Prostate tissues from patients with prostate cancer and benign prostatic hyperplasia (BPH) frequently contain histological inflammation, and a proportion of these patients show evidence of Propionibacterium acnes infection in the prostate gland. We developed a multicolor fluorescent in situ hybridization (FISH) assay targeting P. acnes 23S rRNA along with a 14-kb region of the P. acnes genome. This assay was used to analyze prostate tissues from patients with prostate cancer and BPH. P. acnes infection of the prostate gland was demonstrated in prostatic tissue in 5 of 10 randomly selected prostate cancer patients. FISH analysis and confocal laser microscopy imaging revealed intracellular localization and stromal biofilmlike aggregates as common forms of P. acnes infection in prostate tissues from both prostate cancer and BPH patients. A sequential analysis of prostate tissue from individual patients suggested that P. acnes can persist for up to 6 years in the prostate gland. These results indicate that P. acnes can establish a persistent infection in the prostate gland. Further study is needed to clarify the link between this bacterium and prostatic inflammation which may contribute to the development of BPH and prostate cancer.Clinical benign prostatic hyperplasia (BPH) occurs in half of all men by the age of 80 years (20), while prostate cancer is a leading cause of morbidity and death among men in the United States and Western Europe (13). Chronic or recurrent inflammation has been implicated in the development and progression of both BPH (20) and prostate cancer (9,24,25,30), and various degrees of histological inflammation are seen in the majority of prostate tissue specimens examined. The possibility of an infectious etiology for this prostatic inflammation has been raised in the context of both BPH (20) and prostate cancer, although the accumulation of a larger amount of evidence supports this hypothesis for the latter. For example, epidemiological studies have reported an increased risk of prostate cancer in patients with a history of sexually transmitted infections (29,31,32). The genetic signatures of bacteria were found in prostate tissues from 21 of 107 patients with prostate cancer (21). Moreover, a positive correlation between prostatic inflammation and the detection of bacterial 16S rRNA gene sequences in tissue specimens obtained by radical prostatectomy has been reported (16).Recently, a single bacterial species, Propionibacterium acnes, has been shown to infect a considerable proportion (35%) of prostate glands removed at radical prostatectomy (6). Moreover, the presence of P. acnes was strongly correlated with histological inflammation, suggesting that this bacterium might be linked to cancer development. The possible involvement of P. acnes in prostate pathology was further highlighted by the detection of P. acnes 16S rRNA gene sequences in BPH tissues, with a positive association between the detection of P. acnes DNA and subsequent prostate cancer development reported (1). We have developed ...

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Expression of the gene encoding bacterial hemoglobin (VHb) from Vitreoscilla has been previously used to improve recombinant cell growth and enhance product formation under microaerobic conditions. It is very likely that the properties of VHb are not optimized for foreign hosts; therefore, we used error-prone PCR to generate a number of randomly mutated vhb genes to be expressed and studied in Escherichia coli. In addition, the mutated VHb proteins also contained an extension of eight residues (MTMITPSF) at the amino terminus. VHb mutants were screened for improved growth properties under microaerobic conditions and 15 clones expressing mutated hemoglobin protein were selected for further characterization and cultivated in a microaerobic bioreactor to analyze the physiological effects of novel VHb proteins on cell growth. The expression of four VHb mutants, carried by pVM20, pVM50, pVM104, and pVM134, were able to enhance microaerobic growth of E. coli by approximately 22%, 155%, 50%, and 90%, respectively, with a concomitant decrease of acetate excretion into the culture medium. The vhb gene in pVM20 contains two mutations substituting residues Glu19(A17) and Glu137(H23) to Gly. pVM50 expresses a VHb protein carrying two mutations: His36(C1) to Arg36 and Gln66(E20) to Arg66. pVM104 and pVM134 express VHb proteins carrying the mutations Ala56(E10) to Gly and Ile24(B5) to Thr, respectively. Our experiments also indicate that the positive effects elicited by mutant VHb-expression from pVM20 and pVM50 are linked to the peptide tail. Removal of the N-terminal sequence reduced cell growth approximately 23% and 53%, respectively, relative to wild-type controls. These results clearly demonstrate that it is possible to obtain mutated VHb proteins with improved characteristics for improving microaerobic growth of E. coli by using combined mutation techniques, addition of a peptide tail, and random error-prone PCR.

In several organisms, expression of a gene encoding dimeric hemoglobin (VHb) from the obligate aerobic bacterium Vitreoscilla stercoraria has been shown to increase microaerobic cell growth and enhance oxygen-dependent cell metabolism. In an attempt to further improve these effects of VHb, a gene encoding two vhb genes connected by a short linker of six base pairs was constructed and expressed in Escherichia coli(double VHb). Escherichia coli cells expressing double VHb reached a cell density 19% higher than that of cells expressing native VHb. The protein production per cell remained constant since the increase in cell growth was accompanied by an increase in protein content by 16%. Investigation of ribosome and tRNA content revealed that cells expressing double VHb reached their maximal capacity of protein synthesis later during cultivation than cells expressing native VHb, and furthermore they reached considerably higher levels of ribosome and tRNA compared to that of the VHb-expressing cells.