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Figure 7 | BMC Biotechnology

Figure 7

From: Differences in virulence of pneumolysin and autolysin mutants constructed by insertion duplication mutagenesis and in-frame deletion in Streptococcus pneumoniae

Figure 7

Detection of TGR of WT plyand lytAgene from blood and liver of BALB/c mice with 3h-IDM culture of IDM mutants. (A-D) Cultural media used for IDM mutants growth. IDM-ply mutant was cultivated in plain BHI broth (A) and BHI supplemented with erm (B) whilst IDM-lytA mutant was grown in plain BHI broth (C) and BHI supplemented with cat (D). (I) Detection of the restored WT ply/lytA gene from blood and liver samples following IP injection of 3-h culture of IDM-ply/-lytA mutants at 10min, 1 day and 2 days. WT ply gene was amplified from samples by using the ply-P1/ply-P2 primer set for IDM-ply mutant grown from plain BHI (A,I) and BHI with erm (B,I). Full length of lytA gene was detected by the lytA-P1/lytA-P2 primer set for IDM-lytA mutant grown from BHI (C,I) and BHI with cat (D,I). (II) PCR analysis of IDM-ply/lytA gene from the same samples used in (I). Detection the genes of IDM-ply from IDM-ply mutant grown from BHI (A,II) and BHI with erm (B,II), and IDM–lytA gene from IDM-lytA mutant grown from BHI (C,II) and BHI with cat (D,II) was performed by ply-P1/pVA891-R and lytA-P1/pEVP3-R respectively. The lowest inocula of the 3-h IDM-ply cultures in BHI alone and BHI supplemented with erm for positive TGR of ply were approximately 104 CFU (A) and 103CFU (B) respectively whilst that of the IDM-lytA mutant grown in BHI and BHI supplemented with cat for positive TGR of lytA were 102 CFU (C) and 103 CFU (D) respectively. B, blood; L, liver; erm, erythromycin; cat, chloramphenicol.

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