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SEROLOGICAL DETECTION OF CASEOUS LYMPHADENITIS
IN SHEEP USING ELISA IN AND OUTSIDE BAGHDAD
Faisal Ghazi Habasha* Taha Yassin Ghani Al-Badrawi**
Department of Internal and Preventive Medicine, College of Veterinary Medicine,
University of Baghdad, Baghdad,Iraq
(Received 18 May 2016 , Accepted 26 May 2016)
Keywords: Caseous lymphadenitis, (ELISA), Sheep
ABSTRACT
Caseous lymphadenitis caused by Corynebacterium pseudotuberculosis is a
chronic worldwide disease of sheep and goats resulted in abscess formation in
subcutaneous and internal lymph nodes. Indirect immunosorbent assay (ELISA) were
used for detection of anti PLD IgG antibodies of C. pseudotuberculosis in two
hundred and five serum samples collected from sheep flocks and from Al-shualla
slaughterhouse in Baghdad. The seropositive percentage was 15.12 of the total sera
tested. A non-significant of higher positive percentage of CLA was seen in the female
sheep as comparison to male one A higher seropositive percentage was recorded in
old age group in comparison to younger one.
.INTRODUCTION
Caseous Lymphadenitis (CLA) is a chronic worldwide disease affecting sheep
and goats, characterized by formation of abssesation withgranulomatous, necrotizing
inflammation in one or more lymph nodes casing enlargement of lymph
nodesinfection Corynebacterium pseudotuberculosis. It results, less commonly in
pneumonia, mastitis,hepatitis, orchitis, arthritis, abortion, stillbirth, and subcutaneous
abscesses insheep and goats (1,2,3).
In human it occurs as a subacute to chronic lymphadenitis and pneumonia
(4,5).
Various diagnostic methods developed for diagnosis of CLA in animals. Many
serological tests that used in serodiagnosis included passive hemagglutination,
synergistic hemolysis inhibition, immunodiffusion and indirect ELISA tests (4,6).
The purpose of this study was to detect the IgG antibodies specific for the
Corynebacterium pseudotuberculosis in sera of sheep using enzyme-linked
immunosorbent assay (ELISA) utilized a recombinant form of phospholipase D
(PLD) from C. pseudotuberculosis to detect anti- PLD IgG antibodies in sera of sheep
infected with Caseous Lymphadenitis.
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MATERIALS AND METHODS
SERUM SAMPLES
A total of 205 sheep blood samples were collected from different sheep flocks
and slaughterhouse inside and outside Baghdad province as showed in Table (1).
Table (1) Serum samples that were collected according to area and gender of sheep
Area male female total
Baghdad 14 29 43
Al-Shualaslaughter house 89 0 89
Out of Baghdad 11 62 73
Total samples 114 91 205
All 205 blood samples were collected and transported in col box contains ice
bags and stored in refrigerator until clotted then centrifuged at 2000 rpm for 15
minutes. The separated serum tubes and stored at -18 0Cuntill used. Table(2) showed
the distribution of sheep serum samples according to age groups and gender.
Table (2) Sheep serum samples that were collected according to age and gender
Age Male Female Total
≤ 1 year 80 2 82
1.5-2 year 30 56 86
2.5-3 year 4 26 30
3.5-4 year 0 7 7
total 114 91 205
ELISA Test
The commercial kit ELITEST CLA product of Hyphen BioMed, France was
used for detection of IgG antibodies specific for the causative agent of caseous
lymphadenitis (CLA) in sheep and goat sera. This kit was performed according to the
manufactures instruction.
RESULTS
The results of ELISA for naturally infected and non-infected sheep revealed
the 31sheep out of 205 were carried anti-PLD IgG antibodies in their sera represented
15.12% of total examined serum samples.. Seven (16.28%) positive cases were
recorded in Baghdad out of 43 serum sample examined. Nine (9.9%) represented
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positive serum samples were tested from slaughterhouse, and 15 (25.48%) was
positive cases out of 73 tested serum samples in out of Baghdad as in Table (3)
Table (3) The number and percentage of positive serum samples in different areas
using indirect ELISA test
Areas
No. of Tested
serum
No. of positive
cases
Percentage of
positive cases
Baghdad 43 7 16.28
Slaughterhouse 89 9 9.9
Out of Baghdad 73 15 25.48
Total 205 31 15.12
According to age groups in sheep sera, CLA differed among different age groups
which were higher percentage recorded in sheep aged over 3 yearsmore than those of
other age groups. It was 30%, in 2.5-3 year age and was 28.57% in 3.5-4 year old
while the lowest percentage was recorded in sheep ≤ one year old (6.08%) as in Table
(4). There was significant differences (p<0.01) between young and adult age groups.
Table (4) The number and percentage of positive C. pseudotuberculosis serum
samples according to age groups using ELISA test.
Age / year Tested No. positive Percentage positive
≤ 1 82 5 6.08
1.5-2 86 15 17.44
2.5-3 30 9 30
3.5-4 7 2 28.57
total 205 31 15.12
Chi-square value 11.715*
*(p<0.01)
According to gender, results showed that 13 (11.4%) serum samples were
represented males out of 114 tested, while 18 (19.78%) positive serum samples out of
91 tested were females as in Table (5).
Statistically there were no significant differences between males and females.
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Table (5) The number and percentage of positive C. pseudotuberculosis serum
samples using ELISA test in both sexes
Animal gender
No. of tested serum No. of positive
cases
Percentage of
positive cases
Male 114 13 11.4
Female 91 18 19.78
Total 205 32 15.12
Chi-square value 2.767 NS
NS: Non significant
The optical density (OD) range which represented IgG concentration was
0.538-0.805 with mean (0.686 0.109), in sera samples collected from different sheep
flocks in Baghdad province, while in sera samples collected outside of Baghdad
ranged 0.464-2.069 with mean OD 0.851 0.527as in showed in Table (6). The cut-off
value calculated was 0.431 and the mean absorbance of the blank was 0.060.
Table (6) Optical densities of positive serum samples in different areas
Area Optical density
Mean ± SD Range
Baghdad 0.686 ± 0.109 0.538 - 0.805
Slaughterhouse 0.723 ± 0.196 0.539 – 1.067
Out of Baghdad 0.851 ± 0.527 0.464 – 2.069
DISCUSSION
The total positive percentage of ELISA test on 205 serum samples were
(15.12%) which collected from different areas in and outside Baghdad. The results
confirmed with results obtained by other researcher(7) who reported (17%) were
found positive out of 216 sera tested by AGPT and ELISA. The results of this study
agreed with the results of (8) who indicated the percentage of positive sera of CLA
was (16.9%) out of 879 serum samples tested.
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The seropositive results showed a higher percentage in female than in male
animals and this fact agreed with (9,10). This fact explained that female's stayed
longer time in the flock due to their reproductive activity and resulted in longer
exposure to various risk factors such as cohabitation with infected animal's and
shearing.
Concerning age groups, CLA differed among different age groups where
higher percentage was recorded in sheep of age group 2.5-3 years old. This finding
agreed with results obtained by (11,12, 13,14,10).
التحری السیرولوجی عن مرض التھاب الغدداللمفاویة التجبنی فی الاغنام باستخدام فحص
الالیزا داخل وخارج بغداد
فیصل غازی حباشھ ، طھ یاسین غنی البدراوی
فرع الطب الباطنی و الوقائی کلیة الطب البیطری جامعة بغداد
الخلاصھ
یعتبر مرض التھاب الغدد اللمفاویة التجبنی فی الاغنام والماعز من الامراض المزمنة الواسعة الانتشار
فی العالم مسبا فی تکون الخراج فی الغدد اللمفاویة السطیة والداخلیة. تم استخدم اختبار الالیزا للکشف عن
الاجسام المضادة فی 205 عینة مصل جمعت من قطعان الاغنام المختلفھ ومن الاغنام المجزورة فی مجزرة
الشعلة فی بغداد. سجلت نسبة الاصابھ السیرولوجیة 15.12 % من مجموع المصول المفحوصھ. وکانت نسبة
الاصابة فی الاناث اعلى منھا فی الذکور بینما وجد ان نسبة الاصابة تزداد بازدیاد عمر الحیوان.
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