Objective To evaluate the feasibility of real-time quantitative PCR (RtPCR) with small volume regarding the stability, efficiency and reliability of amplification, and determine the optimal quantity of cDNA template suitable for small PCR volume. Methods The experiment was carried out in 3 groups with 10, 15 and 20μL reaction volume, respectively. In each group, rat β-actin mRNA was detected by RtPCR with 0.1, 0.2, 0.3 or 0.4μL rat cDNA as template, respectively. The amplification curve and melting curve were used to evaluate the reaction stability. The fitting of a curve of gradient templates against threshold cycle numbers was to show the reaction efficiency and the linear correlativity was to estimate the suitability of the template quantity. In addition, in order to estimate reliability, pristane-induced arthritis (PIA) rat model was established, and spleen TNF-α mRNA expression was detected by RtPCR with the selected reaction volumes. Results The amplification of rat β-actin mRNA was specific and stable in 10μL, 15μL and 20μL PCR volume, and had a high efficiency. Furthermore, the standard curves fitted by 0.1-0.4μL gradient templates showed a significant linear correlation in each volume group. When the 10μL and 20μL PCR volumes, and 0.2μL cDNA templates were chosen, the TNF-α mRNA expression in PIA rat spleen showed significant upregulation in both two volume groups as anticipated. Conclusion The experiment shows that it is feasible in the RtPCR amplification to use the small reaction volume of 10μL and 15μL, which has good stability and reliability. And 0.1-0.4μL templates are all suitable for the reaction system. PCR with small volume can not only save the reagents and template, especially rare clinical specimens, but also is helpful for the realization of high-throughput reaction.

Objective:To investigate the attributes and influencing factors of care needs for maternal after perinatal loss, and to provide a reference for promoting maternal physical and mental health and improving the quality of care.Methods:From February to June of 2023, a cross-sectional study was used, 222 maternal after perinatal loss were selected by the convenience sampling method and completed a battery of questionnaires, including a general information questionnaire, the Distress Thermometer (DT), the Perceived Social Support Scale (PSSS), the Attitudes Toward Seeking Professional Psychological Help Scale-Short Form (ATSPPH-SF) and Kano Model-based Questionnaire on Care Needs for Maternal after Perinatal Loss.Results:A total of 207 maternal after perinatal loss were included in the complete data, (31.12±4.55) years old, (68.64 ± 10.70) of PSSS, (16.10 ± 3.43) of ATSPPH-SF. The care needs for maternal after perinatal loss included 4 must-be quality, 8 one-dimensional quality and 13 attractive quality. Multiple linear regression analysis showed that education level ( t=2.28), the PSSS score ( t=2.15) and the ATSPPH-SF score ( t=3.94) were the main influencing factors of care needs for maternal after perinatal loss (all P<0.05). Conclusions:Health care professionals should gradually improve the nursing service system according to the priority division of care needs attributes of maternal after perinatal loss, and develop personalized care according to different influencing factors.

Objective:To analyze the influenza surveillance results and epidemiological characteristics of the disease in Gansu province in 2010-2018, and provide scientific basis for the formulation and adjustment of influenza prevention and control strategies.Methods:Real-time PCR was used to detect the specimens of ILI illness (ILI) cases reported by 19 national influenza surveillance sentinel hospitals in Gansu province. Influenza virus isolation and culture were carried out using canine kidney cells (MDCK).Results:Total of 93 334 specimens from ILI cases were collected from Jan. 2010 to Dec. 2018, and 13 439 samples were positive for influenza virus nucleic acid. The positive rate of viral nucleic acid detection was 14.40%, and the proportion of influenza B virus in positive samples was the highest (37.02%), and seasonal H3N2 influenza virus (35.25%), H1N1 influenza virus (26.78%). The advantages are different in various periods. The peak period of influenza is from Dec. to Mar., and the highest incidence rate is in the age group of 5-15 years (20.34%). The incidence rates of the other age groups are similar. The difference of influenza positive rate in different age groups is statistically significant ( χ2=775.153, P<0.001), but in male and female was not significant. Total of 16 101 ILI specimens positive for influenza virus nucleic acid were subjected to virus isolation and cultured, and 3 892 strains of influenza virus were isolated, of which influenza B virus accounted for the highest proportion (37.82%), followed by seasonal H3N2 influenza virus (33.04%), H1N1 influenza virus (28.96%). Conclusions:The epidemic of influenza in Gansu province had an obvious regularity, and it was undulating and prevalent. It is necessary to strengthen prevention and control measures among school-age children and adolescents.

The strikingly rapidly mutating nature of the severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)genome has been a constant challenge during the coronavirus disease 2019(COVID-19)pandemic.In this study,various techniques,including reverse transcription-quantitative polymerase chain reaction,antigen-detection rapid diagnostic tests,and high-throughput sequencing were analyzed under different scenarios and spectra for the etiological diagnosis of COVID-19 at the population scale.This study aimed to summarize the latest research progress and provide up-to-date understanding of the methodology used for the evaluation of the immunoprotection conditions against future variants of SARS-CoV-2.Our novel work reviewed the current methods for the evaluation of the immunoprotection status of a specific population(endogenous antibodies)before and after vaccine inoculation(adminis-tered with biopharmaceutical antibody products).The present knowledge of the immunoprotection status regarding the COVID-19 complications was also discussed.Knowledge on the immunoprotection status of specific populations can help guide the design of pharmaceutical antibody products,inform practice guidelines,and develop national regulations with respect to the timing of and need for extra rounds of vaccine boosters.

Coronavirus disease 2019 (COVID-19) has been a pandemic for more than a year.With the expanding second wave of the pandemic in winter,the continuous evolution of SARS-CoV-2 has brought new issues,including the significance of virus mutations in infection and the detection of asymptomatic infection.In this review,we first introduced several major SARS-CoV-2 mutations since the COVID-19 outbreak and then mentioned the widely used molecular detection techniques to diagnose COVID-19,primarily focusing on their strengths and limitations.We further discussed the effects of viral genetic variation and asymptomatic infection on the molecular detection of SARS-CoV-2 infection.The review finally sum-marized useful insights into the molecular diagnosis of COVID-19 under the special situation being challenged by virus mutation and asymptomatic infection.

Objective:To investigate the status of grief among maternal spouse after perinatal loss, and analyze its influencing factors, so as to provide some reference for male grief supporting strategic.Methods:Using the convenient sampling method, 180 male spouses of hospitalized women in the Department of Obstetrics from Nanjing Maternity and Child Health Care Hospital from March to October 2022 were recruited. A cross-sectional survey was conducted by the general questionnaire, the Perinatal Grief Scale, the Family Adaptability and Cohesion Scale Ⅱ-Chinese Version, the Social Support Rating Scale, and the Simplified Coping Style Questionnaire.Results:The overall score of the Perinatal Grief Scale in male spouses of women who experienced a perinatal loss was (61.57 ± 14.14) points. The score of the Family Adaptability and Cohesion Scale Ⅱ-Chinese Version was (121 ± 14.42) points, the score of the Social Support Rating Scale was (34.23 ± 7.21) points, and the score of the Simplified Coping Style Questionnaire was (36.08 ± 7.64) points. Multiple linear regression analysis showed that participation in fetal interaction, loss of fetal age, social support and family adaptability were the main factors affecting male grief ( P<0.05). Conclusions:The grief among male spouses of women who experienced a perinatal loss is at a low level. The clinical medical staff can refer to the influencing factors and implement effective support, such as respecting the male's father status, coordinating social support resources, and improving the family's coping ability, in order to alleviate men's grief and help them return to normal life.

MicroRNA-21(miRNA-21)is highly expressed in various tumors.Small-molecule inhibition of miRNA-21 is considered to be an attractive novel cancer therapeutic strategy.In this study,fluoroquinolone de-rivatives Al-A43 were synthesized and used as miRNA-21 inhibitors.Compound A36 showed the most potent inhibitory activity and specificity for miRNA-21 in a dual-luciferase reporter assay in HeLa cells.Compound A36 significantly reduced the expression of mature miRNA-21 and increased the protein expression of miRNA-21 target genes,including programmed cell death protein 4(PDCD4)and phos-phatase and tensin homology deleted on chromosome ten(PTEN),at 10 uM in HeLa cells.The Cell Counting Kit-8 assay(CCK-8)was used to evaluate the antiproliferative activity of A36;the results showed that the IC50 value range of A36 against six tumor cell lines was between 1.76 and 13.0 μM.Meanwhile,A36 did not display cytotoxicity in BEAS-2B cells(lung epithelial cells from a healthy human donor).Furthermore,A36 significantly induced apoptosis,arrested cells at the G0/G1 phase,and inhibited cell-colony formation in HeLa cells.In addition,mRNA deep sequencing showed that treatment with A36 could generate 171 dysregulated mRNAs in HeLa cells,while the expression of miRNA-21 target gene dual-specificity phosphatase 5(DUSP5)was significantly upregulated at both the mRNA and protein levels.Collectively,these findings demonstrated that A36 is a novel miRNA-21 inhibitor.

Objective:To disclose the genome characteristics and mutations of 2019 novel coronavirus (2019-nCoV) strains from the imported cases of Coronavirus Disease 2019 (COVID-19) in Gansu province, thereby to provide scientific reference for the prevention and control of COVID-19 in Gansu province. Methods:The respiratory tract specimens of imported COVID-19 cases from seven countries in Gansu province in 2020 were collected. The virus genome was sequenced by the second-generation sequencing technology, the whole genome sequences were compared and analyzed, and the MEGA software was used to construct a phylogenetic tree based on the neighbor-joining method.Results:A total of 46 2019-nCoV genome sequences with a length of 29 605~29 903 bp were obtained. Compared with the Wuhan reference strain (GenBank ID: NC_045512.2), it was found that the median (minimum to maximum) number of the nucleotide mutations of the 2019-nCoV genome sequence of the imported cases was 10 (7-24). A total of 134 nucleotide mutation sites were found in all 2019-nCoV genome sequences from 7 entry countries in Gansu province, distributed in 11 open reading frames (ORFs). The top three nucleotide mutations in different proteins: ORF1ab (78), S(20), N(12). Among the 134 nucleotide mutations, 82 caused amino acid mutations, and all of them were missense mutations. No insertions or deletions were seen. Types of deletion mutations, the top three amino acid mutations in different proteins: ORF1ab (46), S(11), N(10); the key sites of the receptor binding domain (RBD) of the S protein have not been mutated.Conclusions:No imported cases in Gansu province have been found to carry the reported mutations that can clearly lead to changes in the spread and pathogenicity of 2019-nCoV.

Objective:To evaluate the impact of assisted reproductive technology (ART) on postpartum breastfeeding outcomes.Methods:This paper electronically retrieved the China Biology Medicine disc, China National Knowledge Infrastructure, VIP, WanFang Data, PubMed, Embase, Web of Science, and Cochrane Library. The search period was from database establishment to March 15, 2023. After independent literature search, screening, data extraction, and quality evaluation by two researchers, Meta-analysis was conducted using R 4.2.2 software.Results:A total of 11 articles were included. Meta-analysis showed that compared with naturally conceived mothers, the rates of exclusive breastfeeding at 1th week postpartum ( RR=0.84, 95% CI: 0.73-0.97), exclusive breastfeeding at 6th months postpartum ( RR=0.77, 95% CI: 0.61-0.98), and the incidence of breastfeeding for >6 months postpartum ( RR=0.71, 95% CI: 0.53-0.96) were decreased, and the rate of artificial feeding at 12th months postpartum ( RR=1.09, 95% CI: 1.02-1.17) was increased. However, there were no statistically significant differences in the rate of artificial feeding at 8th months postpartum, the incidence of breastfeeding duration >12 months, and the incidence of breastfeeding difficulties ( P>0.05) . Conclusions:ART reduces the rate of exclusive breastfeeding in postpartum 1th week and 6th months, and the incidence of postpartum breastfeeding duration>6 months, and increases the artificial feeding rate in postpartum 12th months. However, the impact of ART on the incidence of breastfeeding difficulties is not yet clear and still needs to be further demonstrated by high-quality studies.