An autosomal recessive variant in PYGM causes myophosphorylase deficiency in Red Angus composite cattle.

BACKGROUND: Between 2020 and 2022, eight calves in a Nebraska herd (composite Simmental, Red Angus, Gelbvieh) displayed exercise intolerance during forced activity. In some cases, the calves collapsed and did not recover. Available sire pedigrees contained a paternal ancestor within 2-4 generations in all affected calves. Pedigrees of the calves' dams were unavailable, however, the cows were ranch-raised and retained from prior breeding seasons, where bulls used for breeding occasionally had a common ancestor. Therefore, it was hypothesized that a de novo autosomal recessive variant was causative of exercise intolerance in these calves. RESULTS: A genome-wide association analysis utilizing SNP data from 6 affected calves and 715 herd mates, followed by whole-genome sequencing of 2 affected calves led to the identification of a variant in the gene PYGM (BTA29:g.42989581G > A). The variant, confirmed to be present in the skeletal muscle transcriptome, was predicted to produce a premature stop codon (p.Arg650*). The protein product of PYGM, myophosphorylase, breaks down glycogen in skeletal muscle. Glycogen concentrations were fluorometrically assayed as glucose residues demonstrating significantly elevated glycogen concentrations in affected calves compared to cattle carrying the variant and to wild-type controls. The absence of the PYGM protein product in skeletal muscle was confirmed by immunohistochemistry and label-free quantitative proteomics analysis; muscle degeneration was confirmed in biopsy and necropsy samples. Elevated skeletal muscle glycogen persisted after harvest, resulting in a high pH and dark-cutting beef, which is negatively perceived by consumers and results in an economic loss to the industry. Carriers of the variant did not exhibit differences in meat quality or any measures of animal well-being. CONCLUSIONS: Myophosphorylase deficiency poses welfare concerns for affected animals and negatively impacts the final product. The association of the recessive genotype with dark-cutting beef further demonstrates the importance of genetics to not only animal health but to the quality of their product. Although cattle heterozygous for the variant may not immediately affect the beef industry, identifying carriers will enable selection and breeding strategies to prevent the production of affected calves.

Antimicrobial efficacy of a citric acid/hydrochloric acid blend, peroxyacetic acid, and sulfuric acid against Salmonella and background microbiota on chicken hearts and livers.

This study aimed at evaluating the efficacy of a blend of citric acid and hydrochloric acid (CP), peroxyacetic acid (PAA), and sulfuric acid (SA) against Salmonella and mesophilic aerobic plate counts (APC) on chicken hearts and livers. Samples were inoculated with a five-serovar cocktail of Salmonella at ca. 4.8 log CFU/g and treated by immersion with a water control (90 s), CP (5% v/v, 30 s), PAA (0.05% v/v or 500 ppm, 90 s), or SA (2% v/v, 30 s), all at 4°C and with mechanical agitation. Samples were vacuum packed and stored for up to 3 days at 4°C. Three independent replications were performed for each product, treatment, and time combination. The average Salmonella reductions in chicken hearts after 3 days were 1.33 ± 0.25, 1.40 ± 0.04, and 1.32 ± 0.12 log CFU/g for PAA, SA, and CP, respectively. For chicken livers, the values were 1.10 ± 0.12, 1.09 ± 0.19, and 0.96 ± 0.27 for PAA, SA, and CP, respectively. All antimicrobials reduced Salmonella counts in both chicken hearts and livers by more than one log, in contrast to the water control. All treatments effectively minimized the growth of APC for up to 3 days of refrigerated storage, and no differences in objective color values (L, a, or b) were observed. The poultry industry may use these antimicrobials as components of a multifaceted approach to mitigate Salmonella in nonconventional chicken parts.

Environmental monitoring of Nebraska ready-to-eat meat processing establishments resulted in the isolation of Listeria alongside Pseudomonas highly resistant to quaternary ammonia sanitizer.

Robust environmental monitoring for Listeria monocytogenes often may not be feasible for small and very small meat processors in the United States due to limitations in finances, staffing, or expertise. Three small/very small processors in Nebraska were sampled using sponge applicators in non-food contact surface areas to determine if biofilm and sanitizer resistance behaviors of Pseudomonas could relate to the prevalence of L. monocytogenes and Listeria spp. in ready-to-eat meat processing environments. Samples were 3.3% (3/90) positive for L. monocytogenes, and 12.2% (11/90) of samples were positive for Listeria spp. Pseudomonas spp. were also isolated. When Listeria spp. and Pseudomonas spp. were assayed for biofilm production and resistance to a quaternary ammonia sanitizer, multiple isolates belonging to both genera capable of forming biofilms were identified. Four Pseudomonas spp. isolates resisted the 200 ppm manufacturer recommended sanitizer concentration for food contact surface sanitation, and one Pseudomonas spp. isolated from a drain sample that was also positive for L. monocytogenes demonstrated a sanitizer minimum bactericidal concentration of 1000 ppm. These findings further support the need for monitoring of small and very small meat processors for L. monocytogenes as well as highlight the need to identify other bacteria in these processing environments, like Pseudomonas, that are resistant to environmental stressors.

Effects of relative humidity on dry-aged beef quality.

This study was conducted to evaluate the effects of relative humidity (RH) on moisture loss and flavor in dry-aged beef. Sixteen strip loins were assigned to one of the four aging treatments: vacuum (WET), dry-aging at 50% RH, dry-aging at 70% RH, or dry-aging at 85% RH and aged for 42 days at 2 °C. Loins were evaluated for evaporation loss, trim loss, tenderness, sensory, and microbiological characteristics. Results show that lower RH results in accelerated moisture loss during the first 3 days of the aging process without significantly affecting the total amount of moisture loss. Pseudomonadales dominated the aerobically dry-aged loins while Enterobacteriales was the most abundant in the wet-aged samples. Dry-aged samples had increased content of free amino acids in the cooked meat juice compared to the wet-aged counterpart. Dry aging at 50% RH tended to associate with more desirable flavor notes.

Inactivation of Salmonella in nonintact beef during low-temperature sous vide cooking.

Sous vide cooking is a method of food preparation in which food is vacuum sealed and cooked in a water bath that is set to a precise temperature and circulated by a sous vide device. Due to ease of use and affordability, this cooking method has grown increasingly popular in food service kitchens and domestic settings. However, low-temperature, long holding time sous vide cooking recommendations from manufacturers and chefs in popular press raise food safety concerns - specifically those for the preparation of nonintact beef products. The objective of this experiment was to address these concerns by validating a 5 log reduction of Salmonella spp. in sous vide cooked, nonintact beef steaks. Beef semitendinosus sliced into 2.54 cm steaks were internally inoculated to 7 log with Salmonella Typhimurium, Enteritidis, and Heidelberg via a needle inoculation pin pad. Steaks were individually vacuum sealed, and sous vide cooked at 46.1, 51.6, and 54.4°C. The minimum time measured for a 5 log reduction at 51.6 and 54.4°C was 150 and 64.5 min, respectively (P < 0.01). Additionally, a 7.28 log final reduction was achieved at 51.6°C after 322.5 min (P < 0.01). However, 46.1°C was only able to achieve a final reduction of 2.01 log (P < 0.01) after a holding time of 420 min. The results of this experiment validate in sous vide cooked products the time and temperature combinations provided in the USDA-FSIS Appendix A guidance for a 5 log reduction of Salmonella spp. in meat products. Moreover, more research is needed with other relevant foodborne pathogens to determine if sous vide cooking below Appendix A recommendations could lead to unsafe products.

Spoilage Pseudomonas survive common thermal processing schedules and grow in emulsified meat during extended vacuum storage.

Some Pseudomonas species are common meat spoilage bacteria that are often associated with the spoilage of fresh meat. The recently reported ability of these bacteria to also spoil cooked and vacuum packaged meat products has created the need to investigate all potential routes of spoilage they may be able to utilize. The objective of this experiment was to determine if spoilage Pseudomonas spp. survive thermal processing and grow during refrigerated storage under vacuum. Pseudomonas spp. isolates collected from spoiled turkey products were inoculated into a salted and seasoned meat emulsion that was vacuum sealed and thermally treated to final temperatures of 54.4 and 71.1°C to mimic thermal processes commonly used in the meat industry. Samples were stored for a total of 294 days at 4 and 10°C and plated using Pseudomonas spp. specific agar plates. Pseudomonas spp. concentrations were below the detection limit (0.18 log10 CFU/g) immediately after thermal processing and were first recovered from thermally processed samples after 14 days of storage. The final concentration was greater than 2 log10 CFU/g (p < 0.05 compared to post-thermal processing) in thermally processed treatment groups at the end of storage, indicating that these Pseudomonas spp. isolates were able to survive thermal processing and grow during extended vacuum storage. This raises concerns about the ability of spoilage bacteria to survive the thermal processing schedules commonly used in the meat industry and confirms that some Pseudomonas spp. are capable of thriving in products other than aerobically stored fresh meat. Practical Application: Spoilage Pseudomonas spp. can survive traditional thermal processing schedules. Heat resistance should be evaluated for commensal and spoilage bacteria to better understand possible ways spoilage of food products may occur.

The effects of lactate on nitrosylmyoglobin formation from nitrite and metmyoglobin in a cured meat system.

Two experiments were conducted to determine the effects of lactate on nitrite during meat curing. In the first experiment, using a model system, eight reaction components including nitrite and lactate, were used to assess the effect of each component on metmyoglobin reducing activity by excluding one component at a time. Excluding lactate, nicotinamide adenine dinucleotide (NAD), l-lactate dehydrogenase (LDH) or phenazine methosulfate (PMS) resulted in no reducing activity. A second experiment, utilising a meat mixture, investigated the effects of lactate (0%, 2%, 4% or 6%), nitrite (0 or 156ppm), and packaging (oxygen-permeable or vacuum) on residual nitrite, meat colour and pH. Addition of lactate reduced residual nitrite in the meat mixtures. Both experiments support the hypothesis that lactate generates NADH which then reduces metmyoglobin to deoxymyoglobin. The resulting greater concentration of reduced myoglobin subsequently reacted with nitrite to produce more nitric oxide, reducing nitrite concentration and accelerating curing reactions.

Ultimate pH effects on dry-aged beef quality.

This study aimed to evaluate pH effects on moisture loss and meat quality characteristics of dry-aged beef. Strip loins from six normal pH carcasses (pH = 5.47 ± 0.02) and dark cutting (DC) strip loins from six high pH carcasses (pH = 6.69 ± 0.09) were obtained. One strip loin from each carcass was dry aged and one was wet aged, giving four treatments: DRY, DRY-DC, WET, and WET-DC. Loins were aged for 42 d. Ultimate pH did not affect the rate or amount of moisture loss, trim loss, yield, or tenderness in dry-aged beef (P > 0.05). In general, DC steaks had the lowest lightness (L*), redness (a*), and yellowness (b*) values, regardless of aging method (P < 0.05). Discoloration scores and TBARS values for DC steaks remained low throughout retail display. Dry aging significantly reduced bacterial counts mitigating the microbial damages associated with DC. Flavor characteristics of DC were not improved by dry aging when compared to dry-aged loins from carcasses with normal pH.

Influence of sodium chloride reduction and replacement with potassium chloride based salts on the sensory and physico-chemical characteristics of pork sausage patties.

This study evaluated the effects of sodium chloride reduction and replacement with potassium chloride or modified potassium chloride based salts using a weight or molar equivalent basis on the sensory and physico-chemical properties of pork sausage patties. Three independent replications of pork sausage patties were manufactured to compare five treatments: full sodium, reduced sodium, modified potassium chloride weight based replacement, modified potassium chloride molar based replacement, and standard potassium chloride weight based replacement. Salt replacement did not affect (P>0.05) moisture, protein, fat, textural properties, lipid oxidation, or redness. Sausage patties with modified potassium chloride were more acceptable than those with standard potassium chloride (P<0.001). Using modified potassium chloride replaced on a molar equivalent basis resulted in samples with more similar sensory characteristics to the full sodium control than replacement on a weight equivalent basis. The use of modified potassium chloride reduced sodium and improved sodium:potassium ratios while other changes in composition or physico-chemical characteristics were minimal.

Nitrosylation of myoglobin and nitrosation of cysteine by nitrite in a model system simulating meat curing.

Demand is growing for meat products cured without the addition of sodium nitrite. Instead of the direct addition of nitrite to meat in formulation, nitrite is supplied by bacterial reduction of natural nitrate often added as vegetable juice/powder. However, the rate of nitrite formation in this process is relatively slow, and the total ingoing nitrite is typically less than in conventional curing processes. The objective of this study was to determine the impact of the rate of addition of nitrite and the amount of nitrite added on nitrosylation/nitrosation reactions in a model meat curing system. Myoglobin was preferentially nitrosylated as no decrease in sulfhydryl groups was found until maximum nitrosylmyoglobin color was achieved. The cysteine-myoglobin model retained more sulfhydryl groups than the cysteine-only model (p < 0.05). The rate of nitrite addition did not alter nitrosylation/nitrosation reactions (p > 0.05). These data suggest that the amount of nitrite but not the rate of addition impacts the nitrosylation/nitrosation reactions this system.

Inhibition of Listeria monocytogenes using natural antimicrobials in no-nitrate-or-nitrite-added ham.

Consumer demand for foods manufactured without the direct addition of chemical preservatives, such as sodium nitrite and organic acid salts, has resulted in a unique class of "naturally" cured meat products. Formulation with a natural nitrate source and nitrate-reducing bacteria results in naturally cured processed meats that possess traits similar to conventionally cured meats. However, previous research has shown that the naturally cured products are more susceptible to pathogen growth. This study evaluated Listeria monocytogenes growth on ham manufactured with natural curing methods and with commercially available clean-label antimicrobials (cultured sugar and vinegar blend; lemon, cherry, and vinegar powder blend) and assessed impacts on physicochemical characteristics of the product. Hams made with either of the antimicrobials supported L. monocytogenes growth similar to that in the traditionally cured control (P > 0.05). Hams made with prefermented celery juice powder had the lowest residual nitrite concentrations (P < 0.05), and when no antimicrobial was added, L. monocytogenes growth was similar to that of the uncured control (P > 0.05). Aside from residual nitrite and nitrate concentrations, few physicochemical differences were identified. These findings show that ham can be produced with natural curing methods and antimicrobials to provide similar L. monocytogenes inhibition and physicochemical traits as in traditionally cured ham.

Survey of naturally and conventionally cured commercial frankfurters, ham, and bacon for physio-chemical characteristics that affect bacterial growth.

Natural and organic food regulations preclude the use of sodium nitrite/nitrate and other antimicrobials for processed meat products. Consequently, processors have begun to use natural nitrate/nitrite sources, such as celery juice/powder, sea salt, and turbinado sugar, to manufacture natural and organic products with cured meat characteristics but without sodium nitrite. The objective of this study was to compare physio-chemical characteristics that affect Clostridium perfringens and Listeria monocytogenes growth in naturally cured and traditionally cured commercial frankfurters, hams, and bacon. Correlations of specific product characteristics to pathogen growth varied between products and pathogens, though water activity, salt concentration, and product composition (moisture, protein and fat) were common intrinsic factors correlated to pathogen growth across products. Other frequently correlated traits were related to curing reactions such as % cured pigment. Residual nitrite and nitrate were significantly correlated to C. perfringens growth but only for the ham products.

Survival and growth of Clostridium perfringens in commercial no-nitrate-or-nitrite-added (natural and organic) frankfurters, hams, and bacon.

The popularity of "preservative-free" foods among consumers has stimulated rapid growth of processed meats manufactured without sodium nitrite. The objective of this study was to quantify the potential for Clostridium perfringens growth in commercially available processed meats manufactured without the direct addition of nitrite or nitrate. Commercial brands of naturally cured, no-nitrate-or-nitrite-added frankfurters (10 samples), hams (7 samples), and bacon (9 samples) were obtained from retail stores and challenged with a three-strain inoculation (5 log CFU/g) of C. perfringens. Reduced inhibition (P < 0.05) was observed in seven brands of frankfurters, six brands of hams, and four brands of bacon when compared with each respective sodium nitrite-added control. In naturally cured and truly uncured commercial frankfurters, growth over time was approximately 4.7 log, while conventionally cured frankfurters exhibited growth at 1.7 log. Naturally cured ham and bacon products exhibited growth at 4.8 and 3.4 log, respectively, while their conventionally cured counterparts exhibited growth at 2.6 and 2.3 log, respectively. These products also demonstrated variation in growth response. The results indicate that commercially available natural/organic naturally cured meats have more potential for growth of this pathogen than do conventionally cured products. Natural and organic processed meats may require additional protective measures in order to consistently provide the level of safety from bacterial pathogens achieved by conventionally cured meat products, and which is expected by consumers.

Use of natural ingredients to control growth of Clostridium perfringens in naturally cured frankfurters and hams.

A major concern for processed meats marketed as natural/organic is that they do not contain nitrite in concentrations known to be most effective for inhibiting foodborne pathogens. Supplemental treatments to increase the level and consistency of antimicrobial protection in these products may be important to provide consumers with the degree of safety that they have come to expect from conventionally cured meats. Therefore, the objective of this study was to identify and test ingredients that might improve processed meat product safety without altering their natural/organic status. Eight treatments of hams and frankfurters were prepared: (A) uncured control (typical ingredients except nitrite and nitrate); (B) conventionally cured control (erythorbate, nitrite, and a lactate-diacetate blend); (C) natural nitrate cure (including starter culture containing Staphylococcus carnosus); (D) natural nitrate cure (culture and natural antimicrobial A containing a vinegar, lemon, and cherry powder blend); (E) natural nitrate cure (culture and antimicrobial B containing a cultured sugar and vinegar blend); (F) natural nitrite cure without additional antimicrobials; (G) natural nitrite cure with natural antimicrobial A; and (H) natural nitrite cure with antimicrobial B. For the hams, treatments C, D, E, and H impacted growth of Clostridium perfringens to the same extent (P < 0.05) as the conventionally cured control (approximately 2 log less growth over time than uncured control). For frankfurters, treatments D, G, and H had an effect (approximately 1 log) on growth equivalent to that of the conventionally cured control (P < 0.05). These results suggest that natural/organic cured meats have more potential for pathogen growth than conventionally cured products, but supplemental natural ingredients offer safety improvement.