4. Pneumonia: laboratory and instrumental diagnostics.
Pneumonia– acute infectious lesion of the lower respiratory tract, confirmed x-ray, dominant in the picture of the disease and not associated with other known causes.
Laboratory diagnostics:
Complete blood count: leukocytosis, shift of the leukocyte formula to the left, accelerated ESR.
Biochemical blood test: increased amounts of alpha-2 and gamma globulins, seromucoid, sialic acids, fibrinogen, C-reactive protein.
Sputum analysis - a large number of leukocytes, alveolar epithelium, there may be a small number of red blood cells.
Bacteriological examination of sputum (or bronchial washings) before prescribing antibiotics helps to detect the pathogen and determine its sensitivity to antibiotics;
Bacterioscopy (microscopy of sputum smears stained by Gram) - identification of gram-negative or gram-positive microorganisms (important to consider when choosing antibiotics at the time of hospitalization);
In the diagnosis of viral and viral-bacterial pneumonia, virological and serological studies are important.
Instrumental diagnostics:
X-ray studies, CT scan of the lungs - infiltration of lung tissue, pleural reaction.
If necessary, differential diagnosis of pneumonia with tuberculosis and lung cancer is performed by bronchoscopy, as well as pleuroscopy;
Ultrasound – diagnosis of effusion into the pleural cavity;
Indicators of external respiration function - assessment of the state of bronchial patency.
Examination card No. 6
Hypertensive crises: clinic, diagnosis, emergency care
Pulmonary heart: etiology, pathogenesis, clinical picture, diagnosis, treatment.
Sample answers:
Hypertensive crises: clinical picture, diagnosis, emergency care.
A hypertensive crisis is an acute, pronounced increase in blood pressure (BP), accompanied by clinical symptoms, requiring an immediate, controlled reduction in order to prevent or limit target organ damage.
In most cases, a hypertensive crisis develops with a systolic blood pressure >180 mmHg. Art. and/or diastolic blood pressure >120 mm Hg. Art., however, a crisis may develop with a less pronounced increase in blood pressure.
An acute significant increase in blood pressure, accompanied by pronounced clinical manifestations of the brain (encephalopathy), heart (angina pectoris, arrhythmias, left ventricular failure) and kidneys (proteinuria, hematuria, azotemia).
Clinic.
Hypertensive crises are divided into two large groups - complicated (life-threatening) and uncomplicated (non-life-threatening).
A complicated hypertensive crisis is accompanied by life-threatening complications, the appearance or worsening of target organ damage and requires a decrease in blood pressure, starting from the first minutes, over several minutes or hours using parenterally administered drugs.
A hypertensive crisis is considered complicated in the following cases:
Acute hypertensive encephalopathy;
Stroke;
Acute coronary syndrome;
Acute left ventricular failure;
Dissecting aortic aneurysm;
Crisis with pheochromocytoma;
Preeclampsia in pregnant women;
Severe hypertension associated with subarachnoid hemorrhage or traumatic brain injury;
Arterial hypertension in postoperative patients and with the threat of bleeding;
Crisis due to the use of amphetamines, cocaine, etc.
2. An uncomplicated hypertensive crisis, despite pronounced clinical symptoms, is not accompanied by acute clinically significant dysfunction of target organs.
Treatment.
All patients with a hypertensive crisis require a rapid reduction in blood pressure, however, the speed of its reduction is determined by the specific clinical situation.
Complicated hypertensive crisis– the fastest possible hospitalization is required. Blood pressure should be reduced by no more than 25% in the first 1–2 hours. The fastest possible reduction in blood pressure is necessary for dissecting aortic aneurysm and severe acute left ventricular failure (pulmonary edema) - by 25% of the initial value in 5–10 minutes, the optimal time to achieve the target systolic blood pressure level 100–110 mm Hg. Art. is no more than 20 minutes.
Patients with stroke or hypertensive encephalopathy require a special approach, since an excessive and/or rapid decrease in blood pressure leads to an increase in cerebral ischemia. In the acute period of stroke, the question of the need to lower blood pressure and its optimal value is decided together with a neurologist individually for each patient.
For therapy complicated hypertensive crisis apply:
1) Vasodilators:
Enalaprilat: IV infusion over 5 minutes at a dose of 0.625 – 1.25 mg. The onset of the effect is after 15 minutes, the maximum effect is after 30 minutes, the duration of action is 6 hours. Administration of enalaprilat is preferable for ACS (in combination with nitrates), pulmonary edema, cerebrovascular accident (has minimal effect on cerebral blood flow);
Nitroglycerin: 10 mg per 100 ml of 0.9% sodium chloride solution (or perlinganite 10 mg, isoket 10 mg per 150-200 ml of 0.9% sodium chloride solution) intravenously. Preferred for ACS and acute left ventricular failure;
Sodium nitroprusside: the solution is prepared ex temperature, the rate of administration is 1-4 mg/kg/min, depending on the level of maintained blood pressure. It is the drug of choice for hypertensive encephalopathy, but it should be borne in mind that it can increase intracranial pressure.
It is possible to use metoprolol for ACS that has developed against the background of a crisis: 5 mg (5 ml) of the drug intravenously, the administration can be repeated at 2-minute intervals, the maximum dose is 15 mg (15 ml).
Esmolol is a selective ultra-short-acting β 1 -blocker: initial dose 250-500 mcg/kg bolus over 1-3 minutes, repeated bolus administration is possible. It is possible to administer 50-100 mcg/kg/min as an infusion until the effect is achieved. The onset of action is in 60 seconds. Duration of action – 20 minutes. Drug of choice for ACS, tachyarrhythmias, cerebral complications, dissecting aortic aneurysm.
Antiadrenergic drugs:
Urapidil (Ebrantil) is a central α 1-adrenergic blocker,
having a weak β-blocking effect: administered intravenously as a bolus or by long-term infusion. Intravenous bolus: 10-50 mg of the drug is administered slowly under blood pressure monitoring, a decrease in which is expected within 5 minutes after administration. Depending on the effect, repeated administration of the drug is possible.
Intravenous drip or continuous infusion is carried out using a perfusion pump. Maintenance dose: on average 9 mg/h, i.e. 250 mg of the drug (10 ampoules of 5 ml or 5 ampoules of 10 ml) in 500 ml of solution for infusion (1 mg = 44 drops ~ 2.2 ml). Recommended maximum initial rate: 2 mg/min. The rate of drip administration depends on the patient’s blood pressure.
Proxodolol (Albetor) – β 1-2, α 1-adrenergic blocker: administered intravenously by bolus or by infusion.
IV bolus - 10-20 mg (1-2 ml of 1% solution (10 mg/ml) for 1 minute. If necessary, repeat administration at intervals of 5 minutes until the effect appears. Maximum dose - 50-100 mg (5 –10 ml of 1% solution (10 mg/ml).
IV drip - 50 mg (5 ml of 1% solution (10 mg/ml) in 200 ml of 0.9% sodium chloride solution or in 5% glucose solution, at a rate of 0.5 mg/min (2 ml of infusion solution) until a positive reaction is obtained.
4). Diuretics:
Furosemide: 40 – 80 mg IV. It is used as part of a complex of therapeutic measures for acute left ventricular failure.
5). Other drugs:
Neuroleptics (droperidol): 0.25% solution 1-2 ml IV slowly in 20 ml of 5% glucose solution;
Ganglion blockers (pentamine): 5% solution 0.3-1 ml IV slowly in 20 ml of 5% glucose solution.
B. Uncomplicated hypertensive crisis
Treatment of uncomplicated hypertensive crisis is possible on an outpatient basis, using both intravenous and oral or sublingual use of antihypertensive drugs (depending on the severity of the increase in blood pressure and clinical symptoms). Treatment should begin immediately, the rate of reduction in blood pressure should not exceed 25% in the first 2 hours, followed by achieving target blood pressure within several hours (no more than 24–48 hours) from the start of therapy.
1) Captopril: 12.5 – 25 mg sublingually or orally. Repeated use is possible after 90-120 minutes.
2) β-blockers: propranolol 5–20 mg sublingually or metoprolol 25–50 mg sublingually. Drugs of choice for young patients with hypersympathicotonia, during a crisis associated with alcohol intake, in patients with concomitant ischemic heart disease.
3) Clonidine: 0.01% - 0.5 (1.0) ml in 10-20 ml of 0.9% sodium chloride solution, administered intravenously over 5-7 minutes or - 0.75 (1.5 ) ml IM; orally 0.1 - 0.2 mg followed by 0.05 - 0.1 mg every 4 hours. The maximum total dose is 0.7 mg.
Anaphylactic shock: clinic, emergency care.
Anaphylactic shock - an acutely developing, life-threatening process that occurs as a pronounced manifestation of anaphylaxis and is characterized by severe disturbances in the activity of the central nervous system, blood circulation, respiration and metabolism.
Clinic
Shock is manifested by a feeling of fear, anxiety, dizziness, tinnitus, a feeling of heat, lack of air, tightness in the chest, nausea, and vomiting. Urticaria and soft tissue swelling may occur. Acute vascular insufficiency is manifested by sticky cold sweat, sharp pallor of visible mucous membranes and skin, thread-like pulse, and a sharp drop in blood pressure. Consciousness is depressed, breathing is impaired. Further deterioration of the clinical picture is characteristic of developing coma caused by cerebral hypoxia.
Urgent Care:
1. Stop introducing the allergen.
2. Ensure airway patency; if it is impossible to intubate the trachea, conicotomy is performed.
3. Give your legs an elevated position.
4. Inhalation of 100% oxygen (no more than 30 minutes); provide access to the vein.
5. Administer adrenaline - 0.1% 0.3–0.5 ml in 20 ml of 0.9% sodium chloride solution or 5% glucose solution intravenously (repeat if necessary).
6. Start intravenous fluid administration (polyglucin, rheopolyglucin, 0.9% sodium chloride solution, 5% glucose solution).
7. If swelling spreads to the larynx area, administer 2-3 ml of adrenaline endotracheally in 20 ml of 0.9% sodium chloride solution.
8. Introduce glucocorticoid hormones intravenously (prednisolone 90-150 mg or hydrocortisone hemisuccinate 300-600 mg (in a dropper or stream) - if ineffective, repeat.
9. Inject intravenously 2 ml of 1% diphenhydramine solution or 1-2 ml of suprastin solution intravenously.
10. Inject slowly 10 ml of 2.4% aminophylline solution intravenously.
11. For brochospasm - Salbutamol 2.5 mg or Berodual 1 ml inhaled through a nebulizer.
12. After stabilization of the condition - transportation to the hospital.
Kidney amyloidosis: etiopathogenesis, clinical picture, diagnosis, treatment.
Amyloidosis is a systemic disease characterized by extracellular deposition of a special eosinophilic protein of varying origin. In general, the incidence of renal amyloidosis is 75% of all cases of amyloidosis. Moreover, renal failure is the leading cause of death in such patients, second only to heart failure.
The question of what diseases amyloidosis develops is not fully understood, although tuberculosis is still primarily called rheumatoid arthritis. One should remember the continuing possibility of amyloidosis in chronic suppurations - osteomyelitis, bronchiectasis and other chronic pulmonary suppurations, syphilis, as well as lymphogranulomatosis, tumors of the renal parenchyma, lung, ulcerative colitis, Crohn's and Whipple's diseases, prolonged septic endocarditis and other, more rare diseases (for example , medullary thyroid cancer. Recently, more and more attention has been paid to the development of amyloidosis in old age (especially in people over 70-80 years of age).
The study of the biochemical properties of the protein in the 70-80s made it possible to establish that it is heterogeneous in origin and is a derivative of normal and pathological serum proteins and consists of their polypeptide fragments. Such amyloid precursors can be: prealbumin, light chains of immunoglobulins, some hormones of the endocrine glands, etc.
Pathogenesis.
Until now, there is no single concept of its pathogenesis for all forms of amyloidosis. Currently, four main theories of the pathogenesis of amyloidosis are being discussed.
According to the theory of local cellular genesis, amyloid formation can be divided into two phases: pre-amyloid and amyloid itself. In the pre-amyloid phase, plasmacytic infiltration and proliferation in the system of phagocytic macrophages is observed.
The theory of disproteinosis (or organoproteinosis) considers amyloid as a product of perverted protein metabolism. From the standpoint of this theory, the main link in the pathogenesis of amyloidosis is dysproteinemia with accumulation in the plasma of coarse protein fractions and abnormal proteins - paraproteins.
According to the mutation theory, various variants of amyloidosis develop as a result of the mutational formation of a special type of amyloidoblast.
Immune theory of amyloidosis; A particularly important role is played by immune disorders in the amyloid stage - inhibition of the function of cellular immunity and phagocytosis.
Clinic.
Secondary amyloidosis is a disease mainly found in people over 40 years of age. There is a high percentage of kidney damage. 60% of patients have massive proteinuria; nephrotic syndrome occurs quite early; in half of the patients already in the first 3 years. We can talk about a certain stage of the process:
If a person suffers from a chronic cough for a long time, then he is rarely able to independently recognize pneumonia (pneumonia), bronchitis or other acute inflammatory process in the lower respiratory tract. Bronchitis and pneumonia often occur after a cold. Both pathologies can be of either a viral or bacterial nature.
Acute pneumonia and acute bronchitis have many similar signs and symptoms, so it is impossible to confidently determine pneumonia at home without the participation of specialists and without using “gold standards” of diagnosis.
Differential diagnosis
Differential diagnosis of pneumonia is extremely important because, unlike bronchitis, it must be treated completely differently. The differential diagnosis of pneumonia should be based on the patient’s symptoms and laboratory and instrumental criteria:
- The disease usually begins acutely and is accompanied by a febrile state (body temperature exceeds 38°C).
- Sputum with pus and blood impurities appears.
- Auscultation reveals previously absent, local shortening of the pulmonary sound during percussion.
- The leukocyte formula is observed with a shift to the left.
- During the examination, radiology specialists will indicate the main x-ray sign - the accumulation of blood and cells in a certain area of the lung, which creates a certain pattern of darkening.
Diagnosing pneumonia is not an easy task. In addition to the fact that this disease is easily confused with other pathological processes, it itself can be of several types and have a different clinical picture. The classification of pneumonia in adult and pediatric patients depends on the form, etiology, conditions of occurrence, characteristic signs, localization and possible complications.
Based on its form, this pathology is divided into the following groups:
- out-of-hospital (home) - most common;
- inflammatory-infectious process acquired during hospital stay;
- Pneumocystis - observed in patients with immunodeficiency.
Of great importance in treatment tactics is the identification of the pathogen, which may be one of the following pathogens:
- pneumococcus;
- mycoplasma microorganisms;
- chlamydia pneumonia;
- legionella;
- Pfeiffer wand;
- Staphylococcus aureus;
- Friedlander's wand;
- non-cellular infectious agents;
- pathogenic fungi.
- focal (bronchopneumonia) occurs with small localizations or confluent;
- lobar (lower lobe);
- bilateral (both lungs are affected).
Diagnosis and treatment of pneumonia should be given serious attention, since this disease can be fatal without taking appropriate measures. The mortality rate in middle-aged patients who do not suffer from serious somatic diseases is no more than 3%. But pneumonia in older people, aggravated by chronic diseases, has a poor prognosis in every third patient.
Basic diagnostic methods
Patients who value their lives will not take long to figure out how to determine pneumonia at home, and at the first suspicion of pneumonia, they will go to a medical clinic for diagnosis and treatment, or at least call a paramedic at home.
An experienced pulmonologist knows very well how to recognize pneumonia and how to distinguish bronchitis from pneumonia.
Diagnosis of pneumonia includes the following laboratory and instrumental examinations:
- fluoroscopy;
- clinical blood test;
- biochemical blood test;
- urine test;
- examination of sputum and smears;
- assessment of blood gas composition.
In addition, the methodology for examining a person may include the following diagnostic procedures:
- lung biopsy;
- bronchoscopy.
However, before resorting to the methods described above, the pulmonologist begins the examination by assessing the patient’s auscultatory pattern.
Auscultation
When pulmonologists diagnose pathology using auscultation, acute pneumonia can give the following signs of the disease:
- Increased signs of bronchial phonia appear on the sides of the affected area.
- With focal lesions, mixed breathing can be heard. On inhalation, it is characterized as vesicular, and on exhalation, weakened bronchial breathing is observed.
- In the lobar form of the disease, a crunching sound is heard in the initial phase and in the resolution stage.
- Pleuropneumonia is characterized by pronounced noise created by friction of the pleural layers. And when effusion forms, a sharp weakening of breathing is observed.
- In severe cases of the inflammatory process, listening to the heart reveals a pronounced acceleration of the heartbeat.
Especially in elderly patients, a sharp decrease in blood pressure may occur with the development of acute vascular insufficiency.
Diagnosis of the lobar form of the disease always begins with listening. This pathology has 2 informative characteristics:
- Alveolar crackling on inspiration, which does not go away after the patient coughs.
- Moist wheezing on inspiration. They are explained by the fact that when the air flow passes, the bubbly viscous exudate in the bronchi bursts.
With focal pneumonia, a specific crunching sound appears in the first and third stages. And wheezing due to the accumulation of liquid secretions is classified as fine-bubble ringing. Auscultation for lobar pathology helps the pulmonologist determine the stage of inflammation.
Video
Video - What is pneumonia?
X-ray diagnostics
X-ray diagnosis of the disease plays a key role in making an accurate diagnosis. But almost all patients know that when performing an X-ray for pneumonia, radiation exposure to the body cannot be avoided, so they are interested in whether fluorography will show pneumonia.
In fact, the task of fluorographic examination is to prevent serious diseases of the lower respiratory tract. And it occupies a worthy place in radiology. However, if you compare a photo of the lungs on fluorography and a lung x-ray, it becomes obvious that darkening or pathology is better visible on an x-ray.
Whether the disease can be seen on fluorography and how well depends largely on the type of infection that affects the pulmonary structures. In turn, an x-ray of the lungs for pneumonia is not only a way to detect the disease, but it can also be used to monitor treatment and notice positive or negative dynamics.
Lobar pneumonia on an x-ray may look like this:
- extensive darkening;
- one-sided total or partial darkening;
- limited dimming (changes do not extend beyond the segment).
X-rays do not always show focal pneumonia, since in the initial stages the disease is characterized by the appearance of small compactions with biological structures. And yet, an experienced radiologist knows how to identify inflammatory processes even in the absence of compactions, since a focal form on an x-ray can show indirect signs:
- local pathological shadows;
- inflammation of the pleura with the formation of fibrous plaque on its surface or effusion inside it;
- enlargement of pulmonary roots;
- distortion of the pulmonary pattern in a certain area.
If the disease is clearly visualized on an x-ray, but there is a need to detect more subtle details of the pathological process, then CT is used. Computed tomography can detect pneumonia at the earliest stages, since even then it clearly shows pathological changes in tissues.
Blood test
If, when undergoing a preventive examination, patients doubt whether fluorography will show pneumonia, then ESR (erythrocyte sedimentation rate) indicators will definitely help not to miss a developing disease. During a standard examination, they can greatly exceed the norm. And even more so when a general blood test is taken for pneumonia.
As a rule, acute pneumonia has an ESR of 20-25 mm/hour. In severe cases it can reach 40-50 mm/hour. And if ESR indicators are 80 mm/hour or higher, then they may suspect oncology or an autoimmune disease. However, it is worth considering that ESR is not an absolute indicator, especially for patients who are undergoing therapy that suppresses unwanted immune reactions of the body.
In case of lobar pneumonia, diagnosis cannot be done without a blood test. The hemogram in this case has a standard description - acceleration of ESR and a shift of the leukocytosis formula to the left. If the level of leukocytes in the blood reaches more than 10-12x109/liters, then this indicates a high probability of bacterial infection, and if their level becomes below 3x109/liters or above 25x109/liters, then such indicators indicate an unfavorable prognosis.
Biochemistry is also done, but such a blood test for pneumonia does not provide specific information.
Urine and stool examination
If a pathogen is detected in feces and urine, specific treatment is selected. This is especially true in the case of its Klebsiella form. If the bacterial process is not detected in a timely manner and adequate treatment is not begun, the pathology will begin to spread to the gastrointestinal tract, hepatobiliary region, urinary system, and may even affect the brain.
If Klebsiella pneumonia appears in the urine, this indicates that the process has affected the genitourinary system. Any other acute pneumonia shows a small amount of red blood cells in the urine (microhematuria), as well as protein (proteinuria), which does not happen normally. A urine test for pneumonia is not one of the most informative tests.
If Klebsiella pneumonia is detected in the stool, the digestive system is considered to be the source of the lesion. But it should be borne in mind that this bacterium, under normal conditions, is a standard inhabitant of the large intestine and the general microflora of the gastrointestinal tract. Laboratory examination of feces makes it possible to clearly recognize abnormalities in infants. And if it turns out to detect Klebsiella pneumonia in the stool of an adult, then this information will not be so significant.
Sputum examination
At the discretion of the attending physician, who knows exactly how to determine pneumonia, a sputum test for pneumonia may be prescribed. The study of secretions obtained from the respiratory tract is carried out in order to identify the type and type of causative agent of the developed disease. The decisive results are the results of bacterioscopy of a sputum smear, which is stained with Gram and microbiological examination is carried out.
To confirm chlamydial pathology, the most specific and sensitive method is the cultural method of isolating the pathogen, however, it is characterized by a lengthy and labor-intensive process and in practice is usually limited to serotyping. When performing a standard bacterioscopy of a sputum smear for mycoplasma, it is not possible to detect this pathogen, therefore the method of immunofluorescence or enzyme-linked immunosorbent assay is used.
In addition, there is a whole scheme for how to diagnose rare pneumonia caused by Friendlander's bacillus. Microbiological tests are being carried out to inoculate bacteria. The pathogen can be detected in a smear from the cervical canal, in a smear from the throat, and sometimes Klebsiella pneumonia is found in a smear of mucus from the throat. As a rule, the latter case is characteristic of nosocomial infection.
Formulation of diagnosis
After carrying out all the specific tests for pneumonia, assessing what pneumonia looks like in the picture, a diagnosis will be formulated. It will indicate what kind of pathology it is - bronchitis or pneumonia, as well as other details:
- the pathogen that provoked the occurrence of the pathology;
- the area of location of the process and its scale (the lesion affects a segment, a lobe, one side or two at once);
- pneumonia severity criteria;
- characteristics of complications, if any;
- what phase the disease is in (beginning, peak, restoration of normal structure, resolution, final stage, protracted process);
- general picture of somatic diseases that may affect recovery.
An example of a diagnosis: “Lobar acute pneumonia caused by pneumococcus. There is a lower lobe right-sided lesion. The disease is in its peak phase and is severe. Acute partially compensated respiratory failure is observed.”
Patients who have suffered a severe form of lower respiratory tract disease must carefully monitor their health in the future and undergo regular preventive examinations. Even if they feel well, once a year they are required to undergo radiography or fluorography.
State sanitary and epidemiological regulation
Russian Federation
MICROBIOLOGICAL FACTORS
Laboratory diagnostics
community-acquired pneumonia
Guidelines
MUK 4.2.3115-13
Official publication
4.2. CONTROL METHODS. BIOLOGICAL AND
MICROBIOLOGICAL FACTORS
Laboratory diagnosis of community-acquired pneumonia
|
Guidelines |
1. Scope of application
1.1. These guidelines substantiate and define the methodological foundations and algorithms for laboratory diagnosis of pneumonia in the implementation of epidemiological surveillance in relation to community-acquired pneumonia.
1.2. The guidelines are intended for specialists from bodies and institutions of the Federal Service for Supervision of Consumer Rights Protection and Human Welfare, and can also be used by specialists from medical organizations and other interested organizations.
1.3. Guidelines are mandatory when carrying out epidemiological surveillance of community-acquired pneumonia, during anti-epidemic measures and during epidemiological investigations of possible epidemic outbreaks of community-acquired pneumonia.
2. Terms and abbreviations
WHO - World Health Organization.
CAP - community-acquired pneumonia.
LPO is a treatment and preventive organization.
ICD-10 - international classification of diseases.
ARVI is an acute respiratory viral infection.
PCR - polymerase chain reaction.
RT-PCR - real-time polymerase chain reaction.
RIF - immunofluorescence reaction.
ELISA - enzyme immunoassay.
ICA - immunochromatographic analysis.
ABT - antibacterial therapy.
ICU - intensive care unit.
BAL - bronchoalveolar lavage.
3. General information about community-acquired pneumonia
Pneumonia is a group of acute infectious diseases, different in etiology, pathogenesis, and morphological characteristics, characterized by focal damage to the respiratory parts of the lungs with the obligatory presence of intra-alveolar exudation. In the International Classification of Diseases, Injuries and Causes of Death, 10th revision (ICD-10, 1992), pneumonia is clearly separated from other focal inflammatory lung diseases of non-infectious origin. The modern classification of pneumonia takes into account, first of all, the epidemiological conditions of the development of the disease, the characteristics of infection of the lung tissue and the state of the immunological reactivity of the patient’s body. Based on the nature of acquisition, community-acquired pneumonia (CAP) and nosocomial (nosocomial) pneumonia are distinguished. Recently, in addition to the term “nosocomial pneumonia”, a broader term has been used - “pneumonia associated with the provision of medical care” ( healthcare-associated pneumonia). This category, in addition to nosocomial pneumonia, includes pneumonia in persons in nursing homes or other long-term care facilities. It should be emphasized that such a division has nothing to do with the severity of the disease; the main criterion for differentiation is the epidemiological conditions and environment in which pneumonia developed. However, they usually differ from CAP in the etiological structure of pathogens and the profile of antibiotic resistance.
CAP should be understood as an acute illness that occurred in a community setting - that is, outside a hospital or later than 4 weeks after discharge from it, or diagnosed within the first 48 hours of hospitalization, or developed in a patient who was not in a nursing home/long-term medical unit. observation for 14 days or more - accompanied by symptoms of lower respiratory tract infection (fever, cough, sputum production, possibly purulent, chest pain, shortness of breath) and radiological signs of “fresh” focally infiltrative changes in the lungs in the absence of an obvious diagnostic alternative.
The modern classification of CAP, taking into account the state of the immunological reactivity of the patient’s body, allows us to distinguish 2 main groups, suggesting differences in the etiological structure of pneumonia:
Typical CAP (in patients with no significant immune disorders);
CAP in patients with severe immune disorders (acquired immunodeficiency syndrome; other diseases or pathological conditions).
4. Modern ideas about the etiological structure of community-acquired pneumonia
The absolute significance of the etiological role of a particular CAP pathogen can only be determined in relation to a specific region, epidemic focus or epidemiological situation. Broader generalizations make it possible to identify the main trend that determines the significance of a given pathogen in human infectious pathology based on the appropriate level of standardization and frequency of use of laboratory diagnostic methods, as well as the approximate ratio of CAPs caused by the main pathogen of pneumonia - pneumococcus and other pathogens.
According to domestic and foreign researchers S. pneumoniae is the dominant etiological agent of pneumonia, causing from 30 to 80% of CAP in people of all age groups (Pokrovsky V.I. et al., 1995; Zubkov M.N., 2002, Cuhna V.A., 2003, Chuchalin A.G. ., 2006).
Against the backdrop of an increase in populations with severe immune defects (HIV infection, congenital immunodeficiency, oncohematological diseases, etc.), the etiological significance of such opportunistic pathogens of CAP as Pneumocystis juroveci, cytomegalovirus. Taking into account the high level of carriage of these pathogens, diagnosis of the corresponding nosology should be carried out only in groups at risk using modern laboratory research algorithms.
The concept of “viral pneumonia” has not yet found widespread use in diagnosing CAP, however, ICD-10 distinguishes pneumonia caused by influenza viruses, parainfluenza, adenoviruses and others from the group of respiratory tract infection pathogens. At the same time, the viral-bacterial etiology of CAP is quite widely known and described against the background of epidemics of influenza and acute respiratory infections. The domestic standard of specialized medical care for severe pneumonia with complications includes J10.0 “Influenza with pneumonia” (influenza virus identified) and J11.0 “Influenza with pneumonia” (influenza virus not identified) as nosological units.
Viral respiratory tract infections are more severe in children under 5 years of age and elderly people (over 65 years of age), which is reflected in the high level of hospitalizations for pneumonia and mortality among people of this age. In these age groups, viral and viral-bacterial pneumonia are more often recorded.
During influenza epidemics, the risk of developing pneumonia may increase for those age groups in which the level of anamnestic antibodies to the antigenic variant of the influenza virus circulating in a particular epidemic season is below the protective level, as for example, this was observed in the case of pandemic influenza A/H1N1pdm2009 for people from 30 to 60 years. Risk groups for developing pneumonia with influenza also include people suffering from chronic diseases of the cardiovascular system, metabolic disorders (obesity, diabetes), chronic diseases of the bronchopulmonary system, and pregnant women.
The etiological structure of CAP in children differs significantly from the etiology of CAP in adults and varies depending on the age of the child and the severity of the disease, which should be taken into account in the algorithm for diagnosing pneumonia in children. Risk groups for severe pneumonia include children under 5 years of age, frequently ill children, and especially those born at 24–28 weeks of gestation.
Bacterial pathogens of pneumonia are found in 2 - 50% of children, more often in hospitalized children, compared to children undergoing outpatient treatment. The most common bacterial pathogens of community-acquired pneumonia in children over one year of age are considered S. pneumoniae, less often isolated N. influenzae type b, S. pneumoniae is the cause of one third of pneumonia with radiological confirmation in children under 2 years of age. In cases of severe pneumonia requiring intensive care, an infection caused by group A streptococci or S. aureus, which are found in 3 - 7% of cases. Moraxella catarrhalis found in 1.5 to 3.0% of cases of pneumonia in children. Mixed viral-bacterial pneumonia is diagnosed in children, according to various sources, in 8.2 - 33.0% of cases, and when taking into account all mixed: bacterial or viral-bacterial pneumonia in children, their frequency ranges from 8 to 40%. Among pneumococcal pneumonia in children, combination with viral infections is observed in 62% of cases.
In case of CAP in children, it is necessary to take into account the possibility of a mixed bacterial-viral infection, the etiological significance of well-known and recently discovered respiratory viruses: respiratory syncytial, metapneumovirus, bocavirus and rhinoviruses. Various viral pathogens of respiratory infections are found in 30 - 67% of cases of pneumonia in children, and their proportion is higher in young children (up to 80% of cases from 3 months to 2 years), and are much less common in children over 10 years of age. M. pneumoniae And C. pneumoniae predominantly cause pneumonia in school-age children, and are not typical for children from 1 to 5 years of age. These pathogens are more often detected during epidemic rises in incidence in foci of infection.
In endemic regions and according to epidemiological indicators, when etiologically diagnosing CAP, it is necessary to take into account the possibility of the occurrence of zoonotic infections, which are characterized by inflammatory processes in the lungs (Q fever, psittacosis, tularemia, etc.). An important element of the examination of patients with CAP is the exclusion of the etiological role of the causative agent of tuberculosis and other mycobacteria.
5. Logistics support for laboratory research
1. Laminar flow hood, 2nd class biological safety.
2. Binocular microscope with illuminator, set of lenses and eyepieces.
3. Electric thermostats for growing bacteria, maintaining the temperature in the chamber within (37 ± 1) °C.
4. CO 2 incubator, maintaining the temperature in the chamber within (37 ± 1) °C, CO 2 content at 3 - 7%, or an anaerostat.
5. Distiller.
6. Electric autoclave.
7. Refrigerator maintaining a temperature of 4 - 6 °C for storing cultures, biological substrates and reagents.
8. Alcohol lamps and gas burners.
9. Automatic and semi-automatic colony counters for counting colonies.
10. Disposable sterile containers for collecting and transporting sputum, pleural fluid, tracheal aspirate, BAL with a stable base, made of transparent material (preferably plastic to prevent breakage, facilitate disinfection and disposal of the container); the lid must seal the containers tightly and be easy to open; the container should not contain chemicals that negatively affect the viability of bacteria in the sputum.
11. Set of reagents for Gram staining of microslides.
12. Nutrient media for cultivation S. pneumoniae(eg blood agar, CNA agar).
13. Nutrient media for the cultivation of bacteria of the genus Haemophilus(such as chocolate agar), gram-negative bacteria and S. aureus(Endo agar, MacConkey agar, yolk salt agar).
14. Bacteriological dishes (Petri) for growing microbiological cultures.
15. Slides and coverslips of standard sizes for microslides.
16. Racks and trays for test tubes and containers, transportation of Petri dishes, cuvettes and rail racks for fixing and staining smears.
17. Bacteriological loops.
18. Semi-automatic variable volume dispensers.
19. Sterile tips for variable volume dispensers.
20. Drygalsky spatulas are sterile.
21. Laboratory glassware.
22. Plastic Pasteur pipettes for standardizing the volume and transfer of liquids.
23. McFarland turbidity standard or device for determining the concentration of bacterial cells.
24. Discs with antibiotics (optoquin, oxacillin, cefoxitin, etc.).
25. Enzyme immunoassay analyzer included.
26. Fluorescence microscope included.
27. Equipment for a PCR laboratory equipped in accordance with MU 1.3.2569-09
28. Diagnostic reagent kits (test systems) for identifying antigens and DNA/RNA of pneumonia pathogens, as well as specific antibodies to pneumonia pathogens, approved for use in the Russian Federation in the prescribed manner.
6. Diagnosis of community-acquired pneumonia
6.1. Diagnosis of pneumococcal pneumonia
Streptococcus pneumoniae (S. pneumoniae) is the most common bacterial pathogen of CAP. Pneumococcal pneumonia is registered in patients of any age, occurring both in outpatient practice and in hospital (including among those hospitalized in the ICU). An increase in the incidence of CAP of pneumococcal etiology in the Northern Hemisphere is observed in the winter season; pneumococcal pneumonia is more often registered among patients with concomitant chronic diseases - chronic obstructive pulmonary disease, diabetes mellitus, alcoholism, asplenia, immunodeficiency, and often occurs with bacteremia (up to 25 - 30%).
Pneumococcal CAP is usually characterized by an acute onset, high fever, and chest pain. However, clinical, laboratory and radiological manifestations of CAP caused by S. pneumoniae, are not specific enough and cannot be considered an adequate predictor of the etiology of the disease.
To diagnose pneumococcal CAP, culture methods are most often used. Clinical material for research is sputum, venous blood, less often - invasive respiratory samples (BAL, material obtained during bronchoscopy, protected brush biopsy, etc.) and pleural fluid.
When examining sputum, special attention should be paid to the need to assess the quality of the delivered sample. The analysis must begin with the preparation of a smear, since the results of microscopy influence not only the assessment of the suitability of the material, but also the further direction of bacteriological research. The criteria for the suitability of sputum for bacteriological examination are the presence of more than 25 segmented leukocytes and no more than 10 epithelial cells per field of view when viewing at least 20 fields of view of a Gram-stained smear (under magnification × 100). Microscopy of a Gram-stained smear (under magnification × 1,000 using an immersion objective) reveals gram-positive cocci (usually lanceolate diplococci) with a diameter of 0.5 - 1.25 µm, without spores and flagella; most have a polysaccharide capsule.
The study of pleural fluid involves bacterioscopy of a Gram-stained smear followed by a cultural examination. It is performed in the presence of pleural effusion and conditions for safe pleural puncture (visualization on the laterogram of freely displaced fluid with a layer thickness > 1.0 cm). Culture testing of invasive respiratory samples for CAP is recommended for patients with immunodeficiency; this method can be used in some cases with severe CAP, as well as the ineffectiveness of initial antibiotic therapy (ABT).
Microorganisms isolated from BAL in quantities of ≥ 10 4 CFU/ml, from a biopsy obtained using protected brushes - ≥ 10 3 CFU/ml, and sputum - ≥ 10 5 CFU/ml are considered clinically significant in an acute inflammatory process.
To highlight S. pneumoniae from clinical material, it is necessary to use nutrient media enriched with defibrinated animal blood (sheep, horse or goat) at a concentration of 5%. The use of defibrinated human blood gives slightly worse results. Due to the scarcity of defibrinated blood in practical laboratories and its short shelf life, one should remember the possibility of using commercially prepared chocolate agar for the isolation of pneumococci, which is simultaneously used for the isolation of hemophiliacs. Another cultivation condition S. pneumoniae- incubation in an atmosphere with a CO 2 content increased to 3 - 7%, since it is a facultative anaerobe. Probability of allocation S. pneumoniae from respiratory samples increases when using selective media containing additives that inhibit the growth of saprophytic and gram-negative microorganisms (colistin, nalidixic acid, gentamicin).
A key test for differentiating pneumococci from other α-hemolytic streptococci is sensitivity to optochin (the test is based on the ability of optochin to selectively inhibit the growth of pneumococci over other viridans streptococci). However, among S. pneumoniae the number of optoquine-resistant strains is growing, which requires the use of alternative methods for identifying the pathogen (lysis in the presence of bile salts, Neufeld test, agglutination with diagnostic pneumococcal sera).
The informative value of cultural examination of respiratory samples and blood largely depends on compliance with generally accepted rules for their collection, storage and transportation (see appendix). In addition, the likelihood of identifying St. pneumoniae significantly decreases when obtaining clinical samples against the background of systemic ABT. For blood culture, it is preferable to use commercial vials of culture media.
Among non-culture methods for diagnosing pneumococcal pneumonia, the immunochromatographic test, which involves the detection of pneumococcal cell polysaccharide antigen in urine, has become most widespread in recent years. Its main advantage is the ability to use it “at the patient’s bedside” due to the ease of implementation and quick results. The pneumococcal rapid test demonstrates acceptable sensitivity (50 - 80%) and fairly high specificity (> 90%) for CAP in adults compared to traditional methods. Disadvantages of the test include the possibility of obtaining false-positive results in pneumococcal carriage (the test is not recommended for children under 6 years of age) and in persons who have recently had CAP.
Methods for identifying St. pneumoniae in clinical material using PCR. Autolysin genes ( lytA), pneumococcal surface antigen ( psaA) and pneumolysin ( ply) and other target genes. However, these methods are not widely used in clinical practice and their place in the etiological diagnosis of CAP requires clarification.
6.2. Diagnosis of other bacterial pneumonias
An important clinically significant bacterial pathogen of CAP is Haemophilus influenzae (H. influenzae). Community-acquired pneumonia is usually caused by non-typeable strains N. influenzae. According to a number of studies, H. influenzae is more common in patients with concomitant COPD and active smokers; the incidence of infection with this pathogen is higher in patients with non-severe CAP.
Representatives of the family Enterobacteriaceae (Klebsiella pneumoniae, Escherichia coli etc.) and Pseudomonas aeruginosa (P. aeruginosa) are detected in less than 5% of patients with CAP and belong to the category of rare pathogens. However, the importance of these microorganisms may increase in patients with severe CAP, and infection increases the likelihood of an unfavorable prognosis several times.
As epidemiological studies show, the frequency of occurrence of enterobacteria is higher in patients with chronic concomitant diseases, in persons who abuse alcohol, during aspiration, in the case of recent hospitalization and previous ABT. Additional risk factors for infection P. aeruginosa are chronic bronchopulmonary diseases (severe COPD, bronchiectasis), long-term use of systemic steroids, cytostatics.
Another bacterial pathogen is Staphylococcus aureus (S. aureus) - rarely occurs among outpatients with CAP, at the same time, in people with severe disease, its proportion can increase to 10% or more. To infection S. aureus Many factors predispose - old age, living in nursing homes, drug addiction, alcohol abuse. It is known that the relevance S. aureus as a causative agent, CAP increases significantly during influenza epidemics.
There are no specific clinical, laboratory or radiological signs that are typical for CAP caused by these pathogens and that allow it to be distinguished from pneumonia of other etiologies. In some cases, mainly in people with immunosuppression or alcohol abuse, K. pneumoniae can cause lobar pneumonia with localization of the lesion in the upper lobe of the lung, rapid progression of disease symptoms and high mortality.
For the etiological diagnosis of CAP caused by these pathogens, the cultural method of research is of primary importance. N. influenzae, like pneumococcus, belongs to the category of “fastidious” microorganisms that require the presence of factors X, V and 5-7% CO 2 in the nutrient media in the incubation atmosphere for cultivation. To highlight N. influenzae From clinical material, chocolate agar or selective agar is usually used to isolate bacteria of the genus Haemophilus. Culture of clinical material to identify family members Enterobacteriaceae And P. aeruginosa carried out on selective media for the isolation of gram-negative bacteria (Endo, MacConkey agar, etc.), S. aureus- on yolk-salt agar, mannitol-salt agar, etc.
Clinical material for testing may include sputum, venous blood, invasive respiratory specimens and pleural fluid. When examining sputum, as well as for identifying pneumococci, assessing the quality of the collected sample is important. The study of pleural fluid is performed in the presence of pleural effusion and conditions for safe pleural puncture, invasive respiratory samples - only for certain indications.
It should be noted that non-typeable strains H. influenzae And S. aureus are part of the normal microflora of the upper respiratory tract (URT), and the frequency of asymptomatic carriage can be quite high. With age, in the presence of chronic concomitant diseases, as well as recent systemic ABT, the frequency of colonization of the oral cavity and upper respiratory tract with Enterobacteriaceae increases. This fact must be taken into account when clinically interpreting the results of bacteriological examination of respiratory samples, especially sputum.
The informative value of cultural examination of respiratory samples and blood largely depends on compliance with generally accepted rules for their collection, storage and transportation. Identification is based on determining the nutritional requirements of pathogens and the results of biochemical tests. To identify all of these microorganisms, commercial biochemical panels and reagent kits have been developed; automated microbiological analyzers can be used, which reduce the labor intensity of cultural research.
If you suspect CAP caused by S. aureus, it is important not only to isolate and identify the pathogen, but also to determine its sensitivity to oxacillin. Despite the lack of documented evidence of detection of methicillin-resistant S. aureus In patients with CAP on the territory of the Russian Federation, the danger of their appearance and spread is quite real. Among the phenotypic methods for detecting methicillin resistance, the most commonly used are disk diffusion testing with a disk containing 30 μg of cefoxitin or 1 mg of oxacillin, or screening on Mueller-Hinton agar supplemented with 4% NaCl and oxacillin at a concentration of 6 mg/l. To confirm infection with methicillin-resistant S. aureus commercial test systems based on identifying the gene in clinical material have been developed mecA by PCR method.
6.3. Diagnosis of pneumonia caused by Mycoplasma pneumoniae
The causative agent of respiratory mycoplasmosis is Mycoplasma pneumoniae- class representative Mollicutes, uniting wallless bacteria capable of autonomous existence, occupying an intermediate position between bacteria and viruses in terms of the level of structural organization.
Respiratory mycoplasmosis is a common anthropogenic disease. A feature of respiratory mycoplasmosis is the frequency of epidemics at intervals, according to various sources, varying from 3 to 7 years. The spread of infection is facilitated by the frequency and duration of contacts among persons staying in closed and semi-closed groups (military personnel, boarding schools), especially during their formation.
In 3 - 10% of cases of mycoplasma infection, pneumonia is diagnosed radiologically. For pneumonia caused M. pneumoniae, other bacterial or viral pathogens are usually not detected, but in rare cases they are also isolated S. pneumoniae. In 1 - 5% of cases of respiratory mycoplasmosis, hospitalization is required.
Mycoplasma pneumonia is accompanied by frequent, painful and prolonged cough with scanty viscous sputum, which is poorly evacuated, chest pain is noted, and bronchial obstruction may develop. Intoxication is mild. Physical changes in the lungs are absent or mild. The X-ray picture is very variable. In most cases, lesions of the interstitium are detected; in some patients, pneumonia occurs as a focal or segmental type, sometimes inflammatory changes are of a mixed nature. The symptoms of pulmonary failure are not typical for mycoplasma pneumonia. Mycoplasma pneumonia usually has a favorable course, in rare cases the course is very severe.
Diagnosis of mycoplasma pneumonia only on the basis of clinical or radiological data is impossible, since it does not have pathognomonic features. The main role in confirming the mycoplasma etiology of pneumonia is given to laboratory etiological diagnosis. For the etiological diagnosis of mycoplasma pneumonia, the following is used:
DNA detection M. pneumoniae polymerase chain reaction (PCR), the main method for direct DNA detection M. pneumoniae Currently, the standard polymerase chain reaction (PCR) with detection by electrophoretic DNA separation is used, but PCR with real-time detection (RT-PCR) has the greatest specificity and sensitivity;
Detection of mycoplasma antigen in a direct immunofluorescence reaction (RIF);
Serological studies to detect specific IgM and IgG antibodies to M. pneumoniae in blood serum using enzyme-linked immunosorbent assay (ELISA).
Mycoplasma pneumoniae refers to difficult-to-cultivate microorganisms; The isolation process takes 3 to 5 weeks, so the culture method cannot be recommended for use by diagnostic laboratories.
For the purpose of rapid etiological diagnosis of pneumonia, it is recommended to use PCR in the study of biological material obtained from the lower respiratory tract (sputum during deep coughing, aspirates from the trachea, sputum obtained as a result of induction through inhalation of hypertonic sodium chloride solution, bronchoalveolar lavage fluid (BAL) obtained using fiberoptic bronchoscopy).
If a positive PCR result is obtained when examining biological material obtained from the lower respiratory tract, the etiology of pneumonia is considered established. If it is impossible to obtain biological material from the lower respiratory tract for PCR, it is permissible to use smears from the upper respiratory tract (a combined smear from the nasopharynx and posterior pharyngeal wall), and if a positive result is obtained, the etiology of pneumonia should be considered presumptively established. However, obtaining a negative PCR result when examining smears from the upper respiratory tract cannot indicate the absence of mycoplasma infection. In this case, serological diagnosis is recommended, taking into account the totality of results for the detection of specific antibodies of the IgM and IgG classes in paired sera examined simultaneously.
For the purpose of retrospective diagnosis, when the patient is already in the convalescent stage, it is necessary to use serological studies.
The primary immune response is characterized by the synthesis of IgM antibodies 1 to 3 weeks after infection, the detection of which indicates the acute phase of the infection. Immunoglobulins of class G appear by the end of 3 - 4 weeks. The diagnosis of mycoplasma respiratory infection is confirmed by a 4-fold seroconversion of specific antibodies in paired blood sera.
Direct detection of antigens M. pneumoniae in various biosubstrates (smears from the nasopharynx, lavage fluid, biopsies) obtained from patients with respiratory pathology, until now in some diagnostic laboratories they are carried out using RIF. This method, combined with the detection of specific antibodies to mycoplasma in ELISA, makes it possible to confirm the disease caused by Mycoplasma pneumoniae. It should be borne in mind that humoral antibodies persist for several years.
For a reliable and definitive etiological diagnosis of mycoplasma pneumonia, taking into account the possibility of persistence of this pathogen in the human body without pronounced clinical manifestations, additional confirmation of the established diagnosis by any of the methods listed above is recommended.
6.4. Diagnosis of pneumonia caused by Chlamydophila pneumoniae
C. pneumoniae causes pneumonia of varying severity, long-term bronchitis, pharyngitis, and sinusitis. Pneumonia caused by C. pneumoniae usually has a favorable course, in rare cases the course is very severe.
A mixed infection, for example, a combination with pneumococcus or the presence of severe concomitant diseases, especially in older people, complicates the course of the disease and increases the risk of death. Often the infection is asymptomatic.
All ages are at risk, but the incidence of chlamydial pneumonia is higher in school-age children. The incidence among men is higher than among women. Epidemic outbreaks occur every 4 - 10 years. Epidemiological outbreaks in isolated and semi-isolated groups, cases of intra-family transmission of chlamydial infection are described.
None of the currently known methods for diagnosing chlamydial pneumonia provides 100% reliability in identifying the pathogen, which dictates the need to combine at least two methods.
Microbiological isolation C. pneumoniae has limited use due to the fact that it is a long and labor-intensive process, characterized by low sensitivity and is available only to specialized laboratories. However, if a viable pathogen is isolated, the diagnosis can be made with the greatest confidence without the need for confirmatory tests. Culture isolation indicates an active infectious process, since with persistent infection the pathogen becomes unculturable.
The most specific and sensitive method for identifying the pathogen is PCR diagnostics. High sensitivity and the absence of false-positive results can be ensured by using only licensed kits for effective DNA extraction from clinical material and modern-generation PCR kits based on real-time PCR (RT-PCR). The method does not make it possible to differentiate acute and chronic infection.
For the purpose of rapid etiological diagnosis of pneumonia, it is recommended to use PCR in the study of biological material obtained from the lower respiratory tract (sputum during deep coughing, aspirates from the trachea, sputum obtained as a result of induction through inhalation of hypertonic sodium chloride solution, bronchoalveolar lavage fluid (BAL) obtained using fiberoptic bronchoscopy). Serological tests are used for retrospective diagnosis and retrospective analysis of the nature of epidemic outbreaks.
If a positive PCR result is obtained when examining biological material obtained from the lower respiratory tract, the etiology of pneumonia is considered established. However, in cases of pneumonia caused Chlamydophila (Chlamydia) pneumoniae, cough is often nonproductive, in such cases, PCR is recommended to use swabs from the upper respiratory tract (a combined swab from the nasopharynx and posterior pharyngeal wall), and if a positive result is obtained, the etiology of pneumonia should be considered presumptive.
If you receive a negative PCR result when examining smears from the upper respiratory tract if you suspect an infection caused by C. pneumoniae, based on epidemiological or clinical data, serological diagnosis is recommended, taking into account the totality of results for the detection of specific antibodies of the IgM and IgG classes in paired sera examined simultaneously.
For the purpose of retrospective diagnosis, when the patient is in the convalescent stage, it is necessary to use serological tests.
Currently, to detect specific IgM and IgG antibodies to C. pneumoniae use the enzyme-linked immunosorbent assay (ELISA) or immunofluorescence reaction (RIF). Serological criteria for acute C. pneumoniae- infections: a 4-fold increase in titers of IgG antibodies in paired sera or a single detection of IgM antibodies in a titer of ≥ 1:16.
For a reliable and definitive etiological diagnosis of chlamydial pneumonia, taking into account the possibility of persistence of this pathogen in the human body without pronounced clinical manifestations, additional confirmation of the established diagnosis by any of the above methods is recommended.
6.5. Diagnosis of pneumonia caused by Legionella pneumonia
Due to the similarity of clinical manifestations and symptoms of legionellosis and pneumococcal pneumonia, fast and effective laboratory diagnosis becomes crucial for choosing tactics for etiotropic treatment of patients. In 1999, WHO and in 2002, the European Legionellosis Working Group adopted standards as diagnostic criteria, according to which the diagnosis of legionellosis in the case of acute lower respiratory tract infection (clinically and radiologically confirmed) is considered established:
1) when isolating a Legionella culture from the respiratory tract or lung tissue;
2) with a 4-fold or more increase in the titer of specific antibodies to Legionella pneumophila serogroup 1 in the indirect immunofluorescence reaction;
3) when determining a soluble antigen Legionella pneumophila serogroup 1 in urine by enzyme-linked immunosorbent assay (ELISA) or immunochromatographic method (ICA).
In the absence of blood serum taken in the early stages of the disease, detection of a significantly high level of antibodies to Legionella pneumophila serogroup 1 (1:128 and higher) in single serum by indirect immunofluorescence allows the diagnosis of legionellosis to be considered presumptive. Results obtained from the detection of a pathogen or its DNA in respiratory secretions or lung tissue using direct immunofluorescence or PCR are interpreted in a similar way.
Points 2 and 3 of the laboratory diagnostic standards currently apply only to antibodies and antigen determined for Legionella pneumophila serogroups 1. For other serogroups Legionella pneumophila the results obtained from the determination of antibodies or the detection of antigen in urine allow only a presumptive diagnosis to be established. Isolation of a culture of the pathogen remains the only standard method that establishes a definitive diagnosis in case of infection caused by other serogroups Legionella pneumophila or species Legionella spp.. At the same time, it should be noted that more than 80% of sporadic and group cases of legionellosis are caused by strains Legionella pneumophila serogroup 1, and in epidemic outbreaks of community-acquired pneumonia the etiological significance of the strains L. pneumophila serogroup 1 confirmed in 96% of cases.
The main method of standards that currently allows for timely diagnosis and monitoring of Legionella infection is the determination of Legionella antigen in urine using an immunochromatic or enzyme-linked immunosorbent method. The method allows you to finally confirm the diagnosis within 1 - 2 hours. The superiority of this method over other methods included in the standard lies primarily in the timing of the study and the availability of clinical material.
The bacteriological method takes at least 4-5 days, and invasive procedures are required to obtain bronchoscopy and biopsy material, since the pathogen cannot always be isolated from sputum, especially after the start of etiotropic therapy. Detection of a diagnostic increase in antibody titers in the indirect immunofluorescence reaction is possible only in the 3rd week of the disease, when a course of antibiotic therapy has been administered and the outcome of the disease is usually clear. The need to study paired sera determines the retrospective nature of the diagnosis of legionellosis using this method.
The PCR method can be recommended primarily for the study of BAL fluid or biopsy for suspected Legionella pneumonia in immunocompromised patients. If in this category of patients the infection is caused by strains L. pneumophila, not belonging to serogroup 1, then this method is the only one that allows you to quickly establish a diagnosis.
6.6. Diagnosis of pneumonia caused by Pneumocystis jiroveci
Pneumocystosis, as a rule, occurs in the form of acute respiratory diseases, exacerbations of chronic bronchopulmonary diseases, obstructive bronchitis, laryngitis, as well as pneumonia with impaired gas exchange (interstitial pneumonia).
The typical radiological picture of Pneumocystis pneumonia is represented by bilateral hilar interstitial infiltration of the lung tissue with increasing intensity and a large volume of damage in direct proportion to the progression of the disease. Less common are single and multiple compactions of lung tissue, upper lobe infiltrates and pneumothorax. Pleurisy and enlarged intrathoracic lymph nodes are quite rare. In the absence of pathology on radiographs, high-resolution CT may detect ground-glass changes or cellular deformation of the lung pattern.
In adults, Pneumocystis pneumonia usually develops against the background of secondary immunodeficiency. The incubation period is short - from 2 to 5 days, the onset is acute. Pneumocystis pneumonia can develop in patients receiving immunosuppressive therapy (corticosteroids). With drug immunosuppression, this disease manifests itself against the background of a decrease in the dose of corticosteroids. The prodromal period usually lasts 1 - 2 weeks; in AIDS patients - 10 days.
Pneumocystis pneumonia in AIDS is usually characterized by a sluggish chronic process. Initially, auscultatory symptoms are not detected. Respiratory failure associated with a sharp disruption of pulmonary ventilation and gas exchange leads to death. Abscesses, spontaneous pneumothorax and exudative pleurisy are also possible.
Pneumocystosis in children usually develops at 4–6 months of life, when the newborn’s immune system has not yet fully formed. The most susceptible to this disease are premature infants, patients with rickets, with malnutrition and damage to the central nervous system.
In young children, pneumocystis occurs as classic interstitial pneumonia with clear stages of pathological processes.
Based on morphological changes during the manifest course of the disease, three stages of the affected lung are distinguished:
Edema (7 - 10 days);
Atelectatic (up to 4 weeks);
Emphysematous (its duration is variable).
Risk groups for infection Pneumocystis jiroveci are:
Premature children, weakened newborns and young children with hypogammaglobulinemia, malnutrition and rickets;
Patients with leukemia, cancer patients, organ recipients receiving immunosuppressants;
Patients with tuberculosis, cytomegaly and other infections;
HIV-infected.
The most universal method for identifying cysts, trophozoites and sporozoites is the Romanovsky-Giemsa method. Vital staining with neutral red also makes it possible to identify the pathogen in the active phase.
All of the listed staining methods require highly qualified researchers for accurate identification. Pneumocystis jiroveci; Moreover, these methods serve only for indication and are aimed at common fungal polysaccharides of the cyst shell.
The immunofluorescence method (IF) for identifying cysts and trophozoites using monoclonal or polyclonal antibodies in lavage fluid has higher specificity and sensitivity than histochemical staining of preparations.
An immunological method that detects specific antibodies of the IgG and IgM classes (ELISA) also plays a significant role in the diagnosis of pneumocystis, especially when diagnosing when it is impossible to take lavage fluid or sputum from the patient. Antibodies of class G among the healthy population are detected quite often (60 - 80%). Therefore, the study of antibodies should occur over time with mandatory titration of serum. Detection of a 4-fold increase in IgG and/or determination of IgM antibodies against Pneumocystis jiroveci speaks of an acute infectious process caused by this pathogen.
Polymerase chain reaction (PCR) is one of the highly sensitive diagnostic methods that allows the detection of single cells or DNA fragments of the pathogen Pneumocystis jiroveci in sputum or bronchoalveolar lavage.
6.7. Diagnosis of viral and viral-bacterial pneumonia
The viral or viral-bacterial etiology of pneumonia in adults can be suspected during an increase in the incidence of influenza and ARVI, as well as when group cases of the disease occur within a month after the formation of closed and semi-closed groups. The risk group for severe viral pneumonia includes people suffering from heart failure and chronic diseases of the bronchopulmonary system. Concomitant pathologies in severe influenza are also obesity, diabetes mellitus, and pregnancy, especially in the third trimester.
The main causative agents of viral and viral-bacterial pneumonia in immunocompetent adults are considered to be influenza A and B viruses, adenoviruses, PC virus, parainfluenza viruses; Metapneumovirus is less frequently detected. In adults with influenza, complications develop in 10-15% of cases, 80% of which are pneumonia.
It is important to diagnose viral infections during CAP in children, in the etiological structure of which viral infections play a significant role.
Modern methods of etiological diagnosis of acute viral respiratory tract infections are based primarily on: identifying RNA/DNA pathogens using nucleic acid amplification methods, in particular using the most widely used PCR; on the detection of antigens using immunochromatography (ICA), enzyme-linked immunosorbent assay (ELISA), immunofluorescence (RIF). Mainly for retrospective diagnostics, methods for detecting specific antibodies in blood serum (complement fixation reaction (CFR), neutralization reaction (PH), hemagglutination inhibition reaction (HIR), indirect hemagglutination reaction (IRHA), enzyme-linked immunosorbent assay (ELISA)) remain important. Cultivation is possible for influenza A and B viruses, respiratory syncytial virus, parainfluenza viruses types 1 - 3, human metapneumovirus and adenoviruses.
Culture studies are labor-intensive and time-consuming; in routine practice, they are used only for influenza monitoring, while the initial detection of positive samples is carried out by PCR, followed by isolation in culture.
Immunofluorescence reactions allow the detection of antigens of influenza viruses, respiratory syncytial virus, parainfluenza viruses 1 - 3 and adenoviruses. Material for studies using immunofluorescence must be collected no later than three days from the onset of a respiratory infection (in the acute phase of the disease, since the method is most effective when the intracellular content of viral antigens is highest), which makes this method uninformative for the etiological diagnosis of pneumonia. In addition, the method is characterized by subjectivity when interpreting the analysis results.
Serological tests detect antibodies to respiratory syncytial virus (PH, RSK, RNGA, ELISA), parainfluenza viruses 1 - 4 (RTGA, RSK, ELISA), adenoviruses (ELISA), rhinoviruses (RSK); The study is usually retrospective in nature. Compared to RSC, ELISA is more sensitive. When interpreting, the change in the titer of specific antibodies over time in paired sera (obtained with an interval of 2 weeks) is assessed, and their results largely depend on the state of the patient’s immune system.
Recognized evidence of primary viral pneumonia (or mixed viral-bacterial pneumonia) according to international criteria (ESCMID 2011, BTS, 2009 - 2011) is the detection of nucleic acids of an influenza virus or other respiratory virus by PCR. Most often, smears from the nasopharynx and from the back wall of the pharynx are used for diagnosis, while the greatest sensitivity due to the higher content of viruses in the test sample can be achieved with a combination of smears from both loci. For this purpose, smears are taken from the patient with two different probes from the mucous membrane of the lower nasal passage, and then from the back wall of the oropharynx, while tampons from both probes, after taking smears, are sequentially broken off into one tube.
However, in the case of influenza viruses replicating in lung tissue (A/H5N1, A/H1N1pdm2009) in the second week of pneumonia, the concentration of the virus in smears may no longer be sufficient for its detection, especially if the material is collected inadequately. In addition, in order to simultaneously detect both viral and bacterial agents, it is advisable to use material from the lower respiratory tract (sputum from deep coughing, sputum obtained as a result of induction by inhalation of hypertonic sodium chloride solution, tracheal aspirates, bronchoalveolar lavage fluid (BAL) obtained using fiberoptic bronchoscopy).
To identify the most significant pathogens of acute respiratory viral infections: influenza viruses A and B, PC virus, metapneumovirus, parainfluenza viruses 1 - 4, coronaviruses (229E, OS43, NL63, HKUI), rhinoviruses, adenoviruses (B, C, E), bocavirus, PCR reagent kits are available in formats with electrophoretic detection, fluorescence endpoint detection, and real-time detection of accumulation of amplification products (RT-PCR). Tests based on real-time PCR achieve the maximum level of specificity and sensitivity; tests with the simultaneous detection of several pathogens have an advantage. The use of specific conserved regions of the viral genome as targets results in high rates of diagnostic sensitivity and specificity of PCR, approaching 100%, compared to culture studies. When diagnosing influenza, it is possible to determine the subtype of influenza A viruses, including the highly pathogenic avian influenza virus A/H5N1 and the new pandemic variant A/H1N1pdm2009, the so-called swine influenza virus.
Polymerase chain reaction in the format of electrophoretic detection requires special measures to prevent contamination (false-positive results), achieved by carrying out special measures and observing special rules for organizing a laboratory in accordance with MU 1.3.2569-09 “Organization of the work of laboratories using nucleic acid amplification methods when working with material containing microorganisms of I - IV pathogenicity groups.”
The etiology of “pneumonia caused by the influenza virus” should be considered established if RNA of the influenza virus (or in combination with other viruses) is detected by PCR in the material of the lower respiratory tract with a negative result of a bacteriological blood test (or in the absence of DNA of bacterial pathogens of pneumonia in the blood according to PCR results, or when insignificant DNA concentrations are detected in the material of the lower respiratory tract in quantitative PCR). If it is impossible to obtain material from the lower respiratory tract, the influenza etiology of pneumonia can most likely be proven if influenza virus RNA is detected in smears from the nasopharynx and oropharynx.
The etiology of pneumonia caused by other respiratory viruses is considered established if the RNA/DNA PCR method detects one respiratory virus (or several viruses at the same time) in the material of the lower respiratory tract with a negative result of a bacteriological blood test (or in the absence of DNA of bacterial pathogens of pneumonia in the blood according to the results PCR, or when insignificant concentrations of DNA are detected in the material of the lower respiratory tract in quantitative PCR).
The viral etiology of pneumonia is considered presumptively established if the RNA/DNA PCR method detects one respiratory virus (or several viruses at the same time) in smears from the nasopharynx and oropharynx with a negative result of a bacteriological blood test (or in the absence of DNA of bacterial pathogens of pneumonia in the blood according to PCR results, or when insignificant concentrations of DNA are detected in the material of the lower respiratory tract in quantitative PCR), as well as if bacteriological studies have not been carried out.
The viral etiology of pneumonia is considered presumptively established if the RIF method detects antigens of one respiratory virus (or several viruses at the same time) with a negative result of a bacteriological blood test (or in the absence of DNA of bacterial pathogens of pneumonia in the blood according to PCR results, or if insignificant concentrations of DNA are detected in the material of the lower respiratory tract in quantitative PCR), as well as if bacteriological studies were not carried out.
The viral-bacterial etiology of pneumonia is considered established if the RNA/DNA PCR method detects one virus (or several viruses at the same time) in the material of the lower respiratory tract with a positive result of a bacteriological blood test (or detection of significant concentrations of DNA in the blood or in the material of the lower respiratory tract in quantitative PCR).
The viral-bacterial etiology of pneumonia is considered presumptively established if the RIF or ICA method detects antigens of one respiratory virus (or several viruses at the same time) with a positive result of a bacteriological blood test (or detection of significant concentrations of DNA in the blood or in the material of the lower respiratory tract in quantitative PCR).
The results of serological studies allow us to judge the presence or absence of a viral infection against which pneumonia developed.
6.8. Differential diagnosis between zoonotic lung diseases and tuberculosis
Differential diagnosis with zoonotic diseases that cause lung lesions (ornithosis, Q fever, tularemia, etc.) is carried out according to epidemiological indicators and in regions endemic for these pathogens in accordance with the sanitary rules “Prevention of ornithosis”, “Prevention of Q fever”, “Prevention of tularemia " Differential diagnosis with tuberculosis is also an important and necessary component of the examination of patients with severe pneumonia.
7. Algorithm for diagnosing community-acquired pneumonia
The algorithm for laboratory diagnosis of typical CAP (in patients with no severe immune disorders) is different for severe and non-severe pneumonia, for patients with severe immune disorders and children. Timely etiological diagnosis of CAP is especially important for severe pneumonia in patients hospitalized in the ICU.
In case of severe pneumonia, first of all, it is necessary to conduct a bacteriological study for pneumococcus and other bacterial etiological agents, taking into account the spectrum of their sensitivity to antibiotics, and also to exclude legionella etiology using a rapid test for determining the legionella antigen in the urine of patients. During an increase in the incidence of influenza and ARVI, the likelihood of severe pneumonia of a viral or viral-bacterial nature is quite high. In this case, the diagnostic algorithm for severe pneumonia should take into account the possibility of bacterial, viral or viral-bacterial etiology. Underestimation at the stage of laboratory diagnosis of any of the above-mentioned etiological variants of severe pneumonia in ICU patients can lead to fatal outcomes. Mortality in severe CAP can range from 25 to 50%.
The term “non-severe pneumonia” is used for pneumonia that is treated on an outpatient or inpatient basis, but does not require admission to the ICU. In the absence of adequate timely treatment, mild pneumonia can lead to serious complications and chronic diseases of the bronchopulmonary system. The mortality rate can range from 1 to 10%. Along with bacteriological testing for pneumococcus and other bacterial etiological agents, taking into account the spectrum of their sensitivity to antibiotics, the diagnosis of non-severe pneumonia should take into account the possibility of mycoplasma or chlamydial etiology. During an increase in the incidence of influenza and ARVI, there is a high probability of mild viral pneumonia, as well as mixed infections of the mentioned viruses with bacteria, chlamydia or mycoplasma.
Requires an expanded analysis of the etiological structure of CAP in patients with severe immune disorders (acquired immunodeficiency syndrome, other diseases or pathological conditions). In addition to bacteriological testing for pneumococcus, other bacterial etiological agents, taking into account the spectrum of their sensitivity to antibiotics, and legionella, for this group of patients there is a high probability of developing pneumonia caused by “opportunistic etiological agents”, primarily Pneumocystis jiroveci, as well as cytomegalovirus, fungi, herpes virus. Differential diagnosis with tuberculosis and other mycobacterioses is also an important and necessary component of the examination of patients with CAP who have severe immune disorders. To exclude legionella etiology of pneumonia in immunocompromised patients, bronchoalveolar lavage or biopsy is examined using a bacteriological method, or PCR for L. pneumophila serogroups 2 - 15 and Legionella spp..
With CAP in children, the polyetiology is most pronounced, which must be taken into account in the process of laboratory diagnosis. Along with bacteriological testing for pneumococcus and other bacterial etiological agents, taking into account the spectrum of their sensitivity to antibiotics, the diagnosis of pneumonia in children should take into account a wide range of respiratory viruses, and not only during epidemic increases in the incidence of influenza and ARVI (influenza viruses, RS virus, metapneumovirus, parainfluenza viruses, adenoviruses, coronaviruses, bocavirus, rhinoviruses), as well as the etiological role of mycoplasmas and chlamydia. With CAP in children, there is a high probability of a mixed bacterial-viral infection, including a mixed infection with chlamydia and mycoplasma.
8. Quality control of laboratory tests
An obligatory component of modern laboratory diagnostics is a quality system for laboratory research and ensuring its functioning. The quality system includes internal control at the stages of laboratory research and external control.
Internal quality control of microbiological studies is a set of measures and procedures carried out by the laboratory aimed at preventing the adverse effects of factors arising in the process of preparation, performance and evaluation of analysis results that can affect the reliability of the result.
Internal quality control includes:
1. Monitoring compliance with the requirements for the conditions of analysis: (laboratory premises, air environment, temperature conditions for incubation and storage, disinfection and sterilization regimes, etc.).
2. Perform the procedure for maintaining reference bacterial cultures.
3. Quality control of culture media.
4. Quality control of test systems and reagents.
5. Quality control of distilled water.
The structure of the organization of internal quality control, the frequency and frequency of procedures performed is established by the quality management system in force in the laboratory in accordance with GOST ISO/IEC 17025 and GOST R ISO 15189.
Documentation of the results of the control procedures carried out is carried out according to the forms approved by the current quality management system. Registration and storage of control results can be carried out on electronic media.
A mandatory section of internal quality control is a periodic, but at least once a year, analysis of the results of the control procedures performed, taking into account which the quality manual of the testing laboratory is adjusted.
Ensuring internal quality control of molecular genetic (PCR) studies is additionally carried out in accordance with MU 1.3.2569-09 “Organization of the work of laboratories using nucleic acid amplification methods when working with material containing microorganisms of I-IV pathogenicity groups.”
External quality control is carried out in accordance with the requirements of GOST ISO/IEC 17025 and GOST R ISO 15189 in the form of participation in interlaboratory comparative tests (ICT) and/or proficiency testing programs by indicators and with frequency in accordance with established requirements and the needs of laboratories.
9. Safety requirements
Studies of biological (clinical) material are carried out in accordance with current regulatory legal and methodological documents regarding work with microorganisms of III - IV and I - II pathogenicity groups, depending on the type of suspected pathogen.
Application
In order to determine the etiological agent(s) of lower respiratory tract infection when CAP is suspected, sputum obtained by deep coughing, sputum obtained by induction by inhalation of a sterile 5% sodium chloride solution through a nebulizer, sputum obtained by aspiration from the trachea using a surgical vacuum or electrical suction, bronchoalveolar lavage (BAL) obtained using fiberoptic bronchoscopy, as well as blood and pleural fluid.
If it is impossible to obtain material from the lower respiratory tract when testing for respiratory viruses, mycoplasma and chlamydia, it is permissible to use smears from the upper respiratory tract (from the lower nasal passage and from the back wall of the pharynx), which are taken from the patient as soon as possible from the onset of acute respiratory infections symptoms in one tube and tested as one sample.
In hospitalized patients, material for research should be collected as early as possible upon admission (no later than the second day), since at a later date the possibility of superinfection through contact with other patients cannot be excluded. Collection of biological material for bacteriological research should be carried out before prescribing antibiotics.
In case of death, post-mortem (autopsy) material is examined.
Rules for obtaining freely separated sputum for bacteriological and PCR studies
To collect sputum, use sterile, hermetically sealed plastic containers. Before collecting sputum, the patient should be asked to rinse their mouth thoroughly with boiled water. Sputum collection is carried out on an empty stomach or no earlier than 2 hours after eating.
The patient is asked to take several deep breaths, holding the breath for a few seconds, then exhale forcefully, which promotes a productive cough and clears the upper respiratory tract of mucus. The patient is then asked to cough well and collect secretions from the lower respiratory tract (not saliva!) into a sterile container. The volume of the sputum sample should be at least 3 ml for adults and about 1 ml for children.
Should be stored in the refrigerator at a temperature of 4 - 8 °C. The duration of storage of sputum at room temperature should not exceed 2 hours.
For PCR studies It is allowed to store a sputum sample for 1 day at a temperature of 2 to 8 ° C, for a longer period - at a temperature not higher than - 16 ° C.
Rules for obtaining venous blood for bacteriological research
To collect blood for the purpose of bacteriological research, commercial hermetically sealed glass vials or vials made of impact-resistant autoclavable plastic of two types (containing a nutrient medium for isolating aerobes and anaerobes) are used. Blood is drawn with a syringe; the blood is aseptically transferred into a bottle with a transport medium directly through a rubber stopper.
2 samples of venous blood are taken at an interval of 20 - 30 minutes from various peripheral veins - for example, the left and right ulnar vein. One sample will be placed in a bottle to isolate aerobes, the other to isolate anaerobes. The blood volume for each venipuncture should be at least 10 ml for adults and 3 ml for children.
Immediately before venipuncture, the skin at the venipuncture site is disinfected using circular movements from the center to the periphery twice with a 70% alcohol solution or 1 - 2% iodine solution. It is necessary to wait until the disinfectant has completely dried and carry out the manipulation without touching the skin treated area.
After venipuncture, the remaining iodine should be removed from the surface of the skin to avoid burns.
Until transportation from the purpose of bacteriological research the sample along with the direction is stored at room temperature (no more than 2 hours) or in a thermostat.
Rules for obtaining venous blood for PCR testing
Blood should be taken on an empty stomach or 3 hours after a meal from the cubital vein in a sitting position. Blood is drawn into tubes with an anticoagulant (EDTA).
Immediately before venipuncture, the skin at the venipuncture site is disinfected using circular movements from the center to the periphery twice with a 70% alcohol solution or 1 - 2% iodine solution. It is necessary to wait until the disinfectant has completely dried and carry out the manipulation without touching the skin treated area. After venipuncture, the remaining iodine should be removed from the surface of the skin to avoid burns.
After drawing blood, the tube should be gently turned upside down several times so that the blood in the tube is thoroughly mixed. Place the test tube in a rack.
Until transport to the laboratory for the purpose of PCR research the sample along with the direction is stored at a temperature of 20 - 25 ° C for 6 hours from the moment of receiving the material - for the quantitative determination of nucleic acids, and for 12 hours - for the qualitative determination of nucleic acids; at a temperature of 2 - 8 °C - no more than one day for qualitative and quantitative determination of DNA/RNA of infectious objects. Whole blood samples should not be frozen.
Rules for obtaining pleural fluid for bacteriological and PCR studies
Take the material into disposable, tightly screwed tubes with a volume of 10 - 15 ml.
Before the manipulation, the skin is disinfected with 70% ethyl alcohol, then with 1 - 2% iodine solution, excess iodine is removed with a gauze cloth moistened with 70% alcohol to avoid burns to the patient’s skin. Percutaneous aspiration is then performed to obtain a sample of pleural fluid using careful aseptic technique. The sample volume must be at least 5 ml. All air bubbles are removed from the syringe, after which the sample is immediately transferred to a sterile plastic container. The container is tightly closed with a lid.
Before transportation, the sample along with the direction for bacteriological examination stored in the refrigerator at a temperature of 4 - 8 ° C. The duration of storage of pleural fluid at room temperature should not exceed 2 hours.
For PCR studies It is allowed to store the sample for 1 day at a temperature from 2 to 8 °C, for a longer period - at a temperature not higher than -16 °C.
Bronchoscopy is performed under oxygen therapy (oxygen inhalation through nasal catheters, using a Venturi mask or a mask with a reservoir). If it is not possible to ensure sufficient oxygenation of the blood, bronchoscopy is performed under non-invasive ventilation. In patients on mechanical ventilation, the procedure is performed under general anesthesia in conditions of myoplegia through a respirator adapter equipped with a valve for a bronchoscope. Bronchoalveolar lavage is performed according to accepted rules. The fiberoptic bronchoscope is passed into the bronchus until it jams, after which a 0.9% sodium chloride solution heated to 37 °C is injected using disposable syringes, 8 portions of 20 ml each (150 - 160 ml). In order to prevent collapse of the alveoli, suction is carried out at 50 - 80 mm Hg. Art. This procedure allows you to obtain the required number of alveolar macrophages in which the causative agent of CAP multiplies.
During life lung tissue obtained by transbronchial biopsy using a bronchoscope, which allows identifying pneumocystis in 66 - 98%, however, this method of collecting material is not indicated for all patients. Obtaining material for research is also possible with an open lung biopsy or using percutaneous intrathoracic aspiration with a pulmonary needle in patients who are contraindicated for transbronchial biopsy with a progressive course of the disease. The method of open lung biopsy gives the best (100%) results and is equivalent in result to surgical intervention, while obtaining a sufficiently large volume of material for research and a false negative result is completely excluded.
Currently, clinics have begun to actively research bronchoalveolar lavage to identify cysts and trophozoites.
Posthumous material collected during the first day after the death of the patient, smears-imprints of the lung or smears from the foamy contents of the alveoli are prepared.
Rules for obtaining tracheal aspirate for PCR testing
The manipulation is carried out on an empty stomach after brushing the teeth and rinsing the mouth with water. The patient is asked to take several deep breaths, holding the breath for a few seconds, then exhale forcefully. This promotes a productive cough and clears the upper respiratory tract of mucus. After connecting the mucus extractor through an adapter tube to the suction, a catheter for collecting tracheal aspirate was inserted into the pharynx through the oral cavity. Due to irritation of the mucous membrane in the glottis area, a cough reflex is provoked and the tracheal contents are removed through a sterile catheter (size 6 or 7) using suction. The volume of tracheal aspirate should be at least 3 - 5 ml.
Rules for obtaining induced sputum for bacteriological and PCR studies
Before the procedure, patients receive salbutamol (children - 200 mcg) through a metered dose inhaler to prevent bronchospasm. Then, for 15 minutes, oxygen is supplied through a jet nebulizer (aerosol apparatus) at a rate of 5 l/min with 5 ml of a 5% sterile NaCl solution. After this, tapping is carried out on the anterior and posterior walls of the chest to stimulate the discharge of sputum.
The patient is then asked to cough well and collect secretions from the lower respiratory tract (not saliva!) into a sterile container. The volume of the sputum sample should be at least 3 ml for adults and about 1 ml for children.
If sputum is not coughed up, it is recommended to combine the procedure with subsequent tracheal aspirates to suction the contents from the trachea using standard suction using a sterile 6- or 7-gauge catheter.
Before transportation, the sample along with the direction for bacteriological examination stored in the refrigerator at a temperature of 4 - 8 ° C. The duration of storage of sputum at room temperature should not exceed 2 hours.
For PCR studies Storage is allowed for 1 day at a temperature from 2 to 8 °C, for a longer period - at a temperature not higher than -16 °C.
Rules for obtaining swabs from the upper respiratory tract for PCR testing
The material is taken after rinsing the mouth with boiled water at room temperature. If the nasal cavity is filled with mucus, it is recommended to blow your nose before the procedure. For 6 hours before the procedure, you should not use medications that irrigate the nasopharynx or oropharynx or medications for resorption in the mouth.
Smears from the patient are taken with two different probes, first from the mucous membrane of the lower nasal passage, and then from the oropharynx, while the ends of the probes with tampons after taking smears are sequentially placed in one test tube with a volume of 1.5 - 2.0 ml with 0.5 ml of transport medium.
Nasopharyngeal smears in children taken with a dry sterile nasopharyngeal velor swab on a plastic applicator. The probe is inserted with a slight movement along the outer wall of the nose to a depth of 2 - 3 cm to the inferior concha, slightly lowered downwards, inserted into the lower nasal passage under the inferior nasal concha, a rotational movement is made and removed along the outer wall of the nose. The total depth of insertion of the probe should be approximately half the distance from the nostril to the ear opening (3 - 4 cm for children and 5 - 6 cm for adults). After collecting the material, the end of the probe with a swab is lowered into a sterile disposable tube with a transport medium to the point of breakage, while the flexible part of the probe is rolled up into a spiral, then, covering the top of the tube with a lid, the handle of the probe is lowered down, achieving complete breaking off of the upper part of the probe. The test tube is hermetically sealed.
Nasopharyngeal smears in adults It is also permissible to take it with a dry sterile polystyrene probe with a viscose swab. The probe is inserted with a slight movement along the outer wall of the nose to a depth of 2-3 cm to the inferior concha, slightly lowered downwards, inserted into the lower nasal passage under the inferior nasal concha, a rotational movement is made and removed along the outer wall of the nose. The total depth of insertion of the probe should be approximately half the distance from the nostril to the ear opening (5 cm for adults). After collecting the material, the end of the probe with the swab is lowered to a depth of 1 cm into a sterile disposable tube with transport medium and broken off, holding the tube cap. The test tube is hermetically sealed.
Oropharyngeal swabs take a dry sterile polystyrene probe with a viscose swab using rotational movements from the surface of the tonsils, palatine arches and the posterior wall of the oropharynx, gently pressing the patient’s tongue with a spatula. After collecting the material, the working part of the probe with a swab is placed in a sterile disposable tube with a transport medium and a probe with a nasopharyngeal swab. The end of the probe with the swab (1 cm) is broken off, holding the lid of the test tube so that it allows the test tube to be tightly closed. Storage is allowed for three days at a temperature of 2 - 8 ° C, for a longer period - at a temperature not exceeding -16 ° C.
Rules for obtaining material for serological diagnostics
(Detection of specific antibodies)
For a serological study (determination of antibodies), two blood serum samples are required, the 1st sample is taken on the day of the initial diagnosis, the 2nd sample - 2 - 3 weeks after the first. Blood is taken from a vein in a volume of 3 - 4 ml, or from the third phalanx of the middle finger in a volume of 0.5 - 1.0 ml into a disposable plastic tube without an anticoagulant. Blood samples are left at room temperature for 30 minutes or placed in a thermostat at 37 °C for 15 minutes. After centrifugation (10 min at 3000 rpm), the serum is transferred into sterile tubes using a separate aerosol barrier tip for each sample. The shelf life of whole blood is no more than 6 hours; freezing is unacceptable. The shelf life of blood serum at room temperature is for 6 hours, at a temperature of 2 - 8 ° C - for 5 days, for a longer period - at a temperature not exceeding -16 ° C (multiple freezing/thawing is unacceptable).
Rules for obtaining autopsy material for PCR research
Post-mortem material is collected during the first day after the death of the patient using a sterile individual instrument (individually for each organ) from an area of damaged tissue with a volume of 1 - 3 cm 3, placed in disposable sterile plastic containers with a hermetically screwed lid, frozen and stored at a temperature not exceeding - 16 °C.
Rules for obtaining and transporting urine to determine the antigen of Legionella or pneumococcus
Urine samples for research in a volume of 5 - 10 ml are placed in standard plastic containers and stored at room temperature (15 - 30 ° C) for no more than 24 hours after collection before performing the reaction. If necessary, samples can be stored at 2 - 8 °C for up to 14 days or at -20 °C for a long time for initial or repeated testing. Boric acid can be used as a preservative. Before testing, refrigerated or frozen urine samples are tested for the presence of antigen after reaching room temperature.
Requirements for labeling material for laboratory research
The label of the test tubes (containers) with the material indicates: the name and surname of the person being examined, the date the material was taken, and the type of material.
The accompanying document (referral) to the material collected for research in the laboratory must indicate:
Name of the institution that sends the material for research and telephone number;
Last name and first name of the patient being examined;
Age;
Date of illness or contact with the patient;
Probable diagnosis, severity of the disease or reason for examination;
The severity of the disease;
Data on influenza vaccination in the current epidemic season (vaccinated / not vaccinated / no data);
Date and signature of the medical person.
Transporting material produced in thermal containers at the recommended storage temperature. Samples from each patient are additionally packaged in an individual sealed bag with absorbent material.
Processing of biological material before laboratory research
In the laboratory, before starting PCR studies of biological material with a viscous consistency (sputum, tracheal aspirates), it must be pretreated to reduce the viscosity, for example, using a drug like “Mucolysin”, according to the instructions. The autopsy material is subjected to homogenization, followed by the preparation of a 20% suspension using a buffer solution (saline sodium chloride solution or phosphate buffer).
References
1. Community-acquired pneumonia in adults: practical recommendations for diagnosis, treatment and prevention: A manual for doctors / A.G. Chuchalin, A.I. Sinopalnikov et al. M., 2010. 106 p.
2. Pneumonia /A.G. Chuchalin, A.I. Sinopalnikov, L.S. Strachunsky. M., 2006.
3. Standard of specialized medical care for severe pneumonia with complications: Appendix to the order of the Ministry of Health of Russia dated January 29, 2013 No. 741n.
4. Kozlov R.S. Pneumococci: lessons from the past - a look into the future. Smolensk, 2010.
5. SP 3.1.2.2626-10 “Prevention of legionellosis.”
6. SP 3.1.7.2811-10 “Prevention of coxiellosis (Q fever).”
7. SP 3.1.7.2642-10 “Prevention of tularemia.”
8. SP 3.1.7.2815-10 “Prevention of ornithosis.”
10. MU 3.1.2.3047-13 “Epidemiological surveillance of community-acquired pneumonia.”
11. MU 4.2.2039-05 “Technique for collecting and transporting biomaterials to microbiological laboratories” (approved by the Chief State Sanitary Doctor of the Russian Federation on December 23, 2005).
12. Guidelines of the Ministry of Health of the Russian Federation No. 99/168 “Organization of identifying patients with tuberculosis using radiation, clinical and microbiological methods.” 2000.
13. MU 1.3.2569-09 “Organization of the work of laboratories using nucleic acid amplification methods when working with material containing microorganisms of I - IV pathogenicity groups.”
14. MU 2.1.4.1057-01 “Organization of internal quality control of sanitary and microbiological studies of water.”
15. MR No. 01/14633-8-34 “Identification of bacterial antigen Legionella pneumophila serogroup 1 in clinical material using the immunochromatographic method” (approved by the Chief State Sanitary Doctor of the Russian Federation on December 9, 2008).
16. MUK 4.2.1890-04 “Determination of the sensitivity of microorganisms to antibacterial drugs” (approved by the Chief State Sanitary Doctor of the Russian Federation on March 4, 2004).
Pneumonia is called inflammation of the lungs. This is an acute infectious disease that can have a different nature: bacterial, viral, fungal, aspiration. The most common cause of the disease is the bacterium pneumococcus.
The main symptom of pneumonia is suffocation due to high body temperature. When inhaling, a person feels pain in the chest, coughs with copious amounts of purulent sputum, sweats, and shudders. In elderly people, bedridden patients, if a small part of the lungs is affected, the clinical picture may be blurred.
To make a diagnosis, you need to see a general practitioner (pediatrician) or pulmonologist. Diagnosis of pneumonia includes a chest x-ray, sputum examination, and blood test. In the prehospital period, complaints are collected and auscultation is carried out - listening to the lungs with a phonendoscope to determine noises and wheezing.
What symptoms can you use to recognize pneumonia?
The very first reason to suspect pneumonia is the clinical picture characteristic of the disease. The following symptoms can help recognize pneumonia:
- chills and increased sweating;
- fever, body temperature 38-40 degrees;
- suffocating cough;
- difficulty breathing, especially inhalation;
- chest pain;
- coughing up yellow, green, pink, red-brown sputum;
- paleness of lips, nails, fingers - a symptom of oxygen starvation;
- dyspnea;
- body aches;
- fatigue, loss of appetite.
Attention! Pneumonia can be cured if you go to the hospital promptly. If you notice symptoms of pneumonia, you should call a doctor at home or call an ambulance. The disease progresses rapidly. According to statistics, more than 50% of deaths from pneumonia occur in those who self-medicated or failed to recognize the disease in a timely manner. Often the diagnosis of pneumonia is made by a forensic expert.
Picture of the disease in adults, elderly and children
Manifestations of pneumonia may be atypical. Sometimes the disease is practically asymptomatic, without high fever and debilitating cough. However, 3 signs are always present:
- unmotivated weakness, drowsiness;
- mild fever - temperature 37-37.5;
- heavy sweating during sleep.
The latent inflammatory process is typical for people with weakened immune systems, bedridden patients, newborns and elderly people over 65 years of age. An older person may also complain of confusion. On average, children under 5 years of age and the elderly suffer from pneumonia 2-3 times more often than an adult in the prime of life. Diagnosing their disease is more difficult.
Diagnostics
Most often, pneumonia is preceded by an upper respiratory tract disease - acute respiratory infections, colds, flu, tracheitis and others. One disease smoothly flows into another, and even a doctor does not always manage to “catch” this moment. Therefore, it is important to come back for a consultation if treatment for a cold does not help and there is no improvement. To diagnose pneumonia, the following measures are initially carried out.
1. Taking an anamnesis. The patient is questioned about complaints, duration of illness, presence of concomitant diseases, and treatment received. The doctor asks a series of questions. “Do you start to hyperventilate or become out of breath when you rest? When did the cough first appear, and how has it changed over time? Do you cough up yellow, pink, or green mucus? When inhaling or exhaling, do you feel pain in the chest area or not?”
2. Listening to the lungs. The next step is auscultation. The doctor asks you to expose your upper body and breathe correctly and deeply. Using a phonendoscope, he will listen to how the lungs open and what sound is produced. If pneumonia is suspected, auscultation is carried out especially carefully, always from the front and back. The doctor can tap on the back and use a stopwatch to record the time of inhalation and exhalation. Inflammation of the lungs is indicated by shallow bronchial breathing and lag on the affected side. In addition, fine bubbling wheezing, noises, and grinding sounds are clearly audible. Tapping allows you to roughly determine the affected area - directly above it the echo is short and dull.
3. Diagnostic study. To confirm the diagnosis and determine the cause of pneumonia, the doctor will prescribe a series of tests. Detection of pneumonia may include a different set of tests.
Basic methods
The most accessible and reliable way to diagnose pneumonia is to conduct a chest x-ray or fluorography if the patient is over 15 years old. The image allows you to see foci of inflammation, which are called “darkenings”. In fact, they are white spots, because the image is a negative. Diagnostics is carried out in combination with laboratory and instrumental studies, which makes it possible to distinguish pneumonia from diseases with similar symptoms. The standard set of tests required to make a diagnosis includes:
- chest x-ray (fluorography);
- laboratory sputum analysis;
- general blood test.
Pneumonia is indicated by focal shadows in the image. In this case, deformation and strengthening of the vascular-pulmonary pattern are also observed. Spots can have different diameters: up to 3 mm are considered small-focal, 4-7 mm - medium-focal, 8-12 - large-focal. If a child suffers from pneumonia, then the lower sections are involved, and the size of the shadows is 1-2 mm.
Blood in pneumonia has the following deviations from the norm: the number of leukocytes, ESR, neutrophils increases, and a shift in the leukocyte formula to the left is observed. The sputum is inflammatory in nature - leukocytes appear in it. Kurshman spirals, epithelial cells, blood elements, Charcot-Leyden crystals, and alveolar macrophages are found in the secretions. Culture of sputum on nutrient media allows you to identify the pathogen and choose the right antibiotic for treatment.
Attention! In case of pneumonia, x-rays are taken at least twice: upon admission to the hospital and upon discharge.
Additional Methods
If complications of pneumonia are suspected (abscess, pulmonary edema, pleurisy, obstruction, etc.), additional tests and studies are prescribed. In addition, they are necessary if differential analysis is to be carried out. Pneumonia has similar clinical manifestations to a number of other diseases of the respiratory system. It is important to exclude tuberculosis and cancer.
1. CT, MRI (computed tomography, magnetic resonance imaging). Both studies provide three-dimensional images of the lungs and other organs. This allows you to detect various complications, neoplasms, and foreign objects. CT uses X-rays, which are more effective for the differential diagnosis of tuberculosis. MRI scanning is electromagnetic and better shows the condition of soft tissues, blood vessels, and joints. It may be prescribed to differentiate between pneumonia and tumor.
2. Bronchoscopy. An endoscopic research method that allows you to find out about the presence of tumors, inflammation, and foreign bodies in the bronchopulmonary system.
3. Microscopy of sputum smears. If an atypical pathogen or tuberculosis is suspected, smears are stained using the Ziehl-Neelsen method. To differentiate between pneumococci, streptococci, and staphylococci, Gram staining is performed.
4. Biochemical blood test, ultrasound of the heart. Prescribed to exclude complications from other organs.
In conclusion, the diagnosis of pneumonia should be based on the results of x-ray and laboratory tests.
Establishing the etiology of the disease is important. Pneumonia can be pneumococcal, streptococcal, staphylococcal, viral, aspiration (most often when foreign bodies are swallowed by children, vomit enters the respiratory tract).
Diagnostics also allows us to identify the degree and nature of lung damage: focal, lobar, lobar, segmental pneumonia. Treatment is prescribed according to the type of inflammation. The patient often requires hospitalization. Antibiotics for this diagnosis are administered intravenously or intramuscularly. A home treatment regimen is possible if the initial stage of inflammation is diagnosed, and the person takes full responsibility for health.
