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Applied and Environmental Microbiology, July 2007, p. 4351-4353, Vol. 73, No. 13
0099-2240/07/$08.00+0     doi:10.1128/AEM.03001-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Sensitivity of Escherichia albertii, a Potential Food-Borne Pathogen, to Food Preservation Treatments{triangledown}

Manan Sharma,1* Kalmia E. Kniel,2 Alexandra Derevianko,2 Jason Ling,3,{dagger} and Arvind A. Bhagwat3

Food Technology and Safety Laboratory,1 Produce and Quality and Safety Laboratory, Henry A. Wallace Beltsville Agricultural Research Center, Agricultural Research Service, Building 201, 10300 Baltimore Ave., Beltsville, Maryland 20705,3 The Department of Animal and Food Sciences, College of Agricultural and Natural Resources, University of Delaware, Newark, Delaware 197162

Received 28 December 2006/ Accepted 16 April 2007


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ABSTRACT
 
Escherichia albertii is a potential food-borne pathogen because of its documented ability to cause diarrheal disease by producing attachment and effacement lesions. Its tolerances to heat (56°C), acid (pH 3.0), and pressure (500 MPa [5 min]) were evaluated and found to be significantly less than those of wild-type E. coli O157:H7.


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INTRODUCTION
 
Escherichia albertii is a potential food-borne pathogen that may contribute to the burden of food-borne illness in the United States. Previously classified as Hafnia alvei with the ability to produce intimin (2, 11), it was reported to cause diarrheal disease in six children with accompanying symptoms of vomiting, mild dehydration, fever, and abdominal distention (2, 3). Five isolates of H. alvei from six children suffering from diarrhea in Bangladesh were reclassified as E. albertii based on their biochemical properties, genetic homology to Escherichia coli and Shigella flexneri, and unique lipopolysaccharide elements (1, 9, 10, 12). E. albertii isolates produce intimin, a protein that allows enteropathogenic E. coli and enterohemorrhagic E. coli to form attachment and effacement lesions on human intestinal epithelial cells. Commercial systems have misidentified E. albertii as Yersinia ruckeri, Salmonella enterica serovar Enterica, H. alvei, or E. coli (1, 19). The lack of biochemical identification of these strains has limited investigation into the incidence of E. albertii in foods; however, an eae-positive isolate of H. alvei was isolated from minced meat in Norway (14), possibly indicating that this H. alvei strain was actually E. albertii. E. albertii could possibly contribute to the estimated 62,000,000 cases of food-borne illnesses and 3,200 deaths in the United States that have an unknown etiological origin (15). The potential virulence of E. albertii and its possible presence in several foodstuffs led to our evaluation of its tolerance to interventions used in the food industry.

E. albertii 9194, 10457, 10790, 12502, and 19982 and the closely related Shigella boydii 12032 have been described previously (11); E. coli O157:H7 strain 52 (ATCC 43895; nalidixic acid and rifampin resistant) and an rpoS-deficient mutant of strain 52 (strain 55; rpoS::pRR10), along with S. flexneri, were also described previously (4, 6). Heat tolerance was measured by determining thermal decimal reduction times at 56°C (D56°C values) as described by Sharma and Beuchat (18); methods for determination of acid tolerance (pH 3.0 for 2 h at 37°C) were described by Bhagwat and Bhagwat (4). Stationary-phase cultures were treated in a PT-1 hydrostatic press (Avure Technologies, Kent, WA) at 500 MPa for 1 or 5 min at 22°C. Treated cells were enumerated on nonselective media. Among E. albertii strains, strain 19982 showed the most tolerance to heat and pressure treatments, while strain 9194 displayed the greatest acid tolerance.

D56°C values of E. albertii strains 19982 and 10457 were significantly (P ≤ 0.05) greater than those of strains 9194, 10790, and 12502 (Table 1). E. albertii strains are closely related to S. boydii 12032, as determined by multilocus sequence typing of housekeeping genes, differing in 0.72% of nucleotide sites examined (11). However, D56°C values for S. boydii 12032 were significantly higher than those of E. albertii.


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  		TABLE 1. D56°C values for E. albertii, E. coli O157:H7, and Shigella sp. strains

No E. albertii strain tested had a higher D56°C value than any wild-type E. coli O157:H7 strain examined, but all strains had D56°C values similar to those of the rpoS-deficient E. coli O157:H7 55 strain. Variability in wild-type rpoS alleles and other stress response genes of E. albertii strains may also account for differences in the D56°C values of these strains. In a previous study examining enterohemorrhagic E. coli, the acid, heat, and alkali tolerances of strains were influenced by the wild-type rpoS allele present in a strain (6).

E. albertii strains 9194, 10790, and 19982 displayed significantly greater acid tolerances than strains 10457 and 12502 (Fig. 1). Strain 9194 had acid tolerance similar to that of S. boydii 12032. No strain of E. albertii displayed more acid tolerance than wild-type E. coli O157:H7 strain 52. Of all the strains tested, rpoS-deficient E. coli O157:H7 strain 55 was the least acid tolerant. The diversity of acid tolerance responses in strains of E. albertii suggests that the acid response may be based on the functional heterogeneity of stress response genes. Experiments conducted in our work evaluated AR-1, the acid resistance mechanism induced when bacterial cells are oxidatively metabolizing carbon-containing nutrients (8, 13, 16). Previous work has shown that rpoS-deficient E. coli O157:H7 strains do not induce AR1 as effectively as wild-type strains and are more sensitive to oxidative acidic stress (5).


Figure 1
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  		FIG. 1. Acid tolerance, as measured by population reduction (log10 CFU/ml), of E. albertii, E. coli O157:H7, and Shigella spp. after 2 h at pH 3.0 for 37°C. Strains are represented as follows: E. albertii, 9194, 10457, 10790, 12502, and 19982; E. coli O157:H7, 52 and 55; S. boydii, 12032; and S. flexneri. Letters above error bars that are not the same indicate significant (P ≤ 0.05) differences in population reductions (open bars) of bacterial strains (n = 2).

E. albertii strains showed significantly lower pressure tolerances than wild-type E. coli O157:H7 strain 52 after 5 min (Fig. 2). E. albertii strain 19982 showed greater pressure tolerance than strain 12502 after 5 min. There were no significant differences in pressure tolerances between other E. albertii strains and strains 19982 and 12502. Significant differences were not observed in pressure tolerances of wild-type and rpoS-deficient strains of E. coli O157:H7, a result which is not in agreement with previous work which reported a reduction of >2 log10 CFU/ml for the rpoS-deficient compared to wild-type strain results (17). Our work shows a similar reduction in populations of wild-type and rpoS E. coli O157:H7 but lacks statistical significance. The pressure tolerance of S. flexneri and E. coli O157:H7 is in agreement with previous studies which found that S. flexneri and E. coli O157:H7 had the highest decimal reduction pressure value (7). Even after pressure treatment at 500 MPa for 10 min at 22°C, tolerances of E. albertii strains were less than those of wild-type E. coli O157:H7 and S. flexneri strains (data not shown).


Figure 2
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  		FIG. 2. Pressure tolerance, as measured by reductions (log10 CFU/ml) in populations of E. albertii, E. coli O157:H7, and Shigella spp. after treatment at 500 MPa and 22°C for 1 min and 5 min. Strains are represented as follows: E. albertii, 9194, 10457, 10790, 12502, and 19982; E. coli O157:H7, 52 and 55; S. boydii, 12032; and S. flexneri. For each of the indicated treatment times, different letters above error bars indicate significant (P ≤ 0.05) differences in population reductions (open bars) of bacterial strains treated with hydrostatic pressure (n = 2).

This study was the first to evaluate the tolerances of strains of E. albertii for food preservation processes used in industry. Strains of E. albertii were not more tolerant of these stresses than wild-type E. coli O157:H7, indicating that measures used to kill E. coli O157:H7 should be sufficient to inactivate E. albertii. Variations in tolerances of E. albertii strains to heat, acid, and pressure were observed. Culture methods to distinguish E. albertii strains from E. coli O157:H7 should be developed to evaluate the incidence of E. albertii in foods.


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ACKNOWLEDGMENTS
 
This study was supported by USDA-ARS CRIS Project 1265-41000-001-00D and by the overseas Industrial Attachment Program of the School of Life Sciences and Chemical Technology, Ngee Ann Polytechnic, Singapore (J.L.).

We thank the STEC Center (N01-AI-30058), National Food Safety and Toxicology Center, Michigan State University, for supplying strains. We acknowledge the contributions of Cheryl Mudd from FTSL, USDA-ARS, and Keith Lampel and Peter Feng from the FDA, College Park, MD, for their technical expertise.


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FOOTNOTES
 
* Corresponding author. Mailing address: Food Technology and Safety Laboratory, USDA-ARS, ANRI, BARC-EAST, 10300 Baltimore Ave., Bldg. 201, Beltsville, MD 20705. Phone: (301) 504-9198. Fax: (301) 504-8438. E-mail: manan.sharma{at}ars.usda.gov Back

{triangledown} Published ahead of print on 27 April 2007. Back

{dagger} Present address: School of Life Sciences and Chemical Technology, Ngee Ann Polytechnic, 535 Clement Road, Singapore 599489, Singapore. Back


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Applied and Environmental Microbiology, July 2007, p. 4351-4353, Vol. 73, No. 13
0099-2240/07/$08.00+0     doi:10.1128/AEM.03001-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.




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