Complementation

To better understand effects of iron starvation on A. pleuropneumoniae and to identify new potential vaccine targets, we conducted transcript profiling using a DNA microarray containing all 2025 ORFs of the genome of A. pleuropneumoniae serotype 5b strain L20. Upon comparing growth of this pathogen in iron-sufficient versus iron-depleted medium, 210 genes were identified as being differentially expressed. Some genes (92) were identified as being up-regulated; many have confirmed or putative roles in iron acquisition, such as the genes coding for two TonB energy-transducing proteins and the hemoglobin receptor HgbA. Transcript profiling also led to identification of some new iron acquisition systems of A. pleuropneumoniae. Keywords: stress response The bacteria was grown in BHI broth with or without 50 ug/ml EDDHA to generate iron restriction. In the iron-restricted medium, 50 ug/ml FeCl3 was added to the medium 5 min. after the addition of EDDHA. RNA extractions were performed on three different days using the QIAGEN RNeasy Kit, and a total of four hybridization were performed. Data were acquired using a Perkin-Elmer ScanArray Express scanner, and were quantified, normalized and analysed using the SpotFinder, MIDAS and TMEV software from TIGR's TM4 Suite of Microarray Analysis softwares.