Evaluation of Various Lactic Acid Bacteria and Generic E. coli as Potential Nonpathogenic Surrogates for In-Plant Validation of Biltong Dried Beef Processing

Abstract

Validation studies conducted within a food processing facility using surrogate organisms could better represent the manufacturing process than controlled laboratory studies with pathogenic bacteria on precision equipment in a BSL-2 lab. The objectives of this project were to examine potential surrogate bacteria during biltong processing, conduct biltong surrogate validation lethality studies, and measure critical factors and intrinsic parameters during processing. Beef pieces (1.9 cm × 5.1 cm × 7.6 cm) were inoculated with four-strain mixtures of Carnobacterium divergens/C. gallinarum, Pediococcus acidilactici/P. pentosaceous, and Biotype 1 E. coli ATCC BAA (-1427, -1428, -1429, and -1430), as well as a two-strain mixture of Latilactobacillus sakei and other commercially available individual bacterial cultures (P. acidilactici Saga200/Kerry Foods; Enterococcus faecium 201224-016/Vivolac Cultures). Inoculated beef was vacuum-tumbled in marinade and dried in a humidity-controlled oven for 8–10 days (24.9 °C; 55% relative humidity). Microbial enumeration of surviving surrogate bacteria and evaluation of intrinsic factors (water activity, pH, and salt concentration) were performed post inoculation, post marination, and after 2, 4, 6, 8, and 10 days of drying. Trials were performed in duplicate replication with triplicate samples per sampling time and analyzed by one-way RM-ANOVA. Trials conducted with E. faecium, Pediococcus spp., and L. sakei never demonstrated more than 2 log reduction during the biltong process. However, Carnobacterium achieved a >5 log (5.85 log) reduction over a drying period of 8 days and aligned with the reductions observed in previous trials with pathogenic bacteria (Salmonella, E. coli O157:H7, L. monocytogenes, and S. aureus) in biltong validation studies. Studies comparing resuspended freeze-dried or frozen cells vs. freshly grown cells for beef inoculation showed no significant differences during biltong processing. Carnobacterium spp. would be an effective nonpathogenic in-plant surrogate to monitor microbial safety that mimics the response of pathogenic bacteria to validate biltong processing within a manufacturer’s own facility.

Microbial Profiling of Biltong Processing Using Culture-Dependent and Culture-Independent Microbiome Analysis

“…inoculated beef as potential surrogate bacteria during biltong validation studies, whereby Carnobacterium sp. had >5-log process reduction while L. sakei only achieved a 1.8-2.0-log reduction by day 8 of drying [49].…”

“…post-process (Figure 4B) reflects the ability of this organism to survive the biltong process (i.e., only a ~2 log reduction) better than Carnobacterium sp. (i.e., ~5 log reduction) [49]. It would be interesting to know if these bacteria are contributed to raw beef by contact contamination in their processing plants or transferred directly from the animal during slaughter.…”

“…pre-process (Figure 4A) is likely due to their faster growth rate (tha sakei) during refrigerated storage of raw beef (i.e., Figure 2), whereas the prevalence o sakei/Latilactobacillus sp. post-process (Figure 4B) reflects the ability of this organism survive the biltong process (i.e., only a ~2 log reduction) better than Carnobacterium (i.e., ~5 log reduction) [49]. It would be interesting to know if these bacteria are contribu to raw beef by contact contamination in their processing plants or transferred dire from the animal during slaughter.…”

Behavior of the conversion of glucose, galactose and lactose to lactic acid by kefir grains

“…Bacteria have been employed in the lactic acid production. The main ones are Gram-positive cocci and bacilli, facultative anaerobes, non-sporulated and catalase negative, belonging to the genera Lactobacillus (Nagarajan et al, 2022;Sudhakar et al, 2022), Carnobacterium (Karolenko et al, 2022), Leuconostoc (Sarhir et al, 2023;Liu et al, 2022), Pediococcus (Song et al, 2021;Qiu et al, 2020;Yang et al, 2019), Streptococcus (Wu et al, 2021;Yu et al, 2019), Tetragenococcus (Kobayashi et al, 2004), Lactococcus (Taye et al, 2021;Aso et al, 2019), Vagococcus (Rawoof et al, 2021;Ringø et al, 2020), Enterococcus (Abdel-Rahman et al, 2021;Wang et al, 2020;Hassan et al, 2019), Aerococcus (Mostefa et al, 2021) and Weissella (Nagarajan et al, 2022;Montero-Zamora et al, 2022;Cock and Stouvenel, 2005).…”