Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 Prevalence of some protozoa and its effects on biochemical changes in goats in, Marsa, and El-Wadi El-Gadid provinces By Soheir M. El Manyawe*, M.A.M.Abdel Rahman*, A.M.I. Abd El Aal***, Azza M. Kamal**and Samira A.Snousi**** *Parasitology **Chemistry Dept., Animal Health Research Institute, Dokki, Lab. ***, and El-Wadi El-Gadid Lab. **** SUMMARY A comparative study was carried out on 3 goats in,, and El-Wadi El-Gadid provinces. The aim of this work was to determine the prevalence of protozoan parasites infecting goats in the three localities, and to study the effect of these parasites on the biochemical changes in the serum of infected animals. The results indicated the presence of 9 Eimeria species and Cryptosporidium oocysts in the faeces of examined goats. Toxoplasma gondii antibodies were also detected in the serum of the investigated animals. The biochemical analysis of blood serum revealed a significant alteration in total protein, albumin, serum enzymes, electrolytes, trace elements, urea, and creatinine. The importance of parasitic infection control as well as drinking the under ground polluted water with metals and minerals and its effects on animal health, was also discussed. Referred by Prof. Dr. Hussein Omar Prof. Dr. Wahied A. Mossa INTRODUCTION he domestic goat is considered one of the ancient animals known to man, where it is named in the developmental countries the poor man's cow. It produces milk, meat, and mohair which are of great economical importance. Recently, the application of goats as producing animals, attention should be directed to the role of parasitism in the economics of the husbandry of these goats. In T Professor of Parasitology, Fac. Vet. Med., University Professor of Parasitology, Fac. Vet. Med., University Egypt; the climatic conditions are more or less favorable for the development and existence of different parasites. However, little is known about the importance of parasites as health hazards to goats in Egypt. Protozoan parasites cause severe diseases among goats, including Theileria, Babesia, Toxoplasma, Sarcocystis, Eimeria, and Cryptosporidium species (Levine, 1985).. Coccidiosis, is an important enteric 12
Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 disease caused by approximately 12 different species of Eimeria, affecting preweaned or recently weaned animals and those under unsanitary and stress conditions. The parasite causes diarrhea, insufficient weight gain, and occasionally death (Foreyt, 199). Cryptosporidiosis is a prevalent disease in neonatal ruminants and humans. It is caused by Cryptosporidium parvum, which is primarily the disease of lambs and kids less than 3 days of age, and is usually a milder disease than coccidiosis (Fischer, 1984). On the other hand, Toxoplasmosis is one of the major zoonotic diseases, caused by protozoan parasite, Toxoplasma gondii (Dubey, 1986). Infection with T.gondii is extremely common throughout the world, affecting pregnant animal, particularly sheep and goats leading to fetal death, abortion, stillbirth or neonatal death with the subsequent economic losses (Buxton, 1998). Consequently a survey was under taken to determine the prevalence and intensity of protozoan parasitic infection (Eimeria, Cryptosporidium and Toxoplasma) in goats in different environmental localities in Egypt. Also, to study the effect of these parasites on some biochemical parameters in serum of infected animals.. MATERIALS AND METHODS I- Animals:A total of 3 goats, 84 (less than one month), 216 (more than one month) were examined from three different provinces,, Marsa, and El-Wadi ElGadid (1 animals from each province). Blood and faecal samples were collected from each animal.. a- Faecal samples: Fresh faecal samples were collected in plastic bags, labeled and placed in refrigerator at 4 ºC until examined. b- Serum samples: Blood samples were collected in clean and dry test tubes, left to clot at room temperature and then centrifuged at 3 r.p.m. for 1 minutes. The separated sera were kept in sterile labeled tubes at -2ºC until used.. II- Faecal examination: Each faecal sample was examined microscopically using the concentration floatation technique according to Soulsby (1968) for the presence of Eimeria oocysts. All positive samples were subjected for oocysts counting using modified McMaster technique (MAFE, 1977). The detected Eimeria oocysts were identified freshly after sporulation in 2.5% potassium dichromate at 27 ºC. 13
Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 Sporulation time was recorded and description of Eimeria oocysts was done according to Norton (1986)... Faecal samples were also examined for the presence of Cryptosporidium oocysts by direct smear and thin film stained with modified Ziehl-Neelsen technique as described by Henriksen and Pohlenz, (1981) and examined under oil immersion lens of microscope.... III- Serological test: : (Indirect haemaglutination test IHAT) Toxo-IHA-Fast kit was commercially obtained from ABC Diagnostics, New Domietta, Egypt. - The procedures followed the kit instructions. - Antibody titers of 1/8 were considered positive for Toxoplasma gondii. tro- photometrically using standardized test-kits supplied from Bio-Merieux (Bains/ France). -Serum sodium and potassium were determined by using Flam photometer corning 4. -Serum chloride was determined by using chloride analyzer Model 925. -Serum copper and zinc were determined by using Atomic abs o r p t i o n s p e c t r o p h o t o me t e r (Perkin Elmer Model 238 USA). ) ). -Statistical analysis: The obtained data were subjected to software program according to Selvin (1996). RESULTS I- Parasitic examination:a) Eimeria species:the results are shown in tables, (1), (2), (3), and (4). IV- Biochemical examinations: The investigated animals were divided into 3 groups according to the parasitological infections. a) The 1st group was +ve only for Toxoplasmosis: b) The 2nd group was +ve only for Coccidiosis: b) The 3rd group was +ve only for Cryptosporidiosis: Reagents and Kits: -Serum total proteins, albumins, aminotransferases (ALT, AST), alkaline phosphatase, calcium, and magnesium were determined spec14
Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 Table (1): Prevalence of Coccidiosis among goats at three provinces in Egypt. Province Less than 1 month examined More than 1 month examined examined 32 3 93.75 68 59 86.76 1 89 89 % 28 25 89.28 72 56 77.78 1 81 81 % 18 75 76 32 42.1 1 5 5 % 86.9 216 147 68.6 3 22 73.33% El-Wadi El-Gadid 24 84 73 Table (2):Morphological features of Eimeria species detected in goat faeces. Eimeria species 1 E.hirci Average Shape Micropyle Micropyle Sporulation diameter(µ) cap time (day) 23 x 18 Ellipsoidal to sub spherical 1-3 2 E.arloingi 27 x 18 Ellipsoidal to ovoid 2-3 3 E.jolchijevi 32 x 22 Ellipsoidal to ovoid 2-4 4 E.ninakohlyakimovae 5 E.caprina 25 x 2 Ellipsoidal to ovoid Absent 1-2 32 x 22 Ovoid Absent 2-3 6 E.caprovina 31 x 24 Ellipsoidal to sub spherical Absent 2-3 7 E.christenseni 38 x 25 Ovoid 2-4 8 E.alijevi 19 x 17 Spherical to sub spherical Absent Absent 1-5 9 E.apsheronica 3 X 22 Ovoid Absent 1-2 Table (3): Eimeria species detected in goat faeces in different localities. Eimeria species 1 E.hirci 2 E.arloingi 3 E.jolchijevi 4 E.ninakohlyakimovae 5 E.caprina 6 E.caprovina 7 E.christenseni 8 E.alijevi 9 E.apsheronica 79 88.76% 76 85.45 73 82% 76 82.7% 75 92.6% 63 77.8% El-Wadi El-Gadid 41 82% 45 9% 27 54% 64 71.9% 19 38% 59 37 34 24 11 66.3% 41.6% 38.2% 26.9% 12.3% 37 11 7 45.67% 13.6% 8.64% 9 18% 15
Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 Table (4): Mean oocyste count of Eimeria species infecting goats in different localities. Province El-Wadi El-Gadid samples 89 81 5 Degree of infection Light* Middle** 33 (29.37%) 47(41.83%) 35 (31.15%) 41(33.21%) 26 (52%) 2(4%) ***Severe 9(8.1%) 5(4.5%) 4(8%) * Light: Less than 1 oocysts/gm. ** Middle: 11 5, oocysts/gm. *** Severe: Over 5, oocysts/gm. Table (5): Frequency of mixed infections with Eimeria species in goats. of species 2 3 4 5 3 3.37 % 5 5.62 % 37 41.57 % 44 49.44 % 89 89 % 4 4.94 % 7 8.64 % 32 39.51 % 38 46.91 % 81 81 % El-Wadi El-Gadid 2 4% 4 ٨% 18 36 % 26 52 % 5 5 % b) Cryptosporidium species :The results are shown in tables (6). Table (6): Prevalence of Cryptosporidiosis among goats at three provinces in Egypt. Province El-Wadi El-Gadid Less than 1 month exam- ined 32 2 6.25 28 24 84 2 2.38 More than 1 month exam- ined 68 12 17.65 72 9 12.5 76 6 7.89 216 16 27 12.5 examined 1 1 1 3 14 9 6 14 % 9% 6% 29 9.67 %
Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 II-Serological test: Toxoplasma gondii:the results are shown in tables (7), and (8). Table (7): Prevalence of Toxoplasmosis among goats at three provinces in Egypt. Province Less than 1 month exam ined 32 3 9.38 More than 1month 68 34 5 examined 1 examined 37 37 % 28 5 17.86 72 4 55.56 1 45 45 % El-Wadi El-Gadid 24 9 37.5 76 51 67.11 1 6 6 % 84 17 2.24 216 125 57.87 3 142 47.3 % Table (8): Antibody titers of Toxoplasma infected goats using IHAT at three provinces in Egypt. Province 1/8 1/16 37 1/32 6 14 8 45 9 19 El-Wadi El-Gadid 6 13 142 28 1/64 1/128 1/256 6 3 6 5 4 2 18 12 7 6 4 51 26 18 13 6 Table (9): Prevalence of single and mixed infections with Eimeria, cryptosporidium and Toxoplasma species in goats. Province samples Eimeria Single infection Cryptospo. Toxoplasma Mixed infection 14 23 16.43 5 3.57 18 12.86 46 32.86 94 67.14 135 18 13.33 3 2.22 21 15.55 42 31.11 93 68.89 El-Wadi ElGadid 116 1 8.62 2 1.72 24 2.69 36 31.3 8 68.97 391 51 13.4 1 2.56 63 124 31.71 267 68.29 17 16.11
Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 III-Biochemical examinations: Table (1): Biochemical parameters of goat serum infected with Toxoplasma, Eimeria and Cryptosporidium species. Parameter Control Toxoplasma Eimeria Cryptosporidi- protein (gm%) (n=1) 8.1±1.1 gondii(n=63) 6.33±.5* species(n=51) 5.76±1.1** dium(n=1) 6.27±.45* Albumins (gm%) 3.8±.5 2.42±.27** 2.67±.4* 3.2±.2 ALT(u/l) 35±1.5 39.24±1.75* 34.7±.75 34.75±.4 AST(u/l) 4±1.3 68.54±6.67** 41.25±.2 41.5±.5 ALP(u/l) 87.2±.6 86.23±.86 72.76±9.12** 73.26±6.68** Calcium (mg %) 8.4±.3 6.15±.3* 8.7±.2 8.25±.3 Magnesium(mg%) 1.5±.4.73±.7* 1.35±.2 1.25±.2 Sodium (Meq/l) 133.5±2.3 135.2±2.1 99.17±16.6* 18.84±9.7* Potassium (Meq/l) 4.8±.9 2.77±1.** 3.3±1.4* 3.4±.77* Chloride(mg%) 1±2.1 1.15±.1 88.55±4.32** 93.59±5.87* Copper(mg%).81.3±2.4 83.3±1.72 89.2±3.42* 91.7±2.31* Zinc(mg%) 73.4±1.6 71.6±2.32 56.1±3.8** 54.5±2.62* Urea (mg%) 25±.9 39.32±1.73** 32.17±1.51* 31.25±1.47* Creatinine(mg%).43±.3 1.26±.5** 1.19±.3* 1.8±.4 * Data expressed as mean ±SE. * Significantly different from control at (P <.5). *Significantly different from control at (P <.1).* 18
Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 Plate (1): Eimeria and Cryptosporidium species detected in goat faeces. A: Unsporulated oocysts. B: Sporulated oocysts. 19
Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 DISCUSSION heep and goats play an important role in animal production, especially in Egyptian desert which represents more than 96% of Egyptian land. This includes, and El-Wadi El-Gadid provinces. Little is known about parasitic infection, particularly in goats which inhabits this vast area as compared with those farmed in. S The results of the present study indicated that the prevalence of Eimeria infections in goats was 89% in. Similar results were recorded by Otify (1984) and ElManyawe (1999) in Egypt. However, lower infection rates were detected in, and El-Wadi El-Gadid (81% and 5%) respectively. Nine species of Eimeria have been detected in 89 out of 1 faecal samples of goats in, the most prominent species were E.hirci (88%) and E.arloingi (85%), Norton (1986) declared the prominence of the same species. Where as six and five Eimeria species were detected in, and El-Wadi El-Gadid respectively. Single infection with one Eimeria species was not observed, whereas mixed infections with 2 to 5 species were detected. Concerning Cryptosporidiosis, the present study revealed that 14%, 9% and 6% of goat samples were positive for Cryptosporidia infection in the three mentioned provinces of Egypt. Similar results (13.34%) were obtained by ElSayed Nasr(28), where as (2.12%) were only obtained by ElManyawe (1999). The differences of infection rates could be attributed to the variations in the environmental conditions, nature of pasture and management. Kids less than one month of age were more susceptible to Eimeria and Cryptosporidia infections than older animals. The same results were reported by ElManyawe (1999). It was found that multiple parasitisms with different species of Eimeria and Cryptosporidia were occurred in this study. It may lead to serious effects in general conditions of goats as diarrhea, insufficient productivity, loss weight gain and occasionally death, as mentioned by Foreyte, (199).. A total of 37% of goats in were sero-positive for Toxoplasmosis by using IHAT, such rate was close to that recorded (41.67%) by El- Manyawe et al., (21), and lower than 47.4% (Maronpot and Botros, 1972). Where as higher infection rates, 45% and 6% of goats were seropositive in, and El-wadi El-Gadid respectively. Similar results where recorded in warm gov11
Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 ernorates, 57.5% in Qena (Hefnawy et al., 2), 68% in Assiut (Fahmy et al., 1979), and 51.7% in Tabouk, Saudi Arabia (Sanad and Al-Ghabban, 27).. The difference in the infection rates may be related to the environmental conditions, immune status of the animals and method of management. Infection rates increase with age; it could be attributed to the contact rate with the infected materials (Huffman et al., 1981). The highest antibody titer was 1/16 where 14 (37.83%) in, and 19(42.22%) in, and 18(3%) in El-Wadi El-Gadid. The titer 1/16 is probably an indication of a recent and active infection that accompanying the reproductive problem, while titer 1/8 is mainly considered as a beginning of exposure to the disease (Huffman et al., 1981, and Dubey and Welcome, 1988). However, people become infected with Toxoplasma on ingesting the infected goat meat butchered at home and the majority of goat milk is consumed un pasteurized. Moreover, goat meat spread Toxoplasma infection in the environment, since a cat could shed several millions of oocysts after ingesting naturally infected goat meat (Dubey et al., 198). Concerning the biochemical study in the present work, some of the parameters were significantly altered due to protozoan infections. The reduction of serum total protein and albumin concentrations were observed in both Toxoplasma, Eimeria, and Cryptosporidium infections. It has been reported that protein requirement increase in the presence of any lesions on the body and chronic organ disease (Kaneko et al., 1997 and Roussel, et al., 1997). The significant alterations in ALT, AST and ALP as well as significant increase in the level of serum urea and creatinine indicate abnormal hepatic and renal functions. Fluid loss has been observed as one of the most common pathological effects of Eimeria, and Cryptosporidium species in ruminants as mentioned by Holland et al. (1989). In our study, this may be reflected by low level of sodium, potassium and chloride in serum of infected goats. This low level may be also attributed to diarrhea and reduced absorption in the infected tissues of intestine (Cirak et al., 24).. However, coccidial infections were associated with reduced serum zinc and elevated serum copper (Richards and Augastine, 1988). The decrease in serum zinc level was aggravated by drinking polluted water especially in the desert districts. This decrease could be attributed to excessive magnesium intake from the polluted wa111
Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 ter. The excessive dietary magnesium stimulated metalo-proteins production which bound zinc and reduced zinc utilization by the body as recorded by Ibrahim et al. (1992).. On the other hand, Toxoplasma infected goat serum showed a reduction in magnesium, calcium and potassium, where zinc and copper were within normal levels. T.gondii infection might cause changes in the trace elements as confirmed by Geng et al. (21). They also suggested that magnesium and calcium levels were elevated in liver, spleen and brain while their levels were reduced in serum of rat infected with T.gondii. They concluded that trace elements are essential for lymphocyte activity in the tissues. F inally, this study indicated that a great deal of attention must be given to control the protozoan diseases and analyze the under ground water before allowing rural populations to use it and to avoid great economic losses in productivity in these animals. REFERENCES Buxton, D. (1998): "Protozoan i n f e c t i o n s ( T o x o p l a s ma gondii, Neospora caninum, and Sarcocystis species) in sheep and goats: recent advance." Vet. Res. 29(3-4) 289-31. Cirak, V.Y.; Kowalik, S.; Bürger, H.J.; Zahner, H. and Clauss, W. (24): "Effects of Eimeria separata infections on Na+ and Cl- transport in the rat large intestine." Parasitol Res., Apr; 92(6): 49-495. Epub 24 Mar 4. Dubey, J.P. (1986): Toxoplasmosis. JAVMA., 189: 16617. Dubey, J. P. and Welcome, F.L., (1988): Toxoplasma gondiiinduced abortion in sheep. JAVMA., 193(6): 657-66. Dubey, J. P., Sharma, S.P., Lopes, C.W., Williams, J.F., Williams, C.S. and Weisbrode, S.E. (198): "Caprine Toxoplasmosis: abortion, and clinical signs and distribution of Toxoplasma in tissues of goat fed Toxoplasma gondii oocysts." Am. J. Vet. Res., 41, (7): 172-176. El- Manyawe, S.M. (1999): "Intestinal protozoal infection among goats in Egypt." Vet. Med. J., Giza. Vol. 47, 2: 379-388. El- Manyawe, S.M., Khattab, M.H. and Ismail, I., (21): "Serodiagnosis of Toxoplasma antibodies among sheep and goats in the North Coast of Egypt." Alex. J.Vet. Sci, 17(1)35-45. 112
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Egypt. J. Comp. Path. & Clinic. Path. Vol. 23 1 (February) 21; 12-115 ﻧﺴــــﺒﺔ اﻹﺻـﺎﺑﺔ ﺑﺒـﻌﺾ اﻷوﻟﯿــﺎت وﺗﺄﺛﯿـﺮھﺎ ﻋﻠﻰ اﻟﺘﻐﯿــﺮات اﻟﺒﯿﻮﻛﯿﻤﯿﺎﺋﯿﺔ ﻓﻲ اﻟﻤﺎﻋﺰ ﻓﻲ ﻣﻨﺎﻃﻖ اﻟﻘﺎھﺮة وﻣﺮﺳىﻤﻄـﺮوح واﻟﻮادي اﻟﺠﺪﯾــﺪ ﺳــــﮭﯿﺮ ﻣﺤﻤﺪ اﻟﻤﻨﯿـﺎوي* ﻣﺤﻤــﺪ ﻋﺒﺪ اﻟﺮﺣـﻤﻦ ﻣﺤﻤﺪ * أﺣﻤﺪ ﻣﺤﻤﺪ إﺑﺮاھﯿﻢ*** ﻋﺰة ﻣﺤﻤﺪ ﻛﻤﺎل** ﺳﻤﯿﺮة أﺣﻤﺪ ﺳﻨﻮﺳﻲ**** *ﻗﺴﻢ اﻟﻄﻔﯿﻠﯿﺎت ** ﻗﺴﻢ اﻟﻜﯿﻤﯿﺎء ﻣﻌﮭﺪ ﺑﺤﻮث ﺻﺤﺔ اﻟﺤﯿﻮان - ﻣﺮﻛﺰ اﻟﺒﺤﻮث اﻟﺰراﻋﯿﺔ - وزارة اﻟﺰراﻋﺔ - اﻟﺪﻗﻲ ***ﻣﻌﻤﻞ ﻣﻄﺮوح **** ﻣﻌﻤﻞ اﻟﻮادي اﻟﺠﺪﯾﺪ اﻟﻤﻠﺨﺺ اﻟﻌﺮﺑﻰ ٣٠٠ اﻟﻘـﺎھﺮة وﻣﻄﺮوح واﻟﻮادي اﻟﺠﺪﯾــﺪ ٠ اﻟﺘﻐﯿــﺮات اﻟﺒﯿﻮﻛﯿﻤﯿـــﺎﺋﯿـــﮫ ﻓﻲ ﻣﺼـــﻞ اﻟﺤﯿﻮاﻧـﺎت اﻟﻤﺼــــﺎﺑﮫ ٠ ﺖ اﻟﻨﺘ ــ ﺎﺋﺞ ﻋﻠﻰ وﺟـــﻮد ٩ وﻗﺪ دﻟ ٠ وﺟــــﻮد أﺟﺴــــﺎم ﻣﻀــﺎده ﻟﻄـﻔﯿـــﻞ اﻟﺘﻮﻛﺴــــﻮﺑﻼزﻣﺎ ﻓﻲ ﻣﺼــﻞ ھﺬه اﻟﺤﯿﻮاﻧﺎت ٠ واﻟﻤﻌــﺎدن اﻟﺜﻘﯿﻠـــﮫ وﻛــﺬﻟﻚ اﻟﯿﻮرﯾــﺎ واﻟﻜﺮﯾﺎﺗﯿﻨﯿﻦ ٠ ﻟﻠﺤــــﯿﻮاﻧــــﺎت ٠ اﻟﻤﺤﻜﻤﻮن : أ. د. ﺣﺴﯿﻦ ﻋﻤﺮ أﺳﺘﺎذ اﻟﻄﻔﯿﻠﯿﺎت ﻛﻠﯿﺔ اﻟﻄﺐ اﻟﺒﯿﻄﺮى ﺟﺎﻣﻌﺔ اﻟﻘﺎھﺮة أ. د. وﺣﯿﺪ ﻋﻠﻰ ﻣﻮﺳﻰ أﺳﺘﺎذ اﻟﻄﻔﯿﻠﯿﺎت ﻛﻠﯿﺔ اﻟﻄﺐ اﻟﺒﯿﻄﺮى ﺟﺎﻣﻌﺔ اﻟﻘﺎھﺮة 115