Susceptibility of Starved Planktonic and Biofilm Listeria monocytogenes to Quaternary Ammonium Sanitizer as Determined by Direct Viable and Agar Plate Counts

1993 ◽  
Vol 56 (7) ◽  
pp. 573-576 ◽  
Author(s):  
TYH-JENQ REN ◽  
JOSEPH F. FRANK
Keyword(s):  
Listeria Monocytogenes ◽  
Quaternary Ammonium ◽  
Phosphate Buffer ◽  
Benzalkonium Chloride ◽  
Agar Plate ◽  
Viable Count ◽  
Plate Count ◽  
Planktonic Cells ◽  
Plate Counts ◽  
Cleaning Procedures

Effective food plant cleaning procedures remove microbial nutrients from surfaces, which could result in contaminating bacteria being subject to a starvation microenvironment. This research investigated the effect of starvation on the susceptibility of Listeria monocytogenes to benzalkonium chloride (BAC). Cells were starved in phosphate buffer at 21°C for 4 d. Biofilm and planktonic listeriae reacted differently to starvation. When cells were grown in tryptic soy broth (TSB), starvation reduced the susceptibility of planktonic cells to BAC by 2.3- to 4.7-fold but had no effect on the susceptibility of biofilm cells. Planktonic cells grown in diluted TSB were 390 times more resistant than normal TSB-grown cells, but when these cells were starved, they lost their increased resistance. This phenomenon was not observed with biofilm cells. Increased resistance of listeriae grown in diluted TSB was associated with dilution of the salt/buffer components of the medium. Sanitizer-treated cells were enumerated by using tryptic soy agar-yeast extract pour plates and by a direct viable count method. Results indicate that some cells exposed to BAC were not detected by the plate count procedure but were still viable.

A Direct Viable Count Method Suitable for Use With Listeria monocytogenes

1992 ◽  
Vol 55 (9) ◽  
pp. 697-700 ◽  
Author(s):  
JOSEPH F. FRANK ◽  
MUSTAFA A. GASSEM ◽  
REVIS A. N. GILLETT
Keyword(s):  
Listeria Monocytogenes ◽  
Phosphate Buffer ◽  
Membrane Filter ◽  
Test Sample ◽  
Viable Count ◽  
Culture Studies ◽  
Count Method ◽  
Differential Reaction ◽  
Plate Counts ◽  
Direct Microscopic Observation

A method was developed to determine viable Listeria monocytogenes via direct microscopic observation. The test sample is mixed with tryptic soy broth containing 6 g/L yeast extract and 4 μg/L of novobiocin, and incubated for 6 h at 35°C. Cells are then collected on a membrane filter, stained with acridine orange, and elongated cells counted using an epifluorescent microscope. Viable cells enlarged from 0.8 to 1.5 μm to a length of 2.4 to 4.8 μm and did not divide during the incubation. Without novobiocin in the medium, the cell population increased 10-fold. The method, as described, is only useful for pure culture studies since no differential reaction was used. Survival of L. monocytogenes Scott A during starvation at 21°C in 0.8% saline and phosphate buffer diluent was determined using the direct viable count method. Results from using phosphate buffer indicate that the viable count method can detect greater numbers of starved cells than conventional nonselective plate counts.

scholarly journals Advanced Killing Potential of Thymol against a Time and Temperature Optimized Attached Listeria monocytogenes Population in Lettuce Broth

Biomolecules ◽  
2021 ◽  
Vol 11 (3) ◽  
pp. 397
Author(s):  
Dimitra Kostoglou ◽  
Parthena Tsaklidou ◽  
Ioannis Iliadis ◽  
Nikoletta Garoufallidou ◽  
Georgia Skarmoutsou ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Benzalkonium Chloride ◽  
Planktonic Cells ◽  
Antimicrobial Efficiency ◽  
Fresh Vegetables ◽  
Viable Bacteria ◽  
Steel Surfaces ◽  
Polymerase Chain ◽  
Killing Action ◽  
Foodborne Infections

Fresh vegetables and salads are increasingly implicated in outbreaks of foodborne infections, such as those caused by Listeria monocytogenes, a dangerous pathogen that can attach to the surfaces of the equipment creating robust biofilms withstanding the killing action of disinfectants. In this study, the antimicrobial efficiency of a natural plant terpenoid (thymol) was evaluated against a sessile population of a multi-strain L. monocytogenes cocktail developed on stainless steel surfaces incubated in lettuce broth, under optimized time and temperature conditions (54 h at 30.6 °C) as those were determined following response surface modeling, and in comparison, to that of an industrial disinfectant (benzalkonium chloride). Prior to disinfection, the minimum bactericidal concentrations (MBCs) of each compound were determined against the planktonic cells of each strain. The results revealed the advanced killing potential of thymol, with a concentration of 625 ppm (= 4 × MBC) leading to almost undetectable viable bacteria (more than 4 logs reduction following a 15-min exposure). For the same degree of killing, benzalkonium chloride needed to be used at a concentration of at least 20 times more than its MBC (70 ppm). Discriminative repetitive sequence-based polymerase chain reaction (rep-PCR) also highlighted the strain variability in both biofilm formation and resistance. In sum, thymol was found to present an effective anti-listeria action under environmental conditions mimicking those encountered in the salad industry and deserves to be further explored to improve the safety of fresh produce.

Enumeration of Lactobacillus acidophilus with the Agar Plate Count

1978 ◽  
Vol 41 (6) ◽  
pp. 439-442 ◽  
Author(s):  
E. B. COLLINS
Keyword(s):  
Lactobacillus Acidophilus ◽  
Agar Plate ◽  
Plate Count ◽  
Distilled Water ◽  
Anaerobic Conditions ◽  
Dry Ice ◽  
Standard Methods ◽  
Plate Counts

Higher plate counts on MRS agar were obtained under anaerobic conditions for three of four strains of Lactobacillus acidophilus and commercially prepared nonfermented acidophilus milks (products A and B) made with two of the strains. Average values for aerobic counts of products A and B were 87 and 60%, respectively, of those for corresponding anaerobic counts. Incubation of plates poured with MRS agar for 72 ± 3 h at 37 C was sufficient for maximal counts. Two strains and the nonfermented acidophilus milks gave highest counts on Standard Methods Agar (SMA). Greatest numbers of bile-resistant colonies were indicated by MRS agar (with 0.2% oxgall). The average of plate counts for products A and B determined on MRS agar with oxgall was 65% of that for corresponding plate counts determined on MRS without oxgall. Buffered distilled water, 0.9% NaCl, and 1% peptone each served satisfactorily as diluent. Overlaying MRS agar in poured plates with additional medium was not advantageous. Plate counts of samples that had been frozen and stored at −26 C or in dry ice were as high as those of duplicate samples that had been stored at 1.7 C.

Dry Rehydratable Film for Enumeration of Total Aerobic Bacteria in Foods: Collaborative Study

1990 ◽  
Vol 73 (2) ◽  
pp. 242-248
Author(s):  
Michael S Curiale ◽  
Therese Sons ◽  
J Sue Mcallister ◽  
Barbara Halsey ◽  
Terrance L Fox
Keyword(s):  
Collaborative Study ◽  
Agar Plate ◽  
Plate Count ◽  
Relative Standard ◽  
Plate Counts ◽  
Agar Method ◽  
Standard Deviations ◽  
Aerobic Plate Count ◽  
Film Method ◽  
Dry Film

Abstract A rehydratable dry-film plating procedure for aerobic plate counts has been compared to the standard agar plate method (966.23B and C, 15th ed.; 46.014-46.015, 14th ed.) in a collaborative study by 12 laboratories. Each laboratory analyzed the normal microflora of 3 samples in duplicate for 6 products. The aerobic plate counts ranged from 1.0 X 103 to 1.0 X 108 cfu/g. The products were flour, nuts, frozen raw shrimp, spice, frozen raw ground turkey, and frozen and refrigerated vegetables. Repeatability standard deviations of the 2 methods did not differ significantly for 13 of 18 test samples. For 1 shrimp and 2 turkey samples, the dry-film method had lower repeatability variances (P < 0.05) and for 1 spice sample the agar method had lower repeatability variances (P < 0.05). Relative standard deviations of repeatability were between 1.7 and 15.5% for the dry-film method and 1.2 and 16.0% for the agar method. Relative standard deviations of reproducibility ranged from 2.4 to 23.4 % for the dry-film method and 2.3 to 18.8% for the agar method. The dry rehydratable film method has been adopted official first action for determination of the aerobic plate count.

A COMPARISON OF THE EFFECT OF FOUR MILKING MACHINE CLEANING AND SANITIZING PROCEDURES ON THE STANDARD PLATE COUNT OF RAW MILK1

1955 ◽  
Vol 18 (11) ◽  
pp. 284-287
Author(s):  
G. H. Watrous ◽  
E. M. Kesler ◽  
H. V. Atherton
Keyword(s):  
Quaternary Ammonium ◽  
Statistical Difference ◽  
Plate Count ◽  
Standard Plate Count ◽  
Dry Storage ◽  
Standard Plate ◽  
Plate Counts ◽  
Milking Machine ◽  
Water Rinse ◽  
Rack Storage

A study of four milking machine washing and/or sanitizing methods indicates that an adequate washing procedure using the proper alkaline detergent, coupled with dry storage and a warm water rinse before use was as satisfactory as chlorine sanitizing, the use of a quaternary ammonium cleaner sanitizer, or lye rack storage. The plate counts obtained on milk where machines were thus treated showed no statistical difference attributable to the method employed.

Assimilable Organic Carbon Concentrations and Bacterial Numbers in a Water Distribution System

1993 ◽  
Vol 27 (3-4) ◽  
pp. 159-166 ◽  
Author(s):  
R. A. Gibbs ◽  
J. E. Scutt ◽  
B. T. Croll
Keyword(s):  
Seasonal Variation ◽  
Organic Carbon ◽  
Distribution System ◽  
Water Distribution ◽  
Water Distribution System ◽  
Seasonal Pattern ◽  
Agar Plate ◽  
Plate Count ◽  
Spatial And Seasonal Variation ◽  
Plate Counts

A three year study was conducted to investigate bacterial growth in a drinking water distribution system in the UK. Bacterial numbers were estimated using Yeast Extract Agar plate counts. Plate counts in the distribution system showed patterns of spatial and seasonal variation. The spatial pattern was that plate counts increased through the distribution system until approximately 30 to 40 hours retention time and remained constant further through the distribution system. The seasonal pattern was that plate counts were low in the winter and had large peaks in the summer and autumn. Assimi able organic carbon (AOC) concentrations were measured in the second and third years of the study using an adenosine triphosphate (ATP) assay. There appeared to be no relationship tietween AOC concentrations and the spatial and seasonal variation in plate counts. The lack of correlation may have been caused by a lack of sensitivity in the AOC technique. Another explanation is that the increase in plate counts through the distribution system was due to an increase in the culturability of bacteria on plate count media, rather than an increase in bacterial numbers. Bacteria may not have grown through the distribution system and therefore not utilised the AOC.

Controlling Attachment and Growth of Listeria monocytogenes in Polyvinyl Chloride Model Floor Drains Using a Peroxide Chemical, Chitosan-Arginine, or Heat†

2014 ◽  
Vol 77 (12) ◽  
pp. 2129-2132 ◽  
Author(s):  
MARK E. BERRANG ◽  
CHARLES L. HOFACRE ◽  
JOSEPH F. FRANK
Keyword(s):  
Listeria Monocytogenes ◽  
Polyvinyl Chloride ◽  
Hot Water ◽  
Plate Count ◽  
Processing Plant ◽  
Peroxyacetic Acid ◽  
Planktonic Cells ◽  
Log Reduction ◽  
Before And After ◽  
Poultry Processing

Listeria monocytogenes can colonize a poultry processing plant as a resident in floor drains. Limiting growth and attachment to drain surfaces may help lessen the potential for cross-contamination of product. The objective of this study was to compare a hydrogen peroxide-peroxyacetic acid–based chemical to chitosan-arginine or heat to prevent attachment of or destroy existing L. monocytogenes on the inner surface of model floor drains. L. monocytogenes was introduced to result in about 109 planktonic and attached cells within untreated polyvinyl chloride model drain pipes. Treatments (0.13% peroxide-based sanitizer, 0.1% chitosan-arginine, or 15 s of hot water at 95 to 100°C) were applied immediately after inoculation or after 24 h of incubation. Following treatment, all pipes were incubated for an additional 24 h; planktonic and attached cells were enumerated by plate count. All treatments significantly (P < 0.05) lowered numbers of planktonic and attached cells recovered. Chitosan-arginine resulted in approximately a 6-log reduction in planktonic cells when applied prior to incubation and a 3-log reduction after the inoculum had a chance to grow. Both heat and peroxide significantly outperformed chitosan-arginine (8- to 9-log reduction) and were equally effective before and after incubation. Heat was the only treatment that eliminated planktonic L. monocytogenes. All treatments were less effective against attached cells. Chitosan-arginine provided about a 4.5-log decrease in attached cells when applied before incubation and no significant decrease when applied after growth. Like with planktonic cells, peroxide–peroxyacetic acid and heat were equally effective before or after incubation, causing decreases ranging from 7 to 8.5 log for attached L. monocytogenes. Applied at the most efficacious time, any of these techniques may lessen the potential for L. monocytogenes to remain as a long-term resident in processing plant floor drains.

scholarly journals Common Plant-Derived Terpenoids Present Increased Anti-Biofilm Potential against Staphylococcus Bacteria Compared to a Quaternary Ammonium Biocide

Foods ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 697 ◽  
Author(s):  
Dimitra Kostoglou ◽  
Ioannis Protopappas ◽  
Efstathios Giaouris
Keyword(s):  
Quaternary Ammonium ◽  
Benzalkonium Chloride ◽  
Cell Type ◽  
Planktonic Cells ◽  
Common Plant ◽  
Resistance Coefficients

The antimicrobial actions of three common plant-derived terpenoids (i.e., carvacrol, thymol and eugenol) were compared to those of a typical quaternary ammonium biocide (i.e., benzalkonium chloride; BAC), against both planktonic and biofilm cells of two widespread Staphylococcus species (i.e., S. aureus and S. epidermidis). The minimum inhibitory and bactericidal concentrations (MICs, MBCs) of each compound against the planktonic cells of each species were initially determined, together with their minimum biofilm eradication concentrations (MBECs). Various concentrations of each compound were subsequently applied, for 6 min, against each type of cell, and survivors were enumerated by agar plating to calculate log reductions and determine the resistance coefficients (Rc) for each compound, as anti-biofilm effectiveness indicators. Sessile communities were always more resistant than planktonic ones, depending on the biocide and species. Although lower BAC concentrations were always needed to kill a specified population of either cell type compared to the terpenoids, for the latter, the required increases in their concentrations, to be equally effective against the biofilm cells with respect to the planktonic ones, were not as intense as those observed in the case of BAC, presenting thus significantly lower Rc. This indicates their significant anti-biofilm potential and advocate for their further promising use as anti-biofilm agents.

A Comparison of Phosphate Buffered and Distilled Water Dilution Blanks for the Standard Plate Count of Raw-Milk Bacteria1

1975 ◽  
Vol 38 (5) ◽  
pp. 264-268 ◽  
Author(s):  
C. N. HUHTANEN ◽  
A. R. BRAZIS ◽  
W. L. ARLEDGE ◽  
C. B. DONNELLY ◽  
R. E. GINN ◽  
...  
Keyword(s):  
Phosphate Buffer ◽  
Dairy Products ◽  
Raw Milk ◽  
Plate Count ◽  
Sign Test ◽  
Distilled Water ◽  
Standard Plate Count ◽  
Standard Plate ◽  
Plate Counts ◽  
Water Dilution

Raw milk samples were diluted with distilled water or distilled water with added phosphate buffer as recommended by Standard Methods for the Examination of Dairy Products. The standard plate counts were higher in diluent without phosphate buffer with both high and low count milk. The higher counts were significant when analyzed by a nonparametric sign test or a t-test of differences but were not significant with an analysis of variance technique. Reproducibility was not statistically different in the two diluents. It is suggested that the use of phosphate buffer for raw milk bacteria counts be discontinued until information showing definite advantages is provided.

EFFECT OF TIME AND TEMPERATURE OF INCUBATION ON THE PLATE COUNT OF DRY MILKS

1964 ◽  
Vol 27 (8) ◽  
pp. 241-244
Author(s):  
Rafael Pedraja ◽  
Ainis Mengelis
Keyword(s):  
Incubation Time ◽  
Agar Plate ◽  
Plate Count ◽  
Plate Method ◽  
Bacterial Counts ◽  
Grade Classification ◽  
Dry Milk ◽  
Plate Counts ◽  
Nonfat Dry Milk

Bacterial counts of nonfat dry milk of various heat types and of dry buttermilk were determined by the agar plate method using three incubation conditions, 35 C for 48 hours, 32 C for 48 hours, and 32 C for 72 hours. Generally higher plate counts were obtained when incubating samples at 32 C for 72 hours. No outstanding differences were found when samples were incubated at 35 C for 48 hours, as compared to incubation at 32 C for 48 hours. There was no relation between the direct microscopic clump count and differences in plate counts due to varying incubation conditions. Results clearly demonstrate that only a very small percentage of samples were affected in their grade classification as a result of extending the incubation time to 72 hours.

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