Get to Know Your Neighbors: Characterization of Close Bacillus anthracis Isolates and Toxin Profile Diversity in the Bacillus cereus Group.

Unexpected atypical isolates of Bacillus cereus s.l. occasionally challenge conventional microbiology and even the most advanced techniques for anthrax detection. For anticipating and gaining trust, 65 isolates of Bacillus cereus s.l. of diverse origin were sequenced and characterized. The BTyper3 tool was used for assignation to genomospecies B. mosaicus (34), B. cereus s.s (29) and B. toyonensis (2), as well as virulence factors and toxin profiling. None of them carried any capsule or anthrax-toxin genes. All harbored the non-hemolytic toxin nheABC and sphygomyelinase spH genes, whereas 41 (63%), 30 (46%), 11 (17%) and 6 (9%) isolates harbored cytK-2, hblABCD, cesABCD and at least one insecticidal toxin gene, respectively. Matrix-assisted laser desorption ionization-time of flight mass spectrometry confirmed the production of cereulide (ces genes). Phylogeny inferred from single-nucleotide polymorphisms positioned isolates relative to the B. anthracis lineage. One isolate (BC38B) was of particular interest as it appeared to be the closest B. anthracis neighbor described so far. It harbored a large plasmid similar to other previously described B. cereus s.l. megaplasmids and at a lower extent to pXO1. Whereas bacterial collection is enriched, these high-quality public genetic data offer additional knowledge for better risk assessment using future NGS-based technologies of detection.

Hydrogen Peroxide Vapor Decontamination of Hazard Group 3 Bacteria and Viruses in a Biosafety Level 3 Laboratory.

Aim: This study aimed to validate the efficacy of hydrogen peroxide vapor (HPV) decontamination technology set up in a biosafety level 3 (BSL-3) laboratory on surrogates and hazard group 3 (HG3) agents. Methods and Results: The HPV decontamination system (Bioquell) was assessed with both qualitative and quantitative methods on (1) spore surrogates (Geobacillus stearothermophilus, Bacillus atrophaeus, and Bacillus thuringiensis) in the BSL-3 laboratory and in the material airlock and on (2) HG3 agents (Bacillus anthracis; SARS-CoV-2, Venezuelan equine encephalitis virus [VEE], and Vaccinia virus) in the BSL-3 laboratory. Other HG3 bacteria likely to be handled in the BSL-3 laboratory (Yersinia pestis, Burkholderia mallei, Brucella melitensis, and Francisella tularensis) were excluded from the HPV decontamination assays as preliminary viability tests demonstrated the total inactivation of these agents after 48 h drying on different materials. Conclusions: The efficacy of HPV decontamination was validated with a reduction in viability of 5-7 log10 for the spores (surrogates and B. anthracis), and for the enveloped RNA viruses. Vaccinia showed a higher resistance to the decontamination process, being dependent on the biological indicator location in the BSL-3 laboratory.

Haslea silbo, A Novel Cosmopolitan Species of Blue Diatoms.

Specimens of a new species of blue diatoms from the genus Haslea Simonsen were discovered in geographically distant sampling sites, first in the Canary Archipelago, then North Carolina, Gulf of Naples, the Croatian South Adriatic Sea, and Turkish coast of the Eastern Mediterranean Sea. An exhaustive characterization of these specimens, using a combined morphological and genomic approach led to the conclusion that they belong to a single new to science cosmopolitan species, Haslea silbo sp. nov. A preliminary characterization of its blue pigment shows similarities to marennine produced by Haslea ostrearia, as evidenced by UV-visible spectrophotometry and Raman spectrometry. Life cycle stages including auxosporulation were also observed, providing data on the cardinal points of this species. For the two most geographically distant populations (North Carolina and East Mediterranean), complete mitochondrial and plastid genomes were sequenced. The mitogenomes of both strains share a rare atp6 pseudogene, but the number, nature, and positions of the group II introns inside its cox1 gene differ between the two populations. There are also two pairs of genes fused in single ORFs. The plastid genomes are characterized by large regions of recombination with plasmid DNA, which are in both cases located between the ycf35 and psbA genes, but whose content differs between the strains. The two sequenced strains hosts three plasmids coding for putative serine recombinase protein whose sequences are compared, and four out of six of these plasmids were highly conserved.

Complex Relationships between the Blue Pigment Marennine and Marine Bacteria of the Genus Vibrio.

Marennine, the water-soluble blue pigment produced by the marine diatom Haslea ostrearia, is known to display antibacterial activities. Previous studies have demonstrated a prophylactic effect of marennine on bivalve larvae challenged with a pathogenic Vibrio splendidus, suggesting that the blue Haslea is a good candidate for applications in aquaculture as a source of a natural antimicrobial agent. Indeed, the genus Vibrio is ubiquitous in aquaculture ecosystems, and regular events of pathogenic invasion cause some of the biggest losses worldwide. To better characterize the effects of marennine on Vibrios, a panel of 30 Vibrio strains belonging to 10 different species was tested, including bivalve pathogenic species (e.g., Vibrio crassostreae and Vibrio harveyi). Vibrio strains were first exposed to 10 and 25 µg mL-1 of Blue Water (BW), a concentrated culture supernatant of H. ostrearia containing marennine. This screening evidenced a great diversity in responses, from growth stimulation to a total inhibition, at both the interspecific or intraspecific level. In a second series of experiments, 10 Vibrio strains were exposed to BW at concentrations ranging from 5 to 80 µg mL-1. The highest concentrations of BW did not systematically result in the highest growth inhibition as hormetic responses-opposite effects regarding the concentration-were occasionally evidenced. The relationships between marennine and Vibrio strains appear more complex than expected and justify further study-in particular, on the mechanisms of action-before considering applications as a natural prophylactic or antibiotic agent in aquaculture.

Harmful or harmless: Biological effects of marennine on marine organisms.

Marennine is a water-soluble blue-green pigment produced by the marine diatom Haslea ostrearia. The diatom and its pigment are well known from oyster farming areas as the source of the greening of oyster gills, a natural process increasing their market value in Western France. Blooms of blue Haslea are also present outside oyster ponds and hence marine organisms can be exposed, periodically and locally, to significant amounts of marennine in natural environments. Due to its demonstrated antibacterial activities against marine pathogenic bacteria (e.g. Vibrio) and possible prophylactic effects toward bivalve larvae, marennine is of special interest for the aquaculture industry, especially bivalve hatcheries. The present study aimed to provide new insights into the effects of marennine on a large spectrum of marine organisms belonging to different phyla, including species of aquaculture interest and organisms frequently employed in standardised ecotoxicological assays. Different active solutions containing marennine were tested: partially purified Extracellular Marennine (EMn), and concentrated solutions of marennine present in H. ostrearia culture supernatant; the Blue Water (BW) and a new process called Concentrated Supernatant (CS). Biological effects were meanwhile demonstrated in invertebrate species for the three marennine-based solutions at the highest concentrations tested (e.g., decrease of fertilization success, delay of embryonic developmental stages or larval mortality). Exposure to low concentrations did not impact larval survival or development and even tended to enhance larval physiological state. Furthermore, no effects of marennine were observed on the fish gill cell line tested. Marennine could be viewed as a Jekyll and Hyde molecule, which possibly affects the earliest stages of development of some organisms but with no direct impacts on adults. Our results emphasize the need to determine dosages that optimize beneficial effects and critical concentrations not to be exceeded before considering the use of marennine in bivalve or fish hatcheries.

Antimicrobial Compounds from Eukaryotic Microalgae against Human Pathogens and Diseases in Aquaculture.

The search for novel compounds of marine origin has increased in the last decades for their application in various areas such as pharmaceutical, human or animal nutrition, cosmetics or bioenergy. In this context of blue technology development, microalgae are of particular interest due to their immense biodiversity and their relatively simple growth needs. In this review, we discuss about the promising use of microalgae and microalgal compounds as sources of natural antibiotics against human pathogens but also about their potential to limit microbial infections in aquaculture. An alternative to conventional antibiotics is needed as the microbial resistance to these drugs is increasing in humans and animals. Furthermore, using natural antibiotics for livestock could meet the consumer demand to avoid chemicals in food, would support a sustainable aquaculture and present the advantage of being environmentally friendly. Using natural and renewable microalgal compounds is still in its early days, but considering the important research development and rapid improvement in culture, extraction and purification processes, the valorization of microalgae will surely extend in the future.

Pretreatment of the cockroach cercal afferent/giant interneuron synapses with nicotinoids and neonicotinoids differently affects acetylcholine and nicotine-induced ganglionic depolarizations.

We have recently demonstrated that neonicotinoid insecticides were able to act as agonists of postsynaptic nicotinic acetylcholine receptors (nAChRs) expressed at the synapse between the cercal nerve XI and the giant interneurons, in the sixth abdominal ganglion. In this work, we demonstrated that nicotinoids such as nornicotine acted as an agonist of nicotinic acetylcholine receptors expressed at cercal afferent/giant interneurons while cotinine was a poor agonist. Indeed, nornicotine induced a ganglionic depolarization which was blocked by the nicotinic antagonist mecamylamine. In addition, we found that pretreatment of the sixth abdominal ganglion with 1 and 10 µM nornicotine and cotinine had no significant effect on acetylcholine and nicotine-induced depolarization. But pretreatment with 1 and 10 µM acetamiprid and imidacloprid had a strong effect. 1 and 10 µM acetamiprid completely blocked acetylcholine-induced depolarization, whereas imidacloprid had a partial effect. The present work therefore suggests, in agreement with previous studies, that nornicotine and cotinine bind to distinct cockroach postsynaptic nAChRs, whereas acetamiprid and imidacloprid have competitive effects with acetylcholine and nicotine on ganglionic depolarization.