Indeed, this effect was not observed with other classes of antibiotics [19–25]. In the present work and for the first time, an effect similar to that of beta-lactams is reported with tetracycline. Curiously, this antibiotic

induced larger plaques than beta-lactams. In the light of the foregoing discussion, this may be expected since it is well established that tetracycline can cause cell elongation and filamentation, so it is potentially able to increase phage production [34–36]. However, in the this website light of the results obtained, filamentation (or cell size elongation) seems not to be the only determinant of plaque size increase. In fact we observed that tetracycline induced the greatest increase in plaque size, but cells subjected to it were smaller than those incubated with the other antibiotics tested. Indeed, we found no correlation between plaque size and cell size. An unexpected

observation in this work was the conspicuous effect of glycerol in increasing phage plaque size and contrast. Glycerol produced a huge improvement in plaque observations when tetracycline was used. It allowed plaques to be observed that had very little contrast and were difficult to observe when tetracycline alone was used. This difficulty in observing the plaques obtained with tetracycline and no glycerol may explain why the effect of tetracycline, and even of other classes of antibiotics, has not been observed previously. learn more We conclude that glycerol plays a critical role in improving plaque observation. Glycerol may increase phage

diffusion in the medium Galeterone resulting in enhanced plaque size. Since it is a nonfermentative carbon source for these bacteria its presence will result in increased biomass or delay the onset of stationary phase. A plaque is unlikely to increase in size as the lawn cells enter late log growth stage [10, 37–39]. All in all, the influence of antibiotics on burst size, latent period and adsorption rate and the influence of glycerol on the diffusivity of phages in the medium and on bacterial growth seem to act together leading to a great increase in plaque size. Moreover, it was demonstrated here that antibiotics not only have the ability to increase phage plaques, they also do not suppress bacteriophage development at subminimal inhibitory concentrations (sub-MICs). In addition, the present results allow us to conclude that the new method (PAMA) can be applied to both Gram-negative and Gram-positive bacteria with lytic phages. The phages used represent the three families in the order Caudovirales, which include 96% of all observed phages [16]. Obviously, the antibiotic to be used in the PAMA, as well its concentration, have to be optimized for each bacterial host. Conclusion It is well known that some phages in the classical DLA technique produce plaques that are difficult or impossible to observe with the naked eye, leading to erroneous phage enumeration.