Occupational safety assessment of biogenic urea nanofertilisers using in vitro pulmonary, and in vivo ocular models.

Nanomaterials (NMs) are now gaining popularity to be used in agriculture as fertilisers to reduce the dose of conventional fertilisers and enhance nutrient use efficiency. Urea has found its application as a conventional nitrogenous fertiliser since long, however, the nutrient use efficiency of the bulk form of urea is low due to issues related to ammonia volatilisation. This study proposes a biogenic synthesis route to develop urea nanoparticles that can be used as nano-fertiliser for better uptake and hence improved nutrient efficiency. Large scale production and widespread application of these nano-fertilisers to the agricultural fields will enhance the direct exposure to workers and farmers. Therefore, the occupational safety evaluation becomes critical. In this study, we report a new method for synthesis of urea nanoparticles (TNU, absolute size: 12.14 ± 7.79 nm) followed by nano-safety evaluation. Herein, the pulmonary and ocular compatibilities of TNU were investigated in vitro and in vivo respectively. The assay for cellular mitochondrial activity was carried out on human lung fibroblasts (WI-38) under varied TNU exposure concentrations up to 72 h. The acute biocompatibility effect, ocular irritation and sub-lethal effects were measured on New Zealand Rabbit. The results show that TNU do not exhibit any cytotoxicity and detrimental cell mitochondrial activity up to the highest tested concentration of 1000 µg/mL and 72 h of testing. The animal experiment results also show that neither acute nor sub-lethal toxic effects can be detected after TNU ocular instillation up to 21 days when tested up to environmentally relevant concentration of 15 µg/mL. These results suggest the occupational safety of biogenic urea nanoparticles and support its application as nanofertiliser.

Fertilizing benefits of biogenic phosphorous nanonutrients on Solanum lycopersicum in soils with variable pH.

Nanoformulations of Phosphorous (P) have recently been proposed as alternatives to P fertilizers. In this study, the fertilizing efficacies of P-based nanomaterials (NMs), nanohydroxyapatite (nHAP) and nanophosphorus (nP), were examined on Solanum lycopersicum (Pusa Rohini, Indian tomato) in growth room pot experiments. These NMs differed in their mode of synthesis, chemical composition, size and shape. Rock-phosphate (RP), phosphoric acid (PA) and di-ammonium phosphate (DAP) were included as bulk materials for comparison. Three varieties of artificial soils were included in the study, neutral (pH 7.2), acidic (pH 4.3) and basic (pH 9.8). The effects of the NMs on germination, plant growth, and P content were assessed at the 15th and 30th days after treatment. The results showed that P-based NMs enhance the overall germination and plant growth by increasing P levels in all types of soils for the tomato plants in comparison to the bulk P sources. Analysis using X-ray fluorescence revealed enhanced P content in the plants indicating the uptake of P-based NMs. Evaluation of H2O2, total phenolics and total flavonoids contents after NM treatment suggest that there is no stress caused due to the application of NMs to the plant. The results of this study indicate the beneficial role of P-based NMs as fertilizers at the early stages of plant development, which opens a scope for further investigation of underlying metabolic and molecular pathways and field trials.

Abiotic factors and aging alter the physicochemical characteristics and toxicity of Phosphorus nanomaterials to zebrafish embryos.

Nanoscale phosphorus (P)-based formulations are being investigated as potentially new fertilizers to overcome the challenges of conventional bulk P fertilizers in agriculture, including low efficacy rates and high application levels. After agricultural applications, the NMs may be released into aquatic environments and transform over time (by aging) or in the presence of abiotic factors such as natural organic matter or sunlight exposure. It is, therefore, important to investigate the physicochemical changes of NMs in environmentally realistic conditions and assess their potential acute and sublethal toxic effects on aquatic organisms. To investigate this, two separate studies were conducted: 1. the effects of 3-months aged P-based NMs on zebrafish embryos, and 2. the influence of humic acid (HA), UV exposure, or a combination of both on P-based NM toxicity in zebrafish embryos. Four different types of nanohydroxyapatites (nHAPs) and a nanophosphorus (nP) were included in the study. These NMs differed in their physicochemical properties, most prominently their shape and size. Environmental transformations were observed for P-based NMs due to aging or interaction with abiotic factors. The aging of the NMs increased the hydrodynamic diameter (HDD) of rod- and needle-shaped NMs and decreased the size of the platelet and spherical NMs, whereas interactions with HA and UV decreased the NMs' HDD. It was observed that no LC50 (survival) and IC50 (hatch and heart rates) were obtained when the zebrafish embryos were exposed to the aged NMs or when NMs were added in the presence of HA and UV. Overall, these results suggest that P-based NMs cause no acute toxicity and minimal sub-lethal toxicity to zebrafish embryos in environmentally realistic experimental conditions.

Rhizophagus proliferus genome sequence reiterates conservation of genetic traits in AM fungi, but predicts higher saprotrophic activity.

Arbuscular mycorrhizal (AM) fungi are ubiquitous endosymbionts of terrestrial plants. It helps plants to extract more nutrients from the soil and enhances the plant tolerance to various ecological stress factors. The AM fungal genome sequence helps to identify the gene repertoires that are crucial for adaptation to different habitat and mechanisms for interaction with host plant. The present work comprises the first draft of the genome sequence of Rhizophagus proliferus, which is an important AM species present in biofertilizer consortia for agricultural purpose. The estimated genome size of R. proliferus is ~ 110 Mbps and its genomic assembly is 94.35% complete. Genome mining was carried out to identify putative gene families important for biological functions. A total of 22,526 protein-coding genes were estimated in the genome, with an abundance of kinases and reduced number of glycoside hydrolases as compared to other fungal classes. A striking finding in the R. proliferus genome was higher number of carbohydrate esterases (CE), which may suggest towards presence of higher saprotrophic activity in this species as compared to the previously reported AM fungi, which may indicate towards its role as a link between plants and soil mineral nutrients. The genome sequence and annotation of R. proliferus presented here would serve as an important reference for functional genomics studies required for developing biofertilizer formulations in future. In addition, the findings from this work may also prove important in deciphering molecular mechanisms in AM fungi that govern the host-specific interaction and associated agriculture benefits.

Important innate differences in determining symbiotic responsiveness in host and non-hosts of arbuscular mycorrhiza.

Genetic components that regulate arbuscular mycorrhizal (AM) interactions in hosts and non-hosts are not completely known. Comparative transcriptomic analysis was combined with phylogenetic studies to identify the factors that distinguish AM host from non-host. Mycorrhized host, non-mycorrhized host and non-host cultivars of tomato (Solanum lycopersicum) were subjected to RNA seq analysis. The top 10 differentially expressed genes were subjected to extensive in silico phylogenetic analysis along with 10 more candidate genes that have been previously reported for AM-plant interactions. Seven distantly related hosts and four non-hosts were selected to identify structural differences in selected gene/protein candidates. The screened genes/proteins were subjected to MEME, CODEML and DIVERGE analysis to identify evolutionary patterns that differentiate hosts from non-hosts. Based on the results, candidate genes were categorized as highly influenced (SYMRK and CCaMK), moderately influenced and minimally influenced by evolutionary constraints. We propose that the amino acid and nucleotide changes specific to non-hosts are likely to correspond to aberrations in functionality towards AM symbiosis. This study paves way for future research aimed at understanding innate differences in genetic make-up of AM hosts and non-hosts, in addition to the theory of gene losses from the "AM-symbiotic toolkit".

Investigation into the trophic transfer and acute toxicity of phosphorus-based nano-agromaterials in Caenorhabditis elegans.

Biogenic phosphorus (P) based - nanomaterials (NMs) are currently being explored as nanofertilizers. In this study, the acute toxic effects and trophic transfer of multiple types of P-based NMs were examined on soil-dwelling nematode, Caenorhabditis elegans. The study involved four variants of nanohydroxyapatites (nHAPs) synthesized either via a biogenic or a chemical route and another NM, nanophosphorus (nP), biosynthesized from bulk rock phosphate (RP). The pristine NMs differed in their physicochemical properties with each possessing different shapes (biogenic nHAP: platelet-shaped, ˜35 nm; biogenic nP, ˜5-10 nm: dots; chemically synthesized nHAPs: spherical, ˜33 nm, rod, ˜80 nm and needle-shaped, ˜64 nm). The toxic effects of NMs' in C. elegans were assessed using survival, hatching and reproductive cycle as the key endpoints in comparison to bulk controls, calcium phosphate and RP. The interactions and potential uptake of fluorescent-tagged nHAP to E. coli OP50 and C. elegans were investigated using confocal microscopy. The transformation of NMs within the nematode gut was also explored using dynamic light scattering and electron microscopy. C. elegans exposed to all of the variants of nHAP and the nP had 88-100% survival and 82-100% hatch rates and insignificant effects on brood size as observed at the tested environmentally relevant concentrations ranging from 5 to 100 µg.mL-1. Confocal microscopy confirmed the interaction and binding of fluorescent-tagged nHAP with the surface of E. coli OP50 and their trophic transfer and internalization into C. elegans. Interestingly, there was only a small reduction in the hydrodynamic diameter of the nHAP after their uptake into C. elegans and the transformed NMs did not induce any additional toxicity as evident by healthy brood sizes after 72 h. This study provides key information about the environmental safety of agriculturally relevant P-based NMs on non-target species.

Genome-wide identification of Azospirillum brasilense Sp245 small RNAs responsive to nitrogen starvation and likely involvement in plant-microbe interactions.

BACKGROUND: Small RNAs (sRNAs) are non-coding RNAs known to regulate various biological functions such as stress adaptation, metabolism, virulence as well as pathogenicity across a wide range of bacteria, mainly by controlling mRNA stabilization or regulating translation. Identification and functional characterization of sRNAs has been carried out in various plant growth-promoting bacteria and they have been shown to help the cells cope up with environmental stress. No study has been carried out to uncover these regulatory molecules in the diazotrophic alpha-proteobacterium Azospirillum brasilense Sp245 to date. RESULTS: Expression-based sRNA identification (RNA-seq) revealed the first list of ~ 468 sRNA candidate genes in A. brasilense Sp245 that were differentially expressed in nitrogen starvation versus non-starved conditions. In parallel, in silico tools also identified 2 of the above as candidate sRNAs. Altogether, putative candidates were stringently curated from RNA-seq data based on known sRNA parameters (size, location, secondary structure, and abundance). In total, ~ 59 significantly expressed sRNAs were identified in this study of which 53 are potentially novel sRNAs as they have no Rfam and BSRD homologs. Sixteen sRNAs were randomly selected and validated for differential expression, which largely was found to be in congruence with the RNA-seq data. CONCLUSIONS: Differential expression of 468 A. brasilense sRNAs was indicated by RNA-seq data, a subset of which was confirmed by expression analysis. Four of the significantly expressed sRNAs were not observed in nitrogen starvation while 16 sRNAs were found to be exclusively expressed in nitrogen depletion. Putative candidate sRNAs identified have potential mRNA targets primarily involved in stress (abiotic and biotic) adaptability; regulation of bacterial cellular, biological and molecular pathways such as nitrogen fixation, polyhydroxybutyrate synthesis, chemotaxis, biofilm formation and transcriptional regulation. In addition to directly influencing bacteria, some of these sRNAs also have targets influencing plant-microbe interactions through adhesion of bacteria to plant roots directly, suppressing host response, inducing plant defence and signalling.

A new method for biological synthesis of agriculturally relevant nanohydroxyapatite with elucidated effects on soil bacteria.

The study describes a novel and environment friendly route of biosynthesis of nanohydroxyapatite (nHAP). Bacillus licheniformis mediated synthesis of nHAP has been carried out with different phosphate concentrations (2%, 5%, 10% and 20% w/v) of potassium dihydrogen orthophosphate monobasic (K2HPO4). The synthesis is supported by a two-step mechanism - (i) solubilization of P by organic acids extracellularly secreted by the bacterial strain and (ii) gelation of P and Ca. The nHAP particles were characterized using electron microscopy and XRD analysis. Powdered crystalline particles with a size range of 30 ± 5 nm were obtained with shape and size dependent on phosphate concentrations. The particles showed no adverse effect on plant growth-promoting bacteria. Evaluation of nHAP prepared by this route with 2% P source provides scope for a wide range of applications, especially as a nano-fertilizer.

Draft genome sequence of Aspergillus flavus isolate TERIBR1, a highly tolerant fungus to chromium stress.

OBJECTIVES: Aspergillus flavus isolate TERIBR1 was isolated from tannery sludge highly contaminated with chromium. During characterization process, it exhibited capability to adapt and grow in fungal growth media amended with chromium concentration as high as 250 mg/l. In order to understand the genetic underpinnings of the chromium tolerance trait, whole genome sequencing of the TERIBR1 genome was carried out. Information from the current genome will facilitate an understanding of the mechanisms underlying fungal adaptation to heavy metal stress and also heavy metal bioremediation. DATA DESCRIPTION: Here, we report the draft genome sequence along with the assembly and annotation methods used for genome sequence of the A. flavus isolate TERIBR1. The draft genome assembly size is estimated at 37.7 Mb coding for 13,587 genes and has high similarity to the reference genome of A. flavus strain NRRL3357.

Role of Endocrine-Disrupting Engineered Nanomaterials in the Pathogenesis of Type 2 Diabetes Mellitus.

Nanotechnology has enabled the development of innovative technologies and products for several industrial sectors. Their unique physicochemical and size-dependent properties make the engineered nanomaterials (ENMs) superior for devising solutions for various research and development sectors, which are otherwise unachievable by their bulk forms. However, the remarkable advantages mediated by ENMs and their applications have also raised concerns regarding their possible toxicological impacts on human health. The actual issue stems from the absence of systematic data on ENM exposure-mediated health hazards. In this direction, a comprehensive exploration on the health-related consequences, especially with respect to endocrine disruption-related metabolic disorders, is largely lacking. The reasons for the rapid increase in diabetes and obesity in the modern world remain largely unclear, and epidemiological studies indicate that the increased presence of endocrine disrupting chemicals (EDCs) in the environment may influence the incidence of metabolic diseases. Functional similarities, such as mimicking natural hormonal actions, have been observed between the endocrine-disrupting chemicals (EDCs) and ENMs, which supports the view that different types of NMs may be capable of altering the physiological activity of the endocrine system. Disruption of the endocrine system leads to hormonal imbalance, which may influence the development and pathogenesis of metabolic disorders, particularly type 2 diabetes mellitus (T2DM). Evidence from many in vitro, in vivo and epidemiological studies, suggests that ENMs generally exert deleterious effects on the molecular/hormonal pathways and the organ systems involved in the pathogenesis of T2DM. However, the available data from several such studies are not congruent, especially because of discrepancies in study design, and therefore need to be carefully examined before drawing meaningful inferences. In this review, we discuss the outcomes of ENM exposure in correlation with the development of T2DM. In particular, the review focuses on the following sub-topics: (1) an overview of the sources of human exposure to NMs, (2) systems involved in the uptake of ENMs into human body, (3) endocrine disrupting engineered nanomaterials (EDENMs) and mechanisms underlying the pathogenesis of T2DM, (4) evidence of the role of EDENMs in the pathogenesis of T2DM from in vitro, in vivo and epidemiological studies, and (5) conclusions and perspectives.