Phage Biology

T-even phage are especially virulent and are often found among phage selected for therapeutic purposes. Much is known about the molecular aspects of T4, but practically nothing is known about its growth physiology in the wild. Half of the genes in T4 have an unknown function in laboratory conditions, but may have functionality in their natural environments. The purpose of this research is to expand the limited knowledge about phage infections in such conditions. An area in which T-even phage commonly exist is in the mammalian digestive system. The conditions here require anaerobic fermentation and/or respiration. This research will ultimately open a new door into anaerobic bacteriophage infections and possible therapeutic effects or alternative antibiotic uses in agricultural areas as well as human health. Attempting to reproduce the natural conditions of the gut will give a much more accurate determination of the true nature of many bacteriophage infections in vivo.

As a facultative aerobe Eschericia coli has the capability to switch its metabolism in anoxic conditions to anaerobic respiration using an alternative terminal electron acceptor such as KNO₃. The organism can then still use the electron transport system (ETS). Virtually no research has been done to see if phage infect anaerobically grown E. coli. This study compares the ability of T-even phage to infect three strains of E. coli grown in defined media emphasizing anaerobic respiration metabolism. After initial studies in a variety of phage host systems we have focused on T4, Rb33, and LZ4. T4 is the most extensively studied and is fully sequenced, but it has a somewhat limited host range. Rb33 and LZ4 were selected for their ability to infect a broad host range from the ECOR collection as well as other agricultural and clinical isolates including 0157:H7. The bacteria used were B-Drake, ECOR4 (a wild type strain from the ECOR collection), and the sequenced K12 strain MG1655 because they cover a wide spectrum of E. coli. Using a predicted multiplicity of infection (MOI) of 5 to infect at a cell density of about 3x10⁸ CFU ml^-1, we were able to show in many cases efficient infection of the host resulting in burst sizes ranging from 14-110 phage per cell. However, there are cases where phage infect aerobically but are unable to infect anaerobically such as Rb33 in ECOR4. Here we hypothesize under anaerobic conditions that surface receptors are different or altered because of growth physiology. Also, there has been extensive lysis inhibition, up to 27 hours, in anaerobic infections as supposed to aerobic. Total phage production and bacterial survivors were followed to document the ability of the virion to kill its host. The MOI calculated based on the killer titer (KT), or the fraction of cells actually killed, was relatively similar to the approximate MOI when there was a successful infection.

In this novel area of bacteriophage research we have shown that T4, Rb33, and LZ4 have the ability to infect a variety of hosts in an oxygen deprived environment with minimum nutrient availability. Clearly, this aspect of phage biology is important to push therapeutic usage and to understand true phage physiology in conditions that naturally occur.

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Last Updated: May 29, 2008

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