The complexities of inferring symbiont function: Paraburkholderia symbiont dynamics in social amoeba populations and its impact on the amoeba microbiome

The relationship between the social amoeba Dictyostelium discoideum and its endosymbiotic bacteria Paraburkholderia provides a model system for studying the development of symbiotic relationships. Laboratory experiments have shown that any of three species of Paraburkholderia symbiont allow D. discoideum food bacteria to persist through the amoeba lifecycle and survive in amoeba spores, rather than being fully digested. This phenomenon is termed "farming", as it potentially allows spores dispersed to food poor locations to grow their own. The occurrence and impact of farming in natural populations, however, has been a challenge to measure. Here, we surveyed natural D. discoideum populations and found that only one of the three symbiont species, P. agricolaris, remained prevalent. We then explored the effect of Paraburkholdia on the amoeba microbiome, expecting that by facilitating bacterial food carriage it would diversify the microbiome. Contrary to our expectations, Paraburkholderia tended to infectiously dominate the D. discoideum microbiome, in some cases decreasing diversity. Similarly, we found little evidence for Paraburkholderia facilitating the carriage of particular food bacteria. These findings change our understanding of farming and suggest the possibility that Paraburkholderia could be playing multiple roles for its host, as inferred metagenomic analysis indicates a potential role of P. agricolaris in toxin degradation.

Toxin Degradation by Rumen Microorganisms: A Review

Animal feeds may contain exogenous compounds that can induce toxicity when ruminants ingest them. These toxins are secondary metabolites originating from various sources including plants, bacteria, algae and fungi. Animal feed toxins are responsible for various animal poisonings which negatively impact the livestock industry. Poisoning is more frequently reported in newly exposed, naïve ruminants while ‘experienced’ ruminants are observed to better tolerate toxin-contaminated feed. Ruminants can possess detoxification ability through rumen microorganisms with the rumen microbiome able to adapt to utilise toxic secondary metabolites. The ability of rumen microorganisms to metabolise these toxins has been used as a basis for the development of preventative probiotics to confer resistance against the poisoning to naïve ruminants. In this review, detoxification of various toxins, which include plant toxins, cyanobacteria toxins and plant-associated fungal mycotoxins, by rumen microorganisms is discussed. The review will include clinical studies of the animal poisoning caused by these toxins, the toxin mechanism of action, toxin degradation by rumen microorganisms, reported and hypothesised detoxification mechanisms and identified toxin metabolites with their toxicity compared to their parent toxin. This review highlights the commercial potential of rumen inoculum derived probiotics as viable means of improving ruminant health and production.

Microcystis aeruginosa inactivation and microcystin-LR degradation by the photo-Fenton process at the initial near-neutral pH

This article presents the first report of the photo-Fenton process application, at the initial near-neutral pH, aiming at Microcystis aeruginosa inactivation and toxin degradation.

CesH Represses Cereulide Synthesis as an Alpha/Beta Fold Hydrolase in Bacillus cereus

Cereulide is notorious as a heat-stable emetic toxin produced by Bacillus cereus and glucose is supposed to be an ingredient supporting its formation. This study showed that glucose addition benefited on cell growth and the early transcription of genes involved in substrate accumulation and toxin synthesis, but it played a negative role in the final production of cereulide. Meanwhile, a lasting enhancement of cesH transcription was observed with the addition of glucose. Moreover, the cereulide production in ΔcesH was obviously higher than that in the wild type. This indicates that CesH has a repression effect on cereulide production. Bioinformatics analysis revealed that CesH was an alpha/beta hydrolase that probably associated with the cell membrane, which was verified by subcellular localization. The esterase activity against para-nitrophenyl acetate (PNPC2) of the recombinant CesH was confirmed. Although no sign of ester bond cleavage in cereulide or valinomycin was demonstrated in in vitro assays, CesH could reverse the cereulide analogue sensitivity of Bacillus subtilis in vivo, by which toxin degradation was facilitated. Moreover, site directed mutations identified that the conserved catalytic triad of CesH might consist of Serine 86, Glutamate 199, and Histidine 227. These results help us to understand the regulation of cereulide production and provide clues for developing control measurements.

Intracellular Degradation ofHelicobacter pyloriVacA Toxin as a Determinant of Gastric Epithelial Cell Viability

ABSTRACTHelicobacter pyloriVacA is a secreted pore-forming toxin that induces cell vacuolationin vitroand contributes to the pathogenesis of gastric cancer and peptic ulcer disease. We observed that purified VacA has relatively little effect on the viability of AGS gastric epithelial cells, but the presence of exogenous weak bases such as ammonium chloride (NH4Cl) enhances the susceptibility of these cells to VacA-induced vacuolation and cell death. Therefore, we tested the hypothesis that NH4Cl augments VacA toxicity by altering the intracellular trafficking of VacA or inhibiting intracellular VacA degradation. We observed VacA colocalization with LAMP1- and LC3-positive vesicles in both the presence and absence of NH4Cl, indicating that NH4Cl does not alter VacA trafficking to lysosomes or autophagosomes. Conversely, we found that supplemental NH4Cl significantly increases the intracellular stability of VacA. By conducting experiments using chemical inhibitors, stable ATG5 knockdown cell lines, and ATG16L1 knockout cells (generated using CRISPR/Cas9), we show that VacA degradation is independent of autophagy and proteasome activity but dependent on lysosomal acidification. We conclude that weak bases like ammonia, potentially generated duringH. pyloriinfection by urease and other enzymes, enhance VacA toxicity by inhibiting toxin degradation.

Effect of PR toxin on THP1 and Caco-2 cells: an in vitro study

Penicillium roqueforti produces mycotoxins including PR toxin, which is a food and feed contaminant. In this study, PR toxin was purified from culture material of the Penicillium roqueforti F43-1 strain. Toxic effects were evaluated in undifferentiated human Caco-2 intestinal epithelial cells and THP-1 monocytic immune cells. To understand the mechanisms involved in PR-toxin toxicity, cell death and pro-inflammatory gene expression were studied. In addition, PR toxin degradation was assessed. Cytotoxicity studies showed a dose-dependent effect of PR toxin and the calculated mean cytotoxic concentration (IC50) concentrations were for Caco-2 and THP-1 cells >12.5 and 0.83 μM, respectively. Gene expression studies showed that tumour necrosis factor-α expression was significantly increased after 24 h exposure to 312 μM PR toxin. PR toxin induced necrosis on THP-1 cells after 3 h exposure. In the cell culture system, the PR toxin showed a 10-fold reduction in PR toxin concentration within 48 h, indicating that PR toxin was degraded by THP-1. To conclude, PR toxin appears to be one of the most cytotoxic P. roqueforti mycotoxins on Caco-2 and/or THP-1 cells and induces in THP-1 cells both necrosis and an inflammatory response.