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Comparison of nested, multiplex, qPCR; FISH; SeptiFast and blood culture methods in detection and identification of bacteria and fungi in blood of patients with sepsis.

Gosiewski T, Flis A, Sroka A, Kędzierska A, Pietrzyk A, Kędzierska J, Drwiła R, Bulanda M - BMC Microbiol. (2014)

Bottom Line: Blood samples were derived from adult patients with clinical symptoms of sepsis, according to SIRS criteria, hospitalized in the Intensive Care Unit.It was demonstrated that qPCR was significantly more likely to detect microorganisms than the remaining methods; qPCR confirmed the results obtained with the SF kit in all cases wherein bacteria were detected with simultaneous confirmation of Gram-typing.All data collected through the FISH method were corroborated by qPCR.

View Article: PubMed Central - PubMed

ABSTRACT

Background: Microbiological diagnosis of sepsis relies primarily on blood culture data. This study compares four diagnostic methods, i.e. those developed by us: nested, multiplex, qPCR (qPCR) and FISH with commercial methods: SeptiFast (Roche) (SF) and BacT/ALERT® 3D blood culture system (bioMérieux). Blood samples were derived from adult patients with clinical symptoms of sepsis, according to SIRS criteria, hospitalized in the Intensive Care Unit.

Results: Using qPCR, FISH, SF, and culture, microbial presence was found in 71.8%, 29.6%, 25.3%, and 36.6% of samples, respectively. It was demonstrated that qPCR was significantly more likely to detect microorganisms than the remaining methods; qPCR confirmed the results obtained with the SF kit in all cases wherein bacteria were detected with simultaneous confirmation of Gram-typing. All data collected through the FISH method were corroborated by qPCR.

Conclusions: The qPCR and FISH methods described in this study may constitute alternatives to blood culture and to the few existing commercial molecular assays since they enable the detection of the majority of microbial species, and the qPCR method allows their identification in a higher number of samples than the SF test. FISH made it possible to show the presence of microbes in a blood sample even before its culture.

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Picture from a fluorescence microscope, obtained with the use of FISH: (A)visibleK. pneumoniaebacteria (sample no. 67);(B)visibleStreptococcusspp. bacteria (sample no. 35);(C)visibleE. cloacaebacteria (sample no. 13). Magnification 1000 x.
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Fig1: Picture from a fluorescence microscope, obtained with the use of FISH: (A)visibleK. pneumoniaebacteria (sample no. 67);(B)visibleStreptococcusspp. bacteria (sample no. 35);(C)visibleE. cloacaebacteria (sample no. 13). Magnification 1000 x.

Mentions: Neither qPCR nor SF confirmed positive results of blood culture in the same 6 samples (Table 1). All results gathered with the use of FISH (Figure 1) were confirmed by qPCR (Table 1).Figure 1


Comparison of nested, multiplex, qPCR; FISH; SeptiFast and blood culture methods in detection and identification of bacteria and fungi in blood of patients with sepsis.

Gosiewski T, Flis A, Sroka A, Kędzierska A, Pietrzyk A, Kędzierska J, Drwiła R, Bulanda M - BMC Microbiol. (2014)

Picture from a fluorescence microscope, obtained with the use of FISH: (A)visibleK. pneumoniaebacteria (sample no. 67);(B)visibleStreptococcusspp. bacteria (sample no. 35);(C)visibleE. cloacaebacteria (sample no. 13). Magnification 1000 x.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4302608&req=5

Fig1: Picture from a fluorescence microscope, obtained with the use of FISH: (A)visibleK. pneumoniaebacteria (sample no. 67);(B)visibleStreptococcusspp. bacteria (sample no. 35);(C)visibleE. cloacaebacteria (sample no. 13). Magnification 1000 x.
Mentions: Neither qPCR nor SF confirmed positive results of blood culture in the same 6 samples (Table 1). All results gathered with the use of FISH (Figure 1) were confirmed by qPCR (Table 1).Figure 1

Bottom Line: Blood samples were derived from adult patients with clinical symptoms of sepsis, according to SIRS criteria, hospitalized in the Intensive Care Unit.It was demonstrated that qPCR was significantly more likely to detect microorganisms than the remaining methods; qPCR confirmed the results obtained with the SF kit in all cases wherein bacteria were detected with simultaneous confirmation of Gram-typing.All data collected through the FISH method were corroborated by qPCR.

View Article: PubMed Central - PubMed

ABSTRACT

Background: Microbiological diagnosis of sepsis relies primarily on blood culture data. This study compares four diagnostic methods, i.e. those developed by us: nested, multiplex, qPCR (qPCR) and FISH with commercial methods: SeptiFast (Roche) (SF) and BacT/ALERT® 3D blood culture system (bioMérieux). Blood samples were derived from adult patients with clinical symptoms of sepsis, according to SIRS criteria, hospitalized in the Intensive Care Unit.

Results: Using qPCR, FISH, SF, and culture, microbial presence was found in 71.8%, 29.6%, 25.3%, and 36.6% of samples, respectively. It was demonstrated that qPCR was significantly more likely to detect microorganisms than the remaining methods; qPCR confirmed the results obtained with the SF kit in all cases wherein bacteria were detected with simultaneous confirmation of Gram-typing. All data collected through the FISH method were corroborated by qPCR.

Conclusions: The qPCR and FISH methods described in this study may constitute alternatives to blood culture and to the few existing commercial molecular assays since they enable the detection of the majority of microbial species, and the qPCR method allows their identification in a higher number of samples than the SF test. FISH made it possible to show the presence of microbes in a blood sample even before its culture.

Show MeSH
Related in: MedlinePlus