Method of non-invasive express-diagnostics of inflammatory process in intestines of calves

FIELD: veterinary science.

SUBSTANCE: method includes detection of leukocytes, protein, haemoglobin in faeces, and PH-reaction of faeces. A sample of faeces is dissolved in distilled water and applied drop by drop onto appropriate test fields of a test strip designed for urine testing. By variation of colour within 1 minute the reaction is assumed as positive. If pH is less than 7.0 or more than 7.5, and faeces contain soluble protein, haemoglobin and leukocytes at the same time, availability of the inflammatory process in the intestines is confirmed.

EFFECT: invention makes it possible to quickly and accurately diagnose inflammatory process in intestines.

2 cl, 2 ex

 

The invention relates to veterinary medicine and can be used for non-invasive rapid diagnosis of the inflammatory process in the intestine of calves for the simultaneous detection in stool soluble protein, hemoglobin, white blood cells and shift the pH to acidic (less than about 7.0) or alkaline (7.5).

For in vivo diagnosis of inflammatory process in the intestine in calves currently use the complex clinical and laboratory methods of research. Clinical studies include General (inspection, palpation, percussion, etc.) and special methods (rectoscopy, laparoscopy, radiography, ultrasonography). The presence of symptoms of the lesion of the intestine: the symptoms of anxiety, loss of appetite, change of posture of the animal, the frequency of defecation and consistency of stool, change the shape of the abdomen, skin turgor, heart rate and respiratory movements and others noted only in patients with severe inflammatory bowel disease. In the early stages and during the chronic inflammatory bowel diseases (enteritis, colitis) in calves often have erased the clinical picture and asymptomatic.

Significant information content have instrumental methods of investigation: endoscopy (rectoscopy, colonoscopy), histological examination of biopsy specimens of the mucosa, roentgenol�Strastnoi examination of the intestine, ultrasonography (Shabanov A. M. study of the digestive system / A. M. Shabanov // Workshop on clinical diagnosis of animal diseases / M. F. Vasiliev, E. S. Voronin, L. G. Dugin, S. P. Kovalev, G. V. Snot, V. I. Cherkasova, A. M. Shabanov, M. V. Shchukin; ed. E. S. Voronin. - M.: Colossus, 2003. - P. 96-141).

The disadvantages are: the need for special equipment and highly qualified personnel, invasiveness (the trauma) and the complexity of the research procedures, radiation exposure in animals and the impossibility of frequent follow-up examinations to monitor the course of disease and effectiveness of treatment.

Laboratory tests (hematological, immunological, biochemical indices of the blood leukogram, erythrocyte sedimentation rate, classes of immunoglobulins, cytokines, acute phase proteins, etc. - can diagnose the presence of inflammation in the body your calves, but they are nonspecific and vary with the inflammation of any localization (bronchopulmonary inflammation, omphalitis, enteritis, hepatitis, etc.) (Horadagoda, N. U. Acute phase proteins in cattle: discrimination between acute and chronic inflammation / N. U. Horadagoda, K. M. Knox, H. A. Gibbs, S. W. Reid, A. Horadagoda, S. E. Edwards, P. D. Eckersall // Vet. Rec. - 1999. - Vol. 144. - No. 16. - P. 437-441).

The known method for non-invasive diagnosis of inflammatory process in the intestine in animals according to the results of some�moving in stool calprotectin concentration by enzyme immunoassay (Grellet, A. Fecal calprotectin concentrations in adult dogs with chronic diarrhea / A. Grellet, R. M. Heilmann, P. Lecoindre, A. Feugier, M. J. Day, D. Peeters, V. Freiche, J. Hernandez, D. Grandjean, J. S. Suchodolski, J. M. Steiner // Am. J. Vet. Res. - 2013. - Vol. 74. - No. 5. - P. 706-711).

The disadvantages of this method are: the need for specialized laboratory space, equipment, test systems and highly skilled personnel.

As the prototype was taken method for non-invasive diagnosis of inflammatory process in the intestine in calves, including the definition in the stool leukocytes on microscopic examination, protein, hemoglobin (occult blood) and pH (response cal) - chemical study (Smirnov, A. M. a Study of the digestive system / Alexander Smirnov // Clinical diagnosis of internal non-infectious diseases of farm animals / by A. M. Smirnov, P. J. Konopelko, V. S. Postnikov, I. M. Belyakov, G. L. Dugin, R. P. Pushkarev, V. S. Kondrat'ev, N. And. Urazaev. - Leningrad: Kolos. Of Leningrad. -DEP, 1981. - S. 176-257). In the feces of healthy calves leukocytes are absent or occur sporadically, there is no protein and hemoglobin (occult blood), pH 7-7,5 (neutral or slightly alkaline). Inflammatory bowel disease in calves is accompanied by the appearance in the feces is mucus, blood, pus; microscopic examination - find a large number of white blood cells, chemical research - haemoglobin and protein. Inflammatory bowel disease often accompanies�I the development of fermentative or putrefactive processes. If you encounter fermentation processes in the intestine produce large quantities of organic acids and pH of feces is reduced (below 7.0) was, when putrefactive processes due to the formation of an excess of ammonia is increased (over 7.5).

The disadvantages of this method are: long-term (up to 24 hours) the time from specimen collection until results of research, the need for specialized laboratory space, equipment and qualified personnel; the possibility of obtaining false negative results, since the leukocytes in the stool can collapse and not be detected by microscopic examination.

The technical problem of the invention is to provide a method of non-invasive rapid diagnosis of the inflammatory process in the intestine in calves that do not require special laboratory facilities, equipment and highly qualified personnel, allowing studies directly at the location of the animal (in the economy), to reduce the economic and time costs for research and analysis of their results.

To solve the technical problem of the invention in the method of non-invasive rapid diagnosis of the inflammatory process in the intestine in calves, including the definition in the stool leukocytes, protein, hemoglobin and pH response of feces, according to the invention, a stool sample descriptio�car Ried out in distilled water and put a drop on the corresponding test field of the test strip, designed for the analysis of urine, by a colour change in less than 1 minute is considered a positive reaction, and at a pH of less than 7.0 or more than 7.5 and the presence in Calais at the same time the soluble protein, hemoglobin and leukocytes diagnose the presence of inflammation in the intestine. For breeding we take the feces and water in the ratio 1:(5÷7).

In the proposed method using the test strips for urine analysis with at least 4 test fields to determine the above parameters based on the method of "dry chemistry".

The method allows to simplify the diagnosis of inflammatory process in the intestine in calves compared to known methods, to reduce the time and material costs for research and analysis of their results.

Method of non-invasive rapid diagnosis of the inflammatory process in the intestine in calves is as follows.

For non-invasive rapid diagnosis of the inflammatory process in the intestine in calves a stool sample taken directly from the rectum during defecation in a volume of 0.2-0.4 ml in a disposable plastic container with a volume of 2-5 ml, add 5+7-fold volume of distilled water with pH 7.0 and disposable plastic stir stick until a homogeneous state. To determine the protein, hemoglobin, leukocytes, and pH of disposable plastic pipette (volume 1 ml) solution is applied �about one drop per test field test strips for urine analysis. The reaction is considered negative if the color of the test field has not changed within 1 minute, and positive if there is staining of varying intensity. Each test can be assessed qualitatively (Yes - no) or semi-quantitative(+, ++, +++, ++++), contrasting color with the supplied test strips scale.

Chemical reactions underlying the detection of soluble protein, hemoglobin, leukocytes, and pH, laboratory methods identical to those used for the chemical examination of the feces (Kondrakhin I. P. Methods of veterinary clinical laboratory diagnostics: a Handbook / I. P. Kondrakhin, A. Arkhipov, V. I. Levchenko, G. A. Talanov, L. A. Frolov, V. E. Novikov; Under the editorship of Professor I. P. Kondrashina. - M.: Colossus, 2004. - S. 253-309).

Ranges of concentrations of the substances to be detected by the test strips for urine analysis: soluble protein (mainly albumin, to a lesser extent globulin) is 0.15-10.0 g/l, hemoglobin of 0.6-10 mg/l, leukocyte elastase 25-500 leukocytes/l, pH 5 to 9 at intervals of 0.5-1. The definition in the stool leukocyte elastase, unlike microscopic examination reveals a leukocyte count even after destroying them and therefore excludes the possibility of obtaining false negative results.

Method of non-invasive rapid diagnosis of the inflammatory process in the intestine in calves is illustrated �the company.

Example No. 1. Calf 9330, age 8 days, with a diagnosis of acute enteritis (severe). In a clinical study: the fatness unsatisfactory, sucking reflex is absent, the calf lies with half-shut eyes, the neck is extended, the ears drooping, limbs scattered to the side, sunken eyeballs, the mucous membrane of the mouth is dry, cold, cyanotic. Skin dry, significantly reduced turgor - skin fold is straightened in just 9 seconds. The ruffled hair, no Shine. Lower right area of the abdominal wall palpation painful, calf gnashing of teeth, auscultation listen loud rumbling sounds. Profuse diarrhea. Kal liquid, gray, with admixture of air bubbles, with a putrid odor. The sphincter of the anus is relaxed. The root of the tail, perineum and hind legs covered in feces; on the root of the tail alopecia (6 cm). Body temperature 39,9°C, tachycardia (122 beats per minute), sinus arrhythmia, shortness of breath (58 breaths per minute), shallow, mixed dyspnea. Hematological signs of inflammation: leukocytosis (11,3×109/l), shift leukocyte formula to the left. Chemical examination of stool the claimed method using test strips for urinalysis (10 EA production Arkrey, Japan): pH 8.0, the reaction to the protein positive (++), hemoglobin sharply put�individual (++++), the leukocyte - positive (++). Shift the pH of the feces in the alkaline side indicates the presence of putrefactive processes in the intestine. The combination of features is the presence in the feces of calves soluble protein, hemoglobin, leukocytes, and a shift in alkaline pH (7.5) side - indicates the presence of inflammation in the intestine.

From this example it is clear that the clinical signs of enteritis in calves (increased defecation, liquid stools, pain lower right area of the abdominal wall palpation, loud rumbling sounds auscultation of bowel) coincide with the results of chemical examination of the feces (the presence in the feces of soluble protein, hemoglobin, white blood cells and the pH shifts to the alkaline side) indicating the presence of inflammation in the intestine.

Example No. 2. Calf 9336, age 8 days, healthy gastro-intestinal tract. In a clinical study: good fatness, active sucking reflex is well expressed, the mucous membrane of the mouth pale pink, wet, warm. The skin is elastic, good turgor - skin fold is straightened within 1 second. The abdominal wall is painless on palpation, auscultation bowel tapped babbling sounds. A good appetite. Cal designed, dark yellow, with a faint sour smell. The tone of the sphincter of the anus you�Agen well. The body temperature of 38.5°C, heart rate and respiratory movements 70 and 24 min, respectively. Hematological signs of inflammation are absent. Chemical examination of stool the claimed method using test strips for urinalysis (10 EA production Arkrey, Japan): pH 7.0, the reaction to the protein, hemoglobin, leukocytes - negative.

From this example it is clear that the results of clinical trials calf healthy gastro-intestinal tract (normal defecation, feces decorated, the abdominal wall is painless on palpation, auscultation listens for bowel sounds fluid transfusion), hematological signs of inflammation are absent, the results of chemical examination of the feces (absence of soluble protein, hemoglobin, leukocytes and optimum pH) indicate the absence of an inflammatory process in the intestine.

Example No. 3. Calf 9334, age 9 days, clinical signs of intestinal inflammation are absent. Fatness good sucking reflex is well expressed, oppression is absent, the mucous membrane of the mouth pale pink, wet, warm. The eyes are not sunken, the skin elastic, good turgor - skin fold is straightened within 1 second. The abdominal wall is painless on palpation, auscultation - listen�are short and sparse babbling sounds. Kal-decorated, yellow-green, with a faint sour smell. The body temperature of 38.8°C, heart rate and respiratory movements 75 and 26 min, respectively. In hematological examination - signs of inflammation: leukocytosis (9,8×109/l), shift leukocyte formula to the left. Chemical examination of stool the claimed method using test strips for urinalysis (10 EA production Arkrey, Japan): pH 6.5, the reaction to the protein positive (++), hemoglobin - positive (++), leukocytes weakly positive (+). Shift the pH of the feces in the acidic side indicates the presence of fermentation processes in the intestine. The combination of features is the presence in the feces of calves soluble protein, hemoglobin, leukocytes, and a shift of pH in acidic (less than 7.0) was side - indicates the presence of inflammation in the intestine.

When you study in 5 days - clinical signs of enteritis: lower right area of the abdominal wall is painless on palpation, auscultation bowel tapped short and sparse babbling sounds. Kal liquid, light yellow color. Fatness is satisfactory, sucking reflex saved, oppression is absent, the mucous membrane of the mouth pale pink, wet, warm. The eyes are not sunken, the skin is elastic, slightly decreased turgor is the skin fold raspravljati� for 2 seconds. The body temperature of 38.6°C, heart rate and respiratory movements 100 and 38 min, respectively. Hematological signs of inflammation: leukocytosis (15,3×109/l), shift leukocyte formula to the left. Chemical examination of stool the claimed method using test strips for urinalysis (10 Ε And production Arkrey, Japan): pH 6.0, the reaction to the protein positive (++), hemoglobin - sharply positive (++++), leukocytes weakly positive (+). Shift the pH of the feces in the acidic side indicates the presence of fermentation processes in the intestine. The combination of features is the presence in the feces of calves soluble protein, hemoglobin, leukocytes, and a shift of pH in acidic (less than 7.0) was side - indicates the presence of inflammation in the intestine.

From this example it is clear that the inventive method allows for the early diagnosis of enteritis in calves. The results of chemical examination of the feces during the primary survey (the presence of soluble protein, hemoglobin, white blood cells, the pH shifts to the acidic (less than 7.0) was side) indicate the presence of latent inflammatory process in the intestine, manifested clinically in 5 days (second study). The presence of an inflammatory process in the calf primary study indicate the hematologic signs of inflammation (leukocytosis, leukocyte shift to the form�s to the left).

Example No. 4. Calf 9290, age 65 days, with a diagnosis of acute catarrhal enterocolitis (moderately severe). In a clinical study: the fatness satisfactory, decreased appetite, calf lies, sensitivity and reflexes are reduced. Dry skin, decreased turgor - skin fold is straightened within 3 seconds. Wool without Shine, ruffled, the root of the tail and perineum stained feces, on the root of the tail alopecia. The lower part of the abdominal wall and the right iliac region painful on palpation, auscultation listen loud rumbling sounds. Feces thin, frothy, yellow, sour smell, contains a significant amount of mucus and undigested particles of food. The sphincter of the anus moderately relaxed. Body temperature 39,7°C, heart rate and respiratory movements 124 and 40 min, respectively. Hematological signs of inflammation: leukocytosis (12,8×109/l), shift leukocyte formula to the left. Chemical examination of stool the claimed method using test strips for urinalysis (10 EA production Arkrey, Japan): pH 5.0, the reaction to the protein positive (++), hemoglobin - sharply positive (++++), on leukocyte - positive (++). Shift the pH of the feces in the acidic side indicates the presence of fermentation processes in the intestine. The combination of features is the presence in the feces of calves dissolve�imago protein hemoglobin, leukocytes, and a shift of pH in acidic (less than 7.0) was side - indicates the presence of inflammation in the intestine.

When you study after 7 days of treatment the clinical signs of intestinal inflammation are absent. Fatness is satisfactory, the calf is active, the sensitivity and reflexes are good. The mucous membrane of the mouth pale pink, wet, warm. The skin is elastic, good turgor - skin fold is straightened within 1 second. The coat is moderately ruffled. The abdominal wall is painless on palpation, auscultation listens for bowel sounds fluid transfusion. A good appetite. Defecation normal. Kal-decorated, yellow-brown, with a faint sour smell. The tone of the sphincter of the anus good. The body temperature of 38.5°C, heart rate and respiratory movements 80 and 30 min, respectively. In hematological examination - signs of inflammation: leukocytosis (10,4×10%), shift leukocyte formula to the left. Chemical examination of stool the claimed method using test strips for urinalysis (10 EA production Arkrey, Japan): pH 6.5, the reaction to the protein positive (++), hemoglobin - positive (++), leukocytes weakly positive (+). Shift the pH of the feces in the acidic side indicates the presence of fermentation processes in the intestine. With�Etania signs - the presence in the feces of calves soluble protein, hemoglobin, leukocytes, and a shift of pH in acidic (less than 7.0) was side - indicates the presence of inflammation in the intestine.

From this example it is clear that the proposed method allows to detect hidden inflammation in the intestine in calves (in the absence of clinical signs of disease and to monitor the effectiveness of treatment.

Thus, the proposed method is non-invasive rapid diagnosis of the inflammatory process in the intestine of calves for the simultaneous detection in stool soluble protein, hemoglobin, white blood cells and shift the pH to acidic (less than about 7.0) or alkaline (7.5) side using test strips for urine analysis with at least 4 test fields to determine the above parameters based on the method of "dry chemistry" has a high sensitivity, simplicity and reliability, allowing for faster diagnosis of the inflammatory process in the intestine in calves, significantly reduce the cost and time spent on clinical and laboratory studies, can be used for early diagnosis and for monitoring the effectiveness of treatment of inflammatory diseases of the intestine directly at the location of the animal (in the economy).

1. Method of non-invasive rapid diagnosis of the inflammatory process in the intestine in calves, in�Lucasi determination in the stool leukocytes, protein, haemoglobin and pH response of feces, characterized in that the stool sample is diluted in distilled water and put a drop on the corresponding test field test strips, intended for the analysis of urine, by a colour change in less than 1 minute is considered a positive reaction, and at a pH less than 7.0 or more than 7.5 and the presence in Calais at the same time the soluble protein, hemoglobin and leukocytes diagnose the presence of inflammation in the intestine.

2. A method according to claim 1, characterized in that for breeding take the feces and water in the ratio 1:(5÷7).



 

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3 ex, 3 dwg

FIELD: medicine.

SUBSTANCE: CEA, CA 19-9, CA 72-4, M2-PK cancer-specific markers and Ki-67 proliferation index are pre-determined; combined positron emission and computer tomography (PET/CT) are performed. Up to two procedures of neoadjuvant intra-arterial chemoembolisation are conducted with the first one performed by administering a half single systemic dose of irinotecan in lipiodol 4-6 ml; no sooner than 2-3 weeks later, the cancer-specific markers and Ki-67 proliferation index and PET/CT SUVmax are checked; if at least one of this factors tends to increase as compared to references, the intra-arterial chemoembolisation is conducted again by administering at least 1/3 single systemic dose of irinotecan in no more than lipiodol 5 ml. Not later than 10-14 days following the second chemoembolisation or no sooner than 2-3 weeks after the first chemoembolisation, a surgical intervention is required if the tumour keeps its metabolic and proliferative activity on these days.

EFFECT: method enables decreasing the tumour process activity and performing the surgical intervention at the moment of extreme devitalisation of the tumour and micro-disseminates, and thereby improving the remote therapeutic effects.

2 cl, 4 dwg, 3 ex

FIELD: medicine.

SUBSTANCE: invention represents a method for predicting a risk of isolated hysteromyoma involving sampling and studying peripheral venous blood, differing by the fact that DNA is recovered from the peripheral venous blood; genetic polymorphisms of interleukin genes are typed, and combinations of polymorphisms of interleukin-1 (-889 TT IL-1) and interleukin-5 (-703 C IL-5) genes are analysed; a high risk of isolated hysteromyoma is predicted, if observing a combination of -889 TT IL-1 allele and -703 C IL-5 allele in Russian females, native of the Central Black Earth Region.

EFFECT: invention provides obtaining criteria of the risk of isolated hysteromyoma as shown by the combination of the molecular-genetic markers 889 TT IL-1A and -703 C IL-5.

1 ex, 1 tbl, 2 dwg

FIELD: medicine.

SUBSTANCE: method includes the preparation of smear from peripheral blood with preliminary fixation with methyl alcohol, drying, washing with distilled water. After that, the smears are placed in a potassium chloride solution in a ratio of 0.57 g of potassium chloride per 100 ml of distilled water for 20 min and washed with distilled water. Additionally prepared is a mixture of solutions, prepared ex tempore, containing a solution "A" and "B". The solution "A" includes a 50% silver nitrate solution in an amount of 5 g of silver nitrate + 5 ml of distilled water. The solution "B" includes a 2% solution of gelatin on a 1% formic acid solution in an amount of 15.8 ml of distilled water + 0.2 ml of 100% formic acid + 4.0 ml of 10% gelatin. The solutions "A" and "B" are mixed in an amount of 5 ml of each, in darkness, with further submergence of the blood smears for 20 min in darkness in the thermostat at a temperature of 37°C with the further submergence of the smears into distilled water for 2-3 seconds. After that, they are twice subjected to a 8 min exposure in a 5% sodium thiosulphate solution in darkness in the thermostat at a temperature of 37°C. After that, they are washed successively with tap water and distilled water, after-staining is performed in the Romanovskiy dye for 30 min. After that, the smears are washed again with tap water, air-dried, placed in the Canadian balm and covered with a coverslip.

EFFECT: increased quality of smear staining and provision of a possibility to identify and further evaluate parameters of nucleolus organiser regions.

4 tbl, 6 ex

FIELD: medicine.

SUBSTANCE: patients are assessed individually to establish the fact of chronic cardiac insufficiency (CCI) suffered by detecting: age, height, weight, Quetelet index, Brock constant, systolic blood pressure, diastolic blood pressure, triglycerides, total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, low-density lipoprotein cholesterol to total cholesterol ratio, very low-density lipoprotein cholesterol, high-density lipoprotein cholesterol to low-density lipoprotein cholesterol ratio, sum of the very low-density lipoprotein cholesterol and high-density lipoprotein cholesterol to low-density lipoprotein cholesterol ratio, atherogenic index, alanine aminotransferase, aspartate aminotransferase, aspartate aminotransferase to alanine aminotransferase ratio, lactate dehydrogenase, glucose, α-amylase, total protein, albumin, uric acid, carbamide, creatinine, creatinine kinase, alkaline phosphatase, total bilirubin, conjugated bilirubin, ADP-induced thrombocyte aggregation, thrombocyte count, mean thrombocyte volume, erythrocyte count, mean erythrocyte volume, erythrocyte volume distribution, haematocrit, haemoglobin, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, leukocyte count, segmented neutrophil percentage, eosinophil percentage, basophile percentage, lymphocyte percentage, monocyte percentage and displacement constant. The derived discriminator functions are used to establish the fact of the absence of CCI, the first stage of CCI, or the second stage of CCI.

EFFECT: method enables assessing the patients to establish the fact of CCI by determining the significant parameters.

3 dwg, 2 tbl, 1 ex

FIELD: medicine, psychiatry.

SUBSTANCE: one should isolate DNA out of lymphocytes of peripheral venous blood, then due to the method of polymerase chain reaction of DNA synthesis one should amplify the fragments of hSERT locus of serotonin carrier gene and at detecting genotype 12/10 one should predict the risk for the development of hallucino-delirious forms of psychoses of cerebro-atherosclerotic genesis.

EFFECT: more objective prediction of disease development.

3 ex

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