Question

Compare and contrast classical and serological identification of bacterial, viral, fungal and parasitic human pathogens

Answer 1

Classical methods of bacteria identification look at the phenotypic features. For identify a particular organism different tests are done to finally come to a conclusion. Various techniques which look at the phenotypic features are Cultures , Microscopy, staining reactions , Macroscopic morphology of colonies formed on agar, antibiotic susceptibility test, Biochemical tests

Culture: differential media like Mac conkey agar, EMB agar help in identification of gram positve and gram negative strains. For example ; E coli form colonies with metallic green sheen on EMB agar. Mannitol salt agar which is a selective media promotes the growth of pathogenic staphylococci. It also helps to differentiate between pathogenic and non pathogenic strains of staphylococci. Pathogenic strain is able to ferment mannitol whereas non pathogenic is unable to do so.

staining reactions : It helps to differentiate between gram positive and gram negative organism. gram postive appear purple whereas gram negative appear pink under light microsocpe. Acid fast staining help in identification of Mycobacterium. This test is used in diagnosis of Tuberculosis. Malachite green is used to stain endospores. India Ink and crystal voilet is used to identify capsulated bacteria like klebsiella pneumoniae.

Colony morphology: Various parameters used to assess colony morphology are shape ,size, colour, margin and texture. Different form of colonies are punctiform, circular, filamentous, irregular, rhizoid, spindlle. Different bacteria have different forms which helps in their identification. For example Bacillus subtilis forms snowflake like colonies on nutrient poor agar.

Anitbiotic susceptibility test : Microbes are grown in agar containing antibiotic discs. If the bacteria is suceptible, growth will not take place, if its resistant then growth will take place. Different bacteria have different susceptibility and resistance profiles.

Biochemical test: IMViC test is used for identifying and distinguishing members of enterobacteriaceae. Urease test is used to identify Proteus, klebsiella pneumoniae

Disadvantages of classical methods are that all microorganisms are not culturable example Treponema pallidum. Some bacteria are fastidious in nature having complex nutritional requirement e.g Leptospira, Neisseria gonorrhoeae .

Serological methods eliminate such disadvantages as samples can be directly tested for microbes. These are easier and require less time than classic bacterial culture methods, also they are high throughput i.e multiple sample can be tested at once. In these methods antibodies formed in response to bacterial, viral, fungal pathogens are used to detect or identify microbes in sample.

serological test include precipitation test, agglutination test, western blots, immunofluoresence, radio immunoassay, Enzyme linked immunosorbent assay (ELISA).

microbes have antigens on their cell surface. If antibodies in serum are specific for that antigen, antigen antibody complex will form which can detected. In ELISA, these complexes are detected by an second antibody having labels. These labels can be either radioactive elements I¹²⁵ in case of radioimmunoassay ?or fluorophores in case of immunofluoresence . Either antigen or antibody is labelled.