Abstract
Sepsis is one of the leading causes of morbidity and mortality in hospitalized patients worldwide. Molecular technologies for rapid detection of microorganisms in patients with sepsis have only recently become available. LightCycler SeptiFast test Mgrade (Roche Diagnostics GmbH) is a multiplex PCR analysis able to detect DNA of the 25 most frequent pathogens in bloodstream infections. The time and labor saved while avoiding excessive laboratory manipulation is the rationale for selecting the automated MagNA Pure compact nucleic acid isolation kit-I (Roche Applied Science, GmbH) as an alternative to conventional SeptiFast extraction. For the purposes of this study, we evaluate extraction in order to demonstrate the feasibility of automation. Finally, a prospective observational study was done using 106 clinical samples obtained from 76 patients in our ICU. Both extraction methods were used in parallel to test the samples. When molecular detection test results using both manual and automated extraction were compared with the data from blood cultures obtained at the same time, the results show that SeptiFast with the alternative MagNA Pure compact extraction not only shortens the complete workflow to 3.57 hrs., but also increases sensitivity of the molecular assay for detecting infection as defined by positive blood culture confirmation.
Highlights
Sepsis is a clinical condition that is defined by objective signs establishing systemic inflammatory response syndrome (SIRS) together with the physician’s suspicion of infection
Of the different strategies for treating the septic condition that ranges from systemic inflammatory response syndrome (SIRS) to septic shock and multiorganic failure, appropriate antibiotic therapy and prompt initial treatment have the greatest impact on reducing mortality and morbidity associated with sepsis [2,3,4]
To evaluate the functionality of both methods we used 20 reference microbial strains obtained from the American Type Culture Collection (ATCC) (USA): Acinetobacter baumannii (ATCC 19686), Enterobacter cloacae (ATCC 13847), Serratia marcenscens (ATCC 14756), Stenotrophomonas maltophilia (ATCC 51331), Escherichia coli (ATCC 25922) Klebsiella pneumoniae (ATCC 70063), Proteus mirabilis (ATCC 12453), Pseudomonas aeruginosa (ATCC 27853), Staphylococcus aureus (ATCC 25923), Staphylococcus epidermidis (ATCC 12228), Streptococcus pneumoniae (ATCC 49619) Streptococcus agalactiae (ATCC 13813) Enterococcus faecalis (ATCC 29212) Enterococcus faecium (ATCC 35667), Candida albicans (ATCC 90028) Candida tropicalis (ATCC 750) Candida parapsilosis (ATCC 22019), Candida glabrata (ATCC 1526), Candida krusei (ATCC 6258), Aspergillus fumigatus (ATCC 36607)
Summary
Sepsis is a clinical condition that is defined by objective signs establishing systemic inflammatory response syndrome (SIRS) together with the physician’s suspicion of infection. Of the different strategies for treating the septic condition that ranges from systemic inflammatory response syndrome (SIRS) to septic shock and multiorganic failure, appropriate antibiotic therapy and prompt initial treatment have the greatest impact on reducing mortality and morbidity associated with sepsis [2,3,4]. LightCycler SeptiFast test Mgrade (Roche Diagnostics GmbH) is a multiplex real-time PCR based assay which is able, in about six hours, to detect the presence of DNA from 25 important bacterial and fungal species relevant for sepsis and nosocomial infection. The microorganisms covered by the test are the cause of approximately
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