Spore-forming bacteria in gelatin: Characterization, identification by 16S rRNA and MALDI-TOF mass spectrometry (MS), and presence of heat resistance and virulence genes.

Gelatin, a versatile protein derived from collagen, is widely used in the food, pharmaceutical and medical sectors. However, bacterial contamination by spore-forming bacteria during gelatin processing represents a significant concern for product safety and quality. In this study, an investigation was carried out to explore the heat and chemical resistance, as well as the identification and characterization of spore-forming bacteria isolated from gelatin processing. The methodologies involved chemical resistance tests with drastic pH in microplates and thermal resistance tests in capillary tubes of various isolates obtained at different processing stages. In addition, phenotypic and genotypic analyses were carried out to characterize the most resistant isolates of spore-forming bacteria. The findings of this study revealed the presence of several species, including Bacillus cereus, Bacillus licheniformis, Bacillus sonorensis, Bacillus subtilis, Geobacillus stearothermophilus, and Clostridium sporogenes, with some isolates exhibiting remarkable chemical and heat resistances. In addition, a significant proportion of the most resistant isolates showed gelatinase activity (n = 19/21; 90.5 %) and the presence of heat resistance (n = 5/21; 23.8 %), and virulence genes (n = 11/21; 52.4 %). The results of this study suggest that interventions should be done in quality control practices and that process parameter adjustments and effective contamination reduction strategies should be implemented through gelatin processing.

Unveiling the matrix effect on Bacillus licheniformis and Bacillus subtilis spores heat inactivation between plant-based milk alternatives, bovine milk and culture medium.

This study examined the inactivation of spores of Bacillus licheniformis and Bacillus subtilis in four pea-based milk alternatives, semi-skimmed bovine milk and Brain Heart Infusion (BHI) broth to assess the matrix impact on the thermal inactivation of bacterial spores. Heat inactivation was performed with the method of capillary tubes in temperature range 97-110 °C. A four-parameter non-linear model, including initial level, shoulder duration, inactivation rate and tailing, was fitted to the data obtained. D-values were estimated and secondary ZT-value models were developed for both species. A secondary model for the shoulder length of B. licheniformis in a plant-based milk alternative formulation was built too. Models were validated at a higher temperature, 113.5 °C. D-values in the different matrices ranged between 2.3 and 8.2 min at 97 °C and 0.1-0.3 min at 110 °C for B. licheniformis. D-values for B. subtilis ranged between 3.9 and 6.3 min at 97 °C and 0.2-0.3 min at 110 °C. ZT-values in the different matrices ranged between 7.3 and 8.9 °C and 8.9-10.0 °C for B. licheniformis and B. subtilis, respectively. Significant differences in inactivation parameters were found within the pea-based formulations as well as when compared to bovine milk. Heat resistance was higher in pea-based matrices. Shoulders observed were temperature- and matrix-dependent, while no such trend was found for the tailings. These results provide insights, useful on designing safe thermal processing, limiting spoilage in plant-based milk alternatives and thus, reducing global food waste.

Description, life cycle, and development of the myxozoan Myxobolus rasmusseni n. sp. in fathead minnows, Pimephales promelas: A possible emerging pathogen in southern Alberta, Canada.

Morphological, gene sequence, host tissue tropism, and life cycle characteristics were utilized to describe the myxozoan, Myxobolus rasmusseni n. sp. from fathead minnow, Pimephales promelas, collected from reservoirs in southern Alberta. Results from serial histological sections of whole heads showed that myxospores were contained within irregular-shaped and sized coelozoic capsules (=plasmodia). Clusters of membrane-bound, myxospore-filled plasmodia filled the head cavities of juvenile fathead minnows, leading to the development of large, white, disfiguring lesions in mid to late summer. Bilateral exopthalmia (pop-eye disease) was a common outcome of M. rasmusseni n. sp. development. BLASTn search of a 1974 bp sequence of the 18S rDNA gene isolated from myxospores indicated that M. rasmusseni n. sp. was distinct from other coelozoic and histozoic Myxobolus spp. cataloged in GenBank. 18S rDNA gene sequences from triactinomyxon spores released from the oligochaete Tubifex were 100% identical to sequences from myxospores collected from syntopic fathead minnows. Results from a longitudinal survey of the 2020 cohort of fathead minnows showed that young-of-the-year are exposed at 1-5 mo and that 60-90% of these had developed myxospore-filled lesions approximately one year later. Data regarding potential sources and timing of M. rasmusseni n. sp. emergence in fathead minnow populations are needed.

Polyethylene imine-modified photonic crystal microfluidic chip for highly sensitive detection of microbial spores.

To address the lengthy cycles, complex operations, high costs, and insufficient sensitivity of biomarker detection in traditional biological control agents, photonic crystal treated with PEI was developed for highly sensitive detection of Sclerotinia sclerotiorum microbial spores. By incorporating gelatin molecules, photonic crystal is endowed with excellent photothermal stability and high stability in aqueous solutions. The photonic crystal surface is conferred a positive charge by PEI, which can be used to enhance the adsorption of spores. Efficient enrichment of Sclerotinia sclerotiorum and Purpureocillium lilacinum spores is achieved, with coefficients of determination (RYe et al. (2014)2) 0.963 and 0.971, respectively. The detection range is from 102 to 106 spores/ml, and the photonic crystal exhibited good reusability. The prepared photonic crystal enables rapid, non-destructive, and accurate quantitative detection of microbial spores.

Intranasal administration of DSM 32444 Bacillus subtilis spores: safety and tolerability.

Introduction. Administered nasally, spores of the Gram-positive bacterium Bacillus subtilis have been shown to be able to induce innate immunity sufficient to confer protection to influenza and respiratory syncytial virus.Hypothesis. Although members of the aerobiome, intranasal delivery of high numbers of live spores carries potential safety issues.Aim. To address the potential safety risk of using live spores, we assessed the safety of spores that had been completely inactivated using heat sterilization.Methodology. Using autoclaved, and therefore killed, spores of a generally recognized as safe-notified B. subtilis strain (DSM 32444), safety was assessed in vitro (biotype, genome and cell based cytoxicity) and in vivo, using intranasal administration in rodent models and lastly in human volunteers.Results. Using a 15-day, repeat-dose, regimen in a rodent model, no indication of toxicity was observed. In a registered human study (NCT05984004), a formulated preparation of inactivated DSM 32444 spores referred to as SPEROVID was developed, and tolerance in human volunteers was assessed following 7 days of nasal dosing (2-4 times/day).Conclusion. Our study demonstrated that in humans an intranasal dose of up to 3×108 killed spores was safe and well tolerated.

Adsorptive removal of aflatoxin B1 via spore protein from Aspergillus luchuensis YZ-1.

Aflatoxin B1 (AFB1) is the most toxic mycotoxin commonly found in the environment. Finding efficient and environmentally friendly ways to remove AFB1 is critical. In this study, Aspergillus luchuensis YZ-1 demonstrated a potent ability to adsorb AFB1 for the first time, and the binding of AFB1 to YZ-1 is highly stable. Spores exhibited higher adsorption efficiency than mycelia, adsorbing approximately 95 % of AFB1 within 15 min. The spores were comprehensively characterized using scanning electron microscopy, transmission electron microscopy, energy-dispersive X-ray spectroscopy, and atomic force microscopy. Various adsorption kinetic models (pseudo-first and pseudo-second order), adsorption isotherm models (Freundlich and Langmuir), Fourier transform infrared, and X-ray photoelectron spectroscopy were used to investigate the adsorption properties and mechanisms. The adsorption capacity of spores decreased with heating, urea, and SDS treatments, indicating that spore proteins may be the primary substance for AFB1 adsorption. Subsequent experiments showed that proteins with molecular weights greater than 50 kDa played a key role in the adsorption. Additionally, the spores possess excellent storage properties and are valuable for adsorbing AFB1 from vegetable oils. Therefore, the YZ-1 spores hold promise for development into a novel biosorbent for AFB1 removal.

Inhibition and Mechanism of Citral on Bacillus cereus Vegetative Cells, Spores, and Biofilms.

Bacillus cereus causes food poisoning by producing toxins that cause diarrhea and vomiting and, in severe cases, endocarditis, meningitis, and other diseases. It also tends to form biofilms and spores that lead to contamination of the food production environment. Citral is a potent natural antibacterial agent, but its antibacterial activity against B. cereus has not been extensively studied. In this study, we first determined the minimum inhibitory concentrations and minimum bactericidal concentrations, growth curves, killing effect in different media, membrane potential, intracellular adenosine triphosphate (ATP), reactive oxygen species levels, and morphology of vegetative cells, followed by germination rate, morphology, germination state of spores, and finally biofilm clearance effect. The results showed that the minimum inhibitory concentrations and minimum bactericidal concentrations of citral against bacteria ranged from 100 to 800 µg/mL. The lag phase of bacteria was effectively prolonged by citral, and the growth rate of bacteria was slowed down. Bacteria in Luria-Bertani broth were reduced to below the detection limit by citral at 800 µg/mL within 0.5 h. Bacteria in rice were reduced to 3 log CFU/g by citral at 4000 µg/mL within 0.5 h. After treatment with citral, intracellular ATP concentration was reduced, membrane potential was altered, intracellular reactive oxygen species concentration was increased, and normal cell morphology was altered. After treatment with citral at 400 µg/mL, spore germination rate was reduced to 16.71%, spore morphology was affected, and spore germination state was altered. It also had a good effect on biofilm removal. The present study showed that citral had good bacteriostatic activity against B. cereus vegetative cells and its spores and also had a good clearance effect on its biofilm. Citral has the potential to be used as a bacteriostatic substance for the control of B. cereus in food industry production.

Two-year investigation of spore-formers through the production chain at two cheese plants in Norway.

Spore-forming bacteria are the most complex group of microbes to eliminate from the dairy production line due to their ability to withstand heat treatment usually used in dairy processing. These ubiquitous microorganisms have ample opportunity for multiple points of entry into the milk chain, creating issues for food quality and safety. Certain spore-formers, namely bacilli and clostridia, are more problematic to the dairy industry due to their possible pathogenicity, growth, and production of metabolites and spoilage enzymes. This research investigated the spore-forming population from raw milk reception at two Norwegian dairy plants through the cheesemaking stages until ripening. Samples were collected over two years and examined by amplicon sequencing in a culture independent manner and after an anaerobic spore-former enrichment step. In addition, a total of 608 isolates from the enriched samples were identified at the genus or species level using MALDI-TOF analysis. Most spore-forming isolates belong to the genera Bacillus or Clostridium, with the latter dominating the enriched MPN tubes of raw milk and bactofugate. Results showed a great variation among the clostridia and bacilli detected in the enriched MPN tubes. However, B. licheniformis and C. tyrobutyricum were identified in all sample types from both plants throughout the 2-year study. In conclusion, our results shed light on the fate of different spore-formers at different processing stages in the cheese production chain, which could facilitate targeted actions to reduce quality problems.

Impact of pH and High-Pressure Pasteurization on the Germination and Development of Clostridium perfringens Spores under Hyperbaric Storage versus Refrigeration.

This study aimed to evaluate hyperbaric storage at room temperature (75-200 MPa, 30 days, 18-23 °C, HS/RT) on Clostridium perfringens spores in brain-heart infusion broth (BHI-broth) at pH 4.50, 6.00, and 7.50 and coconut water (pH 5.40). Both matrices were also pasteurized by high pressure processing (600 MPa, 3 min, 17 °C, HPP) to simulate commercial pasteurization followed by HS, in comparison with refrigeration (5 °C, RF). The results showed that, at AP/RT, spores' development occurred, except at pH 4.50 in BHI-broth, while for RF, no changes occurred along storage. Under HS, at pH 4.50, neither spore development nor inactivation occurred, while at pH 6.00/7.50, inactivation occurred (≈2.0 and 1.0 logs at 200 MPa, respectively). Coconut water at AP/RT faced an increase of 1.6 logs of C. perfringens spores after 15 days, while for RF, no spore development occurred, while the inactivation of spores under HS happened (≈3 logs at 200 MPa). HPP prior to HS seems to promote a subsequent inactivation of C. perfringens spores in BHI-broth at pH 4.50, which is less evident for other pHs. For HPP coconut water, the inactivation levels under HS were lower (≈2.0 logs at 200 MPa). The Weibull model well described the inactivation pattern observed. These results suggest that HS/RT can be simultaneously used as a tool to avoid C. perfringens spores' development, as well as for its inactivation, without the application of high temperatures that are required to inactivate these spores.

Calamitean Cones and Their In Situ Spores from the Pennsylvanian Limnic Basins of the Czech Republic.

This paper describes the in situ spores of the Calamospora type, macerated from sixty-one specimens of calamitean cones belonging to sixteen species of genera, such as the Palaeostachya, Macrostachya, Calamostachys, and Huttonia from the Pennsylvanian Czech Republic period, specifically from the Moscovian/Kasimovian ages (i.e., Duckmantian-Stephanian). The in situ spores were compared to twenty dispersed species of Calamospora. The majority of spores were microspores; however, some cones yielded both micro- and megaspores. Morphological variations of the in situ spores, including the diameter, labrum, contact area, ontogenetic stages, and secondary folds of the exine, are described, including their importance for the classification of calamospores. The relationships of Elaterites, Pteroretis, Vestispora, and some monopseudosaccate spores are discussed. All Paleozoic Calamospora-producing parent plants are summarized.

[Genome Stability of Bacillus velezensis after Two-Year Exposure in Open Space].

Spore-forming bacteria have a unique resistance to negative environmental conditions, including aggressive space factors, and are an excellent model for studying adaptation mechanisms and survival strategies at the molecular level. The study analyzed the genome of Bacillus velezensis, which remained viable after a 2-year exposure in outer space on the outer surface of the ISS as part of the Test space experiment. A comparative analysis of the draft genomes of the exhibit strain and the ground control did not reveal significant changes; the average nucleotide identity was 99.98%, which indicates the ability of microorganisms to maintain genome stability in space conditions, due to both increased stress resistance of bacterial spores and efficient operation of the system of repair of accumulated changes. The study of a single nucleotide polymorphism in the genome of B. velezensis revealed nine point substitutions, three of which are in intergenic regions, six in protein-coding genes, three of them are missense mutations, two nucleotide deletions leading to a shift in the reading frame, and one synonymous substitution. The profiles of the housekeeping genes were determined during MLST typing and it was found that the allelic profiles obtained for B. velezensis T15.2 and 924 strains do not correspond to any of the previously described sequence types. The presented results indicate the ability of B. velezensis bacteria to maintain the viability of spores and the integrity of the genome for a long time under extreme conditions of outer space, which is important for the problem of planetary protection, as well as the potential possibility of performing biotechnological processes based on B. velezensis during space exploration.

Rapid, Point-of-Care Microwave Lysis and Electrochemical Detection of Clostridioides difficile Directly from Stool Samples.

The rapid detection of the spore form of Clostridioides difficile has remained a challenge for clinicians. To address this, we have developed a novel, precise, microwave-enhanced approach for near-spontaneous release of DNA from C. difficile spores via a bespoke microwave lysis platform. C. difficile spores were microwave-irradiated for 5 s in a pulsed microwave electric field at 2.45 GHz to lyse the spore and bacteria in each sample, which was then added to a screen-printed electrode and electrochemical DNA biosensor assay system to identify presence of the pathogen's two toxin genes. The microwave lysis method released both single-stranded and double-stranded genome DNA from the bacterium at quantifiable concentrations between 0.02 µg/mL to 250 µg/mL allowing for subsequent downstream detection in the biosensor. The electrochemical bench-top system comprises of oligonucleotide probes specific to conserved regions within tcdA and tcdB toxin genes of C. difficile and was able to detect 800 spores of C. difficile within 300 µL of unprocessed human stool samples in under 10 min. These results demonstrate the feasibility of using a solid-state power generated, pulsed microwave electric field to lyse and release DNA from human stool infected with C. difficile spores. This rapid microwave lysis method enhanced the rapidity of subsequent electrochemical detection in the development of a rapid point-of-care biosensor platform for C. difficile.

Current and Future Effects of Climate Change on Airborne Allergens.

PURPOSE OF REVIEW: Delineation of the impact of elevated carbon dioxide and concomitant global warming on airborne allergens is performed. RECENT FINDINGS: European tree pollen trends in general showed earlier start and end dates and increased total pollen release, with some differences both in locale and among species. Earlier flowering was also seen with grasses and weeds. In the case of some boreal trees, flowering was delayed due to a pre-seasonal requirement for necessary accumulated chilling temperature to achieve bud-set. Anthropogenic climate change induced rise in temperature and CO2 levels has resulted in demonstrable increases in aeroallergens. This has been most dramatic in tree pollen annual load, but also seen with grasses and weeds. Collected data is greatest for the Northern Hemisphere, especially the European continent, with supporting data from North America and Australia.

Differential temperature sensitivity of haploid and diploid spores of two red intertidal algae from the Magellan Strait.

Spores have crucial importance in the establishment and development of seaweed populations. When the spore release matches with the low tidal period, they experience an extreme variation in the environmental conditions including the temperature. In this study, we assess the photosynthetic responses and growth of haploid (tetraspores) and diploid (carpospores) spores of two Gigartinales species (Mazzaella laminarioides and Iridaea cordata) from sub-Antarctic populations when exposed to an increasing temperature. In the laboratory, freshly released spores were exposed to a temperature gradient (7 [control], 10, 15, and 20 °C) recreating the temperature increase experienced by these spores during typical spring tides. Germination and further growth of spores previously exposed to temperature treatments were assessed. Carpospores and tetraspores exhibited variation in their photosynthetic response (measured as effective quantum yield; ΦPSII) to temperature increase. In Mazzaella laminarioides, only carpospores exhibited a reduction in ΦPSII (by 7-24% at 15-20 °C), while both types of spores of Iridaea cordata were sensitive to temperature increase (12-24% of ΦPSII reduction at 10-20 °C). Spores previously exposed to temperature treatments and maintained at 7 °C and low PAR germinated and developed in germlings. In general, germlings originated from carpospores pre-treated at high temperatures showed higher growth rates. The different responses to temperature increase exhibited by haploid and diploid propagules of both species highlight their ecophysiological capacity to face high-temperature variation ensuring successful recruitment survival.

Resilience of Canola to Plasmodiophora brassicae (Clubroot) Pathotype 3H under Different Resistance Genes and Initial Inoculum Levels.

In this study, we explored the resilience of a clubroot resistance (CR) stacking model against a field population of Plasmodiophora brassicae pathotype 3H. This contrasts with our earlier work, where stacking CRaM and Crr1rutb proved only moderately resistant to pathotype X. Canola varieties carrying Rcr1/Crr1rutb and Rcr1 + Crr1rutb were repeatedly exposed to 3H at low (1 × 104/g soil) and high (1 × 107/g soil) initial resting spore concentrations over five planting cycles under controlled environments to mimic intensive canola production. Initially, all resistant varieties showed strong resistance. However, there was a gradual decline in resistance over time for varieties carrying only a single CR gene, particularly with Crr1rutb alone and at the high inoculum level, where the disease severity index (DSI) increased from 9% to 39% over five planting cycles. This suggests the presence of virulent pathotypes at initially low levels in the 3H inoculum. In contrast, the variety with stacked CR genes remained resilient, with DSI staying below 3% throughout, even at the high inoculum level. Furthermore, the use of resistant varieties, carrying either a single or stacked CR genes, reduced the total resting spore numbers in soil over time, while the inoculum level either increased or remained high in soils where susceptible Westar was continuously grown. Our study demonstrates greater resistance resilience for stacking Rcr1 and Crr1rutb against the field population of 3H. Additionally, the results suggest that resistance may persist even longer in fields with lower levels of inoculum, highlighting the value of extended crop rotation (reducing inoculum) alongside strategic CR-gene deployment to maximize resistance resilience.

A critical assessment of microbial-based antimicrobial sanitizing of inanimate surfaces in healthcare settings.

The global rise in antimicrobial resistance (AMR) poses a significant public health threat, especially in healthcare settings, where controlling the spread of antimicrobial genes is crucial. While person-to-person transmission remains the primary route for healthcare-associated infections (HAIs), hospital surfaces serve as key reservoirs for antimicrobial-resistant microorganisms. Regular cleaning and disinfection of these surfaces are essential. Microbial-based products for sanitizing hospital surfaces have emerged as promising tools to combat HAIs and AMR. However, a review of 32 publications found inconsistencies and potential risks. A total of 15 publications included hospital-based trials, while the rest were either in vitro or in situ assays, reviews, book chapters, or commentaries. In most of the hospital-based studies, specific strains of applied microorganisms were not identified, and the term "probiotic" was inaccurately used. These products mainly featured spores from Bacillus and Priestia genera, which was mainly hypothesized to work through competitive exclusion. Most hospital-based studies have shown that the application of microbial-based products resulted in a significant reduction in pathogens on surfaces, thereby contributing to a decrease in the incidence of healthcare-associated infections (HAIs). Further research is however needed to understand the effectiveness, mechanisms of action, and safety of microbial-based sanitizing agents. Strain-level identification is crucial for safety assessments, yet many reviewed products lacked this information. Consequently, there is a need for rigorous safety evaluations within existing regulatory frameworks to ensure the efficacy and safety of microbial-based cleaning products in healthcare settings.

Automatic real-time monitoring of fungal spores: the case of Alternaria spp.

We present the first implementation of the monitoring of airborne fungal spores in real-time using digital holography. To obtain observations of Alternaria spp. spores representative of their airborne stage, we collected events measured in the air during crop harvesting in a contaminated potato field, using a Swisens Poleno device. The classification algorithm used by MeteoSwiss for operational pollen monitoring was extended by training the system using this additional dataset. The quality of the retrieved concentrations is evaluated by comparison with parallel measurements made with a manual Hirst-type trap. Correlations between the two measurements are high, especially over the main dispersion period of Alternaria spp., demonstrating the potential for automatic real-time monitoring of fungal spores.

Illuminating the Cryptococcus neoformans species complex: unveiling intracellular structures with fluorescent-protein-based markers.

Cryptococcus neoformans is a fungal pathogen of the top critical priority recognized by the World Health Organization. This clinically important fungus also serves as a eukaryotic model organism. A variety of resources have been generated to facilitate investigation of the C. neoformans species complex, including congenic pairs, well-annotated genomes, genetic editing tools, and gene deletion sets. Here, we generated a set of strains with all major organelles fluorescently marked. We tested short organelle-specific targeting sequences and successfully labeled the following organelles by fusing the targeting sequences with a fluorescence protein: the plasma membrane, the nucleus, the peroxisome, and the mitochondrion. We used native cryptococcal Golgi and late endosomal proteins fused with a fluorescent protein to label these two organelles. These fluorescence markers were verified via colocalization using organelle-specific dyes. All the constructs for the fluorescent protein tags were integrated in an intergenic safe haven region. These organelle-marked strains were examined for growth and various phenotypes. We demonstrated that these tagged strains could be employed to track cryptococcal interaction with the host in phagocytosis assays. These strains also allowed us to discover remarkable differences in the dynamics of proteins targeted to different organelles during sexual reproduction. Additionally, we revealed that "dormant" spores transcribed and synthesized their own proteins and trafficked the proteins to the appropriate subcellular compartments, demonstrating that spores are metabolically active. We anticipate that these newly generated fluorescent markers will greatly facilitate further investigation of cryptococcal biology and pathogenesis.

Fungi: friends or foes-an outreach science initiative for the collection of airborne fungal spores by high school students.

Fungi mostly reproduce through spores that are adapted for airborne dispersal; hence, fungal spores (and fungi) are found virtually everywhere. Fungi can be "friends or foes." Our friends include fungi used in the food and biotech industries, fungi that contribute to the cycling of carbon and nutrients, and those involved in the decontamination of polluted soils and/or water, to mention just a few examples. Many species, however, are foes-they are detrimental to plants, animals, and/or humans. Annually, >1.5 million people die due to invasive fungal infections. With the aim of enhancing microbiology literacy and the understanding of microbial concepts, we set up a project for the collection of airborne spores (the principal agent through which human airways are exposed to fungi). Students from five high schools in the Oeiras municipality partnered with us as citizen scientists; they carried out sampling by collecting fungal spores on adhesive stickers. The fungal spores collected by the students were subsequently processed in the schools and our research laboratory. Results obtained by the students themselves revealed a large variety of fungal species capable of growing in a rich medium at 30°C. In the research laboratory, using selective isolation conditions, 40 thermotolerant fungi were isolated, 32 of which were taxonomically identified as aspergilla, mostly from within the Aspergillus fumigatus taxa, yet exhibiting high genetic heterogeneity. The protocols and results were presented to the students, who were made aware of the local dispersal of airborne fungal spores, including some from potentially pathogenic fungi. Through carrying out scientific activities, the students developed both the interest and the self-confidence needed to implement future environmental investigations.

Characteristic substance analysis and rapid detection of bacteria spores in cooked meat products by surface enhanced Raman scattering based on Ag@AuNP array substrate.

BACKGROUND: Bacterial spores are the main potential hazard in medium- and high-temperature sterilized meat products, and their germination and subsequent reproduction and metabolism can lead to food spoilage. Moreover, the spores of some species pose a health and safety threat to consumers. The rapid detection, prevention, and control of bacterial spores has always been a scientific problem and a major challenge for the medium and high-temperature meat industry. Early and sensitive identification of spores in meat products is a decisive factor in contributing to consumer health and safety. RESULTS: In this study, we developed a novel and stable Ag@AuNP array substrate by using a two-step synthesis approach and a liquid-interface self-assembly method that can directly detect bacterial spores in actual meat product samples without the need for additional in vitro bacterial culture. The results indicate that the Ag@AuNP array substrate exhibits high reproducibility and Raman enhancement effects (1.35 × 105). The differentiation in the Surface enhanced Raman scattering (SERS) spectra of five bacterial spores primarily arises from proteins in the spore coat and inner membrane, peptidoglycan of cortex, and Ca2⁺-DPA within the spore core. The correct recognition rate of linear discriminant analysis for spores in the meat product matrix can reach 100 %. The average recovery accuracy of the SERS quantitative model was at around 101.77 %, and the limit of detection can reach below 10 CFU/mL. SIGNIFICANCE: It provides a promising technological strategy for the characteristic substance analysis and timely monitoring of spores in meat products.