Analysis of multidrug resistance in Streptococcus suis ATCC 700794 under tylosin stress

Figures & data

Figure 1. Growth curve of Streptococcus suis ATCC 700794 in the absence of tylosin and in the presence of 1/4 MIC of tylosin. Data are expressed as means ± standard deviations.

Figure 2. K-means clustering representation of 171 profiles of Streptococcus suis with up-regulations (a) and down-regulations (b). The magnitude of the percentage is represented by a color scale (top right) going from low (green) to high (red).

Table 1. Identification of Differentially Expressed Proteins.

Accession	Proteins	Fold change
G7SBK2	Elongation factor Ts	2.27
G7SMG3	Elongation factor G	3.67
Q9EZW2	Elongation factor Tu	1.74
G5L3D2	Putative histidine kinase of the competence regulon, ComD	0.74
G7SLJ0	Putative competence-damage inducible protein	0.56
A4VZB4	Protein GrpE	0.72

Table 2. Primers used for the quantitative qRT-PCR analysis.

Genes	Primer sequence
grpE	Forward: 5’AGCCGCAGCAGCACAACAAG 3’
	Reverse: 5’CTCGCCATCTACCACATCATCATCTG 3’
cinA	Forward: 5’GGTACAGACAGTAGCAGAGCGATTG 3’
	Reverse: 5’AAGTACGTTGTCCTCTGTTGATTCCG 3’
comD	Forward: 5’AGGTCGTTATAGTCGTCATGTTGAGTC 3’
	Reverse: 5’ GTTCGAATCGCATCAATGTCTTCCTC 3’
fus	Forward: 5’CTGGATGGAGCAAGAGCAAGAGC 3’
	Reverse: 5’ACACCTGATTGTGAGTCAAGAACGG 3’
tuf	Forward: 5’TTGGTGTGCTTCTTCGTGGTGTAC 3’
	Reverse: 5’TACGGAAGTAGAACTGTGGACGGTAG 3’
tsf	Forward: 5’ AGCAGAATTGGCAGCAGAAGGC 3’
	Reverse: 5’ GAAGCGTTCACTGAGTTCAAGTAAGC 3’
16S rRNA	Forward: 5’ GATATATGGAGGAACACCG 3’^[36]
	Reverse: 5’ GACCCAACACCTAGCACT 3’ ^[36]

Figure 3. The network of significantly differentially expressed proteins (ratio>1.2 or <0.8 (p-value<0.05)) was analyzed by STRING. Small nodes represent proteins of unknown 3D structure; large nodes represent proteins with some 3D structure information or predicted. Coloured nodes represent query proteins and first shell of interactors; white nodes represent second shell of interactors. The blue lines represent database evidence; the purple lines represent experimental evidence; yellow lines represent text mining evidence; the black lines represent coexpression evidence; and green lines represent neighbourhood evidence.

Table 3. MICs of resistance strain Streptococcus suis ATCC 700794.

Antibiotic	Sensitive strain (ATCC 700794)	Resistance strain (S-t-R 128)	CLSI
terramycin	0.5 μg/mL	128 μg/mL	S	2	R	8
ofloxacin	1 μg/mL	32 μg/mL	S	2	R	8
enrofloxacin	0.5 μg/mL	2 μg/mL	S	0.5	R	2
chlortetracycline	0.625 μg/mL	32 μg/mL	S	2	R	8
florfenicol	2 μg/mL	2 μg/mL	S	2	R	8
penicillin K	1 μg/mL	1 μg/mL	S	0.5	R	2

Figure 4. (a). Effect of 1/4 MIC of tylosin on mRNA expression of genes in Streptococcus suis ATCC 700794. Data are expressed as means ± standard deviations. The expression was normalized to 16S rRNA. Significantly different (*p < 0.05) compared to untreated control bacteria. (b). Effect on mRNA expression of genes in drug resistance level at 16 μg/mL of tylosin (S-t-R 16) Streptococcus suis ATCC 700794. Data are expressed as means ± standard deviations. The expression was normalized to 16S rRNA. Significantly different (*p < 0.05) compared to control bacteria. (c). Effect on mRNA expression of genes in drug resistance level at 128 μg/mL of tylosin (S-t-R 128) Streptococcus suis ATCC 700794. Significantly different (*p < 0.05) compared to untreated control bacteria. Data are expressed as means ± standard deviations. The expression was normalized to 16S rRNA. Significantly different (*p < 0.05) compared to control bacteria. (d). Effect on mRNA expression of fusA, tuf, tsf in Streptococcus suis ATCC 700794, drug resistance level at 16 μg/mL of tylosin (S-t-R 16) Streptococcus suis ATCC 700794 and drug resistance level at 128 μg/mL of tylosin (S-t-R 128) Streptococcus suis ATCC 700794. Data are expressed as means ± standard deviations. The expression was normalized to 16S rRNA. Significantly different (*p < 0.05) compared to control bacteria. (e). Effect on mRNA expression of comD, cinA, grpE in Streptococcus suis ATCC 700794, drug resistance level at 16 μg/mL of tylosin (S-t-R 16) Streptococcus suis ATCC 700794 and drug resistance level at 128 μg/mL of tylosin (S-t-R 128) Streptococcus suis ATCC 700794. Data are expressed as means ± standard deviations. The expression was normalized to 16S rRNA. Significantly different (*p < 0.05) compared to control bacteria.