Inactivation of Cryptosporidium parvum Oocysts in Cider by Flash Pasteurization

2001 ◽  
Vol 64 (4) ◽  
pp. 523-527 ◽  
Author(s):  
MING QI DENG ◽  
DEAN O. CLIVER
Keyword(s):  
Cryptosporidium Parvum ◽  
Fluorescent Antibody ◽  
Most Probable Number ◽  
Probable Number ◽  
Log Reduction ◽  
Antibody Staining ◽  
Heat Treated ◽  
Infected Cells ◽  
Medical Importance

Cryptosporidium parvum is a well-recognized pathogen of significant medical importance, and cider (apple juice) has been associated with foodborne cryptosporidiosis. This study investigated the effect of flash pasteurization on the viability of contaminant C. parvum oocysts. Cider inoculated with oocysts was heated at 70 or 71.7°C for 5, 10, or 20 s, and oocyst viability was measured by a semiquantitative in vitro infectivity assay. By infecting multiple wells of confluent Madin-Darby bovine kidney cells with serial dilutions of heat-treated oocysts and examining infected cells by indirect fluorescent antibody staining, the most probable number technique was applied to quantify log reduction of oocyst viability. Heating for 10 or 20 s at either temperature caused oocyst killing of at least 4.9 log (or 99.999%), whereas oocyst inactivation after pasteurization for 5 s at 70 and 71.7°C was 3.0 log (99.9%) and 4.8 log (99.998%), respectively. Our results suggested that current practices of flash pasteurization in the juice industry are sufficient in inactivating contaminant oocysts.

A Most Probable Number Method for the Enumeration of Legionella Bacteria in Water

1991 ◽  
Vol 24 (2) ◽  
pp. 143-147 ◽  
Author(s):  
N. A. Grabow ◽  
R. Kfir ◽  
W. O. K. Grabow
Keyword(s):  
Water Samples ◽  
Membrane Filtration ◽  
Quantitative Method ◽  
Fluorescent Antibody ◽  
Most Probable Number ◽  
Probable Number ◽  
Comparative Tests ◽  
Antibody Staining ◽  
Direct Fluorescent Antibody ◽  
Yeast Extract Agar

A new quantitative method for the enumeration of Legionella bacteria in water is described. Appropriate tenfold serial dilutions of water samples concentrated by membrane filtration are plated in triplicate on buffered charcoal yeast extract agar. After incubation for 3 days representative smears from individual plates are tested for the presence of Legionella by direct fluorescent antibody staining. The number of positive plates in each dilution is used to calculate the Legionella count by means of conventional most probable number statistics. In comparative tests on a variety of water samples this method yielded significantly higher counts than previously used procedures.

Efficacy of Gaseous Chlorine Dioxide as a Sanitizer against Cryptosporidium parvum, Cyclospora cayetanensis, and Encephalitozoon intestinalis on Produce

2008 ◽  
Vol 71 (12) ◽  
pp. 2410-2414 ◽  
Author(s):  
YNES R. ORTEGA ◽  
AMY MANN ◽  
MARIA P. TORRES ◽  
VITALIANO CAMA
Keyword(s):  
Cryptosporidium Parvum ◽  
Chlorine Dioxide ◽  
Most Probable Number ◽  
Escherichia Coli O157 ◽  
Probable Number ◽  
Cyclospora Cayetanensis ◽  
E Coli ◽  
Log Reduction ◽  
Encephalitozoon Intestinalis ◽  
Gaseous Chlorine Dioxide

The efficacy of gaseous chlorine dioxide to reduce parasite and bacterial burden in produce was studied. Basil and lettuce leaves were inoculated with Cryptosporidium parvum and Cyclospora cayetanensis oocysts, Encephalitozoon intestinalis spores, and a cocktail of two isolates of nalidixic acid–resistant Escherichia coli O157:H7. The inoculated samples were then treated for 20 min with gaseous chlorine dioxide at 4.1 mg/liter. Cryptosporidium had a 2.6 and 3.31 most-probable-number log reduction in basil and lettuce, respectively. Reduction of Encephalitozoon in basil and lettuce was 3.58 and 4.58 CFU/g respectively. E. coli loads were significantly reduced (2.45 to 3.97 log), whereas Cyclospora sporulation was not affected by this treatment.

Effect of Organic Acids and Hydrogen Peroxide on Cryptosporidium parvum Viability in Fruit Juices

2003 ◽  
Vol 66 (9) ◽  
pp. 1650-1657 ◽  
Author(s):  
KALMIA E. KNIEL ◽  
SUSAN S. SUMNER ◽  
DAVID S. LINDSAY ◽  
CAMERON R. HACKNEY ◽  
MERLE D. PIERSON ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Cell Culture ◽  
Organic Acids ◽  
Cryptosporidium Parvum ◽  
Most Probable Number ◽  
Oocyst Wall ◽  
Probable Number ◽  
Apple Cider ◽  
Infectivity Assay

Cryptosporidium parvum has historically been associated with waterborne outbreaks of diarrheal illness. Foodborne cryptosporidiosis has been associated with unpasteurized apple cider. Infectious oocysts are shed in the feces of common ruminants like cattle and deer in and near orchards. In this study, the ability of organic acids and hydrogen peroxide (H2O2) added to fruit juice to inhibit the survival of C. parvum was analyzed. Oocyst viability was analyzed by a cell culture infectivity assay with the use of a human ileocecal cell line (HCT-8) whose infectivity pattern is similar to that for human oral infectivity. Cell monolayers were infected with 106 treated oocysts or a series of 10-fold dilutions. Parasitic life stages were visualized through immunohistochemistry with 100 microscope fields per monolayer being counted. In vitro excystation assays were also used to evaluate these treatments. Organic acids and H2O2 were added to apple cider, orange juice, and grape juices on a weight/volume basis. Malic, citric, and tartaric acids at concentrations of 1 to 5% inhibited C. parvum's infectivity of HCT-8 cells by up to 88%. Concentrations ranging from 0.025 to 3% H2O2 were evaluated. The addition of 0.025% H2O2 to each juice resulted in a >5-log reduction of C. parvum infectivity as determined with a most-probable-number–based cell culture infectivity assay. As observed with differential interference contrast and scanning electron microscopy, reduced infectivity may be mediated through effects on the oocyst wall that are caused by the action of H2O2 or related oxygen radicals. The addition of low concentrations of H2O2 can represent a valuable alternative to pasteurization.

scholarly journals Fabrication and Characterization of Antimicrobial Magnetron Cosputtered TiO2/Ag/Cu Composite Coatings

Coatings ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 473
Author(s):  
Dilyana Gospodonova ◽  
Iliana Ivanova ◽  
Todorka Vladkova
Keyword(s):  
Antimicrobial Activity ◽  
Composite Coatings ◽  
Surface Characteristics ◽  
Standard Test ◽  
Most Probable Number ◽  
Bacterial Strains ◽  
Probable Number ◽  
Fabrication And Characterization

The aim of this study was to prepare TiO2/Ag/Cu magnetron co-sputtered coatings with controlled characteristics and to correlate them with the antimicrobial activity of the coated glass samples. The elemental composition and distribution, surface morphology, wettability, surface energy and its component were estimated as the surface characteristics influencing the bioadhesion. Well expressed, specific, Ag/Cu concentration-dependent antimicrobial activity in vitro was demonstrated toward Gram-negative and Gram-positive standard test bacterial strains both by diffusion 21 assay and by Most Probable Number of surviving cells. Direct contact and eluted silver/coper nanoparticles killing were experimentally demonstrated as a mode of the antimicrobial action of the studied TiO2/Ag/Cu thin composite coatings. It is expected that they would ensure a broad spectrum bactericidal activity during the indwelling of the coated medical devices and for at least 12 h after that, with the supposition that the benefits will be over a longer time.

Differential Survival of Generic E. coli and Listeria spp. in Northeastern U.S. Soils Amended with Dairy Manure Compost, Poultry Litter Compost, and Heat-Treated Poultry Pellets and Fate in Raw Edible Radish Crops

2021 ◽  
Author(s):  
Marie Limoges ◽  
Deborah A. Neher ◽  
Thomas R. Weicht ◽  
Patricia D. Millner ◽  
Manan Sharma ◽  
...  
Keyword(s):  
Fresh Produce ◽  
Poultry Litter ◽  
Dairy Manure ◽  
Animal Origin ◽  
Most Probable Number ◽  
Post Inoculation ◽  
Probable Number ◽  
E Coli ◽  
Manure Compost ◽  
Heat Treated

Composted or heat-treated Biological Soil Amendments of Animal Origin (BSAAO) can be added to soils to provide nutrients for fresh produce. These products lower the risk of pathogen contamination of fresh produce when compared with use of untreated BSAAO; however, meteorological conditions, geographic location, and soil properties can influence the presence of pathogenic bacteria, or their indicators (e.g., generic E. coli) and allow potential for produce contamination. Replicated field plots of loamy or sandy soils were tilled and amended with dairy manure compost (DMC), poultry litter compost (PLC), or no compost (NoC) over two different field seasons, and non-composted heat-treated poultry pellets (HTPP) during the second field season. Plots were inoculated with a three-strain cocktail of rifampicin-resistant E. coli (rE.coli) at levels of 8.7 log CFU/m2. Direct plating and most probable number (MPN) methods measured the persistence of rE.coli and Listeria spp. in plots through 104 days post-inoculation. Greater survival of rE. coli was observed in PLC plots in comparison to DMC plots and NoC plots during year 1 (P < 0.05). Similar trends were observed for year 2, where rE. coli survival was also greater in HTPP amended plots (P < 0.05). Survival of rE. coli was dependent on soil type, where water potential and temperature were significant covariables. Listeria spp. were found in NoC plots, but not in plots amended with HTPP, PLC or DMC. Radish data demonstrate that PLC treatment promoted the greatest level of rE.coli translocation when compared to DMC and NoC treatments (P  < 0.05). These results are consistent with findings from studies conducted in other regions of the US and informs Northeast produce growers that composted and non-composted poultry-based BSAAO supports greater survival of rE. coli in field soils. This result has the potential to impact the food safety risk of edible produce grown in BSAAO amended soils as a result of pathogen contamination.

Thermal Inactivation of Borrelia burgdorferi, the Cause of Lyme Disease

1990 ◽  
Vol 53 (4) ◽  
pp. 296-299 ◽  
Author(s):  
SI K. LEE ◽  
AHMED E. YOUSEF ◽  
ELMER H. MARTH
Keyword(s):  
Borrelia Burgdorferi ◽  
Thermal Inactivation ◽  
Treatment Time ◽  
Treatment Temperature ◽  
Most Probable Number ◽  
Lag Phase ◽  
Probable Number ◽  
Heat Treated ◽  
Thermal Death ◽  
D Values

Borrelia burgdorferi strain EBNI was cultivated in BSK-II medium at 34°C, then cultures at different physiological states were heat-treated at temperatures in the range of 50 to 70°C. Numbers of survivors were estimated by the Most Probable Number technique. Log MPN was plotted against treatment time, and resulting survivor curves were linear. Estimated D-values for cultures incubated at 34°C for 7 d before heat-treatment were 5.5, 4.3, 2.7, .47, and .14 min at 50, 55, 60, 65, and 70°C, respectively. Spirochetes in the lag phase had greater resistance to heat than those in the stationary phase, with the latter being more resistant to heat than spirochetes in the same phase of growth but refrigerated at 4°C for 3 d. D-values for B. burgdorferi are generally less at 50°C, and greater at 70°C than those reported for other nonsporeforming pathogens. When log10 MPN was plotted against treatment temperature, two linear segments for each thermal death curve were obtained. Our data show the spirochete had higher z-values than most nonsporeforming pathogens. The pH of the medium, in the range of 5.0 to 7.6, did not affect resistance of B. burgdorferi to heat.

scholarly journals Difference in persistent tuberculosis bacteria between in vitro and sputum from patients: implications for translational predictions

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Alan Faraj ◽  
Oskar Clewe ◽  
Robin J. Svensson ◽  
Galina V. Mukamolova ◽  
Michael R. Barer ◽  
...  
Keyword(s):  
Drug Response ◽  
Most Probable Number ◽  
Sputum Smear ◽  
Probable Number ◽  
Phenotypic Resistance ◽  
Phenotypic Differences ◽  
Model Based ◽  
Persistent Bacteria ◽  
The Difference

Abstract This study aimed to investigate the number of persistent bacteria in sputum from tuberculosis patients compared to in vitro and to suggest a model-based approach for accounting for the potential difference. Sputum smear positive patients (n = 25) provided sputum samples prior to onset of chemotherapy. The number of cells detected by conventional agar colony forming unit (CFU) and most probable number (MPN) with Rpf supplementation were quantified. Persistent bacteria was assumed to be the difference between MPNrpf and CFU. The difference in persistent bacteria between in vitro and human sputum prior to chemotherapy was quantified using different model-based approaches. The persistent bacteria in sputum was 17% of the in vitro levels, suggesting a difference in phenotypic resistance, whereas no difference was found for multiplying bacterial subpopulations. Clinical trial simulations showed that the predicted time to 2 log fall in MPNrpf in a Phase 2a setting using in vitro pre-clinical efficacy information, would be almost 3 days longer if drug response was predicted ignoring the difference in phenotypic resistance. The discovered phenotypic differences between in vitro and humans prior to chemotherapy could have implications on translational efforts but can be accounted for using a model-based approach for translating in vitro to human drug response.

scholarly journals Chlorine Dioxide Inactivation ofCryptosporidium parvum Oocysts and Bacterial Spore Indicators

2001 ◽  
Vol 67 (7) ◽  
pp. 2993-3001 ◽  
Author(s):  
Christian P. Chauret ◽  
Chris Z. Radziminski ◽  
Michael Lepuil ◽  
Robin Creason ◽  
Robert C. Andrews
Keyword(s):  
Chlorine Dioxide ◽  
Most Probable Number ◽  
Inactivation Kinetics ◽  
Clostridium Sporogenes ◽  
Probable Number ◽  
Waterborne Pathogen ◽  
Viability Assay ◽  
The Relationship ◽  
Different Sources

ABSTRACT Cryptosporidium parvum, which is resistant to chlorine concentrations typically used in water treatment, is recognized as a significant waterborne pathogen. Recent studies have demonstrated that chlorine dioxide is a more efficient disinfectant than free chlorine against Cryptosporidium oocysts. It is not known, however, if oocysts from different suppliers are equally sensitive to chlorine dioxide. This study used both a most-probable-number–cell culture infectivity assay and in vitro excystation to evaluate chlorine dioxide inactivation kinetics in laboratory water at pH 8 and 21�C. The two viability methods produced significantly different results (P < 0.05). Products of disinfectant concentration and contact time (Ct values) of 1,000 mg � min/liter were needed to inactivate approximately 0.5 log10 and 2.0 log10 units (99% inactivation) of C. parvumas measured by in vitro excystation and cell infectivity, respectively, suggesting that excystation is not an adequate viability assay. Purified oocysts originating from three different suppliers were evaluated and showed marked differences with respect to their resistance to inactivation when using chlorine dioxide.Ct values of 75, 550, and 1,000 mg � min/liter were required to achieve approximately 2.0 log10 units of inactivation with oocysts from different sources. Finally, the study compared the relationship between easily measured indicators, includingBacillus subtilis (aerobic) spores andClostridium sporogenes (anaerobic) spores, and C. parvum oocysts. The bacterial spores were found to be more sensitive to chlorine dioxide than C. parvum oocysts and therefore could not be used as direct indicators of C. parvum inactivation for this disinfectant. In conclusion, it is suggested that future studies address issues such as oocyst purification protocols and the genetic diversity of C. parvum, since these factors might affect oocyst disinfection sensitivity.

scholarly journals Simulation of Cross-Contamination and Decontamination of Campylobacter jejuni during Handling of Contaminated Raw Vegetables in a Domestic Kitchen

2008 ◽  
Vol 71 (12) ◽  
pp. 2448-2452 ◽  
Author(s):  
LAY-CHING CHAI ◽  
HAI-YEN LEE ◽  
FARINAZLEEN MOHD GHAZALI ◽  
FATIMAH ABU BAKAR ◽  
PRADEEP KUMAR MALAKAR ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Tap Water ◽  
Hot Water ◽  
Wash Water ◽  
Most Probable Number ◽  
Cross Contamination ◽  
Probable Number ◽  
Transfer Rates ◽  
Log Reduction ◽  
Potential Risk Factors

Campylobacter jejuni was found to occur at high prevalence in the raw salad vegetables examined. Previous reports describe cross-contamination involving meat; here we investigated the occurrence of cross-contamination and decontamination events in the domestic kitchen via C. jejuni–contaminated vegetables during salad preparation. This is the first report concerning quantitative cross-contamination and decontamination involving naturally contaminated produce. The study was designed to simulate the real preparation of salad in a household kitchen, starting with washing the vegetables in tap water, then cutting the vegetables on a cutting board, followed by slicing cucumber and blanching (heating in hot water) the vegetables in 85° water. Vegetables naturally contaminated with C. jejuni were used throughout the simulation to attain realistic quantitative data. The mean of the percent transfer rates for C. jejuni from vegetable to wash water was 30.1 to 38.2%; from wash water to cucumber, it was 26.3 to 47.2%; from vegetables to cutting board, it was 1.6 to 10.3%; and from cutting board to cucumber, it was 22.6 to 73.3%. The data suggest the wash water and plastic cutting board as potential risk factors in C. jejuni transmission to consumers. Washing of the vegetables with tap water caused a 0.4-log reduction of C. jejuni attached to the vegetables (most probable number/gram), while rapid blanching reduced the number of C. jejuni organisms to an undetectable level.

scholarly journals Biphasic Modulation of Apoptotic Pathways in Cryptosporidium parvum-Infected Human Intestinal Epithelial Cells

2008 ◽  
Vol 77 (2) ◽  
pp. 837-849 ◽  
Author(s):  
Jin Liu ◽  
Mingqi Deng ◽  
Cheryl A. Lancto ◽  
Mitchell S. Abrahamsen ◽  
Mark S. Rutherford ◽  
...  
Keyword(s):  
Cryptosporidium Parvum ◽  
Expression Patterns ◽  
Nuclear Condensation ◽  
Successful Infection ◽  
Cell Gene Expression ◽  
Infected Cells ◽  
Apoptotic Pathways ◽  
Induced Apoptosis ◽  
The Impact

ABSTRACT The impact of Cryptosporidium parvum infection on host cell gene expression was investigated by microarray analysis with an in vitro model using human ileocecal HCT-8 adenocarcinoma cells. We found changes in 333 (2.6%) transcripts at at least two of the five (6, 12, 24, 48, and 72 h) postinfection time points. Fifty-one of the regulated genes were associated with apoptosis and were grouped into five clusters based on their expression patterns. Early in infection (6 and 12 h), genes with antiapoptotic roles were upregulated and genes with apoptotic roles were downregulated. Later in infection (24, 48, and 72 h), proapoptotic genes were induced and antiapoptotic genes were downregulated, suggesting a biphasic regulation of apoptosis: antiapoptotic state early and moderately proapoptotic state late in infection. This transcriptional profile matched the actual occurrence of apoptosis in the infected cultures. Apoptosis was first detected at 12 h postinfection and increased to a plateau at 24 h, when 20% of infected cells showed nuclear condensation. In contrast, experimental silencing of Bcl-2 induced apoptosis in 50% of infected cells at 12 h postinfection. This resulted in a decrease in the infection rate and a reduction in the accumulation of meront-containing cells. To test the significance of the moderately proapoptotic state late in the infection, we inhibited apoptosis using pancaspase inhibitor Z-VAD-FMK. This treatment also affected the progression of C. parvum infection, as reinfection, normally seen late (24 h to 48 h), did not occur and accumulation of mature meronts was impaired. Control of host apoptosis is complex and crucial to the life of C. parvum. Apoptosis control has at least two components, early inhibition and late moderate promotion. For a successful infection, both aspects appear to be required.

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