Foodborne Pathogens Research Articles

Escherichia coli diarrhoea among the adults – prospective research

Introduction: Eschrichia coli is common diarrhea causing agent. This study aimed to identify the bacterial pathogens responsible for diarrhea and abdominal pain in adults. Foodborne illnesses remain a significant public health concern, with Escherichia coli. Methods: The study, conducted from January to May 2021 at GSL Medical College, included adults with diarrhea and abdominal pain. Stool samples were collected, processed, and cultured on selective media to identify bacterial pathogens. Statistical analysis was performed to assess pathogen prevalence and its association with symptoms, using descriptive statistics for findings interpretation. Results: Among 100 adults with diarrhea and abdominal pain, 52% were male, and the majority (40%) were aged 31–45 years. Abdominal pain (80%) and diarrhea (75%) were the most common symptoms. Escherichia coli was the leading isolate (30%), followed by Salmonella spp. (25%) and Staphylococcus aureus (20%). Conclusion: In conclusion, this study highlights Escherichia coli as the primary bacterial cause of diarrhea among adults, followed by Salmonella spp. and Staphylococcus aureus. These findings underscore the importance of targeted food safety measures and public health interventions to address foodborne pathogens and prevent related illnesses in communities. Key words: Escherichia coli, Diarrhea, Foodborne pathogens, Abdominal pain, Bacterial isolation

Supernatant of plant-associated bacteria potency against biofilms formed by foodborne pathogen and food spoilage bacteria.

Food products are often contaminated by pathogens and spoilage bacteria. Most of them can form biofilms, a community of cells embedded in protective extracellular matrix layers resistant to harsh conditions, including antibiotics. Therefore, alternative antibiofilm agents are required to overcome biofilm formation. This study aims to determine and quantify the antibiofilm activity of supernatants from plant-associated bacteria against biofilms of foodborne pathogen and food spoilage bacterium, namely Bacillus cereus and Bacillus subtilis. Plant-associated bacteria (PAB) have shown promising antibiofilm activities against biofilm-forming pathogens in previous studies. Thirteen PAB isolated from Ternate, Indonesia were used in this study. Supernatants of PAB were subjected to antimicrobial activity and quorum quenching detection, both using the well diffusion method. Four supernatants inhibited the growth of B. subtilis, but none affected the growth of B. cereus. Eight supernatants were able to disrupt the quorum sensing system of an indicator bacterium, wild-type Chromobacterium violaceum. Biofilm inhibition and destruction were quantified using 96-well microplates. The highest biofilm inhibition and destruction activities of PAB supernatants against each of B. cereus and B. subtilis biofilms were > 76%, and were later confirmed by light microscope and scanning electron microscope. Brine shrimp lethality assay (BSLA) was conducted and revealed that the selected PAB supernatants were non-toxic. The 16S rRNA gene of PAB were sequenced and they showed similarities to Bacillus, Priestia, and Chryseobacterium. Compounds in the supernatants were determined by GC-MS which revealed contents of fatty acids, ethyl esters, and diketopiperazines. Therefore, PAB supernatants have potential as antibiofilm agents against biofilm formed by Bacillus cereus and Bacillus subtilis.

Characteristics and in vitro properties of potential probiotic strain Fructobacillus tropaeoli KKP 3032 isolated from orange juice.

Fructobacillus, a Gram-positive, non-spore-forming, facultative anaerobic bacterium, belongs to the fructophilic lactic acid bacteria (FLAB) group. The group's name originates from fructose, the favored carbon source for its members. Fructobacillus spp. are noteworthy for their distinctive traits, captivating the interest of scientists. However, there have been relatively few publications regarding the isolation and potential utilization of these microorganisms in the industry. In recent years, F. tropaeoli has garnered interest for its promising role in the food and pharmaceutical sectors, although the availability of isolates is rather limited. A more comprehensive understanding of Fructobacillus is imperative to evaluate their functionality in the industry, given their unique and exceptional properties. Our in vitro study on Fructobacillus tropaeoli KKP 3032 confirmed its fructophilic nature and high osmotolerance. This strain thrives in a 30% sugar concentration, shows resistance to low pH and bile salts, and exhibits robust autoaggregation. Additionally, it displays significant antimicrobial activity against foodborne pathogens. Evaluating its probiotic potential, it aligns with EFSA recommendations in antibiotic resistance, except for kanamycin, to which it is resistant. Further research is necessary, but preliminary analyses confirm the high probiotic potential of F. tropaeoli KKP 3032 and its ability to thrive in the presence of high concentrations of fructose. The results indicate that the isolate F. tropaeoli KKP 3032 could potentially be used in the future as a fructophilic probiotic, protective culture, and/or active ingredient in fructose-rich food.

Foodborne Pathogens Across Different Food Matrices in Sicily (Southern Italy)

Foodborne diseases result from the consumption of foods contaminated with pathogens or their toxins and represent a serious public health problem worldwide. This study aimed to assess the presence of Rotavirus (RoV), Adenovirus (AdV), Norovirus (NoV), Hepatitis A and Hepatitis E viruses (HAV and HEV, respectively), Toxoplasma gondii, Coxiella burnetii and Leptospira spp. across various food matrices in Sicily. The analysis concerned 504 samples, including mussels, farmed meat, game meat, vegetables and bulk milk. Following appropriate pre-treatment, acid nucleic extraction was carried out and amplification of pathogen nucleic acids was carried out by molecular methods. The mussels tested positive for NoVs (3/51, 5.9%) and farm meat resulted positive for T. gondii (1/34, 2.9%). The game offal samples tested positive for HEV, which was detected in 17 out of 222 samples (7.7%), and T. gondii (18/318, 5.7%) and Leptospira spp. (2/318, 0.6%). The milk samples tested positive for C. burnetii (15/ 85, 17.6%), T. gondii (2/85, 2.4%) and Leptospira spp. (1/85, 1.2%). This study highlights the variability in the risk of contamination of different food matrices, confirming the importance of vigilance in the consumption of potentially contaminated food products.

Rosmarinic acid alleviated intestinal barrier damage caused by Escherichia coli by regulating the gut microbiota and inhibiting the NF-κB signalling pathway in mice.

Escherichia coli (E. coli) is a common zoonotic foodborne pathogen that poses a major threat to public health and economic development. Rosmarinic acid (RA) can inhibit intestinal inflammation; however, the protective effect of RA against the intestinal barrier damage induced by E. coli in mice and the underlying mechanism have not been elucidated. In this study, mice were orally administered with RA (20 mg kg-1) by gavage for one week and then were intraperitoneally challenged with E. coli. Mouse colonic epithelial cells (MCECs) were pretreated with RA for 6 h and challenged with E. coli (MOI = 1000) for 3 h. The results revealed that RA alleviated E. coli-induced weight loss in mice; reduced the increase in the levels of TNF-α, IL-6 and IL-1β in the serum; alleviated the decrease in ZO-1 protein expression; and increased intestinal permeability by inhibiting the NF-κB signalling pathway both in vivo and in vitro. Moreover, RA relieved the increase in intestinal permeability, reversed the structural damage to the mouse gut microbiota caused by E. coli, and increased the abundance of beneficial bacteria, including Lachnospiraceae_NK4136_group. Additionally, RA lost its protective function against E. coli infection in a pseudosterile mouse model, suggesting that the protection induced by RA was dependent on the gut microbiota. In conclusion, these results indicate that RA alleviates E. coli-induced inflammatory damage to the intestinal barrier by inhibiting the NF-κB signalling pathway and maintaining gut microbiota homeostasis. These findings provide new ideas and foundations for the application of RA as protection against E. coli.

Genomic Surveillance of Foodborne Pathogens: Advances and Obstacles

Context: The genomic surveillance of foodborne pathogens in the United States has grown exponentially in the past decade, grounded in a powerful combination of novel sequencing technologies, bioinformatic approaches, data-sharing networks, and metadata harmonization efforts. This practice report examines recent advances in genomic epidemiology as applied to food safety programs and delineates State, Tribal, Local, and Territorial infrastructure necessary for continued life-saving improvements in public health. Program: National databases of foodborne pathogen genomes, along with data sharing and evaluation networks such as GenomeTrakr and PulseNet, have transformed how connections are made among isolates and how root causes of outbreaks are determined, allowing much more timely interventions to protect public health. Freely available bioinformatics tools such as GalaxyTrakr and the National Center for Biotechnology Information Pathogen Detection database have allowed laboratories with limited local computing resources to participate in surveillance efforts and contribute to traceback investigations. Implementation: In this report, we describe advances in genomic epidemiology that have occurred over the past decade and examine obstacles to fully implementing this technology within State, Tribal, Local, and Territorial public health systems. Evaluation: Despite a clear return on investment from governmental expenditures on genomic surveillance of foodborne pathogens, we identify significant obstacles to further sustained progress. These obstacles include workforce gaps, ineffective data sharing, and lack of constitutive and sustained funding. Discussion: Many public health laboratories face major obstacles to widespread and routine adoption of genomic surveillance technologies. While whole genome sequencing has become an integral part of routine public health microbiology, the seamless integration of these protocols into the existing practices, laboratory workflows, and information systems remains challenging. Centralized efforts to address these issues include (1) support through the Food and Drug Administration Laboratory Flexible Funding Model, (2) training and proficiency assessments, (3) open-source, standardized protocols for collecting high-quality genomic data, and (4) open access informatics software.

Key roles of two-component systems in intestinal signal sensing and virulence regulation in enterohemorrhagic Escherichia coli.

Enterohemorrhagic Escherichia coli (EHEC) is a foodborne pathogen that infects humans by colonizing the large intestine. Upon reaching the large intestine, EHEC mediates local signal recognition and the transcriptional regulation of virulence genes to promote adherence and colonization in a highly site-specific manner. Two-component systems (TCSs) represent an important strategy used by EHEC to couple external stimuli with the regulation of gene expression, thereby allowing EHEC to rapidly adapt to changing environmental conditions. An increasing number of studies published in recent years have shown that EHEC senses a variety of host- and microbiota-derived signals present in the human intestinal tract and coordinates the expression of virulence genes via multiple TCS-mediated signal transduction pathways to initiate the disease-causing process. Here, we summarize how EHEC detects a wide range of intestinal signals and precisely regulates virulence gene expression through multiple signal transduction pathways during the initial stages of infection, with a particular emphasis on the key roles of TCSs. This review provides valuable insights into the importance of TCSs in EHEC pathogenesis, which have relevant implications for the development of antibacterial therapies against EHEC infection.

Investigating the Antibacterial, Antioxidant, and Cholesterol-lowering Properties of Yogurt Fortified with Postbiotic of Lactobacillus acidophilus and Lactiplantibacillus plantarum in the Wistar Rat Model.

Postbiotics have gained attention in the food industry due to their functional properties and ease of use compared to their live parent cells. Postbiotics are the metabolic byproducts of probiotic microorganisms, offering advantages such as antimicrobial and anti-diabetic effects. The study aimed to explore the potential antibacterial, antioxidant, and cholesterol-lowering effects of postbiotics from Lactobacillus acidophilus (LA) and Lactiplantibacillus plantarum (LbP) through in vitro and in vivo studies. Freeze-dried postbiotics from L. acidophilus BLAC 258 and L. plantarum were used in yogurt to inhibit foodborne pathogens over a 21-day storage period at 4 °C. The cholesterol-lowering effects of the postbiotic yogurt were assessed in Wistar rats fed with Normal Basal Diet (NBD) and High Cholesterol Diet (HCD). All experiments were performed in triplicate, and the collected data were analyzed with a one-way ANOVA using SPSS v.20 (2021) software. The Tukey Honestly Significant Difference (HSD) test was used for means differences at the 95% confidence interval. The results showed that postbiotic-fortified yogurt exhibited significant antioxidant and antibacterial effects. The antioxidant capacity of the yogurt increasingly peaked at 48.81% on day 14. Also, Listeria monocytogenes counts in the postbiotic yogurt decreased by approximately 2 Log10 on day 3. High-cholesterol-fed rats receiving postbiotic yogurt experienced significant reductions in total cholesterol, triglycerides, and LDL levels. Overall results indicate that postbiotics functional yogurt might be a safe and effective strategy for managing cholesterol levels and inhibiting foodborne pathogens.

Conducting polymer functionalized Cu-metal organic framework-based electrochemical immunosensor for rapid and sensitive quantitation of Escherichia coli O157:H7.

Escherichia coli (E. coli) O157:H7 is an important food-borne pathogen that can cause hemorrhagic diarrhea and enteritis in humans and animals. Realizing the rapid quantitation of E. coli O157:H7 is of great significance for the guarantee of food safety and disease control. In this study, an electrochemical immunosensing technique based on a functionalized composite of Cu-metal organic framework (Cu-MOF) and poly (3,4-ethylenedioxythiophene)-poly (styrenesulfonate) (PEDOT:PSS) is developed, achieving rapid and sensitive quantitation of E. coli O157:H7 in food and clinical feces samples. The organic functionalization of Cu-MOF significantly improves the interface conductivity to facilitate electron transfer and provides the sulfonic groups (-SO3H) to conjugate bio-recognizing elements for target determination. The immunosensor delivers a linear detection range of 3 × 102 ~ 3 × 108cfu/mL, a low limit of detection (LOD) of 7.4cfu/mL, and a short analysistime of 40min. In addition, it does not show any cross-reactivity with other common pathogens and exhibits high repeatability with relative standard deviations (RSDs) all lower than 2.09%, providing a promising approach for warranting food safety and control of E. coli O157:H7 disease.

Development of Alginate Hydrogels Incorporating Essential Oils Loaded in Chitosan Nanoparticles for Biomedical Applications

A hybrid alginate hydrogel–chitosan nanoparticle system suitable for biomedical applications was prepared. Chitosan (CS) was used as a matrix for the encapsulation of lavender (Lavandula angustifolia) essential oil (LEO) and Mentha (Mentha arvensis) essential oil (MEO). An aqueous solution of an acidic Natural Deep Eutectic Solvent (NADES), namely choline chloride/ascorbic acid in a 2:1 molar ratio, was used to achieve the acidic environment for the dissolution of chitosan and also played the role of the ionic gelator for the preparation of the chitosan nanoparticles (CS-NPs). The hydrodynamic diameter of the CS-MEO NPs was 130.7 nm, and the size of the CS-LEO NPs was 143.4 nm (as determined using Nanoparticle Tracking Analysis). The CS-NPs were incorporated into alginate hydrogels crosslinked with CaCl2. The hydrogels showed significant water retention capacity (>80%) even after the swollen sample was kept in the aqueous HCl solution (pH 1.2) for 4 h, indicating a good stability of the network. The hydrogels were tested (a) for their ability to absorb dietary lipids and (b) for their antimicrobial activity against Gram-positive and Gram-negative foodborne pathogens. The antimicrobial activity of the hybrid hydrogels was comparable to that of the widely used food preservative sodium benzoate 5% w/v.

The Gene Cluster Cj0423–Cj0425 Negatively Regulates Biofilm Formation in Campylobacter jejuni

Abstract: Campylobacter jejuni (C. jejuni) is a zoonotic foodborne pathogen that is widely distributed worldwide. Its optimal growth environment is microaerophilic conditions (5% O2, 10% CO2), but it can spread widely in the atmospheric environment. Biofilms are thought to play an important role in this process. However, there are currently relatively few research works on the regulatory mechanisms of C. jejuni biofilm formation. In this study, a pan-genome analysis, combined with the analysis of biofilm phenotypic information, revealed that the gene cluster Cj0423–Cj0425 is associated with the negative regulation of biofilm formation in C. jejuni. Through gene knockout experiments, it was observed that the Cj0423–Cj0425 mutant strain significantly increased biofilm formation and enhanced flagella formation. Furthermore, pull-down assay revealed that Cj0424 interacts with 93 proteins involved in pathways such as fatty acid synthesis and amino acid metabolism, and it also contains the quorum sensing-related gene luxS. This suggests that Cj0423–Cj0425 affects fatty acid synthesis and amino acid metabolism, influencing quorum sensing and strain motility, ultimately inhibiting biofilm formation.

Surface monitoring of L. monocytogenes by real-time fluorescence and colorimetric LAMP.

Listeria monocytogenes is a major foodborne pathogen affecting developing, and developed countries. The analysis of food contact surfaces in food industries is key for better controlling this pathogen. The current study focused on the development, optimization, and evaluation of a rapid and simple method for the detection of L. monocytogenes on stainless steel surfaces, suitable for decentralized setups, taking advantage of Loop-mediated isothermal amplification (LAMP). This was accomplished using a general pre-enrichment broth (TSB), with a simple DNA extraction based on a chelating resin, and final isothermal amplification. Two different detection strategies were tested, real-time fluorescence and naked-eye colorimetric, which were evaluated after 5, 7, and 24h of pre-enrichment. Regardless the detection chemistry selected, after 5-7h of pre-enrichment, 103-104CFU/cm2 were needed to obtain a positive result, while after 24h, it was possible to detect concentrations below 10CFU/cm2. Within each given time, all the performance parameters calculated, relative sensitivity, specificity, and accuracy, reached values higher than 80-90%; likewise, a Cohen's k of concordance with a culture-based approach higher than 0.8. Overall, the most sensitive assay can be performed in roughly 25h. This time-to-result outperforms commercial kits with the added value of specifically detecting L. monocytogenes instead of Listeria spp. KEY POINTS: • Real-time fluorescence and naked-eye colorimetric, were compared for the novel assay. • An LOD50 of 3.4CFU/cm2 and 4.2CFU/cm2 was calculated for the two assays. • Three pre-enrichment times were compared providing 24h better results.

CFD Analysis of UV-C Intensity Radiation Distribution and Inactivation of Foodborne Pathogens on Whole and Minimally Processed Mango

Ultraviolet shortwave (UV-C) is a technology for postharvest fruit disinfection. This study aimed to use computational fluid dynamics (CFD) based on the discrete ordinate (DO) radiation model to analyze the distribution of UV-C intensity on whole and minimally processed mangoes within a disinfection chamber and to predict treatments against foodborne pathogens. The mango spears were oriented both parallel and perpendicular to the lamp and positioned at varying distances from the radiation source (50, 75, and 100 mm for spears and 100 mm for whole fruit). CFD simulations integrated with in vitro kinetic parameters enabled predictions of the time and doses needed to inactivate one to three logarithmic units of pathogens on the fruit surface. The highest average radiation intensity values were recorded for the whole mango oriented parallel to the lamp at 100 mm and the spears oriented normally to the lamp at 50 mm. The estimated times to achieve inactivation of one to three logarithmic units of microorganisms ranged from approximately 15 to 6540 s, while the doses necessary for this inactivation were, on average, 1.854, 5.291, and 10.656 kJ/m2, respectively. CFD simulations are valuable for optimizing UV-C treatments in large-scale designing from both microbicide and sustainable perspectives.

Immunogenicity and cross-protective efficacy induced by delayed attenuated Salmonella with regulated length of lipopolysaccharide in mice

ABSTRACT Non-typhoidal Salmonella enterica (NTS) is a major global foodborne pathogen that poses a major public health concern worldwide, and no vaccines are available for protecting against infection of multiple Salmonella serotypes, therefore, the development of Salmonella vaccines to provide broad protection is valuable. In this work, we aimed to regulate lipopolysaccharide (LPS) synthesis of live Salmonella in vivo for exposing conserved protein antigens on the outer membrane while maintaining smooth LPS patterns in vitro to keep their original ability to invade host cells for inducing cross-protection against infection of multiple Salmonella serotypes. We generated a series of mutants defective in genes to affect the length of LPS. These mutants exhibit in vivo regulated-delayed attenuation and altered length of LPS, and all these mutants were derived from SW067 (ΔpagL7 ΔpagP81::Plpp lpxE ΔlpxR9 Δfur9) containing ∆pagP81::Plpp lpxE mutation to reduce their endotoxic activity. Animal experiments demonstrated that all regulated delayed attenuated mutants exhibited reduced ability to colonize the organs of the mice, and SW114 (waaI), SW116 (waaJ), SW118 (waaL), and SW120 (wbaP) induced a significant production of IgG and IgA against OMPs isolated from S. Typhimurium, S. Enteritidis, and S. Choleraesuis. SW114 (waaI), SW116 (waaJ), and SW118 (waaL) were capable of conferring significant protection against infection of wild-type S. Enteritidis and S. Choleraesuis, with SW118 (waaL) triggering significant CD4+ T-cell responses as well as the B220low IgG+ BM cell. In conclusion, regulated delayed attenuated Salmonella vaccines with the whole core oligosaccharides of LPS showed a goo.d ability to expose conserved outer antigens and to trigger strong cross-immune responses against both homologous and heterologous Salmonella infections. These results give new insight into the development of the Salmonella vaccine against multiple serotypes of Salmonella.

Antimicrobial resistance, virulence factors and phylogenetic profiles of Vibrio parahaemolyticus in the eastern coast of Shenzhen

Vibrio parahaemolyticus (V. parahaemolyticus) is a major food-borne pathogen which causes human gastroenteritis. Since the characteristics of V. parahaemolyticus remain unknown, 220 isolates selected from clinical and environmental samples in Dapeng of Shenzhen were tested for the presence of two hemolysin-expressing genes tdh and trh. Among 27 clinical isolates, 26 carrired the tdh gene, and the other one carried both tdh and trh genes, however neither genes were detected in environmental isolates. Meanwhile, antimicrobial susceptibility profiles revealed the isolates with high frequency of resistance to ampicillin (77.73%) and colistin (71.82%) and medium to streptomycin (57.27%). Genetically, by whole genome sequencing (WGS), comparative genomics studies was performed on isolates from various districts and GenBank. Data analysis showed that antimicrobial resistance genes (ARGs) blaCARB, tet(34) and tet(35) were harbored in all genomes and other ARGs was absent in the genomes of 27 clinical isolates. Besides, little regional difference was observed. As for virulence factors, MAM7, T3SS1, T3SS1 secret effector, T3SS2, T3SS2 secret effector, and VpadF were carried by most isolates. Two isolates from other districts were tdh gene positive which clustered with clinical isolates from Dapeng in the same clade, indicating close genetic distance. This study revealed the widely distribution of V. parahaemolyticus in Shenzhen and the diverse ARGs and virulence genes it carried. Furthermore, pathways that pathogen disseminated through were discussed.

Virulence Profiles of Salmonella enterica Isolated from Three Food Matrices Collected from Retail Markets.

Salmonella enterica is one of the most common foodborne pathogens associated with the consumption of contaminated porcine, dairy, and avian products. Nontyphoidal Salmonella is a major cause of bacterial diarrhea, responsible for ∼150 million cases and 60,000 deaths annually. The main goal of this study was to determine the prevalence of Salmonella spp. and to establish the virulence profile (VP) from genes (avrA, invE, ssaD, sseF, ssaQ, ttrC) and plasmid genes (pefA, spvB, spvC) in isolates obtained from cheese, chicken, and pork sold in food markets in Barrancabermeja, Colombia. A survey was conducted on 100 samples each matrix. The detection of Salmonella spp. followed the ISO 6579:2017 standards modified, and isolates were confirmed using the invA gene. In addition, single polymerase chain reaction assays were developed to detect the nine virulence genes. Salmonella spp. was found in 62%, 32%, and 14% of pork, chicken, and cheese samples, respectively. A total of 277 isolates were biochemically, serologically, and molecularly compatible with Salmonella spp. The most representative serogroups were C and B. Forty-seven combinations of virulence gene were detected; 53.5% of the pork isolates, 46.2% of the cheese isolates, and 39% of the chicken isolates were distributed among VP1, VP2, and VP3 suggesting a higher pathogenic potential. In addition, seven isolates harbored plasmid-encoded virulence genes (spvB and spvC), which are associated with increased invasiveness. The results revealed a higher prevalence of Salmonella spp. in pork and chicken compared with other studies conducted in Colombia. The serogroups identified include serovars that more frequently affect humans Salmonella Enteriditis, Salmonella Newport, and Salmonella Typhimurium. The isolations have the majority of the virulence genes studied. These findings highlight the need to improve control measures and educate food handlers to minimize the presence of Salmonella spp. and its potential transmission.

Purification and Characterization of Plantaricin PQ12: A Novel Bacteriocin from Lactiplantibacillus plantarum Isolated from Fermented Cucumbers, with Inhibitory Activity against Foodborne Pathogens

Purification and Characterization of Plantaricin PQ12: A Novel Bacteriocin from <i>Lactiplantibacillus plantarum</i> Isolated from Fermented Cucumbers, with Inhibitory Activity against Foodborne Pathogens

Quality Characteristics and Essential Oil Properties of Thymus capitatus, Mentha piperita, and Sideritis cypria Dried under Different Conditions

The drying of medicinal and aromatic plants (MAPs) is one of the main preservation methods for these products that can prolong their shelf life, if performed properly. The current study aimed to examine the effects of different drying conditions (sun, shade, and oven drying at 42 °C) on the quality characteristics of Thymus capitatus, Mentha piperita, and Sideritis cypria; their essential oil (EO) yield; and their biological properties (antioxidant and antibacterial activities). According to the results of the current study, oven drying resulted in faster moisture loss for all investigated species and slightly darker products. For T. capitatus, sun drying resulted in higher EO carvacrol content, whereas EOs obtained from shade and oven drying (at 42 °C) presented high total phenolic content and great antimicrobial activity. For M. piperita, shade drying resulted in a higher EO yield and higher iso-menthone content, whilst the EO obtained from oven-dried mint plants presented great antibacterial activity against the investigated foodborne pathogens. S. cypria plants dried in an air-ventilated oven produced an EO rich in β-caryophyllene and α-pinene, which also presented great antioxidant and antibacterial activity. The findings of the current study indicate that traditional drying methods, such as sun and shade, can result in good-quality dried MAPs that can yield EOs with significant biological activities, along with minimum energy consumption and lower carbon dioxide production (lower environmental carbon footprint), as opposed to oven drying. However, the drying-process duration could be a limitation at the industrial scale.

Rapid Dual-Gene Detection of Burkholderia gladioli and Subspecies Cocovenenans in Fresh Noodles and Tremella Using CRISPR-Cas12a/Cas13a.

Burkholderia gladioli pv. cocovenenans is a notable foodborne pathogen that poses a significant risk to food safety. Contaminated food requires distinct classification and treatment procedures for non-pathogenic B. gladioli and its pathogenic subspecies cocovenenans. Hence, establishing a rapid and sensitive detection method to distinguish them is necessary. In this study, we aimed to establish a method combining the CRISPR-Cas12a/Cas13a dual system with recombinase aided amplification for rapid, specific and sensitive detection of non-pathogenic B. gladioli and pathogenic subspecies cocovenenans in food. First, a RAA-CRISPR-Cas12a/Cas13a method was developed and its feasibility was assessed. Next, specificity was analyzed using 23 strains of B. gladioli and 5 non-target strains. Following this, sensitivity was evaluated by preparing gradient dilutions of B. gladioli pv. cocovenenans bacterial solutions. At last, real food test samples, including fresh noodles and tremella artificially contaminated with B. gladioli pv. cocovenenans, were utilized for method validation and sensitivity comparison. The established RAA-CRISPR-Cas12a/Cas13a method exhibited high specificity and achieved 100% accuracy in detecting species B. gladioli and its subspecies cocovenenans. This rapid method could be finished within 45 min with a detection limit of 10° CFU/mL for bacterial concentration. Additionally, it achieved detection limits of 102 CFU/g for fresh noodles and 103 CFU/g for tremella. The rapid RAA-CRISPR-Cas12a/Cas13a method demonstrated high specificity and sensitivity in detecting and disgusting species B. gladioli and subspecies cocovenenans in both food test samples and post-cultivation colonies. The RAA-CRISPR-Cas12a/Cas13a method presented in this study offers a novel molecular approach for the rapid, accurate and sensitive detection of B. gladioli and its subspecies cocovenenans in foods.

Locking-Fluorescence Signals Regulated CRISPR/Cas12a Biosensor Based on Metal-Organic Framework for Sensitive Detection of Salmonella typhimurium.

The efficient, sensitive, and rapid detection of Salmonella typhimurium (S. typhimurium) in food and food products is important to ensure food safety and health. This study developed a fluorescence biosensing assay that integrated recombinase-aided amplification (RAA) and CRISPR/Cas12a with a zeolitic imidazolate framework-8@fluorescein sodium (ZIF-8@FLS) nanocomposite for the sensitive detection of S. typhimurium. In this approach, using RAA as a preamplification module, CRISPR/Cas12a-AChE as a target recognition and dual-enzyme cascade amplification module, and the prepared ZIF-8@FLS with high porosity and rapid pH responsiveness as a fluorescence signal explosive amplification module, the RAA-CRISPR/Cas12a-ZIF-8@FLS biosensor was constructed. Under optimal conditions, it exhibited an excellent linear relationship for S. Typhimurium, with a sensitive detection limit as low as 1.3 × 102 CFU/mL and could complete sample detection within 2 h relying on the RAA and ZIF-8@FLS explosive fluorescence rapid response, demonstrating its significant advantages in specificity, sensitivity, and reliability in food-borne pathogens detection.