scholarly journals Biofilm Formation and Sanitizer Resistance of Escherichia coli O157:H7 Strains Isolated from “High Event Period” Meat Contamination†

2014 ◽  
Vol 77 (11) ◽  
pp. 1982-1987 ◽  
Author(s):  
RONG WANG ◽  
NORASAK KALCHAYANAND ◽  
DAVID A. KING ◽  
BRANDON E. LUEDTKE ◽  
JOSEPH M. BOSILEVAC ◽  
...  

In the meat industry, a “high event period” (HEP) is defined as a time period during which commercial meat plants experience a higher than usual rate of Escherichia coli O157:H7 contamination. Genetic analysis indicated that within a HEP, most of the E. coli O157:H7 strains belong to a singular dominant strain type. This was in disagreement with the current beef contamination model stating that contamination occurs when incoming pathogen load on animal hides, which consists of diverse strain types of E. coli O157:H7, exceeds the intervention capacity. Thus, we hypothesize that the HEP contamination may be due to certain in-plant colonized E. coli O157:H7 strains that are better able to survive sanitization through biofilm formation. To test our hypothesis, a collection of 45 E. coli O157:H7 strains isolated from HEP beef contamination incidents and a panel of 47 E. coli O157:H7 strains of diverse genetic backgrounds were compared for biofilm formation and sanitizer resistance. Biofilm formation was tested on 96-well polystyrene plates for 1 to 6 days. Biofilm cell survival and recovery growth after sanitization were compared between the two strain collections using common sanitizers, including quaternary ammonium chloride, chlorine, and sodium chlorite. No difference in “early stage” biofilms was observed between the two strain collections after incubation at 22 to 25°C for 1 or 2 days. However, the HEP strains demonstrated significantly higher potency of “mature” biofilm formation after incubation for 4 to 6 days. Biofilms of the HEP strains also exhibited significantly stronger resistance to sanitization. These data suggest that biofilm formation and sanitization resistance could have a role in HEP beef contamination by E. coli O157:H7, which highlights the importance of proper and complete sanitization of food contact surfaces and food processing equipment in commercial meat plants.

2016 ◽  
Vol 79 (11) ◽  
pp. 1875-1883 ◽  
Author(s):  
RONG WANG ◽  
BRANDON E. LUEDTKE ◽  
JOSEPH M. BOSILEVAC ◽  
JOHN W. SCHMIDT ◽  
NORASAK KALCHAYANAND ◽  
...  

ABSTRACT In the meat industry, a high-event period (HEP) is defined as a time period when beef processing establishments experience an increased occurrence of product contamination by Escherichia coli O157:H7. Our previous studies suggested that bacterial biofilm formation and sanitizer resistance might contribute to HEPs. We conducted the present study to further characterize E. coli O157:H7 strains isolated during HEPs for their potential to cause contamination and to investigate the genetic basis for their strong biofilm-forming ability and high sanitizer resistance. Our results show that, compared with the E. coli O157:H7 diversity control panel strains, the HEP strains had a significantly higher biofilm-forming ability on contact surfaces and a lower susceptibility to common sanitizers. No difference in the presence of disinfectant-resistant genes or the prevalence of antibiotic resistance was observed between the HEP and control strains. However, the HEP strains retained significantly higher copy numbers of the pO157 plasmid. A positive correlation was observed among a strain's high plasmid copy number, strong biofilm-forming ability, low sanitizer susceptibility, and high survival and recovery capability after sanitization, suggesting that these specific phenotypes could be either directly correlated to gene expression on the pO157 plasmid or indirectly regulated via chromosomal gene expression influenced by the presence of the plasmid. Our data highlight the potential risk of biofilm formation and sanitizer resistance in HEP contamination by E. coli O157:H7, and our results call for increased attention to proper and effective sanitization practices in meat processing facilities.


2006 ◽  
Vol 69 (8) ◽  
pp. 1802-1807 ◽  
Author(s):  
K. HARRIS ◽  
M. F. MILLER ◽  
G. H. LONERAGAN ◽  
M. M. BRASHEARS

A study was conducted to determine if acidified sodium chlorite (1,200 ppm) and acetic and lactic acids (2 and 4%) were effective in reducing foodborne pathogens in beef trim prior to grinding in a simulated processing environment. The reduction of Salmonella Typhimurium and Escherichia coli O157:H7 at high (4.0 log CFU/g) and low (1.0 log CFU/g) inoculation doses was evaluated at various processing steps, including the following: (i) in trim just after treatment application, (ii) in ground beef just after grinding, (iii) in ground beef 24 h after refrigerated storage, (iv) in ground beef 5 days after refrigerated storage, and (v) in ground beef 30 days after frozen storage. All antimicrobial treatments reduced the pathogens on the trim inoculated with the lower inoculation dose to nondetectable numbers in the trim and in the ground beef. There were significant reductions of both pathogens in the trim and in the ground beef inoculated with the high inoculation doses. On the trim itself, E. coli O157:H7 and Salmonella Typhimurium were reduced by 1.5 to 2.0 log cycles, with no differences among all treatments. In the ground beef, the organic acids were more effective in reducing both pathogens than the acidified sodium chlorite immediately after grinding, but after 1 day of storage, there were no differences among treatments. Overall, in the ground beef, there was a 2.5-log reduction of E. coli O157:H7 and a 1.5-log reduction of Salmonella Typhimurium that was sustained over time in refrigerated and frozen storage. Very few sensory differences between the control samples and the treated samples were detected by a consumer panel. Thus, antimicrobial treatments did not cause serious adverse sensory changes. Use of these antimicrobial treatments can be a promising intervention available to ground beef processors who currently have few interventions in their process.


2005 ◽  
Vol 68 (4) ◽  
pp. 660-665 ◽  
Author(s):  
S. B. O[apos]BRIEN ◽  
G. DUFFY ◽  
E. CARNEY ◽  
J. J. SHERIDAN ◽  
D. A. McDOWELL ◽  
...  

In this study, we investigated the prevalence and numbers of Escherichia coli O157 on bovine hides. Samples (n = 1,500) were collected over a 17-month period (30 samples per week) by sponge swabbing approximately 122-cm2 areas of the bovine rump of slaughtered cattle at an early stage of carcass processing (first legging). Sponge samples (n = 1,500) were stomached in buffered peptone water supplemented with novobiocin, directly plated on sorbitol MacConkey with Cefixime tellurite (SMAC-CT), enriched for 24 h, extracted by immunomagnetic separation, and plated onto SMAC-CT agar. Presumptive E. coli O157 colonies from SMAC-CT plates were confirmed by PCR for the presence of eaeA, hlyA, fliCh7, vt1, vt2, and portions of the rfb (O-antigen encoding) region of E. coli O157. Overall, E. coli O157 was recovered from 109 samples (7.3%) at concentrations ranging from less than 0.13 to 4.24 log CFU/100 cm2. PCR analysis revealed a wide diversity of genetic profiles among recovered isolates of verocytotoxigenic E. coli. Of the isolates recovered, 99 of 109 contained the attaching and effacing gene (eaeA) and the hemolysin gene (hlyA), and 78 of 109 had the flagellar H7 antigen–encoding gene (fliCh7). Only 6 of 109 isolates contained both verotoxin-producing genes (vt1 and vt2); 91 of 109 contained the vt2 gene only, whereas 1 of 109 contained the vt1 gene only. The remaining 11 of 109 contained neither vt1 nor vt2.


2012 ◽  
Vol 75 (11) ◽  
pp. 1960-1967 ◽  
Author(s):  
IFIGENIA GEORNARAS ◽  
HUA YANG ◽  
GALATIOS MOSCHONAS ◽  
MATTHEW C. NUNNELLY ◽  
KEITH E. BELK ◽  
...  

Studies were conducted to compare the decontamination efficacy of six chemical treatments against Escherichia coli O157:H7 and multidrug-resistant and antibiotic-susceptible Salmonella inoculated on beef trimmings. The inocula, comprising four-strain mixtures of rifampin-resistant E. coli O157:H7 and antibiotic-susceptible or multidrug-resistant (MDR and/or MDR-AmpC) Salmonella Newport and Salmonella Typhimurium, were inoculated (3 log CFU/cm2) separately onto samples (10 by 5 by 1 cm) derived from beef chuck rolls. Samples were left untreated (control), were immersed for 30 s in acidified sodium chlorite (0.1%, pH 2.5), peroxyacetic acid (0.02%, pH 3.8), sodium metasilicate (4%, pH 12.6), Bromitize Plus (0.0225% active bromine, pH 6.6), or AFTEC 3000 (pH 1.2), or were immersed for 5 s in SYNTRx 3300 (pH 1.0). Levels of surviving Salmonella on treated trimmings were not influenced by serotype or antibiotic resistance phenotype and were generally similar (P > 0.05) or lower (P < 0.05) than levels of surviving E. coli O157:H7 regardless of antimicrobial treatment. Overall, depending on chemical treatment (reductions within each chemical treatment were similar among all tested inocula), initial counts of E. coli O157:H7 (2.7 to 3.1 log CFU/cm2) were reduced (P < 0.05) by 0.2 to 1.4 log CFU/cm2. Similarly, initial counts of the tested Salmonella inocula (2.8 to 3.3 log CFU/cm2) were reduced (P < 0.05) by 0.4 to 1.4 (Salmonella Newport, antibiotic susceptible), 0.3 to 1.4 (Salmonella Newport, MDRAmpC), 0.2 to 1.5 (Salmonella Typhimurium, antibiotic susceptible), 0.4 to 1.3 (Salmonella Typhimurium, MDR), and 0.4 to 1.5 (Salmonella Typhimurium, MDR-AmpC) log CFU/cm2, depending on antimicrobial treatment. Reductions obtained with sodium metasilicate were 1.3 to 1.5 log CFU/cm2, regardless of inoculum, and reductions obtained with the five remaining antimicrobial treatments were 0.2 to 0.7 log CFU/cm2 (depending on treatment). Findings of this study should be useful to regulatory authorities and the meat industry as they consider Salmonella contamination on beef trimmings.


2005 ◽  
Vol 71 (1) ◽  
pp. 247-254 ◽  
Author(s):  
Jee-Hoon Ryu ◽  
Larry R. Beuchat

ABSTRACT The resistance of Escherichia coli O157:H7 strains ATCC 43895-, 43895-EPS (an exopolysaccharide [EPS]-overproducing mutant), and ATCC 43895+ (a curli-producing mutant) to chlorine, a sanitizer commonly used in the food industry, was studied. Planktonic cells of strains 43895-EPS and/or ATCC 43895+ grown under conditions supporting EPS and curli production, respectively, showed the highest resistance to chlorine, indicating that EPS and curli afford protection. Planktonic cells (ca. 9 log10 CFU/ml) of all strains, however, were killed within 10 min by treatment with 50 μg of chlorine/ml. Significantly lower numbers of strain 43895-EPS, compared to those of strain ATCC 43895-, attached to stainless steel coupons, but the growth rate of strain 43895-EPS on coupons was not significantly different from that of strain ATCC 43895-, indicating that EPS production did not affect cell growth during biofilm formation. Curli production did not affect the initial attachment of cells to coupons but did enhance biofilm production. The resistance of E. coli O157:H7 to chlorine increased significantly as cells formed biofilm on coupons; strain ATCC 43895+ was the most resistant. Population sizes of strains ATCC 43895+ and ATCC 43895- in biofilm formed at 12�C were not significantly different, but cells of strain ATCC 43895+ showed significantly higher resistance than did cells of strain ATCC 43895-. These observations support the hypothesis that the production of EPS and curli increase the resistance of E. coli O157:H7 to chlorine.


2004 ◽  
Vol 67 (10) ◽  
pp. 2099-2106 ◽  
Author(s):  
J. D. STOPFORTH ◽  
Y. YOON ◽  
K. E. BELK ◽  
J. A. SCANGA ◽  
P. A. KENDALL ◽  
...  

Samples (10 by 20 by 2.5 cm) of beef carcass tissue were inoculated (104 to 105 CFU/cm2) with Escherichia coli O157: H7 that was either non–acid habituated (prepared by incubating at 15°C for 48 h in inoculated filter-sterilized composite [1:1] of hot and cold water meat decontamination runoff fluids, pH 6.05) or acid habituated (prepared in inoculated water fluids mixed with filter-sterilized 2% lactic acid [LA] runoff fluids in a proportion of 1/99 [vol/vol], pH 4.12). The inoculated surfaces were exposed to conditions simulating carcass chilling (−3°C for 10 h followed by 38 h at 1°C). Treatments applied to samples (between 0 and 10 h) during chilling included the following: (i) no spraying (NT) or spraying (for 30 s every 30 min) with (ii) water, (iii) cetylpyridinium chloride (CPC; 0.1 or 0.5%), (iv) ammonium hydroxide (AH; 0.05%), (v) lactic acid (LA; 2%), (vi) acidified sodium chlorite (ASC; 0.12%), (vii) peroxyacetic acid (PAA; 0.02%), (viii) sodium hydroxide (SH; 0.01%), or (ix) sodium hypochlorite (SC; 0.005%) solutions of 4°C. Samples were taken at 0, 10, 24, 36, and 48 h of the chilling process to determine changes in E. coli O157:H7 populations. Phase 1 tested water, SH, PAA, LA, and 0.5% CPC on meat inoculated with non–acid-habituated pathogen populations, whereas phase 2 tested water, SC, AH, ASC, LA, and 0.1% CPC on meat inoculated with acid- and non–acid-habituated populations. Reductions in non–acid-habituated E. coli O157:H7 populations from phase 1 increased in the order NT = water = SH < PAA < LA < CPC. Reductions from phase 2 for acid-habituated cells increased in the order NT = water = SC < ASC = LA = AH < CPC, whereas on non–acid-habituated cells the order observed was NT = water = SC < AH = ASC < LA < CPC. Previous acid habituation of E. coli O157:H7 inocula rendered the cells more resistant to the effects of spray chilling, especially with acid; however, the trend of reduction remained spray chilling with water = non–spray chilling < spray chilling with chemical solutions.


2004 ◽  
Vol 67 (11) ◽  
pp. 2375-2380 ◽  
Author(s):  
ROLANDO J. GONZALEZ ◽  
YAGUANG LUO ◽  
SAUL RUIZ-CRUZ ◽  
JAMES L. McEVOY

Chlorine is widely used as a sanitizer to maintain the microbial quality and safety of fresh-cut produce; however, chlorine treatment lacks efficacy on pathogen reduction, especially when the fresh-cut processing water contains heavy organic loads. A more efficacious sanitizer that can tolerate the commercial processing conditions is needed to maintain microbial safety of fresh-cut produce. This study evaluated the efficacy of Escherichia coli O157:H7 reduction on fresh-cut carrots using new and traditional sanitizers with tap water and fresh-cut processing water scenarios. Fresh-cut carrot shreds inoculated with E. coli O157:H7 were washed in sanitizer solutions including 200 ppm chlorine, citric acid–based sanitizer (Pro-San), 80 ppm peroxyacetic acid-based sanitizer (Tsunami 100), and 1,000 ppm acidified sodium chlorite (SANOVA) prepared in fresh tap water or simulated processing water with a chemical oxygen demand level of approximately 3,500 mg/liter. Samples were packaged and stored at 5°C. Microbial analyses performed at days 0, 7, and 14 indicate that the organic load in the process water significantly affected the efficacy of chlorine on pathogen removal and was especially evident on samples tested during storage. Acidified sodium chlorite provided a strong pathogen reduction even under process water conditions with up to a 5.25-log reduction when compared with the no-wash control. E. coli O157:H7 was not recovered on acidified sodium chlorite–treated samples during the entire 14 days of storage, even following an enrichment step. These results suggest that acidified sodium chlorite holds considerable promise as an alternative sanitizer of fresh-cut produce.


1999 ◽  
Vol 62 (6) ◽  
pp. 580-584 ◽  
Author(s):  
A. CASTILLO ◽  
L. M. LUCIA ◽  
G. K. KEMP ◽  
G. R. ACUFF

The efficacy of a phosphoric acid–activated acidified sodium chloride (PASC) spray and a citric acid–activated acidified sodium chlorite (CASC) spray applied at room temperature (22.4 to 24.7°C) in combination with a water wash was compared with that of a water wash only treatment for reduction of Escherichia coli O157:H7 and Salmonella Typhimurium inoculated onto various hot-boned individual beef carcass surface regions (inside round, outside round, brisket, flank, and clod). Initial counts of 5.5 and 5.4 log CFU/cm2 were obtained after inoculation with E. coli O157:H7 and Salmonella Typhimurium, respectively. Initial numbers for both pathogens were reduced by 3.8 to 3.9 log cycles by water wash followed by PASC spray and by 4.5 to 4.6 log cycles by water wash followed by CASC spray. The sprays consisted of applying 140 ml of the appropriate sanitizing solution for 10 s at 69 kPa. Corresponding reduction values obtained by water wash alone were 2.3 log. The performance of CASC appeared to be consistently better than that of PASC. In general, no effect of the carcass surface region was observed on the log reductions for either pathogen, except for the inside round, which consistently had lower reductions. Both PASC and CASC were capable of effectively reducing pathogens spread to areas beyond the initial contaminated area of the cuts to levels close to or below the counting method detection limit (0.5 log CFU/cm2). However, 30 to 50% of the carcasses treated by these antimicrobial solutions still yielded countable colonies. Results of this study indicate that acidified sodium chlorite sprays are effective for decontaminating beef carcass surfaces.


2005 ◽  
Vol 68 (2) ◽  
pp. 251-255 ◽  
Author(s):  
YASUHIRO INATSU ◽  
LATIFUL BARI ◽  
SUSUMU KAWASAKI ◽  
KENJI ISSHIKI ◽  
SHINICHI KAWAMOTO

Efficacy of acidified sodium chlorite for reducing the population of Escherichia coli O157:H7 pathogens on Chinese cabbage leaves was evaluated. Washing leaves with distilled water could reduce the population of E. coli O157:H7 by approximately 1.0 log CFU/g, whereas treating with acidified chlorite solution could reduce the population by 3.0 log CFU/g without changing the leaf color. A similar level of reduction was achieved by washing with sodium chlorite solution containing various organic acids. However, acidified sodium chlorite in combination with a mild heat treatment reduced the population by approximately 4.0 log CFU/g without affecting the color, but it softened the leaves. Moreover, the efficacy of the washing treatment was similar at low (4°C) and room (25°C) temperatures, indicating that acidified sodium chloride solution could be useful as a sanitizer for surface washing of fresh produce.


2004 ◽  
Vol 67 (10) ◽  
pp. 2123-2131 ◽  
Author(s):  
JEE-HOON RYU ◽  
HOIKYUNG KIM ◽  
LARRY R. BEUCHAT

The influence of exopolysaccharide (EPS) production, nutrient availability, and temperature on attachment and biofilm formation by Escherichia coli O157:H7 strains ATCC 43895 (wild type) and 43895-EPS (extensive EPS-producing mutant) on stainless steel coupons (SSCs) was investigated. Cells grown on heated lettuce juice agar and modified tryptic soy agar were suspended in phosphate-buffered saline (PBS). SSCs were immersed in the cell suspension (109 CFU/ml) at 4°C for 24 h. Biofilm formation by cells attached to SSCs as affected by immersing in 10% tryptic soy broth (TSB), lettuce juice broth (LJB), and minimal salts broth (MSB) at 12 and 22°C was studied. A significantly lower number of strain 43895-EPS cells, compared to strain ATCC 43895 cells, attached to SSCs during a 24-h incubation (4°C) period in PBS suspension. Neither strain formed a biofilm on SSCs subsequently immersed in 10% TSB or LJB, but both strains formed biofilms in MSB. Populations of attached cells and planktonic cells of strain ATCC 43895 gradually decreased during incubation for 6 days in LJB at 22°C, but populations of strain 43895-EPS remained constant for 6 days at 22°C, indicating that the EPS-producing mutant, compared to the wild-type strain, has a higher tolerance to the low-nutrient environment presented by LJB. It is concluded that EPS production by E. coli O157:H7 inhibits attachment to SSCs and that reduced nutrient availability enhances biofilm formation. Biofilms formed under conditions favorable for EPS production may protect E. coli O157:H7 against sanitizers used to decontaminate lettuce and produce processing environments. Studies are under way to test this hypothesis.


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