Deactivation of Ascaris suum eggs using electroporation and sequential inactivation with chemical disinfection

Electroporation has been evaluated as a potential backend wastewater treatment for deactivation of Ascaris suum eggs in buffer solution. Initial results indicate that eggshell permeability is affected by the pulse train electric field strength and duration. Coupling electroporation with chemical exposure, using low concentrations of commercially available disinfectants, allows oxidizing agents to pass through the complex strata of the A. suum eggshell, specifically reaching the innermost embryonic environment, which leads to successful deactivation compared to either method used separately. The aim of this work is to identify and develop an alternative technique that efficiently inactivates helminth eggs present in wastewater.

CSIG-28. ROLE OF ALTERNATIVE SPLICING IN CONFERRING SPATIAL HETEROGENEITY SIGNATURES IN GBM

Therapeutic interventions induce cell stress to trigger apoptosis in tumor cells. Specific cellular stressors, including those initiated by radio-chemotherapy, activate distinct pro-death proteins through a multitude of mechanisms. Apoptosis consists of highly regulated pathways involving post-translational modifications and cleavage of proteins leading to sequential inactivation of the main cellular processes e.g. cell cycle progression. Recently, we demonstrated that apoptotic bodies/ApoEVs in GBM transfer spliceosomal machinery components to neighboring cells, which are subsequently internalized and confer survival advantage through changes in mRNA splicing. Following the validation of the role of RBM11, a spliceosomal component in the said intercellular communication, we extended our findings in the characterization of spatially distinct clonally derived sphere lines from tumor edge and core-located tissues from GBM patients. We analyzed the expression of spliceosomal genes in edge and core-derived clones and found spatially distinct splicing capabilities and that the genes that were affected by alternative splicing events in edge and core cells have different splicing of genes that are involved in ubiquitination and regulate cell cycle progression. Downstream analyses revealed that the most promising target is MDM2. Importantly, this gene regulates cell cycle via ubiquitination. Further, western blot revealed differential expression of both RBM11 and MDM2 in egde and core-derived clones.Collectively, the data so far provides motivation for the possibility of finding cellular vulnerabilities in the splicing machinery and thus actionable targets for intervention that accounts for the bearings of spatial heterogeneity.

FadD19 of Rhodococcus rhodochrous DSM43269, a Steroid-Coenzyme A Ligase Essential for Degradation of C-24 Branched Sterol Side Chains

ABSTRACTThe actinobacterial cholesterol catabolic gene cluster contains a subset of genes that encode β-oxidation enzymes with a putative role in sterol side chain degradation. We investigated the physiological roles of several genes, i.e.,fadD17,fadD19,fadE26,fadE27, andro04690DSM43269, by gene inactivation studies in mutant strain RG32 ofRhodococcus rhodochrousDSM43269. Mutant strain RG32 is devoid of 3-ketosteroid 9α-hydroxylase (KSH) activity and was constructed following the identification, cloning, and sequential inactivation of fivekshAgene homologs in strain DSM43269. We show that mutant strain RG32 is fully blocked in steroid ring degradation but capable of selective sterol side chain degradation. Except for RG32ΔfadD19, none of the mutants constructed in RG32 revealed an aberrant phenotype on sterol side chain degradation compared to parent strain RG32. Deletion offadD19in strain RG32 completely blocked side chain degradation of C-24 branched sterols but interestingly not that of cholesterol. The additional inactivation offadD17in mutant RG32ΔfadD19also did not affect cholesterol side chain degradation. Heterologously expressed FadD19DSM43269nevertheless was active toward steroid-C26-oic acid substrates. Our data identified FadD19 as a steroid-coenzyme A (CoA) ligase with an essentialin vivorole in the degradation of the side chains of C-24 branched-chain sterols. This paper reports the identification and characterization of a CoA ligase with anin vivorole in sterol side chain degradation. The high similarity (67%) between the FadD19DSM43269and FadD19H37Rvenzymes further suggests that FadD19H37Rvhas anin vivorole in sterol metabolism ofMycobacterium tuberculosisH37Rv.