Bacterial Endospores Research Articles

Correlations between available primary amines, endospore coat thickness, and alkaline glutaraldehyde sensitivity for spores of select Bacillus species.

Alkaline glutaraldehyde (GTA) is a high‐level chemical disinfectant/sterilant and has a broad microbial kill spectrum. The precise antimicrobial mechanism of GTA remains debated. GTA kill times are extremely variable across different organisms, illustrating the need for a better understanding of GTA kill mechanisms related to different organisms. A commonly proposed GTA kill mechanism suggests that it works by cross‐linking accessible primary amines on important surface proteins. If true, the antimicrobial activity of GTA may directly correlate to the number of these available functional groups. Bacillus species form highly resistant bacterial endospores that are commonly used as one of the most stringent test organisms for disinfection and sterilization. In this study, we compared the log reduction times of alkaline GTA on spores from 4 Bacillus species to fluorescent profiles generated using Alexa Fluor™ amine‐reactive dyes. GTA kill times were also compared to mean spore coat thicknesses as measured with scanning electron microscopy (SEM). Fluorescence values generated from bound amine‐reactive dye showed a strong, positive correlation to GTA susceptibility, as measured by GTA 6‐log10 reduction times. Spore coat thickness also showed a strong, positive correlation to reduction time values. Results support the hypothesis that GTA kill times are directly related to the number of available primary amines on bacterial endospores. Results also indicated that the killing efficacy of GTA may be influenced by its ability to penetrate the spore coat to reach additional targets, suggesting that damaging important biomolecules beyond surface proteins may be involved in GTA killing mechanisms.

Endocavity Ultrasound Transducers: Why High-Level Disinfection IsNecessary.

As a medical imaging modality, ultrasound is used by a wide cross-section of practitioners including radiologists, obstetricians, gynecologists, gastroenterologists, urologists and cardiologists. The increasing popularity of ultrasound as a diagnostic tool is due not only to the ease of use and portability of systems, but also to the perceived safety aspect of the examination. This latter point needs to be examined. As with any reusable medical device, the ultrasound transducer, also known as a probe, could potentially be a vector for the transmission of pathogenic viruses and fungi between patients if not correctly disinfected after each use. This transmission risk is magnified for an endocavity transducer that has come in contact with the vagina, anal canal or oral cavity, as it could be contaminated with organisms transmitted by blood or mucosal, genital or rectal secretions. Based on the Spaulding system, transducers that come in contact with mucous membranes are classified as semi-critical devices that require high-level disinfection (HLD) after each patient procedure. This HLD process should eliminate all microorganisms except high numbers of bacterial endospores. Only a small number of countries worldwide have implemented transducer reprocessing guidelines that adhere to the Spaulding classification and recommend HLD for endocavity transducers. Overall, there is a lack of conformity among global health agencies regarding the use of HLD for endocavity transducers. This is primarily due to the perception that the infection transmission risk is negligible and that if an endocavity transducer has been covered with a single-use sheath for the procedure, then low-level disinfection provides sufficient protection against pathogen transmission. The objective of this study was to review the published risk of infection transmission from endocavity transducers. By highlighting the outbreaks and case reports that implicate pathogen transmission from transducers, we posit that HLD should be a global standard of practice for the reprocessing of endocavity transducers. It requires substantial time for national health administrations to develop and legislate new recommendations, and for practice changes to be accepted and implemented by healthcare providers. We recommend that Joint Commission International (JCI) and other equivalent organizations enforce the use of HLD of endocavity ultrasound transducers during their accreditation reviews.

Identification of bacterial endospores and targeted detection of foodborne viruses in industrially reared insects for food.

With edible insects being increasingly produced, food safety authorities have called for the determination of microbiological challenges posed to human health. Here, we find that the bacterial endospore fraction in industrially reared mealworm and cricket samples is largely comprised of Bacillus cereus group members that can pose insect or human health risks. Hepatitis A virus, hepatitis E virus and norovirus genogroup II were not detected in the sample collection, indicating a low food safety risk from these viral pathogens.

Microbial Barrier Properties of Cotton Fabric-Influence of Weave Architecture.

The subject of the paper focuses on the effect of weave architecture on microbial barrier properties of woven fabrics or more precisely on identifying crucial elements of weave architecture that dominantly influence bacteria penetration in dry condition. For that purpose, 12 samples of cotton fabrics were woven and examined. In their structure, all samples had the same yarns (36 tex) in warp and weft, same densities of warp (24 yarns/cm), two weft densities (24 and 20 yarns/cm) and six different basic weave structures. Microbial barrier permeability was determined according to a previously developed test method in cooperation with University Hospital Center Zagreb. Bacterial endospores of apathogenic species of the genus Bacillus: Geobacillus stearothermophilus and Bacillus atrophaeus were used. The effect of weave pattern on microbial barrier properties was significant. Weave patterns, decisively determined the number of influencing pores and its sizes in woven fabrics, as well as the yarn floating which jointly almost perfectly correlated with bacteria penetration through the woven fabric. Multiple linear regression of pore numbers and floating threads produced equations which correspond in 99% to the measuring results for densities 24/24 and 24/20, and more than 98% considering both densities of the set. Among compared weave patterns, satin weave had significantly lower permeability of microorganisms (six–seven times) than basket weave (the highest), for both densities.

Plant Root-Exudates Recruit Hyperparasitic Bacteria of Phytonematodes by Altered Cuticle Aging: Implications for Biological Control Strategies.

Phytonematodes are globally important functional components of the belowground ecology in both natural and agricultural soils; they are a diverse group of which some species are economically important pests, and environmentally benign control strategies are being sought to control them. Using eco-evolutionary theory, we test the hypothesis that root-exudates of host plants will increase the ability of a hyperparasitic bacteria, Pasteuria penetrans and other closely related bacteria, to infect their homologous pest nematodes, whereas non-host root exudates will not. Plant root-exudates from good hosts, poor hosts and non-hosts were characterized by gas chromatography-mass spectrometry (GC/MS) and we explore their interaction on the attachment of the hyperparasitic bacterial endospores to homologous and heterologous pest nematode cuticles. Although GC/MS did not identify any individual compounds as responsible for changes in cuticle susceptibility to endospore adhesion, standardized spore binding assays showed that Pasteuria endospore adhesion decreased with nematode age, and that infective juveniles pre-treated with homologous host root-exudates reduced the aging process and increased attachment of endospores to the nematode cuticle, whereas non-host root-exudates did not. We develop a working model in which plant root exudates manipulate the nematode cuticle aging process, and thereby, through increased bacterial endospore attachment, increase bacterial infection of pest nematodes. This we suggest would lead to a reduction of plant-parasitic nematode burden on the roots and increases plant fitness. Therefore, by the judicious manipulation of environmental factors produced by the plant root and by careful crop rotation this knowledge can help in the development of environmentally benign control strategies.

Microbial Profile of the Ventriculum of Honey Bee (Apis mellifera ligustica Spinola, 1806) Fed with Veterinary Drugs, Dietary Supplements and Non-Protein Amino Acids.

The effects of veterinary drugs, dietary supplements and non-protein amino acids on the European honey bee (Apis mellifera ligustica Spinola, 1806) ventriculum microbial profile were investigated. Total viable aerobic bacteria, Enterobacteriaceae, staphylococci, Escherichia coli, lactic acid bacteria, Pseudomonas spp., aerobic bacterial endospores and Enterococcus spp. were determined using a culture-based method. Two veterinary drugs (Varromed® and Api-Bioxal®), two commercial dietary supplements (ApiHerb® and ApiGo®) and two non-protein amino acids (GABA and beta-alanine) were administered for one week to honey bee foragers reared in laboratory cages. After one week, E. coli and Staphylococcus spp. were significantly affected by the veterinary drugs (p < 0.001). Furthermore, dietary supplements and non-protein amino acids induced significant changes in Staphylococcus spp., E. coli and Pseudomonas spp. (p < 0.001). In conclusion, the results of this investigation showed that the administration of the veterinary drugs, dietary supplements and non-protein amino acids tested, affected the ventriculum microbiological profile of Apis mellifera ligustica.

Basidiospores from Wood-Decay Fungi Transform Laccase Substrates in the Absence of Glucose and Nitrogen Supplements.

Preparations of bacterial endospores and fungal conidia are applied in biocontrols, biocatalyses, and lignocellulose fermentations. The biocatalytic abilities of basidiospores from mushrooms of the order Agaricales are unknown. To assess their potential in colonizing recalcitrant substrates solely with their inherent resources, spores of the white-rot fungi Stropharia rugoso-annulata (Stru) and Kuehneromyces mutabilis (Kmt, Strophariaceae) were analyzed for surface-bound and internal total carbohydrates, phenols, proteins, minerals, and oxidoreductases to estimate their chemistry and the preconditions to transform the laccase substrates guaiacol and 2,2′-azinobis-(3-ethylbenzthiazoline-6-sulfonate) (ABTS) independent of external glucose and nitrogen. Surfaces of Stru/Kmt spores released (mg kg−1) hexoses, 7300/9700; phenols, >62/220; proteins, 21/168; and laccases, 42/0–0.15 µmol ABTS•+ kg−1 min−1 that mimicked oxidative activities of the resting spores. Milled-spore extracts contained pentoses, 96,600/6750; hexoses, 160,000/15,130; phenols, 452/767; protein, 12,600/924; true laccase, 688/0.30; and enzyme-protein-activating transition metals such as Cu in concentrations typical of wheat grains. Independent of external N and C supply, spores (<1‰) germinated in bideionized water, supported by their surface resources. Kmt spores germinated, too, at comparable rates in N-free solutions of glucose and the not immediately metabolizable ABTS and guaiacol. The release of proteins and oxidoreductase(s) by Kmt spores starting upon germination was higher in guaiacol-incubated idiophase- than in glucose-incubated trophophase-spores and led to the 3–4-fold formation of guaiacol polymerizates and ABTS•+. Constitutive aromatic ring-cleaving dioxygenases in the dormant spore that could be involved in the intrinsic metabolization of guaiacol were not detected. It is concluded that intrinsic resources enable (germinating) spores to release the highly efficient laccases of basidiomycetes and to transform aromatic compounds in the absence of sugar amendments. Spores show therefore plant seed-like autonomy in nutrient modification and acquisition during the early stages of the colonization of inert substrates.

Can farm weeds improve the growth and microbiological quality of crickets (Gryllus bimaculatus)?

Farming of edible insects has been proposed as a means to reduce current practices of harvesting from the wild. While farming could relieve the pressure on wild populations, as well as on their natural habitats, and generate a continuous supply of edible insects to the consumers, the high cost of commercial chicken feeds is still a challenge to many farmers. The aim of this study was to investigate whether the partial replacement of commercial chicken feed with the farm weed, wandering Jew (Commelina sinensis), would have an impact on weight gain and microbial quality of farmed field crickets (Gryllus bimaculatus). Therefore, we have experimentally compared two types of feed: (1) starter commercial chicken feed only (SO); and (2) starter commercial chicken feed supplemented with fresh wandering Jew weeds (S+W). After a feeding period of thirty days, the final body weight differed significantly (P=0.026) between crickets fed with SO and with S+W, which averaged at 1.11 and 1.39 g, respectively. Generally, high counts of total aerobic,Enterobacteriaceae, lactic acid bacteria, bacterial endospores, yeasts and moulds were observed in both experimental groups. However, bacterial endospore counts of S+W fed crickets (2.7 log cfu/g) were significantly lower (P=0.021) than these of the SO fed ones (3.9 log cfu/g). Metagenetic analyses indicated that Proteobacteria, Firmicutes and Tenericutes were the most abundant phyla, while members of the family Coxiellacea, and the generaLactobacillus, andSpiroplasma were the most abundant operational taxonomic units (OTUs). Concerning the food safety, a few OTUs could be associated with potential food pathogens, such asClostridiaceae,Staphylococcus andEnterobacteriaceae. In summary, the inclusion of fresh wandering Jew weeds in commercial starter chicken feed produced crickets with increased body weight and improved microbial quality.

Mechanism of Bacillus subtilis spore inactivation induced by moderate electric fields

ABSTRACT Bacterial endospores are the key safety targets for inactivation within low-acid foods. Herein, we investigated the inactivation of Bacillus subtilis CGMCC 1.1087 spores (107 CFU/mL) in sterile distilled water using moderate electric fields (MEF, 300 V/cm) under various temperatures ( Industrial relevance Literature in the past few years has shown that moderate electric fields (MEF), typically associated with ohmic heating, have nonthermal effects on bacterial spores, leading to accelerated inactivation. The current work extends the range of temperatures to those well below thermally lethal conditions, and shows that some spore inactivation occurs under MEF, even when temperatures are sublethal. Little or no germination is observed, and spore inner membranes are increasingly compromised over time. The elucidation of such nonthermal effects would be significant to the food industry as it seeks increasingly nonthermal methods for inactivation of spores.

Inactivation of bacterial endospores on surfaces by plasma processed air.

In case of biological hazards and pandemics, personal protective equipment of rescue forces is currently manually decontaminated with harmful disinfectants, primarily peracetic acid. To overcome current drawbacks regarding supply, handling and disposal of chemicals, the use of plasma processed air (PPA) represents a promising alternative for surface decontamination on site. In this study, the sporicidal efficiency of a portable plasma system, designed for field applications, was evaluated. The developed plasma device is based on a dielectric barrier discharge (DBD) and operated with ambient air as process gas. PPA from the plasma nozzle was flushed into a treatment chamber (volume: 300l) and bacterial endospores (Bacillus subtilis and Bacillus atrophaeus) dried on different surfaces were treated under variable conditions. Reductions in spores by more than 4 log10 were found within 3min of PPA exposure. However, the presence of endospores in agglomerates or in an organic matrix as well as the complexity of the respective surface microstructure negatively affected the inactivation efficiency. When endospores were embedded in a dried protein matrix, mechanical wiping with swabs during exposure to PPA increased the inactivation effect significantly. Gaseous ozone alone did not provide a sporicidal effect. Significant spore inactivation was only obtained when water vapour was injected into the PPA stream. The results show that endospores dried on surfaces can be reduced by several orders of magnitude within few minutes in a treatment chamber which is flushed with PPA from of a DBD plasma nozzle. Plasma processed air generated on site by DBD plasma nozzles could be a suitable alternative for the disinfection of various surfaces in closed rooms.

Ultraviolet C light with wavelength of 222 nm inactivates a wide spectrum of microbial pathogens

UVC has been used to inactivate several pathogens. Unlike the conventional 254-nm UVC, 222-nm UVC is harmless to mammalian cells. To investigate the disinfection efficacy of 222-nm UVC against human pathogens which are commonly found in the environment and healthcare facilities. Staphylococcus aureus, Pseudomonas aeruginosa, Escherichia coli, Salmonella enterica subsp. serovar Typhimurium, Campylobacter jejuni, Bacillus cereus (vegetative cells and endospores), Clostridium sporogenes (vegetative cells and endospores), Clostoridioides difficile (endospores), Candida albicans (yeast), Aspergillus niger (hyphae and spores), Trichophyton rubrum (hyphae and spores), feline calicivirus and influenza A virus were irradiated with 222-nm UVC at various doses. The remaining live bacterial and fungal cells, and the viral infectivity were evaluated. The efficiency of 222-nm UVC germicidal effect was compared to that of the conventional 254-nm UVC. The 222-nm UVC showed potent germicidal effect to vegetative bacterial cells, yeast and viruses as efficient as the 245-nm UVC. The 222-nm UVC exhibited more potent germicidal effect to bacterial endospores, compared with the 254-nm UVC. The fungicidal effect of 222-nm UVC against the fungal spores and hyphae was weaker than that of 254-nm UVC. The 222-nm UVC is able to inactivate a wide spectrum of microbial pathogens. In comparison with the conventional 254-nm UVC, the germicidal effect of 222-nm UVC to the fungal hyphae and spores is low, but the 222-nm UVC exhibits strong germicidal effect to the bacterial endospores.

Influence of Washing and Sterilization on Properties of Polyurethane Coated Fabrics Used in Surgery and for Wrapping Sterile Items.

The objective of this work was to determine the influence of washing and sterilization under real hospital conditions on properties of microbial barrier offered by polyurethane coated fabrics used in surgery and for wrapping sterile items. Emphasis was put on the change of surface polyurethane coating by using FTIR analysis. The permeability and durability of the microbial barrier were determined after 0, 10, and 20 washing and sterilization procedures according to previously developed methods. Bacterial endospores of the apathogenic species of the genus Bacillus Geobacillus stearothermophilus and Bacillus atrophaeus were used. Mechanical damage to medical textiles in the washing and sterilization process was determined according to standard HRN EN ISO 13914-1:2008 and associated with changes in physical and mechanical properties. Chemical changes of PU coatings were determined using FTIR analysis. The results showed an exceptionally efficient microbial barrier and its durability in all samples after 0, 10 and 20 washing and sterilization procedures and for a period of one, two and three months.

Antibacterial Surfaces with Activity against Antimicrobial Resistant Bacterial Pathogens and Endospores.

Hospital-acquired bacterial infections are a significant burden on healthcare systems worldwide causing an increased duration of hospital stays and prolonged patient suffering. We show that polyurethane containing crystal violet (CV) and 3-4 nm zinc oxide nanoparticles (ZnO NPs) possesses excellent bactericidal activity against hospital-acquired pathogens including multidrug resistant Escherichia coli (E.coli), Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus (MRSA), and even highly resistant endospores of Clostridioides (Clostridium) difficile. Importantly, we used clinical isolates of bacterial strains, a protocol to mimic the environmental conditions of a real exposure in the healthcare setting, and low light intensity equivalent to that encountered in UK hospitals (∼500 lux). Our data shows that ZnO NPs enhance the photobactericidal activity of CV under low intensity light even with short exposure times, and we show that this involves both Type I and Type II photochemical pathways. Interestingly, polyurethane containing ZnO NPs alone showed significant bactericidal activity in the dark against one strain of E.coli, indicating that the NPs possess both light-activated synergistic activity with CV and inherent bactericidal activity that is independent of light. These new antibacterial polymers are potentially useful in healthcare facilties to reduce the transmission of pathogens between people and the environment.

Light exposure decreases infectivity of the Daphnia parasite Pasteuria ramosa

Abstract Climate change is altering light regimes in lakes, which should impact disease outbreaks, since sunlight can harm aquatic pathogens. However, some bacterial endospores are resistant to damage from light, even surviving exposure to UV-C. We examined the sensitivity of Pasteuria ramosa endospores, an aquatic parasite infecting Daphnia zooplankton, to biologically relevant wavelengths of light. Laboratory exposure to increasing intensities of UV-B, UV-A, and visible light significantly decreased P. ramosa infectivity, though there was no effect of spore exposure on parasitic castration of infected hosts. P. ramosa is more sensitive than its Daphnia host to damage by longer wavelength UV-A and visible light; this may enable Daphnia to seek an optimal light environment in the water column, where both UV-B damage and parasitism are minimal. Studies of pathogen light sensitivity help us to uncover factors controlling epidemics in lakes, which is especially important given that water transparency is decreasing in many lakes.

&amp;lt;i&amp;gt;Bacillus cereus&amp;lt;/i&amp;gt; Group Exhibits More Resistant to Chlorhexidine Rather Than &amp;lt;i&amp;gt;Bacillus subtilis&amp;lt;/i&amp;gt; Group

Chlorhexidine is a widely used antiseptic agent; however, its bactericidal effect against bacterial endospores is limited. The genus Bacillus is spore-forming gram-positive bacteria that are ubiquitously found in the environment and cause opportunistic infection and food poisoning. The susceptibility of bacterial endospores to chlorhexidine was previously evaluated in Bacillus subtilis, but the primary target for disinfection with antiseptic agents, including chlorhexidine, should be harmful strains. We aimed to evaluate the susceptibility of harmful cereus group including Bacillus cereus, and to compare that with harmless Bacillus species, containing B. subtilis. We evaluated the susceptibility of the 15 strains of the cereus group to chlorhexidine in comparison with the 5 other strains, named the subtilis group in this study. Our results indicated that chlorhexidine exerted a bacteriostatic effect against Bacillus species at practical concentrations, especially during long-term exposure. The strains of B. cereus group in this study displayed relatively lower susceptibility to the antiseptic than the B. subtilis group according to the minimum inhibitory and bactericidal concentrations. We concluded that there are intrinsic differences in the susceptibility to chlorhexidine between the groups, but the molecular mechanisms are unknown. The minimum inhibitory or bactericidal concentrations of disinfectants other than chlorhexidine may also need to be clarified in the B. cereus and B. subtilis groups.

Zwitterionic small molecule based fluorophores for efficient and selective imaging of bacterial endospores.

In the emerging scenario of increasing antibiotic resistance and pathogen transmission channels, the grave danger posed by bacterial endospores in critical fields like food industry, health and medicine highlights the urgent need to develop efficient probes for their detection; their sturdy and impermeable multilayer coat makes desirable methods like fluorescence imaging extremely difficult. Selective imaging of the endospores in the presence of the bacteria is even more challenging. Furthermore, it is preferable to maintain the dormant state of endospores through the imaging process, if extended monitoring is required; many of the available techniques involve lethal germination or destruction of the endospores. We show that simple zwitterionic diaminodicyanoquinodimethane (DADQ) molecules with selected functionalities are efficient dyes for fluorescence imaging due to their dipolar structure that facilitates the penetration into the endospore system, and the enhanced fluorescence in their rigid/aggregated state. The facile structural tailorability allows DADQs with various appendage moieties to be synthesized; a derivative with ionic substituents (BT2), and another with optimally long alkyl chains and the resultant hydrophobic character (BHADQ) are shown to be excellent fluorescent probes for endospores. Nanomolar amounts of dyes provide effective staining; while BT2 stains bacteria and endospores, most significantly, BHADQ stains endospores selectively. To the best of our knowledge, this is the first example of selective fluorescence imaging of endospores in their dormant state. A range of spectroscopy, microscopy and calorimetry studies provide insight into the molecular level interactions that enable efficient staining and bright images. DADQ fluorophores are photostable and non-cytotoxic, hence useful in practical applications. The versatile structural tailorability of these dye molecules holds great promise for targeted imaging.

Sterilization of Surgical Tools: Removing Bacterial Endospores with a Combination of Povidone-iodine, Chlorhexidine Gluconate, Ethanol, and Methanol

Sterilization of Surgical Tools: Removing Bacterial Endospores with a Combination of Povidone-iodine, Chlorhexidine Gluconate, Ethanol, and Methanol

Endospore Inactivation by Emerging Technologies: A Review of Target Structures and Inactivation Mechanisms.

Recent developments in preservation technologies allow for the delivery of food with nutritional value and superior taste. Of special interest are low-acid, shelf-stable foods in which the complete control or inactivation of bacterial endospores is the crucial step to ensure consumer safety. Relevant preservation methods can be classified into physicochemical or physical hurdles, and the latter can be subclassified into thermal and nonthermal processes. The underlying inactivation mechanisms for each of these physicochemical or physical processes impact different morphological or molecular structures essential for spore germination and integrity in the dormant state. This review provides an overview of distinct endospore defense mechanisms that affect emerging physical hurdles as well as which technologies address these mechanisms. The physical spore-inactivation technologies considered include thermal, dynamic, and isostatic high pressure and electromagnetic technologies, such as pulsed electric fields, UV light, cold atmospheric pressure plasma, and high- or low-energy electron beam.

Nutrient germination improves DNA recovery from industrial Bacillus subtilis endospores during qPCR enumeration assays

Growth-independent microbial enumeration methods such as quantitative PCR require the efficient extraction of genomic DNA from targeted cells. Bacillus endospores are popular inclusions in commercial products due to their hardiness and metabolic dormancy; however, this hardiness is known to render Bacillus endospores resistant to traditional DNA isolation techniques. Metagenomic studies have sought to address this resistance through nutrient-based germination of bacterial endospores in environmental samples. In the present study, we sought to apply this technique to the enumeration of microbial products using an industrial strain of Bacillus subtilis as a model organism. Germination was induced through incubation of axenic spore suspensions in an AGFK-based rich medium. Total spore count, dipicolinic acid release and OD600 absorbance were monitored over time to track the progression of spore populations through the stages of germination and outgrowth. Aerobic plate counts and flow cytometry were used to monitor cell populations for proliferation during the incubation period. Finally, quantitative PCR with taxon-specific primers was used to examine DNA recovery as a function of time. Results show that customized germination protocols, once appropriately validated for the species and product matrix under consideration, can result in more efficient DNA extraction and thus lower limits of detection for qPCR assays targeting industrial Bacillus endospores in microbial products.

Constraints on CaCO3 precipitation in superabsorbent polymer by aerobic bacteria.

Microbially induced CaCO3 precipitation (MICP) can give concrete self-healing properties. MICP agents are typically bacterial endospores which are coated into shelled granules, infused into expanded clay, or embedded into superabsorbent polymer (SAP). When small cracks appear in the cured concrete, the encapsulation is broken and the metabolic CO2 production from the germinated bacteria causes healing of the cracks by precipitation of CaCO3. Such systems are being tested empirically at large scales, but survival of endospores through preparation and application, as well as germination and growth kinetics of the germinated vegetative cells, remains poorly resolved. We encapsulated endospores of Bacillus subtilis and Bacillus alkalinitrilicus in crosslinked acrylamide-based SAP and quantified their germination, growth, and, in the case of B. alkalinitrilicus, CaCO3 precipitation potential. The endospores survived crosslinking and desiccation inside the polymer matrix. Microcalorimetry and microscopy showed that ~ 80% of the encapsulated endospores of both strains readily germinated after rehydration of freeze-dried SAP. Germinated cells grew into dense colonies of cells inside the SAP, and those of B. alkalinitrilicus calcified with up to 0.3g CaCO3 produced per g desiccated SAP when incubated aerobically. Measurements by planar optodes indicated that the precipitation rates were inherently oxygen limited due to diffusional constraints, rather than limited by electron donor or Ca2+ availability. Such oxygen limitation will limit MICP in all water-saturated and oxygen-dependent systems, and MICP agents based on anaerobic bacteria, e.g., nitrate reducers, should be developed to broaden the applicability of bioactive self-healing concretes to wet and waterlogged environments.