Microbiologic examination of sputum

Microbiological examination is the most important link in the diagnostic search and verification of the causative agent of pneumonia. It involves not only isolating the causative agent, but also studying its properties, including sensitivity to various medications with bactericidal and bacteriostatic action.

For this purpose, the method of sputum seeding on various nutrient media is used. In a sputum sample delivered to the laboratory, purulent lumps are selected and carefully washed in a Petri dish with an isotopic solution of sodium chloride, which to a certain extent allows them to be freed from the microflora of the upper respiratory tract. The seeding of purulent lumps of sputum is carried out on various nutrient media, the composition of which is described in special manuals on microbiology. The media with the seedings are incubated at 37.5 ° C for 24 hours. Pure cultures are isolated from the grown colonies, they are identified by known microbiological methods and their sensitivity to antibiotics is determined.

To determine the quantitative content of microorganisms, the sputum is homogenized, mixed with nutrient broth, and successive tenfold dilutions are prepared from the mixture, which are inoculated onto Petri dishes with blood agar. After 24 hours of incubation at 37.5°C, the results are taken into account, counting colonies of the same type in appearance and taking into account the degree of dilution of the material. Smears are prepared from the colonies and examined under a microscope.

Interpretation of results

Interpretation of the results of microbiological examination of sputum is quite complex, which is explained by a number of factors. It has already been mentioned above about the constant seeding of bronchial contents with the microflora of the upper respiratory tract and oral cavity and the frequent presence in the normal tracheobronchial contents of healthy people of most of the most typical causative agents of respiratory diseases (pneumococci, streptococci, staphylococci, etc.). In this regard, the isolation in sputum during microbiological studies of an association of various microorganisms, most of which in this particular case are opportunistic, makes it extremely difficult to establish the causative agent of the disease. Therefore, to interpret the results of microbiological examination of sputum, it is necessary to take into account the quantitative predominance of a particular type of bacteria (more than 10 ⁶ -10 ⁷ m.c./ml), the appearance of certain microorganisms in the exacerbation phase and their disappearance during the remission period. It is very important to take into account the clinical picture of the disease.

The main and possible causative agents of various clinical forms of pneumonia

Clinical form of pneumonia	Main pathogens	Possible pathogens
Croupous	Pneumococci	Streptococci, Klebsiella
Post-influenza	Staphylococci, pneumococci, klebsiella	Haemophilus influenzae, streptococci
Abscessing	Staphylococci, bacteroids, mixed flora	Klebsiella, Pseudomonas aeruginosa
Aspiration	Bacteroides, anaerobic streptococci	Staphylococci, pneumococci
Postoperative	Staphylococci	Pneumococci, Klebsiella
Interstitial	Mycoplasmas	Pathogens of ornithosis, psittacosis
Secondary pneumonia in hospital patients without previous antibacterial therapy	Staphylococci, pneumococci, klebsiella, bacteroides	Escherichia coli, serratia, etc.
Secondary pneumonia developed against the background of antibacterial therapy	Facultative pathogenic microorganisms	Pseudomonas, Serratia, Klebsiella, Staphylococcus, Proteus, etc.
In patients with chronic bronchitis	Pneumococci, Haemophilus influenzae	Staphylococci, streptococci
In patients with alcoholism	Pneumococci, Haemophilus influenzae, Klebsiella	E. coli, protozoa
In acquired immunodeficiency syndrome	Pneumocystis, fungi	Cytomegaloviruses
In patients whose care is provided by outsiders	Pneumococci, staphylococci, hemophilic papilla	Klebsiella, Escherichia coli

When quantitatively assessing the results of the study of microbial contamination in patients with pneumonia, it is necessary to remember the extremely high sensitivity of this indicator to the prescription of antibiotics. Even short-term treatment with antibacterial drugs can lead to a sharp decrease in microbial contamination, which does not allow an adequate assessment of the results of the sputum study. Therefore, it is advisable to collect sputum before prescribing antibiotic treatment.

It should also be noted that special selective nutrient media are used to cultivate intracellular pathogens of pneumonia (mycoplasma, legionella, chlamydia, rickettsia). Routine microbiological testing using conventional nutrient media (agar-agar) never gives positive results. Therefore, the choice of specific methods of microbiological testing should be carried out with the participation of the attending physician, who is obliged to inform the laboratory physician of his suspicions regarding the possible role of intracellular pathogens in the occurrence of pneumonia in this patient.

It should be added that in real clinical practice, even a technically perfect microbiological examination of sputum allows identifying the pathogen in no more than 40-60% of cases. Therefore, other modern research methods can be used to verify the pathogen. The information content of a bacteriological study can be significantly increased by using tracheobronchial aspirate, fluid obtained during bronchoalveolar lavage (BAL), bronchoscopy, etc., rather than sputum, as the biological material being studied.

In addition, the method of immunofluorescence of various biological materials (bronchoscopy material, blood, pleural contents, etc.), PCR diagnostic methods, and the study of the level of specific antibodies in the blood serum can be used to identify pneumonia pathogens. Unfortunately, these diagnostic methods have not yet found wide clinical application and are currently used only in large specialized centers and laboratories.

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