The biological signal molecule nitric oxide (NO) was found to induce biofilm dispersal across a range of bacterial species, which led to its consideration for therapeutic strategies to treat biofilms and biofilm-related infections. However, biofilms are often not completely dispersed after exposure to NO. To better understand this phenomenon, we investigated the response of biofilm cells to successive NO treatments. When biofilms were first pretreated with a low, noneffective dose of NO, a second dose of the signal molecule at a concentration usually capable of inducing dispersal did not have any effect. Amperometric analysis revealed that pretreated cells had enhanced NO-scavenging activity, and this effect was associated with the production of the flavohemoglobin Fhp. Further, quantitative real-time reverse transcription-PCR (qRT-PCR) analysis showed that expression increased by over 100-fold in NO-pretreated biofilms compared to untreated biofilms. Biofilms of mutant strains harboring mutations in or , encoding a NO-responsive regulator of, were not affected in their dispersal response after the initial pretreatment with NO. Overall, these results suggest that FhpR can sense NO to trigger production of the flavohemoglobin Fhp and inhibit subsequent dispersal responses to NO. Finally, the addition of imidazole, which can inhibit the NO dioxygenase activity of flavohemoglobin, attenuated the prevention of dispersal after NO pretreatment and improved the dispersal response in older, starved biofilms. This study clarifies the underlying mechanisms of impaired dispersal induced by repeated NO treatments and offers a new perspective for improving the use of NO in biofilm control strategies.

This paper presents a hybrid case-based reasoning system for on-line technical support of PC fault diagnosis. HyCase consists of a natural language (keyword) input and the graph-theoretic constraint-net.Natural language or keyword inputs are parsed and then generated into a constraint-net. The constraint-net is validated and its links rationalized and standardized to minimize ambiguity. Cases that partially match either the keywords or the constraint-net are ranked according to their matching scores based on four different preferences, and then retrieved and presented to the user. The case base, which was developed in Microsoft Access 2000, is updated by means of a keyword and constraint-net manager. HyCase was implemented on a PC with a Pentium III processor running at 500 MHz and 128 MB of SDRAM.Twenty typical queries from customers were tested on a collection of 174 cases based on different cut-off overall matching scores. The effectiveness of case retrieval was measured by the proportion of relevant cases retrieved from the case base (recall) and, of these, the proportion directly applicable to the problem at hand (precision). The accuracy of the natural language parser was ascertained to range between 62.5% and 87.7%, while a parsing accuracy of 60% is sufficient to ensure a reasonable recall and precision. A minimum overall matching score of about 0.5 ensures a precision of 0.60. The parsing time gets noticeably longer when there are more than 15 words. Except for minimum overall matching scores exceeding 0.1, the parsing time is largely independent of the minimum score.The merits and limitations of HyCase are also discussed.