Prevention and control of SARS-CoV-2 infection in clinical laboratories: implementation of contingency plan and postpandemic response strategies / 南方医科大学学报

The outbreak of COVID-19 has currently been under control in China, but now the disease has rapidly evolved into a global pandemic. We formulated a prevention and control plan for clinical laboratories responsible for detection of the novel coronavirus infection. We analyzed the implementation of this plan and the problems arising from its clinical practice. We found that the layout of most clinical laboratories (including gene amplification laboratories for clinical samples) was inadequate in response to a major outbreak and did not meet the requirements for biosafety protection and etiology and serology testing; and laboratory staff showed insufficiencies in their awareness regarding biosafety protection; the functions and status of the laboratory in the fever clinic need to be enhanced to increase its detection capacity; the high density of military personnel, the low level of automation of clinical laboratory equipment, and the lack of biosafety cabinets and personal protective equipment all limit the performance of diverse military operations and major overseas missions. In view of these problems, we propose the following strategies and recommendations: the clinical laboratory needs to standardize the design and staff management according to the standards of P2 laboratory; the detection capacity and staffing of fever clinic laboratory in hospitals need to be strengthened, and a separate clinical gene amplification laboratory can be optimal; for those clinical gene amplification laboratories that fail to meet these standards, reconstruction and upgrade should be made according to the requirements of biosafety protection; for the clinical laboratory in the military medical system, in addition to enforcement of biological safety protection of the staff, sufficient supply of medical materials and biological safety equipment should be ensured and biological safety cabinets should be routinely equipped if possible.

Structure and function of B-cell linker and its role in the development of B cell-related diseases / 南方医科大学学报

B cell linker (BLNK) is a key linker protein of B cell receptor (BCR) signaling pathway. BLNK participates in the regulation of PLC-γactivity and the activation of Ras pathway through its typical structure and interaction network with other proteins, and is thus widely involved in the regulation of B cell proliferation, differentiation, apoptosis and signal transduction. Furthermore, it is closely related to anaphylactic diseases, multiple sclerosis, chromosomal aneuploidy, aneuglobulinemia, B lymphocytic leukemia and lymphoma. Herein we review the structure and biological function of BLNK and its role in B cell-related diseases. BLNK can cooperate with a series of effective proteins to activate BCR signaling pathway, thereby regulating the development, maturation and function of B cells. The functional mutation of BLNK can destroy the homeostasis of B cells and affect the development and maturation of B cells, which leads to the occurrence of B cell related diseases. A comprehensive understanding of the biological functions of BLNK not only provides insights into the pathogenesis of B cell-related diseases, but also inspires new ideas and helps to find breakthroughs for the treatment of these diseases with BLNK as the therapeutic target.

Clinical application of serum miR-122-5p and miR-486-5p in the diagnosis of hepatocellular carcinoma / 中华检验医学杂志

Objective Explore the relative expression of miR-122-5p and miR-486-5p in the serum of Hepatocellular carcinoma ( HCC) patients and its clinical value .Methods Case-control study was used in this research.From June of 2016 to March of 2017,60 HCC patients who were hospitalized in Guangzhou General Hospital were selected as HCC group .It also selected 20 hepatitis patients ( hepatitis group ) , 20 cirrhosis patients ( cirrhosis group ) , 20 breast cancer patients ( breast cancer group ) , 20 gastric cancer patients(gastric cancer group)and 20 healthy controls (normal control group) for comparison.The relative expression of miR-122-5p and miR-486-5p was detected by real-time fluorescent quantitative PCR.The specificity and sensitivity of miRNAs for the diagnosis of HCC were analyzed by receiver operating characteristic ( ROC) , and the results were compared with the tumor marker AFP .The effect of miRNA on the diagnosis of hepatocellular carcinoma was evaluated by the area under the ROC curve , which was used to detect the diagnostic efficiency of liver cancer .SPSS22.0 statistical software was used for statistical analysis . The rank sum test was applied in the group comparison .Results Serum levels of miR-122-5p in HCC group, hepatitis group, cirrhosis group, breast cancer group, gastric cancer group and control group were 0.14(0.05-0.51),0.45(0.32-0.58),0.53(0.34-0.67),0.14(0.07-0.28),0.29(0.13-0.36) and 0.73 (0.63-0.95),respectively, and the miR-486-5p were 0.50(0.23-0.77),0.62(0.48-0.82),0.65(0.54-0.85),0.23(0.08-0.40),0.29(0.15-0.45)and 0.76(0.69-1.23).The serum levels of miR-122-5p in hepatitis group , cirrhosis group , HCC group were significantly lower in healthy control group , significance was found (U was 315.37,393.46,429.08, all P<0.01), and the serum levels of miR-486-5p in hepatitis group, cirrhosis group, HCC group were lower in healthy control group , significance was found ( U was 103.67,156.18,207.35, all P<0.05).When using one serum marker to diagnosis HCC , AFP had the highest sensitivity ( 73.7％) and miR-122-5p had the highest specificity ( 95％) .While combined two serum markers, AFP +miR-122-5p had the highest sensitivity and specificity (93％),and miR-122-5p +miR-486-5p had the highest specificity (70％), compared AFP +miR-122-5p to AFP, AUC difference was statistically significant(Z=3.02,P<0.01), while there was no significant difference in AUC with AFP +miR-486-5p, miR-122-5p +miR-486-5p to AFP(Z=1.57,1.39,all P>0.05).The sensitivity and specificity of the three markers were 96.5％and 55％respectively , and the area under the ROC curve was 0.891 (95％CI:0.818-0.964).The combination miR-122-5p, miR-486-5p and AFP were higher than the single test, compared with AFP, miR-122-5p, miR-486-5p, the AUC differences was statistically significant (Z=3.26, 3.72, 4.25, all P<0.01).Conclusion Serum miR-122-5p and miR-486-5p could be used as biological markers for the diagnosis of HCC .

Construction of human kinase knock-out library by using CRISPR/Cas9 technique / 实用医学杂志

Objective CRISPR/Cas9 genome-editing technique provides an novel method for whole genome editing in eukaryotic cells.Recently,we found that gene subtype library with smaller size and focused pur-pose is more economical and practical. In this study,we aimed to target kinases,a group of pivotal cell signal transducers,to construct a kinase knock-out library using CRISPR/Cas9 technique.The construction strategy wll al-so be discussed. Methods 10 sgRNA was designed for each kinase target.After oligo pool synthesis by semicon-ductor chip,the oligos were eluted from the chip. The oligo templates were amplified and cloned into Cas9 vector and transformed into Stble3 competent cells.Monoclonal colonies were selected for DNA sequencing. Results(1) GO analysis of 507 cell kinases showed that the cell kinases took part in a wide range of cell signaling.(2)The sgRNA pool with about 140 bp in length was successfully amplified by using oligo pool as the template and univer-sal PCR primers.(3)In 40 identified library clones,34 clones were sequenced successfully. Among them,the DNA sequencing results of 25 samples were completely consistent with the designed target sequences.But there are some mutations in the primers of 9 samples.Failure in bacteria shaking,DNA sequencing and other factors were ex-isted in the other clones. Conclusion The CRISPR/Cas9 kinase knock-out library can be widely used for screen-ing the important kinases which may mediate cell proliferation,metastasis,drug resistance and autophagy.This li-brary will play an important role in clarifying the development of disease associated with kinases.

Detection of Legionella pneumophila infection in children with lower respiratory infections by ELISA and PCR / 中华急诊医学杂志

Objective To investigate the infection rate of Legionella pneumophila(LP)in children younger than 5 years with lower respiratory infections by ELISA and PCR.Method Serum LP-IgM and IgG were measured by ELISA,and LP DNA in sputum or throat swab were detected by PCR in 300 children less than 5 years with lower respiratory infections.The data were analyzed by chi-square test and the consistency of the two methods was analyzea by NcNemar test.Results Serum antibody detected by ELISA was positive in 17 cases(5.67%),including 10 positive of IgM and 7 positiile of lgG.In these 17 eases,11 were males and 6 were females with similar positive rates(5.76%vs 5.5%).Four cases(2.53%)were positive in children aged from 27 days to 1 year,while 7(7.37%)and 6 cases(12.77%)were positive in children aged 1-3 years and 3-5 years,respectively.Seven cases(5.19%)found in the winter and spring seasons and 10 cases(6.06%)in summer and autumn seasons.Eleven children(11.83%)were from urban area and only 6(2.9%)from countryside.DNA in sputum or throat swab detected by PCR was positive in 16 cases(5.33%),included 10 males and 6 females with similar positive rates(5.24%vs 5.5%).Five cases(3.16%)were positive in children aged from 27 days to 1 year,while 6(6.32%)and 5 cases(10.64%)were positive in children aged 1-3 years and 3-5 years,respectively.Three cases(2.22%)found in the winter and spring seasons and 13 cases(7.88%)in sunmmr and alltumll seasons.Nine children(9.68%)were from urban area and only 7(3.38%)from countryside.McNemar test was used to compare the data of ELISA and PCR results with a Qm of 0.037 and a Pr of 0.8474.Conclusions LP might contribute partly to the lower respiratory infections in children less than 5 years.The infection rate of LP increases with age increasing.Urban children have a higher infection rate than those living in countryside.Both methods have a good consistency.