b) Physical factors such as grazing pattern, management practices, presence of waterbodies, sunshine, humidity, month, area, color, rainfall, precipitation, location and field/slaughter house [20,21]. The results showed that age, sex, breed, medication schedule, grazing pattern, area, month, previous exposure, management practices, presence of water bodies, temperature, precipitation and location are significant risk factors (p < 0.05) in Pakistan. ...
... Blood samples were taken, and serum samples were tested for antibodies against Hypoderma spp. using the enzyme-linked immunosorbent assay (ELISA) method [19]. During the village visit, herd type (dairy, mixed, or beef herd) and size (number of cattle on the premises) were also recorded. ...
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Introduction: Hypodermosis is an ectoparasitic disease of cattle caused by Hypoderma lineatum and Hypoderma bovis. It is an important health problem of cattle, leading to considerable economic losses. There are various factors that are involved in the spread of this disease such as herd size, location, temperature, humidity, and precipitation. Methodology: Blood samples from 112 herds were collected to determine the presence of Hypoderma spp. infestation. For these herds, size and location were determined; temperature, humidity, and precipitation data were obtained from meteorological stations; and topographic features were obtained from existing maps and through field work. A regression analysis was then used to generate a risk factor analysis profile for hypodermosis and geographic information system (GIS) was used to map the risks. Results: The GIS map developed showed the degree of infestation in different geographical locations at district and village levels. Cluster analysis demonstrated that hypodermosis prevalence varied within zones and across zones. The regression analysis showed that the temperature in the months of January, February, March, August, and November, and the precipitation in September and October had significant results (p < 0.05) when all the risks factors were analyzed. Conclusions: It is concluded that different ecological factors have an important impact on the intensity and infestation rate of hypodermosis across the globe. The present study might be used to control and eradicate the hypodermosis across the globe.
... Blood samples were taken, and serum samples were tested for antibodies against Hypoderma spp. using the enzyme-linked immunosorbent assay (ELISA) method [19]. During the village visit, herd type (dairy, mixed, or beef herd) and size (number of cattle on the premises) were also recorded. ...
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Parasitism is the major problem affecting livestock in many parts of the world. Amongst these parasitic problems, the warble fly infestation (hypodermosis) is a notorious and common malady of cattle, buffalo, sheep and goat in Pakistan. The disease is endemic in cattle, buffalo, sheep and goats in semi-hilly and mountainous areas of Pakistan. Keeping in view the importance of buffaloes an epidemiological survey was conducted to find the prevalence of hypodermosis in district Chakwal, Punjab, Pakistan, during the year 2005- 2006. Buffaloes are mainly kept for milk purpose in the study area. Out of 2000 buffaloes examined clinically, for grub infestation in the study area 125 (6.25%) and 37 (5.20%) found to be positive for the warble fly infestation in slaughter house as well as in the field. A prevalence variation among the male and female animals was also observed during the study. The factors of climatic conditions that favor the fly activity and contribute in the onset of disease.
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Cattle are mainly reared for milk and draught purpose in this region. An epidemiological survey was conducted to find out the prevalence of hypodermosis in cattle in district Chakwal, Punjab, Pakistan. Total 4000 cattle were examined clinically in the field (n=2000) and slaughter house (n=2000), for grub infestation in the study area. The prevalence was 35.50 and 23.50% in the slaughter house and field, respectively. Prevalence was higher in males than in the females. In slaughter house, it was recorded as 39.50 and 31.50%, while in the field 26.50 and 20.50% in male and female respectively. Highest prevalence (41.81%) was recorded in calves. Hypodermosis was recorded only from September to December, which suggested that change of climatic conditions in the study area favours the fly activity and contributes towards the onset of disease.
Article
Efficacy of ivermectin against cattle warbles was studied in an endemic area of Pakistan. A total of 229 cattle of three breeds and different ages were divided into two groups; treated and untreated control. Treated group was given ivermectin s/c at the rate of 200 μg/kg body weight during the 1st week of September. None of the treated animal developed warbles. Warbles were seen in 23.8% of untreated control animals. Statistically non-significant differences were seen amongst different breeds and age groups. The seasonality of warbles is discussed.
Article
The objective of the present study was to observe the seasonal pattern of the life cycle and the warble stage in bovines of Dera Ghazi Khan and Rajan Pur Districts of Southern-Punjab, Pakistan. The study was conducted from July 2004 to June 2005. The association was studied between various climatic changes and the fly life cycle. The larvae of warble flies were extracted from the backs of naturally infested animals and identified. The adult fly laid eggs in June, and hatched larvae penetrated the skin and followed intensive internal migration to diaphragm, esophageal wall and subcutaneous connective tissue in the month of July. At the start of August they reached the subcutaneous tissue and emergence began. In both species, the maximum warble emergence was found in mid-December and the lowest in mid-March and mid-August. It was concluded that preventive treatment should be adopted before the month of July to avoid carcass depreciation and decreased leather quality.
Article
In order to detect the presence of Hypoderma lineatum stage I larvae within the esophagus of cattle slaughtered in Chihuahua, Chihuahua, Mexico, a total of five samplings were carried out between July and November 2000. In each instance, a random sample was taken from 10% of the animals slaughtered in a single work shift in each of the two slaughterhouses included in this study. The esophagus were cut longitudinally in order to carry out visual inspection and detect the presence of H. lineatum stage I larvae in the submucosa. The larvae were separated and counted. We identified the presence of H. lineatum stage I larvae in the esophagus for all sampling dates, nevertheless, within the last sampling only one esophagus had them. For all sampling dates the prevalence ranged between 11 and 33%; the latter corresponded to the sampling in October. A total of 287 esophagus was inspected of which 54 were positive with one or more larvae (19%); 233 larvae were obtained from these cases. The number of larvae recovered per sampling ranged from 46 to 74 between July and October, the highest number was found in September's sampling. The largest amount of stage I larvae per esophagus was 22 in the months of July and August. Larvae were always located in the submucosa of the esophagus and all were oriented longitudinally.