A novel, multiplex, real-time PCR–based approach for the detection of the commonly occurring pathogenic fungi and bacteria

Table 1 Melting points of bacterial and fungal amplicons and probes

Microbial strains	T_mP (°C)		T_mA (°C)
Gram positive (G+)	Mean	SD	Mean	SD
Enterococcus faecalis	67.94	0.07	84.14	0.36
Enterococcus faecium	67.84	0.21	84.59	0.78
Listeria monocytogenes	67.80	0.19	86.01	0.36
Staphyloccus aureus	64.85	0.21	83.91	0.54
Staphyloccus epidermidis	64.50	0.30	83.60	0.36
Streptococcus pyogenes	46.54	0.56	84.38	0.78
Gram negative (G-)
Acinetobacter baumannii	66.09	0.15	82.90	0.16
Bacteroides fragilis	48.65	0.18	84.47	0.84
Enterobacter aerogenes	63.95	0.34	83.47	0.48
Enterobacter cloacae	64.98	0.09	84.38	0.24
Escherichia coli	64.69	0.44	84.74	0.54
Haemophilus influenzae	61.99	0.35	84.28	0.30
Klebsiella pneumoniae	65.13	0.23	84.57	0.20
Proteus vulgaris	64.58	0.18	82.87	0.24
Pseudomonas aeruginosa	53.32	0.33	83.00	0.34
Serratia marcescens	64.01	0.30	84.17	0.30
Stenotrophomonas maltophilia	58.10	0.07	84.42	0.15
Fungi
Candida albicans	-	-	87.1	0.33
Candida dubliniensis	-	-	85.5	0.50
Candida quillermondii	-	-	85.1	0.70
Candida krusei	-	-	89.8	0.02
Candida parapsilosis	-	-	85.4	0.88
Candida tropicalis	-	-	84.5	0.75
Aspergillus fumigatus	-	-	91.0	0.38

All the amplicons T_m were measured at the F1 channel (540 nm). The signal was generated by aspecific SybrGreen dye. The G + specific probes produced a signal at the F2 channel (640 nm) the G- probes at the F3 channel (705 nm). The signals were induced with the help of a special FRET technique.