COLLECTION AND DISPATCH OF VIROLOGICAL SAMPLES Dr. Banshi Sharma Senior Veterinary Officer CVL Virological samples may be taken from animals, or their animals, or their environment for disease diagnosis. The samples must be appropriate and adequate in number of the purpose required. Sample must be collected aseptically and care must be taken to avoid cross contamination between samples. The suitable sample must be carefully packed, labeled and transmitted to the laboratory by the fastest practical method. A written note indicating the origin of the material, the relevant history and the tests required, must accompany all samples, The operator should wear protective clothing; overalls, rubber gloves, rubber boots; facemask and goggles. Selection of appropriate tissue or most promising lesions for specific disease is very important. Sterile instruments should be used for sample collection. Disinfectants must not be used on or near tissues to be sampled for virus isolation. The samples should be forwarded in wide mouthed screw-capped bottles (Mc Cartney’s bottle) with wet ice; if they can reach the laboratory within 24 hours. For prolonged time, they can be freezed. Frequent freezing and thawing should be avoided. The samples for viral isolation may be sent n transport media. For histopathological examination, blocks of tissues not more than 0.5ch thick and 1-2 cm 2 are cut and placed in neutral buffered 10% formalin which should be at least 10-20 times the volume of the sample. They should not be frozen. Samples are sent for laboratory as soon as possible. Suitable preservative may be used while transporting the samples as follows: (1) for histopathological examination, 10% neutral buffered formal saline solution; (2) for virological examination, 50% glycerin saline. No preservative is used for viral isolation. Collection of Samples for different Diseases: The sample types may differ from disease to disease. Example, brain and spinal cord in rabies; lungs in pneumonia, liver in hepatitis and kidneys in case of nephritis. The various tissue samples desired for virological examination for various diseases is given in the following table: Diseases Rabies FMD Rinderpest Swine fever Equine Encephalyomyelitis Canine Distemper Infectious Canine Hepatitis Ranikhet Infectious Bursal disease Avian Encephalomyelitis Marek’s Disease Avian Leucosis complex Fowl pox Sample Size Surveillance scheme of a herd/flock to achieve a 85% probability of detecting infection should include 5% sample. For example; if diseases were prevent in 5% of a herd of 500 animals, it would be necessary to sample 56 animals to be 95% confident of finding one positive, assuming that both the sensitivity and specificity of the test is 100% Sample to be collected Brain, saliva (living animal) Epithelial covering of vesicles on the lips, dorsum of tongue, and oral palate in 50% glycerin saline. Blood, large intestine with lesions spleen lymph nodes Heart blood in tubes kept under ice(4ºC), spleen , lymph glands Brain, spinal cord Nasal discharge, lungs, mediastinal lymph node, kidneys, liver Blood, liver, kidneys, spleen Blood, lungs, brain, liver Blood, bursa of fabricious, spleen, kidney, liver , sick bird Brain, spinal cord, pancreas, and proventriculus Sick bird, ovary, testes, brain, nerve Liver, spleen, kidneys, gonads, sick bird Scabs lesion Remarks Do not use glycerin for virus isolation. Procedure for Dispatch of Materials for Histopathological Examination: Collect the sample material soon after death in a suitable fixative (10% buffered formal saline ) in a wide mouthed glass bottle or any suitable container. Include the portion representing the characteristic lesion (1ml + 9ml water = 10% formalin) The volume of fixative should be 10-20 times more than the volume of tissues. Place a pad of cotton in the container and put the tissues sample over it, pour the saline and again give a pad of cotton over the sample tissue. Insert label written with a hard black lead pencil, nicely stopper it, and paste a label on the container also. Seal the mouth of the container; write the address of the laboratory correctly with instruction “ Handle with care” Histological samples should not be frozen. Information to be Send with the Sample: Information and case history always accompany the samples, and ideally should be placed in a plastic envelope. The information should include the following points: Name and address of owner, where disease occurred, with phone, fax. Disease suspected Samples submitted and test required Transport medium Different species on the farm and number, age and sex of each affected animal. Length of time on the farm; if recent arrival, where from Date of first cases and of subsequent cases or losses. Description of the spread of infection in the herd or flock. Number of animals dead, the number showing clinical signs, and their age, sex, and breed. The clinical signs Type and standard of husbandry practice; feed Post mortem findings Description of samples submitted Any medication already given and when given Vaccination record Name and address of sender, with telephone and fax number and date of submission. This information should accompany the package of sample and be attached in an envelope to the outside of the parcel Transport of the Sample: Samples must be carefully packed to avoid any leakage or cross-contamination. They should be delivered within 48 hours and must be kept cool during transit. Screw-capped bottles should be used and should be additionally sealed with adhesive tape or paraffin wax. Samples In individually identified container should be placed in large strong outer container and packed with enough absorbent material to protect from damage.