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SEROPREVALENCE OF BABESIOSIS IN CATTLE IN MOSUL CITY,
IRAQ
H. A. Mohammed, S. D. Hasan, N. G. Fathi, Q. T. Al-Obaidi
Department of Internal and preventive medicine, College of Veterinary Medicine, University of
Mosul, Mosul, Iraq.
(Received 2020,Accepted 18 November 2020)
Keywords: Babesia bigemina, Seroprevalence, I-ELISA.
Correspondence author: qaestalb1976@uomosul.edu.iq
ABSTRACT
Babesia bigemina is considered as one of the most remarkable blood protozoa in cattle
and mainly transmitted via arthropod. This study was conducted on a random group of cows,
they numbered 180 local cows who ranged in age from 3-7 years old, from different localities in
Mosul city north Iraq, comprising both clinically healthy (n=162) and clinically suspected
infected animals (n= 18). In this study, indirect-enzyme immunosorbent Assay (I-ELISA) was
used to detect of babesia antibodies known as B. bigemina in the blood serum. Then, the blood
and biochemical information that existed from both groups was analyzed, so that the two of them
are compared with the control group (n=15). The result showed that the overall seroprevalence of
B. bigemina in cows was 74/180 (41.1%) for clinical and subclinical cows were 10% and 31.1%
respectively. The subclinical infected cows was statistically higher than that of clinically infected
(P<0.05). Clinically infected cows were suffering from acute onset of the disease includes fever,
anorexia, emaciation, drooping in milk yield, jaundice and hemoglobinuria,, with significant
hematological and biochemical parameters alterations. While, subclinically infection cows
appeared healthy with absence of changes in blood and biochemistry tests as compared to control
groups. It has been concluded that significant cases were diagnosed suffering from acute
infection with the B. bigemina with higher prevalence of subclinical cases in Mosul city, Iraq.
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INTRODUCTION
Bovine Babesiosis is list B disease according to OIE classification for the most common
blood parasitic disease of mammals after trypanosome, the disease implicated as a substantial
economic losses due to high morbidity and mortality rates, decrease in animals production and
the cost of treatment (1). It is intraerythrocytic protozoan and belongs to the Babesiidae family
(the genus Babesia, order Piroplasmida, phylum Apicomplexa and subclass piroplamsia), which
responsible to infect domestic animals, wild animals and human (2). However, the term
piroplasmas is considered one of the commonly used terms due to its pear-like form, a form that
expresses the presence of the merozoites present as small parasites within the red blood cells of
the mammals (3, 4).
It is worth noting that there is a major type that acts as a vector for this babesia: The
Rhipicephalus ( formerly Boophilus) ticks (5). There are more common types belonging to this
family and found in cows, and they are: B. bigemina, B. divergens, B. bovis and B. major. It
should be noted that B. bigemina and B. bovis are among the most influential species on
livestock health and productivity, especially those in tropical and subtropical countries such as
Africa, Asia, Australia, and the United States of America (6, 7, 8) In these species, the
transmission of Babesia species is through hard ticks and transovarially passes by the egg from
one generation to the following, and these ticks are: Rhipicephalus microplus and Rhipicephalus
annulatus (9).
On the other hand, there are some symptoms that accompany the infection of babesia
,which include: anorexia, pale mucus membrane, dyspnea, anemia, fever, hemoglobinuria.
Mostly, these clinical signs affect animals that produce low-level parasitemia in a way that they
become carrier(10, 11), Despite this, the livestock may remain infected and have no clinical
symptoms, and this is the main transmission factor (3, 5).
In this regard, there are many techniques that are used to diagnose and detect the presence
of different types of babesia, among which are: microscopic examination of blood film,
serological tests and molecular assays. It must be clarified that a blood smear test is followed in
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severe cases and this technique is classified as a standard technique for routine diagnosis of
babesiosis, and it is rate by some as a gold standard (12).
Besides, serological tests including enzyme-linked immunosorbent assay (ELISA) are
considered able to detect the presence of antibodies in pregnant animals, for later use for
monitoring surveillance and export certification (13, 14, 15). Studies of B. bigemina in Mosul
City, Iraq are very scarce and little information has been reported. The therefore, the aims of this
study were to determine the prevalence of B. bigemina and concurrent infection in clinically
and subclinically infected equids. Moreover, to investigate the clinical ,hematological and
biochemical parameters alterations of clinical and subclinical infection cows.
MATERIAL AND METHODS
Animals and samples collection: From October 2018 to April 2019, the study was conducted
on 180 native cows, 3-7 years old, from different farms in Mosul city north Iraq, comprising both
clinically healthy (n=162) and clinically suspected infected animals (n= 18). Hematological and
biochemical parameters were analyzed from blood and serum samples from clinical and
subclinical cows in comparison to healthy cows (n=15) as control group. Information regarding
age, origin, clinical status and clinical signs of the animals were recorded during sample
collection. Ten-milliliter (ml) blood samples were collected from each cow via jugular vein
puncture using 18G needle into two sterile vacutainers® tubes (5ml each), one with anticoagulant
ethylene diamine tetraacetic acid (EDTA) for hematological analysis. Serum was
separated from the second tube without anti-coagulants for detection B. bigemina antibodies and
for biochemical analysis.
Laboratory processing: In order to perform the direct diagnosis process, these slides were
repaired using methanol70% and were stained with Giemsa for 30 mints to be observed under
the optical microscope (Olympus BX 51, America, Inc.).
The blood was examed to calculate (CBP) : total erythrocyte counts (TRBC), packed cell
volume (PCV), hemoglobin concentration (Hb), total and differential leukocyte counts (TLC),
thin and dried blood smears stained with Giemsa were allocated to calculate the number of
differential leukocyte counts (16).
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In addition, there are a number of Serum biochemical parameters were analyzed using
special cassettes for each in a Chemistry analyzer (Vet Test, Arachem/ USA), which includes:
alanine amino transferase (ALT), aspartate amino transferase (AST), alkaline phosphatase
(ALKP), blood urea nitrogen (BUN), total protein, creatinine, glucose and calcium
Indirect enzyme linked immunosorbent test (I-ELISA): In this study, an Indirect ELISA
examination was performed in order to detect the presence of B. bigemina antibodies present in
the serum .Samples selected in the study, and this was done according to the techniques provided
by (Savona-Sweden). However, absorbance was measured at 405 nm through the use of a microplate
reader known as the BioTek EL-800 micro plate reader. In order to obtain a set of values
for Optical density (OD), these values are then transferred to a Microsoft Excel spread-sheet. Of
these values, Percent sero-positivity (PP) was calculated according to the manufacturer's
instructions equation below, and only when both versions gave PP ≥ 40 an animal was
considered as positive.
The interpretation of results : PP ≤ 25 = Negative , PP 26-39 = Doubtful , PP ≥ 40 = Positive
Statistical analysis: For data analysis, IBM-SPSS statistics (ANOVA) program-version 21 was
used, from which several tests were performed, including: two-sided Chi-square and 95%
confidence interval. In order to compare between the control group and the experimental group
selected in this study, (P <0.05) was determined in order to identify all the important differences.
RESULTS
In the current study, based on indirect ELISA analysis of B. bigemina in cow was 74/180
(41.1%) for the clinical and subclinical cows were 10% and 31.1% respectively. The subclinical
infected cows was statistically higher (P<0.05) than that of clinically infected (Table1).
Mean OD Samples
Mean OD Positive control
PP = X 100
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The clinically infected cows with B. bigemina were suffering from acute form of the
disease and exhibited fever, anorexia, emaciation, drooping in milk yield, jaundice and
hemoglobinuria with different frequency percentages of each signs (Figure 1). While subclinical
infected cows appear healthy with no clinical signs. Further, a statistically significant increase
(P<0.05) in the body temperature (40.5Cº), respiratory rate (58.3 /min), heart rate (89.8 /min)
(Table 2).
Table 1: Clinical status of B. bigemina in cows using indirect ELISA test (n=180).
Values significantly different (P < 0.05) between cows status are labelled with the vertically different letters (a, b).
Figure 1: Frequency and percentage of clinical signs in cows clinically infected with
B. bigemina (n=18).
Clinical Status B. bigemina No. Positive (%)
Clinically infected 18(31.1)a
Subclinical infected 56(10)b
Total 74(41.1)
(77.7%)
(61.1%)
(44.4%)
(72.2%)
(88.8%)
Clinical Signs
Fever Anorexia Milk drooping Jaundice Hemoglobinuria
14
11
8
13
16
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Table 2:Clinical parameters of cows clinically infected with compared with subclinical
infected and healthy group.
Parameters
Healthy group
(n=15)
Mean ± S.E
Subclinical
infected cows
(n=56)
Mean ± S.E
Clinically infected
cows (n=18)
Mean ± S.E
Body temperature °C 37.8 ± 0.22 37.5 ± 0.13 40.5 ± 0.73*
Respiratory rate /1 min 30.5 ± 3.27 31.5 ± 4.45 58.3 ± 5.45*
Heart rate/1 min 57.5 ± 4.33 58.3 ± 5.21 89.8 ± 4.43*
Rumen contraction / 5min 5.82 ± 0.43 5.92 ± 0.23 2.87 ± 0.22*
Mean values ± standard error (S.E.), * (P < 0.05) a significantly different between cows
clinical status.
In the present study the haemo gram of cows clinically infected with B. bigemina
reflected to statistical significant decrease in TEC, Hb, and PCV, along with significant increase
in total TLC, due to neutrophilia and lymphocytosis (P < 0.05), while no statistically significant
alterations of these hematological parameters in the subclinically infected cows compared with
the healthy group (Table 3).
In addition, serum biochemistry analysis of clinically infected cows showed significant
increase in ALT, AST, ALKP, blood urea nitrogen BUN, creatinine and significant decrease in
total protein, while no significant changes were found in the glucose and calcium levels
(P<0.05). No statistically significant alterations of these biochemistry parameters in the
subclinical infected cows compared with the healthy group (Table 4).
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Table 3: Hematological values in cows clinically infected with B. bigemina compared with
subclinical infected and healthy group
Parameters
Healthy group
(n=15)
Mean ± S.E
Subclinically
infected cows
(n=56)
Mean ± S.E
Clinically infected
cows (n=18)
Mean ± S.E
Total erythrocytes counts
×106 μl 7.21 ± 0.84 6.92 ± 0.54 5.29 ± 0.43*
Hemoglobin, mg/ 100ml 11.88 ± 1.64 12.11 ± 1.23 7.83 ± 1.72*
Packed cell volume % 33.25 ± 3.78 32.15 ± 2.67 26.22 ± 2.23*
Total leukocyte counts
x103μl 11.72 ± 1.43 10.98 ± 1.31 13.54 ± 0.9*
Neutrophils x103μl (%) 46.34 ± 3.22 46.25 ± 5.12
(47.62 ± 2.32)*
Lymphocytes x103μl (%) 44.62 ± 3.13 44.12 ± 2.24 (45.94 ± 2.12)*
Monocytes x103μl (%) 3.63 ± 1.01 3.23 ± 1.11 3.73 ± 3.23
Eosinophils x103μl (%) 3.22 ± 1.13
3.13 ± 1.22 3.52 ± 1.22
Basophils x103μl (%) 1.21 ± 0.14 0.91 ± 0.12
0.87 ± 0.14
Mean values ± standard error (S.E.), * (P < 0.05) a significantly different between cows status
Table 4: Biochemical values in cows clinically infected with B. bigemina compared with
subclinical infected and healthy group
Parameters
Healthy group
(n=15)
Mean ± S.E
Subclinically
infected cows
(n=56)
Mean ± S.E
Clinically
infected cows
(n=18)
Mean ± S.E
ALT U/L 40.52 ± 11.83 42.56 ± 12.19 65.18 ± 12.68*
AST U/L 54.55 ± 19.66 57.15 ± 20.15 95.81 ± 22.04*
ALKP U/L 54.35 ± 31.12 56.33 ± 21.11 67.88 ± 23.13*
BUN mg/dL 18.60 ±5.21 19.10 ± 6.23 27.54 ± 7.42 *
Total protein g /dl 7.47 ± 0.22 7.07 ± 0.12 3.27 ± 0.58*
Creatinine mg/dl 0.67 ± 0.19 0.98 ± 0.17 1.87 ± 0.22 *
Glucose mg/dL 58.46 ± 11.86 58.14 ± 10.45 57.21 ± 7.67
Calcium mg/dl 9.14 ±1.88 9.01 ± 1.48 8.9 ±1.72
Mean values ± standard error (S.E.), * (P < 0.05) a significantly different between cows status
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DISCUSSION
This study showed that there is a an increase in the prevalence of B. bigemina infection in
the city of Mosul, which is up to approximately 42%. The reason for this is that there is a large
distribution of types of carrier ticks, as well as the presence of transporting animals that are
either carrier or sub-clinical, in addition to environmental conditions, including farm
managements that have a significant impact on the occurrence of the disease. This is consistent
with what was found in the (17), as found that the main problem in the presence of babesiosis is
the presence of animals that recover from the disease will become chronic carriers that act as a
source of infection for other animals and ticks.
Which means that Bovine Babesiosis occurs mostly in the tropics, subtropics and
temperate regions of the world, and this is what affects many livestock with this microbe (18).
This is consistent with many previous studies that touched on the extent of Bovine Babesiosis
prevalence of this was 27.27% (19), whereas, the prevalence of babesia in Qadisiyah
Governorate reached 47.91%, in Iran 7.10% (20), and in Turkey it was 14.8% (21).
On the other hand, the current study showed that cows infected with B. bigemina was
experiencing fever, anorexia, emaciation, drooping in milk yield, jaundice and hemoglobinuria.
Besides, other symptoms are high body temperature, respiratory rate, heart rate and decreased in
rumen contractions in infected cattle. These results are consistent with the results of some
previous studies who mentioned that the symptoms of infection with this microbe are represented
in pale mucus membranes, jaundice, increased respiratory rate, hemoglobinuria, and fever (22,
23, 24). It is noted when performing a haemogram that the number of erythrocytes and
hemoglobin decreased, while there was a decrease in packed cell volume and total leukocytic
count; Especially when both the infected group were compared to the healthy group. This is
consistent with the results of some previous studies (6, 25, 26).
Regarding the results of the biochemistry of the affected group of cattle, There is a
significant increase in the percentage of AST, ALT, ALKP, BUN, total protein and creatinine, on
the contrary, there were no significant changes in the levels of glucose and calcium in the blood;
This is consistent with the results of some previous studies (24, 25, 26).
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ACKNOWLEDGEMENTS
This work was supported by the Faculty of Veterinary Medicine, university of Mosul.
Mosul, Iraq. The authors are thanks all cattle owners for their kind collaboration.
الانتشار المصلی لداء البابیزیوسز فی الابقار فی مدینة الموصل، العراق
ھدیل عاصم محمد، صدام ظاھر حسن، ناصر غانم فتحی ،قیس طالب العبیدی
فرع الطب الباطنی والوقائی، کلیة الطب البیطری ، جامعة الموصل، الموصل، العراق
الخلاصة
احد اکثر طفیلیات الدم التی یمکن ملاحظتھا فی الماشیة، وینتقل الطفیلی بشکل Babesia bigemina یعد طفیلی
رئیس عن طریق المفصلیات. أجریت الدراسة بصورة عشوائیة على مجامیع الابقار المحلیة ١٨٠ بقرة، وبأعمار تراوحت بین
٧ سنوات ومن مناطق مختلفة فی مدینة الموصل شمال العراق. تضمنت ١٦٢ حیوان سلیم سریریاً و ١٨ حیوان مشکوکة -٣
B. للکشف عن الأجسام المضادة لطفیلی I-ELISA الاصابة. تم اجراء اختبار الممتز المناعی المرتبط بالأنزیم المباشر
فی مصول الدم، وکما وتم اجراء الفحوصات الدمویة والکیموحیویة على کلا المجموعتین ومقارنة النتائج مع bigemina
41.1 ) ، منھا %) ١٨٠/ حیوانات السیطرة ١٥ بقرة. أظھرت نتائج الدراسة ان النسبة الکلیة لانتشار الطفیلی فی الابقار ٧٤
ابقار مصابة سریریا و بقار مصابة تحت السریری ( ١٠ ٪ و ٣١.١ ٪) على التوالی، ھذا یعنی ان نسبة الاصابة التحت
عانت الأبقار المصابة سریریًا من الشکل .(P< السریریة فی الأبقار کانت اعلى معنویا من الابقار المصابة سریریا ( 0.05
الحاد للمرض شملت الحمى وفقدان الشھیة والھزال وتدھور إنتاج الحلیب والیرقان وبیلة الھیموغلوبینیة ، مع وجود
تغییرات فی بعض اختبارات الدم والاختبارات الکیموحیویة، فی حین ظھرت الأبقار المصابة بالشکل التحت السریری بصحة
جیدة مع عدم وجود تغییرات فی اختبارات الدم والاختبارات الکیموحیویة عند مقارنة النتائج مع مجموعة السیطرة. استنتج من
مع انتشار العالی للحالات التحت السریریة فی مدینة B. bigemina الدراسة تشخیص الشکل الحاد من الاصابة بطفیلی
الموصل، العراق.
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