A quantitative, high-throughput urease activity assay for comparison and rapid screening of ureolytic bacteria.

Urease is a dinickel enzyme commonly found in numerous organisms that catalyses the hydrolysis of urea into ammonia and carbon dioxide. The microbially induced carbonate precipitation (MICP) process mediated by urease-producing bacteria (UPB) can be used for many applications including, environmental bioremediation, soil improvement, healing of cracks in concrete, and sealing of rock joints. Despite the importance of urease and UPB in various applications, a quantitative, high-throughput assay for the comparison of urease activity in UPB and rapid screening of UPB from diverse environments is lacking. Herein, we reported a quantitative, 96-well plate assay for urease activity based on the Christensen's urea agar test. Using this assay, we compared urease activity of six bacterial strains (E. coli BL21, P. putida KT2440, P. aeruginosa PAO1, S. oneidensis MR-1, S. pasteurii DSM 33, and B. megaterium DSM 319) and showed that S. pasteurii DSM 33 exhibited the highest urease activity. We then applied this assay to quantify the inhibitory effect of calcium on urease activity of S. pasteurii DSM 33. No significant inhibition was observed in the presence of calcium at concentrations below 10 mM, while the urease activity decreased rapidly at higher concentrations. At a concentration higher than 200 mM, calcium completely inhibited urease activity under the tested conditions. We further applied this assay to screen for highly active UPB from a wastewater enrichment and identified a strain of S. pasteurii exhibiting a substantially higher urease activity than DSM 33. Taken together, we established a 96-well plate-based quantitative, high-throughput urease activity assay that can be used for comparison and rapid screening of UPB. As UPB and urease activity are of interest to environmental, civil, and medical researchers and practitioners, we envisage wide applications of the assay reported in this study.

Syndecan-1 suppresses cell growth and migration via blocking JAK1/STAT3 and Ras/Raf/MEK/ERK pathways in human colorectal carcinoma cells.

BACKGROUND: Syndecan-1 (SDC-1) is a crucial membrane proteoglycan, which is confirmed to participate in several tumor cell biological processes. However, the biological significance of SDC-1 in colorectal carcinoma is not yet clear. An objective of this study was to investigate the role of SDC-1 in colorectal carcinoma cells. METHODS: Expression of SDC-1 in colorectal carcinoma tissues was evaluated by Reverse transcription-quantitative real-time PCR (RT-qPCR) and western blot. After transfection with pcDNA3.1 or pc-SDC-1, the transfection efficiency was measured. Next, SW480, SW620 and LOVO cell viability, apoptosis, migration and adhesion were assessed to explore the effects of exogenous overexpressed SDC-1 on colorectal carcinoma. In addition, the influences of aberrant expressed SDC-1 in Janus kinase 1 (JAK1)/signal transducer and activator of transcription 3 (STAT3) and rat sarcoma virus (Ras)/rapidly accelerated fibrosarcoma (Raf)/mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinase (ERK) pathways were detected by western blot analysis. RESULTS: SDC-1 mRNA and protein levels were down-regulated in human colorectal carcinoma tissues. SDC-1 overexpression inhibited cell proliferation via suppressing CyclinD1 and c-Myc expression, meanwhile stimulated cell apoptosis via increasing the expression levels of B-cell lymphoma-2-associated x (Bax) and Cleaved-Caspase-3. Additionally, SDC-1 overexpression restrained cell migration via inhibiting the protein expression of matrix metallopeptidase 9 (MMP-9), and elicited cell adhesion through increasing intercellular cell adhesion molecule-1 (ICAM-1). Furthermore, SDC-1 overexpression suppressed JAK1/STAT3 and Ras/Raf/MEK/ERK-related protein levels. CONCLUSIONS: In general, the evidence from this study suggested that SDC-1 suppressed cell growth, migration through blocking JAK1/STAT3 and Ras/Raf/MEK/ERK pathways in human colorectal carcinoma cells.

Cytomegalovirus Pneumonia in Inflammatory Bowel Disease: Literature Review and Clinical Recommendations.

Aim: The objective was to elucidate the correlation between CMVP and immunosuppressive therapy in IBD patients, we hope this review could expand on the significance of CMV as an opportunistic pathogen and the potential impact on morbidity and mortality in IBD patients. Methods: Records and clinical trajectories linked to CMVP in IBD patients were extracted from the PubMed database, irrespective of language barriers. The reference lists incorporated in these studies were manually inspected. Conclusions were generated using straightforward descriptive analysis. Results: In total, 18 IBD patients, including Crohn's disease (CD, 67%) and Ulcerative Colitis (UC, 33%), affected by CMVP were identified from 17 published articles. A minority of these patients (17%) exhibited active disease, whereas the majority (83%) presented with quiescent disease. Fever (100%) and dyspnea (44%) emerged as the most prevalent clinical symptoms. All the patients had undergone immunosuppressive therapy. A significant proportion, up to 89%, had received thiopurine treatment prior to the CMVP diagnosis. Interestingly, none of the patients were subjected to biological therapy. Half of the patients manifested with Hemophagocytic Lymphohistiocytosis (HLH). Almost all patients (94%) were administered antiviral treatment and a substantial 83% experienced full recovery. Immunosuppressive agents were either tapered or discontinued altogether. A subset of patients, 17%, suffered fatal outcomes. Conclusion: Our findings underscore the need for heightened suspicion of CMVP in IBD patients who exhibit symptoms such as fever and dyspnea. During the COVID-19 pandemic, CMVP should be considered a potential differential diagnosis. It was observed that CMVP primarily transpires during CD remission. Azathioprine emerged as the predominant immunosuppressant linked to CMV reactivation. The prompt application of effective antiviral therapy can substantially enhance patient outcomes. CMV vaccine might serve as a viable prevention strategy.

[Advances of enzymes related to microbial cement].

Microbial induced calcium carbonate precipitation (MICP) refers to the natural biological process of calcium carbonate precipitation induced by microbial metabolism in its surrounding environment. Based on the principles of MICP, microbial cement has been developed and has received widespread attention in the field of biology, civil engineering, and environment owing to the merits of environmental friendliness and economic competence. Urease and carbonic anhydrase are the key enzymes closely related to microbial cement. This review summarizes the genes, protein structures, regulatory mechanisms, engineering strains and mutual synergistic relationship of these two enzymes. The application of bioinformatics and synthetic biology is expected to develop biocement with a wide range of environmental adaptability and high performance, and will bring the MICP research to a new height.

Status of Gene Methylation and Polymorphism in Different Courses of Ulcerative Colitis and Their Comparison with Sporadic Colorectal Cancer.

BACKGROUND: The objective of this study is to explore the common genetic and epigenetic mechanism of ulcerative colitis (UC) and sporadic colorectal cancer (SCRC) by observing genes methylation level and single nucleotide polymorphisms (SNPs) of different disease courses in UC and SCRC. METHODS: Two hundred subjects were enrolled, including 40 in the healthy control (HC) group, 50 in the short disease course UC group (SUC), 52 in the long disease course UC group (LUC), and 58 in the SCRC group. Methylation-specific polymerase chain reaction was used to detect the methylation of MINT1 and cyclooxygenase 2 (COX-2) gene. Single nucleotide polymorphisms of interleukin (IL)-23R rs10889677 and IL-1ß rs1143627 were detected by Sanger sequencing. RESULTS: Compared with HCs (32.5%), methylation level of MINT1 was significantly increased in SCRC (67.2%; P = 0.001) and was a risk factor for CRC (odds ratio, [OR] 4.26). The methylation ratios of COX-2 were 95.0%, 58.0%, 23.1%, and 24.1% in HC, SUC, LUC, and SCRC, respectively, which were negatively correlated with the disease course of UC (r = -0.290). Hypermethylation of COX-2 was a protective factor for SUC (OR, 0.11), LUC (OR, 0.02), and SCRC (OR, 0.03; P < 0.05). Compared with HCs, rs10889677 allele A was a risk factor for SUC and LUC, and rs1143627 allele T was a protective factor for SUC and LUC. Genotype TT was a protective factor for SUC. CONCLUSION: The hypomethylation of COX-2 gene was a common risk factor and epigenetic modification for UC and SCRC, which might be one of the mechanisms through which UC patients were susceptible to CRC. The hypermethylation of MINT1 was a risk factor for SCRC but not for UC; alleles of IL-23Rrs10889677 and IL-1ßrs1143627 were related to UC but not to SCRC.