Gut microbiota dysbiosis-related susceptibility to nontuberculous mycobacterial lung disease.

The role of gut microbiota in host defense against nontuberculous mycobacterial lung disease (NTM-LD) was poorly understood. Here, we showed significant gut microbiota dysbiosis in patients with NTM-LD. Reduced abundance of Prevotella copri was significantly associated with NTM-LD and its disease severity. Compromised TLR2 activation activity in feces and plasma in the NTM-LD patients was highlighted. In the antibiotics-treated mice as a study model, gut microbiota dysbiosis with reduction of TLR2 activation activity in feces, sera, and lung tissue occurred. Transcriptomic analysis demonstrated immunocompromised in lung which were closely associated with increased NTM-LD susceptibility. Oral administration of P. copri or its capsular polysaccharides enhanced TLR2 signaling, restored immune response, and ameliorated NTM-LD susceptibility. Our data highlighted the association of gut microbiota dysbiosis, systematically compromised immunity and NTM-LD development. TLR2 activation by P. copri or its capsular polysaccharides might help prevent NTM-LD.

The Trend of TIM3 Expression on T Cells in Patients With Nontuberculous Mycobacterial Lung Disease: From Immune Cell Dysfunction to Clinical Severity.

Background: The incidence of nontuberculous mycobacterial lung disease (NTM-LD) is increasing worldwide. Immune exhaustion has been reported in NTM-LD, but T-cell immunoglobulin and mucin domain-containing protein 3 (TIM3), a co-inhibitory receptor on T cells, has been scarcely studied. Methods: Patients with NTM-LD and healthy controls were prospectively recruited from July 2014 to August 2019 at three tertiary referral centers in Taiwan. We examined TIM3 expression on the T cells from the participants using flow cytometry. TIM3 expression was analyzed for different disease statuses and after treatment. The apoptosis and cytokine profiles were analyzed according to the TIM3 expression. Results: Among enrolled subjects (47 patients and 46 controls), TIM3 on CD4+ cells (6.44% vs. 4.12%, p = 0.028) and CD8+ cells (18.47% vs. 9.13%, p = 0.003) were higher in NTM-LD patients than in the controls. The TIM3 level on CD4+ and CD8+ T cells was positively associated with T-cell apoptosis in the NTM-LD patients. In stimulating peripheral blood mononuclear cells using PMA plus ionomycin, a high TIM3 level on T cells correlated with low interleukin-2 and tumor necrosis factor-alpha (TNF-α) on CD4+ cells and interferon-gamma and TNF-α on CD8+ T cells. For clinical manifestation, low body mass index (BMI), positive sputum acid-fast smear, and high radiographic score correlated with high TIM3 expression on T cells. After NTM treatment, TIM3+ decreased significantly on CD4+ and CD8+ T cells. Conclusions: In patients with NTM-LD, TIM3+ expression increased over CD4+ and CD8+ T cells and correlated with cell apoptosis and specific cytokine attenuation. Clinically, TIM3+ T cells increased in patients with low BMI, high disease extent, and high bacilli burden but decreased after treatment.

The Dynamic Change of Immune Checkpoints and CD14+ Monocytes in Latent Tuberculosis Infection.

Controlling latent tuberculosis infection (LTBI) is important for preventing tuberculosis (TB). However, the immune regulation of LTBI remains uncertain. Immune checkpoints and CD14+ monocytes are pivotal for immune defense but have been scarcely studied in LTBI. We prospectively enrolled participants with LTBI and controls from January 2017 to December 2019. We measured their CD14+ monocytes and the expression of immune checkpoints, including programmed death-1 (PD-1), cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), and T cell immunoglobulin mucin domain-containing-3 (TIM3) on T lymphocytes in peripheral blood mononuclear cells before and after LTBI treatment. A total of 87 subjects were enrolled, including 29 IGRA-negative healthy controls (HC), 58 in the LTBI group (19 without chronic kidney disease (non-CKD), and 39 with end-stage renal disease (ESRD)). All PD-1, CTLA-4, and TIM3 on lymphocytes and monocytes were higher in the LTBI group than that in the HC group. Total CD14+ monocytes were higher and PD-L2+CD14+ over monocytes were lower in patients with LTBI-non-CKD than that in the HC group. After LTBI treatment, CD14+ monocytes, TIM3+ on CD4+ and monocytes, and CTLA-4 on lymphocytes decreased significantly. Multivariable logistic regression indicated that CD14+ monocytes was an independent factor for LTBI-non-CKD from the HC group, whereas PD-L2+CD14+ monocytes and TIM3+ monocytes were significant for LTBI-ESRD from the HC group. In conclusion, LTBI status was associated with increasing CD14+ monocytes plus low PD-L2 expression. By contrast, increased expression of immune checkpoints over all immune cells might be due to Mycobacterium tuberculosis related immune exhaustion, which decreased after treatment.

A Dual Repeat Cis-Element Determines Expression of GERANYL DIPHOSPHATE SYNTHASE for Monoterpene Production in Phalaenopsis Orchids.

Phalaenopsis bellina is a scented orchid emitting large amount of monoterpenes. GERANYL DIPHOSPHATE SYNTHASE (PbGDPS) is the key enzyme for monoterpene biosynthesis, and shows concomitant expression with the emission of monoterpenes during flower development in P. bellina. Here, we identified a dual repeat cis-element in the GDPS promoter that is critical for monoterpene biosynthesis in Phalaenopsis orchids. A strong correlation between the dual repeat and the monoterpene production was revealed by examination of the GDPS promoter fragments over 12 Phalaenopsis species. Serial-deletion of the 2-kb GDPS promoter fragments demonstrated that the integrity of the dual repeat was crucial for its promoter activities. By screening the Arabidopsis transcription factors (TFs) cDNA library using yeast one-hybrid assay, AtbZIP18, a member of group I of bZIP TFs, was identified to be able to bind the dual repeat. We then identified PbbZIP4 in the transcriptome of P. bellina, showing 83% identity in the DNA binding region with that of AtbZIP18, and the expression level of PbbZIP4 was higher in the scented orchids. In addition, PbbZIP4 transactivated the GDPS promoter fragment containing the dual repeat in dual luciferase assay. Furthermore, transient ectopic expression of PbbZIP4 induced a 10-fold production of monoterpenoids in the scentless orchid. In conclusion, these results indicate that the dual repeat is a real TF-bound cis-element significant for GDPS gene expression, and thus subsequent monoterpene biosynthesis in the scented Phalaenopsis orchids.

High Glucose Concentration Promotes Vancomycin-Enhanced Biofilm Formation of Vancomycin-Non-Susceptible Staphylococcus aureus in Diabetic Mice.

We previously demonstrated that vancomycin treatment increased acquisition of eDNA and enhanced biofilm formation of drug-resistant Staphylococcus aureus through a cidA-mediated autolysis mechanism. Recently we found that such enhancement became more significant under a higher glucose concentration in vitro. We propose that besides improper antibiotic treatment, increased glucose concentration environment in diabetic animals may further enhance biofilm formation of drug-resistant S. aureus. To address this question, the diabetic mouse model infected by vancomycin-resistant S. aureus (VRSA) was used under vancomycin treatment. The capacity to form biofilms was evaluated through a catheter-associated biofilm assay. A 10- and 1000-fold increase in biofilm-bound bacterial colony forming units was observed in samples from diabetic mice without and with vancomycin treatment, respectively, compared to healthy mice. By contrast, in the absence of glucose vancomycin reduced propensity to form biofilms in vitro through the increased production of proteases and DNases from VRSA. Our study highlights the potentially important role of increased glucose concentration in enhancing biofilm formation in vancomycin-treated diabetic mice infected by drug-resistant S. aureus.

Salient region detection improved by principle component analysis and boundary information.

Salient region detection is useful for several image-processing applications, such as adaptive compression, object recognition, image retrieval, filter design, and image retargeting. A novel method to determine the salient regions of images is proposed in this paper. The L0 smoothing filter and principle component analysis (PCA) play important roles in our framework. The L0 filter is extremely helpful in characterizing fundamental image constituents, i.e., salient edges, and can simultaneously diminish insignificant details, thus producing more accurate boundary information for background merging and boundary scoring. PCA can reduce computational complexity as well as attenuate noise and translation errors. A local-global contrast is then used to calculate the distinction. Finally, image segmentation is used to achieve full-resolution saliency maps. The proposed method is compared with other state-of-the-art saliency detection methods and shown to yield higher precision-recall rates and F-measures.

Vancomycin promotes the bacterial autolysis, release of extracellular DNA, and biofilm formation in vancomycin-non-susceptible Staphylococcus aureus.

Staphylococcus aureus, an important human pathogen, is particularly adept at producing biofilms on implanted medical devices. Although antibiotic treatment of nonsusceptible bacteria will not kill these strains, the consequences should be studied. The present study focuses on investigating the effect of vancomycin on biofilm formation by vancomycin-non-susceptible S. aureus. Biofilm adherence assays and scanning electron microscopy demonstrated that biofilm formation was significantly enhanced following vancomycin treatment. Bacterial autolysis of some subpopulations was observed and was confirmed by the live/dead staining and confocal laser scanning microscopy. A significant increase in polysaccharide intercellular adhesin (PIA) production was observed by measuring icaA transcript levels and in a semi-quantitative PIA assay in one resistant strain. We show that the release of extracellular DNA (eDNA) via cidA-mediated autolysis is a major contributor to vancomycin-enhanced biofilm formation. The addition of xenogeneic DNA could also significantly enhance biofilm formation by a PIA-overproducing S. aureus strain. The magnitude of the development of the biofilm depends on a balance between the amounts of eDNA and PIA. In conclusion, sublethal doses of cell wall-active antibiotics like vancomycin induce biofilm formation through an autolysis-dependent mechanism in vancomycin-non-susceptible S. aureus.

Identification of prophage gene z2389 in Escherichia coli EDL933 encoding a DNA cytosine methyltransferase for full protection of NotI sites.

Pulsed-field gel electrophoresis (PFGE) analysis revealed that the genomes of some pathogenic Escherichia coli O157:H7 strains, including EDL933, were resistant to NotI digestion. An amino acid sequence comparison suggested that the z2389 gene carried on prophage CP-933R in strain EDL933 is likely to encode a C(5)-cytosine methyltransferase. The z2389-equivalent gene was found in the NotI-resistant strains tested, but it was not detected in the NotI-susceptible strains. PFGE analysis of the wild-type EDL933 strain and of a z2389 null mutant revealed that z2389 was associated with full genome protection against NotI digestion and partial protection against EagI digestion. In vitro methylation experiments with purified recombinant protein demonstrated that Z2389 is capable of methylating NotI and EagI sites. Sequencing of bisulfite-treated DNA indicated that the methylation occurred at the first cytosine residue of the NotI recognition sequence, whereas EagI sites remained unmethylated or were methylated at the first cytosine residue. Thus, z2389 encodes a DNA cytosine methyltransferase that confers full protection to NotI sites.

Reaction efficiencies and rate constants for the goethite-catalyzed Fenton-like reaction of NAPL-form aromatic hydrocarbons and chloroethylenes.

The contaminants present as nonaqueous phase liquids (NAPLs) in the subsurface are long-term sources for groundwater pollution. Fenton-like reaction catalyzed by natural iron oxides such as goethite in soils is one of the feasible in situ chemical reactions used to remediate contaminated sites. This research evaluated the Fenton-like reaction of five chlorinated ethylenes and three aromatic hydrocarbons using goethite as the catalyst. The reaction efficiencies and rate constants of these compounds in NAPL and dissolved forms were compared. The content of goethite used in batch experiments was in the range similar to those found in subsurfaces. Low H2O2 concentrations (0.05 and 0.1%) were tested in order to represent the low oxidant concentration in the outer region of treatment zone. The results showed that at the tested goethite and H2O2 ranges, the majority of contaminants were removed in the first 120 s. When aromatics and chloroethylenes were present as NAPLs, their removal efficiencies and reaction constants decreased. The removal efficiencies of 0.02 mmol NAPL contaminants were 26-70% less than those of the dissolved. The measured rate constants were in the order of 10(9) M(-1) s(-1) for dissolved chlorinated ethylenes and aromatic hydrocarbons, but were 25-60% less for their NAPL forms. The initial dosage of H2O2 and NAPL surface areas (18.4-38.2 mm2) did not significantly affect reaction efficiencies and rate constants of chlorinated NAPLs. Instead, they were related to the octanol-water partition coefficient of compounds. For both dissolved and NAPL forms, aromatic hydrocarbons were more reactive than chlorinated ethylenes in Fenton-like reaction. These results indicated that the decrease in reaction efficiencies and rate constants of NAPL-form contaminants would pose more negative impacts on the less reactive compounds such as benzene and cis 1,2-DCE during goethite-catalyzed Fenton-like reaction.