Study on the Clinical Features and Prognosis of <Emphasis Type="Italic">Penicilliosis marneffei</Emphasis> Without Human Immunodeficiency Virus Infection

Objective

To improve the diagnosis and treatment of Penicilliosis marneffei without human immunodeficiency virus infection.

Methods

Analyze and review the clinical features, diagnosis and treatment of six cases of P. marneffei without human immunodeficiency virus infection at The First Affiliated Hospital of Fujian Medical University.

Results

Two cases were diagnosed in the ENT Department, three cases in the respiratory department and one case in the dermatological department. Penicillium marneffei infection was confirmed by sputum culture, blood culture and tissue biopsy. After definite diagnosis, one refused further treatment, and others showed significant improvement.

Conclusion

Penicilliosis marneffei is insidious onset and easy to be escaped and misdiagnosed. To achieve early diagnosis and appropriate treatment, doubtful cases should be alerted for the diagnoses as P. marneffei.

     

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.

     

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">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.

     

<Emphasis Type="Italic">In silico</Emphasis> characterization and Molecular modeling of double-strand break repair protein <Emphasis Type="Italic">MRE11</Emphasis> from <Emphasis Type="Italic">Phoenix dactylifera</Emphasis> v deglet nour

Background

DNA double-strand breaks (DSBs) are highly cytotoxic and mutagenic. MRE11 plays an essential role in repairing DNA by cleaving broken ends through its 3′ to 5′ exonuclease and single-stranded DNA endonuclease activities.

Methods

The present study aimed to in silico characterization and molecular modeling of MRE11 from Phoenix dactylifera L cv deglet nour (DnMRE11) by various bioinformatic approaches. To identify DnMRE11 cDNA, assembled contigs from our cDNA libraries were analysed using the Blast2GO2.8 program.

Results

The DnMRE11 protein length was 726 amino acids. The results of HUMMER show that DnMRE11 is formed by three domains: the N-terminal core domain containing the nuclease and capping domains, the C-terminal half containing the DNA binding and coiled coil region. The structure of DnMRE11 is predicted using the Swiss-Model server, which contains the nuclease and capping domains. The obtained model was verified with the structure validation programs such as ProSA and QMEAN servers for reliability. Ligand binding studies using COACH indicated the interaction of DnMRE11 protein with two Mn²⁺ ions and dAMP. The ConSurf server predicted that residues of the active site and Nbs binding site have high conservation scores between plant species.

Conclusions

A model structure of DnMRE11 was constructed and validated with various bioinformatics programs which suggested the predicted model to be satisfactory. Further validation studies were conducted by COACH analysis for active site ligand prediction, and revealed the presence of six ligands binding sites and two ligands (2 Mn²⁺ and dAMP).

     

Silencing of <Emphasis Type="Italic">Mythimna separata chitinase</Emphasis> genes via oral delivery of <Emphasis Type="Italic">in planta</Emphasis>-expressed RNAi effectors from a recombinant plant virus

Objectives

To evaluate transient expression of RNA interference (RNAi) effectors in Nicotiana benthamiana plants by using recombinant virus vectors and also oral delivery of the effectors for silencing of Mythimna separata endogenous gene expression.

Results

Mythimna separata is a serious pest of corn production in China. To evaluate RNAi approaches to target specific RNAs in M. separate, we cloned fragments of the M. separata chitinase sequences into a virus vector in order to produce RNAi effectors during virus infection and replication in plants. When the infected plants were fed to M. separata, expression levels of target MseChi1 and MseChi2 genes were down-regulated by 76 and 45 %, respectively, and sequence-specific siRNAs were detected in recipient insects. RNAi-based silencing of chitinase genes also led to body weight decreases by 43 %.

Conclusion

Our research demonstrates target mRNA knockdown and suggests a promising application for controlling of M. separata by in planta expression of RNAi effectors using a recombinant plant virus.

     

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.

     

SfP53 and filamentous actin (F-actin) are the targets of viral pesticide AcMNPV-<Emphasis Type="Italic">Bm</Emphasis>K IT (P10/PH) in host <Emphasis Type="Italic">Spodoptera frugiperda</Emphasis> 9 cells

Objectives

To analyze the anti-insect mechanism of viral pesticide AcMNPV-BmK IT(P10/PH) in the host Spodoptera frugiperda 9 (Sf9) cells.

Results

Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV)- mediated expression of BmK IT, regulated by P10 protein promoter (P10) and polyhedrosis promoter (PH), promoted the replication of progeny virus in host Sf9 cells. AcMNPV-BmK IT(P10) could accelerate the budding process (or speed) of budded virus (BV) in Sf9 cells. The impact of AcMNPV-BmK IT(P10) on the nuclear polymerization of filamentous actin (F-actin) participated in regulating the accelerated budding process. Unexpectedly, both AcMNPV-BmK IT(P10) and AcMNPV-BmK IT(PH) delayed the nuclear polymerization of F-actin and promoted the clearance of F-actin in the nucleus. SfP53, an important apoptosis factor, was involved in the regulation of AcMNPV-BmK IT(P10/PH) in Sf9 cells. AcMNPV-BmK IT(P10/PH) could also delay and promote the nuclear recruitment of SfP53 after 27 h post infection (h p.i.).

Conclusion

SfP53 and F-actin are the targets of viral pesticide AcMNPV-BmK IT (P10/PH) in host Sf9 cells, which provides the experimental basis for the development of recombinant baculovirus biopesticides.

     

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.

     

A versatile <Emphasis Type="Italic">Trichoderma reesei</Emphasis> expression system for the production of heterologous proteins

Objectives

To develop a versatile Trichoderma reesei (teleomorph Hypocrea jecorina) expression system for the high-purity production of heterologous proteins.

Results

The versatile T. reesei expression system is based on xyn1 and xyn2 promoters, A824V transition in XYRI, and a bicomponent carbon source strategy. Red fluorescent protein gene rfp and alkaline endoglucanase EGV gene egv3 from Humicola insolens were used as reporter genes to test our versatile expression system

Conclusions

The versatile T. reesei expression system can be applied to produce heterologous proteins with high purity and high yield.

     

Genome sequence analysis of a flocculant-producing bacterium, <Emphasis Type="Italic">Paenibacillus shenyangensis</Emphasis>

Objective

To explore the metabolic process of Paenibacillus shenyangensis that is an efficient bioflocculant-producing bacterium. The biosynthesis mechanism of bioflocculation was used to enrich the genome of Paenibacillus shenyangensis and provide a basis for molecular genetics and functional genomics analyses.

Results

According to the analysis of de novo assembly, a total of 5,501,467 bp clean reads were generated, and were assembled into 92 contigs. 4800 unigenes were predicted of which 4393 were annotated showing a specific gene function in the NCBI-Nr database. 3423 genes were found in the database of cluster of orthologous groups. Among the 168 Kyoto Encyclopedia of Genes and Genomes database, cell growth and metabolism were the main biological processes, and a potential metabolic pathway was predicted from glucose to exopolysaccharide within the starch and sucrose metabolism pathway.

Conclusion

By using the high-throughput sequencing technology, we provide a genome analysis of Paenibacillus shenyangensis that predicts the main metabolic processes and a potential pathway of exopolysaccharide biosynthesis.

     

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.

     

AcMNPV-<Emphasis Type="Italic">Bm</Emphasis>K IT improves the progeny virus production via baculovirus GP64 envelope fusion protein

Objectives

To analyze the mechanisms underlying the impact of recombinant Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV)-mediated BmK IT expression on the function of baculovirus GP64 envelope fusion protein and progeny virus production.

Results

Viral propagation assay indicated that overexpression GP64 could promote replication of AcMNPV. AcMNPV-mediated expression of BmK IT also promoted replication of AcMNPV. Immunofluorescence analysis showed BmK IT, which was regulated by very early promoter IE1 in AcMNPV, could make the GP64 protein move to the cytomembrane soon after transfection. BmK IT, which is regulated by P10 protein promoter (P10) and polyhedrosis promoter (PH), could promote the expression of GP64.

Conclusion

BmK IT, regulated by very early promoter IE1, P10 protein promoter (P10) and PH, accelerated the expression of GP64 protein, promoted its early cytomembrane localization and then triggered virus budding and progeny virus production.

     

The bacterial community associated with rose-scented geranium (<Emphasis Type="Italic">Pelargonium graveolens</Emphasis>) leaves responds to anthracnose symptoms

Background

The fungus Colletotrichum is a plant pathogen that causes the anthracnose disease, resulting in huge losses in various crops including the rose-scented geranium (Pelargonium graveolens). Although the bacterial community associated with plants has an important role in the establishment of plant diseases, little is known about what happens in P. graveolens.

Aims

To increase the knowledge about the bacterial community associated with P. graveolens and its relationship with anthracnose disease symptoms.

Methods

Quantitative PCR and high-throughput sequencing were combined to determine the presence of the fungus Colletotrichum and to reveal the bacterial communities associated with different plant parts – root, stem and leaf – and in the rhizosphere and bulk soil, and also to determine the respective bacterial communities associated with P. graveolens leaves symptomatic and asymptomatic for anthracnose disease.

Results

The fungus Colletotrichum was detected in all plant parts and in the surrounding soil. Bacterial communities varied spatially in plants, and the disease symptoms also influenced the composition of the bacterial community. Abundances of operational taxonomic units (OTUs) assigned to the phylum Actinobacteria and to the genus Streptococcus were greatly increased in asymptomatic leaves.

Conclusions

The bacterial community associated to geranium leaves responds to anthracnose symptoms.

     

Production of caffeoylmalic acid from glucose in engineered <Emphasis Type="Italic">Escherichia coli</Emphasis>

Objectives

To achieve biosynthesis of caffeoylmalic acid from glucose in engineered Escherichia coli.

Results

We constructed the biosynthetic pathway of caffeoylmalic acid in E. coli by co-expression of heterologous genes RgTAL, HpaBC, At4CL2 and HCT2. To enhance the production of caffeoylmalic acid, we optimized the tyrosine metabolic pathway of E. coli to increase the supply of the substrate caffeic acid. Consequently, an E. coli–E. coli co-culture system was used for the efficient production of caffeoylmalic acid. The final titer of caffeoylmalic acid reached 570.1 mg/L.

Conclusions

Microbial production of caffeoylmalic acid using glucose has application potential. In addition, microbial co-culture is an efficient tool for producing caffeic acid esters.

     

Genetic improvement of <Emphasis Type="Italic">Magnetospirillum gryphiswaldense</Emphasis> for enhanced biological removal of phosphate

Objectives

To improve its phosphate accumulating abilities for phosphate recycling from wastewater, a magnetotactic bacterium, Magnetospirillum gryphiswaldense, was genetically modified to over-express polyphosphate kinase.

Results

Polyphosphate kinase was over-expressed in the bacterium. The recombinant strain accumulated ninefold more polyphosphate from synthetic wastewater compared to original wild type. The magnetic property of the recombinant M. gryphiswaldense strain was retained.

Conclusions

The recombinant M. gryphiswaldense can be used for phosphate removal and recovery in bioremediation.

     

Modification of fatty acid selectivity of <Emphasis Type="Italic">Candida antarctica</Emphasis> lipase A by error-prone PCR

Objective

To generate Candida antarctica lipase A (CAL-A) mutants with modified fatty acid selectivities and improved lipolytic activities using error-prone PCR (epPCR).

Results

A Candida antarctica lipase A mutant was obtained in three rounds of epPCR. This mutant showed a 14 times higher ability to hydrolyze triacylglycerols containing conjugated linoleic acids, and was 12 and 14 times more selective towards cis-9, trans-11 and trans-10, cis-12 isomers respectively, compared to native lipase. Lipolytic activities towards fatty acid esters were markedly improved, in particular towards butyric, lauric, stearic and palmitic esters.

Conclusion

Directed molecular evolution is an efficient method to generate lipases with desirable selectivity towards CLA isomers and improved lipolytic activities towards esters of fatty acids.

     

Inhibitory effects of YCW and MOS from <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> on <Emphasis Type="Italic">Escherichia coli</Emphasis> and <Emphasis Type="Italic">Salmonella pullorum</Emphasis> adhesion to Caco-2 cells

Background

For many years, yeast cell walls (YCW) and mannan oligosaccharides (MOS) have been used as alternatives to antibiotics and health feed additives to enhance the growth performance and health of food animals. In the present study, the inhibitory effects of YCWand MOS on the adhesion of enteropathogenic bacteria to intestinal epithelial cells were tested.

Methods

YCW and MOS were extracted from Saccharomyces cerevisiae (XM 0315), and the morphology of YCW and MOS bound to pathogenic bacteria was observed by scanning electron microscopy (SEM). Real-time fluorescent quantitative PCR was used to quantitatively analyze the effects of YCW and MOS on the adhesion of Escherichia coli (CVCC3367) and Salmonella pullorum (CVCC520) to Caco-2 cells.

Results

The results showed that YCW inhibited E. coli and S. pullorum binding to Caco-2 cells by 95% and 74%, respectively, whereas MOS prevented E. coli and S. pullorum binding by 67% and 50%, respectively.

Conclusions

These data suggest that YCW has a stronger ability than MOS to inhibit pathogenic bacteria from adhering to Caco-2 cells in vitro.

     

Plant growth-promoting bacterium <Emphasis Type="Italic">Pseudomonas fluorescens</Emphasis> FR1 secrets a novel type of extracellular polyhydroxybutyrate polymerase involved in abiotic stress response in plants

Objectives

Identification of novel microbial factors contributing to plant protection against abiotic stress.

Results

The genome of plant growth-promoting bacterium Pseudomonas fluorescens FR1 contains a short mobile element encoding a novel type of extracellular polyhydroxybutyrate (PHB) polymerase (PhbC) associated with a type I secretion system. Genetic analysis using a phbC mutant strain and plants showed that this novel extracellular enzyme is related to the PHB production in planta and suggests that PHB could be a beneficial microbial compound synthesized during plant adaptation to cold stress.

Conclusion

Extracellular PhbC can be used as a new tool for improve crop production under abiotic stress.

     

Metabolomics as a tool for understanding the evolution of <Emphasis Type="Italic">Tabebuia sensu lato</Emphasis>

Background

Plant systematic studies have changed substantially in the last years, stimulated by new strategies for phylogenetic studies. In this regard, chemistry data has been a useful tool for understanding plant phylogenetic relationships.

Objective

Our aim was to apply metabolomic approaches, followed by multivariate statistical analysis and dereplication of Tabebuia sensu lato species, and compare our results with classifications based on traditional taxonomy and molecular phylogeny. We also evaluated the application of metabolomics as a chemotaxonomic identification tool, as well as to enlighten plant chemical evolution.

Methods

Metabolomic data was generated through a high-resolution mass spectrometry with electrospray ionization of 27 Tabebuia sensu lato specimens from different populations, consisting of 15 Handroanthus (from four species) and 12 Tabebuia sensu stricto (from three species). Chemometric tools, such as principal component analysis and metabolite heatmaps, were used to scrutinize the metabolic changes among species.

Results

Tabebuia and Handroanthus species presented different secondary metabolite storage capacity. The genus Tabebuia revealed higher levels of glycosylated iridoids esterified with a phenylpropanoid moiety, such as specioside, verminoside, and minecoside, while Handroanthus accumulated iridoids linked to a simple phenol, lignans, and verbascoside derivatives.

Conclusion

These results corroborate splitting the Tabebuia s.l., which was supported by profound changes in secondary metabolism, suggesting metabolomics as an excellent tool for understanding species evolution.