Foodborne diseases are mainly caused by bacterial contamination which can lead to severe diarrhea. This study aimed to detect the presence of Shiga toxin-Producing Escherichia coli O157, Escherichia coli non-O157 and virulence gene in raw vegetables. The samples were purchased from wet market and hypermarket in Selangor. The detections were carried out by using the combination methods of Most Probable Number-Polymerase Chain Reaction (MPNPCR). A total of 37(18.5%) samples were found to be contaminated by STEC. Out of these 37 isolates, four (10.8%) of the isolates were E. coli O157 while 33(89.2%) were E. coli nonO157. However, there was no E. coli O157:H7 detected in all the samples. The occurrence of Shiga toxin-Producing E. coli in edible raw vegetables samples suggests the importance of this pathogen in vegetables. Therefore, more studies are required to remove this pathogen from vegetables.
Silver nanoparticles (AgNPs) used in this study were synthesized using pu-erh tea leaves extract with particle size of 4.06 nm. The antibacterial activity of green synthesized AgNPs against a diverse range of Gram-negative foodborne pathogens was determined using disk diffusion method, resazurin microtitre-plate assay (minimum inhibitory concentration, MIC), and minimum bactericidal concentration test (MBC). The MIC and MBC of AgNPs against Escherichia coli, Klebsiella pneumoniae, Salmonella Typhimurium, and Salmonella Enteritidis were 7.8, 3.9, 3.9, 3.9 and 7.8, 3.9, 7.8, 3.9 μg/mL, respectively. Time-kill curves were used to evaluate the concentration between MIC and bactericidal activity of AgNPs at concentrations ranging from 0×MIC to 8×MIC. The killing activity of AgNPs was fast acting against all the Gram-negative bacteria tested; the reduction in the number of CFU mL-1 was >3 Log10 units (99.9%) in 1-2 h. This study indicates that AgNPs exhibit a strong antimicrobial activity and thus might be developed as a new type of antimicrobial agents for the treatment of bacterial infection including multidrug resistant bacterial infection.
The effects of methanolic extract of Javanese turmeric (Curcuma xanthorrhiza Roxb.) at different level of concentrations on the inactivation of Bacillus cereus, Escherichia coli, Pseudomonas spp. and Staphylococcus aureus in oyster mushroom (Pleurotus sajor-caju) were investigated. This study was conducted principally for the achievement on the best combination between the
susceptibility of C. xanthorrhiza extract on natural microflora and foodborne pathogenic bacteria with the sensory acceptability of the soaked oyster mushroom. Three different concentrations (g/ml), 0.05%, 0.50% and 5.00%, of C. xanthorrhiza extract prepared with dilution method were designed as sanitizing agent in treating the oyster mushroom at 5 minutes and 10 minutes.
There was significance reduction in the survival of microbial load between the untreated fresh oyster mushroom and those soaked with 0.05%, 0.50% and 5.00% rhizome extract (P
Bacterial spores have special significance in foods because they are much more resistant to physical and chemical antimicrobial treatment. Nowadays, there is interest in using natural products such as plant extract for food preservation. In this study, 26 of tropical medicinal plants and spices were screened for their sporicidal activity against the spores of Bacillus cereus. The spores of B. cereus was harvested after incubation at 30°C for 1 week and treated with various plant extracts using the method of Standard Operating Procedure for the AOAC (Association of Official Analytical Chemists) Sporicidal Activity. Glutaraldehyde was used as a positive control. Among them, Indonesian bay leaf (Eugenia polyantha Wight) inactivated more than 3 log of spores/ml of B. cereus (99.99%) at the concentration of 1% and completely killed B. cereus spores at concentration of 2.5%. These results suggest that Indonesian bay leaf extract has strong sporicidal activity against spores of B. cereus.
MPT64 is a specific protein that is secreted by Mycobacterium tuberculosis complex (MTBC). The objective of this study was to obtain optimum culture conditions for MPT64 synthetic gene expression in Escherichia coli BL21 (DE3) by response surface methodology (RSM). The RSM was undertaken to optimize the culture conditions under different cultivation conditions (medium concentration, induction time and inducer concentration), designed by the factorial Box-Bhenken using Minitab 17 statistical software. From the randomized combination, 15 treatments and three center point repetitions were obtained. Furthermore, expression methods were carried out in the flask scale fermentation in accordance with the predetermined design. Then, the MPT64 protein in the cytoplasm of E. coli cell was isolated and characterized using sodium dodecyl sulfate polyacrilamide electrophoresis (SDS-PAGE) then quantified using the ImageJ program. The optimum conditions were two-fold medium concentration (tryptone 20 mg/mL, yeast extract 10 mg/mL, and sodium chloride 20 mg/mL), 5 h of induction time and 4 mM rhamnose. The average concentration of recombinant MPT64 at optimum conditions was 0.0392 mg/mL, higher than the predicted concentration of 0.0311 mg/mL. In conclusion, the relationship between the selected optimization parameters strongly influenced the level of MPT64 gene expression in E. coli BL21 (DE3).
In this study, the Mycobacterium tuberculosis protein 64 (MPT64) protein was constructed without any tags to facilitate the purification using column affinity chromatography, but the MPT64 must be obtained as a pure protein. This study was purpose to ensure the efficient extracting method to purify protein MPT64 directly from the polyacrylamide gel. The crude extract of extracellular protein containing MPT64 protein was separated into single protein band and the targeted protein which is located in the size of 24 kDa was excised. Each of the six bands was collected in a sterile microtube to be eluted using electroelution and the optimized of the passive-elution method. Both the elution methods demonstrated the purity level of the MPT64 protein by detecting a solely band on the gel at the 24 kDa. Among the variety of passive-elution time, the highest MPT64 protein concentration was 0.549 mg/ml after elution for 72 h. However, the electroelution result provided higher MPT64 protein concentration, i.e., 0.683 mg/mL. However, based on the recognition of the purified MPT64 protein on commercial detection kit of MPT64 protein, it showed that the positive result was only showed by the passive-elution extracting protein. Therefore, for purifying the protein MPT64 from the sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels, the efficient method was passive elution.
Listeriosis and salmonellosis are the major foodborne illnesses worldwide. Over the last decade,
increasing reports about the antibiotic resistance of Listeria monocytogenes and Salmonella from diverse sources have prompted public health concerns, especially in developing countries with over reliance or misuse of antibiotic drugs in the treatment of humans and animals. In this study, antibiotic susceptibility profiles of 58 L. monocytogenes and 12 Salmonella Enteritidis strains from vegetable farms and retail markets in Malaysia were testedby the standard disk diffusion method. Listeria monocytogenes isolates were found to exhibit 100% resistance to penicillin G. Also, high resistance patterns were observed for meropenem (70.7%) and rifampicin (41.4%). The multiple antibiotic resistance (MAR) index of L. monocytogenes isolates ranged from 0.11 to 0.56. Besides, the antibiogram results revealed that multidrugresistant (MDR) S. Enteritidis were detected and all the S. Enteritidis isolates demonstrated resistance to at least four antibiotics. Ampicillin, amoxicillin, and trimethoprim failed to inhibit all the S. Enteritidis strains. Salmonella Enteritidis isolates also displayed high resistance to nalidixic acid (75.0%), trimethoprim-sulfamethoxazole (75.0%), and chloramphenicol (66.7%). Findings in this study indicated that vegetables could be potential sources of multidrug resistance of L. monocytogenes and S. Enteritidis, which can be a serious issue and a major concern for public health. Thus, there is a great need for surveillance programs in Malaysia to continuously monitor the antibiotic resistance profiles of important pathogens.
Given the remarkable increase of public interest in organic food products, it is indeed critical to evaluate the microbiological risk associated with consumption of fresh organic produce. Organic farming practices including the use of animal manures may increase the risk of microbiological contamination as manure can act as a vehicle for transmission of foodborne pathogens. This study aimed to determine and compare the microbiological status between organic and conventional fresh produce at the retail level in Malaysia. A total of 152 organic and conventional vegetables were purchased at retail markets in Malaysia. Samples were analyzed for mesophilic aerobic bacteria, yeasts and molds, and total coliforms using conventional microbiological methods. Combination methods of most probable number-multiplex polymerase chain reaction (MPN-mPCR) were used to detect and quantify foodborne pathogens, including Escherichia coli O157:H7, Shiga toxin-producing E. coli (STEC), Listeria monocytogenes, Salmonella Typhimurium, and Salmonella Enteritidis. Results indicated that most types of organic and conventional vegetables possessed similar microbial count (P > 0.05) of mesophilic aerobic bacteria, yeasts and molds, and total coliforms. E. coli O157:H7 and S. Typhimurium were not detected in any sample analyzed in this study. Among the 152 samples tested, only the conventional lettuce and organic carrot were tested positive for STEC and S. Enteritidis, respectively. L. monocytogenes were more frequently detected in both organic (9.1%) and conventional vegetables (2.7%) as compared to E. coli O157:H7, S. Typhimurium, and S. Enteritidis. Overall, no trend was shown that either organically or conventionally grown vegetables have posed greater microbiological risks. These findings indicated that one particular type of farming practices would not affect the microbiological profiles of fresh produce. Therefore, regardless of farming methods, all vegetables should be subjected to appropriate post-harvest handling practices from farm to fork to ensure the quality and safety of the fresh produce.
Foodborne diseases continue to pose a significant global health concern, causing a considerable number of deaths worldwide. In response to concerns surrounding bacterial resistance and the limitations of traditional antibiotics, there is a growing interest in exploring natural antibacterial agents as potential alternatives for addressing foodborne pathogens. Nowadays efforts are being made on exploring the potential of natural antibacterial agents against foodborne pathogens. In this study, the antibacterial efficacy of silver nanoparticles synthesized using S. polyanthum leaves extract (SP-AgNPs) against selected Gram-negative and Gram-positive foodborne pathogens was investigated by using multiple assays, including the well diffusion assay (WDA), minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and time-kill assay. The well diffusion assay demonstrated the inhibitory potential of SP-AgNPs against all tested foodborne pathogens, with inhibition zones ranging from 10.16 + 1.25 to 13.16 + 1.52 mm. Furthermore, the MIC values ranged from 0.008 to 0.125 mg/mL, indicating the potent antibacterial activity of SP-AgNPs across a broad spectrum of foodborne pathogens. The MBC values, ranging from 0.008 to 0.250 mg/mL, further confirming the bactericidal ability of SP-AgNPs against these pathogens. In the time-kill experiment, most foodborne pathogens were entirely killed after 4 h of incubation at 4 × MIC concentration. However, only E. coli, K. pneumoniae, and S. Typhimurium showed a reduction in population to 3 Log10 CFU/mL, indicating a strong bactericidal effect of SP-AgNPs on most tested pathogens. In conclusion, this study provides compelling evidence that SP-AgNPs exhibit potent antibacterial activity against selected foodborne pathogens. The findings suggest that SP-AgNPs synthesized using S. polyanthum leaves extract hold great promise as a novel antibacterial agent for effectively controlling foodborne pathogens. These findings contribute to continuing efforts in developing alternative strategies to prevent foodborne diseases and enhance food safety.
Moringa oleifera Lam, family Moringaceae, is a perennial plant which is called various names, but is locally known in Malaysia as "murungai" or "kelor". Glucomoringin, a glucosinolate with from M. oleifera is a major secondary metabolite compound. The seeds and leaves of the plant are reported to have the highest amount of glucosinolates. M. oleifera is well known for its many uses health and benefits. It is claimed to have nutritional, medicinal and chemopreventive potentials. Chemopreventive effects of M. oleifera are expected due to the existence of glucosinolate which it is reported to have the ability to induce apoptosis in anticancer studies. Furthermore, chemopreventive value of M. oleifera has been demonstrated in studies utilizing its leaf extract to inhibit the growth of human cancer cell lines. This review highlights the advantages of M. oleifera targeting chemoprevention where glucosinolates could help to slow the process of carcinogenesis through several molecular targets. It is also includes inhibition of carcinogen activation and induction of carcinogen detoxification, anti-inflammatory, anti-tumor cell proliferation, induction of apoptosis and inhibition of tumor angiogenesis. Finally, for synergistic effects of M. oleifera with other drugs and safety, essential for chemoprevention, it is important that it safe to be consumed by human body and works well. Although there is promising evidence about M. oleifera in chemoprevention, extensive research needs to be done due to the expected rise of cancer in coming years and to gain more information about the mechanisms involved in M. oleifera influence, which could be a good source to inhibit several major mechanisms involved in cancer development.
The use of nanoparticle delivery systems to enhance intracellular penetration of antibiotics and their retention time is becoming popular. The challenge, however, is that the interaction of nanoparticles with biological systems at the cellular level must be established prior to biomedical applications. Ciprofloxacin-cockle shells-derived calcium carbonate (aragonite) nanoparticles (C-CSCCAN) were developed and characterized. Antibacterial activity was determined using a modified disc diffusion protocol on Salmonella Typhimurium (S. Typhimurium). Biocompatibilittes with macrophage were evaluated using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-Bromo-2'-deoxyuridine (BrdU) assays. Transcriptional regulation of interleukin 1 beta (IL-1β) was determined using reverse transcriptase-polymerase chain reaction (RT-PCR). C-CSCCAN were spherical in shape, with particle sizes ranging from 11.93 to 22.12 nm. Encapsulation efficiency (EE) and loading content (LC) were 99.5% and 5.9%, respectively, with negative ζ potential. X-ray diffraction patterns revealed strong crystallizations and purity in the formulations. The mean diameter of inhibition zone was 18.6 ± 0.5 mm, which was better than ciprofloxacin alone (11.7 ± 0.9 mm). Study of biocompatability established the cytocompatability of the delivery system without upregulation of IL-1β. The results indicated that ciprofloxacin-nanoparticles enhanced the antibacterial efficacy of the antibiotic, and could act as a suitable delivery system against intracellular infections.
Increase of foodborne diseases has promulgated the development of new natural food additive
with high extraction yield to eliminate food pathogenic organisms. One such possibility is the
use of plant product as antibacterial agents with non-conventional method to enhance the yield.
In this study, cinnamon leaves (Cinnamomum zeylanicum Blume) were subjected to ultrasonic
assisted extraction (UAE) using response surface methodology (RSM) to optimise extraction
yield and total phenolic contents. The effect of two independent factors, extraction temperature
(x1: 25-40°C) and extraction time (x2: 15-45 minutes) were investigated. Optimum extraction
yield and total phenolic contents of cinnamon leaves were 27.49 ± 1.59% and 3987 ± 79.10 mg
GAE/g which were closely as predicted using RSM (28.34%, 4048 mg GAE/g), respectively.
The optimum condition of extraction yield (40°C and 45 minutes) showed the maximum zone
of inhibition against all tested foodborne pathogens (7.33 ± 0.50 to 13.22 ± 0.44 mm), whereas
optimum condition of total phenolic contents (33°C and 31 minutes) showed the lowest zone
inhibition (6.78 ± 0.67 mm to 11.67 ± 1.41 mm). The minimal inhibitory concentration (MIC)
values range from 97.65 to 6250.00 μg/mL and minimal bactericidal concentration (MBC)
values from 6.25 to 50.00 mg/mL. These results indicated that UAE method is excellent in
producing significantly the highest of extraction yield, total phenolic contents and act as a
potential natural antibacterial agent even using low extraction temperature and short time.
The implementation of the head-only electrical stunning procedure in poultry processing has been aimed at enhancing eating, ethical, and religious quality. However, inconsistencies in voltage and frequency standardization, along with variations in previous research outcomes, have led to numerous cases of both under-stunned and over-stunned birds. Thus, this study aimed to comprehensively evaluate the effects of varying voltages and frequencies during electrical water bath stunning on carcass quality, meat attributes, and textural properties in broiler chickens. A cohort of 240 healthy female broilers (Cobb 500, 42-days-old, 2 kg ± 0.1 kg) was meticulously selected from a commercial farm. The birds underwent exposure to different stunning voltages (2.5, 10.5, 30, and 40 V) and frequencies (50 and 300 Hz). Subsequent analyses were conducted on meat samples to assess physicochemical properties, carcass quality, and textural attributes. The findings revealed a higher incidence of petechial hemorrhage (P < 0.05) in birds stunned at 10.5 V compared to other voltage. Notably, no broken bones were recorded in birds subjected to high voltages (30 and 40 V). Low frequency (50 Hz) significantly increased the occurrence of petechial hemorrhage and simultaneously resulted in pectoralis major muscle with decreased redness (a*). Birds subjected to the 10.5 V stunning treatment exhibited a lower cooking loss percentage. Significant interactions between voltage and ageing (V × A) were observed. Birds stunned at 30 V and aged for 7 d displayed highest drip loss compared to a one-day ageing period across different voltage levels. This interaction also impacted pH values, with birds subjected to 10.5 V showing significantly lower (P < 0.05) pH at d 7 of ageing. The meat hardness was influenced by the V × A interaction, wherein birds stunned at 10.5 V exhibited lower hardness after one day of ageing compared to other voltage levels. Red wing tips, lightness (L*), adhesiveness, and resilience were also significantly impacted (P < 0.05) by the interaction between frequency and voltage. A notable 3-way interaction was observed for gumminess and chewiness (F × V × A), where the 2-way interaction between frequency and voltage (F × V) affected both parameters differently at various ageing periods. Additionally, there was a significant interaction (P < 0.05) between frequency and voltage influencing shear strength and yellowness.
Bacterial infections are regarded as one of the leading causes of fatal morbidity and death in patients infected with diseases. The ability of microorganisms, particularly methicillin-resistant Staphylococcus aureus (MRSA), to develop resistance to current drugs has evoked the need for a continuous search for new drugs with better efficacies. Hence, a series of non-PAINS associated pyrrolylated-chalcones (1-15) were synthesized and evaluated for their potency against MRSA. The hydroxyl-containing compounds (8, 9, and 10) showed the most significant anti-MRSA efficiency, with the MIC and MBC values ranging from 0.08 to 0.70 mg/mL and 0.16 to 1.88 mg/mL, respectively. The time-kill curve and SEM analyses exhibited bacterial cell death within four hours after exposure to 9, suggesting its bactericidal properties. Furthermore, the docking simulation between 9 and penicillin-binding protein 2a (PBP2a, PDB ID: 6Q9N) suggests a relatively similar bonding interaction to the standard drug with a binding affinity score of -7.0 kcal/mol. Moreover, the zebrafish model showed no toxic effects in the normal embryonic development, blood vessel formation, and apoptosis when exposed to up to 40 µM of compound 9. The overall results suggest that the pyrrolylated-chalcones may be considered as a potential inhibitor in the design of new anti-MRSA agents.
Listeria monocytogenes (L. monocytogenes) is a serious food-borne pathogen for immunocompromised individuals. L. monocytogenes is capable of producing biofilm on the surface of food processing lines and instruments. The biofilm transfers contamination to food products and impose risk to public health. Transfers contamination to food products, and impose risk hazard to public health. The aim of this study was to investigate biofilm producing ability of L. monocytogenes isolates. Microtitre assay was used to measure the amount of biofilm production by ten L. monocytogenes isolates from minced chicken / meat, sausages and burgers. Results showed that all 10 L. monocytogenes isolates were able to form biofilm after 24 h at 20˚C on polystyrene surface (the common surface in food industries). Some strains were capable of forming biofilm more than the others. All strains showed a slight raise in the quantities of attached cells over 48 and 72 h. L. monocytogenes strains isolated from minced chicken, minced meat and burgers were better biofilm-producers comparing to the strains isolated from sausages.
Piper cubeba L. is traditionally recognised as flavouring ingredient in various types of foods and has been used to marinate meat. Scientifically, it has been reported to possess various valuable nutritional and pharmacological properties including antimicrobial potential. The aim of the present work was to determine the antibacterial activity of ethanolic P. cubeba L. extract against Escherichia coli and its effect on the microbiological quality of raw chicken meat during storage. Disc diffusion assay was done and resulted in 8.40 ± 0.10 mm of inhibition zone. The bacteriostatic and bactericidal effects of the extract were determined at 0.63 ± 0.00 mg/mL and 1.25 ± 0.00 mg/mL of concentration by MIC and MBC methods, respectively. The killing time was recorded at 2 × MIC (1.25 mg/mL) for 4 h. The application of the extract on chicken meat samples showed reduction in TPC and E. coli count with the observed optimum condition at 5.00% concentration stored at -18°C for 14 days based on the consistent reduction. Sensory attributes acceptability evaluation by 9-point hedonic scale showed acceptable score for colour, odour, texture and overall acceptability of the treated raw chicken meat samples. The findings implies that P. cubeba L. can be listed as one of the alternatives to reduce the bacterial load of raw chicken meat prior to cooking which is very important in ensuring food safety as well as reducing the occurrence of foodborne poisoning associated with chicken meat.
Natural products, either pure compounds or standardized plant extracts, have provided opportunities for the discovery of new drugs. Nowadays, most of the world's population still relies on traditional medicines for healthcare purposes. Plants, in particular, are always used as traditional medicine, as they contain a diverse number of phytochemicals that can be used for the treatment of diseases. The multicomponent feature in the plants is considered a positive phytotherapeutic hallmark. Hence, ethnopharmacognosy has been the focus for finding alternative treatments for diseases. Melicopelunu-ankenda, also known as Euodialunu-ankenda, is widely distributed in tropical regions of Asia. Different parts of M.lunu-ankenda have been used for treatment of hypertension, menstrual disorder, diabetes, and fever, and as an emmenagogue and tonic. It has also been consumed as salad and as a condiment for food flavorings. The justification of use of M.lunu-ankenda in folk medicines is supported by its reported biological activities, including its cytotoxic, antibacterial, antioxidant, analgesic, antidiabetic, and anti-inflammatory activities. This review summarizes the phytochemical compounds isolated from various parts of M.lunu-ankenda, such as root and leaves, and also its biological activities, which could make the species a new therapeutic agent for some diseases, including diabetes, in the future.
Vibrio parahaemolyticus is a main foodborne disease in seafood and generally seafood is
easily deteriorates in quality of color and flavor. In this study, clove (Syzygium aromaticum)
extract shows potent antibacterial activity against growth of antibiotics resistant Vibrio
parahaemolyticus on seafood samples (cockles and shrimps). Vibrio parahaemolyticus was
artificial contaminates on the samples with 106 CFU/ml. The samples were treated with different
concentration of cloves extract with 10 mg/ml which are 0.5%, 5% and 10% concentration
from methanol food grade extraction in 0 hr, 5 min, 10 min, 15 min, 30 min, 60 min and
120 min. Tab water and deionized water were selected as a negative control. As a result, the
amount of 10 % cloves managed to mitigates the number of V. parahaemolyticus on seafood
samples in 5 minutes and 15 min on both samples. Therefore, our results signify the fact that
cloves can be apply as natural sanitizer which could meet consumer demands for safe and
traditionally consumed either raw without any undesirable effect when applied in the seafood
system industries.
The organic foods’ market is becoming one of the rapidly growing sections in agricultural economies in the world. During the last two decades, food-borne outbreaks associated with fresh produce have rapidly increased. E. coli O57:H7, the caustic agent of acute hemorrhagic diarrhea and abdominal cramps, is mainly associated with meat and poultry product outbreaks but frequent outbreaks linked to the consumption of vegetables have been reported. The aim of this study was to investigate prevalence of E. coli O157:H7 in some organic foods. A total of 230 organic food samples including four-winged bean, tomato, white radish, red cabbage, chinese cabbage, lettuce, cucumber and chicken form retailed groceries and supermarkets in Malaysia were investigated. Low prevalence of E. coli O157:H7 was detected in organic vegetables and chickens. The estimated quantity of E. coli O157:H7 in all samples ranged from 2400 MPN/g. The overall MPN/g estimate of E. coli O157:H7 in the samples from organic groceries was higher than supermarket with the maximum of >2400 MPN/g. Most of the samples from supermarket showed a minimum of
Table eggs are an affordable yet nutritious protein source for humans. Unfortunately, eggs are a vector for bacteria that could cause foodborne illness. This study aimed to investigate the effectiveness of a quaternary ammonium compound (quat) sanitizer against aerobic mesophilic bacteria, yeast, and mold load on the eggshell surface of free-range and commercial farms and the post-treatment effect on microbial load during storage. Total aerobic mesophilic bacteria, yeast, and molds were enumerated using plate count techniques. The efficacy of the quaternary ammonium sanitizer (quat) was tested using two levels: full factorial with two replicates for corner points, factor A (maximum: 200 ppm, minimum: 100 ppm) and factor B (maximum: 15 min, minimum: 5 min). Quat sanitizer significantly (p < 0.05) reduced approximately 4 log10 CFU/cm2 of the aerobic mesophilic bacteria, 1.5 to 2.5 log10 CFU/cm2 of the mold population, and 1.5 to 2 log10 CFU/cm2 of the yeast population. However, there was no significant (p ≥ 0.05) response observed between individual factor levels (maximum and minimum), and two-way interaction terms were also not statistically significant (p ≥ 0.05). A low (<1 log10 CFU/cm2) aerobic mesophilic bacteria trend was observed when shell eggs were stored in a cold environment up to the production expiry date. No internal microbial load was observed; thus, it was postulated that washing with quat sanitizer discreetly (without physically damaging the eggshell) does not facilitate microbial penetration during storage at either room temperature or cold storage. Current study findings demonstrated that the quat sanitizer effectively reduced the microbial population on eggshells without promoting internal microbial growth.