A. Hinton

Nisin, rosemary, and ethylenediaminetetraacetic acid affect the growth of Listeria monocytogenes on ready-to-eat turkey ham stored at four degrees Celsius for sixty-three days

The objectives of this study were to determine the anti-Listeria and general antimicrobial properties of nisin, rosemary, and EDTA alone and in combination on Listeria monocytogenes inoculated on ready-to-eat vacuum-packaged diced turkey ham and to ascertain the effects of the treatments on pH and objective color. The turkey hams were cut into 0.5-cm pieces, inoculated with a L. monocytogenes cocktail containing 5 strains of the bacterium, and treated with either no treatment and no inoculum (negative control), inoculum only (positive control), 0.5% nisin, 20 mM EDTA, 1% rosemary, 0.5% nisin + 20 mM EDTA, 0.5% nisin + 1% rosemary, 0.5% nisin + 20 mM EDTA + 1% rosemary, or 20 mM EDTA + 1% rosemary. All samples were vacuum-packaged, stored for 63 d at 4 degrees C +/- 1 degrees C, and analyzed at 1-wk intervals for total aerobes, L. monocytogenes, lactic acid organisms, pH, and objective color. Nisin, nisin with rosemary, nisin with EDTA, and nisin with rosemary and EDTA treatments reduced (P < 0.05) L. monocytogenes counts by 4.42, 4.20, 3.73, and 4.11 log cfu/g when compared with the positive control, respectively, on d 0. Listeria monocytogenes counts remained less than 2.75 log cfu/g for all hams treated with nisin. The EDTA and rosemary treatments alone and in combination were ineffective in inhibiting growth of L. monocytogenes. Although none of the treatments completely eliminated L. monocytogenes, the results indicated that ready-to-eat turkey ham can have significantly decreased L. monocytogenes when treated with nisin alone or in combination with rosemary or EDTA, or both.

Nisin affects the growth of Listeria monocytogenes on ready-to-eat turkey ham stored at four degrees Celsius for sixty-three days

The objectives of this study were to determine pH, anti-Listeria and general antimicrobial properties of nisin on ready-to-eat vacuum-packaged diced turkey ham inoculated with Listeria monocytogenes, and the usage level that would exert maximum antimicrobial effect during 63 d of storage. Ready-to-eat diced turkey ham was inoculated with a 5-strain L. monocytogenes cocktail; treated with 0.2, 0.3, 0.4, and 0.5% nisin treatment solutions; vacuum-packaged; stored at 4+/-1 degrees C for 63 d; and analyzed at 1-wk intervals for total aerobic counts, pH, L. monocytogenes, and lactic acid bacteria. Antimicrobial effectiveness of nisin increased as concentration increased from 0.2 to 0.5%. Aerobic plate counts for 0.4 and 0.5% nisin were lower (P<0.05) than negative and positive controls. All nisin treatments resulted in 4 log reductions (P<0.05) in L. monocytogenes when compared with the positive control on d 0. Four log reductions were also observed on d 7 for 0.4% nisin treatment and d 7 and 14 for 0.5% nisin treatment when compared with the positive control. Listeria monocytogenes counts decreased from 4.97 log cfu/g on d 0 and remained less than 2 log cfu/g through 63 d of storage for the 0.5% nisin treatment. Lactic acid bacteria counts were lower (P<0.05) for 0.5% nisin treatment when compared with positive and negative controls from 28 through 63 d. Except for d 56 and 63, pH was similar (P<0.05) for all treatments. This study revealed that nisin could be used for postprocessing intervention to control L. monocytogenes in ready-to-eat poultry products.

Acousto-Optic Tunable Filter Hyperspectral Microscope Imaging Method for Characterizing Spectra from Foodborne Pathogens

A hyperspectral microscope imaging (HMI) method, which provides both spatial and spectral information of bacterial cells, was developed for foodborne pathogen detection. The AOTF-based hyperspectral microscope imaging system can be effective for characterizing spectral properties of biofilms formed by Salmonella enteritidis as well as Escherichia coli. The intensity of spectral images and the pattern of intensity distribution of spectra varied with system parameters (integration time as well as gain) of the HMI system. Preliminary results demonstrated that determination of optimum parameter values of the HMI system and the integration time, which must be no more than 250 ms, are important for quality image acquisition from biofilms formed by S. enteritidis. Among the 89 contiguous spectral images between 450 and 800 nm, the intensity of images at 458, 494, 522, 550, 574, 590, and 670 nm were distinctive for biofilm of S. enteritidis, whereas the intensity of spectral images at 546 nm was distinctive for E. coli with dark-field illumination with a metal halide light source. For more accurate comparison of intensity from spectral images, a calibration protocol of quantitative intensity comparison needs to be developed to standardize image acquisition using neutral-density filters and multiple exposures. For the identification or classification of unknown foodborne pathogen samples, ground truth region-of-interest pixels need to be selected for spectrally pure fingerprints from various foodborne pathogens such as E. coli and Salmonella species.

Classification and structural analysis of live and dead Salmonella cells using Fourier transform infrared spectroscopy and principal component analysis.

Fourier transform infrared spectroscopy (FT-IR) was used to detect Salmonella Typhimurium and Salmonella Enteritidis food-borne bacteria and to distinguish between live and dead cells of both serotypes. Bacteria cells were prepared in 10(8) cfu/mL concentration, and 1 mL of each bacterium was loaded individually on the ZnSe attenuated total reflection (ATR) crystal surface (45° ZnSe, 10 bounces, and 48 mm × 5 mm effective area of analysis on the crystal) and scanned for spectral data collection from 4000 to 650 cm(-1) wavenumber. Analysis of spectral signatures of Salmonella isolates was conducted using principal component analysis (PCA). Spectral data were divided into three regions such as 900-1300, 1300-1800, and 3000-2200 cm(-1) based on their spectral signatures. PCA models were developed to differentiate the serotypes and live and dead cells of each serotype. Maximum classification accuracy of 100% was obtained for serotype differentiation as well as for live and dead cells differentiation. Soft independent modeling of class analogy (SIMCA) analysis was carried out on the PCA model and applied to validation sample sets. It gave a predicted classification accuracy of 100% for both the serotypes and its live and dead cells differentiation. The Mahalanobis distance calculated in three different spectral regions showed maximum distance for the 1800-1300 cm(-1) region, followed by the 3000-2200 cm(-1) region, and then by the 1300-900 cm(-1) region. It showed that both of the serotypes have maximum differences in their nucleic acids, DNA/RNA backbone structures, protein, and amide I and amide II bands.

Effect of stomaching on numbers of bacteria recovered from chicken skin

Stomaching of skin samples releases only slightly more bacteria than a single rinse. Successive rinses, however, continue to remove almost as many bacteria as the first rinse. One hypothesis to explain this observation is that relatively violent treatment of skin generates smaller pieces of skin, thus increasing the net surface area and effectively sequestering bacteria in a water film on the skin pieces so that numbers of bacteria suspended in the rinsate do not increase. An experiment was conducted to determine whether inoculated marker bacteria are removed from the rinse liquid as skin pieces are stomached and naturally occurring bacteria are released. In each of 4 replications, 5 prechill broiler carcasses were collected from a commercial processing plant. Two 5-g pieces (n = 40) of breast skin were removed from each carcass and placed in a stomacher bag. An inoculum of 30 mL of 0.85% saline solution containing approximately 10(4) of nalidixic acid-resistant Salmonella enterica serovar Typhimurium per milliliter was added to each sample. Skin samples were hand-massaged for 30 s to mix the inoculum, after which a 1-mL aliquot was removed for enumeration of bacteria. A similar sample was taken after 4 min of vigorous stomaching of the skin sample. Bacterial counts recovered from the 30-s hand-massage were 4.3, 2.7, 2.6, and 3.7 log(10) cfu/mL of rinsate for aerobic bacteria, coliforms, Escherichia coli, and Salmonella, respectively. After stomaching, counts were 4.3, 2.9, 2.8, and 3.8, respectively. There was no difference in aerobic plate counts, but mean coliform and E. coli counts were significantly higher (P < 0.05) after stomaching. Numbers of inoculated Salmonella did not decrease. Breaking up the skin into smaller pieces by stomaching did not reduce the number of inoculated bacteria suspended in the rinsate.

Neutralization of residual antimicrobial processing chemicals in broiler carcass rinse for improved detection of Campylobacter1

SUMMARY The USDA‐Food Safety and Inspection Service (FSIS) has established pathogen reduction performance standards for Campylobacter on broiler carcasses. Processors may apply antimicrobial processing aids as a spray or immersion to lower contamination on carcasses. In the United States, broiler carcasses are generally sampled by whole carcass rinse and the potential exists for residual levels of antimicrobial processing aid to be carried over into the rinsate. It has been shown that, if un‐mitigated, such carryover can interfere with the detection of Salmonella. In the current study, we demonstrate that unmitigated carryover of antimicrobial treatment also can interfere with the detection and recovery of Campylobacter in broiler carcass rinse samples. We tested traditional buffered peptone water and found that it did not offer enough neutralizing capability to counteract residual antimicrobial activity of some post‐chill processing aids (peroxyacetic acid, cetylpyridinium chloride, acidified sodium chloride, or a blend of acids) to allow full recovery of Campylobacter. A recently reported formulation for a neutralizing buffered peptone water (currently being used by FSIS) outperformed the traditional carcass rinse medium and allowed significantly improved recovery of Campylobacter even in the presence of 3 of the 4 tested antimicrobial processing aids. Performance of the new carcass rinse medium with the fourth antimicrobial processing aid (acidified sodium chloride) was not different from the traditional formulation. Neutralizing buffered peptone water represents a significant improvement in the broiler carcass rinse method for detection of Campylobacter.

Development of classification models to detect Salmonella Enteritidis and Salmonella Typhimurium found in poultry carcass rinses by visible-near infrared hyperspectral imaging

Salmonella is a major cause of foodborne disease outbreaks resulting from the consumption of contaminated food products in the United States. This paper reports the development of a hyperspectral imaging technique for detecting and differentiating two of the most common Salmonella serotypes, Salmonella Enteritidis (SE) and Salmonella Typhimurium (ST), from background microflora that are often found in poultry carcass rinse. Presumptive positive screening of colonies with a traditional direct plating method is a labor intensive and time consuming task. Thus, this paper is concerned with the detection of differences in spectral characteristics among the pure SE, ST, and background microflora grown on brilliant green sulfa (BGS) and xylose lysine tergitol 4 (XLT4) agar media with a spread plating technique. Visible near-infrared hyperspectral imaging, providing the spectral and spatial information unique to each microorganism, was utilized to differentiate SE and ST from the background microflora. A total of 10 classification models, including five machine learning algorithms, each without and with principal component analysis (PCA), were validated and compared to find the best model in classification accuracy. The five machine learning (classification) algorithms used in this study were Mahalanobis distance (MD), k-nearest neighbor (kNN), linear discriminant analysis (LDA), quadratic discriminant analysis (QDA), and support vector machine (SVM). The average classification accuracy of all 10 models on a calibration (or training) set of the pure cultures on BGS agar plates was 98% (Kappa coefficient = 0.95) in determining the presence of SE and/or ST although it was difficult to differentiate between SE and ST. The average classification accuracy of all 10 models on a training set for ST detection on XLT4 agar was over 99% (Kappa coefficient = 0.99) although SE colonies on XLT4 agar were difficult to differentiate from background microflora. The average classification accuracy of all 10 models on a validation set of chicken carcass rinses spiked with SE or ST and incubated on BGS agar plates was 94.45% and 83.73%, without and with PCA for classification, respectively. The best performing classification model on the validation set was QDA without PCA by achieving the classification accuracy of 98.65% (Kappa coefficient=0.98). The overall best performing classification model regardless of using PCA was MD with the classification accuracy of 94.84% (Kappa coefficient=0.88) on the validation set.

Identification and characterization of Salmonella serotypes using DNA spectral characteristics by fourier transform infrared

Analysis of DNA samples of Salmonella serotypes were performed using FT-IR spectrometer by placing directly in contact with a diamond attenuated total reflection (ATR) crystal. Spectra were recorded from 4000 cm-1 to 525 cm-1 wavenumber with the resolution of 4 cm-1 and data spacing of 1.928 cm-1. Collected spectra were subtracted from the background spectra of empty diamond crystal surface. Principal Component Analysis (PCA) was conducted at four different spectral regions to differentiate the different serotypes of Salmonella on the basis of difference in their spectral features of DNA structure macromolecules. PCA was used to show the natural clusters in the data set and to describe the difference between the sample clusters. At the region 1800 - 1200 cm-1, PC1 distinguished 93 % and PC2 distinguished 7 % of the serotypes. Therefore, maximum classification of 100 % in total was obtained at this region. For all the Salmonella serotypes, the frequency between 1000-1150 cm-1 and 1170 -1280 cm-1 had higher loading values which showed their significant contribution in the serotype classification.

Rapid detection of salmonella using SERS with silver nano-substrate

Surface Enhanced Raman Scattering (SERS) can detect the pathogen in rapid and accurate. In SERS weak Raman scattering signals are enhanced by many orders of magnitude. In this study silver metal with biopolymer was used. Silver encapsulated biopolymer polyvinyl alcohol nano-colloid was prepared and deposited on stainless steel plate. This was used as metal substrate for SERS. Salmonella typhimurium a common food pathogen was selected for this study. Salmonella typhimurium bacteria cells were prepared in different concentrations in cfu/mL. Small amount of these cells were loaded on the metal substrate individually, scanned and spectra were recorded using confocal Raman microscope. The cells were exposed to laser diode at 785 nm excitation and object 50x was used to focus the laser light on the sample. Raman shifts were obtained from 400 to 2400 cm-1. Multivariate data analysis was carried to predict the concentration of unknown sample using its spectra. Concentration prediction gave an R2 of 0.93 and standard error of prediction of 0.21. The results showed that it could be possible to find out the Salmonella cells present in a low concentration in food samples using SERS.