Assessment of Microbial and Radioactive Contaminations in Korean Cold Duck Meats and Electron-Beam Application for Quality Improvement

Commercial cold duck meats were purchased from three different supermarkets in different regions in Daegu, Korea. Ten different samples (1 kg/package: 3 replications) were purchased having the same manufacturing date within circulation date. Ten samples (C1-C10) were classified according to their product types (whole, boneless whole/BW, and boneless slice/BS) and smoking process (smoked and non-smoked). The samples were stored in a refrigerator (4°C) for 1 wk prior to use and all experiments were completed within each circulation dates.

Microbial counts of the total aerobic bacteria (TAB), yeasts & molds (Y&M), and coliforms were evaluated according to the guidelines of the Association of Official Analytical Chemists (AOAC, 2000) and expressed as colony forming units (CFU)/g. Nine kinds of foodborne pathogens were analyzed according to Korea Food Code (MFDS, 2011). The condition for enrichment, isolation, and confirmation of 9 kinds of foodborne pathogens is given in Table 1. Meat sample was added to each enrichment broth and incubated at optimal conditions. Colonies isolated from each selective medium were finally identified using VITEK2 Compact (BioMerieux Ind., France).

The commercial cold-duck meat samples were analyzed by gamma-ray spectrometry counting system by using standard analysis method suggested by Association of Analytical Communities with method number 996.05 (AOAC, 1998). The minimum detectable activity for radio-nuclides was quantified at 1 Bq/kg per fresh weight after considering the size and the counting time of the sample.

Analysis was performed according to EN 13783 (2001) to screen whether the samples have been pre-pasteurized. Samples of 10 g were added to peptone saline and diluted 10 times. The solution was diluted by logarithmic dilution series (10¹-10⁷). Each diluted solution was transferred to the filtration manifold tower containing 10 μm polypropylene filter above 0.6 μm polycarbonate filter for DEFT. After that, staining and rinsing steps were performed. For APC (aerobic plate count), 1.0 mL of each dilution was transferred to a Petri dish containing plate count agar (Difco, Laboratories, USA). The plates were incubated at 30°C for 72 h and then expressed as log₁₀ CFU/g. The DEFT count (X) per gram of meat was calculated by using the mean number of DEFT units per microscope field (N/n), the dilution factor (DF) of the sample, and the microscope factor (MF) of the sample Eq. (1).

The DEFT count was then converted to a logarithmic value. The difference between the DEFT count and APC count is then obtained by subtracting the APC count (logarithmic value) from the DEFT count (logarithmic value).

E-beam irradiation was carried out using an electron accelerator (High Energy Linear Accelerator, 10 MeV, EB Tech, Korea) at doses of 0, 1, 3, and 7 kGy, which were applied for determining the reduction effect of the initial microbial populations. The absorbed doses were measured using an alanine-electron paramagnetic resonance dosimetry system, with an EMS 104 EPR analyzer (Bruker Biospin, Germany).

Two pathogens, Salmonella typhimurium (KCTC 1916) and Listeria monocytogenes (KCTC 3569), were grown in a tryptic soy broth (Difco, Laboratories, USA) at 30°C for 48 h. The pathogens were cultured to a cell density of approximately 10⁶-10⁷ CFU/mL levels. One gram of sterile meat samples (30 kGy) were inoculated with cell suspension (200 mL) of the two pathogens, respectively. Then, it was kept in a sterile workstation for 1 min to allow it to be absorbed. The inoculated samples in the stomacher bag were e-beam irradiated from 0 to 3.5 kGy. Ten gram sample was aseptically homogenized in a sterile stomacher bag containing 90 mL sterile saline solution. After serial dilutions, 100 μL aliquot from an appropriate dilution was plated on to the medium. Medium used for the microbial count was tryptic soy agar (Difco, Laboratories, USA). Plates were incubated at the optimal growing temperature of the bacteria for 48 h and the CFU per gram were counted at 30-300 CFU per plate. D₁₀ values (the dose required to inactivate 90% of a population) for each of the organisms tested were determined by the linear fit of the logarithmic survivors versus irradiation dose points (Kim et al., 2007).

All experiments were performed in triplicate. Data were analyzed by using SPSS 19.0. Statistical significance was set to p<0.05. The comparison of the means was conducted using the Duncan’s Multiple Range test.

Ten different cold commercial duck meat samples (C₁-C₁₀) were monitored for total aerobic bacteria (TAB), yeasts & molds (Y&M), and coliform counts, which ranged from 5.18 to 7.23 Log CFU/g, 4.85 to 6.56 Log CFU/g, and 3.72 to 6.04 Log CFU/g, respectively, as shown in Table 2. TAB and Y&M posed a serious microbial threat, and maximum microbial contamination was found in the BS (C₄-C₆) duck meat samples. Seven different samples (C₁-C₇) were highly contaminated with coliforms, whereas the smoked BW (C₈-C₁₀) samples were not contaminated. The contamination level in duck meat has been studied by many researchers. Kim et al. (2016) reported that the refrigerated whole raw duck meat showed lower TAB than that showed by sliced duck meat. The results for TAB and coliforms of whole duck were also in consistent with the findings of Chae et al. (2006) and Sung et al. (2013). Szosland-Fałtyn et al. (2014) suggested that yeasts and molds in whole duck meat were up to 3.77 and 4.45 Log CFU/g, respectively.

The nine foodborne pathogens were also monitored from different cold duck meat samples (Table 2). The pathogen, L. monocytogenes, was detected in all BS duck meat samples (C₄-C₆), while Salmonella spp. was only found in BS (C₆). The other pathogens were not detected in all the 10 different cold duck meat samples. Haslia et al. (2015) depicted that L. monocytogenes and Salmonella spp. pathogens were detected in the raw duck meat products, which is consistent with the results of the present study. Some other studies are also consistent with the present study and demonstrated that some samples out of 32 cooked and raw meat samples showed contamination with Salmonella spp. (Jalali et al., 2008). Szosland-Fałtyn et al. (2014) reported that about 25% and 6% of L. monocytogenes and Salmonella spp. pathogens, respectively, were present in the duck meat sample. Higher contamination of microorganisms, including L. monocytogenes and Salmonella spp., in commercial cold duck meat may be very lethal for the safety status of meat and in return may seriously threaten the human health.

The contents of radionuclides, such as ¹³¹I, ¹³⁷Cs, and ¹³⁴Cs, from different commercial cold duck meat samples are given in Table 3. It is evident from the results that there were no radionuclides detected from any of the 10 different samples. The acceptable limit for each radionuclide in foods is different. The maximum permitted concentration of ¹³⁴Cs and ¹³⁷Cs is less than 100 Bq/kg in general foods according to the Korean Food Code (MFDS, 2011). According to the Ministry of Agriculture, Forestry, and Fisheries, Japan (MAFF), concentration less than 10 Bq/kg in drinking water, less than 50 Bq/Kg in milk and infant foods, and less than 100 Bq/Kg in meat and fish are acceptable for human consumption (Manabe et al., 2016). The results are somehow consistent with the findings of Brandhoff et al. (2016), who reported that different food samples, including dairy and meat, were subjected to the analysis of selected radionuclides in meat but are less than the acceptable limit. The results were further supported by the findings of Miyazaki et al. (2013) who depicted that about 300 samples were determined for the presence of radioactive iodine (¹³¹I) and cesium (¹³⁷Cs and ¹³⁴Cs) and it was found that only few samples showed some contamination, but below the regulatory limits.

DEFT shows the number of both viable and non-viable cells from the samples, while APC shows the number of viable cells only from the sample (Akram et al., 2012). DEFT/APC technique confirms the microbial decontamination by some previous processing operations. When there is no decontamination treatment, the DEFT count can be similar to that of APC. However, mostly the APC count will be lesser than the DEFT count after decontamination treatments because heat or non-heat processes have an ability to inhibit the growth of viable microorganisms (Ahn et al., 2013). If DEFT/APC ratio is higher than 4, it can be an indication of some processing treatment (EN 13783, 2001). The DEFT/APC results of commercial cold duck meat products are shown in Table 4.

The results showed that DEFT count (9.41-11.35) of commercial duck meat samples was higher than the APC (5.15-7.08). The ratios of DEFT/APC for commercial duck meat samples ranged from 3.02 to 6.20. The cold duck meats (C₁-C₆) showed DEFT/APC ratios below 3.76; however, the smoked duck meat samples (C₇-C₁₀) presented DEFT/APC ratio higher than 4. Higher DEFT/APC ratios were demonstrated by smoked duck meat samples constituting significantly higher difference in microbial contamination, which was attributed to pre-pasteurization process, such as smoking (EN 13783, 2001). Osman et al. (2013) reported that DEFT/APC is a microbiological screening method for irradiation treatments based on comparison of counts. They discriminated the fresh and deboned chicken meat samples based on microbial population. Previously, Sommers and Boyd (2006) reported that this method clearly distinguishes the samples having different level of APC among different samples, which is consistent with the findings of our study where different duck meat samples have different value of APC and DEFT.

The microbial qualities of 10 (C₁-C₁₀) commercial cold duck meat samples were evaluated and the sample BS (C₆) showed presence of Salmonella typhimurium and Listeria monocytogenes among all the samples as shown in Table 2. We calculated D₁₀ values from graphical analysis and regression equation and determination of D₁₀ values was carried out in accordance with the reported method of Kortei et al. (2014). The sample BS (C₆) was designated as model sample and treated with different doses (0.5, 1, 1.5, 2, 2.5, 3, 3.5 kGy) of e-beam and D₁₀ values (kGy) of S. typhimurium and L. monocytogenes were determined. The results showed that the D₁₀ values for S. typhimurium and L. monocytogenes were 0.65 kGy and 0.42 kGy, respectively. The regression (R²) for L. monocytogenes and S. typhimurium was 0.9859 and 0.9615, respectively. The level of contamination decreased with increasing the dose level of e-beam irradiation. L. monocytogenes could not be detected after increasing the dose to 2 kGy, but Salmonella was detected even up to 3.5 kGy as shown in Fig. 1. The results are consistent with the findings of Medina et al. (2009) who reported that the loads of L. monocytogenes in cold smoked salmon were reduced with increasing e-beam irradiation. In another study, application of 3 kGy e-beam irradiation resulted in the reduction of the Salmonella load in beef by 2-4 Log CFU/g, which supports findings of the present study (Li et al., 2015).

Fig. 1. D₁₀ values (kGy) of Salmonella typimurium and Listeria monocytogenes inoculated into commercial cold duck meats.

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The microbial quality upon e-beam irradiation of the BS (C₆) cold duck meat is shown in Fig. 2. Four doses (0, 1, 3, 7 kGy) of e-beam were applied to the BS (C₆) cold duck meat sample and the findings indicated that TAB, Y&M, and coliforms were significantly reduced from 9.36 to 3.13 Log CFU/g, 9.19 to 3.00 Log CFU/g, and 3.38 Log CFU/g to the non-detected level, respectively. It is obvious that the initial microbial loads of TAB in the meat sample were effectively reduced in a dose-dependent manner and similar patterns were observed for Y&M and coliforms. Lewis et al. (2002) reported that the use of e-beam at 1.8 kGy significantly reduced the total plate count from 4.60 to 1.62 Log CFU/200 mL in poultry meat. This showed that e-beam irradiation significantly reduced the TAB count. The results are in agreement with the findings of Kim et al. (2013) who reported that application of e-beam dose to pork jerky led to the reduction in TAB count from 4.54 to 2.81 Log CFU/g. Recently, the results were further supported by the reports of An et al. (2017) who depicted that there was reduction in TAB and coliforms were obtained in duck meat.

Fig. 2. Microbial qualities of commercial cold duck meat by e-beam irradiation.

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