Whole genome sequencing analysis of seven unknown resistance mechanisms of carbapenem-resistant Klebsiella pneumoniae strains resistance to ceftazidime-avibactam.

AIMS: To investigate alternative resistance mechanisms among seven ceftazidime-avibactam (CZA)-resistant carbapenem-resistant Klebsiella pneumoniae (CRKP) strains lacking common antimicrobial resistance genes (ARGs) using whole genome sequencing. METHODS AND RESULTS: ARG and virulence factors (VFs) were screened using the ARG database CARD and the VF database, respectively, and identified using genomic annotation data with BLAST+. Six strains were ST11 sequence types (STs), and one was ST2123. ST11 strains harbored more ARGs than the ST2123 strains. All seven strains carried multiple ARGs with efflux-mediated antibiotic resistance, including oqxA, oqxB, tet (A), qacEdltal, CRP, H-NS, Kpn-E, F, G, H, acrA, LptD, acrB, acrD, cpxA, mdtB, and mdtC. These efflux-mediated ARGs were identified in most strains and even all strains. Whole genome sequencing revealed that the ST11 strain carried multiple potential prophages, genomic islands, and integrative and conjugative elements, while the ST2123 strain carried an independent potential prophages and a genomic island. CONCLUSIONS: Whole genome sequencing analysis revealed that these seven CZA-resistant CRKP strains lacking common ARGs exhibited efflux-mediated antibiotic resistance-associated ARGs. The main mechanism by which CRKP resists CZA is antibiotic inactivation. Except for tet (A), no ARGs and validation experiments related to efflux were found. This study's results provide a new possibility for the resistance mechanism of CRKP to CZA, and we will verify this conclusion through experiments in the future.

Identification of residues critical for the extension of Munc18-1 domain 3a.

BACKGROUND: Neurotransmitter release depends on the fusion of synaptic vesicles with the presynaptic membrane and is mainly mediated by SNARE complex assembly. During the transition of Munc18-1/Syntaxin-1 to the SNARE complex, the opening of the Syntaxin-1 linker region catalyzed by Munc13-1 leads to the extension of the domain 3a hinge loop, which enables domain 3a to bind SNARE motifs in Synaptobrevin-2 and Syntaxin-1 and template the SNARE complex assembly. However, the exact mechanism of domain 3a extension remains elusive. RESULTS: Here, we characterized residues on the domain 3a hinge loop that are crucial for the extension of domain 3a by using biophysical and biochemical approaches and electrophysiological recordings. We showed that the mutation of residues T323/M324/R325 disrupted Munc13-1-mediated SNARE complex assembly and membrane fusion starting from Munc18-1/Syntaxin-1 in vitro and caused severe defects in the synaptic exocytosis of mouse cortex neurons in vivo. Moreover, the mutation had no effect on the binding of Synaptobrevin-2 to isolated Munc18-1 or the conformational change of the Syntaxin-1 linker region catalyzed by the Munc13-1 MUN domain. However, the extension of the domain 3a hinge loop in Munc18-1/Syntaxin-1 was completely disrupted by the mutation, leading to the failure of Synaptobrevin-2 binding to Munc18-1/Syntaxin-1. CONCLUSIONS: Together with previous results, our data further support the model that the template function of Munc18-1 in SNARE complex assembly requires the extension of domain 3a, and particular residues in the domain 3a hinge loop are crucial for the autoinhibitory release of domain 3a after the MUN domain opens the Syntaxin-1 linker region.

Super-resolution fluorescence microscopic imaging in pathogenesis and drug treatment of neurological disease.

Since super-resolution fluorescence microscopic technology breaks the diffraction limit that has existed for a long time in optical imaging, it can observe the process of synapses formed between nerve cells and the protein aggregation related to neurological disease. Thus, super-resolution fluorescence microscopic imaging has significantly impacted several industries, including drug development and pathogenesis research, and it is anticipated that it will significantly alter the future of life science research. Here, we focus on several typical super-resolution fluorescence microscopic technologies, introducing their benefits and drawbacks, as well as applications in several common neurological diseases, in the hope that their services will be expanded and improved in the pathogenesis and drug treatment of neurological diseases.

Complete Genome Sequence of Staphylococcus epidermidis CCSM0287, Isolated from Healthy Facial Skin.

Staphylococcus epidermidis strain CCSM0287 was isolated from healthy facial skin. The complete genome of CCSM0287 was sequenced using a combination of Pacific Biosciences (PacBio) RS II single-molecule real-time (SMRT) and Illumina sequencing. The assembled 2.5-Mbp genome consisted of one chromosome and three plasmids.

Experimental study on the redistribution law of heavy metals in sludge disintegration.

Aiming at the problems of large output of excess sludge, difficulty in treatment and disposal, and the potential toxicity of heavy metals restricting its resource utilization, this paper studies the redistribution law of heavy metals in the process of sludge disintegration. The dissertation investigates the distribution law of typical heavy metals such as Cu, Pb, Cd, Zn in the process of microwave and citric acid-microwave cracking sludge under different specific energy and fixed specific energy conditions. The Tessier five-step continuous extraction method was used to extract heavy metals, and the changes in their content and chemical forms were analyzed, which provided certain technical support for the subsequent harmless treatment and resource utilization of excess sludge. The main findings of this paper are as follows: The dissolution rate of heavy metals Cu, Pb, Cd, and Zn increased rapidly during the citric acid-microwave cracking process in the TS specific energy range of 0-45000 kJ/kg, and then gradually tended to be gradual. The maximum dissolution rates of Cu, Pb, Cd, Zn were 8.06%, 16.58%, 14.69%, and 24.11%, respectively. The concentrations of Cu, Pb, Cd, and Zn in the sludge were mainly F4; F3, F4; F2, F3. The proportions of stable states of Cu, Pb, Cd, and Zn in sludge increased to 88.6%, 55.91%, 35.7%, and 31.35%, respectively. When the specific energy was 45000 kJ/kg TS, the concentrations of Pb, Zn, and Cd in the solid phase of the sludge appeared to increase under microwave cracking alone and decrease under the combined action of citric acid and microwave. The concentration of Cu in the solid phase of the sludge increased slightly. The dissolution rates of Pb, Cd, and Zn by microwave alone and citric acid-microwave method were 14.23% and 16.58%, 10.34% and 14.69%, 17.53%, and 24.11%, respectively. The dissolution rates of Cu by both methods were lower. The steady state ratios of Pb and Zn in the citric acid-microwave method increased to 55.91% and 31.25%, respectively; the steady state ratio of Cd in the microwave alone method increased to 39.51%; both methods had no significant effect on the stability of Cu.

PTPRM Is Critical for Synapse Formation Regulated by Zinc Ion.

In the nervous system, the trace metal ion zinc is required for normal mammalian brain development and physiology. Zinc homeostasis is essential for the control of physiological and pathophysiological brain functions. Synapses, the junctions between neurons, are the center of the brain's information transmission. Zinc deficiency or excess leads to neurological disorders. However, it is still unclear whether and how zinc ion regulates synapse formation. Here, we investigated the effect of zinc on synapse formation in a cultured neuron system, and found that synapse formation and synaptic transmission were regulated by zinc ions. Finally, we identified that PTPRM is the key gene involved in synapse formation regulated by zinc ions. This study provides a new perspective to understanding the regulation of brain function by zinc ion.

Urothelial carcinoembryonic antigen 1 score for early detection of prostate cancer and risk prediction.

UCA1 score appears useful in detecting nonhigh-risk (including very low-, low-, or intermediate-risk) prostate cancer. Combination of the PSA level and the UCA1 score may significantly reduce the burden of prostate biopsy.

Evaluation of different strategies to minimize the matrix effects on LC-MS/MS analysis of multiple lipophilic shellfish toxins in both acidic and alkaline chromatographic conditions.

Lipophilic shellfish toxins (LSTs) accumulated by shellfish pose a potential threat to consumer health. A mandatory routine monitoring of LSTs has been adopted for seafood products by liquid chromatography-mass spectrometry (LC-MS) in many countries. In this study, two methods developed on liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) under acidic and alkaline chromatographic conditions were assessed for the determination of multiple LSTs. Different strategies including matrix solid-phase dispersion (MSPD), solid phase extraction (SPE) and sample dilution were applied and evaluated the matrix effects of mussel, scallop, clam, and oyster samples on the signal response of mass spectrometry. Results showed that the alkaline method achieved a lower limit of detection (LOD) and more robust compared to the acidic method. The obvious signal suppression of OA and DTX1 (55%-76%) and signal enhancement of PTX2 (27%-34%) occurred in the crude extracts of shellfish under acidic chromatography. In the alkaline method, no remarkable matrix effects of crude extracts were found except for the scallop matrix on the signal intensity of DTX1, AZA3 and GYM-A (121%-130%). Clean-up methods MSPD, SPE and sample dilution obviously reduced the inhibition of shellfish matrices on the signal response of OA and DTX1, however, which were still subject to signal inhibition under acidic condition. Sample dilution was more effective than SPE and MSPD in minimizing the matrix interference in both acidic and alkaline methods. Furthermore, sample dilution in combination with the alkaline chromatography was the most effective method. Bivalve mollusks harvested from Beibu Bay, South China Sea, were generally contaminated by GYM-A and SPX1 at low concentrations.

Growth and toxin production of Azadinium poporum strains in batch cultures under different nutrient conditions.

Azaspiracid-2 (AZA2) is the dominant toxin produced by Azadinium poporum strains AZDY06 and AZFC22 isolated from the South China Sea. Biomass and AZA2-production were examined within batch cultures with variation in experimental concentrations of nitrate (0, 88, 882, and 2647µM) or phosphate (0, 3.6, 36, and 109µM), different nitrogen sources (nitrate and urea) and media (f/2-Si, L1-Si, and K-Si) in the present study. Growth of both strains positively responded to nitrate or phosphate nutrients, but the growth status was significantly repressed by the highest additional level of phosphate (109µM). Both AZDY06 and AZFC22 grew well with higher specific growth rates, but with shorter growth periods, within f/2-Si medium spiked with urea than that within media spiked with nitrate. L1-Si medium with relatively high concentrations of trace metals was relatively favorable to both strains of A. poporum tested here. No obvious change within the toxin profile occurred in all cultures of both strains under the various nutrient conditions, although trace amounts of some suspicious derivatives of AZA2 occurred in some cultures. AZA2 cell quotas within both strains significantly (p<0.05) increased at the stationary phase under lower additional phosphate (0 and 3.6µM). Significant differences were not found within AZA2 cell quotas in cultures with additional nitrate ranging from 0 to 2647µM. The highest AZA2 cell quota and maximum AZA2 quantity per culture volume occurred in batch culture at the stationary phase under phosphate concentrations at 3.6µM. Neither A. poporum strain exhibited significant changes in AZA2 cell quotas within f/2-Si media spiked with urea or nitrate as nitrogen sources. The AZA2 cell quota of strain AZDY06 also did not change remarkably within f/2-Si, L1-Si, and K-Si media, however the AZA2 cell quota of strain AZFC22 within L1-Si medium was significantly (p<0.05) higher than that within f/2-Si medium.

Determination of multiple toxins in whelk and clam samples collected from the Chukchi and Bering seas.

Buccinidae whelk Neptunea varicifera (Dall), Cardiidae clam Serripes laperousii (Deshayes), and two unknown species of whelk and clam were collected from the Arctic Chukchi Sea and sub-Arctic Bering Sea in July 2014. In this study, the mollusk samples were analyzed by different liquid chromatography-tandem quadrupole mass spectrometry (LC-MS/MS) methods for multiple shellfish toxins, including okadaic acid (OA), pectenotoxin (PTX), yessotoxin (YTX), azaspiracid (AZA), cyclic imines (CI), and saxitoxin (STX) groups. PTX2 (≈2.0 µg kg(-1) whole tissues) was detected exclusively in the clam S. laperousii collected from the Chukchi Sea. OA and dinophysistoxin-1 (DTX1) were restricted to mollusk samples collected from the Bering Sea, and OA was the dominant component of the whelk N. varicifera (63 µg kg(-1) digestive gland) and an unknown species of whelk (6.8 µg kg(-1) digestive gland). Spirolide-1 (SPX1) was confirmed in most samples except for the whelk N. varicifera collected from the Bering Sea. The highest content of SPX1 (≈18.5 µg kg(-1) digestive gland) occurred in the whelk N. varicifera collected from the Chukchi Sea, along with the suspected presence of SPX-C, SPX-D and didesMe-SPX-C. YTX, as well as its derivatives 45-OH-YTX and 45,46,47-Trinor-YTX, were found in all samples, with the highest YTX content (66 µg kg(-1) digestive gland) present in the whelk N. varicifera collected from the Chukchi Sea. Interestingly, STX and dcSTX were measured only in the whelk N. varicifera and unknown species of clam collected from the Chukchi Sea. No AZA-group toxins, gymnodimine (GYM), or pinnatoxin G were found in any samples analyzed. Results demonstrated that the mollusk samples were contaminated by multiple shellfish toxins in the Chukchi and Bering seas. This study highlights the need to monitor potentially toxic microalgae in the Arctic and sub-Arctic regions, as well as species of mollusk that may be included in future commercial or subsistence harvests.