ANNOTATED SEQUENCE RECORD
Genomic analysis of bacteriophage ESP2949-1, which is virulent
for Cronobacter sakazakii
Received: 22 July 2011 / Accepted: 11 October 2011 / Published online: 1 November 2011
Ó Springer-Verlag 2011
Abstract Virulent phage ESP2949-1, which was isolated
from sewage, has an icosahedral head, a contractile tail, and
a double-stranded DNA genome with a length of 49,116 bp
with 50.09% G?C content. Phage ESP2949-1 showed 3%
similarity to enterobacteria phage TLS. Bioinformatics
analysis of the phage genome revealed 43 putative open
reading frames (ORFs). Predicted protein products of the
ORFs were determined and described. Based on its mor-
phology, phage ESP2949-1 can be classiﬁed as a member of
the family Myoviridae. To our knowledge, this is the ﬁrst
report of the genomic sequence and characterization of
phage ESP2949-1 isolated from sewage.
Cronobacter sakazakii can be extremely harmful, especially
to infants with sepsis, meningitis, cerebritis and necrotizing
enterocolitis [4, 6, 8]. Infant or baby foods including infant
milk formula are often contaminated with C. sakazakii .
Kucerova et al. analyzed the C. sakazakii genome and
compared it to those of other Cronobacter spp. .
Recently, biocontrol methods for Cronobacter spp. used to
minimize infection risk have included biological treatment
such as lactic acid bacteria or bacteriophages, chemical
disinfectants, and physical treatments [10, 12, 16]. Bacte-
riophages (or phages) have been intensively studied and
have been used in a variety of practical applications such as
phage therapy , the detection of unknown bacteria 
and biocontrol of food-borne pathogens [6, 7]. More
recently, bacteriophages have been exploited in an attempt
to remove bacterial bioﬁlms . In this study, we report
the morphogenetic analysis and genome sequence bioin-
formatics analysis of phage ESP2949-1, which is virulent
for C. sakazakii and was newly isolated from sewage.
Phage ESP2949-1 was isolated from sewage samples.
C. sakazakii ATCC29544 was grown in Luria-Bertani (LB)
broth or agar supplemented with 10 mmol CaCl
37°C overnight in a shaking incubator. To isolate phage
ESP2949-1 of C. sakazakii, sewage samples were analyzed
by plaque assay [13, 18]. To conﬁrm the morphological
characteristics, puriﬁed phage particles were negatively
stained with 2% aqueous uranyl acetate (pH 4.5) on a
carbon-coated grid and examined by transmission electron
microscopy. Phage ESP2949-1 DNA was isolated from
polyethylene-glycol-precipitated phage particles by the
method of Lee et al., and bacteriophage DNA was
stored at -80°C.
The genome sequence was generated by ultra-high-
throughput GS FLX sequencing to 20-fold redundancy on
average. The nucleotide sequences were compared with those
of other genes in GenBank by the BLAST program (http://
www.ncbi.nlm.nih.gov/BLAST/). The open reading frames
(ORFs) were identiﬁed with the ORF Finder at the National
Center for Bioinformatics site (http://www.ncbi.nlm.nih.
gov/gorf.html). The molecular weight and isoelectric point
were calculated using the Compute pI/Mw program (http://
www.expasy.ch/tools/pi_tool.html).The promoter and tRNA
J.-H. Park and H. Chang contributed equally to this work.
Electronic supplementary material The online version of this
article (doi:10.1007/s00705-011-1147-0) contains supplementary
material, which is available to authorized users.
Y.-D. Lee Á J.-Y. Kim Á J.-H. Park (&)
Department of Food Science and Biotechnology,
Kyungwon University, Sungnam-Si, Kyunggi-Do, South Korea
Y.-D. Lee Á H. Chang (&)
School of Life Sciences and Biotechnology, Korea University,
Anam-Dong, Sungbuk-Gu, Seoul, South Korea
Arch Virol (2012) 157:199–202