Value of Multimodal Ultrasound Combined with BRAF Gene in Evaluating Cervical Lymph Node Metastasis of Papillary Thyroid Microcarcinoma.

OBJECTIVE: The aim of the work described here was to explore the predictive value of multimodal ultrasound combined with the BRAF gene in cervical lymph node metastasis (CLNM) of papillary thyroid microcarcinoma (PTMC). METHODS: One hundred six patients (114 lesions) with PTMC confirmed by surgery and pathology at Yantai Yuhuangding Hospital from July 2021 to August 2022 were analyzed retrospectively. Routine ultrasound, contrast-enhanced ultrasound, shear wave elastography examination and BRAF V600E gene detection were performed before surgery. Patients were divided into two groups on the basis of post-operative pathology: non-metastasis group and metastasis group. Univariate and multivariate analyses were used to analyze the risk factors of cervical lymph node metastasis in PTMC. RESULTS: Univariate analysis revealed that there were significant differences in gender, high echo in lesions, enhancement level, peak intensity (PI) and average modulus of elasticity (Eavg) between the two groups (p < 0.05), but there was no significant difference in BRAF gene mutation (p = 0.855). Multivariate analysis revealed that male gender, microcalcification and hyper- or iso-enhancing parametric increased the risk of CLNM in PTMC (p < 0.05), and that sensitivity (92.3%) and accuracy (73.9%) were higher for combined diagnosis than for single diagnosis; the differences were statistically significant (p < 0.05). CONCLUSION: Male gender, microcalcification and hyper- or iso-enhancing parametrics of CEUS are independent risk factors for CLNM in PTMC patients. Combined diagnosis is more effective.

Extrachromosomal circular DNA landscape of breast cancer with lymph node metastasis.

Breast cancer (BC) is a complex disease with diverse manifestations, often resulting in lymph node metastasis (LNM) and impacting patient prognosis. Extrachromosomal circular DNA (eccDNA) has emerged as a key player in tumorigenesis, yet its contribution to BC LNM remains elusive. Here, we examined primary tumors and matched LNM tissues from 19 BC patients using the Circle-Seq method. We identified a median count of 44,682 eccDNA in primary tumor tissues and 38,057 in their paired LNM tissues. Furthermore, a ladder-like size distribution is observed in both primary tumor and LNM tissues. Meanwhile, similar repeat sequence distribution and GC content are identified from both primary tissue and LNM tissues. Finally, we found that eccDNA from both groups are flanked with palindromic trinucleotide motifs. These observations indicate that eccDNA of primary tumor and LNM tissues are from similar chromosomal origins. However, a subset of miRNA-associated eccDNA displayed selective enrichment in metastatic lesions, such as miR-6730 and miR-548AA1 genes. This observation implicates the function of miRNA-related eccDNA in the metastatic cascade. Our study uncovers the potential significance of these unique eccDNA molecules, shedding light on their role in cancer metastasis.

Ultrasound-guided thrombin injection in the management of brachial artery pseudoaneurysm with median nerve compression.

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Coated cysteamine, a potential feed additive for ruminants - An updated review.

For sustainable development, better performance, and less gas pollution during rumen fermentation, there is a need to find a green and safe feed additive for ruminants. Cysteamine (CS) is a biological compound naturally produced in mammalian cells. It is widely used as a growth promoter in ruminants because of its ability to control hormone secretions. It mainly controls the circulating concentration of somatostatin and enhances growth hormone production, leading to improved growth performance. CS modulates the rumen fermentation process in a way beneficial for the animals and environment, leading to less methane production and nutrients loss. Another beneficial effect of using CS is that it improves the availability of nutrients to the animals and enhances their absorption. CS also works as an antioxidant and protects the cells from oxidative damage. In addition, CS has no adverse effects on bacterial and fungal alpha diversity in ruminants. Dietary supplementation of CS enhances the population of beneficial microorganisms. Still, no data is available on the use of CS on reproductive performance in ruminants, so there is a need to evaluate the effects of using CS in breeding animals for an extended period. In this review, the action mode of CS was updated according to recently published data to highlight the beneficial effects of using CS in ruminants.

Evaluation of deproteinised bovine bone matrix combined with absorbable biofilm for the preservation of extraction sites of mandibular impacted wisdom teeth.

BACKGROUND: Bone defects and deep periodontal pockets often exist distal to the second molar after mandibular third molar extraction, seriously threatening the periodontal health of the second molar. OBJECTIVE: To evaluate the effect of socket preservation with bone substitute materials on alveolar bone resorption and prevention of the distal periodontal defect of the adjacent tooth after mandibular impacted third molar extraction compared with natural healing. METHODS: Ninety-nine patients with mandibular impacted teeth, treated in our hospital from January 2018 to December 2020, were randomly divided into the control and experimental groups. The experimental group underwent minimally invasive tooth extraction and socket preservation using the deproteinised bovine bone mineral, Bio-Oss and the bioabsorbable collagen membrane, Bio-Gide. The control group healed naturally after minimally invasive tooth extraction. The alveolar ridge dimension of the extraction sites, the probing depth, tooth mobility and gingival index on the distal aspect of the mandibular second molars were examined and recorded before and six months after the operations. RESULTS: There was a significant difference between the experimental group and the control group in the alveolar bone width (P< 0.05) and height (P< 0.05) before and after surgery. The probing depth of the extraction sites in both groups was reduced. CONCLUSION: Using Bio-Oss and Bio-Gide to preserve extraction sites of impacted teeth can promote recovery more effectively than natural healing on the height of the distal alveolar bone and the width of the alveolar crest of the second molar and thus improve the periodontal status of the adjacent second molar.

The impact of COVID-19 on a Southern Chinese cohort with neuromyelitis optica spectrum disorders.

BACKGROUND: There are few studies on risk factors for coronavirus disease 2019 (COVID-19) infection in patients with Neuromyelitis Optica Spectrum Disorders (NMOSD). The relationship between NMOSD relapse and COVID-19 needs to be evaluated. The objective of our study is to identify the risk factors of COVID-19 infection and NMOSD relapse among NMOSD patients with COVID-19. METHOD: A total of 379 NMOSD patients registered in a NMOSD database were included in this case-control study after the end of the COVID-19 quarantine and restriction policies on December 6, 2022 in China. Data were obtained from the database. Additional information was obtained by questionnaires and the Neurology out-patient clinic. The clinical characteristics of NMOSD patients with COVID-19 were described. Risk factors associated with COVID-19 infection and outcome among patients with NMOSD were analyzed. Risk factors associated with relapse in NMOSD patients with COVID-19 were also identified. RESULTS: 239 (63.1%) NMOSD patients were infected with COVID-19. Patients with NMOSD who were infected with COVID-19, in comparison to those without COVID-19, were younger at the time of interview (median [IQR] age: 43.00 [32.00-55.00] vs 49.50 [35.25-56.00] years, P = 0.029), younger at NMOSD onset (median [IQR] age: 38.00 [27.00-51.00] vs 45.00 [32.00-52.75] years, P = 0.013), had abnormal visual evoked potentials before infection (73.4% vs 54.3% P = 0.029), had lower baseline Activities of Daily Living Scale (ADL) scores (median [IQR] ADL: 14.00 [14.00-16.00] vs 14.00 [14.00-19.00], P = 0.014) or lower baseline modified Rankin Scale (mRS) scores (1.12±0.749 vs 1.33±0.991, P = 0.037), and were less frequently treated with more than 10 mg prednisone or 8 mg methylprednisolone (25.0% vs 36.0%,p = 0.026). All 9 NMOSD patients who had symptomatic cerebral syndrome developed moderate/severe COVID-19. A higher percentage of patients with moderate/severe COVID-19 experienced more than one core clinical NMOSD symptoms (61.5% vs 55.1%, p = 0.044), compared to patients with mild COVID-19. Higher risk of NMOSD relapse among NMOSD patients with COVID-19 was associated with higher Expanded Disability Status Scale (EDSS) scores (median[IQR] EDSS: 2.00 [1.00-3.00] vs 1.50 [1.00-2.25], P = 0.037) and drug treatments disruption (21.6% vs 5.0% P<0.001). CONCLUSIONS: NMOSD patients with younger age, lower baseline ADL or mRS had higher incidence of being diagnosed with COVID-19 during pandemic. Glucocorticoid use may decrease the risk of COVID-19. NMOSD patients with symptomatic cerebral syndrome before the COVID-19 pandemic are associated with worse COVID-19 outcomes. Drug treatment disruption may result in relapse among NMOSD patients with COVID-19.

Rural areas development and pro-environmental behavior in China: implications for rural development and planning.

This research adds to the body of knowledge by examining the asymmetric link between rural development and pro-environmental behavior in rural China. Rural development is assessed via rural income and digital financial inclusion. We use linear and nonlinear autoregressive distributed lag (ARDL) models to assess short- and long-term effects. The linear analysis suggests that a rise in rural income and digital financial inclusion encourages long-term access to clean fuels and technologies (CFT) in rural areas, while they do not have any significant impact in the short run. On the other side, the nonlinear framework illustrates that a positive shock in rural income and digital financial inclusion encourages long-term access to CFT in rural areas. A negative shock in rural income reduces long-term access to CFT in rural areas, and a negative shock in digital financial inclusion does not significantly impact access to CFT. However, in the short run, only the estimates of rural income are significant, while the estimates attached to digital financial inclusion are insignificant, implying that a rise in rural income increases access to CFT and a fall in rural income reduces access to CFT. Thus, government should encourage collaboration between private sector and civil society organizations to promote sustainable rural development and pro-environmental behavior.

Involvement of Glutamate Cysteine Ligase Genes in Tolerance to Emamectin Benzoate in Spodoptera frugiperda and Their Putative Regulatory Mechanisms.

As the rate-limiting enzyme in de novo Glutathione (GSH) biosynthesis, the mammalian glutamate cysteine ligase (Gcl) catalytic (Gclc) and modifier (Gclm) subunits are regulated at multiple levels, whereas the function and regulatory mechanism of insect Gcl remain to be explored. In this study, we identified and characterized SfGclc and SfGclm in Spodoptera frugiperda. SfGclc and SfGclm were highly expressed in the hindgut and relatively less expressed in other tissues. The exposure of the third instar larvae to LC30 of emamectin benzoate (EMB) significantly reduced the GSH content with a concomitant upregulation of SfGclc and SfGclm. Further in vivo pretreatment with L-BSO, the Gcl inhibitor, increased the susceptibility of S. frugiperda to EMB. Consistently, overexpression of SfGclc and SfGclm increased the Sf9 cell viability under EMB treatment. Finally, both RNAi and the dual-luciferase reporter assay in Sf9 cells revealed that SfGclc is regulated by transcription factor CncC. These data provide insights into the function and regulatory mechanism of insect Gcl, and they imply that disruption of the redox homeostasis might be a practical strategy to enhance the insecticidal activity of EMB and other insecticides.

Photoexcited cryptochromes interact with ADA2b and SMC5 to promote the repair of DNA double-strand breaks in Arabidopsis.

Cryptochromes (CRYs) act as blue-light photoreceptors that regulate development and circadian rhythms in plants and animals and as navigating magnetoreceptors in migratory birds. DNA double-strand breaks (DSBs) are the most serious type of DNA damage and threaten genome stability in all organisms. Although CRYs have been shown to respond to DNA damage, whether and how they participate in DSB repair is not well understood. Here we report that Arabidopsis CRYs promote the repair of DSBs through direct interactions with ADA2b and SMC5 in a blue-light-dependent manner to enhance their interaction. Mutations in CRYs and in ADA2b lead to similar enhanced DNA damage accumulation. In response to DNA damage, CRYs are localized at DSBs, and the recruitment of SMC5 to DSBs is dependent on CRYs. These results suggest that CRY-enhanced ADA2b-SMC5 interaction promotes ADA2b-mediated recruitment of SMC5 to DSBs, leading to DSB repair.

GC­MSC­derived circ_0024107 promotes gastric cancer cell lymphatic metastasis via fatty acid oxidation metabolic reprogramming mediated by the miR­5572/6855­5p/CPT1A axis.

Gastric cancer tissue­derived mesenchymal stem cells (GC­MSCs) play a critical role in facilitating gastric cancer metastasis. Recently, circular RNAs (circRNAs) and metabolic reprogramming have been found to be extensively involved in the malignant progression of tumors, including gastric cancer. However, the biological role and potential mechanisms of GC­MSC­derived circRNAs in metabolic reprogramming remain elusive. Herein, the expression profiles of circRNAs and mRNAs were compared between GC­MSCs and bone marrow­derived MSCs (BM­MSCs) using microarray analysis. circ_0024107 was identified to mediate GC­MSCs to promote gastric cancer lymphatic metastasis by inducing fatty acid oxidation (FAO) metabolic reprogramming. Mechanistically, circ_0024107 served as a sponge of miR­5572 and miR­6855­5p to elicit the FAO metabolic reprograming of GC­MSCs by upregulating carnitine palmitoyltransferase 1A (CPT1A). In addition, GC­MSCs promoted metastasis which was dependent on the induction of FAO in gastric cancer cells mediated by circ_0024107. The circ_0024107/miR­5572/6855­5p/CPT1A axis was deregulated in gastric cancer tissues and GC­MSCs, and was associated with lymph node metastasis and the prognosis of patients with gastric cancer. Taken together, the findings of the present study suggest the crucial role of FAO metabolic reprogramming mediated by GC­MSC­derived circ_0024107 in synergistically promoting gastric cancer lymphatic metastasis via miR­5572/6855­5p­CPT1A signaling; this suggests that circ_0024107 may be an attractive target for gastric cancer intervention.

Genome-wide identification and oxacillinase OXA distribution characteristics of Acinetobacter spp. based on a global database.

Objective: To use genomic analysis to identify Acinetobacter spp. and to explore the distribution characteristics of ß-lactamase oxallicinases (blaOXA) among Acinetobacter species globally. Methods: Genomes of global Acinetobacter spp. were downloaded from GenBank using Aspera batch. After quality check using CheckM and QUAST software, the genomes were annotated using Prokka software to investigate the distribution of blaOXAs across Acinetobacter spp.; a phylogenetic tree was constructed to explore the evolutionary relationship among the blaOXA genes in Acinetobacter spp. Average-nucleotide identification (ANI) was performed to re-type the Acinetobacter spp. BLASTN comparison analysis was implemented to determine the sequence type (ST) of Acinetobacter baumannii strain. Results: A total of 7,853 genomes were downloaded, of which only 6,639 were left for further analysis after quality check. Among them, 282 blaOXA variants were identified from the genomes of 5,893 Acinetobacter spp.; blaOXA-23 (n = 3,168, 53.8%) and blaOXA-66 (2,630, 44.6%) were the most frequent blaOXAs, accounting for 52.6% (3,489/6639), and the co-carriage of blaOXA-23 and blaOXA-66 was seen in 2223 (37.7%) strains. The 282 blaOXA variants were divided into 27 clusters according to the phylogenetic tree. The biggest clade was blaOXA-51-family carbapenem-hydrolyzing enzymes composed of 108 blaOXA variants. Overall, 4,923 A. baumannii were identified out of the 6,639 Acinetobacter spp. strains and 291 distinct STs were identified among the 4,904 blaOXA-carrying A. baumannii. The most prevalent ST was ST2 (n = 3,023, 61.6%) followed by ST1 (n = 228, 4.6%). Conclusion: OXA-like carbapenemases were the main blaOXA-type ß-lactamase spread widely across Acinetobacter spp. Both blaOXA-23 and blaOXA-66 were the predominant blaOXAs, among all A. baumannii strains, with ST2 (belonging to CC2) being the main clone disseminated globally.

Gastric cancer cell-originated small extracellular vesicle induces metabolic reprogramming of BM-MSCs through ERK-PPARγ-CPT1A signaling to potentiate lymphatic metastasis.

Tumor microenvironment and metabolic reprogramming are critical for tumor metastasis. Bone marrow-derived mesenchymal stem cells (BM-MSCs) are widely involved in the formation of tumor microenvironment and present oncogenic phenotypes to facilitate lymph node metastasis (LNM) in response to small extracellular vesicles (sEV) released by gastric cancer (GC) cells. However, whether metabolic reprograming mediates transformation of BM-MSCs remains elusive. Herein, we revealed that the capacity of LNM-GC-sEV educating BM-MSCs was positively correlated with the LNM capacity of GC cells themselves. Fatty acid oxidation (FAO) metabolic reprogramming was indispensable for this process. Mechanistically, CD44 was identified as a critical cargo for LNM-GC-sEV enhancing FAO via ERK/PPARγ/CPT1A signaling. ATP was shown to activate STAT3 and NF-κB signaling to induce IL-8 and STC1 secretion by BM-MSCs, thereby in turn facilitating GC cells metastasis and increasing CD44 levels in GC cells and sEV to form a persistent positive feedback loop between GC cells and BM-MSCs. The critical molecules were abnormally expressed in GC tissues, sera and stroma, and correlated with the prognosis and LNM of GC patients. Together, our findings uncover the role of metabolic reprogramming mediated BM-MSCs education by LNM-GC-sEV, which presents a novel insight into the mechanism underlying LNM and provides candidate targets for GC detection and therapy.

A case report of diagnosis of liver metastasis of medullary thyroid carcinoma by multimodal ultrasound.

Medullary thyroid carcinoma is a rare malignant neuroendocrine tumor. Distant metastasis is difficult to detect early. It is most common in lung, liver, bone and brain. This case was reported as liver metastasis of medullary thyroid carcinoma in an elderly woman, but routine ultrasound findings were atypical. After a series of relevant imaging examinations, contrast-enhanced ultrasound and ultrasound-guided puncture biopsy were used to confirm the nature of the intrahepatic lesions. Therefore, we believe that multimodal ultrasound is of great value in the diagnosis of liver metastasis of medullary thyroid carcinoma.

Recent advances in fluorescent probes of peroxynitrite: Structural, strategies and biological applications.

Peroxynitrite (ONOO-), owing to its high oxidative and nitrating stress, is associated with several physiological processes in addition to various pathological processes, including those related to neurodegenerative diseases and cancer. Detection of ONOO- at the cellular level is of great significance to understand its pathogenesis. To this end, a variety of fluorescent probes based on small molecules and nanoparticles (NPs) have been engineered and applied as excellent tools for imaging of ONOO- in cells as well as in their diverse biological applications. In this review, we highlight representative cases of fluorescent probes based on recognition mechanism and emphasize their response type (ratiometric, two-photon, long-wavelength/near-infrared, and targeting) in ONOO- detection in the last five years. We further discuss their design strategy, sensing mechanism, and application in bio-imaging and describe NP-based probes according to diverse nanoplatforms.

Hsa_circ_0073453 modulates IL-8 secretion by GC-MSCs to promote gastric cancer progression by sponging miR-146a-5p.

Gastric cancer associated mesenchymal stem cells (GC-MSCs) have been demonstrated to promote gastric cancer progression in a paracrine manner. IL-8 is highly secreted by GC-MSCs and is crucial for their oncogenic function. However, the mechanism underlying the modulation of IL-8 secretion by GC-MSCs has not been well elucidated. In this study, Shbio-human ceRNA array was used to identify dysregulated mRNAs and circRNAs between GC-MSCs and bone marrow derived mesenchymal stem cells (BM-MSCs). IL-8 was validated to be a critical paracrine cytokine for GC-MSCs promoting migration and invasion of gastric cancer cells. circ_0073453 was identified as a novel GC-MSC-derived circRNA which acted as a sponge of miR-146a-5p, thus increasing IL-8 expression and secretion to promote gastric cancer cell metastasis. Furthermore, circ_0073453 modulated IL-8 secretion by GC-MSCs to enhance gastric cancer cells PD-L1 expression to resist cytotoxic CD8+ T cell-killing. circ_0073453/miR-146a-5p/IL-8 axis was deregulated in gastric cancer tissues and associated with prognosis depending on MSC abundance in cancer tissues. Taken together, our findings suggest that circ_0073453/miR-146a-5p/IL-8 axis is critical for GC-MSCs promoting gastric cancer progression. Hence, hsa_circ_0073453 may be a potential therapeutic target for gastric cancer.

Value of clinical features combined with multimodal ultrasound in predicting lymph node metastasis in cervical central area of papillary thyroid carcinoma.

OBJECTIVE: To explore the clinical features, multimodal ultrasound features and multimodal ultrasound imaging features in predicting lymph node metastasis in the central cervical region of papillary thyroid carcinoma. METHODS: A total of 129 patients with papillary thyroid carcinoma (PTC) confirmed by pathology were selected from our hospital from September 2020 to December 2022. According to the pathological results of cervical central lymph nodes, these patients were divided into metastatic group and non-metastatic group. Patients were randomly sampled and divided into training group (n = 90) and verification group (n = 39) according to the ratio of 7:3. The independent risk factors for central lymph node metastasis (CLNM) were determined by least absolute shrinkage and selection operator and multivariate logistic regression. Based on independent risk factors to build a prediction model, select the best diagnostic effectiveness of the prediction model sketch line chart, and finally, the line chart calibration and clinical benefits were evaluated. RESULTS: A total of 8, 11 and 17 features were selected from conventional ultrasound images, shear wave elastography (SWE) images and contrast-enhanced ultrasound (CEUS) images to construct the Radscore of conventional ultrasound, SWE and CEUS, respectively. After univariate and multivariate logistic regression analysis, male, multifocal, encapsulation, iso-high enhancement and multimodal ultrasound imaging score were independent risk factors for cervical CLNM in PTC patients (p < 0.05). Based on independent risk factors, a clinical combined with multimodal ultrasound feature model was constructed, and multimodal ultrasound Radscore were added to the clinical combined with multimodal ultrasound feature model to form a joint prediction model. In the training group, the diagnostic efficacy of combined model (AUC = 0.934) was better than that of clinical combined with multimodal ultrasound feature model (AUC = 0.841) and multimodal ultrasound radiomics model (AUC = 0.829). In training group and validation group, calibration curves show that the joint model has good predictive ability for cervical CLNM of PTC patients; The decision curve shows that most of the net benefits of the nematic chart are higher than those of clinical + multimodal ultrasound feature model and multimodal ultrasound radiomics model within a reasonable risk threshold range. CONCLUSION: Male, multifocal, capsular invasion and iso-high enhancement are independent risk factors of CLNM in PTC patients, and the clinical plus multimodal ultrasound model based on these four factors has good diagnostic efficiency. The joint prediction model after adding multimodal ultrasound Radscore to clinical and multimodal ultrasound features has the best diagnostic efficiency, high sensitivity and specificity, which is expected to provide objective basis for accurately formulating individualized treatment plans and evaluating prognosis.

Genomic and clinical characteristics of carbapenem-resistant Enterobacter cloacae complex isolates collected in a Chinese tertiary hospital during 2013-2021.

Objective: To analyze the molecular epidemiology of carbapenem-resistant Enterobacter cloacae complex (CREC) by whole-genome sequencing and to explore its clinical characteristics. Methods: Enterobacter cloacae complex isolates collected in a tertiary hospital during 2013-2021 were subjected to whole-genome sequencing to determine the distribution of antimicrobial resistance genes (ARGs), sequence types (STs), and plasmid replicons. A phylogenetic tree of the CREC strains was constructed based on the whole-genome sequences to analyze their relationships. Clinical patient information was collected for risk factor analysis. Results: Among the 51 CREC strains collected, blaNDM-1 (n = 42, 82.4%) was the main carbapenem-hydrolyzing ß-lactamase (CHßL), followed by blaIMP-4 (n = 11, 21.6%). Several other extended-spectrum ß-lactamase-encoding genes were also identified, with blaSHV-12 (n = 30, 58.8%) and blaTEM-1B (n = 24, 47.1%) being the predominant ones. Multi-locus sequence typing revealed 25 distinct STs, and ST418 (n = 12, 23.5%) was the predominant clone. Plasmid analysis identified 15 types of plasmid replicons, among which IncHI2 (n = 33, 64.7%) and IncHI2A (n = 33, 64.7%) were the main ones. Risk factor analysis showed that intensive care unit (ICU) admission, autoimmune disease, pulmonary infection, and previous corticosteroid use within 1 month were major risk factors for acquiring CREC. Logistic regression analysis showed that ICU admission was an independent risk factor for CREC acquisition and was closely related with acquiring infection by CREC with ST418. Conclusion: BlaNDM-1 and blaIMP-4 were the predominant carbapenem resistance genes. ST418 carrying BlaNDM-1 not only was the main clone, but also circulated in the ICU of our hospital during 2019-2021, which highlights the necessity for surveillance of this strain in the ICU. Furthermore, patients with risk factors for CREC acquisition, including ICU admission, autoimmune disease, pulmonary infection, and previous corticosteroid use within 1 month, need to be closely monitored for CREC infection.

The distribution characteristics of global blaOXA-carrying Klebsiella pneumoniae.

OBJECTIVE: To analyze the distribution of blaOXA among global Klebsiella pneumoniae and the characteristics of blaOXA-carrying K. pneumoniae. MATERIALS AND METHODS: The genomes of global K. pneumoniae were downloaded from NCBI by Aspera software. After quality check, the distribution of blaOXA among the qualified genomes was investigated by annotation with the resistant determinant database. The phylogenetic tree was constructed for the blaOXA variants based on the single nucleotide polymorphism (SNP) to explore the evolutionary relationship between these variants. The MLST (multi-locus sequence type) website and blastn tools were utilized to determine the sequence types (STs) of these blaOXA-carrying strains. and sample resource, isolation country, date and host were extracted by perl program for analyzing the characteristics of these strains. RESULTS: A total of 12,356 K. pneumoniae genomes were downloaded and 11,429 ones were qualified. Among them, 4386 strains were found to carry 5610 blaOXA variants which belonged to 27 varieties of blaOXAs, blaOXA-1 (n = 2891, 51.5%) and blaOXA-9 (n = 969, 17.3%) were the most prevalent blaOXA variants, followed by blaOXA-48 (n = 800, 14.3%) and blaOXA-232 (n = 480, 8.6%). The phylogenetic tree displayed 8 clades, three of them were composed of carbapenem-hydrolyzing oxacillinase (CHO). Totally, 300 distinct STs were identified among 4386 strains with ST11 (n = 477, 10.9%) being the most predominant one followed by ST258 (n = 410, 9.4%). Homo sapiens (2696/4386, 61.5%) was the main host for blaOXA-carrying K. pneumoniae isolates. The blaOXA-9-carrying K. pneumoniae strains were mostly found in the United States and blaOXA-48-carrying K. pneumoniae strains were mainly distributed in Europe and Asia. CONCLUSION: Among the global K. pneumoniae, numerous blaOXA variants were identified with blaOXA-1, blaOXA-9, blaOXA-48 and blaOXA-232 being the most prevalent ones, indicating that blaOXA rapidly evolved under the selective pressure of antimicrobial agents. ST11 and ST258 were the main clones for blaOXA-carrying K. pneumoniae.

Housekeeping gene stability in Pesudomonas aeruginosa PAO1 under the pressure of commonly used antibiotics in molecular microbiology assays.

Pseudomonas aeruginosa is an opportunistic human pathogen notorious for its remarkable capacity of multi-drug resistance, and has become one of the most important model bacteria in clinical bacteriology research. Quantitative real-time PCR is a reliable method widely used in gene expression analysis, for which the selection of a set of appropriate housekeeping genes is a key prerequisite for the accuracy of the results. However, it is easy to overlook that the expression level of housekeeping gene may vary in different conditions, especially in the condition of molecular microbiology assays, where tested strains are generally cultured under the pre-set antibiotic selection pressures, and how this affects the stability of commonly used housekeeping genes remains unclear. In this study, the expression stability of ten classic housekeeping genes (algD, gyrA, anr, nadB, recA, fabD, proC, ampC, rpoS, and rpsL) under the pressure of eight laboratory commonly used antibiotics (kanamycin, gentamycin, tetracycline, chloramphenicol, hygromycin B, apramycin, tellurite, and zeocin) were tested. Results showed that the stability of housekeeping gene expression was indeed affected by the types of antibiotics added, and of course the best reference gene set varied for different antibiotics. This study provides a comprehensive summary of the effects of laboratory antibiotics on the stability of housekeeping genes in P. aeruginosa, highlighting the necessity to select housekeeping genes according to the type of antibiotics used in the initial stage of experiment.