Hollow-Fiber Ultrafiltration for the Concentration and Simultaneous Recovery of Multiple Pathogens in Contaminated Foods

2009 ◽  
Vol 72 (12) ◽  
pp. 2547-2552 ◽  
Author(s):  
HEE-YEON KIM ◽  
HYEUN-JIN PARK ◽  
GWANGPYO KO
Keyword(s):  
Hollow Fiber ◽  
Food Samples ◽  
Murine Norovirus ◽  
Viral Pathogens ◽  
Recovery Rates ◽  
Total Recovery ◽  
E Coli ◽  
Reverse Transcription Pcr ◽  
Elution Buffer ◽  
Multiple Pathogens

We investigated the possibility of using hollow-fiber ultrafiltration (HUF) for the simultaneous recovery of multiple microorganisms in food samples. MS2 bacteriophage, E. coli, Bacillus subtilis spores, and murine norovirus (MNV) were each inoculated into 5 liters of either distilled water (DW) or glycine elution buffer and then concentrated using hollow-fiber polysulfone ultrafilters. The resulting concentrates were further analyzed by either cultivation or TaqMan real-time reverse transcription PCR assay. The overall average recovery rates were 7.1% in DW and 17.1% in glycine elution buffer. When the virus, vegetative bacteria, and bacterial spores were simultaneously inoculated into DW, glycine, or Tris-HCl elution buffers, on average 16.8% of inoculated microorganisms were recovered by HUF. The addition of 3% beef extract blocking buffer to HUF increased the total recovery rate to 46.1%, with incremental recovery rates increasing sharply for B. subtilis spores and MNV. Use of HUF resulted in E. coli recovery rates of 68.0% on lettuce and 66.2% on ham and MNV recovery rates of 1.5% on lettuce and 5.8% on ham. Our study demonstrates that HUF can be effective at simultaneously recovering and concentrating diverse bacterial and viral pathogens from foods.

Evaluation of Various Methods for Recovering Human Norovirus and Murine Norovirus from Vegetables and Ham

2010 ◽  
Vol 73 (9) ◽  
pp. 1651-1657 ◽  
Author(s):  
HYEONJIN PARK ◽  
MINJUNG KIM ◽  
GWANGPYO KO
Keyword(s):  
Molecular Weight ◽  
Polyethylene Glycol ◽  
Hollow Fiber ◽  
Significant Positive Correlation ◽  
Recovery Efficiency ◽  
Murine Norovirus ◽  
Human Norovirus ◽  
Ph Values ◽  
Elution Buffer ◽  
Concentration Methods

We evaluated and optimized each step in an analytical method for detecting norovirus from various foods. We characterized the buffers needed for eluting norovirus from foods such as ham and lettuce. Two different concentration methods, polyethylene glycol (PEG) precipitation and hollow fiber ultrafiltration (HUF), were compared using both murine norovirus (MNV) and human norovirus (HuNoV). For PEG precipitation, an elution buffer containing 3% beef extract (pH 7.1) was more suitable than 0.05 M glycine plus 0.14 M NaCl (pH 7.5), and the recovery efficiency increased with increasing molecular weight of PEG. To determine the optimal buffer for concentrating norovirus by HUF, glycine buffers with different pH values and ionic strengths were examined as elution buffers. Overall, HUF was more efficient for norovirus recovery than was PEG precipitation. Because there was a significant positive correlation between MNV and HuNoV results, MNV could be a useful surrogate for detecting HuNoV in foods.

scholarly journals Bioaccumulation, Retention, and Depuration of Enteric Viruses by Crassostrea virginica and Crassostrea ariakensis Oysters

2008 ◽  
Vol 74 (22) ◽  
pp. 6825-6831 ◽  
Author(s):  
Sharon P. Nappier ◽  
Thaddeus K. Graczyk ◽  
Kellogg J. Schwab
Keyword(s):  
Crassostrea Virginica ◽  
Hepatitis A Virus ◽  
Proteinase K ◽  
Murine Norovirus ◽  
Viral Pathogens ◽  
Crassostrea Ariakensis ◽  
Reverse Transcription Pcr ◽  
Bacteriophage Ms2 ◽  
Continuous Filtration ◽  
Male Specific

ABSTRACT Crassostrea ariakensis oysters are under review for introduction into the Chesapeake Bay. However, the human health implications of the introduction have not been fully addressed. This study evaluated rates of bioaccumulation, retention, and depuration of viruses by Crassostrea virginica and C. ariakensis when the two oyster species were maintained in separate tanks containing synthetic seawater of various salinities (8, 12, or 20 ppt). Oyster bioaccumulation tanks were seeded with 103 PFU/ml of hepatitis A virus (HAV), poliovirus, male-specific bacteriophage (MS2), and murine norovirus 1 (MNV-1) and 103 PCR units/ml of human norovirus (NoV). After 24 h, depuration commenced as oysters (n = 255) were placed in pathogen-free seawater under continuous filtration. Oysters (n = 6) were sampled weekly for 1 month from each tank. Viral RNA was recovered using a modified proteinase K, guanidine, and glassmilk method and analyzed by quantitative reverse transcription-PCR. The odds of C. ariakensis oysters harboring NoV, MNV-1, or HAV were statistically greater than the odds of C. virginica oysters harboring the same viruses (MNV-1 odds ratio [OR], 4.5; P = 0.01; NoV OR, 8.4; P < 0.001; HAV OR, 11.4; P < 0.001). Unlike C. virginica, C. ariakensis bioaccumulated and retained NoV, MNV-1, and HAV for 1 month at all salinities. Additionally, the odds of an oyster testing positive for NoV was 25.5 times greater (P < 0.001) when the oyster also tested positive for MNV-1. This research helps assess the threat of C. ariakensis as a vehicle for viral pathogens due to the consumption of raw oysters and validates the role for MNV-1 as a surrogate for NoV.

scholarly journals Disruption of Type III Interferon (IFN) Genes Ifnl2 and Ifnl3 Recapitulates Loss of the Type III IFN Receptor in the Mucosal Antiviral Response

2019 ◽  
Vol 93 (22) ◽  
Author(s):  
Stefan T. Peterson ◽  
Elizabeth A. Kennedy ◽  
Pamela H. Brigleb ◽  
Gwen M. Taylor ◽  
Kelly Urbanek ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Viral Infections ◽  
Murine Norovirus ◽  
Viral Pathogens ◽  
Genetic Ablation ◽  
Type Iii ◽  
Mucosal Surfaces ◽  
Receptor Interactions ◽  
Type Iii Ifn ◽  
Type Iii Interferon

ABSTRACT Type III interferon (IFN), or IFN lambda (IFN-λ), is an essential component of the innate immune response to mucosal viral infections. In both the intestine and the lung, signaling via the IFN-λ receptor (IFNLR) controls clinically important viral pathogens, including influenza virus, norovirus, and rotavirus. While it is thought that IFN-λ cytokines are the exclusive ligands for signaling through IFNLR, it is not known whether genetic ablation of these cytokines phenotypically recapitulates disruption of the receptor. Here, we report the serendipitous establishment of Ifnl2−/− Ifnl3−/− mice, which lack all known functional murine IFN-λ cytokines. We demonstrate that, like Ifnlr1−/− mice lacking IFNLR signaling, these mice display defective control of murine norovirus, reovirus, and influenza virus and therefore genocopy Ifnlr1−/− mice. Thus, for regulation of viral infections at mucosal sites of both the intestine and lung, signaling via IFNLR can be fully explained by the activity of known cytokines IFN-λ2 and IFN-λ3. Our results confirm the current understanding of ligand-receptor interactions for type III IFN signaling and highlight the importance of this pathway in regulation of mucosal viral pathogens. IMPORTANCE Type III interferons are potent antiviral cytokines important for regulation of viruses that infect at mucosal surfaces. Studies using mice lacking the Ifnlr1 gene encoding the type III interferon receptor have demonstrated that signaling through this receptor is critical for protection against influenza virus, norovirus, and reovirus. Using a genetic approach to disrupt murine type III interferon cytokine genes Ifnl2 and Ifnl3, we found that mice lacking these cytokines fully recapitulate the impaired control of viruses observed in mice lacking Ifnlr1. Our results support the idea of an exclusive role for known type III interferon cytokines in signaling via IFNLR to mediate antiviral effects at mucosal surfaces. These findings emphasize the importance of type III interferons in regulation of a variety of viral pathogens and provide important genetic evidence to support our understanding of the ligand-receptor interactions in this pathway.

scholarly journals Comparison of Fecal Coliform Agar and Violet Red Bile Lactose Agar for Fecal Coliform Enumeration in Foods

2002 ◽  
Vol 68 (4) ◽  
pp. 1631-1638 ◽  
Author(s):  
A. Leclercq ◽  
C. Wanegue ◽  
P. Baylac
Keyword(s):  
Escherichia Coli ◽  
Fecal Coliform ◽  
Food Samples ◽  
Fecal Coliforms ◽  
Predictive Values ◽  
Direct Plating ◽  
Hafnia Alvei ◽  
E Coli ◽  
Plating Method ◽  
Mashed Potatoes

ABSTRACT A 24-h direct plating method for fecal coliform enumeration with a resuscitation step (preincubation for 2 h at 37 ± 1°C and transfer to 44 ± 1°C for 22 h) using fecal coliform agar (FCA) was compared with the 24-h standardized violet red bile lactose agar (VRBL) method. FCA and VRBL have equivalent specificities and sensitivities, except for lactose-positive non-fecal coliforms such as Hafnia alvei, which could form typical colonies on FCA and VRBL. Recovery of cold-stressed Escherichia coli in mashed potatoes on FCA was about 1 log unit lower than that with VRBL. When the FCA method was compared with standard VRBL for enumeration of fecal coliforms, based on counting carried out on 170 different food samples, results were not significantly different (P > 0.05). Based on 203 typical identified colonies selected as found on VRBL and FCA, the latter medium appears to allow the enumeration of more true fecal coliforms and has higher performance in certain ways (specificity, sensitivity, and negative and positive predictive values) than VRBL. Most colonies clearly identified on both media were E. coli and H. alvei, a non-fecal coliform. Therefore, the replacement of fecal coliform enumeration by E. coli enumeration to estimate food sanitary quality should be recommended.

Sequence of Colonization Determines the Composition of Mixed Biofilms by Escherichia coli O157:H7 and O111:H8 Strains†

2015 ◽  
Vol 78 (8) ◽  
pp. 1554-1559 ◽  
Author(s):  
RONG WANG ◽  
NORASAK KALCHAYANAND ◽  
JAMES L. BONO
Keyword(s):  
Escherichia Coli ◽  
Foodborne Pathogens ◽  
Early Stage ◽  
Critical Role ◽  
The United States ◽  
Food Samples ◽  
E Coli ◽  
Cell Percentage ◽  
Composition And Structure ◽  
Biofilm Development

Bacterial biofilms are one of the potential sources of cross-contamination in food processing environments. Shiga toxin–producing Escherichia coli (STEC) O157:H7 and O111:H8 are important foodborne pathogens capable of forming biofilms, and the coexistence of these two STEC serotypes has been detected in various food samples and in multiple commercial meat plants throughout the United States. Here, we investigated how the coexistence of these two STEC serotypes and their sequence of colonization could affect bacterial growth competition and mixed biofilm development. Our data showed that E. coli O157:H7 strains were able to maintain a higher cell percentage in mixed biofilms with the co-inoculated O111:H8 companion strains, even though the results of planktonic growth competition were strain dependent. On solid surfaces with preexisting biofilms, the sequence of colonization played a critical role in determining the composition of the mixed biofilms because early stage precolonization significantly affected the competition results between the E. coli O157:H7 and O111:H8 strains. The precolonizer of either serotype was able to outgrow the other serotype in both planktonic and biofilm phases. The competitive interactions among the various STEC serotypes would determine the composition and structure of the mixed biofilms as well as their potential risks to food safety and public health, which is largely influenced by the dominant strains in the mixtures. Thus, the analysis of mixed biofilms under various conditions would be of importance to determine the nature of mixed biofilms composed of multiple microorganisms and to help implement the most effective disinfection operations accordingly.

scholarly journals Effects ofibeADeletion on Virulence and Biofilm Formation of Avian PathogenicEscherichia coli

2010 ◽  
Vol 79 (1) ◽  
pp. 279-287 ◽  
Author(s):  
Shaohui Wang ◽  
Chunling Niu ◽  
Zhenyu Shi ◽  
Yongjie Xia ◽  
Muhammad Yaqoob ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Wild Type Strain ◽  
Chicken Embryo Fibroblast ◽  
Neonatal Meningitis ◽  
Pcr Analysis ◽  
E Coli ◽  
Reverse Transcription Pcr ◽  
Animal System ◽  
The Brain

ABSTRACTTheibeAgene is located on a genomic island, GimA, which is involved in the pathogenesis of neonatal meningitisEscherichia coli(NMEC) and avian pathogenicE. coli(APEC). The prevalence ofibeAin the APEC collection in China was investigated, and 20 of 467 strains (4.3%) were positive. In addition, analysis of the association of theE. colireference (ECOR) groups with positive strains revealed thatibeAwas linked to group B2. TheibeAgene in DE205B was analyzed and compared to those of APEC and NMEC, which indicated that the specificity ofibeAwas not consistent along pathotypes. The invasion of chicken embryo fibroblast DF-1 cells by APEC DE205B and RS218 was observed, which suggested that DF-1 cells could be a model to study the mechanism of APEC invasion. The inactivation ofibeAin APEC DE205B led to the reduced capacity to invade DF-1 cells, defective virulencein vivo, and decreased biofilm formation compared to the wild-type strain. In addition, strain AAEC189 expressingibeAexhibited enhanced invasion capacity and biofilm formation. The results of the quantitative real-time reverse transcription-PCR (qRT-PCR) analysis and animal system infection experiments indicated that the loss ofibeAdecreased the colonization and proliferation capacities of APEC in the brain during system infection.

Simultaneous Detection of Four Foodborne Viruses in Food Samples Using a One-Step Multiplex Reverse Transcription PCR

2018 ◽  
Vol 28 (2) ◽  
pp. 210-217 ◽  
Author(s):  
Shin-Young Lee ◽  
Mi-Ju Kim ◽  
Hyun-Joong Kim ◽  
KwangCheol Casey Jeong ◽  
Hae-Yeong Kim
Keyword(s):  
Reverse Transcription ◽  
Simultaneous Detection ◽  
Food Samples ◽  
Foodborne Viruses ◽  
Reverse Transcription Pcr ◽  
One Step

Distribution of Coliphages in Various Foods

1986 ◽  
Vol 49 (12) ◽  
pp. 944-951 ◽  
Author(s):  
J. E. KENNEDY ◽  
C. I. WEI ◽  
J. L. OBLINGER
Keyword(s):  
Escherichia Coli ◽  
Food Samples ◽  
Fecal Coliforms ◽  
Bacterial Indicators ◽  
Processed Meats ◽  
E Coli ◽  
Plate Counts ◽  
The Relationship

The distribution of coliphages in various foods and the relationship between the incidences of coliphages and bacterial indicators were investigated. A total of 120 food samples comprising twelve products and including fresh meats, shellfish, vegetables and processed meats, were analyzed for indigenous coliphages using Escherichia coli hosts C, C-3000 and B. Bacterial analyses included enumeration of E. coli, fecal coliforms and coliforms, as well as aerobic plate counts and Salmonella analyses. Coliphages were detected (≥10 PFU/100 g) in 56% of samples and eleven of twelve products. Coliphages, E. coli, fecal coliforms and coliforms were recovered at a level of at least 30 organisms per 100 g in 43, 43, 68 and 81% of samples, with overall mean recoveries of 13, 19, 93 and 4300 organisms/100 g, respectively. Highest and lowest recoveries of coliphages and E. coli were from fresh meats and vacuum-packaged processed meats, respectively. Significant nonparametric correlations between coliphages, E. coli, fecal coliforms and coliforms were found among all food samples.

Simultaneous Enrichment of Shiga Toxin–Producing Escherichia coli O157 and O26 and Salmonella in Food Samples Using Universal Preenrichment Broth

2009 ◽  
Vol 72 (10) ◽  
pp. 2065-2070 ◽  
Author(s):  
MASASHI KANKI ◽  
KAZUKO SETO ◽  
JUNKO SAKATA ◽  
TETSUYA HARADA ◽  
YUKO KUMEDA
Keyword(s):  
Escherichia Coli ◽  
Shiga Toxin ◽  
Food Samples ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Significant Difference ◽  
Peptone Water ◽  
Radish Sprouts ◽  
Pork Samples ◽  
Better Than

Universal preenrichment broth (UPB) was compared with modified Escherichia coli broth with novobiocin (mEC+n) for enrichment of Shiga toxin–producing E. coli O157 and O26, and with buffered peptone water (BPW) for preenrichment of Salmonella enterica. Ten strains each of the three pathogens were inoculated into beef and radish sprouts following thermal, freezing, or no treatment. With regard to O157 and O26, UPB incubated at 42°C recovered significantly more cells from inoculated beef than UPB at 35°C and from radish sprout samples than UPB at 35°C and mEC+n. With regard to Salmonella, UPB incubated at 42°C was as effective as UPB at 35°C and BPW at recovering cells from beef and radish sprout samples. No significant difference was noted between the effectiveness of UPB at 42°C and UPB at 35°C or BPW in the recovery of Salmonella from 205 naturally contaminated poultry samples. By using UPB at 42°C, one O157:H7 strain was isolated from the mixed offal of 53 beef samples, 6 cattle offal samples, and 50 pork samples all contaminated naturally, with no pathogen inoculation. The present study found that UPB incubated at 42°C was as effective as, or better than, mEC+n for enrichment of O157 and O26 and comparable to BPW for preenrichment of Salmonella. These findings suggest that a great deal of labor, time, samples, and space may be saved if O157, O26, and Salmonella are enriched simultaneously with UPB at 42°C.

scholarly journals Extended-Spectrum Beta-Lactamase (ESBL)-Producing Escherichia coli Isolated from Flies in the Urban Center of Berlin, Germany

2019 ◽  
Vol 16 (9) ◽  
pp. 1530 ◽  
Author(s):  
Wibke Wetzker ◽  
Yvonne Pfeifer ◽  
Solvy Wolke ◽  
Andrea Haselbeck ◽  
Rasmus Leistner ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Continuous Monitoring ◽  
Companion Animals ◽  
Multidrug Resistant ◽  
Beta Lactamase ◽  
Extended Spectrum Beta Lactamase ◽  
E Coli ◽  
Extended Spectrum ◽  
Ciprofloxacin Resistance ◽  
Multiple Pathogens

Background: The monitoring of antimicrobial resistance (AMR) in microorganisms that circulate in the environment is an important topic of scientific research and contributes to the development of action plans to combat the spread of multidrug-resistant (MDR) bacteria. As a synanthropic vector for multiple pathogens and a reservoir for AMR, flies can be used for surveillance. Methods: We collected 163 flies in the inner city of Berlin and examined them for extended-spectrum β-lactamase (ESBL)-producing Escherichia coli genotypically and phenotypically. Results: The prevalence of ESBL-producing E. coli in flies was 12.9%. Almost half (47.6%) of the ESBL-positive samples showed a co-resistance to ciprofloxacin. Resistance to carbapenems or colistin was not detected. The predominant ESBL-type was CTX-M-1, which is associated with wildlife, livestock, and companion animals as a potential major source of transmission of MDR E. coli to flies. Conclusions: This field study confirms the permanent presence of ESBL-producing E. coli in an urban fly population. For continuous monitoring of environmental contamination with multidrug-resistant (MDR) bacteria, flies can be used as indicators without much effort.

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