Phage Biology

Gautam Dutta, Naomi Hoyle, Matt Robison, Mike Dyen, Raul Raya, Burton Guttman, Andrew Brabban Elizabeth Kutter. The Evergreen State College. Olympia, WA. 98505

T-even phages are key components of many therapeutic cocktails used in the Republic of Georgia and elsewhere. Using T4-like phages isolated from sheep resistant to E. coli O157 and from pediatric diarrheal patients in Bangladesh, we are now studying phage infection in conditions reflecting the mammalian gut. Aerobic infections in nitrate-glycerol (NG) medium, where nitrate is the final electron acceptor, were used as controls for anaerobic respiration studies with T4-like phages. One of the phages, LZ4, showed normal patterns of rapid infection both aerobically in TSB and anaerobically in NG. However, aerobically in NG most cells remain uninfected until an hour after infection, when there is a sudden rapid killing of the bacteria, along with a large burst of phage (see fig.); this was seen with E. coli B, K12 and ECOR4. The most likely explanation is that under these conditions the receptor used by LZ4 is present at too low a level to allow efficient infection, but the newly-synthesized progeny phage from the few cells that were infected have some special property that allows them to now infect efficiently. This in turn led us to papers from the '50s and '60s showing that newly-made so-called "nascent phage" can infect in the absence of an otherwise-essential cofactor (Wollman & Stent 1952, BBA 9:538; Anderson, 1948, J. Bact. 55:637).

Relevant observations include:

The baseplate is assembled on the membrane, attached by 6 300-Å fibers, and remains attached throughout assembly and cell lysis (fig.: Simon 1969, Virology38: 285); gp12 is not involved.
Nascent phage carry a membrane fragment that is lost within 30 min, stabilized by chilling, and destroyed by lysozyme; cofactorless infection parallels this membrane presence (fig.: Brown & Anderson 1969, J. Virol. 4:94).
A predicted membrane-spanning domain 126 aa from gp 7's C-terminus may be involved in the membrane attachment. Gp7 helps initiate assembly and is large enough (1032aa) to both occupy its observed corner position and form this tether.

We are trying in various ways to explore whether this "nascent phage" phenomenon might explain our observations. If indeed newly-synthesized phage can often infect related bacteria even without the normal receptor, this would have profound implications for such applications as phage therapy, potentially helping explain the lack of widespread selection for phage-resistant mutants during the extensive Eastern European applications.

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Last Updated: May 29, 2008

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