اختبار التاثير التثبيطي لبروتين المناعة IgG لحليب الابل ضد السم العصبي A المفصول من عزنة محلية لبكتريا Clostridium botulinum == Testing the inhibitory effect of immunoglobulin IgG for camel?s milk against the neurotoxin type A separated from a local Isolate of Clostridium botulinum
Author name:
نورس محمد حسن عبد الصمد التميمي
Supervisor name:
امال كاظم غضبان الاسدي | حيدر ابراهيم علي
General topic:
Agricultural sciences
Specific topic:
Food
Degree:
Master
University:
University Of Basrah - Faculty Of Agriculture
Language:
Arabic
University location:
Basrah
First pages:
31T1877 - p.pdf
Abstract:
Sixteen local isolates of Clostridium were isolated from sources : soil , meat and honey .Those sources were obtained from different districte in Basrah government.Characterization of isolates were made after purification by using morphological and biochemical tests which revealed that the isolates were Clostridium botulinum.The microscopic test appeared that all isolates are bacilli Gram positive, obligate , anaerobic ,forming spores and motile. There are colonies had wide spread form with ir regular border when grown of blood agar and egg yolk agar and looked transparent to gray. The biochemical tests revealed that the isolates are β - heamolysis, lipid hydrolyzed, produced H2S, most of them were negative in indole production, nitrate reduction, starch hydrolysis and catalase production. They were grown in pH(4 - 6),temperature(10,30,42) °C and NaCl (4 - 10)% . They not fermented lactose, galactose, amygdalin, Ribose, Inositol, Manose, Melibiose, Sorbitol, Salcin, Rafinose, Xaylose and Rhamnose. But they were fermentated Glucose, Maltose and Cellubiose. They were sensitive for some antibiotics like Erethromycin, Refampin, Metronidazol, Clindamicin, Tetracyclin, Penicillin and Chloramphnichol , but they were resistant for Gentamycin , Nalidixic acid and Trimethoprim.Used for Polymerase chain reaction (PCR) for the bacterial isolates by extraction of DNA and electrophoresis by using agarose, seven isolates of Clostridium botulinum were elected because they appeared difine band of DNA. After that the gene wich responsible on neurotoxin A was detacted by using Polymerase chain reaction (PCR) and electrophoresis using agarose, the isolate Cl.5 of Clostridium botulinum wich was isolated from the soil of Agriculture College field, was elected because of its clear band which was (101) bp with the band of the primer of the gene of neurotoxin A.Neurotoxin A was produced by the isolate Cl.5 using the inoculums medum : (casein hydrolysis, yeast extract, glucose ,dis. water) and production medium; (casein hydrolysis, yeast extract, dis. water) with cold glucose solution 10% and using anaerobic conditions with 37°C for 4 days. The protein of extracted neurotoxin was (0.14 - 0.93) , then the extract was precipitated by ammonium sulphate %60 and the protein was (0.18 - 0.22) ,finally the neurotoxin was purified by ion exchange using DEAE Sephadex A - 50 only one protein peak wasappeared in the void volume , the protein was(0.27) . The activity of the three samples of neurotoxin A the crude ,precipitated and purified was assayed by bioassay using mouse , all the samples revaled high activity by appearing of of intoxication then death . But the purified neurotoxin appeared the highest activity because the mouse died after 3 hr . while the precipitated toxin after 8 hr using and after 10 hr using the crud toxin Stadying minimal leathal dose (MLD) by using concentrations of purified toxin , the concentration 0.1 revealed minimal activity because the mouse died after 3.41 hr .Finally the inhibitory effect of immunoglobulins of camel milk against neurotoxin A was studied by using passive haemagglotination and bioassay , immunoglobulin had agreat activity to inhibite the neurotoxin A.
