Structural insights into thermostable direct hemolysin of Vibrio parahaemolyticus using single-particle cryo-EM

Thermostable direct hemolysin (TDH) is a ~19 kDa, hemolytic pore-forming toxin from the gram-negative marine bacterium Vibrio parahaemolyticus, one of the causative agents of seafood-borne acute gastroenteritis and septicemia. Previous studies have established that TDH exists as a tetrameric assembly in physiological state; however, there is limited knowledge regarding the molecular arrangement of its disordered N-terminal region (NTR)—the absence of which has been shown to compromise TDH's hemolytic and cytotoxic abilities. In our current study, we have employed single-particle cryo-electron microscopy to resolve the solution-state structures of wild-type TDH and a TDH construct with deletion of the NTR (NTD), in order to investigate structural aspects of NTR on the overall tetrameric architecture. We observed that both TDH and NTD electron density maps, resolved at global resolutions of 4.5 and 4.2 Å, respectively, showed good correlation in their respective oligomeric architecture. Additionally, we were able to locate extra densities near the pore opening of TDH which might correspond to the disordered NTR. Surprisingly, under cryogenic conditions, we were also able to observe novel supramolecular assemblies of TDH tetramers, which we were able to resolve to 4.3 Å. We further investigated the tetrameric and inter-tetrameric interaction interfaces to elaborate upon the key residues involved in both TDH tetramers and TDH super assemblies. Our current structural study will aid in understanding the mechanistic aspects of this pore-forming toxin and the role of its disordered NTR in membrane interaction.     

Phenotypic and biochemical alterations in relation to <Emphasis Type="Italic">MT2</Emphasis> gene expression in <Emphasis Type="Italic">Plantago ovata</Emphasis> Forsk under zinc stress

Plantago ovata Forsk is an annual herb with immense medicinal importance, the seed and husk of which is used in the treatment of chronic constipation, irritable bowel syndrome, diarrhea since ancient times. Zinc, an essential metal, is required by plants as they form important components of zinc finger proteins and also aid in synthesis of photosynthetic pigments such as chlorophyll. However, in excess amount Zn causes chlorosis of leaf and shoot tissues and generate reactive oxygen species. The present study is aimed at investigating the changes in expression levels of MT2 gene in Plantago ovata under zinc stress. Data show up to 1.66 fold increase in expression of PoMT2 in 1000 µM ZnSO₄·7H₂O treated sample. Our study also describes alteration of MT2 gene expressions in Plantago ovata as observed through Real time PCR (qPCR) done by \(2^{{ - \Delta \Delta}} C_T\) method. In this study we have observed an upregulation (or induction) in the PoMT2 gene expression level in 500 and 800 µM ZnSO₄·7H₂O treated samples but found saturation on further increasing the dose to 1000 µM of ZnSO₄·7H₂O. Determination of the phenotypic and biochemical changes in Plantago ovata due to exposure to zinc stress of concentrations 500, 800 and 1000 µM revealed oxidative stress. The enhanced expression of MT2 gene in Plantago ovata has a correlation with the increased total antioxidant activity and increased DPPH radical scavenging activity.     

Geographical spread and structural basis of sulfadoxine-pyrimethamine drug-resistant malaria parasites

The global spread of sulfadoxine (Sdx, S) and pyrimethamine (Pyr, P) resistance is attributed to increasing number of mutations in DHPS and DHFR enzymes encoded by malaria parasites. The association between drug resistance mutations and SP efficacy is complex. Here we provide an overview of the geographical spread of SP resistance mutations in Plasmodium falciparum (Pf) and Plasmodium vivax (Pv) encoded dhps and dhfr genes. In addition, we have collated the mutation data and mapped it on to the three-dimensional structures of DHPS and DHFR which have become available. Data from genomic databases and 286 studies were collated to provide a comprehensive landscape of mutational data from 2005 to 2019. Our analyses show that the Pyr-resistant double mutations are widespread in Pf/PvDHFR (P. falciparum ～61% in Asia and the Middle East, and in the Indian sub-continent; in P. vivax ～33% globally) with triple mutations prevailing in Africa (～66%) and South America (～33%). For PfDHPS, triple mutations dominate South America (～44%), Asia and the Middle East (～34%) and the Indian sub-continent (～27%), while single mutations are widespread in Africa (～45%). Contrary to the status for P. falciparum, Sdx-resistant single point mutations in PvDHPS dominate globally. Alarmingly, highly resistant quintuple and sextuple mutations are rising in Africa (PfDHFR-DHPS) and Asia (Pf/PvDHFR-DHPS). Structural analyses of DHFR and DHPS proteins in complexes with substrates/drugs have revealed that resistance mutations map proximal to Sdx and Pyr binding sites. Thus new studies can focus on discovery of novel inhibitors that target the non-substrate binding grooves in these two validated malaria parasite drug targets.     

Perspectives and challenges of micro/nanoplastics‐induced toxicity with special reference to phytotoxicity

Plastic pollution has become a global concern for ecosystem health and biodiversity conservation. Concentrations of plastics are manifold higher in the terrestrial system than the aquatic one. Micro/nanoplastics (M/NP) have the ability to alter soil enzymatic system, soil properties and also affect soil borne microorganisms and earthworms. Despite, the knowhow regarding modulatory effects of plastics are acquired from the study on aquatic system and reports on their phytotoxic potentials are limited. The presence of cell wall that could restrict M/NP invasion into plant roots might be the putative cause of this limitation. M/NP inhibit plant growth, seed germination and gene expression; and they also induce cytogenotoxicity by aggravating reactive oxygen species generation. Dynamic behavior of cell wall; the pores formed either by cell wall degrading enzymes or by plant–pathogen interactions or by mechanical injury might facilitate the entry of into roots M/NP. This review also provides a possible mechanism of large sized microplastics‐induced phytotoxicity especially for those that cannot pass through cell wall pores. As M/NP affect soil microbial community and soil parameters, it is hypothesized that they could have the potential to affect N₂ fixation and research should be conducted in this direction. Reports on M/NP‐induced toxicity mainly focused only on one polymer type (polystyrene) in spite of the toxicological relevancies of other polymer types like polyethylene, polypropylene etc. So, the assessment of phytotoxic potential of M/NP should be done using other plastic polymers in real environment as they are known to intract with other environmental stressors as well as can alter the the soil–microbe–plant interaction.     

Functional interplay between plastic polymers and microbes: a comprehensive review

Biodegradation - Escalated production of plastic, their worldwide distribution and persistent nature finally results into their environmental accumulation causing severe threats to the ecological...     

The effect of gradual increase in salinity on the biomass productivity and biochemical composition of several marine,halotolerant<Emphasis Type="Bold">,</Emphasis> and halophilic microalgae

Open ponds are the preferred cultivation system for large-scale microalgal biomass production. To be more sustainable, commercial scale biomass production should rely on seawater, as freshwater is a limiting resource, especially in places with high irradiance. If seawater is used for both pond fill and evaporative volume makeup, salinity of the growth media will rise over time. It is not possible for any species to achieve optimum growth over the whole saline spectrum (from seawater salinity level up to salt saturation state). In this study, we investigated the effects of gradual salinity increase (between 35 and 233 ppt) on biomass productivity and biochemical composition (lipid and carbohydrate) of six marine, two halotolerant, and a halophilic microalgae. A gradual and slow stepped salinity increase was found to expand the salinity tolerance range of tested species. A gradual reduction in biomass productivity and maximum photochemical efficiency was observed as a consequence of increased salinity in all tested species. Among the marine microalgae, Tetraselmis showed highest biomass productivity (32 mg L^?1 day^?1) with widest salinity tolerance range (35 to 109 ppt). Halotolerant Amphora and Navicula were able to grow from 35 ppt to 129 ppt salinity. Halophilic Dunaliella was the only species capable of growing between 35 and 233 ppt and showed highest lipid content (56.2%) among all tested species. This study showed that it should be possible to maintain high biomass in open outdoor cultivation utilizing seawater by growing Tetraselmis, Amphora, and Dunaliella one after another as salinity increases in the cultivation system.     

Generation of a highly diverse panel of antagonistic chicken monoclonal antibodies against the GIP receptor

Raising functional antibodies against G protein-coupled receptors (GPCRs) is challenging due to their low density expression, instability in the absence of the cell membrane's lipid bilayer and frequently short extracellular domains that can serve as antigens. In addition, a particular therapeutic concept may require an antibody to not just bind the receptor, but also act as a functional receptor agonist or antagonist. Antagonizing the glucose-dependent insulinotropic polypeptide (GIP) receptor may open up new therapeutic modalities in the treatment of diabetes and obesity. As such, a panel of monoclonal antagonistic antibodies would be a useful tool for in vitro and in vivo proof of concept studies. The receptor is highly conserved between rodents and humans, which has contributed to previous mouse and rat immunization campaigns generating very few usable antibodies. Switching the immunization host to chicken, which is phylogenetically distant from mammals, enabled the generation of a large and diverse panel of monoclonal antibodies containing 172 unique sequences. Three-quarters of all chicken-derived antibodies were functional antagonists, exhibited high-affinities to the receptor extracellular domain and sampled a broad epitope repertoire. For difficult targets, including GPCRs such as GIPR, chickens are emerging as valuable immunization hosts for therapeutic antibody discovery.