Influence of macrolides on guanosine diphospho-D-mannose dehydrogenase activity in Pseudomonas biofilm

The formation of biofilm is regarded as a major cause of intractable infectious disease. Our studies were done to elucidate the action of a 14-membered-ring macrolide (erythromycin; EM) and a 16-membered-ring macrolide (midecamycin; MDM) on guanosine diphospho-d-mannose dehydrogenase (GDP-mannose dehydrogenase; GMD), one of the constituents of bacterial biofilm that is known to produce alginate. The mucoid type of Pseudomonas aeruginosa PT-1578 and the non-mucoid type of P. aeruginosa PAO1 were grown with nutrient-rich and nutrient-poor media. Comparative measurements were made of their GMD enzyme activities, with glucose-6-phosphate dehydrogenase (G6PDH), a cell membrane enzyme, used as a control. It was found that the GMD enzyme activity of mucoid type of Pseudomonas bacteria increased when they were grown on nutrient-poor media. Measurements were also made to determine the effects of EM and MDM against GMD and G6PDH enzyme activities. In media with either EM or MDM added, the production of G6PDH was not inhibited, irrespective of the concentration of EM or MDM. However, EM was effective against the production of GMD, showing a concentration-dependent effect. Scanning electron microscopy studies were also carried out to determine the effects of both macrolides on bacterial alginate production. It was found that reduction of alginate content occurred after the addition of EM. When environmental conditions for bacteria deteriorate, GMD enzyme is activated, production of alginate is initiated, and then biofilm is formed. Our results suggest that EM may have an inhibitory effect on the GMD production cycle, hence inhibiting the formation of biofilm. This may explain the differences in the clinical usefulness of 14-membered-and 16-membered-ring macrolides against biofilm disease.