A spiropyran with low pK_a for tracking DNA G-quadruplexes and revealing the dissipation of ΔΨm with senescence using an in-situ switching strategy

The in-situ fluorescence triggering of bioprobes only using endogenous bioforces is an ideal non-destructive real-time detection method, which is of particular interest to improve the accuracy of clinical diagnosis and treatment. We have recently reported a strategy of spiropyran in-situ switching triggered by endogenous biological forces in vivo to develop optical probes for this purpose. However, such probes, as with all spiropyrans, are sensitive to lysosomal acidity. We here present a spiropyran-based fluorescent probe TANG with low pKa, which can recognize intranuclear DNA G4s in situ without the aid of exogenous light or chemicals and is as stable to lysosomal acidity due to a decreased pKa value (4.3). Interestingly, despite the stability to lysosomal pH environment, the TANG spiropyran can be opened in situ by the negative membrane potential of extranuclear mitochondria (ΔΨm), causing a ratiometric change in fluorescence signals and providing the in-situ and real-time tracking of ΔΨm. Of note, ratiometric imaging using TANG indicates that ΔΨm decreases gradually with cellular senescence, which is to the best of our knowledge the first visualization of such mitochondrion-related aging processes using a ratiometric imaging approach.</p

A spiropyran with low pK_a for tracking DNA G-quadruplexes and revealing the dissipation of ΔΨm with senescence using an in-situ switching strategy

The in-situ fluorescence triggering of bioprobes only using endogenous bioforces is an ideal non-destructive real-time detection method, which is of particular interest to improve the accuracy of clinical diagnosis and treatment. We have recently reported a strategy of spiropyran in-situ switching triggered by endogenous biological forces in vivo to develop optical probes for this purpose. However, such probes, as with all spiropyrans, are sensitive to lysosomal acidity. We here present a spiropyran-based fluorescent probe TANG with low pKa, which can recognize intranuclear DNA G4s in situ without the aid of exogenous light or chemicals and is as stable to lysosomal acidity due to a decreased pKa value (4.3). Interestingly, despite the stability to lysosomal pH environment, the TANG spiropyran can be opened in situ by the negative membrane potential of extranuclear mitochondria (ΔΨm), causing a ratiometric change in fluorescence signals and providing the in-situ and real-time tracking of ΔΨm. Of note, ratiometric imaging using TANG indicates that ΔΨm decreases gradually with cellular senescence, which is to the best of our knowledge the first visualization of such mitochondrion-related aging processes using a ratiometric imaging approach.</p

First Report of Coexistence of blaSFO–1 and blaNDM–1 β-Lactamase Genes as Well as Colistin Resistance Gene mcr-9 in a Transferrable Plasmid of a Clinical Isolate of Enterobacter hormaechei

Many antimicrobial resistance genes usually located on transferable plasmids are responsible for multiple antimicrobial resistance among multidrug-resistant (MDR) Gram-negative bacteria. The aim of this study is to characterize a carbapenemase-producing Enterobacter hormaechei 1575 isolate from the blood sample in a tertiary hospital in Wuhan, Hubei Province, China. Antimicrobial susceptibility test showed that 1575 was an MDR isolate. The whole genome sequencing (WGS) and comparative genomics were used to deeply analyze the molecular information of the 1575 and to explore the location and structure of antibiotic resistance genes. The three key resistance genes (blaSFO–1, blaNDM–1, and mcr-9) were verified by PCR, and the amplicons were subsequently sequenced. Moreover, the conjugation assay was also performed to determine the transferability of those resistance genes. Plasmid files were determined by the S1 nuclease pulsed-field gel electrophoresis (S1-PFGE). WGS revealed that p1575-1 plasmid was a conjugative plasmid that possessed the rare coexistence of blaSFO–1, blaNDM–1, and mcr-9 genes and complete conjugative systems. And p1575-1 belonged to the plasmid incompatibility group IncHI2 and multilocus sequence typing ST102. Meanwhile, the pMLST type of p1575-1 was IncHI2-ST1. Conjugation assay proved that the MDR p1575-1 plasmid could be transferred to other recipients. S1-PFGE confirmed the location of plasmid with molecular weight of 342,447 bp. All these three resistant genes were flanked by various mobile elements, indicating that the blaSFO–1, blaNDM–1, and mcr-9 could be transferred not only by the p1575-1 plasmid but also by these mobile elements. Taken together, we report for the first time the coexistence of blaSFO–1, blaNDM–1, and mcr-9 on a transferable plasmid in a MDR clinical isolate E. hormaechei, which indicates the possibility of horizontal transfer of antibiotic resistance genes

Assessment of the Xpert MTB/RIF Ultra assay on rapid diagnosis of extrapulmonary tuberculosis

Objective: To evaluate the diagnostic performance of Xpert MTB/RIF Ultra for EPTB (Extrapulmonary Tuberculosis) patients on different types of extrapulmonary specimens from different anatomic sites. Methods: Patients with suspected EPTB were prospectively included, extrapulmonary specimens were collected and subjected to culture, Xpert and Xpert Ultra assays in accordance with relevant guidelines. Results: A total of 225 cases were included which contained 200 EPTB cases (43 culture-positive EPTB, 157 culture-negative EPTB which were diagnosed based on pathological results and a satisfied response to anti-TB treatment) and 25 non-EPTB cases. Sensitivities of Xpert Ultra and Xpert for culture-positive cases were 83.7% (95%CI, 68.7–92.7) and 67.4% (95% CI, 51.3–80.5) respectively. Specificities of Xpert Ultra and Xpert were 92.0% (95% CI, 72.5–98.6) and 96.0% (95% CI, 77.7–99.8) respectively. The sensitivities of Xpert Ultra, Xpert and culture for 200 EPTB cases were 52.5% (105/200, 95% CI, 45.4–59.6), 34.0% (68/200, 95% CI, 27.6–41.1) and 21.5% (43/200, 95% CI, 16.2–28.0) respectively. By comparison among different types of specimens, Xpert Ultra can detect 78.9% (56/71) of EPTB on fine-needle aspiration (FNA) tissues which was higher than that on pleural fluid (43.7% (45/103), p < 0.05. Conclusions: Xpert Ultra assay had a higher sensitivity than those of Xpert and culture on extrapulmonary specimens, which could be a promising approach for rapid EPTB diagnosis. Keywords: Mycobacterium tuberculosis, Xpert MTB/RIF Ultra, Xpert MTB/RIF, Extrapulmonary tuberculosis, Diagnosi

SesI May Be Associated with the Invasiveness of Staphylococcus epidermidis

Staphylococcus epidermidis is a commensal bacterium which widely colonizes in human skin and mucous membrane and rarely causes clinically manifested infections. S. epidermidis surface protein I (SesI) is considered to be the major virulence factor of S. epidermidis infection, but its pathogenesis is not clear. Here, we demonstrated that the prevalence of sesI among S. epidermidis invasive isolates (20.8%, 26/125) was significantly higher than that among colonizing isolates (3.8%, 4/106). The positive rates of biofilm-associated genes (aap, icaA, IS256) and resistance-associated genes mupA among the sesI-positive isolates were significantly higher than those among sesI-negative isolates (p &lt; 0.05). And antimicrobial susceptibility testing showed that the resistance rates of sesI-positive isolates to ciprofloxacin, gentamicin and trimethoprim/sulfamethoxazole were significantly higher than those among sesI-negative isolates. Interestingly, 80.8% (21/26) of sesI-positive isolates belong to ST2 determined by MLST, while ST2 was not found among any of the 99 sesI-negative invasive isolates, indicating that there is a strong association between carriage of sesI and ST2 clone. In order to further study the role of sesI gene in pathogenesis, the sesI gene mutant (S. epidermidis RP62AΔsesI) and complementary expression strain (S. epidermidis RP62AΔsesI-C) were successfully constructed. All experimental data indicated that sesI may promote S. epidermidis to adhere and aggregate, but it had no obvious effect on the mature stage of biofilm formation. Taken together, these results suggest that sesI, along with antimicrobial and other biofilm-associated genes enables S. epidermidis easier for colonization and adhesion and contributes to the spread of S. epidermidis, especially ST2 clone

Assessment of the Cepheid 3-gene Host Response Fingerstick Blood Test (MTB-HR) on rapid diagnosis of tuberculosis

ABSTRACTThe World Health Organization has identified high-priority target product profiles for new TB diagnostics which include rapid biomarker-based, non-sputum-based diagnostic testing, using an easily accessible sample. The Cepheid 3-gene Host Response Fingerstick Blood Prototype Test (MTB-HR) quantifies relative mRNA levels of a 3-gene signature (GBP5, DUSP3, and KLF2) from a whole-blood sample on the GeneXpert platform. The objective of the present study was to evaluate the performance of the MTB-HR to distinguish between active tuberculosis (ATB), latent Mycobacterium tuberculosis infection (LTBI), other pulmonary diseases, and healthy volunteers at a tertiary care centre. Among 653 participants enrolled in this study, 192 were diagnosed as having ATB, and the remaining 461 were classified as non-ATB, including 137 cases of LTBI, 224 cases of other pulmonary diseases, and 100 healthy volunteers. The corresponding AUCs of the MTB-HR in distinguishing untreated ATB from non-ATB, LTBI, other pulmonary diseases, and healthy volunteers were 0.814 (95% CI, 0.760-0.868, sensitivity 76.1%, specificity 71.6%), 0.739 (95% CI, 0.667-0.812, sensitivity 59.7%, specificity 78.1%), 0.825 (95% CI, 0.770-0.880, sensitivity 82.1%, specificity 65.6%), 0.892 (95% CI, 0.839-0.945, sensitivity 76.1%, specificity 88.0%), respectively. When only samples with TAT of less than 1 h were included, the AUC of the MTB-HR in distinguishing untreated ATB from non-ATB was largest, 0.920 (95% CI, 0.822-1.000, sensitivity 81.3%, specificity 87.7%). In conclusion, the MTB-HR assay shows potential as a rapid, blood-based screening and triage test for ATB, especially for untreated ATB, with the advantage of increased diagnostic yield since blood is more readily available

EGCG treats ICH via up-regulating miR-137-3p and inhibiting Parthanatos

Intracranial hemorrhage (ICH) causes high mortality and disability without effective treatment in the clinical setting. (−)-Epigallocatechin-3-gallate (EGCG) exerts an essential role in the central nervous system and offers a promising therapeutic agent for the treatment of oxidative damage-related diseases. MiR-137 can inhibit the oxidative stress and apoptosis to attenuate neuronal injury. However, the role of EGCG in regulating miR-137-3p and neuronal Parthanatos remains to be unclear. In the present study, we build the ICH mice model to investigate the antioxidant effects of EGCG via upregulating miR-137-3p and inhibiting neuronal Parthanatos. We revealed that EGCG upregulated miR-137-3p and inhibited neuronal Parthanatos, and promoted the functional recovery, alleviated ICH-induced brain injury, and reduced oxidative stress in mice following ICH. However, following the inhibition of miR-137-3p and activation of Parthanatos, EGCG was unable to exert neuroprotective roles. These combined results suggest that EGCG may upregulate miR-137-3p and inhibit neuronal Parthanatos to accelerate functional recovery in mice after ICH, laying the foundation for EGCG to be a novel strategy for the treatment of neuronal injuries related to Parthanatos