Cat Gene Research Articles

Comparative analysis of two L-carnitine preparations and their concentration effects on CAT expression in healthy human peripheral blood lymphocyte cultures


 
 CAT gene encodes catalase, a key antioxidant enzyme in the body against oxidative stress. This enzyme plays an important role in the molecular mechanisms of inflammation, apoptosis, mutagenesis and tumorigenesis. Anti-oxidant L-carnitine is used in food supplementation, medical co-treatment and bodyweight regulation. We aimed to investigate molecular basis of L-carnitine commercial preparations supplementation in reducing oxidative stress with customized CAT gene assay in vitro. Human lymphocytes cell culture was established using standard procedure and treated with range of concentrations of L-carnitine in two preparations. We tested two preparations: 500 mg tablets of L-carnitine and liquid L-carnitine with vitamin B6. L-carnitine significantly reduced the expression of CAT gene in cultured lymphocytes at concentrations of 50 μmol/l and 250 μmol/l compared to negative control, (p = 0,001; p = 0,001; respectively). The L-carnitine liquid supplement with vitamin B6 also reduced the transcription of CAT gene at concentrations of 50 μmol/l and 250 μmol/l as compared to the negative control (p = 0,018; p = 0,006; respectively). Selected L-carnitine preparations modulated the transcriptional activity of the antioxidant enzyme gene in human lymphocyte culture, indicating its possible effects in inhibition of pro-inflammatory processes that involve catalase activity.
 
 
 
 

Effects of Genetic Polymorphisms of Cathepsin A on Metabolism of Tenofovir Alafenamide

Cathepsin A (CatA) is important as a drug-metabolizing enzyme responsible for the activation of prodrugs, such as the anti-human immunodeficiency virus drug Tenofovir Alafenamide (TAF). The present study was undertaken to clarify the presence of polymorphisms of the CatA gene in healthy Japanese subjects and the influence of gene polymorphism on the expression level of CatA protein and the drug-metabolizing activity. Single-strand conformation polymorphism method was used to analyze genetic polymorphisms in healthy Japanese subjects. Nine genetic polymorphisms were identified in the CatA gene. The polymorphism (85_87CTG>-) in exon 2 was a mutation causing a deletion of leucine, resulting in the change of the leucine 9-repeat (Leu9) to 8-repeat (Leu8) in the signal peptide region of CatA protein. The effect of Leu8 on the expression level of CatA protein was evaluated in Flp-In-293 cells with a stably expressed CatA, resulting in the expression of CatA protein being significantly elevated in variant 2 with Leu8 compared with Leu9. Higher concentrations of tenofovir alanine (TFV-Ala), a metabolite of TAF, were observed in the Leu8-expressing cells than in the Leu9-expressing cells using LC/MS/MS. Our findings suggest that the drug metabolic activity of CatA is altered by the genetic polymorphism.

3.5-GHz radiofrequency electromagnetic radiation promotes the development of Drosophila melanogaster

With the rapidly increasing popularity of 5G mobile technology, the effect of radiofrequency radiation on human health has caused public concern. This study explores the effects of a simulated 3.5 GHz radiofrequency electromagnetic radiation (RF-EMF) environment on the development and microbiome of flies under intensities of 0.1 W/m2, 1 W/m2 and 10 W/m2. We found that the pupation percentages in the first 3 days and eclosion rate in the first 2 days were increased under exposure to RF-EMF, and the mean development time was shortened. In a study on third-instar larvae, the expression levels of the heat shock protein genes hsp22, hsp26 and hsp70 and humoral immune system genes AttC, TotC and TotA were all significantly increased. In the oxidative stress system, DuoX gene expression was decreased, sod2 and cat gene expression levels were increased, and SOD and CAT enzyme activity also showed a significant increase. According to the 16S rDNA results, the diversity and species abundance of the microbial community decreased significantly, and according to the functional prediction analysis, the genera Acetobacter and Lactobacillus were significantly increased. In conclusion, 3.5 GHz RF-EMF may enhance thermal stress, oxidative stress and humoral immunity, cause changes in the microbial community, and regulate the insulin/TOR and ecdysteroid signalling pathways to promote fly development.

Pharmacological Effect of Caffeine on Drosophila melanogaster: A Proof-of-Concept in vivo Study for Nootropic Investigation.

A comprehensive investigation into drug candidates with nootropic activity using a proper and high throughput yet economical model organism is an important issue to consider. This proof-of-concept study was carried out to determine whether Drosophila melanogaster can be used as an in vivo screening platform to assess the nootropic activity of certain candidates for the treatment of neurodegenerative diseases. To test this, caffeine was used as a nootropic compound and a Drosophila mutant line lacking PGRP-LB with hyperactivation of NF-κB leading to early death with neurodegenerative phenotype was used as a model organism. Caffeine was orally administered via food to the PGRP-LB mutant of D. melanogaster at different concentrations (0.4 mM, 0.08 mM, 0.016 mM) prior to phenotypical observations of the survival and locomotor activity, as well as gene expression analysis, to assess the expression level of sod1, sod2, and cat genes. The results pointed out that the lifespan of D. melanogaster treated with 0.016 mM caffeine was dramatically increased; nonetheless, no changes were observed in the locomotor activity. Phenotypical analysis using a T-maze vial test demonstrated a good cognitive improvement in response to caffeine administration. Molecular analysis revealed that caffeine at a concentration of 0,016 mM induced the expression of the endogenous antioxidant genes sod1 and cat, but not sod2, signifying that the increased lifespan may be associated with a marked improvement in cytoplasmic antioxidant function. In general, the findings of the present study are in line with those previously observed in the mammalian model organism. Therefore, it can be concluded that D. melanogaster can be used as a model organism in preliminary investigation and screening of nootropic candidates prior to further testing in its mammalian counterparts.

Putative regulatory functions of SNPs associated with bronchial asthma, arterial hypertension and their comorbid phenotype.

Linkage disequilibrium (LD) of single nucleotide polymorphisms (SNPs) of TLR4/AL160272.2 (rs1927914, rs1928298, rs7038716, rs7026297, rs7025144) was estimated in the Slavs of West Siberia. We further investigated an association of SNPs in TLR4/AL160272.2 (rs1927914, rs7038716, rs7025144), SERPINA1 (rs1980616), ATXN2/BRAP (rs11065987), IL2RB (rs2284033), NT5C2 (rs11191582), CARD8 (rs11669386), ANG/RNASE4 (rs1010461), and ABTB2/ САТ (rs2022318) genes with bronchial asthma (BA), arterial hypertension (AH) and their comorbidity. Then, the disease-associated SNPs were annotated in silico in relation to their potential regulatory functions. Strong LD was detected between rs1928298 and rs1927914, as well as rs7026297 and rs7038716 in the Slavs of West Siberia. It was found that the rs1927914 G allele of the TLR4 gene and the rs1980616 C allele of the SERPINA1 gene are associated with the predisposition to BA. These SNPs can affect binding affinity of transcription factors of the Pou and Klf4 families, as well as the expression levels of the TLR4 and SERPINA1 genes. The rs11065987 allele A of the ATXN2/BRAP genes, the rs11669386 A allele of the CARD8 gene, the rs2284033 allele G of the IL2RB gene, and the rs11191582 allele G of the NT5C2 gene were associated with the risk of AH. These variants can alter binding affinity of the Hoxa9, Irf, RORalpha1 and HMG-IY transcription factors, as well as the expression levels of the ALDH2, CARD8, NT5C2, ARL3, and SFXN2 genes in blood cells/vessels/heart, respectively. The risk of developing a comorbid phenotype of AD and AH is associated with the A allele of rs7038716 and the T allele of rs7025144 of the TLR4/AL160272.2 genes, the A allele of rs1010461 of the ANG gene and the C allele of rs2022318 of the ABTB2/CAT genes. Variants rs7038716 and rs7025144 can change the expression levels of the TLR4 gene in blood cells, while rs1010461 and rs2022318 influence the expression levels of the ANG and RNASE4 genes as well as the CAT and ABTB2 genes in blood cells, lungs/vessels/heart.

Regulation of iron homeostasis by peroxide-sensitive CatR, a Fur-family regulator in Streptomyces coelicolor.

CatR, a peroxide-sensing transcriptional repressor of Fur family, can de-repress the transcription of the catA gene encoding catalase upon peroxide stress in Streptomyces coelicolor. Since CatR-regulated genes other than catA and its own gene catR have not been identified in detail, the understanding of the role of CatR regulon is very limited. In this study, we performed transcriptomic analysis to identify genes influenced by both ΔcatR mutation and hydrogen peroxide treatment. Through ChIP-qPCR and other analyses, a new consensus sequence was found in CatR-responsive promoter region of catR gene and catA operon for direct regulation. In addition, vtlA (SCO2027) and SCO4983 were identified as new members of the CatR regulon. Expression levels of iron uptake genes were reduced by hydrogen peroxide and a DmdR1 binding sequence was identified in promoters of these genes. The increase in free iron by hydrogen peroxide was thought to suppress the iron import system by DmdR1. A putative exporter protein VtlA regulated by CatR appeared to reduce intracellular iron to prevent oxidative stress. The name vtlA (VIT1-like transporter) was proposed for iron homeostasis related gene SCO2027.

Protective Effect of Selenium on the Oxidative Damage of Kidney Cells Induced by Sodium Nitrite in Grass Carp (Ctenopharyngodon idellus).

The present study was conducted to investigate the protective effects of selenium on the oxidative damage of kidney cells (CIK) caused by nitrite exposure in grass carp (Ctenopharyngodon idella). Cells were pre-incubated by Na2SeO3 (10μmol/L) for 12h and then exposed to NaNO2 (25mg/L) for 24h, the cell viability, apoptosis, gene expression, and antioxidant enzyme activity were assayed. The results show that nitrite reduced cell viability and induced apoptosis, and the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) as well as the sod, cat, and gpx genes reduced (p < 0.05), while the intracellular calcium ion concentration increased (p < 0.05). Interestingly, selenium treatment significantly alleviated the nitrite induced changes in cell growth, apoptosis, and calcium influx. The cell viability after low-concentration selenium treatment is higher than that of normal cells (p < 0.05). CIK cells were pre-incubated with Na2SeO3 and then exposed to NaNO2, the antioxidant indicators could be maintained at normal levels. And compared with nitrite exposure, intracellular calcium ion concentration and apoptotic rate of selenium-incubated still decreased. The expressions of Nrf2 and Keap1 genes increased significantly in CIK cells treated with sodium selenite for 12h, and the same trend as the enzyme activities of this group. The results show that the supplement of selenium can enhance the cell's resistance to sodium nitrite exposure to a certain extent, by alleviating the antioxidant imbalance, high apoptosis rate, and intracellular calcium ion disturbance caused by nitrite exposure. And the Nrf2-Keap1 pathway may play an important role in the process.

Effect of Chrysophyllum albidum fruit pulp powder on antioxidant and proinflammatory genes in non-diabetic and type 2 diabetic rats.

Diabetes mellitus (DM) is a metabolic disorder characterized by chronic hyperglycemia resulting from insulin deficiency or dysfunction. The imbalance between free radicals and antioxidants known as oxidative stress has been implicated in the pathogenesis and complications associated with DM. Chrysophyllum albidum is a seasonal fruit found to be rich in natural antioxidants. DM was induced by high-fat diet dietary supplementation for 14days followed by intraperitoneal injection of streptozotocin (35mg/kg). Thirty-five experimental rats were then divided into seven groups viz.: non-diabetic control; diabetic control; metformin; diabetic and non-diabetic fed with 5 and 10% C. albidum fruit pulp powder (CAFPP). Fasting blood glucose was done with an automatic auto-analyzer and weights were monitored at three-day intervals. The expressions of Nrf2, SOD, CAT, GST, TNF-α, DPP4, and insulin were investigated using RT-PCR. Schrödinger suites was used for docking of C. albidum phytocompounds with insulin. Diabetic rats fed with CAFPP for thirteen days have their blood glucose lowered significantly (p < 0.05) and gained weight compared to diabetic control. CAFPP significantly (p < 0.05) up-regulated Nrf2, CAT, GST, SOD, and insulin genes expression in the diabetic group relative to diabetic control with concomitant down-regulation of TNF-α and DPP4 genes expression. Molecular docking of compounds previously characterized from C. albidum revealed that they are potent ligands of insulin receptors. The study revealed that CAFPP could be effective in the management of DM-related oxidative stress by up-regulating antioxidant and down-regulating pro-inflammatory genes expression. It also positively modulates genes associated with glucose metabolism. The online version contains supplementary material available at 10.1007/s40200-021-00921-0.

Developmental titanium dioxide nanoparticle exposure induces oxidative stress and neurobehavioral changes in zebrafish.

The widespread commercial application of titanium dioxide nanoparticles (TiO2 NPs) leads to ubiquitous presence of TiO2 NPs in the aquatic environment, which highlights the necessity to determine their potential adverse effects on aquatic organisms. The developing nerve system is particularly susceptible to environment perturbation. However, few studies have explored the developmental neurobehavioral toxicity of TiO2 NPs, especially at smaller particle size ranges (≤20nm) that have relatively longer retention time in the water column. In this study, zebrafish embryos were exposed to non-teratogenic concentrations of 0.1 and 1mg/L TiO2 NPs (average size of 14-20nm) from 8 to 108h post-fertilization (hpf) followed by various assessments at different time points up to 12 days post-fertilization (dpf). Our findings revealed that 1mg/L TiO2 NPs perturbed the motor and social behaviors in larval zebrafish. These behavioral changes were characterized by decreased swimming speed in a locomotor response test at 5dpf, increased travel distance in a flash stimulus test at 5dpf, increased preference to the light zone in a light/dark preference test at 10dpf, and increased mirror attack and percent time spent in the mirror zone in a mirror stimulus response assay at 12dpf. Mechanistic examinations at 5dpf revealed elevated cell apoptosis and oxidative stress. Cell apoptosis was characterized by increased acridine orange (AO) positive cells in the olfactory region and neuromasts of the lateral line system. Oxidative stress was characterized by increased lipid peroxidation, increased ROS production, and upregulated catalase (cat) gene expression. In addition, TiO2 NP exposure also upregulated genes associated with the developmental nervous system such as the growth associated protein 43 (gap43) and proliferating cell nuclear antigen (pcna). Our results suggest that the neurobehavioral changes in larvae exposed to 1mg/L TiO2 NPs during early development may result from cell apoptosis and oxidative stress induced neuronal damages.

Screening and validation of RT-qPCR reference genes of Viola yedoensis under different light intensities

Screening the reference genes that were stably expressed under different light intensities for Viola yedoensis could provide reference for the related molecular research. In this study, 11 candidate reference genes were detected by RT-qPCR for tissues of V. yedoensis treated with different light intensities. Ge Norm, Norm Finder, Best Keeper, and Ref Finder website were used to comprehensively evaluate the reference genes, and verify the stability of the reference gene based on CAT1. Finally, the ideal reference gene was determined. The results showed that CYP, Actin, and SAMDC had small Ct value ranges and stable expression. Ge Norm demonstrated that CYP, SAMDC, and Actin were suitable reference genes. Norm Finder showed that the expression of α-TUB was the most stable. Best Keeper recommended CYP, Actin, and SAMDC as reference genes. Ref Finder assessed that SAMDC, CYP, α-TUB, and Actin had better stability, while GAPDH had the worst stability. The expression trend of CAT1 gene was consistent when calibrated with SAMDC, CYP, and Actin, while it was quite different from that calibrated with GAPDH. In summary, SAMDC, CYP, and Actin can be used as ideal reference genes for the gene expression profiling of V. yedoensis under different light intensities.

Interactive effects of dietary quercetin nanoparticles on growth, flesh antioxidant capacity and transcription of cytokines and Aeromonas hydrophila quorum sensing orchestrating genes in Nile tilapia (Oreochromis niloticus)

Recently, the concept of incorporating natural products into nanocarriers has been intended to promote fish growth and health via modulating their stability and bioavailability. In this concern, the potential role of reformulated quercetin into nanocarriers was examined, for the first time, on Nile tilapia's performance and immunity, flesh quality and antioxidant indices and disease resistance. Five hundred fish assigned into five experimental groups with formulated diets containing quercetin nanoparticles (QT-NPs) at levels of 0, 100, 200, 300 and 400 mg/kg were challenged with Aeromonas hydrophila (A. hydrophila) after 12 weeks feeding trial. Fish final body weight gain and feed efficiency were significantly maximized in groups enriched with 300 and 400 mg/kg of QT-NPs. Significant reduction in total saturated fatty acids and an elevation in polyunsaturated fatty acids’ contents were noticed in fish fed higher QT-NPs doses. The levels of Hydrogen peroxide, reactive oxygen species and malondialdehyde, the markers of meat antioxidant capacity, were reduced by higher inclusion levels of QT-NPs. Accordingly, serum activities and transcriptional levels of GSH-Px, CAT and SOD genes were increased with elevated QT-NPs levels. Immune responses mediated by upregulation of IL-10 and TGF-β and downregulation of IL-1β, IL-8 and TNF-α mRNA levels were found to be positively affected by QT-NPs. Dietary QT-NPs downregulated the expression of ahyI and ahyR quorum sensing genes conferring protection against A. hydrophila challenge. This study concluded that supplementation of quercetin in encapsulated nanoparticles could improve its efficacy making it as a compelling approach to improve fish performance and as a promising drug candidate against A. hydrophila virulence.

Cylindrospermopsin impairs vascular smooth muscle cells by P53-mediated apoptosis due to ROS overproduction

Cylindrospermopsin (CYN) is a toxic secondary metabolite from cyanobacteria that can cause cardiovascular disease. However, the study of CYN-induced cardiovascular toxicity in vitro is very limited and the mechanism is remain to be clarified. Vascular smooth muscle cells (VMSCs) have an important function in maintaining the structural and functional integrity of the aortic wall, and are an important in vitro model for cardiovascular research. Thus, the effects of CYN exposure (2, 20, 200, and 2000 nM) on VMSCs were analyzed. In vitro study, results showed that CYN exposure decreased VMSCs viability, inhibited VMSCs migration, induced DNA damage, destroyed cytoskeleton, changed cell morphology, promoted VMSCs apoptosis, and increased intracellular reactive oxygen species (ROS) levels. In addition, CYN could induce the activities of SOD, CAT and GPX, and promote the expressions of SOD1, CAT, GPx1, p53 and Bax genes and inhibit the expression of Bcl-2 gene, leading to a higher ratio of Bax/Bcl-2. Taken together, CYN may induce ROS overproduction, leading to increased p53 expression and ultimately promoting VSMC apoptosis. Therefore, the present study demonstrates that CYN could impair VMSCs, leading to vascular developmental defects and angiocardiopathy.

Genomic Analysis of Ciprofloxacin-Resistant Salmonella enterica Serovar Kentucky ST198 From Spanish Hospitals.

Salmonella enterica serovar Kentucky (S. Kentucky) with sequence type (ST) 198 and highly resistant to ciprofloxacin (ST198-CipR) has emerged as a global MDR clone, posing a threat to public health. In the present study, whole genome sequencing (WGS) was applied to characterize all CipR S. Kentucky detected in five Spanish hospitals during 2009–2018. All CipR isolates (n = 13) were ST198 and carried point mutations in the quinolone resistance-determining regions (QRDRs) of both gyrA (resulting in Ser83Phe and Asp87Gly, Asp87Asn, or Asp87Tyr substitutions in GyrA) and parC (with Thr57Ser and Ser80Ile substitutions in ParC). Resistances to other antibiotics (ampicillin, chloramphenicol, gentamicin, streptomycin, sulfonamides, and tetracycline), mediated by the blaTEM–1B, catA1, aacA5, aadA7, strA, strB, sul1, and tet(A) genes, and arranged in different combinations, were also observed. Analysis of the genetic environment of the latter resistance genes revealed the presence of multiple variants of SGI1 (Salmonella genomic island 1)-K and SGI1-P, where all these resistance genes except catA1 were placed. IS26 elements, found at multiple locations within the SGI1 variants, have probably played a crucial role in their generation. Despite the wide diversity of SGI1-K- and SGI1-P-like structures, phylogenetic analysis revealed a close relationship between isolates from different hospitals, which were separated by a minimum of two and a maximum of 160 single nucleotide polymorphisms. Considering that S. enterica isolates resistant to fluoroquinolones belong to the high priority list of antibiotic-resistant bacteria compiled by the World Health Organization, continuous surveillance of the S. Kentucky ST198-CIPR clone is required.

MS-275 (Entinostat) Promotes Radio-Sensitivity in PAX3-FOXO1 Rhabdomyosarcoma Cells.

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma of childhood. About 25% of RMS expresses fusion oncoproteins such as PAX3/PAX7-FOXO1 (fusion-positive, FP) while fusion-negative (FN)-RMS harbors RAS mutations. Radiotherapy (RT) plays a crucial role in local control but metastatic RMS is often radio-resistant. HDAC inhibitors (HDACi) radio-sensitize different cancer cells types. Thus, we evaluated MS-275 (Entinostat), a Class I and IV HDACi, in combination with RT on RMS cells in vitro and in vivo. MS-275 reversibly hampered cell survival in vitro in FN-RMS RD (RASmut) and irreversibly in FP-RMS RH30 cell lines down-regulating cyclin A, B, and D1, up-regulating p21 and p27 and reducing ERKs activity, and c-Myc expression in RD and PI3K/Akt/mTOR activity and N-Myc expression in RH30 cells. Further, MS-275 and RT combination reduced colony formation ability of RH30 cells. In both cell lines, co-treatment increased DNA damage repair inhibition and reactive oxygen species formation, down-regulated NRF2, SOD, CAT and GPx4 anti-oxidant genes and improved RT ability to induce G2 growth arrest. MS-275 inhibited in vivo growth of RH30 cells and completely prevented the growth of RT-unresponsive RH30 xenografts when combined with radiation. Thus, MS-275 could be considered as a radio-sensitizing agent for the treatment of intrinsically radio-resistant PAX3-FOXO1 RMS.

NRN1 and CAT Gene Polymorphisms, Complex Noise, and Lifestyles interactively Affect the Risk of Noise-induced Hearing Loss

ObjectiveThe effects of interactions between genetic and environmental factors on the noise-induced hearing loss (NIHL) are still unclear. This study aimed to assess interactions among gene polymorphisms, noise metrics, and lifestyles on the risk of NIHL. MethodsA case-control study was conducted using 307 patients with NIHL and 307 matched healthy individuals from five manufacturing industries. General demographic data, lifestyle details, and noise exposure levels were recorded. The Kompetitive allele-specific polymerase chain reaction (KASP) was used to analyze the genotypes of 18 SNPs. ResultsGMDR model demonstrated a relevant interaction between NRN1 rs3805789 and CAT rs7943316 (P = 0.0107). Subjects with T allele of rs3805789 or T allele of rs7943316 had higher risks of NIHL than those with the SNP pair of rs3805789-CC and rs7943316-AA (P < 0.05). There was an interaction among rs3805789, rs7943316, and kurtosis (P = 0.0010). Subjects exposed to complex noise and carrying both rs3805789-CT and rs7943316-TT or rs3805789-CT/TT and rs7943316-AA had higher risks of NIHL than those exposed to steady noise and carrying both rs3805789-CC and rs7943316-AA (P < 0.05). The best six-locus model involving NRN1 rs3805789, CAT rs7943316, smoking, video volume, physical exercise, and working pressure for the risk of NIHL was found to be the interaction (P = 0.0010). An interaction was also found among smoking, video volume, physical exercise, working pressure, and kurtosis (P = 0.0107). ConclusionConcurrence of NRN1 and CAT constitutes a genetic risk factor for NIHL. Complex noise exposure significantly increases the risk of NIHL in subjects with a high genetic risk score. Interactions between genes and lifestyles as well as noise metrics and lifestyles affect the risk of NIHL.

Combinatorial pathway balancing provides biosynthetic access to 2-fluoro-cis,cis-muconate in engineered Pseudomonas putida

SummaryThe wealth of bio-based building blocks produced by engineered microorganisms seldom include halogen atoms. Muconate is a platform chemical with a number of industrial applications that could be broadened by introducing fluorine atoms to tune its physicochemical properties. The soil bacterium Pseudomonas putida naturally assimilates benzoate via the ortho-cleavage pathway with cis,cis-muconate as intermediate. Here, we harnessed the native enzymatic machinery (encoded within the ben and cat gene clusters) to provide catalytic access to 2-fluoro-cis,cis-muconate (2-FMA) from fluorinated benzoates. The reactions in this pathway are highly imbalanced, leading to accumulation of toxic intermediates and limited substrate conversion. By disentangling regulatory patterns of ben and cat in response to fluorinated effectors, metabolic activities were adjusted to favor 2-FMA biosynthesis. After implementing this combinatorial approach, engineered P. putida converted 3-fluorobenzoate to 2-FMA at the maximum theoretical yield. Hence, this study illustrates how synthetic biology can expand the diversity of nature's biochemical catalysis.

Effect of progressive drought stress on physio-biochemical responses and gene expression patterns in wheat.

The online version contains supplementary material available at 10.1007/s13205-021-02991-6.

Antimicrobial resistance and genetic diversity of Enterococcus faecalis from yolk sac infections in broiler chicks

Despite restrictions on the use of antibiotics in poultry, the percentage of multidrug resistant bacteria, isolated from both adult birds and chicks, remains high. These bacteria can spread between countries via hatching eggs or chicks. Antibiotic resistant bacteria can also pose a threat to hatchery and farm workers or to consumers of poultry. The aim of the study was to perform a phenotypic and genotypic analysis of the drug resistance of E. faecalis isolates from yolk sac infections in broiler chicks from Poland and the Netherlands and to determine their genetic diversity. The tests revealed resistance to antibiotics from category D, that is, tetracycline (69.7%); category C – lincomycin (98.7%), erythromycin (51.3%), aminoglycosides (high-level streptomycin and kanamycin resistance – 10.5% and 3.95%, respectively), and chloramphenicol (7.9%); and category B – ciprofloxacin (25% with resistance or intermediate resistance). No resistance to penicillin, ampicillin, high-level gentamicin, tigecycline, or linezolid was noted. Various combinations of the erm(B), tet(M), tet(L), tet(O), ant(6)-Ia, aph(3′)-IIIa, ant(4′)-Ia, cat, and msr(A/B) genes were detected in all isolates (irrespective of the drug-resistance phenotype). Among isolates that carried the tet(M) and/or the tet(L) gene, 28% also had the Int-Tn gene, in contrast with isolates possessing tet(O). There were 28 sequence types and 43 PFGE restriction patterns. About 60% of isolates were of sequences types ST59, ST16, ST116, ST282, ST36, and ST82. Nine new sequence types were shown (ST836-ST844). In conclusion, broiler chicks can be a source of drug-resistant sequence types of E. faecalis that are potentially hazardous for people and animals. Restrictive programs for antibiotic use in broiler breeding flocks should be developed to decrease drug resistance in day-old chicks and reduce economic losses during rearing.

Bulk and nanoparticles of zinc oxide exerted their beneficial effects by conferring modifications in transcription factors, histone deacetylase, carbon and nitrogen assimilation, antioxidant biomarkers, and secondary metabolism in soybean.

Nanoscience paves the way for producing highly potent fertilizers and pesticides to meet farmer’s expectations. This study investigated the physiological and molecular responses of soybean seedlings to the long-time application of zinc oxide nanoparticles (ZnO NPs) and their bulk type (BZnO) at 5 mg L-1 under the two application methods (I- foliar application; II- soil method). The ZnO NPs/BZnO treatments in a substance type- and method-dependent manner improved plant growth performance and yield. ZnO NPs transactionally upregulated the EREB gene. However, the expression of the bHLH gene displayed a contrary downward trend in response to the supplements. ZnO NPs moderately stimulated the transcription of R2R3MYB. The HSF-34 gene was also exhibited a similar upward trend in response to the nano-supplements. Moreover, the ZnONP treatments mediated significant upregulation in the WRKY1 transcription factor. Furthermore, the MAPK1 gene displayed a similar upregulation trend in response to the supplements. The foliar application of ZnONP slightly upregulated transcription of the HDA3 gene, while this gene showed a contrary slight downregulation trend in response to the supplementation of nutrient solution. The upregulation in the CAT gene also resulted from the nano-supplements. The concentrations of photosynthetic pigments exhibited an increasing trend in the ZnONP-treated seedlings. The applied treatments contributed to the upregulation in the activity of nitrate reductase and the increase in the proline concentrations. ZnO NPs induced the activity of antioxidant enzymes, including peroxidase and catalase by averages of 48.3% and 41%, respectively. The utilization of ZnO NPs mediated stimulation in the activity of phenylalanine ammonia-lyase and increase in soluble phenols. The findings further underline this view that the long-time application of ZnO NPs at low concentrations is a safe low-risk approach to meet agricultural requirements.

Evaluation of inducible promoter-riboswitch constructs for heterologous protein expression in the cyanobacterial species Anabaena sp. PCC 7120.

Cyanobacteria are promising chassis for synthetic biology applications due to the fact that they are photosynthetic organisms capable of growing in simple, inexpensive media. Given their slower growth rate than other model organisms such as Escherichia coli and Saccharomyces cerevisiae, there are fewer synthetic biology tools and promoters available for use in model cyanobacteria. Here, we compared a small library of promoter–riboswitch constructs for synthetic biology applications in Anabaena sp. PCC 7120, a model filamentous cyanobacterium. These constructs were designed from six cyanobacterial promoters of various strengths, each paired with one of two theophylline-responsive riboswitches. The promoter–riboswitch pairs were cloned upstream of a chloramphenicol acetyltransferase (cat) gene, and CAT activity was quantified using an in vitro assay. Addition of theophylline to cultures increased the CAT activity in almost all cases, allowing inducible protein production with natively constitutive promoters. We found that riboswitch F tended to have a lower induced and uninduced production compared to riboswitch E for the weak and medium promoters, although the difference was larger for the uninduced production, in accord with previous research. The strong promoters yielded a higher baseline CAT activity than medium strength and weak promoters. In addition, we observed no appreciable difference between CAT activity measured from strong promoters cultured in uninduced and induced conditions. The results of this study add to the genetic toolbox for cyanobacteria and allow future natural product and synthetic biology researchers to choose a construct that fits their needs.