Genetic variants of glutamate receptor gene family in Taiwanese Kawasaki disease children with coronary artery aneurysms

Background

Patients with Kawasaki disease (KD), a pediatric systemic vasculitis, may develop coronary artery aneurysm (CAA) as a complication. To investigate the role of glutamate receptors in KD and its CAA development, we performed genetic association studies.

Methods and results

We examined the whole family of glutamate receptors by genetic association studies in a Taiwanese cohort of 262 KD patients. We identified glutamate receptor ionotropic, kainate 1 (GRIK1) as a novel susceptibility locus associated with CAA formation in KD. Statistically significant differences were noted for factors like fever duration, 1st Intravenous immunoglobulin (IVIG) used time (number of days after the first day of fever) and the GRIK1 (rs466013, rs425507, and rs38700) genetic variants. This significant association persisted even after using multivariate regression analysis (Full model: for rs466013: odds ratio =2.12; 95% CI =1.22-3.65; for rs425507: odds ratio =2.16; 95% CI =1.26-3.76; for rs388700: odds ratio =2.16; 95% CI =1.26-3.76).

Conclusions

We demonstrated that GRIK1 polymorphisms are associated CAA formation in KD, even when adjusted for fever duration and IVIG used time, and may also serve as a genetic marker for the CAA formation in KD.

     

Metabolic changes of genetically engineered grapes (<Emphasis Type="Italic">Vitis vinifera</Emphasis> L.) studied by <Superscript>1</Superscript>H-NMR,metabolite heatmaps and <Emphasis Type="Italic">i</Emphasis>PLS

Introduction

The Deficiens Homologue 9-iaaM (DefH9-iaaM) gene is an ovule-specific auxin-synthesizing gene which is expressed specifically in placenta/ovules and promotes auxin-synthesis. It was introduced into the genome of two grape cultivars Thompson Seedless and Silcora and both transgenic cultivars had an increased number of berries per bunch.

Objectives

This study investigates the down-stream metabolic changes of Silcora and Thompson seedless grape cultivars when genetically modified through the insertion of the DefH9-iaaM gene into their genome.

Methods

The effects of the genetic modification upon the grape metabolome were evaluated through ¹H-NMR and exploratory data analysis. Chemometric tools such as Interval Partial Least Squares regression and metabolite heatmaps were employed for scrutinizing the changes in the transgenic metabolome as compared to the wild type one.

Results

The results show that the pleiotropic effect on the grape metabolome as a function of the gene modifications is relatively low, although the insertion of the transgene caused a decrement in malic acid and proline and an increment in p-coumaric acid content. In addition, the concentration of malic acid was successfully correlated with the number of inserted copies of transgene in the Silcora cultivar, proving that the increased production of berries, promoted by the inserted gene, is achieved at the expense of a decrement in malic acid concentration.

Conclusion

NMR together with chemometrics is able to identify specific metabolites that were up- or down regulated in the genetically engineered plants allowing highlighting alterations in the down-stream metabolic pathways due to the up-stream genetic modifications.

     

Iridoid and phenylethanoid/phenylpropanoid metabolite profiles of <Emphasis Type="Italic">Scrophularia</Emphasis> and <Emphasis Type="Italic">Verbascum</Emphasis> species used medicinally in North America

Introduction

Botanicals containing iridoid and phenylethanoid/phenylpropanoid glycosides are used worldwide for the treatment of inflammatory musculoskeletal conditions that are primary causes of human years lived with disability, such as arthritis and lower back pain.

Objectives

We report the analysis of candidate anti-inflammatory metabolites of several endemic Scrophularia species and Verbascum thapsus used medicinally by peoples of North America.

Methods

Leaves, stems, and roots were analyzed by ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) and partial least squares-discriminant analysis (PLS-DA) was performed in MetaboAnalyst 3.0 after processing the datasets in Progenesis QI.

Results

Comparison of the datasets revealed significant and differential accumulation of iridoid and phenylethanoid/phenylpropanoid glycosides in the tissues of the endemic Scrophularia species and Verbascum thapsus.

Conclusions

Our investigation identified several species of pharmacological interest as good sources for harpagoside and other important anti-inflammatory metabolites.

     

<Emphasis Type="Italic">PiggyBac</Emphasis> transposon-mediated mutagenesis and application in yeast <Emphasis Type="Italic">Komagataella phaffii</Emphasis>

Objective

Around one-fourth of the Komagataella phaffii genes encode hypothetical proteins with unknown functions. However, lack of powerful tools for genetic screening in K. phaffii significantly limits the functional analysis of these unknown genes. Transposon mutagenesis has been utilized as an insertional mutagenesis tool in many other organisms and would be extremely valuable if it could be applied in K. phaffii.

Results

In this study, we investigated in K. phaffii the transposition activity and efficiency of piggyBac (PB) transposon, a DNA transposon from the cabbage looper moth Trichoplusia ni through the integrated-plasmid system. We also designed a binary-plasmid system which could generate stable mutants. Finally we evaluated the quality of this mutagenesis system by a simple screening for functional genes involved in K. phaffii carbon catabolite repression.

Conclusions

Our results demonstrate that PB-mediated mutagenesis could be a feasible and useful tool for functional gene screening in K. phaffii.

     

<Emphasis Type="Italic">massPix</Emphasis>: an R package for annotation and interpretation of mass spectrometry imaging data for lipidomics

Introduction

Mass spectrometry imaging (MSI) experiments result in complex multi-dimensional datasets, which require specialist data analysis tools.

Objectives

We have developed massPix—an R package for analysing and interpreting data from MSI of lipids in tissue.

Methods

massPix produces single ion images, performs multivariate statistics and provides putative lipid annotations based on accurate mass matching against generated lipid libraries.

Results

Classification of tissue regions with high spectral similarly can be carried out by principal components analysis (PCA) or k-means clustering.

Conclusion

massPix is an open-source tool for the analysis and statistical interpretation of MSI data, and is particularly useful for lipidomics applications.

     

Role of genetic variations of <Emphasis Type="Italic">chitinase 3</Emphasis>-<Emphasis Type="Italic">like 1</Emphasis> in bronchial asthmatic patients

Background

Single nucleotide polymorphisms (SNPs) in chitinase 3-like 1 (CHI3L1) are associated with bronchial severity and pulmonary function. CHI3L1 proteins are involved in both innate and adaptive immune responses; however, to date, the correlation of these SNPs and their age of onset of bronchial asthma has not been demonstrated.

Methods

To address the role of these genetic variations, 390 patients with well-controlled bronchial asthma and living in Japan were recruited, genotyped, and had a pulmonary function test performed on them in this study. To analyze the concentration levels of CHI3L1 protein, bronchial lavage fluids were examined.

Results

Forced expiratory volume in one second, %predicted (%FEV1), was significantly decreased in homozygotes of rs1214194 compared to heterozygotes and wild type. The age of onset of adult bronchial asthma was significantly younger in GG homozygotes of rs4950928 and AA homozygotes of rs1214194 than in the other two genotypes. The concentration of CHI3L1 protein in bronchial lavage fluid increased in both homozygotes of rs4950928 and rs1214194.

Conclusions

Our study demonstrated that the homozygotes of rs4950928 and rs1214194 of CHI3L1 might predict an early onset of bronchial asthma and have the propensity to promote airway remodeling.Trial registration JMA-IIA00045 remodeling-ICS

     

Effects of <Emphasis Type="Italic">ADIPOQ</Emphasis> polymorphisms on PCOS risk: a meta-analysis

Background

Whether adiponectin (ADIPOQ) polymorphisms are associated with the risk of polycystic ovary syndrome (PCOS) remain controversial. Therefore, we performed this study to better explore correlations between ADIPOQ polymorphisms and PCOS risk.

Methods

Literature retrieve was conducted in PubMed, Medline and Embase. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated.

Results

Eighteen studies were enrolled for analyses. Pooled overall analyses showed that rs1501299 polymorphism was significantly associated with PCOS risk (recessive model: p?=?0.02, OR?=?0.77, 95%CI 0.62–0.95; allele model: p?=?0.001, OR?=?1.15, 95%CI 1.06–1.26). Further subgroup analyses according to ethnicity of participants revealed that rs1501299 and rs2241766 polymorphisms were both significantly correlated with PCOS risk in Caucasians. In addition, rs1501299 polymorphism was also significantly correlated with PCOS risk in East Asians.

Conclusions

Our findings indicated that rs1501299 and rs2241766 polymorphisms might serve as genetic biomarkers of PCOS in certain ethnicities.

     

Improvement of 1,3-propanediol production in <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis> by moderate expression of <Emphasis Type="Italic">puuC</Emphasis> (encoding an aldehyde dehydrogenase)

Objective

To improve 1,3-propanediol production in Klebsiella pneumoniae, the effects of puuC expression in lactate- and lactate/2,3-butanediol-deficient strains were assessed.

Results

Overexpression of puuC (encoding an aldehyde dehydrogenase) inhibited 1,3-propanediol production and increased 3-hydroxypropionic acid formation in both lactate- and lactate/2,3-butanediol-deficient strains. An improvement in 1,3-propanediol production was only achieved in a lactate-deficient strain via moderate expression of puuC; at the end of the fermentation, 1,3-propanediol productivity increased by 14 % compared with the control. Further comparative analysis of the metabolic flux distributions in different strains indicated that 3-hydroxypropionic acid formation could play a considerable role in cell metabolism in K. pneumoniae.

Conclusion

An improvement in 3-hydroxypropionic acid formation would be beneficial for cell metabolism, which can be accomplished by enhancing 1,3-propanediol productivity in a lactate-deficient strain via moderate expression of puuC.

     

Development of a novel engineered antibody targeting <Emphasis Type="Italic">Neisseria</Emphasis> species

Objectives

A Neissaria bacterial pilus sugar, bacillosamine, was synthesized and, for the first time, used as a probe to screen a single-chain variable fragment (scFv).

Results

Four Neisseria, Neisseria gonorrhoeae, Neisseria meningitidis, Neisseria sicca and Neisseria subflava, and two negative controls, Streptococcus pneumoniae and Escherichia coli, were tested through ELISA, immunostaining and gold nanoparticle immunological assay. All results indicated that the selected scFv is feasible for the specific detection of Neisseria species via the recognition of bacillosamine.

Conclusions

The recombinant scFv could detect Neisseria strains at 10⁶ CFU/ml.

     

The role of <Emphasis Type="Italic">IFITM3</Emphasis> in the growth and migration of human glioma cells

Background

Interferon induced transmembrane protein 3 (IFITM3) is transcribed in most tissues and highly interferon-inducible. However, the role of IFITM3 in cancer is still poorly understood.

Methods

Expression levels ofIFITM3were analyzed in 60 glioma patients by immunohistochemistry (IHC). Following closely, we investigated the phenotype of IFITM3 knockdown on glioma cell growth and tumorigenesis in vitro using lentivirus-mediated loss-of-function strategy.

Results

Depletion of IFITM3in U251 cells dramatically inhibited cell proliferation and colony formation, which demonstrated that reduced IFITM3 protein levels could cause inhibition of tumorigenesis. Knockdown of IFITM3 also induced cell cycle arrest in G0/G1 phase, especially in the sub-G1 phase representing apoptotic cells. In addition, the migration of U251 cells was visibly weakened after IFITM3 knockdown, as determined by Transwell assay.

Conclusions

Our findings provide new evidence that IFITM3 plays an important role in glioma cell growth and migration, suggesting that silencing of IFITM3 by RNA interference (RNAi) may be a potential approach to suppress glioma growth.

     

Metabolic response of porcine colon explants to in vitro infection by <Emphasis Type="Italic">Brachyspira hyodysenteriae</Emphasis>: a leap into disease pathophysiology

Introduction

Swine dysentery caused by Brachyspira hyodysenteriae is a production limiting disease in pig farming. Currently antimicrobial therapy is the only treatment and control method available.

Objective

The aim of this study was to characterize the metabolic response of porcine colon explants to infection by B. hyodysenteriae.

Methods

Porcine colon explants exposed to B. hyodysenteriae were analyzed for histopathological, metabolic and pro-inflammatory gene expression changes.

Results

Significant epithelial necrosis, increased levels of l-citrulline and IL-1α were observed on explants infected with B. hyodysenteriae.

Conclusions

The spirochete induces necrosis in vitro likely through an inflammatory process mediated by IL-1α and NO.

     

Degradation of carbazole,dibenzothiophene and polyaromatic hydrocarbons by recombinant <Emphasis Type="Italic">Rhodococcus</Emphasis> sp.

Objectives

With the view of designing a single biocatalyst for biorefining, carbazole dioxygenase was cloned from Pseudomonas sp. and expressed in Rhodococcus sp.

Results

The recombinant, IGTS8, degraded both carbazole and dibenzothiophene at 400 mg/l in 24 h. Maximum carbazole degradation was in 1:1 (v/v) hexadecane/aqueous phase. Anthracene, phenanthrene, pyrene, fluoranthene and fluorine were also degraded without affecting the aliphatic component.

Conclusions

Recombinant Rhodococcus sp. IGTS8 can function as a single biocatalyst for removing major contaminants of fossil fuels viz. dibenzothiophene, carbazole and polyaromatic compounds.

     

Synthesis of rosmarinic acid analogues in <Emphasis Type="Italic">Escherichia coli</Emphasis>

Objectives

To produce rosmarinic acid analogues in the recombinant Escherichia coli BLRA1, harboring a 4-coumarate: CoA ligase from Arabidopsis thaliana (At4CL) and a rosmarinic acid synthase from Coleus blumei (CbRAS).

Results

Incubation of the recombinant E. coli strain BLRA1 with exogenously supplied phenyllactic acid (PL) and analogues as acceptor substrates, and coumaric acid and analogues as donor substrates led to production of 18 compounds, including 13 unnatural RA analogues.

Conclusion

This work demonstrates the viability of synthesizing a broad range of rosmarinic acid analogues in E. coli, and sheds new light on the substrate specificity of CbRAS.

     

Determination and optimization of a strong promoter element from <Emphasis Type="Italic">Bacillus amyloliquefaciens</Emphasis> by using a promoter probe vector

Objective

To construct a promoter probe vector, pBE-bgaB, to screen strong promoters from Bacillus amyloliquefaciens.

Results

266 colonies containing active promoter elements from the genomic DNA of B. amyloliquefaciens were identified. Among these, promoter P41 exhibited the strongest β-Gal activity in Escherichia coli and B. amyloliquefaciens. Sequence analysis showed that promoter P41 contained P _ykuN , a ykuN gene encoding flavodoxin. Optimization of the ribosome-binding site from P41 to P₃₈₂ improved β-Gal activity by ~ 200%.

Conclusion

A new strong promoter for protein expression and genetic engineering of Bacillus species.

     

Seroprevalence of <Emphasis Type="Italic">Borrelia burgdorferi</Emphasis> sensu lato and <Emphasis Type="Italic">Anaplasma phagocytophilum</Emphasis> in Danish horses

Background

Borrelia burgdorferi sensu lato and Anaplasma phagocytophilum are able to infect horses. However, the extend to which Danish horses are infected and seroconvert due to these two bacteria is unknown. The aim of the present study was to evaluate the seroprevalence of B. burgdorferi sensu lato and A. phagocytophilum in Danish horses.

Methods

A total of 390 blood samples collected from all major regions of Denmark and with a geographical distribution corresponding to the density of the Danish horse population were analyzed. All samples were examined for the presence of antibodies against B. burgdorferi sensu lato and A. phagocytophilum by the use of the SNAP^®4DX ^® ELISA test.

Results

Overall, 29.0% of the horses were seropositive for B. burgdorferi sensu lato whereas 22.3% were seropositive for A. phagocytophilum.

Conclusions

Antibodies against B burgdorferi sensu lato and A. phagocytophilum are commonly found among Danish horses thus showing that Danish horses are frequently infected by these organisms.

     

Enhanced succinic acid production in <Emphasis Type="Italic">Corynebacterium glutamicum</Emphasis> with increasing the available NADH supply and glucose consumption rate by decreasing H<Superscript>+</Superscript>-ATPase activity

Objectives

To enhance succinic acid production in Corynebacterium glutamicum by increasing the supply of NADH and the rate of glucose consumption by decreasing H⁺-ATPase activity.

Results

A mutant of C. glutamicum NC-3-1 with decreased H⁺-ATPase activity was constructed. This increased the rate of glycolysis and the supply of NADH. Fermentation of C. glutamicum NC-3-1 gave 39 % higher succinic acid production (113 and 81 g/l), a 29 % higher succinic acid yield (0.94 and 0.73 g succinic acid/g glucose) and decreased by-products formation compared to that of C. glutamicum NC-3 in 5 l bioreactor.

Conclusion

The point mutation in C. glutamicum NC-3-1 increased the rate of glycolysis and resulted in higher succinic acid production, higher succinic acid yield and significantly decreased formation of by-products.

     

<Emphasis Type="Italic">Streptococcus pyogenes</Emphasis>-purpura fulminans as an invasive form of group A streptococcal infection

Background

Streptococcus pyogenes is an uncommon pathogen of purpura fulminans, and the pathogenesis of S. pyogenes-purpura fulminans remains unclear because of paucity of cases. We reported a pediatric case of S. pyogenes-purpura fulminans with literature review of the disease.

Case presentation

A 3-year-old boy showed limping, lethargy and acral gangrene within 24 h. A diagnosis of S. pyogenes-purpura fulminans was made for bacterial isolation from throat and peripheral blood. Intensive therapy led to a survival with amputation of the left distal metatarsal bone, and normal development. The isolated M12 carried no mutation of csrS/R or rgg. Thrombophilia or immunodeficiency was excluded.

Discussion

Twelve-reported cases (9 pediatric and 3 elderly) of S. pyogenes-purpura fulminans started with shock and coagulopathy. Five patients age <?8 years had no underlying disease and survived. One youngest and two immunocompromised patients died.

Conclusion

Streptococcus pyogenes-acute infectious purpura fulminans is a distinctive rare form of aggressive GAS infections.

     

Anaerobic microplate assay for direct microbial conversion of switchgrass and Avicel using <Emphasis Type="Italic">Clostridium thermocellum</Emphasis>

Objective

To develop and prototype a high-throughput microplate assay to assess anaerobic microorganisms and lignocellulosic biomasses in a rapid, cost-effective screen for consolidated bioprocessing potential.

Results

Clostridium thermocellum parent Δhpt strain deconstructed Avicel to cellobiose, glucose, and generated lactic acid, formic acid, acetic acid and ethanol as fermentation products in titers and ratios similar to larger scale fermentations confirming the suitability of a plate-based method for C. thermocellum growth studies. C. thermocellum strain LL1210, with gene deletions in the key central metabolic pathways, produced higher ethanol titers in the Consolidated Bioprocessing (CBP) plate assay for both Avicel and switchgrass fermentations when compared to the Δhpt strain.

Conclusion

A prototype microplate assay system is developed that will facilitate high-throughput bioprospecting for new lignocellulosic biomass types, genetic variants and new microbial strains for bioethanol production.