Method of taking of biological material for diagnostics of leptospirosis of wild animals

G01N1/00 - INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (separating components of materials in general B01D, B01J, B03, B07; apparatus fully provided for in a single other subclass, see the relevant subclass, e.g. B01L; measuring or testing processes other than immunoassay, involving enzymes or micro-organisms C12M, C12Q; investigation of foundation soil in situE02D0001000000; monitoring or diagnostic devices for exhaust-gas treatment apparatus F01N0011000000; sensing humidity changes for compensating measurements of other variables or for compensating readings of instruments for variations in humidity, seeG01D; or the relevant subclass for the variable measuredtesting or determining the properties of structures G01M; measuring or investigating electric or magnetic properties of materials G01R; systems in general for determining distance, velocity or presence by use of propagation effects, e.g. Doppler effect, propagation time, of reflected or reradiated radio waves, analogous arrangements using other waves G01S; determining sensitivity, graininess, or density of photographic materials G03C0005020000; testing component parts of nuclear reactors G21C0017000000)

FIELD: veterinary medicine.

SUBSTANCE: method comprises collection of urine after natural urination of animal into a sterile container. At that frozen urine samples are taken with the snow in winter, with the outdoor temperature is 10-50°C below zero.

EFFECT: use of the proposed method enables to extend the range of the animals tested to carrying the pathogenic leptospira, to provide the most long-term storage of the biological material selected, and to improve the accuracy of determining the source of leptospirosis.

 

The invention relates to the field of veterinary medicine, in particular to methods for biological material for the diagnosis of leptospirosis in wild animals.

There is a method of sampling to determine the causative agent of leptospirosis in the urine of animals, according to which the urine is collected in natural urine in a clean test tube and mikroskopiruyut directly in the farms. When this urine microscopy should be completed at a temperature of 30-40°C for 3 h, 25-30°C for 4-5 h, 20-25°C for 6-8 h, 16-20°C for 10-12 h after capture (agricultural Animals. Methods of laboratory diagnosis of leptospirosis. GOST 25386-91. M., 1992).

In accordance with the instruction "On the prevention and recovery of animals from leptospirosis" dated June 23, 1992, No. 22-3/40, urine is collected and mikroskopiruyut directly in the farms, with pathological material should be taken and investigated for leptospirosis within 6 hours after taking in the summer time, 10-12 hours after taking in the winter time or when stored in a refrigerated condition.

The above methods involve sampling of biological material - urine for the diagnosis of leptospirosis in animals during natural urination directly in the farms, which makes it impossible for its application in the study of wild animals, and also eliminates the possibility of the awn long-term storage of samples of biological material.

Perhaps keeping the samples at a temperature of not higher than 16°C, which carry urine into a test tube and there make glycerin in a ratio of 1/9 (glycerin/urine), thoroughly mix, freeze and store (the website of the research center for Biosafety and environmental control of resources APK Dnipropetrovsk state agrarian University http: biosafety-center.dp.ua/viddil-imunohimichniy-molekulyarno-genetichniy-analiz/Bolezni/Leptospirosis.php).

This method is also, as mentioned above, involves the study of samples taken from animals directly in the farms, which limits its wide application, in particular, for the diagnosis of leptospirosis in wild animals, in addition, leads to an appreciation of the process laboratory study, a set of application glycerin.

There is a method of determining stereogenic hormones in the feces and urine, collected from snow to assess the stress levels in wild rodents (L. A. Gerlinsky, M. P. Moshkin, C. I. Evsikov W. Ecology, 1993. No. 1, S. 97 and online http: w-3/rfbr.EN/default.asp?doc_id-4762).

However, this method is not for the diagnosis of infectious diseases, such as leptospirosis wild mammals

The technical result achieved in the solution of the task is to obtain a method of sampling biological material is urine, frozen in winter soderzhaniya diagnosis of leptospirosis in wild animals.

This object is achieved in that in the proposed method the selection of biological material - urine is produced in the winter when the outside air temperature of minus 10 to 50°C frozen with snow.

The technical result of the invention is achieved as follows.

Sampling of biological material for the diagnosis of leptospirosis in wild animals is carried out in winter when the outside temperature is minus 10-50°C in winter pasture keeping of wild animals in the reserve or Park areas, as well as the migration routes establish observation of animals. Within 1-12 hours after natural urination animal frozen urine with snow collected in a sterile container and frozen and sent to the laboratory. In the laboratory of biological material examined conventional methods - urine microscopy or polymerase chain reaction. In addition, the proposed method allows you to store sampled within 1-2 months at a temperature of minus 20°C.

The inventive method of sampling of biological material for the diagnosis of leptospirosis in wild animals can also be used for examination of the urine of horses herd of contents.

Animals-leptospirosis excrete leptospires in the environment with urine and infect the water, feed the, grassland, soil, litter and other objects of the external environment, through which infect healthy animals. Water transmission of the pathogen main. While Leptospira very resistant to low temperatures, they remain viable after a long freeze in ice. At zero temperature Leptospira perish.

When the outside temperature is minus 10 to 50°With animal urine freeze within 1 hour, with Leptospira in the urine, remain viable and are determined in laboratory studies. Since the selection of biological material is produced after natural bladder of the animal, the material is frozen urine taken in a sterile container with snow. Urine samples are taken at temperatures of minus 10°C, 20°C, 30°C, 40°C and 50°C (5 to 6 samples in each temperature interval).

Selected biological material in frozen form sent to study in the laboratory.

At temperatures above minus 10°From the collection of frozen urine has certain difficulties. Because it is not formed frozen "lump", the urine is spread on the surface and in deep snow. Sampling decreases the concentration of leptospires. Therefore, reducing the reliability of diagnosis.

Urine can be investigated directly after thawing or after centrifuged who I am. Transparent urine, not containing crystals, salts, cereals, sperm and other foreign particles, centrifuged at 10000-12000 rpm for 30 minutes, decant the supernatant, the precipitate resuspended in the remaining drop of urine and mikroskopiruyut. In the urine of patients and carriers of the pathogen detect Leptospira in the samples collected in any temperature interval. Urine containing significant amounts of foreign particles, purified by centrifugation at 3000 rpm for 10 minutes, then decant the supernatant and centrifuged for deposition of leptospires at 10000-12000 rpm for 30 minutes.

Additionally, you can establish the diagnosis for leptospirosis study urine polymerase chain reaction. The result of the detection of the disease is 100% (leptospirosis).

Thus, the claimed method of sampling biological material is urine, frozen with snow at minus 10-50°C in the natural habitat of wild mammals without their removal and trapping, allows you to conduct an operational diagnosis of leptospirosis and cost that provides timely special preventive measures for the identification of patients and carriers of leptospires.

Method of sampling of biological material for the diagnosis of leptospirosis in wild animals, including the collection of urine after natural is the first urination of the animal in a sterile container, characterized in that the frozen samples of urine taken together with snow in the winter when the outside temperature is minus 10 to 50°C.



 

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