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@#Lung adenocarcinoma has become the most common type of lung cancer. According to the 2015 World Health Organization histological classification of lung cancer, invasive lung adenocarcinoma can be divided into 5 subtypes: lepidic, acinar, papillary, solid, and micropapillary. Relevant studies have shown that the local lobectomy or sublobectomy is sufficient for early lepidic predominant adenocarcinoma, while lobectomy should be recommended for tumors containing micropapillary and solid ingredients (≥5%). Currently, the percentage of micropapillary and solid components diagnosed by frozen pathological examination is 65.7%, and the accuracy of diagnosis is limited. Therefore, to improve the accuracy of diagnosis, it is necessary to seek new methods and techniques. This paper summarized the characteristics and rapid diagnosis tools of early lung adenocarcinoma subtypes.
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The standardization and validation of a multiplex assay requires the combination of important parameters such as sensitivity and specificity, acceptable levels of performance, robustness, and reproducibility. We standardized a multiparametric Dot-blot aimed at the serological screening of paracoccidioidomycosis, histoplasmosis, and aspergillosis. A total of 148 serum were evaluated: 10 from healthy subjects, 36 from patients with paracoccidioidomycosis, 62 from patients with histoplasmosis, and 40 from patients with aspergillosis. It was found that the multiparametric Dot-blot showed a high percentage of cross-reactivity. However, when evaluated individually, in the serological screening of histoplasmosis, a good performance was observed when compared to the double immunodiffusion assay, considered the gold standard test, with 100% co-positivity and 83.3% co-negativity. The performance of serological screening for aspergillosis was not satisfactory when compared to double immunodiffusion, showing 71.4% co-positivity and 100% co-negativity. The evaluation of the stability of nitrocellulose membranes showed that membranes sensitized with H. capsulatum antigen remained stable for 90 days and those sensitized with A. fumigatus antigen for 30 days. We conclude that the use of crude antigens was not suitable for the standardization of the multiparametric Dot-blot assay, due to the high cross-reactivity, and that further tests should be performed with purified proteins (AU).
A padronização e validação de um ensaio multiplex requer a combinação de parâmetros importantes, como sensibilidade e especificidade, níveis aceitáveis de desempenho, robustez e reprodutibilidade. Este trabalho padronizou um Dot-blot multiparamétrico visando a triagem sorológica da paracoccidioidomicose, histoplasmose e aspergilose. Foram avaliadas 148 amostras de soro: 10 de indivíduos saudáveis, 36 de pacientes com paracoccidioidomicose, 62 de pacientes com histoplasmose e 40 de pacientes com aspergilose. Verificou-se que o Dot-blot multiparamétrico apresentou elevado percentual de reatividade cruzada. Entretanto, quando avaliado individualmente, na triagem sorológica da histoplasmose observou-se bom desempenho quando comparado ao ensaio de imunodifusão dupla, considerado o teste padrão ouro, com 100% de co-positividade e 83,3% de co-negatividade. O desempenho da triagem sorológica da aspergilose não foi satisfatório quando comparado a imunodifusão dupla, apresentando 71,4% de co-positividade e 100% de co-negatividade. A avaliação da estabilidade das membranas de nitrocelulose mostrou que membranas sensibilizadas com antígeno de H. capsulatum permaneceram estáveis por 90 dias e as sensibilizadas com antígeno de A. fumigatus, por 30 dias. Concluímos que o uso de antígenos brutos não foi adequado para a padronização do ensaio de Dot-blot multiparamétrico, devido ao alto índice de reatividade cruzada, e que novos testes devem ser realizados com proteínas purificadas (AU).
Subject(s)
Paracoccidioidomycosis , Aspergillosis , Reference Standards , Immunologic Tests , Public Health , Methodology as a Subject , Histoplasmosis , Mycoses/diagnosisABSTRACT
Serological techniques can detect antibodies against Sarcocystis spp., Neospora caninum and Toxoplasma gondii antigens in single or mixed infections. Immunofluorescent antibody tests (IFAT) is considered the gold standard technique for Sarcocystosis diagnostic in cattle serum and a positive IFAT result reflects Sarcocystis spp. infection. Therefore, the aims of the present study were to compare IFAT and Dot-blot for sarcocystosis diagnostic in experimentally infected mice and to investigate serological cross-reactions with N. caninum and T. gondii in these methods. Mice (Mus musculus) were inoculated intraperitoneally with bradizoites of Sarcocystis spp. or tachyzoites of N. caninum or T. gondii. Serum samples were obtained and analyzed by IFAT and Dot-blot for the three protozoa. Serum from N. caninum and T. gondii experimentally infected mice were tested by IFAT and reacted only to N. caninum or T. gondii antigens, respectively. Specific antibodies against Sarcocystis spp. were present in all animals experimentally infected with this protozoan, with IFAT titers from 10 to 800. Serum samples from mice experimentally infected with Sarcocystis spp., N. caninum and T. gondii and tested by Dot-blot demonstrated no cross reaction between protozoa. A Dot-blot using Sarcocystis spp. antigen appears to be a good alternative to IFAT in the serological diagnosis of Sarcocystosis.(AU)
As técnicas sorológicas podem detectar anticorpos contra os antígenos de Sarcocystis spp., Neospora caninum e Toxoplasma gondii em infecções únicas ou mistas. O teste de anticorpos imunofluorescentes (IFAT) é considerado a técnica padrão-ouro para o diagnóstico de sarcocistose no soro de bovinos e um resultado positivo de IFAT reflete Sarcocystis spp. infecção. Portanto, os objetivos do presente estudo foram comparar IFAT e Dot-blot para diagnóstico de sarcocistose em camundongos infectados experimentalmente e investigar reações cruzadas sorológicas com N. caninum e T. gondii nesses métodos. Os camundongos (Mus musculus) foram inoculados intraperitonealmente com bradizoítos de Sarcocystis spp. ou taquizoítos de N. caninum ou T. gondii. As amostras de soro foram obtidas e analisadas por IFAT e Dot-blot para os três protozoários. O soro de N. caninum e T. gondii infectados experimentalmente foram testados por IFAT e reagiram apenas aos antígenos de N. caninum ou T. gondii, respectivamente. Anticorpos específicos contra Sarcocystis spp. estavam presentes em todos os animais experimentalmente infectados com este protozoário, com títulos de IFAT de 10 a 800. Amostras de soro de camundongos infectados experimentalmente com Sarcocystis spp., N. caninum e T. gondii e testadas por Dot-blot não demonstraram reação cruzada entre protozoários. Um Dot-blot usando Sarcocystis spp. O antígeno parece ser uma boa alternativa ao IFAT no diagnóstico sorológico da sarcocistose.(AU)
Subject(s)
Animals , Male , Mice , Cattle/parasitology , Serologic Tests/methods , Cattle Diseases , Sarcocystis , Sarcocystosis/diagnosis , Sarcocystosis/veterinary , Serologic Tests/veterinary , Fluorescent Antibody Technique, IndirectABSTRACT
Cercospora kikuchii, fitopatógeno usual en plantas de soja, ocasiona deterioro en la scosechas. Su identificación precoz y correcta evitaría el uso indebido de plaguicidas y permitiría iniciar un tratamiento adecuado. Una técnica rápida, económica y de fácil ejecución es el Dot blot, capaz de reconocer la presencia de una proteína específica del género conocida como CFP (Cercosporin Facilitator Protein). El objetivo de este trabajo fue validar dicha técnica para garantizar la fiabilidad del resultado. Para ello, se procesaron 29 plantas de soja infectadas y 31 plantas sanas teniendo en cuenta una confianza deseada del 95% y un error permitido del 5%. La técnica presentó una sensibilidad diagnóstica del 93,3% y una especificidad diagnóstica del 96,7%. La eficacia fue del 95% y los valores predictivos positivo y negativo del 96,6 y el 93,5%, respectivamente. Estos resultados la postulan como una herramienta útil para detectar precozmente c. kikuchii en plantas de soja.
Cercospora kikuchii is a common pathogen in soybean plants that causes crop spoilage. Its early and precise identification would prevent the misuse of pesticides and allow the initiation of an appropriate treatment. A quick, economical and easy-to-execute technique is the Dot blot, capable of recognizing the presence of a genus-specific protein called CFP (Cercosporin Facilitator Protein). The objective was to validate this technique to guarantee the reliability of the results. For that purpose, 29 infected soybean plants and 31 healthy plants were processed, taking into account a 95% desired confidence level and a permissible error of 5%. The technique provided a diagnostic sensitivity of 93.3% and a diagnostic specificity of 96.7%. The efficiency was 95% and positive and negative predictive values were 96.6% and 93.5%, respectively. These results postulate it as a useful resource for the early detection of C. kikuchii in soybean plants.
Subject(s)
Ascomycota , Soybeans , Ascomycota/isolation & purification , Soybeans/microbiology , Reproducibility of ResultsABSTRACT
Objective:To access the performance of the Tellgenplex human papillomavirus (HPV) DNA test compared to the polymerase chain reaction-reverse dot blot (PCR-RDB) assay for the HPV genotyping.Methods:Sixty cervical swab samples were genotyped by the Tellgenplex HPV DNA test and the PCR-RDB assay. The Tellgenplex HPV DNA test and the PCR-RDB assay can detect 26 and 23 HPV genotypes, respectively. Each sample showed discrepancy was genotyped using sequencing.Results:The percent agreement between the two tests ranged from 83.3% to 100.0% according to different genotype. This showed perfect agreement (>0.81) for high-risk HPV genotypes (35, 39, 45, 53, 56, 59, 66, 68, and 82), substantial agreement (>0.65) for high-risk HPV genotypes (16, 18, 33, 52, and 58) and low-risk HPV genotype 43 between the two assays by the kappa analysis. The positive rates of the two assays for frequent HPV genotypes (16, 35, 39, 45, 52, 53, 58, 59, 66, and 82) were not statistically different, but the PCR-RDB assay showed higher positive rates than the Tellgenplex HPV DNA test for HPV genotypes 81 (P<0.05). As for more than 10 positive results by the Tellgenplex HPV DNA test and/or the PCR-RDB assay, the PCR-RDB assay showed higher relative sensitivity and specificity than the Tellgenplex HPV DNA test for the three HPV genotypes (16, 52, and 81). All HPV genotypes that can be detected by only the Tellgenplex HPV DNA test (HPV genotypes 44 and 55) were confirmed by sequencing.Conclusions:In conclusion, our results demonstrated that the PCR-RDB assay which can detect more multiple HPV genotypes in each specimen shows higher relative sensitivity and specificity than the Tellgenplex HPV DNA test, which makes it a better option for routine clinical use.
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Objective To analyze the variation characteristics of rpoB,katG,inhA,rpsL and embB related genes of Mycobacterium tuberculosis(MTB)in Qinzhou,Guangxi.Methods PCR reverse point hybridization was used to detect 5 common resistance mutants of Mycobacterium tuberculosis in 237 MTB-DNA positive sputum samples.Results Among 237 cases of tuberculosis patients,72 cases(30.38%)were resistant to the four kinds of anti-TB drugs.The resistance mutation rate of rifampin,isoniazid and streptomycin was 2.53%, 13.92%,3.80%.The top 5 gene mutation detection loci of Mycobacterium tuberculosis were-15M,S531L and 43M.Conclusion The main drug-resistant strains are isoniazid resistance,and the mutation of inhA gene were the major one in Qinzhou,Guangxi.
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Objective To explore the application value of PCR－reverse dot blot hybridization (PCR－RDB) gene membrane chip technique in genetic diagnosis of hereditary non－syndrome deafness in children.Methods The blood samples(2 mL)were collected from 38 children with congenital deafness,excluding high－risk factors for deaf-ness at Dongguan Rehabilitation School,and then genomic DNA extracted.By using self－designed multiplex－PCR combined with PCR－RDB gene chip technology,20 hot－spot mutations of 4 pathogenic genes of common deafness in Chinese population were detected.Sanger sequencing was used as the gold standard to corroborate the positive samples. Results Among 38 subjects,deafness gene mutations were detected in 16 cases,with a detection rate of 42.11%,and they were all verified by family study.Among 16 cases,6 cases of GJB2 gene mutation(3 cases of homozygote,3 cases of heterozygous),4 cases of SLC26A4 mutation,2 cases of MTRNR (m.1555A＞G)mutation,4 cases of compound muta-tion,but none of GJB3 gene mutations.And their detection rates were 15.79%,10.53%,5.26%,10.53%,and 0,re-spectively.DNA samples from 16 children with deafness gene mutation were corroborated by Sanger sequencing,and the compliance rate was 100%.Conclusions For 20 hot－spot mutations of 4 common deafness pathogenic genes,the matc-hing PCR－RDB gene membroine chip technology was designed and the susceptible gene of congenital deafness children was detected.This technique has some advantages like high detection rate,fast,accurate and economical.It is an ideal method for gene screening on hereditary non－syndrome deafness children and has good clinical application prospects.
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Objective: To access the performance of the Tellgenplex human papillomavirus (HPV) DNA test compared to the polymerase chain reaction-reverse dot blot (PCR-RDB) assay for the HPV genotyping. Methods: Sixty cervical swab samples were genotyped by the Tellgenplex HPV DNA test and the PCR-RDB assay. The Tellgenplex HPV DNA test and the PCR-RDB assay can detect 26 and 23 HPV genotypes, respectively. Each sample showed discrepancy was genotyped using sequencing. Results: The percent agreement between the two tests ranged from 83.3% to 100.0% according to different genotype. This showed perfect agreement (>0.81) for high-risk HPV genotypes (35, 39, 45, 53, 56, 59, 66, 68, and 82), substantial agreement (>0.65) for high-risk HPV genotypes (16, 18, 33, 52, and 58) and low-risk HPV genotype 43 between the two assays by the kappa analysis. The positive rates of the two assays for frequent HPV genotypes (16, 35, 39, 45, 52, 53, 58, 59, 66, and 82) were not statistically different, but the PCR-RDB assay showed higher positive rates than the Tellgenplex HPV DNA test for HPV genotypes 81 (P<0.05). As for more than 10 positive results by the Tellgenplex HPV DNA test and/or the PCR-RDB assay, the PCR-RDB assay showed higher relative sensitivity and specificity than the Tellgenplex HPV DNA test for the three HPV genotypes (16, 52, and 81). All HPV genotypes that can be detected by only the Tellgenplex HPV DNA test (HPV genotypes 44 and 55) were confirmed by sequencing. Conclusions: In conclusion, our results demonstrated that the PCR-RDB assay which can detect more multiple HPV genotypes in each specimen shows higher relative sensitivity and specificity than the Tellgenplex HPV DNA test, which makes it a better option for routine clinical use.
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El polisacárido Vi (PsVi) de Salmonella Typhi es un antígeno T-independiente y ha demostrado ser protector en adultos jóvenes. Sin embargo, para aumentar la respuesta de anticuerpos y conferir propiedades T-dependientes al polisacárido, se ha conjugado a proteínas. Dentro de los controles exigidos por los organismos regulatorios para estas vacunas está la identidad antigénica de sus componentes y para eso se recomiendan el uso de técnicas de Resonancia Magnética Nuclear o técnicas serológicas. El objetivo del presente trabajo, fue establecer las condiciones óptimas de trabajo de un Dot Blot que permitiera determinar, rápidamente, la identidad de los antígenos en vacunas conjugadas contra S. Typhi. Para ello, se estudiaron los tiempos de incubación, las concentraciones óptimas de anticuerpo monoclonal (AcM) y del ingrediente farmacéutico activo (IFA), así como los volúmenes de aplicación óptimos para las IFAs y formulaciones vacunales, tanto para el PsVi como para el toxoide diftérico (TD). Los resultados mostraron que para la determinación de la identidad antigénica fueron suficientes 5 µL de muestras de los conjugados monovalentes en una dilución de 1/10 (vol/vol) e igual volumen para las formulaciones vacunales. Quedó demostrado que la concentración de 2,5 µg/mL para el AcM contra el PsVi y a 2 µg/mL para el AcM contra TD fueron suficientes para la determinación; mientras que los tiempos de incubación fueron ajustados a 15 min con incubación a 37 ºC. Como conclusión del trabajo se puede decir que quedaron establecidas las condiciones óptimas de trabajo para la determinación rápida de la identidad antigénica del PsVi y del TD presentes en IFA y formulaciones vacunales conjugadas(AU)
Vi polysaccharide from Salmonella Typhi is a T-independent antigen that has proven to be protective in young adults. However, it has been conjugated to proteins in order to confer T-dependent properties to the polysaccharide, and improving the antibody response. The regulatory agencies require knowing the identity of antigens included in vaccines. The Nuclear Magnetic Resonance spectroscopy and serological techniques are recommended. The aim of this work was to establish the optimal working conditions of a Dot Blot that would allow to determine quickly the identity of the antigens in conjugate vaccines against S. Typhi. The incubation times, optimum concentrations of monoclonal antibodies (MAb) and active pharmaceutical ingredient (API), as well as optimum application volumes for APIs and vaccine formulations were studied for both, PsVi and diphtheria toxoid (DT). It was proven that 5 µL of samples of the monovalent conjugates were sufficient at a dilution of 1/10 (vol/vol) and an equal volume for the vaccine formulations. It was demonstrated that the concentration of 2.5 µg/mL for the MAb against PsVi and 2 µg/mL for the MAb against DT were suitable. The incubation times were adjusted to 15 min with incubation at 37 ºC. It was established a simple and rapid method for the specific identification of PsVi and DT present in API and conjugate vaccines(AU)
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Salmonella typhi , Diphtheria Toxoid , Magnetic Resonance Spectroscopy/methods , Vaccines, ConjugateABSTRACT
Objective To understand the various HPV types and the factors influencing their distribution among women with normal cervical cytology in the Shanghai area, to provide basic data for cervical cancer prevention and vaccine use.Methods A questionnaire-based survey was conducted among 3 372 married women in the Shanghai area. A PCR reverse dot blot (PCR-RDB) method was adopted for HPV genotyping of cervical exfoliated cell samples from 3 206 women. Odds ratio (OR) for HPV infection were analyzed using logistic regression. Results Of the 3 206 women investigated, 669 (20.87%) were positive for HPV infection. The highest incidence of HPV infections was seen in the age groups of 55-59 years, 50-54 years and 45-49 years, with the rates of positive detection being 27.67%, 21.65%, and 21.55%, respectively. While 70.4% of the positive cases had a single infection, 29.6% showed multiple infections. In cases with multiple infections, double infection was predominant (20.63%). The top five high-risk gene types were HPV52(3.65%),HPV53(2.71%),HPV51(2.03%),HPV58(1.87%),and HPV16 (1.40%). The top three low-risk gene types were HPV81 (2.03%), HPV42 (1.43%), and HPV55 (1.31%). Among the women with HPV infections,there were 483 having medium-high-risk HPV infection.There were significant differences in the age,the age at first sexual activity,sexual activities per week,education,and alcohol consumption between HPV positive and HPV negative patients. Age stratification showed that the infection rates in the 55-59 years old group were significantly higher than that in the other age groups(χ2=15.349, P=0.000). Both single factor and multivariate non-conditional logistic regression analyses showed that higher education and the start of sexual activity at a later age were protective factors for medium-high-risk HPV infection,with regression coefficients of-0.165 and-0.08,respectively (P<0.01) in the multivariate analysis. The risk factors included age between 55- 59 years, menstrual status (menopause), sexual activity (≥3 times per week) and alcohol consumption. High-risk HPV infections also had the same risk factors,and the odds ratios were 1.558,1.275,and 1.678,respectively(P<0.01).However, 55-59 years of age and alcohol consumption are independent risk factors for medium-high risk HPV infection. Conclusions High-risk HPV in women of Shanghai is commonly caused by HPV52, HPV53, HPV51,HPV58,and HPV16.The high-risk group includes women who are 55-59 years old.While drinking is an independent risk factor for medium-high risk HPV infection, a moderate sex life and delayed age at first sex can reduce the risk of high-risk HPV infections.
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Objective To investigate the exfoliative cell HPV infection situation,genotype distibution and charactristics among female population in Kashgar area.Methods The cervical exfoliated cells specimens were collected from 1548 women,and 23 genotypes of HPV were detected by using the PCR reverse dot blot hybridization method.The HPV infection situation and its genotypes were analyzed.Results The HPV infection positive rate was 33.33% (485/1 548),in which the HPV-16 infection rate was 10.3%;HPV52 infection rate was 9.9%;HPV58 infection rate was 7.6%,HPV53 infection rate was 7.2%;the single subtype infection rate was was 66.8%,the multiple infection rate was 33.2%;the HPV infection rate was highest(43.3%) in female population aged 41-50 years old,which was extremely higher than that in other age groups(P<0.05).Conclusion The HPV infection among female population of Kashgar area is dominated by HPV16,the HPV infection rate is highest in female population aged aged 41-50 years old,which provides a theoretical basis for the HPV prevention and control work of related departments in Kashgar area.
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Objective To evaluate the application value of PCR-reverse dot blot hybridization in the identification of Candida and the detection of Candida albicans drug-resistant genes.Methods The vaginal secretion samples from 285 patients with candidal vaginitis and 50 healthy women were collected.The identification of Candida species and their drug susceptibility were detected by the bioMérieux Yeast identification cards and MIC method(Zhengzhou Antu kit),respectively.The identification of Candida species and the mutation of Candida albicans,drug-resistant genes were also detected by the Shenzheng Yaneng test kit(PCR-reverse dot blot hybridization).The drug-resistant genes were also identified by PCR and nucleic acid sequencing.Based on the culture identification,MIC method and nucleic acid sequencing as the contrast methods,the sensitivity,specificity and accuracy of PCR-reverse dot blot hybridization in the identification of Candida species and the mutation detection of Candida albicans drug-resistant genes were evaluated.Results Compared with the bioMérieux Yeast identification method,the sensitivity,specificity,positive predictive value,negative predictive value and total coincidence rate of PCR-reverse dot blot hybridization for detecting six kinds of Candida species,including Candida albicans,Candida glabrata,Candida tropicalis,Candida parapsilosis,Candida krusei and Candida guilliermondii,were above 95％,96％,96％,98％ and 97％,respectively.There was no significant difference in detecting six kinds of Candida species between the two methods (x2 =0.44,0,0,0,0 and 0,respectively,P ＞ 0.05),and there was good consistency between them (Kappa ＞ 0.9).Compared with the MIC method,the sensitivity,specificity,positive predictive value,negative predictive value and total coincidence rate of PCR-reverse dot blot hybridization for detecting the drug resistance of Candida albicans were 98％,88％,98％,88％ and 96％,respectively.There was no significant difference in detecting the drug resistance of Candida albicans between the two methods (x2 =0.17,P ＞ 0.05),and there was good consistency between them (Kappa ＞ 0.8).The results of PCR-reverse dot blot hybridization in detecting the mutation sites of six kinds of Candida albicans drug-resistant genes were 100％ of coincidence with that of the nucleic acid sequencing method.Conclusion The PCR-reverse dot blot hybridization has high consistency with the culture method and the nucleic acid sequencing method in the identification of Candida species and the mutation detection of Candida albicans drug-resistant genes,which is more early and rapid than the traditional detection methods,and may be applied to the auxiliary diagnosis of vulvovaginal candidiasis (VVC).
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Objective:To investigate the diagnostic value of different detection methods for Mycobacterium tuberculosis in bronchoalveolar lavage fluid (BALF) from patients with pulmonary tuberculosis.Methods:BALF from l00 patients in Changsha Central Hospital from January 2013 to December 2015 was collected.Among 100 patients,65 cases were clinically diagnosed as tuberculosis,and 35 cases served as control.BALF smear method,polymerase chain reaction (PCR) and membrane reverse dot blot (RDB) were used for synchronous detection of Mycobacterium tuberculosis.Results:The positive rates by BALF smear method,PCR and RDB were 43.08％,73.84％ and 92.31％,respectively (P＜0.05).Sensitivity,specificity,accuracy,and negative predictive value for BALF smear were 43.08％,88.57％,59.00％,and 45.59％,respectively;for PCR were 73.85％,100％,83.00％,and 67.31％,respectively;for RDB were 92.31％,100.00％,95.00％,and 87.50％,respectively.Conclusion:The technique of membrane RDB can not only accurately diagnose Mycobacterium tuberculosis,but also can rapidly and easily identify the resistance of Mycobacterium tuberculosis to streptomycin (SM),rifampicin (RFP) and isoniazid (INH) genotypes.It possesses high clinical value.
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Objective To evaluate the necessity and feasibility by using two different PCR-based techniques for prenatal diagnosis of thalassemia. Methods 509 specimens for prenatal diagnosis of thalassemia were detected respectively by single tube multiplex PCR(STMP),reverse dot blot(RDB)or probe melting curve assay(PMCA)for commonα-thalassemia orβ-thalassemia mutations in double-blind tests. Samples with different detection results were confirmed with DNA sequencing analysis. Prenatal diagnosis of thalassemia results were verified or followed up after birth. Results In detectingα-thalassemia andβ-thalassemia,there was one case in STMP + RDB and another case PMCA indicating differentiating results. The detection sensitivity of STMP + RDB was higher than that of PMCA,and its difference can be used as an indication for maternal blood contamination. Conclusion The two PCR methods with different principles were necessary and feasible for the prenatal diagnosis of thalassemia. The two methods were complementary to each other ,which can ensure the reliability of the prenatal diagnosis results and reduce the defects of single technique. It is worthy to be popularized in clinical application.
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Objective To investigate the infection status and genotype distribution of human papillomavirus (HPV) infection in Changzhou district ,and to provide a theoretical basis for the prevention ,development and clinical diagnosis and treatment of HPV . Methods From October 2015 to July 2016 ,1 718 cases of female cervical epithelial cells were collected ,and 28 kinds of gene typing were detected by PCR‐reverse dot blot hybridization .Results The infection rate:1 718 cases of women were collected ,the positive HPV infections were 34 .23% .The infection types :single infection rate was 23 .57% (405/1 718) .The high‐risk HPV subtype in‐fections accounted for 17 .17% (295/1 718) and the low‐risk HPV subtype infections accounted for 5 .18% (89/1 718) ,suspected high‐risk infection was 1 .22% (21/1 718) .Multiple infection rate was 10 .94% (188/1 718) .HPV52 was the most common infec‐tion among high‐risk HPV infection ,the positive rate was 16 .16% (95/588) .HPV61 was the most common infection among low‐risk HPV infection ,the positive rate was 4 .08% (24/588) .There was no significant difference between age and HPV positive rate . The 61-70 age group had the highest HPV multiple infection rate in all age groups .Conclusion The high HPV infection is ob‐served in Changzhou district ,among which single HPV52 infection and the high‐risk HPV infection are the most common infec‐tions .There is difference in HPV infection among different age groups .
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Objective To investigate the infection of 23 kinds of human papillomavirus ( HPV) subtypes in female cervical epithelium samples and to analyze their relationships with age and results of cyto-logic test in Haikou area. Methods A total of 4 037 local healthy women were enrolled in this study from July 2013 to August 2016, 1 967 of whom received cervical cytology test. Cervical cell samples collected from those women were detected for HPV typing by using PCR-reverse dot blot hybridization. Results (1) The total positive rate of HPV in 4 037 samples was 22. 15% (894 cases), and the detection rates of carci-nogenic, possibly carcinogenic and non-carcinogenic HPV were 16. 13%, 3. 99% and 5. 55% (651, 161 and 224 cases), respectively. The positive rates of 6 genotypes were high, which were HPV52, 53, 81, 51, 16 and 58 in turn. (2) The detection rates of carcinogenic, possibly carcinogenic HPV and some of the gen-otypes (HPV18, 52, 53, 66) increased with age (all P<0. 05). (3) Multiple infection in HPV-positive women accounted for 24. 38% (218/894), in which the infection rates of carcinogenic types declined with age and the numbers of HPV genotypes in carcinogenic infections were negatively correlated with age ( both P<0. 05). (4) Only 2. 49% (49/1 967) of the samples were positive for cervical cytologic test, classified into the ≥ASC-US ( atypical squamous cells of undetermined significance ) group. The detection rates of eight kinds of carcinogenic and two kinds of possibly carcinogenic HPV in≥ASC-US group were significantly higher than those in the negative (no intraepithelial lesion or malignant lesion, NILM) group (all P<0. 05). Conclusion This study indicates that Haikou women have higher rates of HPV infection, especially the eld-erly group. HPV subtype infections present some regional characteristics and are mainly single type infec-tion. Cervical cancer screening should be combined with tests for HPV and cytology analysis to improve its effectiveness.
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Objective To analyze the genotypes and age distributions of human papillomavirus in affiliated hospital of Chifeng u‐niversity ,to provide theory basis for the screening and prevention of cervical cancer in Chifeng area .Methods The cervical exfolia‐ted cell specimens from 1 368 gynecological outpatients and female inpatients in affiliated hospital of Chifeng university were collect‐ed from January to June 2016 .Using PCR‐reverse dot blot technology ,25 HPV subtypes was performed ,a statistical analysis was conduted combing with age .Results Among 1 368 specimens ,546 specimens were found with positive HPV ,the positive rate was 39 .91% .The 24 genotypes were detected .The top three subtypes of HPV infection were HPV16(12 .08% ) ,HPV 58(8 .05% )and HPV 52(7 .61% ) ,HPV73 genotype was not detected .Masculine gender rate in the groups with various age had significance differ‐ence after the chi‐square test ,among different age groups ,the prevalence rate was the highest in the patients with higher than 56 years old(52 .5% ) ,followed by the rate in patients with lower than 25 years old(52 .38% ) .Conclusion The subtype‐specific and age‐specific distributions of HPV infection in Chifeng area have obvious heterogeneity ,which indicates that HPV genotypes and age should be considered in screening ,prevention and treatment of cervical cancer .
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Objective To investigate the distribution characteristics of hepatitis B virusgene types and the situation of drug re-sistance gene in Chuxiong area and to benefit the rational clinical antiviral therapy and the prognostic evaluation.Methods The DNA content,hepatitis B virus gene types and drug resistance associated sites in serum of hepatitis B patients were detected by the technology of fluorescence quantitative PCR and the gene chip of reverse dot blot.Results Among 103 serum samples,detection of DNA content in 85 samples was positive while DNA content of 18 samples was negative.85 positive samples were typed for hepati-tis B virus DNA,of which 63 samples were classified as type C(74.1%),21 samples were classified as type B(24.7%)and 1 sample was classified as mixed type B and C (1.2%).57 samples had the wild-type gene loci(67%)and 28 samples had the mutant gene locus(33%).Among these samples,1 7 cases of mutation were placed in rt180M,12 cases were placed in rt204V,6 cases were placed in rt204I,3 cases were placed in rt207I,3 cases were placed in rt181V and 3 cases were placed in rt236T.Among 28 samples of drug resistance gene mutation,5,22,24 and 1 1 samples were sensitive to lamivudine,telbivudine,adefovir and Entecavi,respectively.Con-clusion The main hepatitis B virus genotype is type C in Chuxiong area.The most important mutant site is rt180M and the second is rt204V.The most sensitive drugs are telbivudine and adefovir in clinical antiviral therapy and the second sensitive drug is Ente-cavi While the most commonly resistant drug is lamivudine.
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Objective To understand the gene carrying rate ,gene mutation types and distribution characteristics of thalassemia in the northeast area of Chongqing .Methods 28 633 specimens collected from the patients and individuals with physical examina‐tion in our hospital from January to December 2013 were performed the RBC parameters detection and hemoglobin electrophoresis screening .The specimens with phenotype positive were definitely verified the thalassemia type by using Gap‐PCR and reverse dot blot(RDB) .Results Among 28 633 specimens ,1 358 specimens were finally diagnosed as thalassemia with the thalassemia carrying rate of 4 .74% ,including 589 cases(2 .06% ) of α‐thalassemia and 741 cases (2 .59% ) of β‐thalassemia cases .Among the α‐thalasse‐mia genotypes ,‐αα/‐‐SEA genotype(1 .38% ) was most common ,the next was ‐αα/‐α3 .7 genotype (0 .37% ) and αα/‐α4 .2 genotype (0 .20% ) .Among the β‐thalassemia genotypes ,CD41‐42 genotype (1 .27% ) had the highest constituent ratio ,followed by IVS‐2‐654 genotype(1 .27% ) and CD17 genotype(0 .30% ) .28 cases were found to be the double heterozygote with α‐thalassemia and β‐thalassemia .Conclusion The northeast area of Chongqing is a region with the high incidence rate of thalassemia and complicated heredity .Therefore this research provides the reference information for the prevention of thalassemia ,genetic counseling and prena‐tal diagnosis .
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<p><b>OBJECTIVE</b>A PCR-reverse dot blot hybridization (RDBH) assay was developed for rapid detection of rpoB gene mutations in 'hot mutation region' of Mycobacterium tuberculosis (M. tuberculosis).</p><p><b>METHODS</b>12 oligonucleotide probes based on the wild-type and mutant genotype rpoB sequences of M. tuberculosis were designed to screen the most frequent wild-type and mutant genotypes for diagnosing RIF resistance. 300 M. tuberculosis clinical isolates were detected by RDBH, conventional drug-susceptibility testing (DST) and DNA sequencing to evaluate the RDBH assay.</p><p><b>RESULTS</b>The sensitivity and specificity of the RDBH assay were 91.2% (165/181) and 98.3% (117/119), respectively, as compared to DST. When compared with DNA sequencing, the accuracy, positive predictive value (PPV) and negative predictive value (NPV) of the RDBH assay were 97.7% (293/300), 98.2% (164/167), and 97.0% (129/133), respectively. Furthermore, the results indicated that the most common mutations were in codons 531 (48.6%), 526 (25.4%), 516 (8.8%), and 511 (6.6%), and the combinative mutation rate was 15 (8.3%). One and two strains of insertion and deletion were found among all strains, respectively.</p><p><b>CONCLUSION</b>Our findings demonstrate that the RDBH assay is a rapid, simple and sensitive method for diagnosing RIF-resistant tuberculosis.</p>