Application of Amplon in combination with peroxyacetic acid for the reduction of nalidixic acid-resistant Salmonella Typhimurium and Salmonella Reading on skin-on, bone-in tom turkey drumsticks.

Peroxyacetic acid (PAA) has become an important component of pathogen reduction in poultry processing, but there are potential concerns for continued exposure. The objective was to evaluate the effects of PAA and Amplon (AMP) used alone or in the combination. Bone-in tom turkey drumsticks (N = 100, n = 10, k = 5, 0 and 24 h) per study were obtained and inoculated with either nalidixic acid-resistant Salmonella Typhimurium or Salmonella Reading (64 μg/mL). The inocula were allowed to adhere to the drums at 4°C for 60 min for a final attachment of 108 and 107 cfu/g per S. Typhimurium and S. Reading, respectively. Drumsticks were treated with a no-treatment control; tap water, pH 8.5 (TW); TW+500 ppm PAA, pH 3.5 (PAA); TW+500 ppm AMP, pH 1.3 (AMP); TW + PAA + AMP (PAA + AMP). Treatments were applied as short duration dips (30 s) and allowed to drip for 2 min. After treatment, drums were stored at 4°C until microbial analyses at 0 and 24 h. Drums were rinsed in neutralizing buffered peptone water and spot plated for total aerobes and Salmonella. Bacterial counts were log10 transformed and analyzed using n-way ANOVA. All treatments reduced S. Reading on turkey legs at both 0 and 24 h (P < 0.0001; P < 0.0001). At 24 h, drums treated with PAA + AMP (3.92 log10 cfu/g) had less S. Reading than no-treatment control, TW, and AMP. Treatment by time interactions were observed for total aerobes among drums in both studies (P < 0.0001, P < 0.0001) and Salmonella among drums inoculated with S. Typhimurium (P < 0.0001). During the S. Reading and S. Typhimurium study, all treatments reduced Salmonella and total aerobes on drums. During the S. Typhimurium study, drums treated with PAA + AMP had the lowest numerical load of S. Typhimurium and total aerobes. The combination of AMP + PAA may exhibit a synergistic effect in reducing Salmonella on turkey drums, thus increasing the safety of turkey products for consumers.

Introduction

Nearly fifty million people are affected by foodborne illness in the United States yearly, with some cases being fatal (CDC, 2019a). Salmonella is the etiological agent of Salmonellosis and results in significant bacterial foodborne disease outbreaks annually (CDC, 2019a). Salmonella is most commonly transmitted through the consumption of poultry products with approximately 4 of 5 Salmonella outbreaks associated with meat products in 2018 in poultry-related products (Chai et al. 2017; CDC, 2019b). Thus, federal regulatory oversight agencies that safeguard the US food supply, such as the Center for Disease Control, have set objectives for reducing Salmonella related infections. Specifically, the Center for Disease Control aimed to lower Salmonella-related infections to 11.4 cases per 100,000 people by 2020, while currently developing more stringent goals for 2030 (Healthy People, 2020). One tool the poultry industry uses to help reduce Salmonella and improve food safety on poultry products is antimicrobials applied at the processing plant.

Because different chemicals have various mechanisms that they exhibit on particular bacteria, the combination of the sanitization agents may ultimately have a better outcome for reducing pathogens (Bacon et al., 2000; Stopforth et al., 2007). Furthermore, the combination of various processing antimicrobials such as organic and inorganic acids may benefit processors by eliminating the potential for noxious residues and carcass discoloration as compared with other antimicrobials (Kim et al., 2017; Landrum et al. 2019). The industrial use of up to 2,000 ppm of peroxyacetic acid (PAA) in poultry manufacturing has been officially permitted by the USDA and Food Safety and Protection Services and is regarded as generally recognized as safe (21 CFR 173.370; FSIS, 2019; Kim et al., 2017; Moore et al., 2017). The antimicrobial PAA is an organic acid and a combination of acetic acid and hydrogen peroxide that degrades into acetic acid, oxygen, and water without imparting a reduction in meat quality (Kim et al., 2017; Moore et al., 2017). Although organic acids are efficient and function by reducing the pH in the immediate bacterial environment, gram-negative microorganisms can develop resistance (Dittoe et al., 2019).

Salmonella is capable of adapting to and surviving acidification from organic acids owing to the presence of lipopolysaccharides in the outer membrane as well as other mechanisms (Ricke, 2003; Dittoe et al., 2019). However, acid resistance acquired by bacteria from organic acid acidified environments does not exhibit resistance to inorganic acids, as Salmonella requires the production of additional proteins to control the pH homeostasis (Foster et al., 1991; Dittoe et al., 2019). Consequently, inorganic acids remain strong sanitization agent alternatives to PAA (Dittoe et al., 2019). A registered inorganic mix of sulfuric acid and sodium sulfate sold as Amplon by Zoetis (Florham Park, NJ) is used as an accepted antimicrobial, acidifier, and processing aid in meat manufacturing that shows the ability to reduce pathogen contamination in poultry-associated products while keeping its organoleptic properties (Kim et al., 2017). Similar to PAA, the constituents of Amplon are considered generally recognized as safe under the Food and Drug Administration (FSIS 7120.1), and are registered for the antimicrobial use throughout poultry processing via spray, wash, or dip application (Kim et al., 2017; Moore et al., 2017).

While extensive research has been conducted with chicken processing and potential interventions, less is known for turkey processing. Therefore, it is an objective of the present study to examine the potential of Amplon as an antimicrobial intervention in turkey production by determining the efficacy of Amplon alone (500 ppm) or in combination with PAA (500 ppm) on the frequency of Salmonella Reading and Salmonella Typhimurium on skin-on, bone-in turkey drumsticks. Commonly, processing aids and antimicrobials are tested against S. Typhimurium as an industry standard. However, S. Reading is becoming a significant problem linked to raw turkey products (Tanguay et al., 2017; CDC, 2019b). Consequently, we evaluated the efficacy of Amplon-acidified PAA as a short-duration antimicrobial part dip by demonstrating the reduction of S. Typhimurium and S. Reading. The study aimed to provide the industry with further intervention strategies to mitigate poultry-associated Salmonellosis.

Materials and methods

Drum Leg Procurement and Indigenous Pathogen Screening

One hundred bone-in, skin-on drum tom turkey legs were obtained for the study evaluating S. Typhimurium (study 1; N = 100, n = 10, k = 5, 0 and 24 h) and another 100 for the studies evaluating S. Reading (study 2; N = 100, n = 10, k = 5, 0 and 24 h). The turkey drumsticks, with an average weight of 1,023.59 g (SEM = 15.20) and 1,118.12 (SEM = 92.16), were obtained from a poultry facility immediately after processing. These turkey legs were acquired from a commercial partner and were killed as a component of normal industrial turkey processing and were, therefore, institutional animal care and use committee exempted. All industry and federal guidelines were followed. Immediately after processing, the drum turkey legs were shipped to the University of Arkansas Center for Food Safety, where 1 turkey leg was screened for the presence of indigenous Salmonella per study. The rest were stored at 4°C refrigeration for 3 to 4 h (4 h maximum) until the onset of the study.

Inoculum Preparation and Inoculation

To assure the validity of the antimicrobials against both Salmonella species, the studies were performed on separate d: In study 1, only S. Typhimurium was used; in study 2, only S. Reading was used. Between 12 and 16 h before each study started, the isolated cultures were inoculated using the following methodology and grown overnight. For simplification purposes, all methodologies in the following are described uniformly, as the methods used were the same across both studies.

Preparation of Nalidixic Acid–Resistant Salmonella Species

Frozen and pure stock cultures of S. Typhimurium (ATCC 19585) and S. Reading were streaked to isolation under aerobic conditions at 37°C for 24 h on Xylose Lysine Dextrose (XLD; HiMedia, West Chester, PA) media before the study. The S. Reading culture was part of the Center for Food Safety's collection of stock cultures, and the culture was initially isolated from turkey. To prepare 64 μg/mL nalidixic acid (NA)–resistant strains, 10 mL of sterile 1X PBS (8 g of NaCl, 0.2 g of KCl, 1.44 g of Na2HPO4, and 0.24 g of KH2PO4 per 1 L, with the pH adjusted to 7.4 with HCl) and 64 mg/mL of NA (higher than the breaking point for resistance) were combined in a sterile 15-mL conical tube to prepare 64 μg/mL stock solution. The solution was then serially diluted (1:10) in 9 mL of sterile Mueller Hinton Broth (Hardy Diagnostics, Irving, TX). These tubes were then inoculated with either S. Typhimurium or S. Reading and grew overnight at 37°C. The following day, serial dilutions were performed again, with the highest dilution with NA-resistant surviving strains inoculated into higher NA dilutions and incubated overnight at 37°C. The method was repeated until the cultures survived in 64 μg/mL of NA.

Once cultures grew in Mueller Hinton Broth with 64 μg/mL of NA, the resistance was confirmed by streaking on XLD plates prepared with 64 μg/mL of NA (XLD + NA). One colony was isolated from the incubated plates and streaked to isolation again using the methods described previoulsy. This was repeated once more to confirm the presence of Salmonella.

Inoculation

Isolated colonies from each plate were transferred to 10 capped, sterile 500-mL bottles containing 400 mL (for a total of 4 L for each Salmonella serovar) each of Mueller Hinton Broth (Hardy Diagnostics, Irving, TX). The cultures were placed in a shaking incubator at 37°C for 12 to 16 h at 200 rpm. Immediately after the overnight incubation of the cultures, 40 mL of the cultures aliquoted to separate 50-mL conical tubes and centrifuged at 18,000 g for 3 min to collect the pellet, decanted, washed, and resuspended in 1X PBS (8 g of NaCl, 0.2 g of KCl, 1.44 g of Na2HPO4, and 0.24 g of KH2PO4 per 1 L, with the pH adjusted to 7.4 with HCl) and recentrifuged. A total of 2 washes were performed. After the final wash, the pellets were resuspended in 40 mL of sterile PBS.

The drum turkey legs were inoculated using 550 mL of inoculum per 10 legs (1 mL of inocula per 20 g of turkey). To determine the effects of the treatments against both strains, the first 100 legs were inoculated with only S. Typhimurium (study 1), while the remaining 100 legs were inoculated with S. Reading (study 2). The inocula were allowed to adhere at 4°C for 60 min for a final attachment level of 108 cfu/g. After the attachment period, the weights of each leg were recorded, and the treatments were administered. In poultry rinse bags, a drum turkey leg and 500 mL of the corresponding treatment were transferred and agitated to allow full coverage for 30 s by shaking using a 90° arcing motion. The treatments were decanted, and the legs were allowed to drip for 2 to 3 min before proceeding to the next step. To reduce the risk of cross contamination, each step of the treatment occurred in individual sterile poultry rinse bags. The treatments used were a no-treatment control (NTC); tap water (TW, pH 8.5); TW + 500 ppm PAA (PAA, pH 3.5); TW + 500 ppm Amplon (AMP, pH 1.3); TW + PAA + AMP (PAA + AMP). The commercial PAA used in the present study was Actrol Max (22% PAA; Zoetis, Florham Park, NJ).

Microbial Analysis

After the allotted resting period, 400 mL of neutralizing Buffered Peptone Water (pH 7.7; 20.0 g of buffered peptone, 7 g of refined soy lecithin or equivalent, 1.0 g of sodium thiosulfate, 12.5 g of sodium bicarbonate, per 1 L of DI water; USDA Food Safety and Inspection Service, 2016) was poured into each bag on top of the drum turkey legs. The legs were manually agitated for 1 min using a 180° arcing motion. The legs were aseptically removed from the bags and discarded, while the rinsates were used for downstream analysis. Exactly 1 mL of each biological replicate was aliquoted to 2-mL microcentrifuge tubes with subsequent 20 μL being serially diluted (1:10) to 10−7 in 180 μL of 1X PBS in a flat-bottom 96-well plate. Using the drop method for plating, 10 μL of the rinsate was spot plated on XLD + NA and Tryptic Soy Agar (EMD Millipore Corporation, Billerica, Massachusetts) and allowed to dry completely before inverting (Thomas et al., 2015). The plates were inverted and incubated aerobically at 37°C for 24 h. On XLD + NA, only black colonies were identified as Salmonella isolates.

Statistical Analysis

Each turkey leg was randomly assigned to a treatment and a time point before analyses. Duplicate plates were averaged before analyses. CFU/mL was transformed on a cfu/g basis with the following equation as described by Dittoe et al. (2019).

The cfu/g of Salmonella were log10 transformed and reported on a log10 cfu of S. Typhimurium and S. Reading per g of turkey drumstick (log10 CFU/g of drumstick). Data were analyzed using JMP 14 (Cary, NC) and analyzed using a two-way ANOVA (treatment and time). If no interaction existed between treatment and time, 1-way ANOVA was performed to evaluate the effect of treatment and time, separately. In addition, a 1-way ANOVA of treatments at each time (0 and 24 h) was performed if no interaction existed (treatment and time). Data were considered significant at a P value of less than or equal to 0.05.

Results

In the present study, we evaluated whether or not Amplon-acidified PAA reduces NA-resistant S. Typhimurium and S. Reading on poultry drumsticks when applied as a short-duration dip. Results of the present study demonstrated the efficacy of combining AMP and PAA. As such, all treatments were capable of reducing Salmonella on turkey drums inoculated with NA-resistant S. Reading at 0 h (Figure 1A: P < 0.0001) and 24 h (Figure 1B: P < 0.0001). Drums treated with PAA, AMP, and PAA + AMP were not different from one another at 0 h (4.04, 4.44, and 3.84 log10 cfu/g, respectively). Although not significantly different, at 24 h, PAA + AMP resulted in the lowest recoverable load of NA-resistant S. Typhimurium than AMP (3.92 and 4.73 log10 cfu/g, respectively), as well as PAA when used alone (4.40 log10 cfu/g). However, all drums treated with the 3 treatments were different than the controls, NTC, and TW at 0 and 24 h (6.63 and 5.87 log10 cfu/g at 0 h and 6.40 and 5.88 at 24 h log10 cfu/g). Therefore, treatments had an effect overall, but that effect was not unique for each treatment group over time. The lowest load of NA-resistant Salmonella on treated drumsticks was observed among those treated with PAA + AMP at both 0 and 24 h (3.84 and 3.92 log10 cfu/g) compared with the controls (6.63 and 6.40 log10 cfu/g).

Figure 1

The effect of treatment at 0 (A) and 24 (B) h on Salmonella when 64 μg/mL nalidixic acid–resistant S. Reading was artificially inoculated on tom turkey drumsticks. Drumsticks were treated with the following short-duration dip treatments: no -reatment control (NTC), tap water (TW); TW + 500 ppm PAA (PAA); TW + 500 ppm AMP, pH 1.3 (AMP); TW + PAA + AMP (PAA + AMP). Drumsticks were agitated in 500 mL of the treatments for 30 s and allowed to drip for 2–3 min before microbial analysis (A: P < 0.0001, N = 50, n = 10, k = 5; B: P < 0.0001, N = 50, n = 10, k = 5). Baseline attachment of Salmonella Reading at 107 cfu/g or 6.64 log10 cfu/g as represented by NTC at 0 h. Abbreviations: AMP, Amplon; PAA, peroxyacetic acid.

To monitor the overall bacterial load, aerobic plate counts, were taken alongside NA-resistant S. Reading. All treatments reduced bacterial load on inoculated drumsticks (Figure 2, P = 0.02). Although there was not an interaction between time and treatment for Salmonella load (P > 0.05), overall bacterial load was significantly different (P = 0.02) for drums inoculated with NA-resistant S. Reading. Drumsticks rinsed with TW at 0 h (5.89 log10 cfu/g) had less aerobic bacteria than those not treated, NTC, at 0 and 24 h (7.79 and 6.83 log10 cfu/g). Drums treated with PAA + AMP had the lowest load of total aerobic bacteria (4.70 and 4.55 log10 cfu/g at 0 and 24 h) with over a 2 og10 reduction as compared with the controls (7.79 and 6.83 log10 cfu/g at 0 and 24 h). Drums treated with PAA, AMP, and PAA + AMP were not different from one another (4.90 and 4.81; 4.91 and 5.05; 4.70 and 4.55 log10 cfu/g, respectively).

Figure 2

The interaction of treatment and time (0 and 24 h) on the load of total aerobes when 64 μg/mL nalidixic acid–resistant Salmonella Reading was artificially inoculated on tom turkey drumsticks. Drumsticks were treated with the following short-duration dip treatments: no-treatment control (NTC), tap water (TW); TW + 500 ppm PAA (PAA); TW + 500 ppm AMP, pH 1.3 (AMP); TW + PAA + AMP (PAA + AMP). Drumsticks were agitated in 500 mL of the treatments for 30 s and allowed to drip for 2–3 min before microbial analysis (P = 0.02, N = 100, n = 10, k = 5). Solid colored bars represent the mean log10 cfu of aerobes per g at 0 h, whereas diagonal hashed bars represent the mean log10 cfu of aerobes per g at 24 h. Abbreviations: AMP, Amplon; PAA, peroxyacetic acid.

A treatment by time interaction was observed for the load of Salmonella on turkey drums inoculated with NA-resistant S. Typhimurium where all treatments reduced Salmonella (Figure 3, P < 0.0001). The lowest load of NA-resistant S. Typhimurium was observed among turkey legs treated with PAA + AMP and PAA at 0 h (3.79 and 3.49 log10 cfu/g, respectively) compared with those treated with NTC (7.81 log10 cfu/g). Compared with the mean of Salmonella for the inoculated untreated control turkey legs, NTC and TW, all experimental treatments reduced the total load of Salmonella. The drums treated with PAA had the lowest recoverable load of NA-resistant S. Typhimurium at 0 h (3.49 log10 cfu/g), but those treated with PAA + AMP exhibited the sustained mitigation of Salmonella (3.79 and 3.87 log10 cfu/g at 0 and 24 h, respectively), which was not different than the reduction that drums treated with PAA exhibited at 0 h. Therefore, PAA + AMP potentially exhibited a synergistic effect.

Figure 3

The interaction of treatment and time (0 and 24 h) on the load of Salmonella when an antibiotic resistant Salmonella Typhimurium was artificially inoculated on tom turkey drumsticks. Drumsticks were treated with the following short-duration dip treatments: no-treatment control (NTC), tap water (TW); TW + 500 ppm PAA (PAA); TW + 500 ppm AMP, pH 1.3 (AMP); TW + PAA + AMP (PAA + AMP). Drumsticks were agitated in 500 mL of the treatments for 30 s and allowed to drip for 2–3 min before microbial analysis (P < 0.0001, N = 100, n = 10, k = 5). Baseline attachment of S. Typhimurium at 108 cfu/g or 7.81 log10 cfu/g as represented by NTC at 0 h. Solid colored bars represent the mean log10 cfu of Salmonella per g of drumstick at 0 h, whereas diagonal colored hashed bars represent the mean log10 cfu of Salmonella per g of drumstick at 24 h. Abbreviations: AMP, Amplon; PAA, peroxyacetic acid.

There was an interaction between time and treatment on the load of total aerobic bacteria on artificially inoculated turkey drums in the current experiment (Figure 4, P < 0.0001). Drums treated with PAA had the lowest load of aerobic bacteria at 0 h (6.22 log10 cfu/g) compared with the controls, NTC and TW (9.49 and 8.82 log10 cfu/g, respectively) and was not statistically different from the PAA + AMP treatment (7.29 log10 cfu/g). Although PAA showed the greatest bacterial reduction at 0 h for both NA-resistant S. Typhimurium and associated aerobes (3.49 and 6.21 log10 cfu/g, respectively), it exhibited a rebound in numbers of S. Typhimurium and aerobic bacteria at 24 h (4.83 and 7.95 log10 cfu/g). However, the PAA + AMP treatment was able to sustain the aerobic bacterial populations from 0 h (7.28 log10 cfu/g) to 24 h (6.27 log10 cfu/g).

Figure 4

The interaction of treatment and time (0 and 24 h) on the load of total aerobes when 64 μg/mL nalidixic acid–resistant Salmonella Typhimurium was artificially inoculated on tom turkey drumsticks. Drumsticks were treated with the following short-duration dip treatments: no-treatment control (NTC), tap water (TW); TW + 500 ppm PAA (PAA); TW + 500 ppm AMP, pH 1.3 (AMP); TW + PAA + AMP (PAA + AMP). Drumsticks were agitated in 500 mL of the treatments for 30 s and allowed to drip for 2 to 3 min before microbial analysis (P < 0.0001, N = 100, n = 10, k = 5). Solid colored bars represent the mean log10 cfu of aerobes per gram of drumstick at 0 h, whereas diagonal hashed bars represent the mean log10 cfu of aerobes per gram of drumstick at 24 h. Abbreviations: AMP, Amplon; PAA, peroxyacetic acid.

Discussion

In the present study, Amplon-associated treatments reduced the concentration of NA-resistant S. Typhimurium and S. Reading lower than the representative infectious dose, 106 to 108 cfu (Wickham et al., 2007; Chen et al., 2013; Dittoe et al., 2019; FSIS, 2019). The log10 reduction of 4.02 cfu/g of S. Typhimurium and 2.79 cfu/g of S. Reading in this research demonstrated the feasibility of Amplon-acidified PAA to decrease the number of such Salmonella serovars. Thus, the data presented in this study suggest further research into whether or not treated poultry parts with Amplon-acidified PAA could reduce Salmonella contamination in raw turkey parts.

The effects of antimicrobials depend on their dilution, length of interaction with the surface area of the product, and its environmental conditions to demonstrate the antimicrobial effects (Kim et al. 2017; Moore et al., 2017; Landrum et al., 2019). Poultry manufacturers target a 1 log10 cfu/mL bacterial decline throughout each processing step (Mixon, 2020). In the study performed by Scott et al. (2015) on the advantage of Amplon and its antimicrobial effects on chicken wings, the use of Amplon (pH 1.1, 20 s) produced approximately 1 log10 reduction of Salmonella. An analogous study performed with sulfuric acid and sodium sulfate mix (pH 1.5 and 1.0) applied on beef surfaces exhibited significant antimicrobial effects against various strains of Escherichia coli and Salmonella, displaying its effects against various bacterial genera as well as diverse protein matrices, with a significant impact of solution pH on Salmonella load (Scott-Bullard et al., 2017). Alternatively, Bauermeister et al. (2008) investigated the use of PAA (85 ppm) alone and observed the reduction of Salmonella-positive chicken carcasses. In another study carried out by Nagel et al. (2013), the application of PAA in postchill tanks resulted in the most reduction of S. Typhimurium (2 log10 cfu/mL) on whole chicken carcasses than other antimicrobials used in the study.

Furthermore, using any antimicrobials that can produce a 2-log10 decrease should successfully reduce pathogens persisting on carcasses after chilling (Nagel et al., 2013). Rosenquist et al. (2003) determined that a 2 log10 reduction of Campylobacter on poultry carcasses results in a 30-fold decrease in the risk associated with foodborne disease. These findings suggest a possibility that the combination of the 2 antimicrobials might lead to greater results in sustained reduction of bacterial pathogens, thus averting foodborne-related illness.

Amplon and PAA exhibited the same trend throughout the study and were not different from one another. Peroxyacetic acid had an initial reduction but in most cases resulted in a rebounding of aerobic plate counts. S. Reading and S. Typhimurium were reduced differently, although PAA + AMP, AMP alone, and PAA were all effective. The additive effect of PAA may be due to the increased acidification of PAA owing to the combination with Amplon as was seen by Dittoe et al. (2019) who acidified PAA with an inorganic acid, sodium bisulfate. The grouping of Amplon with PAA exhibited analogous trends in reducing Salmonella as previous studies investigating the use of PAA alone (Scott et al., 2015). Similarly, the use of Amplon alone has been demonstrated to be an effective antimicrobial agent against Salmonella in former studies (Schmidt et al., 2012; Scott et al., 2015). The use of the sulfuric acid and sodium sulfate mix on beef trimmings produced a reduction of S. Typhimurium and Salmonella Newport, showing the antimicrobial mixture does not appear to be serovar specific (Geornaras et al., 2012). Dittoe et al. (2019) noted that the combination of PAA and SBS produced the most sustained reduction of Salmonella Enteritidis compared with the use of either PAA or SBS alone. A simultaneous multihurdle approach may hinder the ability of Salmonella to develop resistance to the mixture of inorganic and organic compounds. As determined in the present research, the significant decrease of pathogenic counts produced by Amplon-acidified PAA has the potential to successfully moderate foodborne pathogens. However, its use may be impeded owing to the complex composition of the poultry integument.

Although Amplon-acidified PAA proved a plausible antimicrobial combination in the present study, there is a potential shielding influence of poultry meat and skin that may obstruct the capability of sanitizer on poultry cuts (Dittoe et al., 2019). Poultry integument exhibits a greater buffering effect than its offcuts or the fat tissue itself (Tan et al., 2014). Consequently, the ability of Amplon-acidified PAA may be restrained. In future studies, the application of Amplon-acidified PAA should be investigated using ground meat products to explore the potentially beneficial reduction of bacterial counts owing to the lack of integument.

Conclusions

As determined in the present research, the significant and sustainable decrease of S. Reading counts produced by Amplon-acidified PAA has the potential to successfully moderate foodborne pathogens. Turkey parts treated with Amplon (500 ppm)-acidified PAA (500 ppm) resulted in a 4.02 and 2.79 log10 cfu/g decrease in Salmonella load (S. Thyphimurium and S. Reading), compared with controls. The application of Amplon and PAA alone showed a 2 log10 cfu/g reduction of S. Reading, which nevertheless suggests the potential to be an advantageous tool to further regulate the contamination of poultry parts.

The combinatorial application of these antimicrobial compounds could restrict the ability of Salmonella to adapt and develop resistance. To establish if the antimicrobial activity is similar throughout all general poultry-associated serovars, Amplon-acidified PAA must be tested with other Salmonella serovars. Although the present study did not evaluate the change of pH in the solutions, it will be necessary and economically beneficial for the industry to know the effect of the pH to formulate the most cost-effective and safe antimicrobial mixture. Finally, studies that improve the function of Amplon to reduce Salmonella and establish other possible synergistic mixtures need elucidation. With further studies, continued advances in antimicrobial development for use in poultry processing can be conducted that will diminish the spread of pathogens to the food supply.