Methods for diagnosing helminthiasis at the present stage. Microscopic examination of stool

Methods helminthological research are divided into direct and indirect. Direct methods: detection of helminths themselves, their fragments, eggs, larvae in feces, urine, duodenal secretions, sputum, nasal and vaginal mucus, contents of subungual spaces, biopsied pieces of tissue. Indirect methods: identification of secondary changes that occur in the human body as a result of the vital activity of the parasite, serological reactions, general research blood, urine. The most common methods for examining feces are helminth-ovoscopic and protozooscopic. When diagnosing, it is impossible to identify eggs or larvae of all types of helminths living in digestive system person. Thus, when using the flotation method, trematode eggs and, in some cases, unfertilized roundworm eggs do not float into the surface film (due to their high specific gravity). In feces it is very rare to find pinworm eggs and taeniid oncospheres, which are detected special methods research: scraping from perianal folds for pinworms and taeniaids, sedimentation methods for trematodes (opisthorchid eggs, etc.). Therefore, for a targeted examination of a patient for helminth infections, the doctor in the referral must indicate which helminths should be focused on (diagnosis), which will allow the laboratory assistant to choose the appropriate technique for identifying this type of helminth. Feces taken from different places stool in an amount of at least 50 grams (teaspoon) in a clean glass container must be sent to the laboratory no later than 24 hours after defecation and examined on the day of receipt. If it is necessary to preserve feces until the next day, it is placed in a cold place (0-4°C) or filled with one of the preservatives. Before examination, the feces are mixed with a stick so that the helminth eggs are evenly distributed in the total mass. If eggs of any helminth are found in the preparation, viewing is not stopped, because there may be double or triple invasion. Monitoring the effectiveness of treatment of helminth infections is carried out by examining feces for helminth eggs 2-3 weeks or 2-3 months after treatment, depending on the detected helminth. Macroscopic methods are used to detect whole mature helminths or their fragments in feces with the naked eye or using a hand magnifying glass. Often, actively crawling pinworms can be seen on the surface of feces after defecation; roundworms are excreted in feces; Sometimes people themselves notice the passage of helminths. In patients with diphyllobothriasis, fragments of tapeworm strobili may be excreted (in the form of “noodles”), and in those infected with taeniids (pork or bovine tapeworm), segments of helminths often leave with feces (in the form of “white clippings”) or they actively crawl out of the anus. The macroscopic method is the main one for the differential diagnosis of taeniasis and taeniarynchosis (in combination with a survey). Of the special macroscopic methods, the method of sequential washing of feces is used. Feces are mixed in water to obtain a uniform suspension, after which, under good lighting, they are carefully examined in separate small portions in black photographic cuvettes or against a dark background in Petri dishes. Using tweezers or a dissecting needle, remove all suspicious white particles, large formations suspicious for fragments of helminths, and examine them under a magnifying glass between two slides. Small helminths or cestode heads are examined under a magnifying glass in a drop of glycerin or under a microscope. When using this method for diagnosing the segments of the pork, bovine tapeworm, and wide tapeworm, the washed segments are placed between two glasses and, looking at the light under a magnifying glass or low magnification microscope, the species is determined by the structure of the uterus (in a mature segment pork tapeworm 8-12 lateral branches extend from the central trunk, and bovine tapeworm 18-32, more often 28-32, in the broad tapeworm the segments are wider and the uterus in the center is in the form of a “rosette”). If the uterus is hard to see, then it can first be kept for some time in a 50% glycerin solution, after which even the empty trunks of the uterus can be clearly seen. When identifying these cestodes by the structure of the detached heads, they are carefully placed with the neck in a drop of glycerin between slides (or covered with a coverslip) and, without squeezing, examined under a microscope at low magnification.

Microscopic methods are divided into simple, complex and special.

Simple ones include the methods of native smear, native smear with Lugol's solution, methods of thick smear under cellophane according to Kato, twisting (according to Shulman) and perianal scraping.

Complex methods are more effective and are based on the concentration of eggs in the preparations. They involve pre-treatment of feces with liquid reagents, as a result of which helminth eggs either precipitate or float to the surface of the liquid.

Complex methods include enrichment methods:

a) flotation (when specific gravity eggs are less than the specific gravity of the saline solution and the eggs float to the surface film);

b) sedimentation (when the specific gravity of eggs is greater than the specific gravity of saline solutions and the eggs settle into sediment).

Special methods for detecting eggs and larvae of helminths, cysts and vegetative forms of protozoa are methods of scraping, flotation, sedimentation, larvoscopy, protozooscopy, bile examination and methods of staining smears of feces, sputum, etc.

Ershova I.B., Osychnyuk L.M., Mochalova A.A., State Institution "Lugansk State Medical University", Department of Pediatrics with Childhood Infections.
The article was published in the journal “Actual Infectology”, No. 2 (3), 2014.
Information resource “Zaslavsky Publishing House” www.mif-ua.com

The article describes methods for diagnosing helminth infestations: microscopic, enzyme immunoassay, serological, polymerase chain reaction, bioresonance diagnostics, hemoscanning, as well as instrumental (ultrasound and X-ray examination, computed tomography) and laboratory, having indirect meaning (clinical analysis blood, blood test for liver function tests, stool test for dysbacteriosis). The advantages of the methods, their disadvantages, and the reliability of the data obtained are described. Questionnaires for patients are proposed to identify the risk of helminth infection. The advantages of the methods, their disadvantages, and the reliability of the data obtained are described. Questionnaires for patients are proposed to identify the risk of helminth infection.

Helminths have a negative effect on the human body. They lead to allergization, the development of polyhypovitaminosis, macro- and microelementosis, impaired hematopoiesis and vascular permeability, hormonal imbalance. Helminth infections contribute to the formation chronic diseases(cholecystitis, cholelithiasis, pancreatitis, colitis, diabetes mellitus, bronchial asthma, atopic dermatitis), psycho-emotional disorders (chronic fatigue, irritability, anxiety, hyperactivity in children), anemia, etc. With prolonged helminth infestation, secondary immunodeficiency may develop.

The alertness of doctors regarding helminth infections among the population is currently insufficient, and prevention is reduced to the treatment of identified infested patients.

Diagnosis of helminthiases is based on clinical, epidemiological and laboratory data. Signs such as asthenic syndrome, recurrent urticaria, impaired regeneration of the skin and mucous membranes, difficult-to-treat atopic dermatitis and broncho-obstructive syndrome, polylymphadenopathy and hepatosplenomegaly of unknown origin, adenoid vegetations of degree II–III, “geographical” tongue, reduced or selective appetite, unstable stool, may indicate the presence helminths.

At serological study determine the presence of antibodies to helminths (reliability - about 60%): if echinococcosis, cysticercosis, trichinosis, toxocariasis is suspected, indirect hemagglutination reactions, latex agglutination, complement fixation, and immunofluorescence are widely used.
Not in all cases, methods for determining specific antibodies have sufficient specificity and reliability. The antigenic composition of the helminth depends not only on the species, but also on the stage; While going through a complex development cycle from egg to adult, helminths change their antigenic composition. In addition, somatic antibodies are used in immunodiagnostic reactions, and in the host’s body antibodies are produced mainly to the excreta and secretions of the helminth. Nonspecific sensitization of the body, the commonality of some antigens of trematodes, protozoa and humans create a high proportion of false-positive reactions in titers below reliably diagnostic ones.

Method for determining helminths using polymerase chain reaction is highly specific and highly sensitive, but due to its high cost and complexity it cannot be used for screening when, for example, you need to examine a group of children from a children's institution.

The immune system does not always react (recognize and destroy) to the presence of helminths in the body. This is because some helminths have a durable and chemically resistant capsule or are coated with a substance that is not recognized by the immune system; localized in tissues that are most protected from inflammatory reactions, for example in the spinal cord; many species of them secrete anti-enzymes in the digestive tract, which saves them from death; have longer duration life (for years, and sometimes until the death of the person himself); feed on glycolysis of net carbohydrates; have devices such as suction cups, hooks, etc., which facilitate fixation inside the body; in many species there is sexual reproduction, in which genetic information is exchanged, which leads to an increase in the heterogeneous population and a decrease in vulnerability; have a high level of fertility.

At ultrasonic , x-ray examination organs abdominal cavity , computed tomography can be identified indirect signs helminth infections: hepatosplenomegaly, unevenness of the liver and spleen parenchyma due to small hyperechoic signals, enlarged lymph nodes in the hilum of the spleen and the helminths themselves (echinococci, balls of intestinal helminths, etc.).

Hemoscanning - qualitative blood examination using a powerful dark-field microscope, you can see the state of blood cells (shape, size, activity, color, etc.), the presence of nonspecific elements and substances - all this directly or indirectly indicates the presence of helminths in the body. The image is displayed on the monitor screen using the built-in video camera in the microscope. This diagnostic method is highly reliable.

Indirect laboratory signs helminthiasis there may be anemia, basophilia, eosinophilia, increased levels of aspartate aminotransferase. Thus, with toxocariasis, a leukemoid reaction of eosinophils (more than 20%) is detected against the background of a persistent allergic syndrome (atopic dermatitis with severe itching and resistance to traditional therapy, severe bronchial asthma). Oppression of normal coli a stool test for dysbacteriosis can also indicate possible helminthiasis.

Taking into account the prevalence of helminthiases, we offer questionnaire to determine the risk of helminth infection.

• Swimming in freshwater bodies of water.
• Do not wash your hands before eating with soap and hot water.
• Drink water from unverified sources.
• You eat homemade lard with streaks of meat.
• Do you eat lightly salted fish?
• You eat medium-cooked meat (with blood).
• Do you eat lightly salted, non-factory prepared caviar?
• Don't wash chicken eggs with soap.
• Do not wash bananas, oranges, tangerines before eating.
• Fertilize your garden with manure.
• Eat vegetables straight from the garden.
• You eat fruits and berries straight from the garden.
• Eat fallen fruits.
• Do not pour boiling water over all your greens to make salads.
• Store carrots in sand taken from the yard.
• Walk barefoot on the grass.
• Family members had helminthic infestations.
• The family has a dog or cat.

For each answer " Yes" - 2 points, " Sometimes" - 1, " No" - 0. With a score of 0–5, the probability of infection is negligible, 6–12 - infection is possible, 13–25 - high probability, more than 25 points - very high. With the last two results, regular examination and, possibly, preventive treatment are necessary.

Since the clinical manifestations of helminth infections are not always specific, and in the initial stages they are nonspecific, we offer a questionnaire for patients to self-diagnosis helminthiasis.

• There is itching in the anus in the morning.
• Nausea in the morning when brushing teeth.
• Peeling of fingers or toes with peeling of layers of skin.
• Allergic skin rash, itchy skin.
• Peeling and swelling in the eyelid area.
• Increased fatigue, lethargy, drowsiness.
• Increased feeling of hunger.
• Feeling of discomfort in the stomach.
• Loss of body weight.
• The presence of several chronic diseases of the gastrointestinal tract, joints, and bronchopulmonary system.
• Poor health, lack of official diagnosis, long-term ineffective treatment.
• Periodic rise in temperature, accompanied by muscle and joint pain.

Answer " Yes» speaks about at least 2–3 questions high probability helminth infections.

CONCLUSIONS

1. On modern stage No laboratory methods tests for helminthiases, which are 100% reliable.

2. The polymerase chain reaction and bioresonance diagnostics have the greatest reliability in diagnosing helminthiases.

BIBLIOGRAPHY

Introduction

Chapter I. Classification of helminth infections

Chapter II. Pathogenesis of helminthiases

1 The main phases of helminthiases in pathogenesis and clinic

2 Factor of influence of the pathogen on the host’s immune system

Chapter III. Basic principles for diagnosing helminthiasis

Chapter IV. Macroscopic research methods

Chapter V. Qualitative Research Methods

1 Native smear

2 Kato method

3 Methods based on the principle of egg deposition

4 Enrichment methods based on the principle of floating eggs

5 Method for detecting helminth eggs in feces using the enrichment method

6 Flotation methods

7 Tape method

Chapter VI. Quantitative Research Methods

1 Stoll method

2 Krasilnikov-Volkova method

ChapterLAVA VII. Methods for diagnosing protozoal infestations

1 Native smear

Conclusion

Bibliography

INTRODUCTION

Small business development in Food Industry with insufficient technological control led to a decrease in the quality and safety of food products. Illegal livestock slaughter points and clandestine production of meat products that have not passed veterinary control have appeared. The deterioration of the situation is also facilitated by the issuance of veterinary control certificates for food products on a “commercial basis”, a decrease in the quality of medical examinations, and the hiring of persons without health certificates.

Every year, the number of domestic animals increases worldwide. According to the Russian Canine Federation, about 5 million. purebred dogs are registered in Russia. In general, there are supposedly about 30 million dogs in our country, and many of them are stray. According to various studies, up to 80% of domestic dogs are infected with worms. Pollution problem environment the feces of these animals is becoming increasingly acute. Surveys carried out in various countries have established significant soil contamination in populated areas helminth eggs with fluctuations of up to 60% of positive samples. The places most contaminated with helminth eggs are around garbage containers, courtyards, sandboxes of kindergartens, markets, city veterinary hospitals, and basements.

Today, due to the current problem, the diagnosis of helminthiasis is of great importance, which is especially important in the early stages of the disease.

The purpose of our work is a theoretical study of methods for modern diagnosis of the most common helminthiases used today in clinical practice.

Job objectives:

identifying the most effective methods research on helminth infections in clinical practice;

Familiarity with WHO statistics;

identifying the advantages and disadvantages of various methods;

identifying the most effective methods;

studying the history of helminthiasis development

study of current and promising methods for identifying helminthiasis.

CHAPTER I. CLASSIFICATION OF HELMINTOSISES

The second group includes helminths (Trichinella, trematodes), infection of which can occur through animal meat and fish.

When feeding, helminths secrete toxic substances in the host's body, which are instantly absorbed into the blood, distributed throughout the host's tissues, affecting his nervous system and all vital functions. important organs. .

Helminthiases are characterized by the development of eggs and larvae of pathogens only in external environment without the participation of intermediate hosts. The development of geohelminth eggs occurs in the soil or on vegetables and is determined by factors such as temperature, humidity and soil aeration.

Tenidosis is a disease associated with the consumption of raw meat. In these diseases, humans are the definitive host of helminths and the only source of infection. A person becomes infected by eating meat infected with the larval stage of the biohelminth. There are two known varieties of tapeworm: bovine tapeworm and pork tapeworm. When eating meat contaminated with bovine tapeworm larvae, a person develops a disease called taeniarinchiasis. When eating meat contaminated with pork tapeworm larvae, taeniasis develops.

Trichinosis is a biohelminthiasis characterized by fever, muscle pain and allergic manifestations. Human infection occurs through consumption of meat containing encapsulated Trichinella larvae.

Diphyllobothriasis is a biohelminthiasis characterized by damage to the gastrointestinal tract and has a chronic course. Human infection occurs when eating insufficiently heat-treated or lightly salted fish and caviar containing tapeworm larvae.

Opisthorchiasis is a biohelminthiasis characterized by damage to the liver and pancreas and has a chronic course. Human infection occurs through consumption of lightly salted, lightly dried, raw or insufficiently heat-treated fish containing cat fluke larvae.

Geohelminthiases are helminthiases whose pathogens develop without the participation of an intermediate host. The eggs or larvae of geohelminths released from the body develop to invasive stage in the soil. Representatives of living nature (biotic environment) here can only play the role of mechanical carriers of invasive larvae. For example, flies may accidentally carry eggs or larvae on their proboscis or stalk. Geohelminthiases include: ascariasis, trichuriasis, hookworm disease, strongyloidiasis and other diseases. oh, dogs - on their limbs, hair.

CHAPTER II. PATHOGENESIS OF HELMINTOSISES

According to WHO experts, helminthiases have now, to some extent, become “forgotten diseases” - there is an underestimation of their medical and social significance throughout the world. Even in endemic countries, they receive insufficient attention from both health authorities and the population.

2.1 The main phases of helminthiases in pathogenesis and clinic

In the pathogenesis and clinical picture of helminth infections, two main phases are distinguished: acute - the first 2-3 weeks after invasion, and in severe cases - up to 2 months or more, and chronic - lasting from several months to many years.

.2 Factor of influence of the pathogen on the host’s immune system

The influence of the pathogen on the host’s immune system continues to play a significant role in the chronic phase of invasion. One of the important causes of organ and systemic lesions, especially with tissue helminthiases, is the formation immune complexes, which activate mediator systems (complement, cytokines, etc.). Along with stimulating the immune response, helminths have an immunosuppressive effect, which promotes their survival in the host body. The state of immunodeficiency due to helminth infections negatively affects a person’s resistance to bacterial, viral and other infections, contributes to their protracted course and the formation of carriage, and reduces the effectiveness of preventive vaccinations. This is well shown in the frequency of typhoid carriage, the incidence of tuberculosis and other chronic diseases. infectious diseases among the population of hyperendemic foci of opisthorchiasis.

In clinically manifest forms of helminthiases, the first signs appear in different terms after infection: with ascariasis, manifestations of the acute phase are observed already on the 2-3rd day, with most other helminthiasis - after 2-3 weeks, with filariasis incubation period lasts 6-18 months. In the early acute phase of helminthiases, manifestations are characteristic: allergic reactions: fever, recurrent itchy skin rashes, swelling - from local to generalized, swollen lymph nodes, myalgia, arthralgia, in peripheral blood - leukocytosis with hypereosinophilia. Against this background, pulmonary syndrome (from minor catarrhal phenomena to asthmatic conditions, pneumonia and pleurisy) and abdominal syndrome (abdominal pain and dyspeptic disorders) often develop. The liver and spleen increase in size, symptoms and syndromes of damage to the central nervous system(CNS). With some helminthiases, specific symptoms are also observed: with trichinosis, in typical cases, from the first days of the disease, a symptom complex is observed, including fever, muscle pain, swelling of the eyelids and face; with liver trematodes (opisthorchiasis, fascioliasis) - icteric syndrome, enlarged liver and spleen. Even among helminthiases caused by similar types of pathogens, there are significant differences in the severity of the course and the nature of the manifestations of the acute period: for example, with Japanese schistosomiasis, it develops much more often and is more severe than with genitourinary and intestinal schistosomiasis.

Greater polymorphism clinical manifestations Characterized by strongyloidiasis, in which, along with a variety of allergic and dyspeptic symptoms, patients often exhibit signs of dysfunction of the biliary tract. With liver trematodes (opisthorchiasis, clonorchiasis, fascioliasis), chronic cholecystocholangitis, hepatitis, pancreatitis develop, damage to various parts of the gastrointestinal tract is possible, and neurological disorders are also observed. A characteristic feature Urogenital schistosomiasis is “terminal hematuria” (the appearance of a drop of blood at the end of urination) and dysuric disorders. In patients with filariasis, the allergic syndrome is expressed to one degree or another; lymphatic filariasis (wuchereriosis and brugiosis) is characterized by lymphadenopathy, lymphangitis and lymphostasis; with onchocerciasis, along with these symptoms, serious eye damage is noted.

Intestinal cestodiasis (diphyllobothriasis, teniarhynchosis, taeniasis, hymenolepiasis) in many cases are asymptomatic, manifesting only by the passage of mature helminth segments during defecation or independently (only with teniarhynchosis). Patients with diphyllobothriasis develop anemia caused by vitamin B12 deficiency. Among helminthiases, a special place is occupied by larval cestodiases: echinococcosis, alveococcosis, cysticercosis. They can also be asymptomatic for a long time, even in the presence of fairly large cysts. At the same time, rupture or suppuration of even a small echinococcal bladder leads to serious consequences: the development anaphylactic shock, purulent peritonitis, pleurisy, etc. As a result of compression of the portal and inferior vena cava by a growing bladder or alveococcus, portal hypertension with all the characteristic manifestations and consequences.

Cysticercosis of the central nervous system occurs in the form of cerebral and spinal lesions with corresponding varied symptoms; Localization of the helminth in the ventricles of the brain is accompanied by signs intracranial hypertension. Toxocariasis, registered in our country mainly in children, is clinically expressed by abdominal and pulmonary syndromes, neurological disorders, eye damage, and severe eosinophilia in the peripheral blood. .

IN last years Great progress has been made in studying the mechanisms of development of the pathological process during helminthiasis.

CHAPTER III. BASIC PRINCIPLES OF DIAGNOSIS OF HELMINTISES

According to WHO, the accuracy of diagnoses based on stool tests does not exceed 5-10%.

Main method laboratory diagnostics of these infestations is the detection of eggs or larvae of helminths.

Stools for analysis must be delivered to the laboratory no later than one day after their isolation, and if strongyloidiasis is suspected, immediately

Currently, low-intensity invasions predominate, especially with geohelminthiasis, therefore, during coproovoscopy it is necessary to use enrichment methods.

However, even before the laboratory examination, diagnostics should be based on mandatory information obtained from studying the patient’s epidemiological, geographic, social and disease history.

Despite, at first glance, it would seem asymptomatic Many helminthic-protozoal infestations; in most of those infested, a thorough medical examination can reveal important symptoms.

Therefore, the results of a patient interview, the most complete anamnestic data, the presence of even mild symptoms in most cases determine the possibility of making a diagnosis with such invasions as enterobiasis, taeniasis, diphyllobothriasis, and often with ascariasis and opisthorchiasis with a 79-80% probability. And most importantly, these data determine specific indications for laboratory diagnostics: the correct choice of technique, appropriate preparation of the patient, collection of the necessary biological material (liquids or excreta) for examination.

Immunological diagnostic methods, as well as PCR, to detect the DNA of pathogens, are very popular among the main research methods today. Immunological methods for diagnosing helminth infections are based on the detection of specific antibodies to certain helminths in the blood serum. For immunological research, the method of indirect hemagglutination is used, enzyme immunoassay, immunoelectrophoresis, immunoabsorption and others serological methods blood tests.

Immunological research methods are used to diagnose alveococcosis, echinococcosis, cysticercosis, ascariasis, schistosomiasis and other helminthiases as auxiliary ones in combination with clinical and instrumental diagnostic methods. These research methods can be considered effective only when the helminths are located directly in the tissues human body.

Biological material for research on the presence of helminths, their fragments, larvae and eggs are feces, urine, duodenal contents, bile, sputum, rectal and perianal mucus, blood, muscle. Given the predominant localization of most of the most common helminths in the gastrointestinal tract, feces are most often the object of research. Macroscopic methods are used to detect isolated helminths or their fragments: heads, strobila fragments or individual segments. The purpose of microscopic examination is to detect eggs and larvae. Currently, thick smears according to Kato-Miura, sedimentation methods, and flotation methods are recommended for use.

Native and concentrated preparations of feces: eggs of roundworm helminths, whipworm, dwarf tapeworm, broad tapeworm, toxocara, cat trematode, Siberian fluke, etc.

In native unprocessed feces or liquid feces when washed with the naked eye, one can detect whole or fragments of adult roundworms, pinworms, segments of the bovine tapeworm, pork tapeworm, and fragments of the strobila of the broad tapeworm.

In urine samples: schistosome eggs, trichomonas, diactophyme eggs, daughter capsules of echinococcus, with echinococcosis of the kidneys.

In skin scrapings from the perianal area: pinworm eggs, tennid eggs (oncospheres).

In sputum: larvae of strongylides, ascaris, toxocar, daughter capsules of echinococcus in pulmonary echinococcosis, amoebas in pulmonary amebiasis.

In samples of duodenal contents (duodenum): larvae and even eggs of strongylids, trophozoids of Giardia, eggs of trematodes (feline, Chinese, liver, lanceolate bifolia).

In cerebrospinal fluid samples: trypanosomes, larvae of angiostrongylid nematodes.

In smears from vaginal mucus: trichomanas, trophozoites of intestinal amoeba, sometimes eggs of pinworms and even an adult.

In mother's milk in special cases: strongylid larvae.

Native tissue preparations (aspiration material from the bronchi, bone marrow, lymph nodes, abscesses, scrapings from mucous membranes, biopsy) by processing with special methods is used in addition to or instead of histological studies for the diagnosis of trypanosomiasis, pneumocystis, trichinosis.

Feces for fecal analysis to detect helminths must be collected correctly, following the recommendations:

Before collection, it is forbidden to do an enema or use any laxatives;

When defecating, feces should be collected on plastic wrap or in a tray. In this case, the sample must not be allowed to come into contact with urine, secretions, water, personal hygiene items, etc.;

If it is not possible to immediately send the tests to the laboratory, then the material must be stored at a temperature of 4-8 degrees Celsius. At the same time, the tests must reach the laboratory on the same day.

If possible, several stool samples should be collected from different bowel movements during the same day.

CHAPTER IV. MACROSCOPIC METHODS OF RESEARCH

Inspect feces with a magnifying glass or with the naked eye.

Progress of the study. Feces are mixed with water until a uniform suspension is obtained, after which, under lighting, they are carefully examined in small proportions in black photographic cuvettes or against a dark background in Petri dishes. If suspicious particles are detected or when the presence of small forms of helminths (for example pinworms) is obviously suspected, all suspicious white particles are transferred to a glass slide with tweezers or dissecting needles in a drop of glycerol, isotonic solution sodium chloride or water and examined with the naked eye under a magnifying glass or microscope. If suspicious formations are detected, you should examine them under a magnifying glass, after squeezing them between two glass slides.

Settling method. Progress of the study. All studied material (in in this case fresh feces) are placed in tall jars, diluted with a strong stream of water and left to settle. The cloudy layer above the sediment is carefully poured into another vessel, the sediment is diluted again and the mixture is allowed to settle. These operations are performed until the water above the sediment becomes clear. The water is drained, and the sediment is examined in small parts in Petri dishes against a dark background.

Evaluation of the results obtained. Differential diagnosis between helminths is based on their anatomical and morphological characteristics. Determining the identity of nematodes, as a rule, is possible only from whole individuals, less often - from sufficiently large fragments that have retained their characteristic diagnostic features. Cestodes are most often diagnosed by mature or hermaphroditic scolex and segments.

Microscopic research methods

The purpose of microscopic methods is to identify helminth eggs and larvae. These methods are divided into qualitative and quantitative.

CHAPTER V. QUALITATIVE RESEARCH METHODS

.1 Native smear

Method proposed by Daven in 1853. - one of the oldest and most simple methods this group, but there are numerous statements in the literature indicating its low effectiveness compared to other methods.

METHODS FOR HELMINTHS DETECTION.

Ascariasis can be detected using the following methods: coprooscopic smear on Kato, unified enrichment methods of Fulleborn, Kalantaryan and other flotation techniques.

5.2 Kato method

The Kato method (thick smear with cellophane) is a successor to the native smear. This method very convenient for mass examinations; smears can be prepared on the spot. This method is based on identifying helminths in a smear of feces cleared with glycerin and stained with malachite green. To implement this method, you need a Kato cellophane mixture consisting of 6 milliliters of a 3% solution of malachite green in water, 500 milliliters of glycerin, 500 milliliters of a 6% solution of phenol.

Progress of the study. Cellophane strips smaller than a glass slide are cut out of hydrophilic cellophate and pre-treated by immersing them in Kato's solution: glycerin softens the cellophane, lightens the preparation and protects it from drying out; phenol is used to protect laboratory technicians from pathogenic bacteria; malachite green reduces eye strain. The strips are placed in Kato solution so that they are adjacent to each other (3-5 ml per 100 strips). After 24 hours, the strips are ready for further action. They can be stored for a very long time in the specified mixture in an airtight container.

A milligram of feces is placed on a glass slide in a thick layer, covered with a strip of cellophane and pressed onto the slide with a rubber stopper so that the feces are not squeezed out from under the cellophane.

The smear examination should be carried out no later than an hour after its preparation. In a short period of time (within an hour) at room temperature, the smear becomes more transparent.

If there are no reagents for preparing the Kato mixture, it can be replaced with a 50% aqueous solution of glycerin.

Detection of eggs is carried out under a microscope in the entire thick smear.

Diagnostic value. The Kato method is the most effective for diagnosing teniarynchosis. You can also identify eggs of roundworms, whipworms, trematodes and others. But this method is not effective in diagnosing hymenolepiasis, hookworm disease and trichostrongyloidiasis.

.3 Methods based on the principle of egg deposition

These methods are currently not recommended as a unified method due to their low efficiency or cumbersomeness. From this group of methods stands out the method of examining feces for helminth eggs using synthetic detergents (synthetic detergents"Lotus" type). Under the influence of surfactants included in detergents, helminth eggs are released from feces and concentrated in the sediment.

A 1% solution of washing powder is used as a reagent (the powder is dried in an oven at 100 °C for 1-2 hours, 10 g of powder is dissolved in 1 liter of tap water).

There are two methods:

method - 20-30 ml of detergent solution is poured into a bottle with a capacity of 30-50 ml, and a portion of feces the size of a hazelnut is placed there. It is advisable to place feces for examination in a detergent solution no later than 1 hour after defecation. The ratio of solution to feces is approximately 1:2. Feces should be in the solution for at least a day. During this time, a two- or three-layer sediment forms at the bottom of the bottle. The bottom layer consists of coarse heavy particles; helminth eggs are concentrated in the middle layer, on which light flakes sometimes settle. A Pasteur pipette with a high end is inserted into the middle layer of the sediment, possibly lower, but without touching the bottom of the flask, one to three drops of liquid are collected and transferred to a glass slide. The drop is covered with a cover glass or cellophane plate according to Kato and examined under a microscope. Two preparations should be prepared on one glass.

method - Feces are placed in a detergent solution in a ratio of approximately 1: 10 and mixed until a suspension is formed. After 30 minutes, the contents of the tube are shaken for 1-2 minutes and centrifuged for 5 minutes at 1000-1500 rpm. Two preparations are prepared from the sediment on one slide.

Both preparations are examined completely under a microscope, taking into account all detected helminth eggs.

Diagnostic value: This method can detect eggs of all types of helminths.

Fulleborn method. This method is based on the floating of helminth eggs in a saturated NaCl solution with a high relative density. To do this, dissolve 400 g of NaCl in 1 liter of water while boiling. The relative density of the solution is 1.18-1.22. The solution is stored in a closed bottle.

To carry out the analysis, 2-3 g of feces are placed in a jar with a volume of 30-50 ml and, while stirring with a stick, a saturated solution of sodium chloride is added almost to the top. A strip of paper is used to quickly remove large floating particles.

After 45-60 minutes. After settling with a wire loop, remove the surface film and transfer it to a glass slide in a drop of 50% aqueous glycerol solution. Several preparations are being prepared. Additionally, 2-4 preparations from the sediment are examined, collecting it with an eye pipette onto 2 glass slides.

The need to study the sediment is due to the fact that the eggs of trematodes and taeniids float very poorly and can remain in the sediment. Eggs of nematodes (with the exception of unfertilized roundworm eggs), dwarf tapeworms and tapeworms float well.

The advantages of this method include its low cost and accessibility, but the disadvantages are the need to view preparations from the surface film and sediment, as well as the duration of settling.

.4 Enrichment methods based on the egg floating principle

The methods of this group are based on the floating of helminth eggs in a solution that has a high specific gravity in comparison with them.

Kalantaryan method . Feces are suspended in a flotation solution, which has a higher relative density than helminth eggs. In this case, helminth eggs float to the surface, and the resulting film is examined under a microscope.

As a reagent, use a flotation solution according to Kalantaryan (1 kg of sodium nitrate is dissolved in 1 liter of water, boil the mixture until a film forms and pour without filtering into dry bottles; relative density of the solution is 1.38) or a flotation solution according to Brudastov - Krasnonos (900 g of nitrate sodium and 400 g of potassium nitrate are dissolved when heated in 1 liter of water; relative density of the solution is 1.47-1.48).

The disadvantage of this method is the high cost of sodium nitrate.

Goryachev method . This method (precipitation method) is used to diagnose opisthorchiasis. The specific gravity of opisthorch eggs is high, so they do not float in saline solutions. 70-100 ml of saturated sodium chloride solution is poured into a cylinder with a diameter of 2-3 cm.

Separately, carefully stir 0.5 g of feces in 20-25 ml of water and carefully filter through a funnel with two layers of gauze into a cylinder on saline solution, avoiding mixing. Opisthorchid eggs slowly settle to the bottom of the cylinder.

In 2-3 hours upper layer the feces are sucked out with a pipette, and the remaining saline solution is left to stand for 12-20 hours or centrifuged. The sediment is pipetted onto a glass slide, covered with a coverslip and examined under a microscope. This method is also applicable for the diagnosis of other trematodes.

.5 Method for detecting helminth eggs in feces using the enrichment method

In beakers, thoroughly stir 5-10 g of feces and 100-200 ml of one of the flotation solutions with a glass rod. Immediately after the end of stirring, remove large particles that float to the surface with a glass rod. A glass slide is placed on the surface of the saline solution. If there is empty space between the mixture and the slide, add saline until the mixture comes into complete contact with the slide.

Leave to settle for 20-30 minutes, after which the slide is removed, placed under the microscope with the film facing up, and all the film adhering to the surface of the slide is examined without a cover glass. To avoid drying out during the study, the film can be mixed with two to three drops of a 50% glycerin solution.

All helminth eggs found in the preparation are taken into account.

Diagnostic value: the described method can detect infection with roundworms, whipworms, hookworms, taeniids, trematodes, tapeworms and other types of helminths.

5.6 Flotation methods

Currently, for the diagnosis of opisthorchiasis (clonorchiasis), the methods of Kato and Kalantaryan are recommended as quite effective and technically simpler.

5.7 Tape method

The sticky tape method is used to diagnose enterobiasis. A piece of adhesive transparent polyethylene tape 4-5 cm long is applied in a sticky layer through the anus to the perianal folds, immediately removed and glued to a glass slide. The preparations obtained in this way are examined microscopically.

The advantages of this method over scraping from the perianal folds are the speed and possibility of fairly long storage drugs.

CHAPTER VI. QUANTITATIVE RESEARCH METHODS

Quantitative research methods are used to determine the intensity of invasion, assess the effectiveness of various anthelmintic drugs, determine the quality of deworming, monitor ongoing mass treatment and preventive measures, etc.

Quantitative determination of helminth eggs is studied by two methods: the Stoll method and the method of Krasilnikov and Volkova (1974).

.1 Stoll method

The table method is used for ascariasis. Necessary equipment: microscope, glass flask with 56 and 60 milliliter markings, graduated cylinder, glass beads, rubber flask stopper, graduated pipettes, glass slides and 0.4% sodium hydroxide solution.

Progress of the study. Decinormal sodium hydroxide solution is poured into a flask with a graduated cylinder to the 56 milliliter mark and feces are added until the liquid level rises to the 60 milliliter mark (4 milliliters of feces is obtained). This mixture is shaken with glass beads for one minute, after closing the flask with a rubber stopper (you can also stir with a stick). Immediately after shaking, take 0.075 milliliters of the mixture with a graduated pipette (it contains 0.005 milliliters of feces), transfer it to a glass slide and count the number of eggs in the preparation under a microscope. In order to determine the number of eggs in 1 gram of feces, the detected number is multiplied by 200.

Comparison of the number of eggs in a sample found in a patient before and after treatment allows us to talk about the effectiveness of deworming.

This method is quite simple and gives comparable results for all helminthiases, the pathogens of which systematically release eggs into the patient’s intestines. However, the method has a significant drawback - relatively low sensitivity, especially with low intensity of invasion.

.2 Krasilnikov-Volkova method

When studying this method, take at least 1 gram of feces and mix it in a glass flask or large test tube with a 1% Lotus solution (you can take a 1.5% Ekastra solution) in a ratio of 1: 10. The suspension is thoroughly shaken until a homogeneous suspension is formed. , immediately after this, take 0.1 milliliter of suspension with a graduated pipette (approximately equal to 0.01 grams of feces) and transfer it to a glass slide. This preparation is covered with a cover glass or cellophane plate (20 x 30 mm), kept for at least one day at 50% aqueous solution glycerin.

The number of eggs in the entire preparation is counted under a microscope. To calculate the number of eggs in one gram of feces, the resulting number is multiplied by 100.

This method is more effective than the Stoll method. Firstly, it is more sensitive and allows you to detect helminths when weak degree infestations. Secondly, it is very convenient for mass examinations, since detergent solutions, which are preservatives for helminth eggs, make it possible to conduct research on not entirely fresh material. However prerequisite This involves collecting feces directly into a detergent solution.

For quantitative research, you can use any of the described unified qualitative methods based on the principle of floating eggs. But in this case, the same amount of feces and the same volume of flotation solution must be taken for analysis. The degree of infestation can be calculated by knowing the number of eggs in 1 gram of feces.

CHAPTER VII. METHODS FOR DIAGNOSTICS OF PROTOZOAL INVASIONS

.1 Native smear

Apply 1-2 drops of isotonic solution to a glass slide in two places with a short gap.

Using a wooden stick, select a small piece from the feces sample and transfer it into a drop on a glass slide, stirring until a homogeneous mass is formed. Next, a cover slip is placed on top and examined under a microscope.

Through a well-prepared preparation, printed text should be visible.

When examining stool in patients, luminal forms are detected - cysts. In the acute period of the disease, large vegetative forms of amoebas are found - erythrophages (hematophages), which quickly die in the external environment. The data is given below:

Type of infestationNumber of eggs in 1g of fecesNumber of helminths in the intestinesDegree of infestationAscariasis1-10001-10Weak10001-5000011-50Moderate50001-1900051-200SevereOver 190000Over 200Very severeTrichocephalosis1-20001 -25Very weak2001-750026-100Weak7501-37500101-500Moderate37501-75000501-1000SevereOver 75000Over 1000Very severeHookworm1-25001-25Very weak2501-1000026-100С labaya10000 -50000101-500Moderate50001-100000501-1000SevereOver 100000Over 1000Very SevereNecatorosis1-6001-25Very Weak601-210026-100Weak2101-11000101-500Moderate11101-22 100501-1000HeavyOver 22100Over 1000Very heavy

During a serological study, the presence of antibodies to helminths is determined (reliability is about 60%): if echinococcosis, cysticercosis, trichinosis, toxocariasis is suspected, indirect hemagglutination reactions, latex agglutination, complement fixation, and immunofluorescence are widely used.

Not in all cases, methods for determining specific antibodies have sufficient specificity and reliability. The antigenic composition of the helminth depends not only on the species, but also on the stage; While going through a complex development cycle from egg to adult, helminths change their antigenic composition. In addition, somatic antibodies are used in immunodiagnostic reactions, and in the host’s body antibodies are produced mainly to the excreta and secretions of the helminth. Nonspecific sensitization of the body, the commonality of some antigens of trematodes, protozoa and humans create a high proportion of false-positive reactions in titers below the reliably diagnostic ones.

The method of determining helminths using polymerase chain reaction is highly specific and highly sensitive, but due to its high cost and complexity it cannot be used for screening when, for example, it is necessary to examine a group of children from a children's institution.

The immune system does not always react (recognize and destroy) but the presence of helminths in the body. This is because some helminths have a durable and chemically resistant capsule, or are coated with a substance that is not recognized by the immune system; localized in tissues that are most protected from inflammatory reactions, for example in the spinal cord; many species of them secrete anti-enzymes in the digestive tract, which saves them from death; have a long life expectancy (years, and sometimes until the death of the person); feed on glycolysis of net carbohydrates; have devices such as suction cups, hooks, etc., which facilitate fixation inside the body; in many species there is sexual reproduction, in which genetic information is exchanged, which leads to an increase in the heterogeneous population and a decrease in vulnerability; have a high level of fertility. These research methods can be considered effective only when the helminths are located directly in the tissues of the human body. In this case, apply: skin and intradermal test, ring precipitation reaction, indirect hemagglutination and so on.

In the blood of infected individuals, the titers of specific antibodies increase, first of the IgM and then of the IgG classes. If this is not reinfestation, then in the body of a patient with allergic manifestations there are still no helminths in the reproductive stage of development, and diagnosis of helminthiasis is impossible. The period of helminth maturation can be quite long and depends on many factors. !!! At this stage, diagnosis of the disease is possible only by identifying antibodies to helminth antigens. The use of a highly sensitive enzyme immunoassay method for this purpose has proven to be very effective.

Immunological methods are used to detect schistosomiasis, alveococcosis, cysticercosis, echinococcosis, ascariasis and other types of helminths.

Accompanying examinations can also confirm helminthiases. Helminthiases are often revealed in tests for dysbacteriosis and a general blood test (low hemoglobin, increased ESR, eosinophilia).

CONCLUSION

Currently, there is no simple, accessible and reliable method for diagnosing helminthiases. The choice of one method or another depends on the type of helminth, as well as the stage of the disease.

In conclusion, I would like to remind you that the diagnosis of any

Important Today, due to the current situation, prevention should be given priority.

Prevention of helminthiasis includes a set of measures to identify patients, treat them, ensure living conditions, everyday life and production that exclude the spread of these diseases, protect and improve the environment from pathogens. The volume and nature of measures taken to reduce the incidence of the most common geohelminthiases among the population of the Russian Federation are determined by the level of incidence, climatic conditions, characteristics of life and economic activity population and the results of sanitary and helminthological monitoring, since geohelminthic infections are primarily a sanitary problem. The basis for the prevention of trichinosis, teniarhynchosis, taeniosis is to ensure the safety of meat products for human health, and the prevention of opisthorchiasis, diphyllobothriasis, and other helminthiasis transmitted through fish, crustaceans, mollusks and reptiles is to ensure the guaranteed safety of fish and other relevant products. Prevention and control of echinococcosis and alveococcosis is carried out using measures aimed at preventing infection of humans, farm animals, and dogs; health education, regular medical examination risk contingents (reindeer herders, fur breeders, hunters). In the prevention of helminthiases transmitted by contact (enterobiasis), measures aimed at breaking the mechanism of transmission of their pathogens are of primary importance, and it should be taken into account that these helminthiases predominantly affect children in organized groups.

It is necessary to observe personal preventive measures - wash your hands before eating, after visiting the toilet, returning home from the street, after contact with animals. Berries, vegetables, fruits, herbs should be thoroughly washed with running water and rinsed boiled water. You should not try raw minced meat or fish. Fish, seafood, and meat should be well fried, stewed or boiled. It is necessary to remember about your pets - do not feed them raw fish, meat, internal organs animals, conduct them periodically preventive treatment(deworming).

BIBLIOGRAPHY

1. M. V. Severin, D. N. Ponamarev, V. M. Borzunov, T. B. Tretyakova. Methods for diagnosing the most common protozooses and helminthiasis Ekaterinburg 1996. -71s.

A. M. Bronshtein, N. A. Malyshev. “Helminth infections of humans” Moscow 2010 -109 p.

Methodical manual Helminth infections in pediatric practice Moscow 2008. -30s.

Http://doctorspb. ru/ medical portal for doctors and students.

Biology, edited by Academician of the Russian Academy of Medical Sciences, Professor V. N. Yarygin: Volume 2. Moscow publishing group "GEOTAR-Media", 2012 - 553 pp.

Slyusarev A. A., Zhukova S. V. - K.: Vishchashk. Head publishing house, 1987-415p.

Http://www. Pasteur-nii. spb. ru/ Helminthology

12. http: //www. rusnauka. com/17_APSN_2013/Biologia/10_140855. doc. htm

clinical and pathogenetic features and the current state of diagnosis, treatment of human helminth infections

Http://fersirs. ucoz. ru/news/klassifikacija_gelmintov_klassifikacija_gelmintozov_po_voz/2013-12-19- Classification of helminths.

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Protozoa are divided into 4 classes:

When encysted, the microorganism acquires a round shape and is covered with a protective shell. In the form of a cyst, the protozoa becomes less susceptible to unfavorable environmental factors.

The following may be subject to research:

Note:There are many types of diagnostics; we will consider those types that are most common in clinical laboratory practice.

Private types of diagnostics

In each specific case, the laboratory assistant is given the task of finding specific pathogen, sometimes others are discovered along with the main one.

There are 6 species of this microorganism capable of living in the human intestine. Clinical significance Only the dysenteric amoeba, which occurs in vegetative form and in the form of cysts, has.

Additionally, immunological methods are used:

• indirect immunofluorescence;
• indirect agglutination (INA);
• radial immunodiffusion.

Note: serological methods are not very informative and are used only as an addition to the main ones in doubtful cases.

Diagnostics of ciliated (ciliates)

The pathogenic form of microorganisms of this genus is balantidium. This is a microbe that causes balantidiasis, a disease accompanied by an ulcerative process of the large intestine. The pathogen is detected in the native smear in the form of a vegetative form and a cyst. The material for the smear (feces and mucus) is taken during a sigmoidoscopy examination and sown on special media.

Diagnosis of flagellates (Leishmania, Giardia, Trypanosomes, Trichomonas)

Leishmania, trypanosomes, lamblia, and trichomonas are dangerous for humans.

Leishmania– microbes that cause leishmaniasis are examined in blood smears, bone marrow materials, and scrapings from skin infiltrates. In some cases, when diagnosing leishmania, culture on nutrient media is used.

Trypanosomes– pathogens sleeping sickness(American/African trypanosomiasis, or Chagas disease).

The African variant is determined in the initial period during the study of peripheral blood. As the disease progresses, pathological microbes are found in the material of lymph node punctures, and in advanced stages - in the cerebrospinal fluid.

To diagnose trypanosomes if Chagas disease is suspected, the material being examined is examined under a microscope at low magnification. In this case, the smears and thick drop are pre-stained.

Trichomonas(intestinal, oral,) are detected by microscopy of materials taken from the affected mucous membranes.

Identification of sporozoans (malarial plasmodium, causative agent of coccidosis, etc.)

The most common and dangerous species for humans is the malarial plasmodium, which has 4 main types of pathogen: the causative agent of three-day malaria, four-day malaria, tropical malaria and malaria ovale.

Sexual development of Plasmodium (sporogony) takes place in Anopheles mosquitoes. Asexual (tissue and erythrocyte schizogony) - in human liver tissue and erythrocytes. These features life cycle must be taken into account when diagnosing malarial plasmodium.

Thus, in the blood of a newly ill patient, germ cells of the sporogony cycle can be detected. But at the height of malarial attacks, schizonts appear in large numbers in the blood.

Moreover, in different phases of malarial fever, various shapes plasmodium:

• during the chill period, the blood fills with merozoites, a type of schizont;
• at elevated temperatures, ring-shaped trophozoites accumulate in erythrocytes;
• a decrease in temperature is characterized by a predominance of amoeba-like trophozoites;
• during periods of normal condition, the blood contains adult forms of schizonts.

The study of the causative agent of malaria (malarial plasmodium) is carried out in a smear and in a thick drop.

Note:Diagnosis of malaria by examination of smears and thick drops of blood is sometimes erroneous. Blood platelets in some cases may be mistakenly attributed to the malarial pathogen. Also sometimes fragments of leukocytes and other cells simulate plasmodium.

Basic methods for studying protozoa

Let's briefly look at the most common research methods for the presence of protozoa.

Diagnosis of protozoa using a native smear and a smear stained with Lugol’s solution (in stool)

The drug is prepared from an emulsion of feces in an isotonic solution. Two drops of sodium chlorine and Lugol's solution are applied to a glass slide. The test material is added to both compositions with a wooden stick and, after covering with glass, is viewed at different microscope resolutions.

The protozoa found are recorded based on certain characteristics. For accuracy, prepare 2-3 preparations from the same material. In doubtful cases, the analysis is repeated several times over 2-3 weeks.

The method can detect vegetative and cystic forms:

• lamblia;
• balantidium;
• dysenteric amoeba.

Along with pathogenic forms, non-pathogenic protozoa are also identified. Also in healthy carriers there are luminal and cystic forms.

Important:research should be carried out repeatedly to avoid inaccuracies and errors.

The result of diagnosing protozoa using the native and stained smear method should contain a description of the form of the pathogen (luminal, cyst, tissue).

Research requirements:

• the material taken for analysis (liquid feces) is examined no later than 30 minutes after defecation;
• formalized stool must be diagnosed within 2 hours after defecation;
• the material should not contain impurities (disinfectants, water, urine);
• to work with the material, use only wooden sticks; glass ones are not suitable due to the slipping of mucus;
• Sticks must be burned immediately after use.

Preservation method (stool examination) for diagnosing protozoa

The study is carried out by fixing protozoa with a preservative. The difference between this method and the previous one is that preservatives allow you to preserve the drug for a long period.

Preservatives used:

• Barrow. Contains preservative ingredients: 0.7 ml sodium chloride, 5 ml formalin, 12.5 ml 96% alcohol, 2 g phenol and 100 ml distilled water. Coloring composition: 0.01% solution of thionin (azura).
• Safarliev's solution. Ingredients: 1.65 g zinc sulfate, 10 ml formalin, 2.5 g crystalline phenol, 5 ml acetic acid, 0.2 g methylene blue, 100 ml water. This preservative is used in cases where the material must be stored for more than a month.

Empty bottles are filled with a preservative, the material is transferred into them in a 3:1 ratio, then dye is added if necessary. The results are assessed by studying 2-3 drugs.

Formalin-ether enrichment method (analysis for the presence of protozoa in feces)

This diagnostic method allows you to separate and concentrate protozoan cysts. The following ingredients are needed for the analysis: formalin (10 ml), 0.85 g of isotonic solution, distilled water, sulfuric ether, Lugol's solution.

The mixture of biomaterial with the listed liquids is mixed and centrifuged. The sediment obtained at the bottom of the test tube is stained with Lugol's solution and examined for the presence of cysts and vegetative forms.

Method for detecting Leishmania (bone marrow smear)

To diagnose leishmaniasis, the following reagents are used: Nikiforov’s mixture (sulfuric ether and ethanol), phosphate buffer, Azur-eosin according to Romanovsky.

The bone marrow substance is very carefully placed on a glass slide after special preparation. A microscope with an immersion system is used.

IN acute period disease, a large number of leishmania are found in the punctate.

Note:Sometimes the blood cells may resemble processed Leishmania, so it is very important for the laboratory technician to be careful and have sufficient experience to examine them independently.

Method for detecting leishmania in a smear from skin infiltrate

The required reagents are similar to the previous analysis.

The test material is obtained from the existing tubercle or ulcerative contents. If leishmaniasis is suspected, scraping is done very carefully with a scalpel, without blood. Then the preparation is prepared on glass. To ensure the accuracy of the results obtained, several preparations are simultaneously examined.

In the presence of the disease, leishmania is also detected among the macrophages, fibroblasts, and lymphoid cells present in the test material.

Method for isolating a pure culture of Leishmania obtained by scraping pathological tissues

With this method of diagnosing protozoa, tissue scrapings are placed in a special nutrient medium, in which active reproduction of Leishmania occurs.

Before taking the scraping, the skin is thoroughly treated with alcohol, then an incision is made into the tubercle, from the bottom of which the contents are removed and placed in a test tube with medium. The material is taken several times, after which it is placed in different test tubes. Then cultivation occurs in a thermostat at a temperature of 22-24 degrees. The results are assessed under a microscope. This method is used when other, cheaper and quick ways Diagnosis of protozoa is ineffective.

You can see how tests for the presence of protozoa using a drop of blood are deciphered in practice by watching the video review:

Lotin Alexander, medical columnist

To detect fragments of helminths, feces are examined with the naked eye, then mixed with water and examined in small portions in a Petri dish against a dark background. All suspicious particles are placed on a glass slide in a drop of water and examined under a magnifying glass. You can place a daily portion in a cylinder with the addition of 5-10 times the amount of water. After stirring, the vessel is left until the suspended particles settle completely. The surface layer of liquid is drained and clean water is poured. The washed sediment is examined in small portions with the naked eye or under a magnifying glass. Microscopic examination methods are used to detect eggs.

Native smear method. A small amount of feces from different places in the test portion is ground on a glass slide in a drop of 50% glycerin solution, isotonic sodium chloride solution or water. The mixture is covered with a coverslip and viewed under a microscope.

Fulleborn floating method. One part of the feces is mixed with 20 parts of a saturated solution of sodium chloride (specific gravity 1.18), added in small portions. Large particles that float to the surface are immediately removed, and the mixture is left for 45 minutes. During this time, helminth eggs, having a lower specific gravity than the sodium chloride solution, float to the surface. The surface film is removed with a wire loop about 1 cm in diameter and transferred to a glass slide for examination under a microscope.

Kalantaryan method. The effectiveness of the floating method increases when replacing sodium chloride with a saturated solution of sodium nitrate. In this case, the mixture is kept for 10-15 minutes.

The surface film formed after settling a mixture of feces with a solution of sodium chloride or sodium nitrate can also be removed with a glass slide. For this purpose, a jar filled to the brim with a mixture of feces and a salt solution is covered with a glass slide so that its lower surface is in contact with the liquid. After settling, the glass is removed and, quickly turning upward with the surface on which the film is located, is examined under a microscope.

Scraping of the sperianal folds (to identify pinworm eggs and bovine tapeworm oncospheres) is done in the morning before toileting. Using a wooden spatula soaked in water or a 50% glycerin solution, scrape around the anus. The resulting material is transferred to a glass slide in a drop of water or 50% glycerol solution and viewed under a microscope. The spatula can be replaced with a damp cotton swab, which is used to wipe the perianal area, then rinse well in water. The water is centrifuged and the sediment is examined under a microscope.

Behrmann method (for identifying larvae). A metal mesh with 5-6 g of feces applied to it is fixed on a glass funnel inserted into a tripod. A rubber tube with a clamp is placed on the lower end of the funnel. The funnel is filled with water heated to t° 50°, so that Bottom part the mesh containing feces came into contact with the water. The larvae actively move into the water and accumulate in the lower part of the rubber tube. After 4 hours, the liquid is drained into centrifuge tubes, centrifuged, and the sediment is examined under a microscope.

Analysis of sputum, nasal mucus and vaginal discharge to identify eggs of the pulmonary trematode paragonimus, roundworm and hookworm larvae, pinworm eggs, and fragments of echinococcal bladder. The test portion of mucus (discharge) is smeared onto glass and viewed macroscopically against a black and white background, and then under a microscope. You can add a 25% antiformin solution to the test material, shake thoroughly and leave for 1-1.5 hours to dissolve the mucus. The mixture is centrifuged and the precipitate is examined under a microscope.

Analysis of duodenal and gastric juice for identifying eggs of liver flukes, hookworms, Strongyloides larvae. All three portions of duodenal contents obtained with are centrifuged and the sediment is examined under a microscope. They are also researching.

Tissue research. To identify Trichinella larvae, pieces of biopsied muscle are carefully split into fibers, squeezed between compressor glasses (thick glasses with screws) and examined under a microscope with a darkened light. To identify cysticerci, the muscles are dissected with dissecting needles, the isolated vesicle is cleared of surrounding tissue, squeezed between two glass slides and examined under a magnifying glass.

Blood test (to detect filaria larvae). Examine the hanging drop on a cover glass edged with Vaseline. You can mix 0.3 ml of blood with 10 times the amount of 3% solution. The mixture is centrifuged and the precipitate is examined under a microscope. To enrich the preparations, add 3 ml of 2% formalin solution or 5 times the amount of liquid consisting of 95 ml of 5% formalin solution, 5 ml of acetic acid and 2 ml of concentrated alcohol solution hematoxylin. The mixture is centrifuged, the precipitate is washed with distilled water and examined under a microscope. For differentiation different types filariae are examined by smears stained using the Giemsa-Romanovsky method.

Immunological diagnostic methods. Allergic diagnostic tests (see) with the corresponding type of helminth are also used (agglutination, complement fixation).

Helminthological research methods. Rice. Helminth eggs. 1-10 - eggs of roundworms (nematodes): 1 - 3 - roundworms (1 - fertilized egg, 2 - fertilized egg without albumen, 3 - unfertilized egg); 4 - cat roundworms; 5 - carnivorous roundworms; 5 - pinworms; 7 - whipworm; 8 - tominx; 9 - hookworm; 10 - trichostrongylid. 11-15 - eggs of tapeworms (cestodes): 11 - bovine tapeworm; 12 - dwarf tapeworm; 13 - rat tapeworm; 14 - pumpkin tapeworm; 15 wide tape. 16 - 24 - eggs of flukes (trematodes): 16 - trematodes (schistosomes) Japanese; 17 - trematodes (schistosomes) urine - sexual; 18 - trematodes (schistosomes) Munson; 19 - trematodes (parogonymus) pulmonary; 20 - trematodes (opisthorchis) Siberian (feline); 21 - trematodes (clonorchis) chinensis; 22 - intestinal trematodes (metagonimus); 23 - trematodes (fasciolas) of the liver; 24 - trematodes (dicrocelium) lanceolate.