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As the second largest vector of infectious diseases, ticks carry and transmit various pathogens that cause human infections and pose a serious public health hazard. In recent years, there has been an ongoing epidemic of tick-borne fever with thrombocytopenia syndrome virus (SFTSV), as well as the occurrence of human infections by brand-new viruses such as Jingmen tick virus (JMTV) and Alongshan virus (ALSV) in China. This paper will review the advancement of disease clinical characteristics, laboratory methods of virus isolation, immunology and molecular biology of these emerging tick-borne viral diseases in China.
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In recent years, with the change of natural and social factors, such as climate warming, urbanization, land reclamation, human population growth, change of life customs, and convenient transportation, the human infectious diseases transmitted by insect vectors, well known as insect-borne infectious diseases, has increased significantly, causing a serious threat to public health. This paper focuses on the epidemiology, clinical characteristics, especially laboratory diagnosis of insect-borne infectious diseases, and emphasizes that medical institutions should pay attention to the rapid and accurate laboratory diagnosis of insect-borne infectious diseases. Metagenomic next generation sequencing has potential value in the diagnosis of insect-borne infectious diseases with unknown causes.
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Objective:To investigate the difference and characteristics of autoantibodies expression in patients infected by 2019-nCoV with various severity, and explore the associations between expression profile of autoantibodies and prognosis of COVID-19 patients.Methods:This retrospective study was conducted on patients with COVID-19 admitted to Zhongnan Hospital, Wuhan University from January 30, 2020 to March 16, 2020. Data on medical records, expression of autoantibodies including antinuclear antibody profile (ANA), anticardiolipin antibody (ACA), inflammatory factor and other laboratory indexes were collected and analyzed. The age and sex matched disease controls (cases of pulmonary infection unrelated to 2019-nCoV infection and autoimmune disease) and healthy controls (healthy check-up individuals) were also included. Following groups were established, ANA test groups: 72 cases of COVID-19 group (including 17 critical and severe cases, and 55 mild cases), 37 disease controls and 44 healthy controls; ACA test groups: 111 cases of COVID-19 group (including 37 critical and severe cases, and 74 mild cases), 37 disease controls and 40 healthy controls. The difference of positive rate or expression level of autoantibodies among various groups was analyzed, and the difference of inflammatory biomarkers and other parameters were compared between patients with ANA positive results and negative results. The Spearman correlation test was applied to determine the relationship between ACA and other parameters. Kaplan-Meier estimation was used to plot survival curves, the log-rank analysis was utilized to explore the association between antibodies and outcome of COVID-19 patients.Results:The positive rate of antibodies was significantly higher in the COVID-19 group than disease and healthy control groups, the ANA fluorescence: 22.22% (16/72), 5.41% (2/37), 6.82% (3/44); ANA spectrum:26.39% (19/72), 8.11% (3/37), 9.09% (4/44); and ACA:37.84% (42/111), 8.11% (3/37), 5.00% (2/40); all P<0.05. The positive rate of ANA, ACA-IgM and the expression level of ACA-IgM were significantly higher in severe COVID-19 subgroups (critically and severe COVID-19 patients) than in the mild COVID-19 patients (the ANA fluorescence: 47.06% [8/17] vs. 14.55% [8/55], ANA spectrum:66.67% [9/17] vs. 18.18% [10/55], ACA-IgM:30.43% [10/37] vs. 9.46% [7/74]; all P<0.05). There were significant differences in the number of red blood cells, hemoglobin concentration, hematocrit, activated partial thromboplastin time, C-reactive protein, interleukin-6 and serum amyloid A between COVID-19 ANA-positive group and COVID-19 ANA-negative group (all P<0.05). The level of ACA-IgM was positively correlated with white blood cell count ( r=0.354, P<0.001), neutrophil count ( r=0.344, P<0.001), platelet count ( r=0.198, P=0.038), D-Dimer ( r=0.260, P=0.009), glutamic-pyruvic transaminase ( r=0.214, P=0.024), γ-glutamyl transpeptidase ( r=0.283, P=0.003), blood urea nitrogen ( r=0.223, P=0.019), and negatively correlated with superoxide dismutase ( r=-0.228, P=0.020). Survival analysis showed that cumulative survival rate of event-free survival (EFS) was lower in patients with positive ANA/ACA-IgM results than in patients with negative ANA/ACA-IgM results ( P<0.05). Conclusions:ANA and ACA autoantibodies can be detected in COVID-19 patients. The positive rate and the expression level of ANA and ACA increase in proportion with the severity of COVID-19 patients. ANA and ACA-IgM could be used as risk stratification determinants for predicting survival of COVID-19 patients.
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The pandemic of 2019 novel coronavirus (2019-nCoV) infection since 2020 caused Coronavirus Disease 2019 (COVID-19) leads the serious threaten to global public health. It is urgent to diagnose COVID-19, guide epidemiological measures, control the infection rates, research/develop the antiviral treatment and promote the vaccine research. The application of nano-material based biosensors (the nano-biosensors) has achieved the high-performance detection of a variety of biomarkers due to their small device size, label free detection, high sensitivity, good specificity, short detection time, and has been considered as great potential to become a point-of-care testing tool for detecting 2019-nCoV. Therefore, by summarizing the working principle and classification of nano-biosensors, and focusing on the research progress of nano-biosensors in the detection of 2019-nCoV reported in the recent years, our review provides the challenges and future development prospects of the nano-biosensor in clinical laboratory.
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SARS-CoV-2 and its variants are raging worldwide. Unfortunately, the global vaccination is not efficient enough to attain a vaccine-based herd-immunity and yet no special and effective drug is developed to contain the spread of the disease. Previously we have identified CD147 as a novel receptor for SARS-CoV-2 infection. Here, we demonstrated that CD147 antibody effectively inhibits infection and cytokine storm caused by SARS-CoV-2 variants. In CD147KO VeroE6 cells, infections of SARS-CoV-2, its variants (B.1.1.7, B.1.351) and pseudovirus mutants (B.1.1.7, B.1.351, B.1.525, B.1.526 (S477N), B.1.526 (E484K), P.1, P.2, B.1.617.1, B.1.617.2) were decreased. Meanwhile, CD147 antibody effectively blocked the entry of variants and pseudomutants in VeroE6 cells, and inhibited the expression of cytokines. A model of SARS-CoV-2-infected hCD147 transgenic mice was constructed, which recapitulated the features of exudative diffuse alveolar damage and dynamic immune responses of COVID-19. CD147 antibody could effectively clear the virus and alveolar exudation, resolving the pneumonia. We found the elevated level of cyclophilin A (CyPA) in plasma of severe/critical cases, and identified CyPA as the most important proinflammatory intermediate causing cytokine storm. Mechanistically, spike protein of SARS-CoV-2 bound to CD147 and initiated the JAK-STAT pathway, which induced expression of CyPA. CyPA reciprocally bound to CD147, triggered MAPK pathway and consequently mediated the expression of cytokine and chemokine. In conclusion, CD147 is a critical target for SARS-CoV-2 variants and CD147 antibody is a promising drug to control the new wave of COVID-19 epidemic.
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Autoantibodies play an important role in aiding to the diagnosis, as well as judging diseases activity and monitoring treatment of autoimmune disorders. The combined detection of multiple autoantibodies is the current mainstream detection method. However, results are frequently reported as qualitative or semi-quantitative. The newly released diagnostic/classification standards for autoimmune diseases require quantitative detection data about the level of autoantibodies. The higher the level of autoantibodies is, the more likely the patient is to be diagnosed as an autoimmune disease, and it may be more relevant to the activity of the autoimmune disease. This article makes an introduction about the clinical value of accurate quantitative detection of autoantibodies as well as the current statues and challenges at the moment.
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Since the outbreak of the novel coronavirus pneumonia (COVID-19), it is urgently to develop the effective strategies for its clinical test or treatments world widely. Extracellular vesicles (EV)/exosomes could function as effective carriers for intercellular communication. Increasing evidence revealed that mesenchymal stem cell-derived EV/exosomes could be considered as an alternative cell-free therapy against the acute respiratory distress syndrome. Moreover, the EV-related metabolomics, proteomics, relapsing, deep-vein-thrombosis, myocardial-toxicity, liquid-biopsy and bio-therapeutic researches focusing on the COVID-19 will not only extended our knowledge for the diagnosis, but also provide novel ideas for treatment of this fatal disease. This article will systemically review the progresses of EV/exosomes in the diagnosis and treatment of COVID-19.
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BackgroundNovel coronavirus (SARS-CoV-2) infects human lung tissue cells through angiotensin-converting enzyme-2 (ACE2), and the body sodium is an important factor for regulating the expression of ACE2. Through a systematic review, meta-analysis and retrospective cohort study, we found that the low blood sodium population may significantly increase the risk and severity of SARS-CoV-2 infection. MethodsWe extracted the data of serum sodium concentrations of patients with COVID-19 on admission from the articles published between Jan 1 and April 28, 2020, and analyzed the relationship between the serum sodium concentrations and the illness severity of patients. Then we used a cohort of 244 patients with COVID-19 for a retrospective analysis. ResultsWe identified 36 studies, one of which comprised 2736 patients.The mean serum sodium concentration in patients with COVID-19 was 138.6 mmol/L, which was much lower than the median level in population (142.0). The mean serum sodium concentration in severe/critical patients (137.0) was significantly lower than those in mild and moderate patients (140.8 and 138.7, respectively). Such findings were confirmed in a retrospective cohort study, of which the mean serum sodium concentration in all patients was 137.5 mmol/L, and the significant differences were found between the mild (139.2) and moderate (137.2) patients, and the mild and severe/critical (136.6) patients. Interestingly, such changes were not obvious in the serum chlorine and potassium concentrations. ConclusionsThe low sodium state of patients with COVID-19 may not be the consequence of virus infection, but could be a physiological state possibly caused by living habits such as low salt diet and during aging process, which may result in ACE2 overexpression, and increase the risk and severity of COVID-19. These findings may provide a new idea for the prevention and treatment of COVID-19.
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An outbreak of new coronavirus SARS-CoV-2 was occurred in Wuhan, China and rapidly spread to other cities and nations. The standard diagnostic approach that widely adopted in the clinic is nuclear acid detection by real-time RT-PCR. However, the false-negative rate of the technique is unneglectable and serological methods are urgently warranted. Here, we presented the colloidal gold-based immunochromatographic (ICG) strip targeting viral IgM or IgG antibody and compared it with real-time RT-PCR. The sensitivity of ICG assay with IgM and IgG combinatorial detection in nuclear acid confirmed cases were 11.1%, 92.9% and 96.8% at the early stage (1-7 days after onset), intermediate stage (8-14 days after onset), and late stage (more than 15 days), respectively. The ICG detection capacity in nuclear acid-negative suspected cases was 43.6%. In addition, the consistencies of whole blood samples with plasma were 100% and 97.1% in IgM and IgG strips, respectively. In conclusion, serological ICG strip assay in detecting SARS-CoV-2 infection is both sensitive and consistent, which is considered as an excellent supplementary approach in clinical application.
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ImportanceA large number of healthcare workers (HCWs) were infected by SARS-CoV-2 during the ongoing outbreak of COVID-19 in Wuhan, China. Hospitals are significant epicenters for the human-to-human transmission of the SARS-CoV-2 for HCWs, patients, and visitors. No data has been reported on the details of hospital environmental contamination status in the epicenter of Wuhan. ObjectiveTo investigate the extent to which SARS-CoV-2 contaminates healthcare settings, including to identify function zones of the hospital with the highest contamination levels and to identify the most contaminated objects, and personal protection equipment (PPE) in Wuhan, China. DesignA field investigation was conducted to collect the surface swabs in various environments in the hospital and a laboratory experiment was conducted to examine the presence of the SARS-CoV-2 RNA. SettingSix hundred twenty-six surface samples were collected within the Zhongnan Medical Center in Wuhan, China in the mist of the COVID-19 outbreak between February 7 - February 27, 2020. ParticipantsDacron swabs were aseptically collected from the surfaces of 13 hospital function zones, five major objects, and three major personal protection equipment (PPE). The SARS-CoV-2 RNAs were detected by reverse transcription-PCR (RT-PCR). Main Outcomes and MeasuresSARS-CoV-2 RNAs ResultsThe most contaminated zones were the intensive care unit specialized for taking care of novel coronavirus pneumonia (NCP) (31.9%), Obstetric Isolation Ward specialized for pregnant women with NCP (28.1%), and Isolation Ward for NCP (19.6%). We classified the 13 zones into four contamination levels. The most contaminated objects are self-service printers (20.0%), desktop/keyboard (16.8%), and doorknob (16.0%). Both hand sanitizer dispensers (20.3%) and gloves (15.4%) were most contaminated PPE. Conclusions and RelevanceMany surfaces were contaminated with SARS-CoV-2 across the hospital in various patient care areas, commonly used objects, medical equipment, and PPE. The 13 hospital function zones were classified into four contamination levels. These findings emphasize the urgent need to ensure adequate environmental cleaning, strengthen infection prevention training, and improve infection prevention precautions among HCWs during the outbreak of COVID-19. The findings may have important implications for modifying and developing urgently needed policy to better protect healthcare workers during this ongoing pandemic of SARS-CoV-2. Key PointsO_ST_ABSQuestionC_ST_ABSWhat was the hospital setting contamination status, the most contaminated objects and PPE of SARS-CoV-2 during the outbreak of COVID-19 in Wuhan, China? FindingsThe most contaminated zones were the intensive care unit for novel coronavirus pneumonia (NCP) (31.9%), Obstetric Isolation Ward specialized for pregnant women with NCP (28.1%), and Isolation Ward for NCP (19.6%). The most contaminated objects and PPE are self-service printers (20.0%), hand sanitizer dispensers (20.3%), and gloves (15.4%). MeaningThe findings may have important implications for modifying and developing urgently needed policy to better protect healthcare workers during this ongoing pandemic of SARS-CoV-2.
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BackgroundThe clinical presentation of 2019 Novel Coronavirus (2019-nCov) infected pneumonia (NCIP) resembles that of other etiologies of community-acquired pneumonia (CAP). We aimed to identify clinical laboratory features to distinguish NCIP from CAP. MethodsWe compared the ability of the hematological and biochemical features of 84 patients with NCIP at hospital admission and 316 patients with CAP. Parameters independently predictive of NCIP were calculated by multivariate logistic regression. The receiver operating characteristic (ROC) curves were generated and the area under the ROC curve (AUC) was measured to evaluate the discriminative ability. ResultsMost hematological and biochemical indexes of patients with NCIP were significantly different from patients with CAP. Nine laboratory parameters were identified to be highly predictive of a diagnosis of NCIP by multivariate analysis. The AUCs demonstrated good discriminatory ability for red cell distribution width (RDW) with an AUC of 0.88 and Hemoglobin (HGB) with an AUC of 0.82. Red blood cell (RBC), albumin (ALB), eosinophil (EO), hematocrit (HCT), alkaline phosphatase (ALP), and white blood cell (WBC) had fair discriminatory ability. Combinations of any two parameters performed better than did the RDW alone. ConclusionsRoutine laboratory examinations may be helpful for the diagnosis of NCIP. Application of laboratory tests may help to optimize the use of isolation rooms for patients when they present with unexplained febrile respiratory illnesses.
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Objective:In view of the shortage of new coronavirus nucleal acid test in the early COVID-19 outbreak, the application value of chest CT in screening COVID-19 patients was explored.Methods:Retrospective analysis was performed on the data of patients with fever who received chest CT and new coronavirus nucleal acid test during January 25, 2020 to February 2, 2020 in Zhongnan Hospital of Wuhan University. A total of 587 patients were enrolled, including 290 males and 297 females, aged from 11.0 to 96.0 (51.3±17.1) years old. Taking the nucleic acid test results as the gold standard, the sensitivity, specificity and rate of misdiagnosis of CT screening were calculated.Results:Among the 587 patients, there were 433 positive cases (73.8%, 433/587) and 154 negative cases (26.2%, 154/587) of novel coronavirus nucleic acid test. Using CT screening, 494 cases (84.2%, 494/587) were positive and 93 cases (15.8%, 93/587) were negative. The sensitivity of CT screening was 97.7% (423/433), specificity was 53.9% (83/154) and rate of misdiagnosis was 2.3% (10/433).Conclusions:In the early COVID-19 outbreak, CT screening has the advantages of high sensitivity and low rate of misdiagnosis, which can compensate for the shortage of new coronavirus nucleal acid test and can be used as a rapid screening for early prevention and control.
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Massive open online courses (MOOC) is an emerging teaching mode based on website, which has been developed rapidly in China since 2013. Contemporary medical education, assisted by MOOC, diverges greatly with various modes. In this paper, the current situation and advantages of MOOC on medical education in the information age were analyzed, and suggestions for improvement of MOOC on modern medical education were proposed in combination with foreign case study, which is conducive to the reform and innovation of medical education model.
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In December 2019, a cluster of patients with pneumonia of unknown cause were linked to a seafood wholesale market in Wuhan, China. Some studies found that the virus was a new kind of virus which had never been found in the human body. Then, the virus was named 2019 Novel Coronavirus (2019-nCoV) by the World Health Organization (WHO). 2019-nCoV nucleic acid detection is one of the essential indicators of NCP (Novel Coronavirus Pneumonia). Recently, some false-negative cases in China-Japan Friendship Hospital and Hangzhou Hospital led the clinical doctors to question the value of the nucleic acid detection. In this paper, more than 3 000 results of 2019-nCoV detection in Zhongnan Hospital, Wuhan University were analyzed. Attention should be paid to the root cause of false-negative results and the related countermeasures should be taken.
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Objective@#In view of the difficulty of the shortage of new coronavirus nucleal acid test in the early COVID-19 outbreak, to explore the application value of chest CT in screening COVID-19 patients.@*Methods@#Retrospective analysis was performed on the data of patients with fever who received chest CT and new coronavirus nucleal acid test during January 25, 2020 to February 2, 2020 in Zhongnan Hospital of Wuhan University. A total of 587 patients were enrolled, including 290 males and 297 females, aged from 11.0 to 96.0 (51.3±17.1) years old. Take the nucleic acid test results as the gold standard, the sensitivity, specificity and rate of missed diagnosis of CT screening COVID-19 were calculated.@*Results@#Among the 587 patients, there were 433 positive cases (73.8%, 433/587) and 154 negative cases (26.2%, 154/587) of novel coronavirus nucleic acid test. Using CT screening, 494 cases (84.2%, 494/587) were positive and 93 cases (15.8%, 93/587) were negative. The sensitivity of CT screening COVID-19 was 97.7% (423/433), specificity was 53.9% (83/154) and rate of missed diagnosis was 2.3% (10/433).@*Conclusions@#In the early COVID-19 outbreak, CT screening has the advantages of high sensitivity and low rate of missed diagnosis of COVID-19, which can compensate for the shortage of new coronavirus nucleal acid test and can be used as the basis for rapid screening for early prevention and control of COVID-19 outbreak.
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Objective:The matria-assisted laser desorption/ionization time of flight mass spectrometry(MALDI-TOF MS) and sequencing methods were performed to assess the methodology (biochemistry methods) for identifying the Pandoraea sputorum and provide the more preferred approach to identify Pandoraea species. Methods:This paper is a study on performance evaluation of identification method. Ten lines of Pandoraea sputorum were isolated from blood cultures of inpatients were collected at Zhongnan Hospital of Wuhan University from July to October 2018 and were confirmed by the cultural characteristics, colonial morphology and Gram′s stain. Further identification was carried out by using the manual biochemical method (API 20NE), automatic biochemistry systems(BioMerieuxVITEK 2 Compact, BD Phoenix-100andThemo ARIS 2X), MALDI-TOF MS (BioMerieuxVITEK MS and Bruker MALDI Biotyper) and the sequencing methods of the 16S rRNA to identify the Pandoraea sputorum. Results:Pandoraea sputorum was non-fermented gram-negative bacteria that are non-motile, oxidase positive, and catalase positive. Ten lines of Pandoraea sputorum were identified as Achromobacter denitrificans, Alcaligenes faecalis or Cupriavidus pauculus and the accuracy rate of genus and species identification was 0 by API 20NE. Among all the samples, six lines were identified as the Pandoraea spp. with the accuracy rate of genus identification was 6/10 by VITEK 2 Compact; whereas the other four lines were identified as the Burkholderia cepacia, Sphingomonas paucimobilis, Ralstonia pickettii, or "Low Discrim" . All of these were identified as "No Identification" by Phoenix-100, which the accuracy rate of genus and species identification was 0. Seven isolates were identified by ARIS 2X as Stenotrophomonas maltophilia, Acinetobacter lwoffii, Sphingomonas paucimobilis, Pseudomonas luteola, Acinetobacter baumannii/Acinetobacter haemolyticus; whereas the other three lines were identified as rare species, thus, the accuracy rate of genus and species identification was 0. Both VITEK MS and MALDI Biotyper indicated all the isolates were Pandoraea sputorum with the accuracy rate of genus and species identification was 10/10. 16S rRNA sequencing for the 10 isolates showed they have 100% of similarity to Pandoraea sputorum by blasting with Genebank. Conclusions:Methods based on biochemical reactions often failed to accurately identify the Pandoraea sputorum to species level. MALDL-TOF MS and 16S rRNA sequencing technology identify Pandoraea sputorum efficiently and precisely enough.
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Isothermal amplification technology is a new class of nucleic acid amplification technology that performs amplification under constant temperature conditions. With the advantages of simple operation, high sensitivity and specificity, this kind of technology shows a good application prospect in clinical detection and scientific research. The principle, characteristics and applications of loop-mediated isothermal amplification(LAMP), crossing priming amplification (CPA),strand displacement amplification(SDA), recombinase polymerase amplification (RPA), nucleic acid sequence-based amplification (NASBA), rolling circle amplification (RCA) and helicase-dependent amplification (HDA) are reviewed in this paper.
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In December 2019, a cluster of patients with pneumonia of unknown cause were linked to a seafood wholesale market in Wuhan, China. Some studies found that the virus was a new kind of virus which had never been found in the human body. Then, the virus was named 2019 novel coronavirus (2019-nCoV) by the World Health Organization (WHO). 2019-nCoV nucleic acid detection is one of the essential indicators of COVID-19. Recently, some false-negative cases in China-Japan Friendship Hospital and Hangzhou Hospital led the clinical doctors to question the value of the nucleic acid detection. In this paper, more than 3 000 results of 2019-nCoV detection in Zhongnan Hospital, Wuhan University were analyzed. Attention should be paid to the root cause of false-negative results and the related countermeasures should be taken.
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Objective To compare the clinical outcomes of gefitinib and erlotinib treating non-small-cell lung cancer (NSCLC)with epidermal growth factor receptor (EGFR)mutation in either exon 19 or 21 . Methods A total of 242 patients diagnosed as NSCLC with EGFR mutation in either exon 19 or 21 from May 201 3 to December 2014 in our hospital were chosen in this study.According to age,sex,smoking history,eastern cooperative oncology group performance status and types of EGFR mutation,all the patients were matched to 121 pairs,and randomly divided into group A and B.Patients in group A received gefitinib treatment,and those in group B received erlotinib treatment.Based on the response evaluation criteria in solid tumors (RECIST),overall response rate (ORR),disease control rate (DCR),progression-free survival (PFS)were assessed.To assess the independent risk factors for PFS by univariate and multivariate Cox regression analysis.The subgroup analysis was performed for the 63 NSCLC patients using these two drugs as the first-line treatment.To evaluate the adverse drug reactions and quality of life between A and B groups.Results The median PFS of group A and B were 11 .6 months and 9.5 months,respectively,with no significant difference (HR =0.39,P >0.05).The ORR and DCR in the two groups were 76.9%,74.4% (χ2 =1 .03,P =0.58)and 90.1 %,86.8% (χ2 =1 .46,P =0.31 ). The independent risk factors of poor PFS were ECOG PS≥2 (HR =2.60,95%CI:1 .54 -4.43,P =0.001 )and non-adenocarcinoma (HR =3.61 ,95%CI:1 .54-8.66,P =0.003).For patients receiving these two drugs as the first-line treatment,there was no significant difference between two groups in overall response rates (76.6% vs. 90.2%,χ2 =0.83,P =0.12)and median PFS (11 .6 months vs.14.4 months,HR =0.59,P >0.05).The adverse drug reactions were significant differences in emotion function (F =10.27,P =0.03),diarrhea (F =10.24,P =0.03)and pain (F =9.02,P =0.04).After receiving drug treatment,the quality of life scores were improved,and most of the differences were statistically significant between A and B groups(P <0.05). Conclusion As for NSCLC with EGFR mutation in either exon 1 9 or 21 ,both gefitinib and erlotinib are well tolerated and have similar clinical effectiveness.
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Objective To investigate the relation between a set of single nucleotide polymorphisms (SNP) in core gene of HBV in chronic hepatitis B patients and HBV DNA levels. Methods PCR restriction fragment length polymorphism(PCR-RFLP) assay and restriction enzyme Tsp509I were adopted to determine HBV SNP in HBV core gene. Nucleotide sequences of core gene were determined using the dideoxy chain termination method. HBV DNA levels were quantitated with real-time PCR. Results Five typical RFLP patterns, RFLP-C, RFLP-D, RFLP-E, RFLP-G and RFLP-C/G mixture were found and the distribution of HBV RFLP patterns was as follows: C, 61. 5% ; D, 2. 6% ; E, 9.6%; G, 16.7%; C/G mixture, 9.6%. Five SNPs, A165T, A336C, A336T, T337C and T385C, were found to be associated with RFLP patterns change and only SNP A336C or A336T caused the substitution of Glu-83 with Asp in HBcAg. The serum HBV DNA levels in RFLP-C group were higher than that in RFLP-G (P =0. 02) and RFLP-C/G group(P = 0. 006) , respectively, furthermore, the positive rate of serum ALT in RFLP-C/G group was lower than that in RFLP-C, RFLP-E and RFLP-G group, respectively. Under the condition of HBeAg-positive, the serum HBV DNA levels in RFLP-C group were higher than that in RFLP-G (P = 0. 015) and RFLP-C/G group(P =0.008) , respectively. Conclusion PCR-RFLP used in this study can be adopted to determine HBV SNPs, not genotypes in Chinese patients with chronic hepatitis B. The serum HBV DNA level in RFLP-C group higher than that in RFLP-G or RFLP-C/G group maybe associated with amino acid mutation, Glu83 Asp.
