Since first isolate of Candida ( C. ) auris was recognized in 2018 in China, over 60 cases of C. auris colonization or infection have been reported up to 2022, with at least two distinct genetic clade. Most isolates from South Asian clade and South African clade were sensitive and resistant to fluconazole, respectively. Here, we report an isolate of fluconazole-resistant South Asian clade C. auris (PUSH159) from blood. The efflux pump genes MDR1 , CDR1 , CDR4 , SNQ2, and ARB1 , and ergosterol biosynthesis gene ERG11 were detected in strain PUSH159. Single nucleotide polymorphism (SNP) analysis revealed that strain PUSH159 carried multiple mutations in drug resistance-associated genes, including a hot-spot mutation (Y132F) in ERG11 , missense mutation of E709D in CDR1 , and missense mutations of K52N and E464K in SNQ2. Our findings support the genetic diversity and differences in antifungal susceptibility within the South Asian clade, and provide early warning for the emergence of fluconazole-resistant South Asian clade isolates in China.

Respiratory infections caused by influenza A viruses (IAVs) pose a global threat annually, leading to significant mortality in severe cases. IAVs are classified into various subtypes, and a combination of multiplex reverse transcription-multiple cross displacement amplification (mRT-MCDA) and lateral flow biosensor (LFB) has been developed to detect three subtypes (H1N1, H3N2, and H7N9) that have been frequently observed in recent years. This technology simultaneously reverse transcribes and amplifies two target genes, hemagglutinin (HA) and neuraminidase (NA), at a constant temperature of 63 °C in just 40 min. The final results can be read using a dual-channel LFB. In practice, the entire process-from sample collection, RNA extraction, and mRT-MCDA amplification to result interpretation-can be completed in less than 1 h. This method is named "multiplex reverse transcription-multiple cross displacement amplification-lateral flow biosensor" (IAVs-mRT-MCDA-LFB) technology. The IAVs-mRT-MCDA-LFB technology demonstrated higher sensitivity compared to conventional reverse transcription-polymerase chain reaction (RT-PCR) and showed no cross-reactivity with other genes present in the samples. Therefore, the IAVs-mRT-MCDA-LFB technique developed in this study is a highly valuable molecular diagnostic tool for detecting IAV-subtypes due to its simplicity, time efficiency, cost-effectiveness, and high specificity and sensitivity.

ObjectiveHelicobacter pylori (H. pylori) plays a major role in causing and advancing gastrointestinal illnesses. Our aim is to analyze the unique makeup and functional changes in the gastric microbiota of patients with chronic non-atrophic gastritis (CNAG), regardless of the presence of H. pylori, and to determine the potential signaling pathways.MethodsWe performed metagenomic sequencing on gastric mucosa samples collected from 17 individuals with non-atrophic gastritis, comprising 6 cases were infected with H. pylori (H. pylori-infected case group) and 11 cases without (control group). The species composition was evaluated with DIAMOND software, and functional enrichment was assessed utilizing the Kyoto Encyclopedia of Genes and Genomes (KEGG) database. We analyzed antibiotic resistance patterns using the Comprehensive Antibiotic Resistance Database as a reference (CARD).ResultsThe presence of H. pylori colonization in CNAG patients was associated with increased diversity in the gastric microbiota. The Phylum Firmicutes was found to be less prevalent, while the Phylum Proteobacteria showed an increase. Functionally, pathways associated with metabolic pathways, including vitamins, auxiliaries, amino acid residue, carbon hydrate, and metabolic energy pathways, were enriched in CNAG patients with H. pylori infection. Additionally, antibiotic resistance genes correlated with antibiotic efflux pump were enriched.ConclusionsFrom a holistic genomic perspective, our findings offer fresh perspectives into the gastric microbiome among CNAG patients carrying H. pylori, which is valuable for future research on CNAG.

Hypertrophic cardiomyopathy (HCM) is a genetic myocardial disease, with an estimated incidence of 0.2%-6%, and is the main cause of sudden cardiac death (SCD) in young athletes. Left ventricular apical aneurysm (LVAA) is a rare subtype of HCM, accounting for about 5% of HCM patients, and has a higher incidence of cardiovascular adverse events. In cases of hypertrophic obstructive cardiomyopathy with LVAA (HOCM-LVAA) that do not respond adequately to optimized medical therapy, the echocardiography-guided percutaneous intra-myocardial septal radiofrequency ablation (PIMSRA, Liwen procedure) emerges as a promising and effective novel therapeutic approach. In this case report, we present for the first time a comprehensive application of echocardiographic techniques, including TTE, 2-D STE, and contrast enhancement, in the diagnosis, treatment, surgical guidance, and assessment of therapeutic outcomes in a case of HOCM-LVAA.

Helicobacter pylori (H. pylori) is a gram-negative, microaerobic bacterium that colonizes the gastric mucosa in about half of the world's population. H. pylori infection can lead to various diseases. Chronic infection by H. pylori exposes the gastric mucosa to bacterial components such as lipopolysaccharide (LPS), outer membrane vesicles (OMVs), and several toxic proteins. Infected with H. pylori activates the release of pro-inflammatory factors and triggers inflammatory responses that damage the gastric mucosa. As the only microorganism that permanently colonizes the human stomach, H. pylori can suppress host immunity to achieve long-term colonization. Toll-like receptors (TLRs) play a crucial role in T-cell activation, promoting innate immune responses and immune tolerance during H. pylori infection. Among the 10 TLRs found in humans, TLR2, TLR4, TLR5, and TLR9 have been thoroughly investigated in relation to H. pylori-linked immune regulation. In the present review, we provide a comprehensive analysis of the various mechanisms employed by different TLRs in the induction of immune tolerance upon H. pylori infection, which will contribute to the research of pathogenic mechanism of H. pylori.

Meningococcal disease caused by Neisseria meningitidis remains a major global public health concern. Serogroup A, B, C and W135 were the major disease-causing serogroups. It is vital to timely and efficiently detect and differentiate these four serogroups. Herein, we developed multiple cross displacement amplification-lateral flow biosensor (MCDA-LFB) assays targeting ctrA, sacB, siaD, siaD and synG gene respectively for detection and subtyping of four N. meningitidis serogroups. This assay utilizes LFB to detect FITC and biotin-labeled target amplicons produced by MCDA through double antibody sandwich principle, to allow sensitive and specific detection under a constant temperature. The detection limit was as low as 10 fg or 100 fg genomic DNA in pure cultures and 5.5 CFUs or 36 CFUs in spiked cerebrospinal fluid (CSF) specimens, which were overall 100 to 1000-fold more sensitive than conventional PCR. High specificity of these assays was also validated through type strains and clinical isolates, with no cross-reactions. MCDA-LFB testing procedure can be finished within 1 h. In conclusion, the N. meningitidis- and serogroup-MCDA-LFB assays established in this study are simple, rapid and efficient, providing valuable molecular methods for diagnosis and surveillance of meningococcal disease, especially in resource-limited regions and when specimen culture fails.

Background: Epidemiological studies showed that diabetes patients are more prone to developing cholelithiasis. Although composition of gut microbiota in type 2 diabetes or cholelithiasis have been studied respectively, the underlying role of gut microbiota in developing from diabetes to cholelithiasis remains unclear. By 16S rRNA gene sequencing, the gut microbial composition of 33 healthy subjects, 53 type 2 diabetes, 31 cholelithiasis and 32 type 2 diabetes complicated with cholelithiasis patients were studied. Results: Microbial diversity significantly decreased in type 2 diabetes complicated with cholelithiasis patients. In type 2 diabetes patients, phylum Proteobacteria class Gammaproteobacteria and order Lactobacillales were significantly increased. In cholelithiasis patients, phylum Bacteroidetes, class Bacteroidia order Bacteroidales family Bacteroidaceae and genus Bacteroides were significantly increased. There were also significant increases of phylum Proteobacteria, class Gammaproteobacteria order Lactobacillales family Lactobacillaceae and genus Lactobacillus in type 2 diabetes complicated with cholelithiasis patients accompanied by elevated serum triglyceride and total bile acids. Conclusions: The results show similar but more intricate gut microbiota dysbiosis in type 2 diabetes complicated with cholelithiasis compared with type 2 diabetes, which might partially explain the mechanism of type 2 diabetes as the risk factor of cholelithiasis from the perspective of gut microbiota.

Streptococcus pneumoniae is an important clinical pathogenic bacterium that is the primary cause of meningitis, septicemia and community-acquired pneumonia. The mortality rate of pneumococcal disease is high, especially in children younger than 5-years-old. Rapid and accurate detection of S.pneumoniae is critical for clinical diagnosis. A ply gene-based multiple cross displacement amplification (MCDA) assay, amplifying DNA under 65°C for 40 min, was established to detect S.pneumoniae. Antarctic thermal sensitive uracil-DNA-glycosylase (AUDG) was applied to prevent carryover contamination. A lateral flow biosensor (LFB) was used to indicate the MCDA results. The ply-MCDA assay could detect as low as 10 fg of S. pneumoniae DNA and 447 colony forming units (CFU)/mL of spiked sputum samples. The analytical sensitivity of the ply-MCDA assay to detect clinical specimens was 100 times higher than that of PCR. The specificity of the ply-MCDA assay was evaluated using 15 S.pneumoniae strains and 25 non-S. pneumoniae strains, which confirmed the high selectivity of the ply-MCDA assay for S.pneumoniae. The AUDG enzyme could effectively eliminate carryover contamination and thus prevented false-positive results. In conclusion, ply-AUDG-MCDA-LFB is a simple, rapid and accurate method to detect S.pneumoniae.

Background Helicobacter himalayensis was isolated from Marmota himalayana in the Qinghai-Tibet Plateau, China, and is a new non- H. pylori species, with unclear taxonomy, phylogeny, and pathogenicity . Results A comparative genomic analysis was performed between the H. himalayensis type strain 80(YS1) T and other the genomes of Helicobacter species present in the National Center for Biotechnology Information (NCBI) database to explore the molecular evolution and potential pathogenicity of H. himalayensis . H. himalayensis 80(YS1) T formed a clade with H. cinaedi and H. hepaticus that was phylogenetically distant from H. pylori . The H. himalayensis genome showed extensive collinearity with H. hepaticus and H. cinaedi . However, it also revealed a low degree of genome collinearity with H. pylori . The genome of 80(YS1) T comprised 1,829,936 bp, with a 39.89% GC content, a predicted genomic island, and 1769 genes. Comparatively, H. himalayensis has more genes for functions in “cell wall/membrane/envelope biogenesis” and “coenzyme transport and metabolism” sub-branches than the other compared helicobacters, and its genome contained 42 virulence factors genes, including that encoding cytolethal distending toxin (CDT). Conclusions We characterized the H. himalayensis 80(YS1) T genome, its phylogenetic position, and its potential pathogenicity. However, further understanding of the pathogenesis of this potentially pathogenic bacterium is required, which might help to manage H. himalayensis -induced diseases.

Background: Streptococcus pneumoniae is an important clinical pathogenic bacterium, which is the primary cause of meningitis, septicemia and community-acquired pneumonia. The mortality rate of pneumococcal disease was high, especially in children under 5 years of age. Rapid and accurate detection of S. pneumoniae is critical. Methods: A ply gene-based multiple cross displacement amplification (MCDA), which amplifies DNA under isothermal conditions (65 °C) for 40 min, was established for accurate and rapid detection of S. pneumoniae. Antarctic thermal sensitive uracil-DNA-glycosylase (AUDG) was applied for eliminating carryover contamination. Lateral flow biosensor (LFB) was used to indicate the amplification results. Results: The ply-MCDA assay can detect as little as 10 fg of S. pneumoniae DNA, as well as 447 CFU/mL of spiked sputum samples. The sensitivity of ply-MCDA assay in clinical samples was 100 times that of PCR. The specificity of MCDA primer targeting the ply gene was validated using 15 S. pneumoniae and 25 non-S. pneumoniae, suggesting that ply gene-based MCDA assay was highly selective for S. pneumoniae. Moreover, the ply-MCDA coupled with AUDG can effectively eliminate carryover contamination, and then prevent false-positive results. Conclusion: The ply-MCDA assay coupled with AUDG was a simple, rapid and accurate method in the diagnosis of S. pneumoniae infection.

Candida albicans is an opportunistic pathogenic yeast that predominantly causes invasive candidiasis. The conventional diagnosis of C. albicans infection depends on time-consuming, culture-based gold-standard methods. Here, a multiple cross displacement amplification (MCDA) assay, combined with a gold nanoparticle-based lateral flow biosensor (LFB) visualization method, was developed for the rapid detection of C. albicans. The internal transcribed spacer II, a region between 5.8 and 28 S fungal ribosomal DNA, is a C. albicans species-specific sequence that was used as the MCDA assay target. As an isothermal amplification method, the MCDA reaction with optimized conditions could be completed within only 40 min at a constant temperature (64°C). Then, the amplification reaction products could be visibly detected by a LFB without special equipment. The developed MCDA-LFB assay for C. albicans detection was a specific and accurate method, and could distinguish C. albicans from other pathogens. Just 200 fg of genomic DNA template from pure cultures of C. albicans could be detected using the MCDA-LFB method. The limit of detection (LOD) of the new method was more sensitive than that of both qPCR and loop-mediated isothermal amplification (LAMP). Of 240 clinical sputum samples, all of the C. albicans-positive (87/240) samples identified by the gold-standard method were successfully detected by the MCDA-LFB assay. Moreover, the true positive rate of the newly developed assay was not only higher than that of qPCR (100 vs. 86.2%), but also higher than that of LAMP (100 vs. 94.3%). Thus, the MCDA-LFB assay might be a simple, specific, and sensitive method for the rapid diagnosis of C. albicans in clinical samples.

Acinetobacter baumannii is a frequent cause of the nosocomial infections. Herein, a novel isothermal amplification technique, multiple cross displacement amplification (MCDA) is employed for detecting all A. baumannii strains and identifying the strains harboring blaOXA-23-like gene. The duplex MCDA assay, which targets the pgaD and blaOXA-23-like genes, could identify the A. baumannii isolates and differentiate these isolates harboring blaOXA-23-like gene. The disposable lateral flow biosensors (LFB) were used for analyzing the MCDA products. A total of sixty-eight isolates, include fifty-three A. baumannii strains and fifteen non-A. baumannii strains, were employed to optimize MCDA methods and determine the sensitivity, specificity and feasibility. The optimal reaction condition is found to be 63 °C within 1 h, with limit of detection at 100 fg templates per tube for pgaD and blaOXA-23-like genes in pure cultures. The specificity of this assay is 100%. Moreover, the practical application of the duplex MCDA-LFB assay was evaluated using clinical samples, and the results obtained from duplex MCDA-LFB method were consistent with conventional culture-based technique. In sum, the duplex MCDA-LFB assay appears to be a reliable, rapid and specific technique to detect all A. baumannii strains and identify these strains harboring blaOXA-23-like gene for appropriate antibiotic therapy.

Background Streptococcus pneumoniae (S. pneumoniae), as an important human pathogenic bacterium, causes high mortality and morbidity, especially among children younger than 5 years. Accurate and rapid detection of S. pneumoniae is critical important. Methods A ply gene-based multiple cross displacement amplification (MCDA), which amplifies DNA under isothermal conditions (65 °C) for 40 min, was established for accurate and rapid detection of S. pneumoniae. The amplicons were indicated using lateral flow biosensor (LFB). Results The MCDA assay could detect as little as 10 fg purified S. pneumoniae genomic DNA, and 447 CFU/mL spiked sputum samples with a sensitivity 100 times that of PCR. The specificity of MCDA primer targeting ply gene was validated using 15 S. pneumoniae strains and 25 non-S. pneumoniae strains, indicating that ply gene-based MCDA assay was highly selective for S. pneumoniae. Moreover, the MCDA assay coupled with antarctic thermal sensitive uracil-DNA-glycosylase (AUDG) can effectively eliminate the carryover contamination, and then preventing false-positive results. Conclusion MCDA assay in combination with AUDG was a simple, rapid, sensitive, specific and reliable method for the diagnosis of S. pneumoniae infection, which appears to be a valuable tool for clinical screening.

Two strains (NLN63T and NLN82) of Gram-stain-negative, oxidase- and catalase-positive, bacilli-shaped organisms were isolated from the faecal samples of two separate Rattus norvegicus in Baisha county of Hainan Province, Southern PR China. Phylogenetic analysis based on the near full-length 16S rRNA sequences revealed that strain NLN63T belongs to the genus Pelistega, having maximum similarity to Pelistega suis CCUG 64465T (97.1 %), Pelistega europaea CCUG 39967T (96.2 %) and Pelistega indica DSM 27484T (96.2 %), respectively. The phylogenomic tree built on 553 core genes from genomes of 20 species in the genus Pelistega and other adjacent genera further confirmed that strains NLN63T and NLN82 form a distinct subline and exhibit specific phylogenetic affinity with P. europaea CCUG 39967T. In digital DNA-DNA hybridization analyses, strain NLN63T showed low estimated DNA reassociation values (21.4-22.6 %) with the type strains of the species in the genus Pelistega. The DNA G+C contents of strains NLN63T and NLN82 were 37.3 and 37.1 mol%, respectively. Strain NLN63T had a unique MALDI-TOF MS profile, contained Q-8 as the major quinone and C16 : 0, summed feature 8 (C18 : 1  ω7c/C18 : 1  ω6c or both) and summed feature 3 (C16 : 1  ω7c/C16 : 1  ω6c or both) as the dominant fatty acids. Based upon these polyphasic characterization data obtained from the present study, a novel species of the genus Pelistega, Pelistega ratti sp. nov., is proposed with NLN63T (=GDMCC 1.1697T=JCM 33788T) as the type strain.

AIM:A high-risk strategy has been implemented for lipid-lowering therapy in the primary prevention of cardiovascular disease. However, atherosclerosis and cardiovascular events are common among individuals with low cardiovascular risk. This study aimed to determine whether the small dense low-density lipoprotein cholesterol (sdLDLC) level can predict carotid atherosclerosis progression and identify high-risk individuals. METHODS:Baseline sdLDLC and low-density lipoprotein cholesterol (LDLC) were measured in 808 participants from the Chinese Multi-provincial Cohort Study, aged 45-74 years. Adjusted relative risk was calculated using a modified Poisson regression model to assess the relationship between sdLDLC and 5-year atherosclerosis progression, as indicated by the progression, incidence, and multi-territorial extent of carotid plaque. RESULTS:The 5-year atherosclerosis progression increased significantly with increased sdLDLC. Baseline sdLDLC was significantly associated with the short-term risk of plaque progression after multivariable adjustment, even in participants with low LDLC or a 10-year estimated cardiovascular risk. sdLDLC predicted plaque progression (relative risk 2.05; 95% confidence interval 1.43-2.93) in participants with LDLC ＜130 mg/dL. Furthermore, participants with the highest sdLDLC but intermediate or low cardiovascular risk (accounting for 16% of the cohort) had double the risk of plaque progression, which was comparable to those with the same sdLDLC and high cardiovascular risk, relative to those with the lowest sdLDLC levels and low cardiovascular risk. CONCLUSIONS:sdLDLC is independently associated with the progression of carotid atherosclerosis, which may provide a basis for clinicians to reclassify individuals believed to be at low cardiovascular risk into the high-risk category, and those with high sdLDLC may benefit from more aggressive cholesterol-lowering treatment.

Pseudomonas aeruginosa causes nosocomial infections of burn patients and other immunocompromised individuals, but the conventional diagnosis of P. aeruginosa infection depends on time-consuming culture-based methods. Hence, a simple, fast, sensitive technique for detection of P. aeruginosa using multiple cross displacement amplification (MCDA) and gold nanoparticle-based lateral flow biosensors (LFB) was developed. By using this technique, the reaction could be completed at an optimized constant temperature (67°C) within only 40 min. The reaction product could be detected visually using an LFB, eliminating the need for special equipment. The P. aeruginosa-MCDA-LFB method was highly specific, and accurately distinguished P. aeruginosa from other pathogens. Just 10 fg of genomic DNA template (from pure culture) could be detected. The assay could also detect P. aeruginosa in clinical sputum samples and showed the same sensitivity and specificity as the reference (culture-biochemical) method. In the future, this rapid, simple and accurate P. aeruginosa-MCDA-LFB technique might be applied in clinical practice.

Klebsiella pneumoniae (K. pneumoniae) is a frequent pathogen causing nosocomial infections and outbreaks. We developed a multiple cross displacement amplification (MCDA) assay for the detection of K. pneumoniae, which can get the positive results within 40 minutes' isothermal amplification. Gold-nanoparticle lateral flow biosensor (LFB) and colorimetric indicators were used for the rapid readouts of MCDA amplification. The detection limit of this assay was 100 fg per reaction at 65°C, which was confirmed to be the optimal amplification temperature according to the real time turbidimeters. For specificity, all of the 30 clinical-source K. pneumoniae strains were positive for the MCDA, and all of the non-K. pneumoniae strains belonging to 31 different species were negative for this MCDA assay. To evaluate the practical applicability of this method, we assessed its detection limit for K. pneumoniae strains in sputum samples (24 CFU per reaction), and DNA templates of 100 sputum samples further underwent the MCDA-LFB tests. All of the sputum samples being positive for K. pneumoniae (30/100) with the culture method were successfully identified with the MCDA assay, the detection power of which was higher than that of polymerase chain reaction (PCR) (25/100). Thus, the MCDA test for K. pneumoniae combined with the gold nanoparticle LFB as the results readout scheme, are simple, specific, and sensitive methods for the rapid diagnosis of K. pneumoniae in clinical samples.

AIMS:To investigate the association of serum ferritin levels and ferritin level changes with the 10-year risk of new-onset type 2 diabetes mellitus (T2DM). METHODS:Among 2359 subjects without T2DM at baseline in 2002, 1956 subjects were re-examined in 2007, and 1660 subjects were invited to be re-examined in 2012. Serum ferritin (ng/ml) levels were measured by latex-enhanced turbidimetric immunoassay. Five-year serum ferritin changes were categorized into four groups using the median as the cut-off point. Multivariate logistic regression was performed to examine the independent association of serum ferritin levels and 5-year ferritin level changes with 10-year new-onset T2DM. RESULTS:At the 10-year follow-up, 205 (12.3%) subjects had developed new-onset T2DM. After adjusting for traditional risk factors and high-sensitivity C-reactive protein, 10-year new-onset T2DM risk was significantly increased in subjects in the highest tertile of baseline serum ferritin levels [odds ratio (OR)=1.80, 95% confidence interval (CI): 1.17-2.79] and in subjects with high serum ferritin levels in both 2002 and 2007 (OR=1.54, 95% CI: 1.01-2.34). After adjusting for baseline fasting blood glucose, the effect was attenuated and became borderline or non-significant. CONCLUSIONS:Serum ferritin levels and ferritin level changes were associated with 10-year new-onset T2DM risk in the Chinese population, whereas the independent effect awaits validation from studies with larger sample sizes.