Further methods of microbial identification

Biochemical methods (e.g. API systems)

The use of miniaturized biochemical systems is a standard procedure in microbiological labs. Correct identification requires the availability of a pure culture and prior determination of the cultural characteristics (gram characteristics, catalase test, oxidase test, sporulation capacity, microscopic image).

ifp uses API identification systems (Analytical Profile Index), for instance, which cover relevant groups of bacteria and yeasts. These systems determine the usability of certain carbohydrates or the formation of specific metabolic products, the evaluation of which is used to create a numerical profile. The profile is then compared to profiles in databases, which ultimately allows for identification of the microorganism. Results are ready after 4 – 72 hours, depending on which system was used.

Sequencing (16S- or 18S- or ITS-rDNA)

This molecular biological method of identification is based on multiplying a specific highly variable section of the ribosomal DNA (rDNA) of the microorganism by means of PCR (polymerase chain reaction) and subsequent sequencing of the amplificate.

The following steps need to be performed for this:

• isolation of the genomic DNA from bacteria, yeasts or fungi (pure culture)
• running the PCR: 16S-rDNA (bacteria) or 18S- or ITS-rDNA (yeasts and fungi)
• purification of the PCR product
• sequencing the PCR product
• evaluating the sequence e.g. via NCBI BLAST

The result of the molecular biological identification will be ready after approx. 3 days.

Cultural methods (fungi)

Classic identification of fungi is based on macroscopic and microscopic analysis of pure cultures on suitable media. The morphological characteristics of the colonies and of the microscopic image are compared with data from the literature.