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Dipicolinic acid (DPA) is a major constituent of spores and reportedly provides protection against inactivation by various thermal processes; however, the relationship between DPA and resistance towards pressure-assisted thermal processing is not well understood. Thermal and pressure-assisted thermal inactivation studies of Clostridium botulinum nonproteolytic strains QC-B and 610-F, proteolytic strain Giorgio-A, and thermal surrogate Clostridium sporogenes PA3679 spores suspended in ACES buffer (0.05 M, pH 7.0) were performed to determine if a relationship exists between DPA release and log reduction of spores. Thermal inactivation at 80, 83, and 87 °C for nonproteolytic strains and 101, 105, and 108 °C for the proteolytic strain and thermal surrogate were conducted. Pressure-assisted thermal inactivation for nonproteolytic strains at 83 °C/600 MPa and for the proteolytic strain and thermal surrogate at 105 °C/600 MPa were performed. Surviving spores were enumerated by 5-tube MPN method for log reductions and analyzed for released DPA by liquid chromatography-tandem mass spectrometry. The correlation between MPN log reductions, released DPA, and D-values were calculated. A positive correlation between released DPA and log reduction of spores was observed for QC-B and 610-F at 80 and 83 °C (r = 0.6073 - 0.7755; P < 0.01). At 87 °C, a positive correlation was detected for 610-F (r = 0.4242, P < 0.05) and no correlation was observed for QC-B (r = 0.1641; P > 0.05). A strong, positive correlation (r = 0.8359 - 0.9284; P < 0.05) between released DPA and log reduction of spores was observed for Giorgio-A at 101, 105, and 108 °C, and a strong, positive correlation (r = 0.8402; P < 0.05) was observed for PA3679 at 101 °C. A positive correlation (r = 0.5646 - 0.6724; P < 0.01) was observed for QC-B, 610-F, and Giorgio-A after pressure-assisted thermal treatment. No correlation (r = 02494; P > 0.05) was found for PA3679 after pressure-assisted thermal treatment. These results suggest a correlation exists between DPA release and heat resistance; however, the level of correlation varied between strains and temperatures. The findings from this research may aid in the development of spore inactivation strategies targeting the thermal resistance profiles of various strains of C. botulinum spores., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier Inc.)

Bacillus licheniformis is one of the major spore-forming bacteria with great genotypic diversity in raw milk, dairy ingredients, and final dairy products; it is found throughout the dairy-processing continuum. Although being widely used as a probiotic strain, this species also serves as a potential risk in the dairy industry based on its roles in foodborne illness and dairy spoilage. Biofilm formation of B. licheniformis, combined with the heat resistance of its spores, make it impossible to prevent the presence of B. licheniformis in final dairy products by using traditional cleaning and disinfection procedures. Despite the extensive efforts to identify B. licheniformis in various dairy samples, no reviews have been written on both hazards and benefits of this sporeformer. This review discusses the prevalence of B. licheniformis from raw milk to commercial dairy products, biofilm formation and spoilage potential of B. licheniformis, and possible prevention methods. In addition, the potential benefits of B. licheniformis in the dairy industry are also summarized., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

SUMMARYIn the 2018-revised Clostridium perfringens typing classification system, isolates carrying the enterotoxin ( cpe ) and alpha toxin genes but no other typing toxin genes are now designated as type F. Type F isolates cause food poisoning and nonfoodborne human gastrointestinal (GI) diseases, which most commonly involve type F isolates carrying, respectivefooly, a chromosomal or plasmid-borne cpe gene. Compared to spores of other C. perfringens isolates, spores of type F chromosomal cpe isolates often exhibit greater resistance to food environment stresses, likely facilitating their survival in improperly prepared or stored foods. Multiple factors contribute to this spore resistance phenotype, including the production of a variant small acid-soluble protein-4. The pathogenicity of type F isolates involves sporulation-dependent C. perfringens enterotoxin (CPE) production. C. perfringens sporulation is initiated by orphan histidine kinases and sporulation-associated sigma factors that drive cpe transcription. CPE-induced cytotoxicity starts when CPE binds to claudin receptors to form a small complex (which also includes nonreceptor claudins). Approximately six small complexes oligomerize on the host cell plasma membrane surface to form a prepore. CPE molecules in that prepore apparently extend β-hairpin loops to form a β-barrel pore, allowing a Ca 2+ influx that activates calpain. With low-dose CPE treatment, caspase-3-dependent apoptosis develops, while high-CPE dose treatment induces necroptosis. Those effects cause histologic damage along with fluid and electrolyte losses from the colon and small intestine. Sialidases likely contribute to type F disease by enhancing CPE action and, for NanI-producing nonfoodborne human GI disease isolates, increasing intestinal growth and colonization., Competing Interests: The authors declare no conflict of interest.

The spore-forming, anaerobic bacterium, Clostridium botulinum, can cause intestinal toxemia (colonization) botulism in adults and infants by colonizing the gut and producing botulinum neurotoxin in situ. In 2006, peanut butter was identified as a lab-confirmed source of C. botulinum spores for an adult colonization botulism case in Canada. It is recommended for infants to be exposed to peanut butter at an early age to help prevent the development of a peanut allergy, yet the prevalence of C. botulinum in retail peanut butters is currently unknown. This report details a survey that was conducted in 2007 for the presence of viable C. botulinum spores in 92 peanut butters and 12 other nut butter spreads obtained from retail grocery stores in Ottawa, Canada. Samples were tested for viable C. botulinum spores by detecting botulinum neurotoxin in enrichment cultures by mouse bioassay. Three of the peanut butters from the entire survey of nut butter spreads (3/104, 3%) produced cultures containing botulinum neurotoxin. Whole genome sequencing performed on one isolate from this survey, as well as a clinical isolate and peanut butter isolates associated with the 2006 adult colonization case revealed that all C. botulinum isolates contained a full-length chromosomal bont/A1 gene within an ha-orf + cassette. This study identifies retail peanut butters as a potential source of viable C. botulinum spores at the time of sampling. Whether peanut butter represents a food category that may be contributing to the incidence of infant botulism has yet to be determined., (© 2024. Crown.)

Infant botulism is now the most common form of human botulism in Canada and the United States. Infant botulism is a severe neuroparalytic disease caused by ingestion of the spore-forming neurotoxic clostridia, including Clostridium botulinum that colonize the large intestine and subsequently produce botulinum neurotoxin in situ. It has been over a century since the first surveys documenting the ubiquitous prevalence of C. botulinum in soils around the world. Since then, honey has been identified as the only well-known risk factor for infant botulism despite a multitude of international environmental surveys isolating C. botulinum spores from ground soil, aquatic sediments, and commonly available infant foods. Associations of infant botulism cases with confirmed sources of C. botulinum exposure have primarily implicated outdoor soil and indoor dust, as well as commonly ingested foods including honey, dry cereals, and even powdered infant formula. Yet the origin of infection remains unknown for most infant botulism cases. This review summarizes the various surveys from around the world for C. botulinum in environmental soils and sediments, honey, and other infant foods, as well as laboratory-confirmed associations with documented infant botulism cases. Additional factors are also discussed, including the composition of infant gut microbiota and the practice of breastfeeding. We make several recommendations to better identify sources of exposure to C. botulinum spores that could lead to effective preventive measures and help reduce the incidence of this rare but life-threatening disease., (© 2024. Crown.)

Quantifying spore germination and outgrowth heterogeneity is challenging. Single cell level analysis should provide supplementary knowledge regarding the impact of unfavorable conditions on germination and outgrowth dynamics. This work aimed to quantify the impact of pH on spore germination and outgrowth, investigating the behavior of individual spore crops, produced under optimal and suboptimal conditions. Bacillus mycoides (formerly B. weihenstephanensis) KBAB4 spores, produced at pH 7.4 and at pH 5.5 were incubated at different pH values, from pH 5.2 to 7.4. The spores were monitored by microscopy live imaging, in controlled conditions, at 30 °C. The images were analyzed using SporeTracker, to determine the state of single cells. The impact of pH on germination and outgrowth times and rates was estimated and the correlation between these parameters was quantified. The correlation between germination and outgrowth times was significantly higher at low pH. These results suggest that an environmental pressure highlights the heterogeneity of spore germination and outgrowth within a spore population. Results were consistent with previous observations at population level, now confirmed and extended to single cell level. Therefore, single cell level analyses can be used to quantify the heterogeneity of spore populations, which is of interest in order to control the development of spore-forming bacteria, responsible for food safety issues., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:Clement Trunet reports financial support was provided by Erasmus Plus. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Bacillus species, which have two cell-type forms (vegetative cells and spores), demonstrate a variety of probiotic functions in animal feed additives and human nutrition. We previously found that the probiotic effect of Bacillus subtilis S-2 spores with high germination response to L-alanine was specifically enhanced by the L-alanine pretreatment. The germination response of Bacillus is highly associated with the germination receptors of spores. However, how L-alanine-induced germination of spores exerts anti-infectious effect in epithelial cells remains unclear. In this study, we constructed the mutant strain of B. subtilis S-2 with germination receptor gerAA knockout to further explore the role of spore germination in resisting pathogen infection to cells. The differential probiotic effects of B. subtilis S-2 and S-2 ΔgerAA spores pretreated with L-alanine were evaluated in intestinal porcine epithelial cells (IPEC-J2) or Caco2 cells infected with enterotoxigenic Escherichia coli (ETEC) or following IL-1β stimulation. The results showed that the germination response of the S-2 ΔgerAA spores to L-alanine was significantly reduced. Compared with the S-2 ΔgerAA spores, the L-alanine-induced germination of B. subtilis S-2 spores significantly increased the activity of anti-adhesion of ETEC to IPEC-J2 cells and reduced the expression of inflammatory factors and cell receptors. L-alanine induction also significantly promoted the expression of autophagy-related proteins in the B. subtilis S-2 spores. These findings demonstrate that the gerAA germination receptor is essential for the probiotic function of Bacillus spores and that L-alanine treatment promotes the anti-infectious properties of the germinated spores in porcine intestinal epithelial IPEC-J2 cells. The result suggests the importance of germination receptor gerAA in helping spore germination and enhancing anti-infectious activity. The findings in the study benefit to screening of potential Bacillus probiotics and increasing probiotic efficacy induced by L-alanine as an adjuvant., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Long-term and excessive use of tetracycline hydrochloride (TC) can lead to its accumulation in the environment, which can cause water contamination, bacterial resistance, and food safety problems. 2,6-Pyridine dicarboxylic acid (DPA) is a major biomarker of Bacillus anthracis spores, and its rapid and sensitive detection is of great significance for disease prevention and counter-terrorism. A bifunctional ratiometric fluorescent nanoprobe has been fabricated to detect DPA and TC. 3,5-dicarboxyphenylboronic acid (BOP) was intercalated into layered europium hydroxide (LEuH) by the ion-exchange method and exfoliated into nanosheets as a fluorescent nanoprobe (PNP). DPA and TC could significantly enhance the red fluorescence of Eu 3+ through the antenna effect under different excitation wavelengths, while the fluorescence of BOP can be used as a reference based on the constant emission intensity, realizing ratiometric detection. A low limit of detection (LOD) for the target (DPA: 9.7 nM, TC: 21.9 nM) can be achieved. In addition, visual detection of DPA and TC was realized using color recognition software based on the obvious color changes. This is the first ratiometric fluorescent nanoprobe based on layered rare-earth hydroxide (LRH) for the detection of DPA and TC simultaneously, which opens new ideas in the design of multifunctional probes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The oral administration of probiotics is nowadays recognized as a strategy to treat or prevent the consequences of unhealthy dietary habits. Here we analyze and compare the effects of the oral administration of vegetative cells or spores of Shouchella clausii SF174 in counteracting gut dysfunctions induced by 6 weeks of high fructose intake in a rat model. Gut microbiota composition, tight junction proteins, markers of inflammation and redox homeostasis were evaluated in ileum and colon in rats fed fructose rich diet and supplemented with cells or spores of Shouchella clausii SF174. Our results show that both spores and cells of SF174 were effective in preventing the fructose-induced metabolic damage to the gut, namely establishment of "leaky gut", inflammation and oxidative damage, thus preserving gut function. Our results also suggest that vegetative cells and germination-derived cells metabolize part of the ingested fructose at the ileum level., Competing Interests: Declarations of competing interests ER acts as a consultant for Gruppo Savio (Italy) that has the rights for the commercialization of strain SF174. Gruppo Savio (Italy) had no role in the design of the study, in the collection, analyses or interpretation of data or in the writing of the manuscript., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

The Wide Area Demonstration (WAD) was a field exercise conducted under the U.S. EPA's Analysis of Coastal Operational Resiliency program, in conjunction with the U.S. Department of Homeland Security and the U.S. Coast Guard. The purpose of the WAD was to operationalize at field scale aspects of remediation activities that would occur following an outdoor release of Bacillus anthracis spores, including sampling and analysis, decontamination, data management, and waste management. The WAD was conducted in May 2022 at Fort Walker (formerly known as Fort A.P. Hill) and utilized Bacillus atrophaeus as a benign simulant for B. anthracis . B. atrophaeus spores were inoculated onto the study area at the beginning of the study, and air samples were collected daily during each of the different phases of the WAD using Dry Filter Units (DFUs). Ten DFU air samplers were placed at the perimeter of the study area to collect bioaerosols onto two parallel 47-mm diameter polyester felt filters, which were then subsequently analyzed in a microbiological laboratory for the quantification of B. atrophaeus . The study demonstrated the use of DFUs as a rugged and robust bioaerosol collection device. The results indicated that the highest B. atrophaeus spore air concentrations (up to ~ 5 colony forming units/m 3 ) occurred at the beginning of the demonstration (e.g. during inoculation and characterization sampling phases) and generally downwind from the test site, suggesting transport of the spores was occurring from the study area. Very few B. atrophaeus spores were detected in the air after several weeks and following decontamination of exterior surfaces, thus providing an indication of the site decontamination procedures' effectiveness. No B. atrophaeus spores were detected in any of the blank or background samples. Implications : Following an incident involving a release of Bacillus anthracis spores or other biological threat agent into the outdoor environment, understanding the factors that may affect the bioagent's fate and transport can help predict viable contaminant spread via the ambient air. This paper provides scientific data for the first time on ambient air concentrations of bacterial spores over time and location during different phases of a field test in which Bacillus atrophaeus (surrogate for B. anthracis) spores were released outdoors as part of a full-scale study on sampling and decontamination in an urban environment. This study advances the knowledge related to the fate and transport of bacterial spores (such as those causing anthrax disease) as an aerosol in the outdoor environment over the course of three weeks in a mock urban environment and has exposure and health risk implications. The highest spore air concentrations occurred at the beginning of the study (e.g. during inoculation of surfaces and characterization sampling), and in the downwind direction, but diminished over time; few B. atrophaeus spores were detected in the air after several weeks and following decontamination. Therefore, in an actual incident, potential reaerosolization of the microorganism and subsequent transport in the air during surface sampling and remediation efforts should be considered for determining exclusion zone locations and estimating potential risk to neighboring communities. The data also provide evidence suggesting that the large-scale decontamination of outdoor surfaces may reduce air concentrations of the bioagent, which is important since exposure of B. anthracis via inhalation is a primary concern.

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 3×10 8 killed spores was safe and well tolerated.

Chronic intestinal inflammation is associated with strong alterations of the microbial composition of the gut. Probiotic treatments and microbiota-targeting approaches have been considered to reduce the inflammation, improve both gut barrier function as well as overall gastrointestinal health. Here, a murine model of experimental colitis was used to assess the beneficial health effects of Bacillus subtilis SF106 and Bacillus clausii (recently renamed Shouchella clausii) SF174, two spore-forming strains previously characterised in vitro as potential probiotics. Experimental colitis was induced in BALB/c mice by the oral administration of dextran sodium sulphate (DSS) and groups of animals treated with spores of either strain. Spores of both strains reduced the DSS-induced inflammation with spores of B. clausii SF174 more effective than B. subtilis SF106. Spores of both strains remodelled the mouse gut microbiota favouring the presence of beneficial microbes such as members of the Bacteroidetes and Akkermansia genera.

Bacillus licheniformis formulations are effective for environmental remediation, gut microbiota modulation, and soil improvement. An adequate spore quantity is crucial for the activity of B. licheniformis formulations. This study investigated the synergistic effects of carbon/nitrogen source consumption and concentration on B. licheniformis BF-002 cultivation, with the aim of developing an automatic co-feeding strategy to enhance spore production. Initial glucose (10 g/L) and amino nitrogen (1.5 g/L) concentrations promote cell growth, followed by reduced glucose (2.0 g/L) and amino nitrogen (0.5 g/L) concentrations for sustained spore generation. The spore quantity reached 2.59 × 10 10 CFU/mL. An automatic co-feeding strategy was developed and implemented in 5 and 50 L cultivations, resulting in spore quantities of 2.35 × 10 10 and 2.86 × 10 10 CFU/mL, respectively, improving by 6.81% and 30.00% compared to that with a fixed glucose concentration (10.0 g/L). The culture broth obtained at both the 5 and 50 L scales was spray-dried, resulting in bacterial powder with cell viability rates of 85.94% and 82.68%, respectively. Even after exposure to harsh conditions involving high temperature and humidity, cell viability remained at 72.80% and 69.89%, respectively. Employing the automatic co-feeding strategy increased the transcription levels of the spore formation-related genes spo0A, spoIIGA, bofA, and spoIV by 7.42%, 8.46%, 8.87%, and 9.79%, respectively. The proposed strategy effectively promoted Bacillus growth and spore formation, thereby enhancing the quality of B. licheniformis formulations., (© 2024 Wiley Periodicals LLC.)

The field of hybrid engineered living materials seeks to pair living organisms with synthetic materials to generate biocomposite materials with augmented function since living systems can provide highly-programmable and complex behavior. Engineered living materials have typically been fabricated using techniques in benign aqueous environments, limiting their application. In this work, biocomposite fabrication is demonstrated in which spores from polymer-degrading bacteria are incorporated into a thermoplastic polyurethane using high-temperature melt extrusion. Bacteria are engineered using adaptive laboratory evolution to improve their heat tolerance to ensure nearly complete cell survivability during manufacturing at 135 °C. Furthermore, the overall tensile properties of spore-filled thermoplastic polyurethanes are substantially improved, resulting in a significant improvement in toughness. The biocomposites facilitate disintegration in compost in the absence of a microbe-rich environment. Finally, embedded spores demonstrate a rationally programmed function, expressing green fluorescent protein. This research provides a scalable method to fabricate advanced biocomposite materials in industrially-compatible processes., (© 2024. The Author(s).)

The urgency for energy efficient, responsive architectures has propelled smart material development to the forefront of scientific and architectural research. This paper explores biological, physical, and morphological factors influencing the programming of a novel microbial-based smart hybrid material which is responsive to changes in environmental humidity. Hygromorphs respond passively, without energy input, by expanding in high humidity and contracting in low humidity. Bacillus subtilis develops environmentally robust, hygromorphic spores which may be harnessed within a bilayer to generate a deflection response with potential for programmability. The bacterial spore-based hygromorph biocomposites (HBCs) were developed and aggregated to enable them to open and close apertures and demonstrate programmable responses to changes in environmental humidity. This study spans many fields including microbiology, materials science, design, fabrication and architectural technology, working at multiple scales from single cells to 'bench-top' prototype.Exploration of biological factors at cellular and ultracellular levels enabled optimisation of growth and sporulation conditions to biologically preprogramme optimum spore hygromorphic response and yield. Material explorations revealed physical factors influencing biomechanics, preprogramming shape and response complexity through fabrication and inert substrate interactions, to produce a palette of HBCs. Morphological aggregation was designed to harness and scale-up the HBC palette into programmable humidity responsive aperture openings. This culminated in pilot performance testing of a humidity-responsive ventilation panel fabricated with aggregated Bacillus HBCs as a bench-top prototype and suggests potential for this novel biotechnology to be further developed., (Creative Commons Attribution license.)

Bacterial vegetative cells turn into metabolically dormant spores in certain environmental situations. Once suitable conditions trigger the germination of spores belonging to the pathogenic bacterial category, public safety and environmental hygiene will be threatened, and lives will even be endangered when encountering fatal ones. Instant identification of pathogenic bacterial spores remains a challenging task, since most current approaches belonging to complicated biological methods unsuitable for onsite sensing or emerging alternative chemical techniques are still inseparable from professional instruments. Here we developed a polychromatic fluorescent nanoprobe for ratiometric detection and visual inspection of the pathogenic bacterial spore biomarker, dipicolinic acid (DPA), realizing rapidly accurate screening of pathogenic bacterial spores such as Bacillus anthracis spores. The nanoprobe is made of aminoclay-coated silicon nanoparticles and functionalized with europium ions, exhibiting selective and sensitive response toward DPA and Bacillus subtilis spores (simulants for Bacillus anthracis spores) with excellent linearity. The proposed sensing strategy allowing spore determination of as few as 0.3 × 10 5 CFU/mL within 10 s was further applied to real environmental sample detection with good accuracy and reliability. Visual quantitative determination can be achieved by analyzing the RGB values of the corresponding test solution color via a color recognition APP on a smartphone. Different test samples can be photographed at the same time, hence the efficient accomplishment of examining bulk samples within minutes. Potentially employed in various on-site sensing occasions, this strategy may develop into a powerful means for distinguishing hazardous pathogens to facilitate timely and proper actions of dealing with multifarious security issues.

Two growth modes have been described for the filamentous Streptomyces bacteria. Their classic developmental life cycle culminates in the formation of dormant spores, where movement to new environments is mediated through spore dispersal. In contrast, exploratory growth proceeds as a rapidly expanding vegetative mycelium that leads to extensive surface colonization and is associated with the release of volatile compounds that promote alkalinization (and reduced iron bioavailability) of its surrounding environment. Here, we report that exploratory growth in Streptomyces venezuelae can proceed in tandem with classic sporulating development in response to specific nutritional cues. Sporulating exploration is not accompanied by a rise in environmental pH but has the same iron acquisition requirements as conventional exploration. We found that mutants that were defective in their ability to sporulate were unaffected in exploration, but mutants undergoing precocious sporulation were compromised in their exploratory growth and this appeared to be mediated through premature activation of the developmental regulator WhiI. Cell envelope integrity was also found to be critical for exploration, as mutations in the cell envelope stress-responsive extracytoplasmic function sigma factor SigE led to a failure to explore robustly under all exploration-promoting conditions. Finally, in expanding the known exploration-promoting conditions, we discovered that the model species Streptomyces lividans exhibited exploration capabilities, supporting the proposal that exploration is conserved across diverse streptomycetes., Importance: Streptomyces bacteria have evolved diverse developmental and metabolic strategies to thrive in dynamic environmental niches. Here, we report the amalgamation of previously disparate developmental pathways, showing that colony expansion via exploration can proceed in tandem with colony sporulation. This developmental integration extends beyond phenotype to include shared genetic elements, with sporulation-specific repressors being required for successful exploration. Comparing this new exploration mode with previously identified strategies has revealed key differences (e.g., no need for environmental alkalinization), and simultaneously allowed us to define unifying requirements for Streptomyces exploration. The "reproductive exploration" phenomenon reported here represents a unique bet-hedging strategy, with the Streptomyces colony engaging in an aggressive colonization strategy while transporting a protected genetic repository., Competing Interests: The authors declare no conflict of interest.

Bacillus cereus is a prevalent foodborne pathogen that induces food poisoning symptoms such as vomiting and diarrhea. Its capacity to form spores and biofilm enables it to withstand disinfectants and antimicrobials, leading to persistent contamination during food processing. Consequently, it is necessary to develop novel and efficient antimicrobial agents to control B. cereus, its spores, and biofilms. Peptidoglycan hydrolases have emerged as a promising and eco-friendly alternative owing to their specific lytic activity against pathogenic bacteria. Here, we identified and characterized a Lysozyme-like cell wall hydrolase Lys14579, from the genome of B. cereus ATCC 14579. Recombinant Lys14579 specifically lysed B. cereus without affecting other bacteria. Lys14579 exhibited strong lytic activity against B. cereus, effectively lysing B. cereus cell within 20 min at low concentration (10 μg/mL). It also inhibited the germination of B. cereus spores and prevented biofilm formation at 12.5 μg/mL. Moreover, Lys14579 displayed good antimicrobial stability with negligible hemolysis in mouse red blood cells and no cytotoxicity against RAW264.7 cells. Notably, Lys14579 effectively inhibited B. cereus in boiled rice and minced meat in a dose-dependent manner. Furthermore, bioinformatics analysis and point mutagenesis experiments revealed that Glu-47 was the catalytic site, and Asp-57, Gln-60, Ser-61 and Glu-63 were active-site residues related with the cell wall lytic activity. Taken together, Lys14579 could be a promising biocontrol agent against vegetative cells, spores, and biofilm of B. cereus in food industry., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that would appear to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Bacillus subtilis spores are important food spoilage agents and are occasionally involved in food poisoning. In foods that are not processed with intense heat, such bacterial spores are controlled by a combination of different hurdles, such as refrigeration, acidification, and low water activity (a w ), which inhibit or delay germination and/or growth. Sporulation temperature has long been regarded as a relevant factor for the assessment of germination in chemically defined media, but little is known about its impact on food preservation environments. In this study, we compared germination dynamics of B. subtilis spores produced at optimal temperature (37 °C) with others incubated at suboptimal (20 °C) and supraoptimal (43 °C) temperatures in a variety of nutrients (rich-growth medium, L-alanine, L-valine, and AGFK) under optimal conditions as well as under food-related stresses (low a w , pH, and temperature). Spores produced at 20 °C had a lower germination rate and efficiency than those incubated at 37 °C in all the nutrients, while those sporulated at 43 °C displayed a higher germination rate and/or efficiency in response to rich-growth medium and mostly to L-alanine and AGFK under optimal environmental conditions. However, differences in germination induced by changes in sporulation temperature decreased when spores were activated by heat, mainly due to the greater benefit of heat for spores produced at 20 °C and 37 °C than at 43 °C, especially in AGFK. Non-heat-activated spores produced at 43 °C still displayed superior germination fitness under certain stresses that had considerably impaired the germination of the other two populations, such as reduced temperature and a w . Moreover, they presented lower temperature and pH boundaries for the inhibition of germination in rich-growth medium, while requiring a higher NaCl concentration threshold compared to spores obtained at optimal and suboptimal temperature. Sporulation temperature is therefore a relevant source of variability in spore germination that should be taken into account for the accurate prediction of spore behaviour under variable food preservation conditions with the aim of improving food safety and stability., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Aims: Indole and mucin are compounds found in the host environment as they are produced by the host or by the host-associated microbiota. This study investigated whether indole and mucin impact Clostridium perfringens growth and sporulation, as well as enterotoxin production and biofilm formation., Methods and Results: There was no impact on growth of Cl. perfringens for up to 400 µM indole and 240 mg/l mucin, and neither indole nor mucin affected sporulation. Reverse-transcriptase qPCR showed that mucin strongly upregulated the expression of Cl. perfringens enterotoxin (up to 121-fold increase), whereas indole had a much more modest effect (2-fold). This was also reflected in increased Cl. perfringens enterotoxin levels in mucin-treated Cl. perfringens (as assessed by a reversed passive latex agglutination assay). Finally, mucin and indole significantly increased biofilm formation of Cl. perfringens, although the effect size was relatively small (less than 1.5 fold)., Conclusion: These results indicate that Cl. perfringens can sense its presence in a host environment by responding to mucin, and thereby markedly increased enterotoxin production., (© The Author(s) 2024. Published by Oxford University Press on behalf of Applied Microbiology International.)

Persistent diarrhea is a severe gastroenteric disease with relatively high risk of pediatric mortality in developing countries. We conducted a randomized, double-blind, controlled clinical trial to evaluate the efficacy of liquid-form Bacillus clausii spore probiotics (LiveSpo CLAUSY; 2 billion CFU/5 mL ampoule) at high dosages of 4-6 ampoules a day in supporting treatment of children with persistent diarrhea. Our findings showed that B. clausii spores significantly improved treatment outcomes, resulting in a 2-day shorter recovery period (p < 0.05) and a 1.5-1.6 folds greater efficacy in reducing diarrhea symptoms, such as high frequency of bowel movement of ≥ 3 stools a day, presence of fecal mucus, and diapered infant stool scale types 4-5B. LiveSpo CLAUSY supportive treatment achieved 3 days (p < 0.0001) faster recovery from diarrhea disease, with 1.6-fold improved treatment efficacy. At day 5 of treatment, a significant decrease in blood levels of pro-inflammatory cytokines TNF-α, IL-17, and IL-23 by 3.24% (p = 0.0409), 29.76% (p = 0.0001), and 10.87% (p = 0.0036), respectively, was observed in the Clausy group. Simultaneously, there was a significant 37.97% decrease (p = 0.0326) in the excreted IgA in stool at day 5 in the Clausy group. Overall, the clinical study demonstrates the efficacy of B. clausii spores (LiveSpo CLAUSY) as an effective symptomatic treatment and immunomodulatory agent for persistent diarrhea in children.Trial registration: NCT05812820., (© 2024. The Author(s).)

Clostridioides difficile is an anaerobic gastrointestinal pathogen that spreads through the environment as dormant spores. To survive, replicate, and sporulate in the host intestine, C. difficile must adapt to a variety of conditions in its environment, including changes in pH, the availability of metabolites, host immune factors, and a diverse array of other species. Prior studies showed that changes in intestinal conditions, such as pH, can affect C. difficile toxin production, spore formation, and cell survival. However, little is understood about the specific genes and pathways that facilitate environmental adaptation and lead to changes in C. difficile cell outcomes. In this study, we investigated two genes, CD2505 and CD2506, that are differentially regulated by pH to determine if they impact C. difficile growth and sporulation. Using deletion mutants, we examined the effects of both genes (herein smrR and smrT ) on sporulation frequency, toxin production, and antimicrobial resistance. We determined that SmrR is a repressor of smrRT that responds to pH and suppresses sporulation and toxin production through regulation of the SmrT transporter. Further, we showed that SmrT confers resistance to erythromycin and lincomycin, establishing a connection between the regulation of sporulation and antimicrobial resistance.IMPORTANCE Clostridioides difficile is a mammalian pathogen that colonizes the large intestine and produces toxins that lead to severe diarrheal disease. C. difficile is a major threat to public health due to its intrinsic resistance to antimicrobials and its ability to form dormant spores that are easily spread from host to host. In this study, we examined the contribution of two genes, smrR and smrT , on sporulation, toxin production, and antimicrobial resistance. Our results indicate that SmrR represses smrT expression, while production of SmrT increases spore and toxin production, as well as resistance to antibiotics., Competing Interests: The authors declare no conflict of interest.

Bacterial spores have outstanding properties from the materials science perspective, which allow them to survive extreme environmental conditions. Recent work by [S. G. Harrellson et al. , Nature 619 , 500-505 (2023)] studied the mechanical properties of Bacillus subtilis spores and the evolution of these properties with the change of humidity. The experimental measurements were interpreted assuming that the spores behave as water-filled porous solids, subjected to hydration forces. Here, we revisit their experimental data using literature data on vapor sorption on spores and ideas from polymer physics. We demonstrate that upon the change of humidity, the spores behave like rubber with respect to their swelling, elasticity, and relaxation times. This picture is consistent with the knowledge of the materials comprising the bacterial cell walls-cross-linked peptidoglycan. Our results provide an interpretation of the mechanics of bacterial spores and can help in developing synthetic materials mimicking the mechanical properties of the spores., Competing Interests: Competing interests statement:The authors declare no competing interest.

During thermal processing of braised sauce beef, the lipid content of circularly used sauce increased accordingly because of lipid migration from beef to sauce, which may impact the bacterial heat resistance in the products. This study aims to characterize the heat resistance of Clostridium sporogenes spores in braised sauce beef, and investigate the effects of oil on the spore surface characteristics and microstructure. The results indicated that the heat resistance of C. sporogenes spores in beef was significantly higher than that in sauce. Oil addition remarkably enhanced the spore heat resistance in sauce, with D 95°C value three times more than that without oil added, and even higher than that in beef. The results of spore surface characteristics indicated that oil addition led to an increase of hydrophobicity and a decrease of zeta potential, which ultimately increased spore heat resistance. Microstructure analysis indicated that exosporium maintenance and cortex expansion induced by oil addition might contribute to the increase of spore heat resistance. This study has sufficiently verified the importance of oil content on the heat resistance of C. sporogenes spores, which should be taken into consideration when developing thermal processes for controlling the spores in food matrices., Competing Interests: Declaration of competing interest There are no conflicts of interests to declare., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Alicyclobacillus acidoterrestris has received much attention due to its unique thermo-acidophilic property and implication in the spoilage of pasteurized juices. The objective of this study was to evaluate the sterilization characteristics and mechanisms of pulsed light (PL) against A. acidoterrestris vegetative cells and spores in apple juice. The results indicated that bacteria cells in apple juice (8-20°Brix) can be completely inactivated within the fluence range of 20.25-47.25 J/cm 2 , which mainly depended on the soluble solids content (SSC) of juice, and the spores in apple juice (12°Brix) can be completely inactivated by PL with the fluence of 54.00 J/cm 2 . The PL treatment can significantly increase the leakage of reactive oxygen species (ROS) and proteins from cells and spores. Fluorescence studies of bacterial adenosine triphosphate (ATP) indicated that the loss of ATP was evident. Scanning electron microscopy and confocal laser scanning microscope presented that PL-treated cells or spores had serious morphological damage, which reduced the integrity of cell membrane and led to intracellular electrolyte leakage. In addition, there were no significant negative effects on total sugars, total acids, total phenols, pH value, SSC and soluble sugars, and organic acid content decreased slightly during the PL treatment. The contents of esters and acids in aroma components had a certain loss, while that of alcohols, aldehydes and ketones were increased. These results demonstrated that PL treatment can effectively inactivate the bacteria cells and spores in apple juice with little effect on its quality. This study provides an efficient method for the inactivation of A. acidoterrestris in fruit juice., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier B.V.)

The threat of bioterrorism has spurred research on the decontamination and containment of different agents. Anthrax [causative agent Bacillus anthracis (Ba)] is a disease that can lead to severe infections within human and animals, particularly when inhaled. This research investigated the use of spore-contaminated simulated runoff events into stormwater control measures (SCMs), which are designed to retain and improve the quality of runoff and may have the potential to filter and contain the spores. In this study, the effectiveness of a bioretention cell (BRC) and high flow media filter (HFMF) in Huron, Ohio, were evaluated for removal of Bacillus globigii (Bg) spores (a harmless cognate of Ba). Three 4-8 mm simulated runoff events were created for each SCM using a fire hydrant and Bg spores were injected into the runoff upstream of the SCM inlets. The BRC significantly (p < 0.001) outperformed the HFMF in reducing Bg concentrations and loads, with an average load reduction of 1.9 log (∼99% reduction) compared to 0.4 (∼60% reduction), respectively. A probable critical design factor leading to these differences was the infiltration rate of the media and subsequent retention time within the filters, which was supported by similar disparities in suspended solids reductions. Differences in spore removal may also have been due to particle size distribution of the HFMF, which was more gravelly than the bioretention cell. At 3 and 6 months after the-simulated runoff tests, soil samples taken from both SCMs, yielding detectable Bg spores within the top 15 cm of media, with increased spore concentrations where ponding occurred for longer durations during the tests. This suggests that forebays and areas near inlets may be hotspots for spore cleanup in a real-world bioterrorism incident., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Bacillus cereus is a foodborne pathogen widely distributed in the large-scale catering industry and produces spores. The study explored the antibacterial activity, potential mechanism of eugenol against B. cereus , and spores with germination rate. The minimum inhibitory concentration (MIC; 0.6 mg/mL) of eugenol to six B. cereus strains was compared with the control; B. cereus treated with eugenol had a longer lag phase. Eugenol at a concentration of more than 1/2MIC decreased viable B. cereus (∼5.7 log colony-forming unit [CFU]/mL) counts below detectable limits within 2 h, and eugenol of 3MIC reduced B. cereus (∼5.9 log CFU/mL) in skim milk below detectable limits within 30 min. The pH values of skim milk were unaffected by the addition of eugenol. The Δ E values below 2 show that the color variations of skim milk were not visible to the human eye. For sensory evaluation, eugenol did not significantly affect the color or structural integrity of the skim milk. It had a negative impact on the flavor and general sensory acceptance of the treated milk. Eugenol hyperpolarized B. cereus cell membrane, decreased intracellular ATP concentration, and increased intracellular reactive oxygen species contents and extracellular malondialdehyde contents, resulting in the cell membrane of B. cereus being damaged and permeabilized, and cell morphology being changed. In addition, according to the viable count, confocal laser scanning microscopy, and spore morphology changes, eugenol reduced the germination rate of B. cereus spores. These findings suggest that eugenol can be used as a new natural antibacterial agent to control B. cereus and spores in the food production chain.

Retail stores maintain fresh rice noodles (FRNs) at room temperature because refrigeration negatively impacts FRNs' texture. The room temperature and high water activity of FRNs help spore-forming Bacillus cereus to grow and produce toxins. In this study, the effect of steam cooking on survival and different storage temperatures on the growth and enterotoxins production of B. cereus in FRNs were investigated. White rice flour was used to make FRNs. Three treatments of FRNs were used in this study; uninoculated, inoculated (with 4.0 log CFU/ml of B. cereus spores), and autoclaved as a negative control. A slurry of rice flour, cornstarch, and water was steam cooked for 4 min at 90°C and incubated for 168 h at 4°C, and for 72 h at 22 and 32°C. Incubated FRNs were tested for pH, B. cereus growth, and enterotoxins production. Steam cooking reduced the total number of B. cereus spores by 0.7 ± 0.3 log CFU/g. Surviving B. cereus spores in inoculated and uninoculated FRNs germinated over 72 h of storage. No B. cereus was detected in negative controls. An interaction was observed across storage temperatures and time (p < 0.05). The B. cereus population in uninoculated FRNs increased by more than 7.0 log CFU/g at 22 and 32°C over 72 h, while inoculated FRNs showed a 5.0 log bacterial increase at these storage temperatures. No growth was observed at 4°C in both inoculated and uninoculated FRNs. The pH of inoculated FRNs was reduced from 6.9 ± 0.1 to 5.7 ± 0.0 at 32°C and to 6.2 ± 0.1 at 22°C, and the pH of uninoculated FRNs was reduced from 7.0 ± 0.1 to 5.8 ± 0.2 at 32°C and to 6.5 ± 0.0 at 22°C, indicative of FRNs spoilage. B. cereus in inoculated FRNs produced enterotoxins after 12 h of storage at 32°C, and over 24 h of storage at 22°C, while no toxin was detected at 4°C. Our findings show that storing FRNs at room temperature for 24 h leads to enterotoxin production, emphasizing the importance of proper FRN storage and their potential risk to consumers. Nevertheless, further research should investigate the effect of other foodborne pathogens on these products., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Identifying and distinguishing dormant and active bacterial spores are vital for biosecurity, food safety, and space exploration. Yet, there is a lack of simple, quick, and nondestructive methods to achieve this. The common Schaeffer-Fulton method is both sample-destructive and requires significant operator involvement. In this study, we employed lanthanide-beta-diketonate complexes to directly observe both dormant and germinated single spores. Staining is instantaneous and requires minimal sample processing. The complex stains areas outside the core of dormant spores, leaving the core hollow and nonfluorescent. However, upon germination, the complex enters the core, making it brightly fluorescent. This difference was noted in five bacterial species including Bacillus , Clostridium , and Clostridioides . Various lanthanides and beta-diketonates can be mixed to form a range of spore-visualizing complexes. Due to their low toxicity, these complexes allow for live imaging of single germinating spores. We demonstrate low-cost imaging using a USB microscope as well as imaging of spores in milk matrices. This method provides a valuable tool for studying bacterial spores.

In laboratory testing, a mobile enclosed disinfection cabinet using ultraviolet-C light and aerosolized hydrogen peroxide was effective for disinfection of hard and soft surfaces. The addition of aerosolized hydrogen peroxide to ultraviolet-C light resulted in improved disinfection of soft surfaces and Clostridioides difficile spores.

The tailing phenomenon, where the survival curve of bacteria shows a slow tailing period after a rapid decline, is a ubiquitous inactivation kinetics process in the advanced plasma sterilization field. While classical models suggest that bacterial resistance dispersion causes the tailing phenomenon, experiments suggest that the non-uniform spatial distribution of spores (clustered structure) is the cause. However, no existing inactivation kinetics model can accurately describe spatial heterogeneity. In this paper, we propose a lattice model based on percolation theory to explain the inactivation kinetics by considering the non-uniform spatial distribution of spores and plasma. Our model divides spores into non-clustered and clustered types and distinguishes between short-tailing and long-tailing compositions and their formation mechanisms. By systematically studying the effects of different spore and plasma parameters on the tailing phenomenon, we provide a reasonable explanation for the kinetic law of the plasma sterilization survival curve and the mechanism of the tailing phenomenon in various cases. As an example, our model accurately explains the 80-second kinetics of atmospheric pressure plasma inactivation of spores, a process that previous models struggled to understand due to its multi-stage and long-tail phenomena. Our model predicts that increasing the spatial distribution probability of plasma can shorten the complete killing time under the same total energy, and we validate this prediction through experiments. Our model successfully explains the seemingly irregular plasma sterilization survival curve and deepens our understanding of the tailing phenomenon in plasma sterilization. This study offers valuable insights for the sterilization of food surfaces using plasma technology, and could serve as a guide for practical applications., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

Fungi outbreaks in water will include a series of processes, including spore aggregation, germination, biofilm, and finally present in a mixed state in the aquatic environment. More attention is paid to the control of dispersed fungal spores, however, there was little knowledge of the control of germinated spores. This study investigated the inactivation kinetics and mechanism of ultraviolet (UV) treatment for fungal spores with different germination percentages compared with dormant spores. The results indicated that the inactivation rate constants (k) of spores with 5%-45% germination were 0.0278-0.0299 cm 2 /mJ for Aspergillus niger and 0.0588-0.0647 cm 2 /mJ for Penicillium polonicum, which were lower than those of dormant spores. It suggested that germinated spores were more tolerant to UV irradiation than dormant spores, and it may be due to the defensive barrier (upregulated pigments) and some reductive substance (upregulated enoyl reductase) by absorbing UV or reacting with reactive oxygen species according to transcriptome analysis. Compared to dormant spores, the k- UV of germinated spores decreased by 18.17%-26.56% for Aspergillus niger, which was less than k-chlorine (62.33%-69.74%). A slighter decrease in k- UV showed UV irradiation can efficiently control fungi contamination, especially when dormant spores and germinated spores coexisted in actual water systems. This study indicates that more attention should be paid to germinated spores., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Inosine could potentially become a novel antibacterial agent against Alicyclobacillus acidoterrestris as low doses of inosine can prevent its contamination. However, until now the antibacterial mechanism of inosine targeting A. acidoterrestris is still unknown. In this study, to unravel the mechanism of inosine against A. acidoterrestris puzzle, the effects of inosine on bacterial surface hydrophobicity, intracellular protein content, cell membrane damage extent, and permeability of the A. acidoterrestris were investigated. The results showed that inosine can effectively inhibit the growth and reproduction of A. acidoterrestris by destroying the integrity of cell membrane and increasing its permeability, causing the leakage of intracellular nutrients. Furthermore, the interaction networks of inosine target proteins were analyzed. The interaction networks further revealed that damage to bacterial cell membranes might be relevant to inosine's effect on bacterial DNA replication and cell energy metabolism through regulating nucleotide synthesis and metabolism and the activity of translation initiation factors. Finally, the antibacterial mechanism of inosine against A. acidoterrestris was proposed., (© 2024 Institute of Food Technologists.)

Clostridium butyricum is a probiotic that forms anaerobic spores and plays a crucial role in regulating gut microbiota. However, the total viable cell count and spore yield of C. butyricum in industrial production are comparatively low. To this end, we investigated the metabolic characteristics of the strain and proposed three distinct pH regulation strategies for enhancing spore production. In addition, precise measurement of fermentation parameters such as substrate concentration, total viable cell count, and spore concentration is crucial for successful industrial probiotics production. Nevertheless, online measurement of these intricate parameters in the fermentation of C. butyricum poses a considerable challenge owing to the complex, nonlinear, multivariate, and strongly coupled characteristics of the production process. Therefore, we analyzed the capacitance and conductivity acquired from a viable cell sensor as the core parameters for the fermentation process. Subsequently, a robust soft sensor was developed using a seven-input back-propagation neural network model with input variables of fermentation time, capacitance, conductivity, pH, initial total sugar concentration, ammonium ion concentration, and calcium ion concentration. The model enables the online monitoring of total viable biomass count, substrate concentrations, and spore yield, and can be extended to similar fermentation processes with pH changes as a characteristic feature., (© 2023 Wiley Periodicals LLC.)

Heat resistance of spores of Clostridium perfringens 8238 (Hobbs Serotype 2), Bacillus cereus NCTC 11143 (4810/72), and Bacillus subtilis PS533, an isogenic derivative of strain PS832 (a 168 strain) was determined in ground beef at 95 °C. Spore purification was by centrifugation and washing with sterile distilled water (dH2O), followed by sonication and then Histodenz centrifugation for B. subtilis and C. perfringens, and centrifugation and washing with sterile dH2O followed by Histodenz centrifugation for B. cereus. Bags containing inoculated beef samples were submerged in a temperature-controlled water bath and held at 95 °C for predetermined lengths of time. Surviving spore populations were enumerated by plating on mannitol egg yolk polymyxin agar (MYP) agar plates for B. cereus and B. subtilis, and on tryptose-sulfite-cycloserine agar (TSC) agar plates for C. perfringens. Survivor curves were fitted to linear, linear with tail, and Weibull models using the USDA Integrated Pathogen Modeling Program (IPMP) 2013 software. The Weibull model provided a relatively better fit to the data since the root mean square error (RMSE), mean square error (MSE), sum of squared errors (SSE), and Akaike information criterion (AIC) values were lower than the values obtained using the linear or the linear with tail models. Additionally, the Weibull model accurately predicted the observed D-values at 95 °C for the three spore-formers since the accuracy factor (A f ) values ranged from 1.03 to 1.08 and the bias factor (B f ) values were either 1.00 or 1.01. Times at 95 °C to achieve a 3-log reduction decreased from 206 min for C. perfringens spores purified with water washes alone to 191 min with water washes followed by sonication and Histodenz centrifugation, from 7.9 min for B. cereus spores purified with water washes alone to 1.4 min with water washes followed by Histodenz centrifugation, and from 20.6 min for B. subtilis spores purified with water washes alone to 6.7 min for water washes followed by sonication and Histodenz centrifugation. Thermal-death-time values reported in this study will assist food processors to design thermal processes to guard against bacterial spores in cooked foods. In addition, clearly spore purity is an additional factor in spore wet heat resistance, although the cause of this effect is not clear., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

Recovery of rare earth elements (REEs) from wastewater with Bacillus subtilis (B. subtilis) during culture is promising due to its environmental benefits. However, the effects of REEs in the culture media on B. subtilis are poorly understood. This study aims to investigate the effects of the terbium (Tb(III)), a typical rare earth element, on the cell growth, sporulation, and spore properties of B. subtilis. Tb(III) can suppress bacterial growth while enhancing spore tolerance to wet heat. Spore germination and content of dipicolinic acid (DPA) were promoted at low concentrations of Tb(III) while inhibited at a high level, but an inverse effect on initial sporulation appeared. Scanning electron microscope and energy dispersive spectrometer detection indicated that Tb(III) complexed cells or spores and certain media components simultaneously. The germination results of the spores after elution revealed that Tb(III) attached to the spore surface was a key effector of spore germination. In conclusion, Tb(III) directly or indirectly regulated both the nutrient status of the media and certain metabolic events, which in turn affected most of the properties of B. subtilis. Compared to the coat-deficient strain, the wild-type strain grew faster and was more tolerant to Tb(III), DPA, and wet heat, which in turn implied that it was more suitable for the recovery of REEs during cultivation. These findings provide fundamental insights for the recovery of rare earths during the culture process using microorganisms., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Despite the variety of pathogens that are transmitted via the airborne route, few data are available on factors that influence the tenacity of airborne pathogens. In order to better understand and thus control airborne infections, knowledge of these factors is important. In this study, three agents, S. aureus, G. stearothermophilus spores and the MS2 bacteriophage, were aerosolized at relative humidities (RH) varying between 30% and 70%. Air samples were then analyzed to determine the concentration of the agents. S. aureus was found to have significantly lower survival rate in the aerosol at RH above 60%. It showed the lowest recovery rates of the three agents, ranging from 0.13% at approximately 70% RH to 4.39% at 30% RH. G. stearothermophilus spores showed the highest tenacity with recovery rates ranging from 41.85% to 61.73% with little effect of RH. For the MS2 bacteriophage, a significantly lower tenacity in the aerosol was observed with a recovery rate of 4.24% for intermediate RH of approximately 50%. The results of this study confirm the significant influence of the RH on the tenacity of airborne microorganisms depending on the specific agent. These data show that the behavior of microorganism in bioaerosols is varies under different environmental conditions., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Siller et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)