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Minufiya Veterinary Journal ISSN: 1110-9750 Minufiya Veterinary Journal ISSN: 1110-9750 Volume 9 September 2015 ISSN (prINt): 1110-9750 published by Faculty oF Veterinary medIcine University oF Sadat City egypt Email: [email protected] Minufiya Veterinary Journal ISSN: 1110-9750 Vol. 9; September 2015 ﻣﺠﻠﺲ إدارة اﻟﻤﺠﻠﺔ رﺋﯿﺲ ﻣﺠﻠﺲ إدارة اﻟﻤﺠﻠﺔ م اﻹﺳﻢ اﻟﻮظﯿﻔﺔ اﻟﺼﻔﺔ ۱ ا.د/ ﺷﻌﺒﺎن ﻣﺤﻤﺪ ﺟﺎد ﷲ ﻋﻤﯿﺪ اﻟﻜﻠﯿﺔ رﺋﯿﺲ ﻣﺠﻠﺲ اﻹدارة ھﯿﺌﺔ اﻟﺘﺤﺮﯾﺮ م اﻹﺳﻢ اﻟﻮظﯿﻔﺔ اﻟﺼﻔﺔ ۱ أ.د/ أﺣﻤﺪ ﻋﺒﺪ اﻟﻤﻨﻌﻢ زﻏﺎوة أﺳﺘﺎذ اﻷﻣﺮاض اﻟﺒﺎطﻨﺔ واﻟﻤﻌﺪﯾﺔ رﺋﯿﺲ ﺗﺤﺮﯾﺮ اﻟﻤﺠﻠﺔ ۲ أ.د/ ﺳﻌﺪ ﻋﺒﺪ اﻟﻔﺘﺎح ﻋﻤﺎره أﺳﺘﺎذ اﻟﺨﻠﯿﺔ و اﻷﻧﺴﺠﺔ ﻋﻀﻮا ً ۳ ا.د/ ﻣﺤﻤﺪ ﻋﺎطﻒ ھﻼل أﺳﺘﺎذ اﻟﺮﻋﺎﯾﺔ وﺗﻨﻤﯿﺔ اﻟﺜﺮوة اﻟﺤﯿﻮاﻧﯿﺔ ﻋﻀﻮا ً ٤ ا.د/ ﺣﻤﺎده ﺿﺎﺣﻲ ﺣﺴﯿﻦ أﺳﺘﺎذ اﻟﺮﻋﺎﯾﺔ وﺗﻨﻤﯿﺔ اﻟﺜﺮوة اﻟﺤﯿﻮاﻧﯿﺔ ﻋﻀﻮا ً ٥ أ.د/ ﺧﺎﻟﺪ ﻣﺤﻤﻮد ﻣﺤﻤﺪ ﺟﻌﻔﺮ أﺳﺘﺎذ اﻟﺘﻐﺬﯾﺔ واﻟﺘﻐﺬﯾﺔ اﻹﻛﻠﯿﻨﯿﻜﯿﺔ ﻋﻀﻮا ً ٦ د/ ﻋﻤﺎد ﻣﺤﻤﻮد ﻣﺤﻤﺪ ﻋﺒﺪ اﻟﺮازق أﺳﺘﺎذ ﻣﺴﺎﻋﺪ اﻟﺘﻮﻟﯿﺪ واﻟﺘﻨﺎﺳﻞ واﻟﺘﻠﻘﯿﺢ اﻻﺻﻄﻨﺎﻋﻲ ﻋﻀﻮا ً ۷ د. ﺷﺮﯾﻒ ﻋﺒﺪ ﷲ زﯾﺪان أﺳﺘﺎذ ﻣﺴﺎﻋﺪ ﺑﻘﺴﻢ اﻟﺼﺤﺔ واﻷﻣﺮاض اﻟﻤﺸﺘﺮﻛﺔ ﻋﻀﻮا ً ۸ د/ ﻣﺒﺮوك ﻋﻄﯿﺔ ﻋﺒﺪ اﻟﺪاﯾﻢ ﻣﺪرس اﻟﻜﯿﻤﯿﺎء اﻟﺤﯿﻮﯾﺔ وﻛﯿﻤﯿﺎء اﻟﺘﻐﺬﯾﺔ ﻋﻀﻮا ً ۹ د/ ﻣﺤﻤﺪ أﺑﻮ اﻟﻌﺰ ﻧﺎﯾﻞ ﻣﺪرس اﻷﻣﺮاض اﻟﺒﺎطﻨﺔ واﻟﻤﻌﺪﯾﺔ ﻋﻀﻮا ً ۱۰ د/ أﺣﻤﺪ ﻣﺤﻤﻮد ﻋﺒﺪ اﻟﻮاﺣﺪ اﻟﺼﯿﻔﻲ ﻣﺪرس اﻷﻣﺮاض اﻟﺒﺎطﻨﺔ واﻟﻤﻌﺪﯾﺔ ﻋﻀﻮا ً ۱۱ د/ أﻛﺮم أﺣﻤﺪ ﺣﺴﻨﯿﻦ ﺳﻼﻣﺔ ﻣﺪرس اﻷﻣﺮاض اﻟﺒﺎطﻨﺔ واﻟﻤﻌﺪﯾﺔ ﻋﻀﻮا ً ۱۲ د/ ﻣﺼﻄﻔﻰ ﻋﺒﺪ اﻟﺠﺎﺑﺮ ﻣﺪرس اﻟﺒﺎﺛﻮﻟﻮﺟﯿﺎ ﻋﻀﻮا ً ۱۳ د/ رﺑﯿﻊ اﻟﺤﺴﯿﻨﻲ اﻣﺒﺎرك ﻣﺪرس اﻟﺮﻗﺎﺑﺔ اﻟﺼﺤﯿﺔ ﻋﻠﻰ اﻷﻏﺬﯾﺔ ﻋﻀﻮا ً ۱٤ ط.ب / ﻋﺒﺎس ﻓﺘﺤﻲ ﻋﺒﺎس ﻣﺼﺮي ﻣﻌﯿﺪ اﻟﺠﺮاﺣﺔ واﻟﺘﺨﺪﯾﺮ واﻷﺷﻌﺔ ﻋﻀﻮا ً ۱٥ ط.ب/ ﻋﺒﺪ اﻟﻔﺘﺎح ﻣﺤﻤﺪ ﻋﺒﺪ اﻟﻔﺘﺎح ﻣﻌﯿﺪ اﻟﺒﺎﺛﻮﻟﻮﺟﯿﺎ اﻹﻛﻠﯿﻨﯿﻜﯿﺔ ﻋﻀﻮا ً ۱٦ أ. ﺷﯿﻤﺎء ﻋﺒﺪ اﻟﺤﻤﯿﺪ ﻣﻮظﻒ إدارى ﺳﻜﺮﺗﯿﺮا ً Minufiya Veterinary Journal ISSN: 1110-9750 Vol. 9; September 2015 Chairman of Journal Board Prof. Dr. Shaaban Gad Allah Chief Editor Prof. Dr. Ahmed A. Zaghawa Associate Editors Prof. Dr. Hamada D.H. Mahboub Dr. Ahmed M. Elsify Dr. Rabee A. Ombarak Minufiya Veterinary Journal ISSN: 1110-9750 Vol. 9; September 2015 Contents S. No Title and authors Page 1 Molecular Diagnosis and Genotyping of Bovine Viral Diarrhea Virus 1 Bazid A. I., Nayel M. A., Ahmed M. Elsify, Khalil S. A. and Zaghloul A. H. 2 A Study on Antiviral Efficacy of Brucella abortus Ether Extract Against Experimental Rabies Infection in Mice 14 Nermeen, G. Shafik, Hanan, I. Mahmoud, Albehwar, A. M. and Hemmat, S. El-Emam. 3 Efficacy of Combined Local Prepared Vaccine Against Mannheimia Haemolytica and Clostridial Diseases 23 Hanan A. Ahmed, Sayed, M. L., Hala, EL-Sawy, Eman M. Elrawy, Mohamed A. Abdelfatah, and Makharita M. Ali. 4 Preparation and Conjugation of Staphylococcal Protein A “SPA” and Streptococcal Protein G “SG” with Fluorescein Isothiocyanate as Non-Species Specific Diagnostic Kits 32 Omnia, F. H., Khodeir, M. H., and Saad, M. A. M. 5 Clinicopathological Studies on Chicken Infectious Anemia Disease Live Virus Vaccines 40 Tamer S. Allam, Nahed S. Saleh, Salah S. El-Ballal and Hesham A. Sultan. 6 Microscopical Identification and Seasonal Dynamics of Gastrointestinal Nematodes in Small Ruminants at Menoufia Province 58 Nasr M. Elbahy, Ahmed O. Elkhtam, Mahmoud AbouLaila, and Amer R. Abdelaziz 7 Prevalence of Different Flatworms Infecting Ruminants in Menoufia Governorate 66 Nasr M. Elbahy, Ahmed O. Elkhtam, Mahmoud AbouLaila, and Amer R. Abdelaziz. 8 Prevalence of Clinical and Subclinical Mastitis in Menoufia Governorate with Reference of PCR 78 Walid S. Mousa , Ahmed A. Zaghawa , Mohammed A. Nayel, Eman E. Abdeen , Ahmed M. Elsify, Akram A. Salama and Mahmoud A. Aly. 9 Clostridium perfringens Toxins With Special Reference to Beta 2 Toxin 85 Ahmed M. Elsify 10 Bacteriological Quality and Safety of Raw Cow’s and Buffalo’s Milk Sold in Menoufia Governorate, Egypt 101 Rabee A. Ombarak and Abdel-Rahman M. Elbagory 11 Studies on The Potential Protective Effect of Curcumin and Zinc against Nickel- Induced Nephrotoxicity in Rats 114 Salah S. El-Ballal, Ashraf M. Morgan and Nermeen B. EL-Borai. 12 Effects of Feeding Rates on Behavior and Growth Performance of Nile Tilapia (Oreochromis Niloticus) 132 Hamada D. H. Mahboub, Sameh G. A. Ramadan, Mohamed A.Y. Helal, Walaa S. H. Ahmed Minufiya Veterinary Journal ISSN: 1110-9750 Molecular diagnosis and genotyping of Bovine viral diarrhea virus Bazid A.I. a, Nayel M.A. b, Ahmed M. Elsify b*, Zaghloul A.H. c, Khalil S.A. d a Department of Virology- Faculty of veterinary medicine – University of Sadat City- Egypt. b Department of animal medicine and infectious diseases - Faculty of veterinary medicine - University of Sadat City- Egypt. c Department of Theriogenology - Faculty of veterinary medicine - University of Sadat City- Egypt. d Department of Microbiology - Faculty of veterinary medicine - Alexandria University – Egypt. * Corresponding Author: [email protected] Accepted: 11/8/2015 Abstract: In the present study isolation, biotyping, antigenic and genomic characterization of BVDV isolates was carried out on buffy coat samples obtained from apparently healthy cattle. The samples were inoculated in MDBK cells for three successive passages and the CPE was recorded indicating that 48%, 62% and 52% were positive CPE in the 1st, 2nd and 3rd passages respectively. The viral antigen was detected in cell culture after propagation also by direct FAT and reveled that (62%) showed positive result. The mean virus titer was recorded after each passage showing values of <102,102 and 104 st nd rd TCID50/ml by 1 , 2 and 3 passages respectively. The virus was identified by VNT using the reference anti-BVDV-1 serum on cell cultures infected fluids at the third passage. It was found that all samples showed a characteristic CPE of BVDV were neutralized by the used antiserum confirming that they are BVDV-1. Nine buffy coat samples were selected to carry out the nested RT-PCR for detection and genotyping of suspected BVDV. The results indicated the presence of BVDV genotype-I. The positive samples for FAT, VNT and RT- PCR in infected cell culture were stained by H&E for biotyping indicating the presence of the two biotypes (CP& NCP) in the samples. The SNT was carried out on serum samples from the same animals indicating the prevalence of antibodies in 62% by mean titer ranged from < ½ to 1/8. In conclusion, the present study reports the identification of BVDV genotype – I in apparently healthy cattle and presence of P.I. animals. Keywords: Bovine viral diarrhea virus, biotyping, reverse transcriptase polymerase chain reaction (RT- PCR). 1 Bazid et al. / Minufiya Vet. J. 2015, 9: 1-13 Introduction: BVDV strains have two biotypes, cytopathic (cp) Bovine viral diarrhea (BVD) is one of the most and non-cytopathic (ncp) which can be imperative worldwide diseases in domestic and distinguished on the basis of their effect on wild ruminants, leading to substantial damage in cultured bovine cells (Gillespie et al., 1960). The infected herds as well as extensive economic two biotypes are important for the occurrence of losses for the cattle industry (Goyal and Ridpatb, MD. The MD is either induced by super infection 2005; Ah- med and Zaher, 2008). Causing of persistently infected animals with an multiple disease and clinical syndromes including antigenically closely related cp BVDV or by embryonic mortalities, abortion, fetal generation of a cp mutant from the persisting ncp mummification, stillbirths, congenital deformities, virus (McClurkin et al., 1985). respiratory disease (Flores et al., 2002) and According to the analysis of the basis of the 5' hemorrhagic syndrome (Walz et al., 1999). UTR, BVDV was segregated into two genotypes, An important condition for maintenance of BVDV BVDV-1and BVDV-2. Sequence homology in bovine populations is the immunotolerant and within each genotype was over 93%, while persistent infection (PI) that result from between genotype 1 and 2 it dropped to 74 % transplacental infection of the fetus before onset of (Ridpath et al., 1994). immunological maturity. Animals persistently According to the phylogenetic tree constructed infected with BVDV not only transmit the virus from 420 nt of the E2 glycoprotein gene fragment; effectively to susceptible in-contact animals, but the BVDV-1 can be subdivided into BVDV-1 a, c, they harbour clones of BVDV which through d, e, f, g and 1 b which is divided into 1b1 and 1b2 serial transplacental passages may be able to and the BVDV-2 is subdivided into BVDV-2a ,2b replicate for years without immunological and 2c. (Motoshi et al., 2001) selective pressure (Toplak et al., 2004). It is well Due to the complex pathogenesis of BVDV established that persistently vireamic animals may infections and presence of many genovares, later succumb to fatal mucosal disease (Brownlie, laboratory diagnosis of BVDV becomes important 1985). in the strategy of control and prevention BVDV BVDV is a small enveloped RNA virus which infections. Moreover, the knowledge of the type of together with classical Swine fever virus (CSFV) strains occurring in the field can help to establish and Border disease virus (BDV) form the genus effective vaccine and effective control (Kabango, pestivirus in the family Flaviviridae (Pringle, 2005). 1999). BVDV consists of a single stranded There are several methods used for diagnosis of positive-sense RNA genome, of approximately BVDV infection which include virus isolation 12.5Kb long (Collett, 1992). (Dubovi, 1990), virus neutralization (Brock 1995), immunoperoxidase (Castro et al., 1997), ELISA 2 Bazid et al. / Minufiya Vet. J. 2015, 9: 1-13 (Chu et al., 1985), FAT (Dubovi, 1990) and A total of 750 apparently healthy Friesian cattle AGPT (Gutekunst and Malmquist, 1963) but all housed under bad conditions in two farms present these tests suffer several disadvantages where in Behera Governorate were investigated. virus isolation may requires as long as three weeks Samples: - especially when more than one passage in cell Two sets of blood samples were collected from 50 culture is required to recover the virus.
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