Characterization and Genome Sequencing of a Novel Coliphage Isolated from Engineered <i>Escherichia coli</i>

Abstract

Objectives: To characterize morphological, physicochemical and genomic features of a novel virulent coliphage which was isolated from an engineered Escherichia coli culture and termed engineered E. coli phage (EEP). Methods and Results: Electron microscopy revealed that EEP has an icosahedral head (62 nm in diameter) and a long, flexible tail (138 nm in length). EEP was able to infect all 10 engineered E. coli strains kept in our laboratory, showing a strong ability to lyse engineered E. coli. Sequencing of the EEP genome revealed a double-stranded DNA (39.8 kb) with 54.72% GC content. Fifty-two open reading frames were predicted to be coding sequences, 18 of which were functionally defined and organized in a modular format, which includes modules for DNA replication, DNA packaging, structural proteins and host cell lysis. This phage could not be inactivated at 90° for 45 min and was resistant to ethanol and alkali treatment. EEP is assigned to the Siphoviridae family based on its morphological, genomic and physicochemical properties. Conclusions: A novel coliphage was isolated from engineered E. coli strains, and its morphological, genomic and physicochemical properties were characterized, which will improve our knowledge of bacteriophage diversity.

Resistance of foodborne pathogen coliphages to thermal and physicochemical treatments applied in food manufacture

“…In addition, Lee et al (2013) evaluated viability in LB broth, and, although coliphage ECP4 showed heat resistance at temperatures below 65°C, ECP4 was reduced by 6-log 10 PFU ml −1 after 10 min, and was not detected after 20 min at 70°C. Similar results were obtained by Li and coworkers at 63 and 72°C (Li et al, 2010). Moreover, several authors have found high resistance at 63°C and moderate resistance at 72°C for lactic acid bacteria (LAB) phages (Mercanti, Guglielmotti, Patrignani, Reinheimer, & Quiberoni, 2012;Pujato et al, 2014).…”

“…refrigeration -4°C-and abusive -50°C-temperatures) and remain biologically active (Coffey et al, 2011) either at refrigeration temperatures or when the cold chain is lost. On the contrary, other authors found that all phage activity was destroyed after 10 min at 60°C (Li et al, 2010). The slightly reduced activity observed in this work at 50°C may be due to disorganization and/or partial denaturation of phage receptors on the bacterial cell surface, thus hindering phage multiplication.…”

“…On the other hand, the six phages evaluated exhibited high resistance to basic pH; however, no viable particles were found at pH 12. Several authors evaluated phage viability at pH range from 2 to 12 and found similar results in coliphages (Coffey et al, 2011;Li et al, 2010), LAB phages (Mercanti et al, 2012) and mycophages (Endersen et al, 2013). Overall, the six phages evaluated seemed to be more resistant in an alkaline than in an acidic environment, in agreement with Feng, Ong, Hu, Tan, and Ng (2003) who found similar results for one (Qβ) of the two phages evaluated.…”

“…However, phage particles of DT2 (namely 3.1 × 10 2 PFU ml −1 on RSM and 1.35 × 10 3 PFU ml −1 on TMG; resulting in 4.7-and 4.1-log 10 reductions, respectively) and DT6 (1.25 × 10 3 PFU ml −1 on TMG; resulting in a 4.2-log 10 reduction) were detected after the treatment, suggesting a high resistance at this extreme temperature. Likewise, heating at 90°C was unable to complete coliphage EEP inactivation (Li et al, 2010). These results proved high thermal resistance of phages evaluated, remaining useful as biocontrol tools after physical treatments applied in food industry.…”

“…Although many authors have studied phage viability and activity at various physicochemical conditions such as thermal treatments (Lee, Kim, & Park, 2013;Li et al, 2010), varying pH (Dini & De Urraza, 2010;Kerby et al, 1949;Sharp, Hock, Taylor, Beard, & Beard, 1946) and cation (Adams, 1949) concentrations, only one report has exhaustively evaluated lytic coliphages in several physicochemical conditions found in food matrices (Coffey et al, 2011). Furthermore, suspension media such as reconstituted skim milk (RSM) and Buffer Trismagnesium-gelatin (TMG) have never been evaluated in order to assess their effect as potential phage protectants during thermal treatments.…”