Bacterial and Fungal Identification by Molecular Methods

The genes encoding the rRNA have been for many years, and still are, the primary targets of molecular taxonomic studies, with several other genomic regions playing a minor role.

Integration of nucleic acid sequencing into routine laboratory practice and use of the BLAST analysis significantly reduced the number of organisms that could not be identified by phenotypic methods. Using the BLAST analysis the turnaround time is shortened, and the results can be obtained faster. This method also enables the identification of unusual species. It offers the promise of being the most accurate method available and provides the highest sensitivity and specificity. 

Hy Laboratories Ltd. is using the above approach for Bacterial & Fungal identification.

Method:

• Extraction of genomic DNA from an isolated colony
• Amplifying the rRNA genes using proprietary primers
• Sequencing the PCR product/s.
• Identification to the genus or species level by comparing the sequence to GenBank public database (and other public or in-house databases) using BLAST Search.

Sample source:

• Isolated colony (on agar plates or liquid broth)
• Directly from specimen - swabs, tissues, food, liquids etc (applicable on a single or dominant strain contaminated specimen only)

 Advantages:

• It is widely considered to be the best advanced and accurate genotypic method for microbial identification
• More reliable and precise compared to error-prone conventional morphological and metabolic identification techniques
• Enables identification up to species level as well as strain identification (when available)
• Enables identification of bacteria or fungi directly from clinical specimens (no need of culturing)
• Eliminates the need for prolonged incubation (especially for fungi) since usually very small amounts of visible growth is needed
• No need of viable colonies
• Turn around time of two working days for most samples
• Eliminates the need to use any other test

In the few cases where 16S rDNA bacterial sequencing analyses can't differ between clinical isolates, we offer the following services:

• RAPD - Ranom Amplified Polymorphic DNA PCR
• MLST - Multi Locus SequenceTyping sequencing.
• PCR Ribotyping - based on polymorphism in the 16S-23S intergenic spacer region
• DNA fingerprinting - protocols by PCR (including antibiotic resistance/susceptibility)*
• HSP 65 sequencing
• PFGE - Pulsed Field Gel Electrophoresis by restriction enzyme digest of chromosomal DNA.

*For cases like molecular typing of methicillin-resistant Staphylococcus aureus (MRSA).