This study was aimed to explore the features of clustered regularly interspaced short palindromic repeats (CRISPR) structures in Shigella by using bioinformatics. We used bioinformatics methods, including BLAST, alignment and RNA structure prediction, to analyze the CRISPR structures of Shigella genomes. The results showed that the CRISPRs existed in the four groups of Shigella, and the flanking sequences of upstream CRISPRs could be classified into the same group with those of the downstream. We also found some relatively conserved palindromic motifs in the leader sequences. Repeat sequences had the same group with corresponding flanking sequences, and could be classified into two different types by their RNA secondary structures, which contain "stem" and "ring". Some spacers were found to homologize with part sequences of plasmids or phages. The study indicated that there were correlations between repeat sequences and flanking sequences, and the repeats might act as a kind of recognition mechanism to mediate the interaction between foreign genetic elements and Cas proteins.
Base Sequence ; Clustered Regularly Interspaced Short Palindromic Repeats ; Computational Biology ; Genome, Bacterial ; Plasmids ; Shigella ; genetics

Objective To explore the image quality and diagnostic efficiency of coronary CTA (CCTA) in patients without heart rate (HR) control by optimizing acquisition phase with auto-ECG-gating technology using 256-row wide-volume detector.Methods Totally 200 patients with suspected coronary artery disease (CAD) were selected and underwent CCTA with auto-ECG-gating on a 256 row wide detector CT (Revolution CT).Patients were divided into 4 groups according to the real-time HR,group A (n=50):HR≤69 bpm;group B (n=50):HR 70-80 bpm;group C (n=50):HR 81-90 bpm;group D:HR≥91 bpm (n=50).CCTA images quality and diagnostic rate were assessed by two experienced radiologists blindly,and effective radiation dose were compared among 4 groups.Results There were no significant differences of age,sex,and body mass index among 4 groups (all P＞0.05),and there was significant difference of image quality score among 4 groups (P＜0.05).Totally,800 coronary arteries and 2 575 segments in 200 patients were assessed.There had no significant difference of diagnostic rate among four groups derived from the segment,coronary artery and patients (all P＞0.05).The effective radiation dose in A D groups were (1.05t0.48)mSv,(2.41± 1.20)mSv,(1.27±0.55) mSv,(2.66±1.12)mSv,and the difference was significant (F=29.22,P＜0.001).Conclusion It is feasible to perform CCTA in single cardiac cycle in patients with arbitrary heart rate by auto-ECG-gating using Revolution CT equipped widevolume detector.And it can improve image quality and success ratio in patients with moderate and high heart rate.

AIM:To explore the effects of galectin-3 (GAL-3) on the viability, migration and inflammation of human umbilical vein endothelial cells (HUVECs) and the mechanisms.METHODS:The HUVECs were cultured in vitro and treated with GAL-3 recombinant protein at 2 mg/L or GAL-3 short hairpin RNA (shRNA).The HUVECs were divided into normal group, recombinant GAL-3 group, shControl group and GAL-3-shRNA group.The mRNA expression of GAL-3, monocyte chemotactic protein (MCP)-1, IL-6, matrix metalloproteinase (MMP)-9 and cyclin D1 was detected by real-time quantitative PCR,and the protein expression of GAL-3, IL-6 and MCP-1 was detected by Western blot.The secretion levels of MCP-1 and IL-6 in the culture medium were measured by ELISA.The viability and the ability of migration of the HUVECs were examined by CCK-8 assay and wound healing assay.The protein levels of heat shock protein 90 (HSP90), ERK1/2, p-ERK1/2, JNK and p-JNK were determined by Western blot.RESULTS:The expression of GAL-3, MCP-1 and IL-6 at mRNA and protein levels, the mRNA expression of MMP-9 and cyclin D1, and the secretion levels of MCP-1 and IL-6 in the culture medium were significantly higher than those in normal group (P<0.05) after the HUVECs were treated with GAL-3 recombinant protein.However, these molecules mentioned above in GAL-3-shRNA group were significantly lower than those in normal group and negative control group (P<0.05).Compared with normal group, the viability and migration ability of the HUVECs in recombinant GAL-3 group were significantly increased, but the viability and migration ability of the HUVECs in GAL-3-shRNA group were lower than those in normal group and shControl group (P<0.05).In addition, the protein levels of p-ERK1/2 and HSP90 in recombinant GAL-3 group were higher than those in normal group (P<0.05), but those in GAL-3-shRNA group were lower than those in normal group and shControl group (P<0.05).The protein level of p-JNK was not oviously changed among the 4 groups.CONCLUSION:GAL-3 is involved in regulating the cell growth, migration and the release of inflammatory cytokines in vascular endothelial cells, which may be mediated by HSP90-ERK1/2 signaling pathway.

Objective To detect the distribution of clustered regularly interspaced short palindromic repeat(CRISPR)associated protein genes cas1 and cas2 in Shigella and to understand the characteristics of CRISPR with relationship between CRISPR and related characteristics on drug resistance. Methods CRISPR associated protein genes cas1 and cas2 in Shigella were detected by PCR,with its products sequenced and compared.Results The CRISPR-associated protein genes cas1 and cas2 were found in all the 196 Shigella isolates which were isolated at different times and locations in China. Consistencies showed through related sequencing appepared as follows:cas2,cas1 (a) and cas1(b)were 96.44%,97.61%and 96.97%,respectively. There were two mutations including 3177129 site(C→G)and 3177126 site(G→C)of cas1(b)gene in 2003135 strain which were not found in the corresponding sites of Z23 and 2008113. Results showed that in terms of both susceptibility and antibiotic-resistance,strain 2003135 was stronger than Z23 and 2008113. Conclusion CRISPR system widely existed in Shigella,with the level of drug resistance in cas1(b) gene mutant strains higher than in wild strains. Cas1(b)gene mutation might be one of the reasons causing the different levels of resistance.

OBJECTIVETo detect the molecular characteristics of Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR) in Shigella and to analyze the distribution of CRISPR related to the time of isolation.

METHODSOf the 52 Shigella strains, 41 were isolated from Henan, 6 from Jiangxi and 5 isolated from Beijing. Both CRISPR locus of S1, S2, S3 and S4 in Shigella were detected by polymerase chain reaction (PCR). The PCR products were sequenced and compared.

RESULTSThe positive rates of CRISPR locus in Shigella were 33.69% (S1), 50.00% (S2), 82.69% (S3) and 73.08% (S4), respectively. Two subtypes were discovered in S1 and S3 locus. Three subtypes were discovered in S2 locus. Four different subtypes were discovered in S4 locus. The isolates from Henan strains were divided into two groups by the time of isolation. Distributions of S1 were different, before or after 2004, on Shigella. S1 could not be detected after 2004. There were no statistical differences of S2, S3 and S4 in two groups.

CONCLUSIONDifferent CRISPR subtypes or Shigella were discovered. A significant correlation was noticed between the CRISPR S1 related to the time of isolation but not between S2, S3 or S4 on the time of isolation.

Clustered Regularly Interspaced Short Palindromic Repeats ; Polymerase Chain Reaction ; Shigella ; genetics

OBJECTIVETo detect the distribution of clustered regularly interspaced short palindromic repeat (CRISPR) associated protein genes cas1 and cas2 in Shigella and to understand the characteristics of CRISPR with relationship between CRISPR and related characteristics on drug resistance.

METHODSCRISPR associated protein genes cas1 and cas2 in Shigella were detected by PCR, with its products sequenced and compared.

RESULTSThe CRISPR-associated protein genes cas1 and cas2 were found in all the 196 Shigella isolates which were isolated at different times and locations in China. Consistencies showed through related sequencing appeared as follows: cas2, cas1 (a) and cas1 (b) were 96.44%, 97.61% and 96.97%, respectively. There were two mutations including 3177129 site(C→G)and 3177126 site (G→C) of cas1 (b) gene in 2003135 strain which were not found in the corresponding sites of Z23 and 2008113.

RESULTSshowed that in terms of both susceptibility and antibiotic-resistance, strain 2003135 was stronger than Z23 and 2008113.

CONCLUSIONCRISPR system widely existed in Shigella, with the level of drug resistance in cas1 (b) gene mutant strains higher than in wild strains. Cas1 (b) gene mutation might be one of the reasons causing the different levels of resistance.

Bacterial Proteins ; genetics ; CRISPR-Associated Proteins ; genetics ; Clustered Regularly Interspaced Short Palindromic Repeats ; Drug Resistance, Bacterial ; genetics ; Mutation ; Shigella ; genetics

Objective: To understand the etiological and clinical characteristics of children with severe hand, foot and mouth disease (HFMD) in Xi′an in 2018, and to provide the evidence for clinical diagnosis and treatment. Methods: The children with severe HFMD admitted at Xi′an Children′s Hospital from January to December 2018 were selected as the research objects.Clinical data were collected, and the anal swab were detected by adopting real time(RT)-polymerase chain reaction(PCR). Results: Ninety-five cases of HFMD were treated in Xi′an Children′s Hospital in 2018, of which 92 cases were severe and 3 cases were critical.Eighty-seven cases were positive for enterovirus nucleic acid, 30 cases were enterovirus 71(EV71)(31.6%), 39 cases were coxsackievirus A6(CA6) (41.0%), 3 cases were CA16(3.2%), 2 cases were CA10(2.1%) and 13 cases were other enteroviruses (13.7%). Among 95 patients, the ratio of male to female was 1.1∶1.0; the peak period of incidence of HFMD was from May to July, and the main age of onset of severe HFMD was under 3 years old.The main clinical manifestations were mental retardation, vomiting, irritability, lethargy and convulsion.Severe cases of CA6 are prone to convulsion.The main form of rash in CA6 cases was bullous rash, and demethylation may occur in recovery period.The rash in EV71 cases was small, thick, hard and few.After active treatment, only one child with EV71 infection died because of severe cerebral dysfunction, frequent convulsions and neurogenic pulmonary edema.The other child was discharged with hemiplegia and language dysfunction.The other severe children were cured and discharged from hospital. Conclusions In 2018, CA6 was the main pathogen of severe HFMD in Xi′an, with bullae was the main manifestation of skin rash, and nail removal could occur during convalescence.Critical and death cases were still caused by EV71.