Next Article in Journal
Prevalence of invA Gene of Salmonella spp. in Fish and Fishery Resources from Manila Bay Aquaculture Farms Using Real-Time PCR
Previous Article in Journal
Enhancing Human Superorganism Ecosystem Resilience by Holistically ‘Managing Our Microbes’
 
 
Search for Articles:
Title / Keyword
Author / Affiliation / Email
Journal
Article Type
 
 
Advanced Search
 
Section
Special Issue
Volume
Issue
Number
Page
 
Journals
Applied Microbiology
Volume 1
Issue 3
10.3390/applmicrobiol1030032
applmicrobiol-logo
Submit to this Journal Review for this Journal Propose a Special Issue
► Article Menu

Article Menu

share Share announcement Help format_quote Cite question_answer Discuss in SciProfiles thumb_up
...
Endorse textsms
...
Comment
first_page
settings
Order Article Reprints
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Review

Spore Formers as Beneficial Microbes for Humans and Animals

by
Anella Saggese
1,
Loredana Baccigalupi
2 and
Ezio Ricca
1,*
1
Department of Biology, Federico II University of Naples, 80126 Naples, Italy
2
Department of Molecular Medicine and Medical Biotechnologies, Federico II University of Naples, 80126 Naples, Italy
*
Author to whom correspondence should be addressed.
Appl. Microbiol. 2021, 1(3), 498-509; https://doi.org/10.3390/applmicrobiol1030032
Submission received: 29 September 2021 / Revised: 22 October 2021 / Accepted: 23 October 2021 / Published: 29 October 2021
Browse Figures
Versions Notes

Abstract

:
Microorganisms efficiently colonize the external and internal surfaces of the animal body establishing mutually beneficial interactions and forming site- and individual-specific microbiota. The degradation of complex polysaccharides in the animal gut, the production of useful compounds, protection against pathogenic microorganisms and contribution to the development of an efficient immune system are the main beneficial effects of a balanced microbiota. A dysbiosis, an imbalanced composition of the microbiota, has been associated with a large number of diseases from gastro-intestinal or urogenital disorders to allergies, cardiovascular and autoimmune diseases and even to the onset of certain cancers. A growing body of evidence has indicated that probiotic treatments, aimed at maintaining or rebalancing the microbiota, are useful to treat/prevent those illnesses. Lactic Acid Bacteria and Bifidobacteria are the most common microbes used in probiotic preparations; however, other bacteria and yeast cells are also widely used in commercial products. Here we focus on the use of bacterial spore formers as probiotics. Spore formers have been marketed as probiotics for over 50 years and are now extensively used for the treatment of intestinal disorders and as dietary supplements in humans, as growth promoters and competitive exclusion agents in animals.

1. Introduction

Spore formers are bacteria able to form endospores (spores), quiescent cells able to survive at conditions lethal for other cells. Spore production (sporulation) is induced by a variety of environmental conditions that impair cell growth, making the sporulation process a survival strategy. Most spore formers belong to two genera of the Firmicutes phylum: The strict anaerobic Clostridia and the aerobic/facultative anaerobic Bacilli [1]. Recently, a variety of bacteria belonging to other poorly characterized genera of the Firmicutes phylum [2] have been shown to either form spore-like structures [3] or contain sporulation-specific genes [4,5], thus widening the spore formers group.
The sporulation process and the spore structure have been studied in detail in Bacillus subtilis, the model system for spore formers. These bacteria grow by binary fission as long as water and nutrients are available and the environmental conditions allow cell growth (vegetative cycle, Figure 1). When growth is no longer possible, cells enter the sporulation cycle, a microbial example of differentiation: (i) Asymmetric cell division occurs creating two cells of different sizes, a small prespore and a large mother cell; (ii) the mother cell engulfs the prespore, which becomes a protoplast in the mother cell cytoplasm; (iii) both cells contribute to the maturation of the prespore into the mature spore by progressive dehydration of the prespore cytoplasm and by forming a series of protective layers; (iv) the mature spore is released in the environment by the lysis of the mother cell (sporulation cycle, Figure 1). In the environment, the quiescent spore can survive for an extremely long time in the absence of water and nutrients and is resistant to the presence of UV irradiation, toxic chemicals and lytic enzymes and extreme temperatures and pH conditions [6]. The spore, however, continuously senses the environment and responds to the presence of water and nutrients, germinating and creating a new cell able to grow (Figure 1) [7].
Survival of the spore in harsh conditions is due to the structure of the mature spore: a core, the innermost part of the spore, contains a dehydrated cytoplasm with proteins, stable RNAs and DNA and is surrounded by the cortex, a thick peptidoglycan-like layer, and by the coat, a proteinaceous multi-layered structure (Figure 2). In some species, the coat is surrounded by a further structure, the exosporium, which has an irregular shape, is loosely attached to the coat and is formed of glycoproteins [8]. In B. subtilis, which does not have an exosporium, an additional layer is present, namely the crust, made of proteins and glycoproteins and only visible by transmission electron microscopy (TEM) after a ruthenium red staining [6].
While anaerobic spore-formers belonging to the Clostridium genus are well recognized and abundant components of the animal gut microflora [9,10], members of the Bacillus genus are generally considered soil bacteria. Indeed, for their exceptional longevity and resistance, Bacillus spores can be isolated from a variety of different environments including rocks, dust and aquatic environments [11,12]. Bacillus spores are also commonly isolated from the gut of various insects and animals, including humans [13,14]. In recent years, it has been suggested that Bacillus spores enter the animal body as food, water and air contaminants, then temporarily colonize the new habitat, becoming allochthonous gut inhabitants [14,15].
Bacillus spores have been shown to be able to perform an entire life cycle in the animal gut, entering as spores through the oral/nasal route, safely passing the gastric barrier [13,16] and germinating in the intestine [17]. In the intestine, the germination-derived cells proliferate and then re-sporulate in the colon [18,19]. The ability to perform an entire life cycle in the gut is not an exclusive property of members of the B. subtilis species. Other Bacillus species have been shown to germinate and proliferate in the gut of insects [20,21], poultry and pigs [22,23].
The common presence of Bacillus spores in the gastro-intestinal tract (GIT) of animals, the ability of spores and germination-derived cells to interact with intestinal cells and the ability to have beneficial effects for the animal hosts offers a rationale for the use of spore formers in probiotic products. It is, however, likely that such use was originally based on traditional foods containing spores, common in many cultures and considered to have health beneficial effects. The best-known example in this context is Natto, a Japanese traditional healthy food based on soybeans fermented with the B. subtilis var. natto and containing spores. B. subtilis var. natto produces the nattokinase, a strong fibrinolytic enzyme known to reduce systolic and diastolic blood pressure [24], whose intake may be relevant in preventing and treating hypertension [25]. Similar fermented foods are traditionally used in Korea (Chongkukjang), India (kinema), Thailand (thuo nao), Myanmar (pepok), Cambodia and Laos (sieng) and all contain spores of various Bacillus species, such as B. subtilis, B. licheniformis and B. amyloliquefaciens [26]. In addition to these species, other members of the Bacillus genus are currently marketed as ingredients of probiotic preparations for humans and animals. The wide commercial success of spores of Bacillus as probiotics or food ingredients is indubitably also due to the extreme stability of spores and their resistances to the heat treatments used for food preparation as well as long-term storage at room temperature without any loss of viability. Spore resistance overcomes problems associated with the reduction of live bacteria often encountered with probiotic preparations containing lactic acid bacteria and/or Bifidobacteria [27,28].
An overview of the use of Bacillus spores as probiotics is presented here, focusing on the interactions between spores and spore-derived cells with intestinal cells and their beneficial effects.

2. Bacillus Spores and Vegetative Cells Interact with Epithelial and Immune Cells

Both spores and cells of various Bacillus species interact with intestinal and immune cells through complex mechanisms still far from being entirely understood. Over the years, several reports have been published and some representative examples are summarized below.

2.1. Interaction of Bacillus Spores with Epithelial and Immune Cells

A series of in vitro and in vivo reports indicate that the interaction with spores of various Bacillus species modulates the immune response. Duc and collaborators [29] and Ceragioli and collaborators [30] independently analyzed the in vitro interaction of spores of B. subtilis with murine and human macrophages, respectively. In both cases, the efficiency of phagocytosis was modest (about 2.5%), as internalized spores rapidly germinated inside the macrophages and derived bacterial cells were quickly eliminated by both murine and human macrophages [29,30]. In an in vivo study with a murine model, orally administered spores of B. subtilis of a laboratory wild-type strain and an isogenic mutant unable to germinate were both able to induce similar levels of spore-specific fecal sIgA and serum IgG, therefore demonstrating the interaction of spores with immune cells [31]. In a different study, spores of three different Bacillus species, B. subtilis, B. licheniformis and B. flexus, orally dosed to mice, promoted active lymphocyte proliferation within Peyer’s patches and the production of cytokines in mesenteric lymph nodes (MLN) (IL-1α, IL-5, IL-6, IFN-γ and TNF-α) and in the spleen (IFN-γ and TNF-α) [32]. In a rabbit model, spores of B. subtilis and B. anthracis were able to bind IgM through a superantigen-like binding site and drive B cell development in the Gut-Associated Lymphoid Tissue (GALT) [33].
A more recent study showed that B. subtilis spores protected in vitro human keratinocytes from oxidative stress and other chemically induced injuries [34]. The same study also proposed a molecular mechanism for those effects, showing that spores adhered to the keratinocyte cell surface, inducing the nuclear translocation of the transcriptional factor Nrf-2, which in turn activated stress-response genes [34]. Antioxidant activity has also been associated with B. megaterium spores, both in vitro on Caco-2 cells and in vivo on a murine model of dextran sodium sulfate (DSS)-induced oxidative stress [35].

2.2. Interaction of Bacillus Vegetative Cells with Epithelial and Immune Cells

The interaction of vegetative cells of various Bacillus species with model intestinal cells has also been investigated. An early study showed that B. subtilis cells, in combination with cells of Bacteroides fragilis, interacted with intestinal immune cells contributing to the GALT maturation and the development of the pre-immune antibody repertoire in rabbits [36]. Interestingly, such an ability was observed with cells of a wild-type strain of B. subtilis and with cells of isogenic mutants impaired in general stress responses, flagellar movement or biofilm formation but not with cells of isogenic mutants unable to sporulate, suggesting that molecules produced during sporulation were essential for interaction [36]. In the same study, other spore formers such as B. licheniformis or B. pumilus were unable to affect GALT development [36]. A different in vitro study showed that vegetative cells of B. subtilis upregulated the expression of the Toll-like receptors TLR2 and TLR4 [32].
Fujita and co-workers [37] showed that the quorum-sensing pentapeptide ERGMT of B. subtilis, known as CSF (Competence and Sporulation Factor), induced synthesis of the heat-shock (HS) proteins in Caco-2 cells, which in turn prevented oxidant-induced intestinal epithelial cell injuries and loss of barrier function. CSF also showed immunomodulation and cytoprotective activities in vivo in DSS-treated mice, leading the authors to conclude that the pentapeptide has anti-inflammatory properties and that it is potentially effective for the treatment of intestinal inflammatory disorders [38]. Although CSF is a B. subtilis-specific molecule, not produced by other species of the genus, other Bacillus species, such as B. megaterium, B. pumilus and B. clausii, produce and secrete other molecules able to induce HS proteins [39]. The CSF-like molecules of B. megaterium have been characterized as a peptide smaller than 3 kDa that was also able to induce p38 MAPK phosphorylation in HT29 cells, associated with a protective response against the excess of inflammation [39]. The same strain was also able to produce a different molecule, larger than 3 kDa, responsible for strong induction of PKB/Akt phosphorylation, altogether indicating the SF185 strain of B. megaterium was able to alert epithelial cells against stressful conditions through secreted molecules [39].
CSF of B. subtilis and other molecules with a similar effect produced by various Bacillus species, being produced and secreted by live cells, belong to the class of post-biotics. The production of post-biotics by Bacillus cells is a still largely unexplored field that is likely to become the object of intense future research activities.

3. Beneficial Effects of Bacillus Probiotics

Although a variety of potential beneficial effects have been associated with Bacillus probiotics, clinical data on humans or tests on farmed animals are still limited. Most studies have been performed on animal models and have confirmed the potential of Bacillus spores and cells as probiotics. An example comes from an in vivo study performed with a mouse model of infection in which B. subtilis spores were shown to reduce the susceptibility to the mouse pathogen Citrobacter rodentium [40]. C. rodentium is an enteric pathogen that colonizes the mouse distal colon causing epithelial lesions (crypt hyperplasia, mucosal thickening with T-cell infiltration, a highly polarized Th1 immune response and epithelial cell proliferation) similar to those caused in humans by enteropathogenic (EPEC) and enterohemorrhagic (EHEC) strains of E. coli [41]. Oral administration of B. subtilis spores one day before infection with C. rodentium was effective in preventing the enteropathy and drastically reducing the mortality rate in mice [40]. The reduced susceptibility to enteric pathogens caused by the ingested spores has been explained by either a “competitive exclusion” effect, with spores physically blocking the interaction between the pathogen and the intestinal cells, or the ability of spores and germination-derived cells to interact with immune cells stimulating the GALT and an adaptive immune response [40].
Many different species and strains of spore formers have been evaluated for their beneficial effects on humans and animals. In some cases, species/strains with additional beneficial properties have also been considered, such as spores of Bacillus strains that produce gastric-stable and strongly antioxidant carotenoids. Cells (but not spores) of a strain of B. indicus, producing a yellow/orange carotenoid [42], reduced plasma markers of inflammation and oxidative markers in a rat model of diet-induced metabolic syndrome, indicating that the use of carotenoid-producing strains may add additional benefits to the probiotic properties [43].
Bacillus probiotics have also been found to be effective against viral infections. In early work, heat-inactivated spores of B. subtilis intranasally administered to mice were able to partially protect (60% survival) the host in a challenge experiment with the influenza virus H5N2 (5 LD50) indicating the role of innate immunity and its stimulation by spores in protecting the host [44]. Killed spores were shown to stimulate TLR-mediated expression of NF-kB, control cytokine production and recruit NK cells into lungs, inducing the maturation of dendritic cells DCs [44]. More recently, it has been reported that a mixture of B. clausii strains is able to protect against Rotavirus infections in vitro [45].
Those reported above are examples of tests performed in vitro or in vivo on animal models. In the following sections, examples of beneficial effects due to Bacillus probiotics on farmed animals and on humans are reported. These selected examples are clearly not exhaustive of the reports present in the literature but show that a variety of species/strains have been used on a variety of hosts, analyzing a variety of different phenotypes. Therefore, in most cases, it is difficult to compare the results of the different studies. Analysis of the literature highlights the lack of a systematic approach to study the effect of Bacillus probiotics and underlines the need for research efforts focused on the understanding of the scientific bases of the observed effects.

3.1. Safety of Bacillus Probiotics

In Europe, a number of criteria set by the European Food Safety Authority (EFSA) must be satisfied for a Bacillus strain in order to be considered and commercialized as a probiotic. First, it has to belong to a list of approved QPS (Qualified Presumption of Safety) species (Table 1).
In addition, each specific strain belonging to the species listed in Table 1 has to carry no significant resistance to a panel of eight antibiotics (Table 2) and exhibit no apparent toxicity in in vitro assays with cell lines (for example, HT29, Vero or Caco2 cells). A strain that meets all these requirements can be assigned the QPS status and commercialized in Europe.
In the USA, in order for a strain to be considered as a probiotic and commercialized, it must be recognized as being Generally Recognized As Safe (GRAS), a status that can be obtained through the submission of a self-affirmed dossier to the Food and Drugs Administration (FDA).

3.2. Bacillus Probiotics for Animal Use

Many countries have strictly limited the use of antibiotics, totally banning their use as growth promoters for farmed animals. As a consequence, the search for alternatives to antibiotics has greatly increased, and probiotics, including Bacillus probiotics, have been identified as a potential solution [46]. The use of biological agents (probiotics) as feed ingredients is strictly regulated as indicated above, and Table 3 and Table 4 report examples of spore-based products authorized to be marketed for animal use and aquaculture, respectively.
Examples of Bacillus probiotics tested in farmed animals include studies performed on chickens infected with Escherichia coli O78:K80, Salmonella enterica or Clostridium perfringens that were protected when pre-dosed with B. subtilis spores [47,48]. In a different study with chickens, diet supplementation with a mixture of B. licheniformis and B. subtilis spores resulted in increased weight gain and improved feed conversion [49]. The same mixture of B. licheniformis and B. subtilis spores was also used to feed pigs for a period of 23 weeks and was shown to reduce the incidence of diarrhea and mortality compared with a control group [50]. B. cereus var. toyoi, a non-toxigenic and non-pathogenic strain of B. cereus, reduced diarrhea and morbidity in piglets challenged with Salmonella [51] and improved weight gain after 6 months of treatment [52].
Bacillus probiotics have been also used in aquaculture, as a feed supplement or directly added to the water, to improve disease resistance and increase farming productivity [53]. The supplementation of spores during egg hatching and the first stages of larvae development influenced the microbial gut composition allowing for manipulation of the microbiota. A strain of B. subtilis was shown to improve the survival of white shrimp, Litopenaeus vannamei, larvae [54] and to increase weight gain in a dose-dependent manner and colonize Epinephelus coioides, protecting it against bacterial (Streptococcus sp.) and viral (Iridovirus) infections [55]. A strain of B. amyloliquefaciens protected Catla catla in a challenge experiment with the pathogen Edwardsiella tarda, improving systemic and mucosal immunological parameters [56].

3.3. Bacillus Probiotics for Human Use

Probiotics for human use are obviously strictly regulated by international agencies as indicated in Section 3.1, and Table 5 reports examples of spore-based products authorized to be marketed for human use.
Furthermore, in this case, a large number of reports are present in the literature, but in this review, we will limit the discussion to a few, select examples.
Piewngam and collaborators [57], upon screening human populations from rural and urbanized areas, observed an inverse correlation between the presence of Bacillus species (mainly B. subtilis) and the absence of the pathogen Staphylococcus aureus. In particular, S. aureus was never detected in fecal samples of adults when Bacillus species were present. Interestingly, such pathogen exclusion was not limited to the site of interaction (the gut) but was also observed in other colonization sites. Indeed, S. aureus never colonized the nasal mucosa in the presence of intestinal Bacillus, even if Bacillus was not present in the nose [57]. In the same study, the authors showed that the cyclic lipopeptide fengycin, produced by most B. subtilis strains, competed with the AIP signal molecule of the Agr quorum-sensing system of S. aureus, essential for the virulence of the pathogen [57]. Moreover, the oral administration of a B. subtilis strain, but not an isogenic mutant not producing fengycin, completely blocked S. aureus colonization in mice, elegantly demonstrating the mechanism of action of B. subtilis protection against S. aureus infections [57].
Another example comes from a recent study in which a carotenoid-producing strain of B. indicus was shown to deliver its carotenoid to the human body causing an increased systemic carotenoid accumulation and improving the barrier functions [58]. In this study, over 60 healthy overweight or obese volunteers showed increased concentrations of bacterial carotenoid in plasma samples after three and six weeks of daily supplementation of the carotenoid-producing probiotic [58].

4. Future Development

Although the use of bacterial spores in commercial probiotic products is already common practice (Table 3, Table 4 and Table 5), new studies addressing the health beneficial effects of Bacillus probiotics, focused on the isolation of novel strains and on the mechanisms by which they exert their beneficial effects on humans and animals, are continuously arising.
Some of these studies are opening new fields in the Bacillus probiotic world, proposing the Bacillus spore as a platform to deliver beneficial molecules, thus combining the probiotic and additive beneficial effects. Other studies have highlighted that Bacillus probiotics may have beneficial effects on neurodegenerative diseases, opening the possible use of probiotics for new therapeutic targets. Examples of such applications and new therapeutic targets are summarized below.

4.1. Functional Spores as Probiotics

A strategy has recently been developed to functionalize spores by adsorbing active molecules on their surface [59]. The strategy has been initially developed using a laboratory collection strain of B. subtilis and later tested on a probiotic strain of B. subtilis [60] and B. megaterium [61]. This strategy allowed a non-recombinant display of enzymes that are stabilized by the interaction with the spore and keep their activity at conditions that also inactivate the free enzymes [59,61]. Spore adsorption is based on the spontaneous binding of molecules to the spore surface without the need for chemical treatment (crosslink) and has been recently reviewed [62]. Being a non-recombinant strategy, spore adsorption allows the functionalization of spores of probiotic strains, adding new beneficial effects, due to the adsorbed molecule, without posing any safety issues. A simple example could be the adsorption of enzymes for the degradation of detrimental molecules, such as food components that induce allergies or intolerances in specific groups of people. However, functional spores have not been tested yet on animal models and their development is still in progress [62].

4.2. Bacillus Probiotics and the Nervous System

Two recent papers have shown that Bacillus probiotics may have beneficial effects on neurodegenerative diseases [63,64], opening a new frontier to the beneficial effects associated with Bacillus probiotics. Goya et al. showed that both spores and cells of various B. subtilis strains, through secreted molecules and biofilm formation, inhibit α-synuclein aggregation and clear preformed aggregates [64]. Cogliati et al. identified the Competence and Sporulation Factor (CSF) of B. subtilis, together with biofilm formation, as involved in the anti-Alzheimer’s Disease (AD) effects observed in a C. elegans model of AD [63]. In such a model, B. subtilis delayed age-related neurodegeneration and cognitive damage and alleviated paralysis defects and behavioral deficits, all induced in transgenic worms by the expression of the Aβ peptide [63]. These two studies, although performed on a C. elegans model, offer exciting possibilities for probiotic treatments and the development of drug therapies based on Bacillus molecules to target neurodegenerative diseases.

5. Conclusions

This review summarizes select examples of the interaction of spores and/or cells of various Bacillus species with epithelial and immune cells, and the beneficial effects exerted by Bacillus probiotics in in vitro and in vivo model animals, in farmed animals and in humans. Rather than being exhaustive of what is present in the literature, these selected examples aim to provide an overview of what is known about various Bacillus species/strains, various hosts and various phenotypes, underlining the need for a more systematic approach to study Bacillus probiotics. Focusing on a single species/strain, a single host and looking at selected phenotypes would it make possible to compare results and address the molecular basis of the effects asserted.
Particularly promising is the possibility of using spores of probiotic strains to deliver health-beneficial molecules to the intestinal or nasal mucosa. Such non-recombinant delivery would combine the beneficial effects of the probiotic to those of the added molecule, with the possibility to deliver active enzymes to degrade food components with allergenic potentials or anti-inflammatory or antioxidant properties. Although these functionalized spore probiotics have not been tested yet in vivo, spores displaying antigens have long been used as oral and nasal vaccines able to induce protective antigen-specific immune responses [62], therefore providing a solid base for their safety and beneficial potentials.

Author Contributions

A.S., L.B. and E.R. wrote, edited and revised the manuscript draft. All authors have read and agreed to the published version. All authors have read and agreed to the published version of the manuscript.

Funding

No specific funding was received to support this work.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Fritze, D. Taxonomy and systematics of the aerobic endospore forming bacteria: Bacillus and related genera. In Bacterial Spore Formers; Ricca, E., Henriques, A.O., Cutting, S.M., Eds.; Horizon Biosience: Norfolk, UK, 2004; pp. 17–34. [Google Scholar]
  2. Yutin, N.; Galperin, M.Y. A genomic update on clostridial phylogeny: Gram-negative spore formers and other misplaced clostridia. Environ. Microbiol. 2013, 15, 2631–2641. [Google Scholar] [CrossRef] [PubMed]
  3. Browne, H.P.; Foster, S.C.; Anonye, B.O.; Kumar, N.; Neville, B.A.; Stares, M.D.; Goulding, D.; Lawley, T.D. Culturing of ‘unculturable’ human microbiota reveals novel taxa and extensive sporulation. Nature 2016, 533, 543–546. [Google Scholar] [CrossRef] [Green Version]
  4. Galperin, M.Y.; Mekhedov, S.L.; Puigbo, P.; Smirnov, S.; Wolf, Y.I.; Rigden, D.J. Genomic determinants of sporulation in Bacilli and Clostridia: Towards the minimal set of sporulation-specific genes. Environ. Microbiol. 2012, 14, 2870–2890. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  5. Egan, M.; Dempsey, E.; Ryan, A.C.; Ross, P.R.; Stanton, C. The Sporobiota of the human gut. Gut Microbes 2021, 13, e1863134. [Google Scholar] [CrossRef]
  6. McKenney, P.T.; Driks, A.; Eichenberger, P. The Bacillus subtilis endospore: Assembly and functions of the multilayered coat. Nat. Rev. Microbiol. 2013, 11, 33–44. [Google Scholar] [CrossRef]
  7. Christie, G.; Setlow, P. Bacillus spore germination: Knowns, unknowns and what we need to learn. Cell. Signal. 2020, 74, 109729. [Google Scholar] [CrossRef]
  8. Henriques, A.O.; Moran, C.P., Jr. Structure, assembly, and function of the spore surface layers. Ann. Rev. Microbiol. 2007, 61, 555–588. [Google Scholar] [CrossRef] [PubMed]
  9. Eckburg, P.B.; Bik, E.M.; Bernstein, C.N.; Purdom, E.; Dethlefsen, M.; Sargent, L.; Gill, S.R.; Nelson, K.E.; Relman, D.A. Diversity of the human intestinal microbial flora. Science 2005, 308, 1635–1638. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  10. Mahowald, M.A.; Rey, F.E.; Seedorf, H.; Turnbaugh, P.J.; Fulton, R.S. Characterizing a model human gut microbiota composed of members of its two dominant bacterial phyla. Proc. Natl. Acad. Sci. USA 2009, 106, 5859–5864. [Google Scholar] [CrossRef] [Green Version]
  11. Nicholson, W.L. Roles of Bacillus endospores in the environment. Cell. Mol. Life Sci. 2002, 59, 410–416. [Google Scholar] [CrossRef]
  12. Nicholson, W.L. Ubiquity, longevity, and ecological roles of Bacillus spores. In Bacterial Spore Formers; Ricca, E., Henriques, A.O., Cutting, S.M., Eds.; Horizon Biosience: Norfolk, UK, 2004; pp. 1–15. [Google Scholar]
  13. Fakhry, S.; Sorrentini, I.; Ricca, E.; De Felice, M.; Baccigalupi, L. Characterization of spore forming Bacilli isolated from the human gastrointestinal tract. J. Appl. Microbiol. 2008, 105, 2178–2186. [Google Scholar] [CrossRef]
  14. Hong, H.A.; To, E.; Fakhry, S.; Baccigalupi, L.; Ricca, E.; Cutting, S.M. Defining the natural habitat of Bacillus spore-formers. Res. Microbiol. 2009, 160, 375–379. [Google Scholar] [CrossRef] [PubMed]
  15. Cutting, S.M.; Hong, H.A.; Baccigalupi, L.; Ricca, E. Oral Vaccine Delivery by Recombinant Spore Probiotics. Int. Rev. Immunol. 2009, 28, 487–505. [Google Scholar] [CrossRef] [PubMed]
  16. Spinosa, M.R.; Braccini, T.; Ricca, E.; De Felice, M.; Morelli, L.; Pozzi, G.; Oggioni, M.R. On the fate of ingested Bacillus spores. Res. Microbiol. 2000, 151, 361–368. [Google Scholar] [CrossRef]
  17. Casula, G.; Cutting, S.M. Bacillus probiotics: Spore germination in the gastrointestinal tract. Appl. Environ. Microbiol. 2002, 68, 2344–2352. [Google Scholar] [CrossRef] [Green Version]
  18. Hoa, T.-T.; Duc, L.H.; Isticato, R.; Baccigalupi, L.; Ricca, E.; Van, P.H.; Cutting, S.M. Fate and dissemination of Bacillus subtilis spores in a murine model. Appl. Environ. Microbiol. 2001, 67, 3819–3823. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  19. Tam, N.K.; Uyen, N.Q.; Hong, H.A.; Duc, L.H.; Hoa, T.T.; Serra, C.R.; Henriques, A.O.; Cutting, S.M. The intestinal life cycle of Bacillus subtilis and close relatives. J. Bacteriol. 2006, 188, 2692–2700. [Google Scholar] [CrossRef] [Green Version]
  20. Margulis, L.; Jorgensen, J.Z.; Dolan, S.; Kolchinsky, R.; Rainey, F.A.; Lo, S.C. The Arthromitus stage of Bacillus cereus: Intestinal symbionts of animals. Proc. Natl. Acad. Sci. USA 1998, 95, 1236–1241. [Google Scholar] [CrossRef] [Green Version]
  21. Feinberg, L.; Jorgensen, J.; Haselton, A.; Pitt, A.; Rudner, R.; Margulis, L. Arthromitus (Bacillus cereus) symbionts in the cockroach Blaberus giganteus: Dietary influences on bacterial development and population density. Symbiosis 1999, 2, 109–123. [Google Scholar]
  22. Jadamus, A.; Vahjen, W.; Simon, O. Growth behaviour of a spore forming probiotic strain in the gastrointestinal tract of broiler chicken and piglets. Arch. Anim. Nutr. 2001, 54, 1–17. [Google Scholar] [CrossRef]
  23. Jadamus, A.; Vahjen, W.; Schafer, K.; Simon, O. Influence of the probiotic strain Bacillus cereus var. toyoi on the development of enterobacterial growth and on selected parameters of bacterial metabolism in digested samples of piglets. J. Anim. Physiol. Anim. Nutr. 2002, 86, 42–54. [Google Scholar] [CrossRef]
  24. Sumi, H.; Hamada, H.; Tsushima, H.; Mihara, H.; Muraki, H. A novel fibrinolytic enzyme (nattokinase) in the vegetable cheese Natto; a typical and popular soybean food in the Japanese diet. Experientia 1987, 43, 1110–1111. [Google Scholar] [CrossRef]
  25. Kim, J.Y.; Gum, S.N.; Paik, J.K.; Lim, H.H.; Kim, K.C.; Ogasawara, K.; Inoue, K.; Park, S.; Jang, Y.; Lee, J.H. Effects of nattokinase on blood pressure: A randomized, controlled trial. Hypertens. Res. 2008, 31, 1583–1588. [Google Scholar] [CrossRef] [Green Version]
  26. Chang, H.C. Healthy and safe Korean traditional fermented foods: Kimki and chongkukjang. J. Ethnic Foods 2018, 5, 161–166. [Google Scholar] [CrossRef]
  27. Binda, S.; Hill, C.; Johansen, E.; Obis, D.; Pot, B.; Sanders, M.E.; Tremblay, A.; Ouwehand, A.C. Criteria to qualify microorganisms as “Probiotic” in foods and dietary supplements. Front. Microbiol. 2020, 11, 1662. [Google Scholar] [CrossRef]
  28. Fenster, K.; Freeburg, B.; Hollard, C.; Wong, C.; Laursen, R.R.; Ouwehand, A.C. The Production and Delivery of Probiotics: A Review of a Practical Approach. Microorganisms 2019, 7, 83. [Google Scholar] [CrossRef] [Green Version]
  29. Duc, L.H.; Hong, A.H.; Nguyen, Q.U.; Cutting, S.M. Intracellular fate and immunogenicity of B. subtilis spores. Vaccine 2004, 22, 1873–1885. [Google Scholar] [CrossRef] [PubMed]
  30. Ceragioli, M.; Cangiano, G.; Esin, S.; Ghelardi, E.; Ricca, E.; Senesi, S. Phagocytosis, germination and killing of Bacillus subtilis spores presenting heterologous antigens in human macrophages. Microbiology 2009, 155, 338–346. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  31. Duc, L.H.; Hong, H.A.; Fairweather, N.; Ricca, E.; Cutting, S.M. Bacterial spores as vaccine vehicles. Infect. Immun. 2003, 71, 2810–2818. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  32. Huang, J.-M.; La Ragione, R.; Nunez, A.; Cutting, S.M. Immunostimulatory activity of Bacillus spores. FEMS Immunol. Med. Microbiol. 2008, 53, 195–203. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  33. Severson, K.M.; Mallozzi, M.; Driks, A.; Knight, K.L. B cell development in GALT: Role of bacterial superantigen-like molecules. J. Immunol. 2010, 184, 6782–6789. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  34. Petruk, G.; Donadio, G.; Lanzilli, M.; Isticato, R.; Monti, D.M. Alternative use of Bacillus subtilis spores: Protection against environmental oxidative stress in human normal keratinocytes. Sci. Rep. 2018, 8, 1745. [Google Scholar] [CrossRef]
  35. Mazzoli, A.; Donadio, G.; Lanzilli, M.; Saggese, A.; Guarino, A.M.; Rivetti, M.; Crescenzo, R.; Ricca, E.; Ferrandino, I.; Iossa, S.; et al. Bacillus megaterium SF185 spores exert protective effects against oxidative stress in vivo and in vitro. Sci. Rep. 2019, 9, 12082. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  36. Rhee, K.-J.; Sethupathi, P.; Driks, A.; Lanning, D.K.; Knight, K.L. Role of commensal bacteria in development of gut-associated lymphoid tissue and preimmune antibody repertoire. J. Immunol. 2004, 172, 1118–1124. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  37. Fujita, M.; Musch, M.W.; Nakagawa, Y.; Hu, S.; Alverdy, J.; Kohgo, Y.; Schneewind, O.; Jabri, B.; Chang, E.B. The Bacillus subtilis quorum-sensing molecule CSF contribute to intestinal homoestasis via OCTN2, a host cell membrane transporter. Cell Host Microbe 2007, 1, 299–308. [Google Scholar] [CrossRef] [Green Version]
  38. Okamoto, K.; Fujita, M.; Nata, T.; Ueno, N.; Inaba, Y.; Ishikawa, C.; Ito, T.; Moriichi, K.; Tanabe, H.; Mizukami, Y.; et al. Competence and sporulation factor derived from Bacillus subtilis improves epithelial cell injury in intestinal inflammation via immunomodulation and cytoprotection. Int. J. Colorectal Dis. 2012, 27, 1039–1046. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  39. Di Luccia, B.; D’Apuzzo, E.; Varriale, F.; Baccigalupi, L.; Ricca, E.; Pollice, A. Bacillus megaterium SF185 induces stress pathways and affects the cell cycle distribution of human intestinal epithelial cells. Benef. Microbes 2016, 7, 609–620. [Google Scholar] [CrossRef] [Green Version]
  40. D’Arienzo, R.; Maurano, F.; Mazzarella, G.; Luongo, D.; Stefanile, R.; Ricca, E.; Rossi, M. Bacillus subtilis spores reduce susceptibility to Citrobacter rodentium-mediated enteropathy in a mouse model. Res. Microbiol. 2006, 157, 891–897. [Google Scholar] [CrossRef] [PubMed]
  41. Schauer, D.B.; Falkow, S. The eae gene of Citrobacter freundii biotype 4280 is necessary for colonization in transmissible murine colonic hyperplasia. Infect. Immun. 1993, 61, 4654–4661. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  42. Khaneja, R.; Perez-Fons, L.; Fakhry, S.; Baccigalupi, L.; Steiger, S.; To, E.; Sandmann, G.; Dong, T.C.; Ricca, E.; Fraser, P.D.; et al. Carotenoids Found in Bacillus. J. Appl. Microbiol. 2010, 108, 1889–1902. [Google Scholar] [PubMed]
  43. Crescenzo, R.; Mazzoli, A.; Cancelliere, R.; Bucci, A.; Naclerio, G.; Baccigalupi, L.; Cutting, S.M.; Ricca, E.; Iossa, S. Beneficial effects of carotenoid-producing cells of Bacillus indicus HU16 in a rat model of diet-induced metabolic syndrome. Benef. Microbes 2017, 8, 823–831. [Google Scholar] [CrossRef] [PubMed] [Green Version]
  44. Song, M.; Hong, H.A.; Huang, J.-M.; Colenutt, C.; Khang, D.D.; Nguyen, T.V.A.; Park, S.-M.; Shim, B.-S.; Song, H.H.; Cheon, I.S.; et al. Killed Bacillus subtilis spores as a mucosal adjuvant for an H5N1 vaccine. Vaccine 2012, 30, 3266–3277. [Google Scholar] [CrossRef]
  45. Paparo, L.; Tripodi, L.; Bruno, C.; Pisapia, L.; Damiano, C.; Pastore, L.; Berni Canani, R. Protective action of Bacillus clausii probiotic strains in an in vitro model of Rotavirus infection. Sci. Rep. 2020, 10, 12636. [Google Scholar] [CrossRef]
  46. Mingmongkolchai, S.; Panbangred, W. Bacillus probiotics: An alternative to antibiotics for livestock production. J. Appl. Microbiol. 2018, 124, 1334–1346. [Google Scholar] [CrossRef] [PubMed]
  47. La Ragione, R.M.; Casula, G.; Cutting, S.M.; Woodward, M.J. Bacillus subtilis spores competitively exclude Escherichia coli O78:K80 in poultry. Vet. Microbiol. 2001, 79, 113–142. [Google Scholar] [CrossRef]
  48. La Ragione, R.M.; Woodward, M.J. Competitive exclusion by Bacillus subtilis spores of Salmonella enterica serotype Enteritidis and Clostridium perfringens in young chickens. Vet. Microbiol. 2003, 94, 245–256. [Google Scholar] [CrossRef]
  49. Šabatková, J.; Kumprecht, I.; Zoba, P.; Suchý, P.; Čermák, B. The Probiotic BioPlus 2B as an Alternative to Antibiotics in Diets for Broiler Chickens. Acta Vet. Brno 2008, 77, 569–574. [Google Scholar]
  50. Alexopoulos, C.; Georgoulakis, I.E.; Tzivara, A.; Kyriakis, C.S.; Govaris, A.; Kyriakis, S.C. Field evaluation of the effect of a probiotic containing Bacillus licheniformis and Bacillus subtilis spores on the health status, performance, and carcass quality of grower and finisher pigs. J. Vet. Med. A Physiol. Pathol. Clin. Med. 2004, 51, 306–312. [Google Scholar] [CrossRef] [PubMed]
  51. Scharek-Tedin, L.; Pieper, R.; Vahjen, W.; Tedin, K.; Neumann, K.; Zentek, J. Bacillus cereus var. Toyoi modulates the immune reaction and reduces the occurrence of diarrhea in piglets challenged with Salmonella Typhimurium DT104. J. Anim. Sci. 2013, 91, 5696–5704. [Google Scholar] [CrossRef]
  52. Williams, L.D.; Burdock, G.A.; Jimenez, G.; Castillo, M. Literature review on the safety of Toyocerin, a non-toxigenic and nonpathogenic Bacillus cereus var. toyoi preparation. Toxicol. Pharmacol. 2009, 55, 236–246. [Google Scholar]
  53. Mohapatra, S.; Chakraborty, T.; Kumar, V.; Deboeck, G.; Mohanta, K.N. Aquaculture and stress management: A review of probiotic intervention. J. Anim. Physiol. Anim. Nutr. 2013, 97, 405–430. [Google Scholar] [CrossRef] [PubMed]
  54. Liu, K.F.; Chiu, C.S.; Shiu, Y.L.; Cheng, W.; Liu, C.H. Effects of the probiotic, Bacillus subtilis E20, on the survival, development, stress tolerance, and immune status of white shrimp, Litopenaeus vannamei larvae. Fish Shellfish Immunol. 2010, 28, 837–844. [Google Scholar] [CrossRef]
  55. Liu, C.; Chiu, C.; Wang, S.; Cheng, W. Dietary administration of the probiotic, Bacillus subtilis E20, enhances the growth, innate immune responses, and disease resistance of the grouper, Epinephelus coioides. Fish Shellfish Immunol. 2012, 33, 699–706. [Google Scholar] [CrossRef] [PubMed]
  56. Das, A.; Nakhro, K.; Chowdhury, S.; Kamilya, D. Effect of potential probiotic Bacillus amyloliquefaciens FPTB16 on systemic and cutaneous mucosal immune responses and desease resistance of catla (Catla catla). Fish Shellfish Immunol. 2013, 35, 1547–1553. [Google Scholar] [CrossRef]
  57. Piewngam, P.; Zheng, Y.; Nguyen, T.H.; Dickey, S.W.; Joo, H.S.; Villaruz, A.E.; Glose, K.A.; Fisher, E.L.; Hunt, R.L.; Li, B.; et al. Pathogen elimination by probiotic Bacillus via signaling interference. Nature 2018, 562, 532–537. [Google Scholar] [CrossRef]
  58. Stevens, Y.; Pinheiro, I.; Salden, B.; Duysburgh, C.; Bolca, S.; Degroote, J.; Majdeddin, M.; Van Noten, N.; Gleize, B.; Caris-Veyrat, C.; et al. Effect of a carotenoid-producing Bacillus strain on intestinal barrier integrity and systemic delivery of cartenoids: A randomised trial in animals and humans. J. Funct. Foods 2021, 80, 104445. [Google Scholar] [CrossRef]
  59. Sirec, T.; Strazzulli, A.; Isticato, R.; De Felice, M.; Moracci, M.; Ricca, E. Adsorption of beta-galactosidase of Alicyclobacillus acidocaldarius on wild type and mutant spores of Bacillus subtilis. Microb. Cell Factories 2012, 11, 100. [Google Scholar] [CrossRef] [Green Version]
  60. Sirec, T.; Cangiano, G.; Baccigalupi, L.; Ricca, E.; Isticato, R. Human intestinal isolates of Bacillus subtilis: Characterization of the spore surface and use as display systems. FEMS Microbiol. Lett. 2014, 358, 194–201. [Google Scholar] [CrossRef] [Green Version]
  61. Lanzilli, M.; Donadio, G.; Fusco, F.A.; Sarcinelli, C.; Limauro, D.; Ricca, E.; Isticato, R. Display of the peroxiredoxin Bcp1 of Sulfolobus solfataricus on probiotic spores of Bacillus megaterium. New Biotechnol. 2018, 46, 38–44. [Google Scholar] [CrossRef]
  62. Ricca, E.; Baccigalupi, L.; Isticato, R. Spore-adsorption: Mechanism and applications of a non-recombinant display system. Biotechnol. Adv. 2021, 47, 107693. [Google Scholar] [CrossRef]
  63. Cogliati, S.; Clementi, V.; Francisco, M.; Crespo, C.; Arfanaraz, F.; Grau, R. Bacillus subtilis delays neurodegeneration and behavioral impairment in the Alzheimer’s disease model Caenorhabditis elegans. J. Alzheimer’s Dis. 2020, 73, 1035–1052. [Google Scholar] [CrossRef] [PubMed]
  64. Goya, M.A.; Xue, F.; Sampedro-Torres-Quevedo, C.; Arnaouteli, S.; Riquelme-Dominquez, L.; Romanowski, A.; Brydon, J.; Ball, K.L.; Stanley-Wall, N.R.; Doitsidou, M. Probotic Bacillus subtilis protects against alpha-synucleinaggregation in C. elegans. Cell Rep. 2020, 30, 367–380. [Google Scholar] [CrossRef] [PubMed]
Applmicrobiol 01 00032 g001 550
Figure 1. Vegetative and sporulation cycles of spore formers. The vegetative cycle occurs via binary fission: A cell divides symmetrically creating two identical daughter cells that grow and divide again. The sporulation cycle starts with asymmetrical cell division that creates a large mother cell and a small prespore. Then the prespore becomes a protoplast included in the mother cell cytoplasm. The formation of a series of protective layers (cortex, coat, exosporium or crust) leads to a mature spore and its release in the environment. The spore can germinate, creating a new vegetative cell able to grow and eventually sporulate again.
Figure 1. Vegetative and sporulation cycles of spore formers. The vegetative cycle occurs via binary fission: A cell divides symmetrically creating two identical daughter cells that grow and divide again. The sporulation cycle starts with asymmetrical cell division that creates a large mother cell and a small prespore. Then the prespore becomes a protoplast included in the mother cell cytoplasm. The formation of a series of protective layers (cortex, coat, exosporium or crust) leads to a mature spore and its release in the environment. The spore can germinate, creating a new vegetative cell able to grow and eventually sporulate again.
Applmicrobiol 01 00032 g001
Applmicrobiol 01 00032 g002 550
Figure 2. A B. subtilis spore observed by transmission electron microscopy. Core, cortex, inner and outer coat are indicated. The crust is not visible since this sample has not been stained with ruthenium red.
Figure 2. A B. subtilis spore observed by transmission electron microscopy. Core, cortex, inner and outer coat are indicated. The crust is not visible since this sample has not been stained with ruthenium red.
Applmicrobiol 01 00032 g002
Table
Table 1. List of approved QPS Bacillus species *.
Table 1. List of approved QPS Bacillus species *.
Bacillus amyloliquefaciens Bacillus atrophaeusBacillus megaterium
Bacillus circulansBacillus mojavensis
Bacillus clausiiBacillus paralicheniformis Bacillus pumilus
Bacillus coagulansBacillus smithii
Bacillus flexusBacillus subtilis
Bacillus fusiformisBacillus vallismortis
Bacillus lentusBacillus velenzensis
Bacillus licheniformis
* Updated by EFSA on 7 July 2021.
Table
Table 2. Panel of antibiotics and relative breakpoints for Bacillus probiotics *.
Table 2. Panel of antibiotics and relative breakpoints for Bacillus probiotics *.
AntibioticMicrobiological Breakpoint (mL/L)
Vancomycin4
Gentamycin4
Kanamycin8
Streptomycin8
Erythromycin4
Clindamycin4
Quinupristin + Dalfopristin4
* data from The EFSA Journal (2008) 732, 1–15.
Table
Table 3. Bacillus probiotics for veterinary use.
Table 3. Bacillus probiotics for veterinary use.
ProductManufacturerSpecies (Spores/Dose) *Animal
AlCareAlpharma Inc. (Australia)B. licheniformis NCTC 13123 (109–1010)Swine
BioGrowProvita Eurotech Ltd. (UK)B. licheniformis1 (1.6 × 109) and B. subtilis 1 (1.6 × 109)Poultry, calves and swine
BioPlus 2BChristian Hansen Hoersholm (Denmark)B. licheniformis DSM 5749(1.6 × 109) and B. subtilis DSM 5750 (1.6 × 109)Piglets, chickens, turkeys
Esporafeed PlusNorel, S.A. (Spain)B. cereus CECT 953 (109)Swine
LactopurePharmed Medicare (India)B. coagulans1Poultry, calves and swine
Neoferm BS 10Sanofi Sante Nutrition Animale (France)B. clausii CNCM MA23/3V and CNCM MA66/4MPoultry, calves and swine
ToyocerinAsahi Vet S.A. (Japan)B. cereus var. toyoi NCIMB-40112/CNCM-1012 (>1010)Calves, poultry, rabbits and swine.
1 No information available on strain name. * Spore /dose indicated where information is available.
Table
Table 4. Bacillus probiotics for aquaculture.
Table 4. Bacillus probiotics for aquaculture.
ProductManufacturerSpecies (Spores/Dose) *
BiostartMicrobial Solutions (South Africa); Advanced Microbial Systems (USA)B. megaterium1, B. licheniformis1, P. polymyxa1 and B. subtilis 1
BioZyme-AquaSino-Aqua Corp. (Taiwan)B. subtilis Wu-S and Wu-T (108)
LiqualifeCargill (USA)Bacillus ssp. 1
PromarineSino-Aqua Corp. (Taiwan)B. subtilis1
Sanocare SanolifeINVE TechnologiesBacillus ssp. 1
SanoguardBelgiumBacillus ssp. 1
1 No information available on strain name. * Spore/dose indicated where information is available.
Table
Table 5. Bacillus probiotics for human use.
Table 5. Bacillus probiotics for human use.
ProductManufacturerSpecies (Spores/Dose) *
BactisubtilMarion Merrell (France); Casella-Med (Germany)B. cereus ATCC 14893 (109)
BibactylUPHACE (Vietnam)B. subtilis var. natto (107–108)9)
BidisubtilisBidiphar (Vietnam)B. cereus1 (106)9)
Bio-AciminViet-Duc Pharm. (Vietnam)B. cereus1 and other bacteria
Bio-KultProbiotics international Ltd. (UK)B. subtilis1 and other bacteria
BiobabyIldong Pharma (Korea)B. subtilis1 (3 × 106), C. butyricum 1 (107), B. coagulans 1 (5 × 107)
BiosubtylBiophar Company (Vietnam)B. cereus1 (106–107)
Biosubtyl DLIVAC (Vietnam)B. subtilis1 (107–108) and other bacteria
Biosubtyl I and IIBiophar Company (Vietnam)B. pumilus1 (106–107)
BiosporinBiofarm (Ukraine)/Garars (Russia)B. subtilis 2335 and B. licheniformis 2336 (ratio is 3:1)
BiovicerinGeyer Medicamentos S.A. (Brazil)B. cereus GM (106)
BispanBinex Co. (Korea)B. polyfermenticus SCD (1.7 × 107)
DomuvarBioProgress SpA (Italy)B. clausii1 (109)
EnterogerminaSanofi Winthrop SpA (Italy)B. clausii1 (106)
Flora-BalanceFlora-Balance (USA)B. laterosporus BOD (>106)
Flora3USAS. boulardii1 and B. coagulans 1
GanedenBC30USAB. coagulans GBI-30
Ildong BiovitaIldong Pharma (Korea)B. subtilis1 (3 × 106), C.butyricum 1 (107), L. sporogenes 1 (5 × 107)
Just ThriveUSAB. indicus HU36, B. coagulans 1, B. clausii 1, B. subtilis HU58
Lacbon, LacrisUni- Sankyo (Japan)B. coagulans1
Lactipan PlusIstituto Biochimico Italiano SpA (Italy)B. subtilis1 (2 × 109)
LactosporeSabinsa Corp. (USA)B. coagulans1 (109)
Latero-FloraGHC (USA)B. laterosporus BOD (>106)
LifeinULHC Lesaffre Human Care Ltd. (France)B. subtilis CU1
Medilac-VitaHanmi Pharmaceutical Co. (China)B. subtilis RO179 (108) and other bacteria
MegaSporeBioticUKB. indicus1, B. subtilis1, B. coagulans1
Nature’s First FoodNature’s First Law (California)B. laterosporus1, B. polymyxa1, B. subtilis1, B. pumilus1
NeolactofloreneNewpharma S.r.l. (Italy)B. coagulans1 and other bacteria
NutriCommitUSAB. subtilis1 and B. coagulans 1
PastylbioPasteur Institute (Vietnam)B. subtilis1 (108)
Primal DefenseGarden of Life (USA)B. subtilis1 (108) and other bacteria
SubtylMekophar (Vietnam)B. cereus vietnami (106–107)
SunnyGreen CleansingUSAB. coagulans1
SustenexGaneden Biotech Inc. (USA)B. coagulans Ganeden BC30
THORNEUSAB. coagulans1
1 No information available on strain name. * Spore/dose indicated where information is available.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Share and Cite

MDPI and ACS Style

Saggese, A.; Baccigalupi, L.; Ricca, E. Spore Formers as Beneficial Microbes for Humans and Animals. Appl. Microbiol. 2021, 1, 498-509. https://doi.org/10.3390/applmicrobiol1030032

AMA Style

Saggese A, Baccigalupi L, Ricca E. Spore Formers as Beneficial Microbes for Humans and Animals. Applied Microbiology. 2021; 1(3):498-509. https://doi.org/10.3390/applmicrobiol1030032

Chicago/Turabian Style

Saggese, Anella, Loredana Baccigalupi, and Ezio Ricca. 2021. "Spore Formers as Beneficial Microbes for Humans and Animals" Applied Microbiology 1, no. 3: 498-509. https://doi.org/10.3390/applmicrobiol1030032

APA Style

Saggese, A., Baccigalupi, L., & Ricca, E. (2021). Spore Formers as Beneficial Microbes for Humans and Animals. Applied Microbiology, 1(3), 498-509. https://doi.org/10.3390/applmicrobiol1030032

Article Metrics

Back to TopTop