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RAPD pattern of B. cytotoxicus strains using t markers (200 bp to 10 kb). D: donor strain E28.3 cytK−1-KO-A (buffer only); Lanes 1 to 5: candidate transconjug transconjugant containing the desired genetic loci (i.e., named E8.1 cytK−1-KO-B .
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The thermotolerant representative of the Bacillus cereus group, Bacillus cytotoxicus, reliably harbors the coding gene of cytotoxin K-1 (CytK-1). This protein is a highly cytotoxic variant of CytK toxin, initially recovered from a diarrheal foodborne outbreak that caused the death of three people. In recent years, the cytotoxicity of B. cytotoxicus...
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... kanamycin-resistant (Kan R ) candidate transconjugants were then verified by RAPD profiling [23] to confirm that they displayed the same pattern as their parental strain E8.1. As shown in Figure 2, the clone present in Lane 2 displayed the RAPD pattern identical to that of its parental strains E8.1. Additional PCR confirmed that this clone (named E8.1 cytK−1-KO-B ) had acquired the locus where cytK-1 was replaced by the Kan R gene (data not shown). ...
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... kanamycin-resistant (Kan R ) candidate transconjugants were then verified by RAPD profiling [23] to confirm that they displayed the same pattern as their parental strain E8.1. As shown in Figure 2, the clone present in Lane 2 displayed the RAPD pattern identical to that of its parental strains E8.1. Additional PCR confirmed that this clone (named E8.1 cytK−1-KO-B ) had acquired the locus where cytK-1 was replaced by the Kan R gene (data not shown). ...
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... trials to construct mutants of other highly cytotoxic B. cytotoxicus strains, such as CH_213 [18], should certainly be considered. Given that viable mutants of B. cytotoxicus obtained, it can be suggested that this gene is not worth mentioning that attempts to create a simi NVH 391-98 through electroporation, as well as v Figure 2. RAPD pattern of B. cytotoxicus strains using the OPA9 primer. ...
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Abstract Background microRNAs (miRNAs) are small noncoding RNAs that negatively regulate gene expression. They are found within cells and in body fluids. Extracellular miRNAs have been shown to associate with the surrounding tissues. Therefore, we predicted that miRNAs in tears may contribute to regulate corneal epithelial cell function. However, i...
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... strains by treating human intestinal cells (Caco-2 cell line) with bacterial culture supernatants. The results showed that a broad range of cytotoxicity values could be observed, in line with the literature [54][55][56][57]. Bacillus paranthracis (14SBCL566) and B. cytotoxicus (14SBCL15) induced the highest levels of cytotoxicity, while the non-cytotoxic strains included one "environmental" Bt, a B. weihenstephanensis (species assumed to be of low virulence [58]) and three non-Bc Bacillus spp., including B. amyloliquefaciens, classified as a "Qualified presumption of safety" organism by the European Food Safety Authority (EFSA) [59]. ...
Bacillus cereus (Bc) is a wide group of Gram-positive and spore-forming bacteria, known to be the etiological agents of various human infections, primarily food poisoning. The Bc group includes enteropathogenic strains able to germinate in the digestive tract and to produce enterotoxins such as Nhe, Hbl, and CytK. One species of the group, Bacillus thuringiensis (Bt), has the unique feature of producing insecticidal crystals during sporulation, making it an important alternative to chemical pesticides to protect crops from insect pest larvae. Nevertheless, several studies have suggested a link between the ingestion of pesticide strains and human cases of food poisoning, calling their safety into question. Consequently, reliable tools for virulence assessment are worth developing to aid decision making in pesticide regulation. Here, we propose complementary approaches based on two biological models, the human intestinal Caco-2 cell line and the insect Drosophila melanogaster, to assess and rank the enteric virulence potency of Bt strains in comparison with other Bc group members. Using a dataset of 48 Bacillus spp. strains, we showed that some Bc group strains, including Bt, were able to induce cytotoxicity in Caco-2 cells with concomitant release of IL-8 cytokine, a landmark of pro-inflammatory response. In the D. melanogaster model, we were able to sort a panel of 39 strains into four different classes of virulence, ranging from no virulence to strong virulence. Importantly, for the most virulent strains, mortality was associated with a loss of intestinal barrier integrity. Interestingly, although strains can share a common toxinotype, they display different degrees of virulence, suggesting the existence of specific mechanisms of virulence expression in vivo in the intestine.
... strains by treating human intestinal cells (Caco-2 cell line) with bacterial culture supernatants. The results showed that a broad range of cytotoxicity values could be observed, in line with the literature [54][55][56][57]. Bacillus paranthracis (14SBCL566) and B. cytotoxicus (14SBCL15) induced the highest levels of cytotoxicity, while the non-cytotoxic strains included one "environmental" Bt, a B. weihenstephanensis (species assumed to be of low virulence [58]) and three non-Bc Bacillus spp., including B. amyloliquefaciens, classified as a "Qualified presumption of safety" organism by the European Food Safety Authority (EFSA) [59]. ...
... They are often considered significant contributors to diarrhea, but their pathogenic mechanism in animal intestines remains unclear (22). Although they have been demonstrated to be cytotoxic in vitro, B. cereus carrying solely nonhemo lytic enterotoxin genes is incapable of causing diarrhea or histopathological changes in pigs (23,24). Therefore, cytotoxin K and nonhemolytic enterotoxins may not be the primary factors responsible for diarrhea, and further investigation is needed to assess their synergistic effects with other virulence factors (25). ...
Cellulolytic bacteria ferment dietary fiber into short-chain fatty acids, which play an important role in improving fiber utilization and maintaining intestinal health. Safe and effective cellulolytic bacteria are highly promising probiotic candidates. In this study, we isolated three strains of Bacillus cereus, which exhibited cellulolytic properties, from Kele pig feces. To assess the genetic basis of cellulose degradation by the isolates, whole-genome sequencing was used to detect functional genes associated with cellulose metabolism. Subsequently, we identified that the B. cereus CL2 strain was safe in mice by monitoring body weight changes, performing histopathologic evaluations, and determining routine blood indices. We next evaluated the biological characteristics of the CL2 strain in terms of its growth, tolerance, and antibiotic susceptibility, with a focus on its ability to produce short-chain fatty acids. Finally, the intestinal flora structure of the experimental animals was analyzed to assess the intestinal environment compatibility of the CL2 strain. In this study, we isolated a cellulolytic B. cereus CL2, which has multiple cellulolytic functional genes and favorable biological characteristics, from the feces of Kele pigs. Moreover, CL2 could produce a variety of short-chain fatty acids and does not significantly affect the diversity of the intestinal flora. In summary, the cellulolytic bacterium B. cereus CL2 is a promising strain for use as a commercial probiotic or in feed supplement.
IMPORTANCE
Short-chain fatty acids are crucial constituents of the intestinal tract, playing an important and beneficial role in preserving the functional integrity of the intestinal barrier and modulating both immune responses and the structure of the intestinal flora. In the intestine, short-chain fatty acids are mainly produced by bacterial fermentation of cellulose. Therefore, we believe that safe and efficient cellulolytic bacteria have the potential to be novel probiotics. In this study, we systematically evaluated the safety and biological characteristics of the cellulolytic bacterium B. cereus CL2 and provide evidence for its use as a probiotic.
... The protein encoded by this new reading frame has a high level of homology to LukDv, a poreforming toxin protein described in S. pneumoniae. This protein can form pores in the membranes of eukaryotic cells, unbalancing the partial pressure inside the host cells, which results in cell lysis, release of the nutrients into the environment, and ultimately in the death of these cells 15 . The novel protein is encoded by a gene with a high homology to cytK, which suggests a possibility of horizontal transfer of this gene among various gram-positive microorganisms that are in a physical contact. ...
Three stable microbial consortia, each composed of Bacillusparanthracis and Staphylococcushaemolyticus strains, were isolated from milk of cows diagnosed with mastitis in three geographically remote regions of Russia. The composition of these consortia remained stable following multiple passages on culture media. Apparently, this stability is due to the structure of the microbial biofilms formed by the communities. The virulence of the consortia depended on the B. paranthracis strains. It seems plausible that the ability of the consortia to cause mastitis in cattle was affected by mutations of the cytK gene of B. paranthracis.
... The protein encoded by this new reading frame has a high level of homology to LukDv, a pore-forming toxin protein described in S. pneumoniae. This protein can form pores in the membranes of eukaryotic cells, unbalancing the partial pressure inside the host cells, which results in cell lysis, release of the nutrients into the environment, and ultimately in the death of these cells [15]. The novel protein is encoded by a gene with a high homology to cytK, which suggests a possibility of horizontal transfer of this gene among various gram-positive microorganisms that are in a physical contact. ...
Bovine mastitis is one of the most economically important diseases in cattle. Infectious mastitis has a viral or bacterial origin. The most common bacterial agents are Staphylococcus , Streptococcus , coliforms, and Bacillus species. These are opportunistic microorganisms that can switch to a pathogenic form, using various virulence factors with follow various pathologies, both animal and human. Three stable microbial consortia, each composed of Bacillus paranthracis and Staphylococcus haemolyticus strains, were isolated from milk of cows diagnosed with mastitis in three geographically remote regions of Russia. The composition of these consortia remained stable following multiple passages on culture media. Apparently, this stability is due to the structure of the microbial biofilms formed by the communities. The virulence of the consortia depended on the B. paranthracis strains. It seems plausible that the ability of the consortia to cause mastitis in cattle was affected by mutations of the cytK gene of B. paranthracis .
... Seuls quelques isolats de cette espèce ont été caractérisés en ce qui concerne leurs effets cytotoxiques. Cependant, le rôle de la cytotoxine K-1 en tant qu'agent causal d'infections alimentaires reste méconnu (Guinebretière et al., 2013 ;Koné et al., 2021 (Cairo et al., 2021). ...
Bacillus thuringiensis (Bt) est une bactérie entomopathogène appartenant au groupe Firmicutes sporulants Bacillus cereus sensu lato. Ce groupe comprend entre autres Bacillus cereus sensu stricto, responsable de toxi-infections alimentaires ou encore Bacillus anthracis, responsable de la maladie du charbon.Bt est capable de réaliser un cycle d’infection complet chez son hôte, ici la larve de l’insecte Galleria mellonella, fausse teigne de la cire. Bt tue son hôte, survit dans son cadavre et sporule, en activant séquentiellement des gènes de virulence, de nécrotrophisme et de sporulation (processus aboutissant à la formation de spores, formes cellulaires déshydratées dormantes et résistantes aux conditions défavorables). Néanmoins, la population bactérienne isolée des cadavres d’insectes est hétérogène et des cellules non-sporulantes, capables de survivre dans un état physiologique inconnu, ont été identifiées.Ces travaux présentent la caractérisation de ces bactéries à l'aide de méthodes de génétique moléculaire et d'imagerie cellulaire ainsi que d'une analyse transcriptomique par RNA-Seq.Les résultats démontrent que ces cellules sont transitoirement nécrotrophes, puis qu’elles subissent un ralentissement métabolique tout en restant viables et particulièrement adaptées à l'environnement du cadavre d'insecte jusqu’à au moins 14 jours post-infection. Par ailleurs, le RNA-Seq révèle que de nombreux gènes exprimés lors de la survie de Bt dans le cadavre de son hôte, sont impliqués dans l’homéostasie du fer ou encore dans la réponse au stress oxydatif. Ces derniers sont plus particulièrement exprimés au cours de la phase tardive de survie dans le cadavre de l’insecte.Cette étude est la première caractérisant l'état physiologique à long terme de Bt lors d'une infection. Elle met en évidence l'existence d'une voie de persistance alternative à la sporulation et montre l’adaptation de ces bactéries au cadavre de l’insecte, un environnement particulier et méconnu.
Horizontal gene transfer (HGT) is a major driving force in shaping bacterial communities. Key elements responsible for HGT are conjugation-like events and transmissible plasmids. Conjugative plasmids can promote their own transfer as well as that of co-resident plasmids. Bacillus cereus and relatives harbor a plethora of plasmids, including conjugative plasmids, which are at the heart of the group species differentiation and specification. Since the first report of a conjugation-like event between strains of B. cereus sensu lato (s.l.) 40 years ago, many have studied the potential of plasmid transfer across the group, especially for plasmids encoding major toxins. Over the years, more than 20 plasmids from B. cereus isolates have been reported as conjugative. However, with the increasing number of genomic data available, in silico analyses indicate that more plasmids from B. cereus s.l. genomes present self-transfer potential. B. cereus s.l. bacteria occupy diverse environmental niches, which were mimicked in laboratory conditions to study conjugation-related mechanisms. Laboratory mating conditions remain nonetheless simplistic compared to the complex interactions occurring in natural environments. Given the health, economic and ecological importance of strains of B. cereus s.l., it is of prime importance to consider the impact of conjugation within this bacterial group.
pXO16, the 350 kb-conjugative plasmid from Bacillus thuringiensis sv. israelensis promotes its own transfer at high efficiency, triggers the transfer of mobilizable and non-mobilizable plasmids, as well as the transfer of host chromosomal loci. Naturally found in B. thuringiensis sv. israelensis, pXO16 transfers to various strains of Bacillus cereus sensu lato (s.l.) at a wide range of frequencies. Despite this host diversity, a paradox remains between the relatively large host spectrum and the natural occurrence of pXO16, so far restricted to B. thuringiensis sv. israelensis. Proposing first insights exploring this paradox, we investigated the behaviour of pXO16 amongst different members of the B. cereus group. We first looked at the transfer of pXO16 to two new host clusters of B. cereus s.l., Bacillus mycoides and Bacillus anthracis clusters. This examination brought to light the impairment of the characteristic rhizoidal phenotype of B. mycoides in presence of pXO16. We also explored the stability of pXO16 at different temperatures as some B. cereus group members are well-known for their psychro- or thermo-tolerance. This shed light on the thermo-sensitivity of the plasmid. The influence of pXO16 on its host cell growth and on swimming capacity also revealed no or limited impact on its natural host B. thuringiensis sv. israelensis. On the contrary, pXO16 affected more strongly both the growth and swimming capacity of other B. cereus s.l. hosts. This reinforced the running hypothesis of a co-evolution between pXO16 and B. thuringiensis sv. israelensis, enabling the plasmid maintenance without impairing the host strain development.
