GP5+/6+ SYBR Green methodology for simultaneous screening and quantification of human papillomavirus

Background: Detection and quantification of human papillomavirus (HPV) may help in predicting the evolution of HPV infection and progression of associated lesions. Objectives: We propose a novel protocol using consensus primers GP5+/6+ in a SYBR Green quantitative real-time (Q-RT) polymerase chain reaction (PCR). The strategy permits screening for HPV infection and viral load quantification simultaneously. Study design: DNA from 153 archived cervical samples, previously tested for HPV detection by GP5+/6+ PCR and typed by EIA-RLB (enzyme immunoassay-reverse line blot) or sequence analysis, was analysed using SYBR Green Q-RT PCR. Melting temperature assay (T(m)) and cycle threshold (C(t)) were used to evaluate HPV positivity and viral load. The T(m) in the range of 77-82 degrees C was considered to be positive for HPV-DNA. HPV results generated through GP5+/6+ conventional PCR were considered the gold standard against which sensitivity and specificity of our assay were measured. Results: Out of 104 HPV positive samples, 100 (96.2%) were also determined as positive by SYBR Green Q-RT PCR; of the 49 HPV-negative samples, all were determined as negative. There was an excellent positivity agreement (K = 0.94) between the SYBR Green Q-RT and the previous methods employed. The specificity and sensitivity were 100% and 96.2%, respectively. Comparison of SYBR Green Q-RT and TaqMan oligo-probe technologies gave an excellent concordance (pc = 0.95) which validated the proposed strategy. Conclusions: We propose a sensitive and easy-to-perform technique for HPV screening and viral load quantification simultaneously. (C) 2009 Elsevier B.V. All rights reserved.

Craniocervical posture and hyoid bone position in children with mild and moderate asthma and mouth breathing

Introduction The objective of the present study was to assess the craniocervical posture and the positioning of the hyoid bone in children with asthma who are mouth breathers compared to non-asthma controls. Methods The study was conducted on 56 children, 28 of them with mild (n = 15) and moderate (n = 13) asthma (14 girls aged 10 79 +/- 1 31 years and 14 boys aged 9 79 +/- 1.12 years), matched for sex, height, weight and age with 28 non-asthma children who are not mouth breathers The sample size was calculated considering a confidence interval of 95% and a prevalence of 4% of asthma in Latin America. Eighteen variables were analyzed in two radiographs (latero-lateral teleradiography and lateral cervical spine radiography), both obtained with the head in a natural position The independent t-test was used to compare means values and the chi-square test to compare percentage values (p < 0 05) Intraclass correlation coefficient (ICC) was used to verify reliability. Results. The Craniovertebral Angle (CVA) was found to be significantly smaller in asthma than in control children (106.38 +/- 766 vs. 111 21 +/- 7.40. p = 0 02) and the frequency of asthma children with an absent or inverted hyoid triangle was found to be significantly higher compared to non-asthma children (36% vs 7%, p = 0.0001). The values of the inclination angles of the superior cervical spine in relation to the horizontal plane were significantly higher in moderate than in mild asthma children (CVT/Hor 85 10 +/- 725 vs. 90 92 +/- 6.69, p = 0 04 and C1/Hor. 80 93 +/- 5.56 vs 85 00 +/- 4 20, p = 0 04) Conclusions These findings revealed that asthma children presented higher head extension and a higher frequency of changes in hyoid bone position compared to non-asthma children and that greater the asthma severity greater the extension of the upper cervical spine. (C) 2010 Elsevier Ireland Ltd. All rights reserved.