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log 2 (ratio)<br />

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Maren Depke<br />

Results<br />

Pathogen Gene Expression Pr<strong>of</strong>iling<br />

For the new transcripts included in the 2.5 h internalization signature, 6 cluster groups were<br />

defined (Fig. R.5.37 A). Cluster group 1 contained 26 transcripts. Expression was increased in the<br />

2.5 h samples and still slightly stronger in the 6.5 h samples. The serum/CO 2 controls at both time<br />

points also exhibited increase <strong>of</strong> expression. Cluster group 2 harbored 32 transcripts which<br />

featured increase in expression at the 2.5 h and to a lesser extent at the 6.5 h internalization time<br />

points and in the later 6.5 h serum/CO 2 control whereas the 2.5 h serum/CO 2 control exhibited in<br />

average no change. Transcripts with decreased expression in the 2.5 h internalized and less<br />

strongly in the 6.5 h sample belonged to cluster group 3 which included 42 transcripts. The 2.5 h<br />

serum/CO 2 control exhibited no apparent trend <strong>of</strong> regulation whereas the 6.5 h control exhibited<br />

a decrease in expression. In cluster group 4, 26 transcripts were classified, which exhibited a<br />

decrease <strong>of</strong> expression in both internalization samples, a slight increase in the 2.5 h serum/CO 2<br />

controls and a decline <strong>of</strong> expression to the reference level in the 6.5 h serum/CO 2 controls.<br />

Internalization samples in cluster group 5 showed an increase <strong>of</strong> expression 2.5 h after start <strong>of</strong><br />

infection, which again declined in samples after 6.5 h. Contrarily, expression in the serum/CO 2<br />

controls was induced at the 2.5 h time point and further increased at the 6.5 h time point. This<br />

cluster contained 49 sequences. Finally, cluster group 6 contained 25 sequences which were<br />

increased in both internalized samples, and possessed a variable expression pattern in the 2.5 h<br />

serum/CO 2 control with increased, repressed and unchanged values, while the 6.5 h serum/CO 2<br />

control samples exhibited a trend <strong>of</strong> decreased expression.<br />

A<br />

B<br />

cluster group 1<br />

cluster group 2<br />

cluster group 1<br />

cluster group 2<br />

6<br />

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2.5 h<br />

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cluster group 3<br />

cluster group 4<br />

cluster group 3<br />

cluster group 4<br />

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cluster group 5<br />

cluster group 6<br />

cluster group 5<br />

cluster group 6<br />

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co.<br />

Fig. R.5.37: K-means clustering <strong>of</strong> gene expression data from newly identified transcripts, which exhibited differential expression in<br />

the 2.5 h internalized sample (A) or in the 6.5 h internalized sample (B) in comparison to the 1 h serum/CO 2 control with statistical<br />

testing and multiple testing correction (p* < 0.05) and a minimal absolute fold change cut<strong>of</strong>f <strong>of</strong> 2. For k-means clustering, the<br />

following parameters were applied: Transcripts should be assigned to the user-defined number <strong>of</strong> 6 groups and sorted <strong>by</strong> deviation<br />

using the metric type cosine correlation. No additional data preparation or statistical cuts were applied.<br />

The ratio data set (experimental condition normalized to the 1 h serum/CO 2 control) employed for clustering consisted <strong>of</strong> the five<br />

experimental conditions <strong>of</strong> non-adherent (1 h), internalized (2.5 h and 6.5 h), and serum/CO 2 control (2.5 h and 6.5 h) staphylococci.<br />

Although not being continuous data, values were depicted as line graphs instead <strong>of</strong> bar charts for better facility <strong>of</strong> inspection.<br />

(1 h n. ad. – non-adherent staphylococci after 1 h <strong>of</strong> infection; 2.5 h int. − 2.5 h internalization; 6.5 h int. − 6.5 h internalization;<br />

2.5 h co. − 2.5 h serum/CO 2 control; 6.5 h co. − 6.5 h serum/CO 2 control)<br />

Correspondingly, also for the new transcripts included in the 6.5 h internalization signature,<br />

6 cluster groups were defined (Fig. R.5.37 B). Here, cluster group 1 contained 19 transcripts with<br />

168

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