Technology

Pathogen DNA Enrichment and Universal Detection/Identification for in-vitro Diagnostics

Several kits are available for the pre-analytic treatment of and pathogen DNA isolation from such diverse specimens like blood, CSF, BAL, synovial and pleural aspirates, swabs, tissues and biopsies. All the specimens contain human DNA which generally is in large excess to pathogen DNA. In unspecific reactions, PCR primers directed to conserved regions of the 16S and 18S rRNA genes, respectively, can interfere with human DNA during the PCR amplification reaction in a way that false-positive results may arise. In addition, primers can be diluted by the unspecific binding to human DNA with the result of false negative results. Molzym’s patented technology of pre-analytic removal of human DNA from clinical specimens, MolYsis, enables the enrichment of bacterial and fungal DNA and hence the increase in detection sensitivity of pathogens (click here for more detailed information).
Molzym’s kits, SepsiTest™, UMD-Universal and the automated systems SepsiTest-SelectNA™ and UMD-SelectNA™ rely on the universal 16S and 18S rRNA gene-targeted PCR or Real-Time PCR amplification and sequencing identification. The regions amplified contain highly variable sequences allowing identification of the pathogens to the species level. The universal nature of the PCR reaction allows the detection and identification of any pathogen present in a sample. Molzym has developed an online tool, SepsiTest™ BLAST which displays the identification results in a plain way. All components of the kits are manufactured under strict quality control thereby guaranteeing absence of contaminating microbial DNA in the reagents and consumables supplied with the kits.

 

Automated Solution for in-vitro Diagnostics

SepsiTest-SelectNA™ stands for simplicity and reliance in molecular analysis of blood samples for etiological agents of sepsis. The automated part, SelectNA™, comprises of a sophisticated, high precision instrument specially designed for the isolation of DNA from a broad-range of organisms. SelectNA™ guarantees a contained environment avoiding air-borne immissions of bacterial DNA during processing samples. The strong UV source moves in the interior thereby inactivating microbes and DNA.
The kit delivered, SepsiTest-SelectNA™ enables the automated extraction and purification of DNA from pathogens enriched from blood by the sample pre-treatment. Subsequently, the eluate is included in supplied PCR or Real-Time PCR assays for the universal detection of bacteria and fungi. First results of the infectious state of patients (presence/absence of bacteria and/or fungi) are gained after approximately 4h with only 75 min hands-on time (4 samples).

 

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Sequencing analysis of positive samples is the method of identifying an organism. Sequencing primers are supplied with each kit. This process follows usual standards of operation with Sanger-directed sequence analysis systems. The total processing time is approx. 260 min with 70 min hands-on time. Typically, sequencing reactions are performed over night with next morning identification results available.

 

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Clinical evaluation

Molzym is continuously performing clinical studies for the evaluation of SepsiTest™, UMD, SepsiTest-SelectNA™ and UMD-SelectNA™ kits using diverse specimens mainly from humans but also from animals. Molzym’s efforts are directed to comparing the kits’ performance with standard microbiological culturing methods. All kits are marked for the in-vitro diagnosis of infectious agents (98/79 EC).
Click here to download a list of organisms detectable, including representatives of genera identified by sequencing in the studies so far.
Latest number of species identified:
Bacteria (224)
Fungi (24)
Protist (1)
Date: October, 2012

 

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