[Epidemiological characteristic of viral encephalitis in children and adolescents in Henan Province, 2012-2023].

Objective: To understand the epidemiological characteristics and spatiotemporal distribution of viral encephalitis in children and adolescents in Henan Province from 2012 to 2023. Methods: The information about viral encephalitis cases from October 1, 2012 to July 26, 2023 were collected from Zhengzhou Children's Hospital (National Children's Regional Medical Center),Henan Provincial Children's Hospital for the analyses on temporal distribution the cases, the severe illness rate, age distribution, pathogen type and imaging findings of the cases. Results: A total of 6 276 cases of viral encephalitis were included in this study after excluding cases with incomplete information. The cases mainly originated from Zhengzhou (38.96%), followed by Zhoukou (9.93%), Xuchang (8.68%), Zhumadian (7.90%) and Pingdingshan (7.39%). The cases in boys accounted for 62.13% and the cases in girls accounted for 37.87%. Most cases (72.45%) occurred in age group 7-13 years. The overall rate of severe illness cases was 4.51% from 2012 to 2023. There were significant differences in severe illness cases among different areas and years (χ2=5.33,P=0.021; χ2=48.14,P<0.001). Enteroviruses were mainly detected (31.57%), in which Coxsackie virus was predominant (58.37%). Imaging findings showed that cerebral hemisphere damage was most common in children and adolescents with viral encephalitis (54.93%). Conclusions: From 2012 to 2023, more cases of viral encephalitis occurred in boys in Henan. Children and adolescents aged 7-13 years were the main affected group. The prevention of enteroviruses infection, especially Coxsackie virus, needs to be strengthened. Special attention should be paid to the prevention of cerebral hemisphere damage after viral encephalitis diagnosis.

[Epidemiological investigation on the local epidemic situation in Zhengzhou High-Tech Zone caused by SARS-CoV-2 Delta variant].

This study collected epidemic data of COVID-19 in Zhengzhou from January 1 to January 20 in 2022. The epidemiological characteristics of the local epidemic in Zhengzhou High-tech Zone caused by the SARS-CoV-2 Delta variant were analyzed through epidemiological survey and big data analysis, which could provide a scientific basis for the prevention and control of the Delta variant. In detail, a total of 276 close contacts and 599 secondary close contacts were found in this study. The attack rate of close contacts and secondary close contacts was 5.43% (15/276) and 0.17% (1/599), respectively. There were 10 confirmed cases associated with the chain of transmission. Among them, the attack rates in close contacts of the first, second, third, fourth and fifth generation cases were 20.00% (5/25), 17.86% (5/28), 0.72% (1/139) and 14.81% (4/27), 0 (0/57), respectively. The attack rates in close contacts after sharing rooms/beds, having meals, having neighbor contacts, sharing vehicles with the patients, having same space contacts, and having work contacts were 26.67%, 9.10%, 8.33%, 4.55%, 1.43%, and 0 respectively. Collectively, the local epidemic situation in Zhengzhou High-tech Zone has an obvious family cluster. Prevention and control work should focus on decreasing family clusters of cases and community transmission.

MicroRNA-147b alleviates inflammation and apoptosis in acute lung injury via inhibition of p38 MAPK signaling pathway.

OBJECTIVE: The pathogenesis of acute lung injury (ALI) is complicated, the condition is developing rapidly, and the mortality rate is high. It is a common acute and critical illness in clinic. Here, we aimed to demonstrate the function and molecular mechanism of microRNA-147b (miR-147b) in ALI. MATERIALS AND METHODS: MiR-147b mimic or miR-147b inhibitor was transfected into A549 cells to upregulate or downregulate miR-147b. The inflammatory response of A549 cells was observed by measuring the levels of inflammatory cytokines (TNF-α, IL-6, IL-1ß) and chemokines (CCL2, CCL4) by enzyme-linked immunosorbent assay (ELISA) assay. The detection of apoptosis in A549 cells relies on Cell Counting Kit-8 (CCK-8) assay, caspase-3 activity assay, and flow cytometry. Quantitative Real Time-Polymerase Chain Reaction (RT-PCR) and Western blot were employed to detect the expression of miRNA and protein. RESULTS: MiR-147b was downregulated in lipopolysaccharide (LPS)-induced ALI rats and LPS-treated A549 cells. Upregulation of miR-147b markedly suppressed LPS-induced inflammation and apoptosis of A549 cells, which was manifested by the reduction of inflammatory cytokines (TNF-α, IL-6, IL-1ß) and chemokines (CCL2, CCL4), the reduction of LDH contents, the increase of cell viability, and the decrease of caspase-3 activity and apoptosis rate of A549 cells. The downregulation of miR-147b further induced inflammation and apoptosis of A549 cells caused by LPS, which was alleviated by inhibition of p38 MAPK pathway. CONCLUSIONS: Taken together, miR-147b was downregulated in ALI, and the overexpression of miR-147b inhibited LPS-induced inflammation and apoptosis in A549 cells via inhibition of p38 MAPK signaling pathway.

[Preliminary analysis on COVID-19 case spectrum and spread intensity in different provinces in China except Hubei province].

Objective: To analyze the characteristics of COVID-19 case spectrum and spread intensity in different provinces in China except Hubei province. Methods: The daily incidence data and case information of COVID-19 were collected from the official websites of provincial and municipal health commissions. The morbidity rate, severity rate, case-fatality rate, and spread ratio of COVID-19 were calculated. Results: As of 20 March, 2020, a total of 12 941 cases of COVID-19 had been conformed, including 116 deaths, and the average morbidity rate, severity rate and case-fatality rate were 0.97/100 000, 13.5% and 0.90%, respectively. The morbidity rates in Zhejiang (2.12/100 000), Jiangxi (2.01/100 000) and Beijing (1.93/100 000) ranked top three. The characteristics of COVID-19 case spectrum varied from province to province. The first three provinces (autonomous region, municipality) with high severity rates were Tianjin (45.6%), Xinjiang (35.5%) and Heilongjiang (29.5%). The case-fatality rate was highest in Xinjiang (3.95%), followed by Hainan (3.57%) and Heilongjiang (2.70%). The average spread ratio was 0.98 and the spread intensity varied from province to province. Tibet had the lowest spread ratio (0), followed by Qinghai (0.20) and Guangdong (0.23). Conclusion: The intervention measures were effective in preventing the spread of COVID-19 and improved treatment effect in China. However, there were significant differences among different regions in severity, case-fatality rate and spread ratio.

D-dimer is associated with the risk of mortality in Coronavirus Disease 2019 patients.

This study aims to investigate the association between D-dimer and the risk of mortality in coronavirus disease 2019 (COVID-19) patients using a meta-analysis. We found that the D-dimer levels in non-survival patients were significantly higher than those in survival patients (SMD = 0.91, 95% CI = 0.79 to 1.03). In conclusion, the elevated D-dimer levels were associated with an increased risk of mortality in COVID-19 patients.

[Analysis on the epidemic factors for COVID-19].

As a new infectious disease, the epidemic process of COVID-19 has a series of special influencing factors and conditions. In this paper, some obvious characteristics of this widespread epidemic are discussed, including the new pathogen making people feel confused, the slow onset bringing confusion to the clinic, the miscellaneous source of infection also causing confusion to prevention and control work, the easy route of transmissions leading to a sharp increase of confirmed cases, the high susceptibility of the population leading to a high incidence, and the natural epidemic process coupled with the complexity of natural factors and the superposition of social factors. The positive and effective prevention and control strategies and measures adopted by China have greatly changed the natural epidemic process and trajectory of this epidemic, which has been highly affirmed by the expert group of the World Health Organization and praised by many countries and international organizations. However, to sum up carefully and think deeply, it will be a long-term and arduous work to plan and realize public health security in China and even the world in the future.

[The preliminary analysis on the characteristics of the cluster for the COVID-19].

Since December 2019, COVID-19, a new emerging infection disease, has spread in 27 countries and regions. The clusters of many cases were reported with the epidemic progresses. We collected currently available information for 377 COVID-19 clusters (1 719 cases), excluded the hospital clusters and Hubei cases, during the period from January 1 to February 20, 2020. There were 297 family clusters (79%), case median was 4; 39 clusters of dining (10%), case median was 5; 23 clusters of shopping malls or supermarkets (6%), case median was 13; 12 clusters of work units (3%), case median was 6, and 6 clusters of transportation. We selected 325 cases to estimate the incubation period and its range was 1 to 20 days, median was 7 days, and mode was 4 days. The analysis of the epidemic situation in a department store in China indicated that there was a possibility of patients as the source of infection during the incubation period of the epidemic. From February 5 to 21, 2020, 634 persons were infected on the Diamond Princess Liner. All persons are susceptible to the 2019 coronavirus. Age, patients during the incubation period and the worse environment might be the cause of the cases rising. The progress of the two typical outbreaks clearly demonstrated the spread of the early cases in Wuhan. In conclusion, screening and isolating close contacts remained essential other than clinical treatment during the epidemic. Especially for the healthy people in the epidemic area, isolation was the key.

[Identification and evaluation on methods with upstream flank sequences of CRISPR1, regarding Escherichia coli and Shigella].

Objective: To analyze the effect of the identification and evaluation of Escherichia (E.) coli and Shigella, based on the upstream flanking sequences of CRISPR1. Methods: Both CRISPR and cas sequences were obtained through the BLAST with repeating sequences against the publicly complete genome in GenBank that related to E. coli and Shigella. Clustal X was used to perform multi-sequences alignment of the flanking sequences. PCR method was used to amplify the upstream flanking sequences of CRISPR1 in order to appraise the effect of identification and evaluation of upstream flanking sequences on E. coli and Shigella, which were based on the upstream flanking sequences of CRISPR1. Results: The results showed that 73.4% of the strains containing the I-E CRISPR/Cas that belonged to the phylogroups A, B1, D while 8.4% strains carried the I-F CRISPR/Cas. Another 17.2% of the strains owned CRISPR3-4 (non-CRISPR/Cas) only belonged to the phylogroups B2. All the Shigella strains carried I-E CRISPR/Cas. More than 99% of similarity the CRISPR1 upstream-flanking sequences was seen in E. coli (except B2) and Shigella and E. coli (B2). Both sensitivity and specificity were greater than 91% after PCR amplification in the region to identify the E.coli and Shigella. Conclusion: The upstream of CRISPR1 could achieve a preliminary identification effect on E.coli and Shigella.

[Association between phage-mediated shiga toxin and molecular distribution of CRISPR in Escherichia coli O26 ∶ H11 or NM].

Objective: To investigate the association between phage-mediated shiga toxin and molecular distribution of CRISPR in Escherichia (E.) coli O26∶H11 or NM. Methods: A total of 135 E. coli O26 ∶ H11 or NM strains were collected from NCBI database. Software CRT and CRISPR Finder were used to extract CRISPR and Excel was used to assign the spacer of unique number and type CRISPR. And the relationship between CRISPR and stx phage was analyzed. Results: All the 135 E. coli O26 ∶ H11 or NM strains had the CRISPR. For CRISPR1, CRISPR2.1, CRISPR2.2 and CRISPR3-4, 19, 22, 1 and 1 subtypes were found, respectively. According to the four CRISPR sites, the strains could be divided into 40 subtypes. Stx-phage was only observed in the group C of CRISPR. Compared with E. coli of stx-phage negative, E. coli with stx-phage harbored more spacers. Conclusions: CRISPR loci was extensively existed in E. coli O26∶H11 or NM, and many subtypes were found in these strains. The presence of stx-phage was related to the molecular distribution of CRISPR in E. coli O26∶H11 or NM. CRISPR might be a valuable biomarker to identify strains with high virulent potential.

[Changes of resistant phenotype and CRISPR/Cas system of four Shigella strains passaged for 90 times without antibiotics].

Objective: To explore the stability of resistant phenotypes and changes of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) gene system on four Shigella strains in the absence of antibiotics. Methods: Four clinical isolated Shigella strains that resistant to different antibiotics were consecutive passaged for 90 times without antibiotics. Agar dilution method was used to determine the minimum inhibitory concentration of Shigella strains. After sequence analysis with PCR, CRISPR Finder and Clustal X 2.1 were applied to identify the changes of CRISPR loci in the Shigella strains. Results: After the consecutive transfer of 90 generations, sensitivity to certain antibiotics of four Shigella strains with different drug resistant spectrums increased. Mel-sf1998024/zz resistance to ampicillin, cephalexin, cefotaxime, chloramphenicol decreased, mel-s2014026/sx resistance to norfloxacin, trimethoprim decreased, mel-sf2004004/sx drug resistance to ampicillin, cefuroxime, cefotaxime, chloramphenicol, trimethoprim decreased and mel-sf2013004/bj resistance to chloramphenicol decreased. The spacer of which matched gene codes Cas and its upstream repeat in 3'end of CRISPR3 got lost in mel-sf1998024/zz and mel-sf2013004/bj. Conclusions:Shigella strains could reduce or lose their resistance to some antibiotics after consecutive transfers, without the interference of antibiotics. CRISPR3 locus had dynamic spacers in Shigella strains while CRISPR3 locus and cas genes might have been co-evolved.

[Opportunity and challenge on molecular epidemiology].

Molecular epidemiology, a branch of epidemiology, combines the theories and methods, both in epidemiology and molecular biology. Molecular epidemiology mainly focuses on biological markers, describing the distribution, occurrence, development and prognosis of diseases at the molecular level. The completion of Human Genome Project and rapid development of Precision Medicine and Big Data not only offer the new development opportunities but also bring about a higher demand and new challenge for molecular epidemiology.

[A surveillance study on CRISPR/Cas molecular biomarker in Escherichia coli].

OBJECTIVE: A new method related to molecular biomarker with CRISPR/Cas (clustered regularly interspaced short palindromic repeats-cas) in Escherichia (E.) coli was developed and used for surveillance programs. METHODS: CRISPR/Cas sequence that containing 135 strains with complete sequence and 203 strains with whole genome shotgun sequence of E. coli in GenBank by BLAST and 361 strains of E. coli (including 38 strains of E. coli O157∶H7) in laboratory were identified by PCR and analyzed with the CRISPR Finder. Spacers were compared with DANMAN and the phylogenetic trees of cas gene were constructed under Clustal Ⅹ and Mega 5.1. RESULTS: With new perspective, a descriptive method was developed targeting on the position of CRISPR/cas in E. coli. The CRISPR1 was detected in 77.04%, 100.00% and 75.62% and the CRISPR2 was detected in 74.81%, 100.00% and 92.24% and the CRISPR3 and CRISPR4 were detected in 11.85%, 0 and 1.39% for 135 strains with complete sequence, 203 strains with whole genome shotgun sequence and 361 strains in the laboratory, respectively. One strain downloaded in GenBank with whole genome sequencing and 2 strains in the our laboratory were identified that containing four CRISPR locus. The other E. coli strain was with insertion sequence in downstream of the non-cas CRISPR1. The unique CRISPR was found in 8 strains of O55∶H7, in 180 strains of O157∶H7, in 8 strains of O157∶HNM, in 40 strains of O104∶H4, in 4 strains of O145∶H28, in all the 699 E. coli strains. The phylogenetic tree could be divided into two groups-cas with type I-E or type I-F. CONCLUSIONS: CRISPR/Cas might be used as a valuable molecular biomarker in epidemiological surveillance studies to identify the high virulent strains or new strains of E. coli. Phage night be related to the missing or obtaining of spacers.

Molecular mechanism of multi-drug resistance in Shigella isolates from rural China.

Diarrhoeal diseases and enteric infections are major causes of morbidity and mortality in the developing countries. Shigellosis was sixth in the rank of death caused by infectious disease in China. Present study assess the patterns of antimicrobial susceptibility and mutations in marA genes of Shigella isolates and its association. One hundred isolates of Shigella spp were tested to evaluate the antimicrobial susceptibility and mutation of marA through PCR-SSCP. The antimicrobial resistance profiles were: Amoxicilline-clavum (85%), Nalidixic acid (100%), Piperacilline (100%), Ciprofloxacin (25%) and Polymixin B (1.1%). S. flexneri isolates were more resistant than those ofS. sonnei to Amoxicilline-clavum (96.6% versus 0.0%, P<0.001) and Ciprofloxacin (27.3% versus 8.3%, P=0.141). S. flexneri isolates were more resistant than those of S. sonnei to three or more antimicrobial agents (96.6% versus 8.3%, P<0.001). Overall mutation rate in marA was 19%. Nucleotide sequencing showed mutations in three positionsÿcodon 6 ( Delation of C), 319 (Ala Gly) and 374 (Addition of C). The mutation of marA may play a minor role and other mechanisms may contribute to the drug resistance of Shigella spp.

A natural vaccine candidate strain against cholera.

E1 Tor Vibrio cholerae (EVC) strains may be classified into two kinds-epidemigenic (EEVC) strains and non-epidemigenic (NEEVC) strains-based on a phage-biotyping system. A large number of EEVC strains have been screened for toxigenic and putative colonization attributes. One such naturally occurring strains (designated IEM101) has been found which is devoid of genes encoding cholera toxin (CT), accessory cholera enterotoxin (ACE), zonula occludens toxin (ZOT), but possesses RS1 sequences and toxin-coregulated pilus A gene (icpA) although icpA is poorly expressed. It expresses type B pili but does not possess type C pili. It is an E1 Tor Ogawa strain and does not cause fluid accumulation in rabbit ilcal loop tests. Active immunization of rabbits with strain IEM101 elicited good protection against challenge with virulent strains of V. cholerae O1. Oral administration caused no side effects in 15 human volunteers, colonized the gut for four to ten days and elicited good immune responses.