Human papillomavirus (HPV) infections are strongly associated with the development of cervical intraepithelial neoplasias and invasive cervical cancer. Polymorphisms in cytokine-encoding genes and behavioural cofactors could play an important role in protecting an individual against viral infections and cancer. Here, we investigated whether IL-6 -174 G>C, IL-8 +396 G>T, and TGF-β1 +869 G>C and +915 G>C polymorphisms were associated with susceptibility to HPV infection in women from north-east (Pernambuco) Brazil. We analysed 108 healthy uninfected women (HC) and 108 HPV-positive women with cervical lesions. Genetic polymorphisms were assessed using Sanger sequencing and polymerase chain reaction-restriction fragment length polymorphism. Comparison of the distribution of the genotypic and allelic frequencies of the IL-18 +396 T>G polymorphism between HPV infected woman an uninfected controls showed that the GG genotype and G allele were both more frequent in the HC group, and were associated with protection from HPV infection (p = 0.0015; OR = 0.29 CI95% = 0.13-0.61; p = 0.0005; OR = 0.45 CI95% 0.29-0.7, respectively). Individuals from the control group could have previously had HPV infection that was spontaneously eliminated; however, it was undetectable at the time of sample collection. Based on our findings, we hypothesize that the IL-8 +396 G>T polymorphism could interfere with susceptibility to HPV infection, by modulating the ability of immune system to fight the virus.

Polymorphisms in chemokine receptors play an important role in the progression of cervical intraepithelial neoplasia (CIN) to cervical cancer (CC). Our study examined the association of CCR2-64I (rs1799864) andCCR5-Δ32 (rs333) polymorphisms with susceptibility to develop cervical lesion (CIN and CC) in a Brazilian population. The genotyping of 139 women with cervical lesions and 151 women without cervical lesions for the CCR2-64I and CCR5-Δ32 polymorphisms were performed using polymerase chain reaction-restriction fragment length polymorphism. The individuals carrying heterozygous or homozygous genotypes (GA+AA) for CCR2-64I polymorphisms seem to be at lower risk for cervical lesion [odds ratio (OR) = 0.37, p = 0.0008)]. The same was observed for the A allele (OR = 0.39, p = 0.0002), while no association was detected (p > 0.05) with CCR5-Δ32 polymorphism. Regarding the human papillomavirus (HPV) type, patients carrying the CCR2-64Ipolymorphism were protected against infection by HPV type 16 (OR = 0.35, p = 0.0184). In summary, our study showed a protective effect ofCCR2-64I rs1799864 polymorphism against the development of cervical lesions (CIN and CC) and in the susceptibility of HPV 16 infection.

OBJETIVO: comparar três métodos para detecção do HPV e determinar a prevalência dos genótipos encontrados. MÉTODOS: um total de 120 amostras de raspagem da região cervical de mulheres portadoras de neoplasia intraepitelial cervical foram analisadas pela reação em cadeia da polimerase convencional, usando os sistemas de primers MY09/11, GP05+/06+ e pela Nested-PCR. As amostras foram submetidas à extração de DNA e, logo após, amplificadas com os primers GH20 e PC04 (β-globina) para verificação da qualidade do DNA obtido e pela reação em cadeia da polimerase convencional e Nested-PCR. Os fragmentos amplificados foram visualizados em gel de agarose a 1,2%, corados com Blue Green Loading Dye I. As amostras positivas foram sequenciadas usando o sequenciador automático de DNA "MegaBACE 1000". Para análise estatística foram utilizados os teste do Χ2 e o de Fisher com nível de significância de 5%. RESULTADOS: quinze amostras não se amplificaram para os primers de β-globina, sendo eliminadas do estudo. Das amostras restantes, 40% (42/105) foram positivas para os primers MY09/11, 98% (103/105) para os primers GP05+/06+ e 92% (97/105) para Nested-PCR. Considerado as técnicas MY09/11 e GP05+/06+, foi possível observar 100% de amostras positivas para o HPV. Neste estudo, a prevalência dos genótipos foi de 58, 23, 5, 4 e 3% para HPVs 16, 18, 31, 33 e 56, respectivamente. Os HPV 67 e 83 apresentaram 2% e os HPV 6, 11, 58 e candHPV85, 1% cada. A prevalência dos genótipos neste estudo está de acordo com o reportado em todo o mundo (IC95%=0,4657-0,8976). CONCLUSÕES: para obter resultados mais confiáveis, é necessário o uso de mais que um sistema de primers para detecção do HPV. Acredita-se que as três técnicas estudadas são importantes e adequadas para o diagnóstico clínico do HPV quando apropriadamente combinadas.

PURPOSE: to compare three methods for the detection of HPV infection and to determine the prevalence of the genotypes found. METHODS: a total of 120 cervical scrape samples from patients with cervical intraepithelial neoplasia were analyzed by the conventional polymerase chain reaction using the MY09/11 and GP05+/06+ primers, and by the Nested polymerase chain reaction. The samples were subjected to DNA amplification with the GH20 and PC04 primers (β-globin) to verify DNA quality and also by polymerase chain reaction and Nested polymerase chain reaction. The amplicons were visualized in 1.2% agarose gel stained with Blue Green Loading Dye I. Positive samples also were sequenced using the automatic DNA sequencer "MegaBACE 1000". The Χ2 and Fisher tests were used for statistical analysis with the level of significance set at 5%. RESULTS: fifteen samples were eliminated from the study because they failed to amplify the β-globin gene. Of the remaining samples, 40% (42/105) were positive using primers MY09/11, 98% (103/105) using primers GP05+/06+, and 92% (97/105) using Nested-PCR. With the MY09/11 and GP05+/06+ techniques, it was possible to obtain 100% HPV-positive samples. In this study, the prevalence of the genotypes found was 57, 23, 5, 4 and 3% for HPV genotypes 16, 18, 31, 33 and 56, respectively. HPVs 67 and 83 were present in 2%, and genotypes 6, 11, 58 and candHPV85 were present in 1% each. The prevalence of the more common genotypes (HPV 16 and 18) in this study agrees with that reported worldwide (IC95%=0.4657-0.8976). CONCLUSIONS: to obtain more reliable results, it is necessary the use of more than one primer system to detect HPV infections. We believe that the three techniques studied are important and suitable for the clinical diagnosis of HPV, when they are appropriately combined.

OBJETIVO: Avaliar a associação entre o polimorfismo no exon-1 do gene MBL2 e a doença periodontal em pacientes diabéticos tipo 2. MÉTODO: A amostra foi composta por 100 pacientes que foram submetidos a um exame clínico periodontal que avaliou seis sítios por dente a profundidade de sondagem (PS), sangramento à sondagem (SS), perda de inserção clínica (PIC), índice de placa (IP) e o número de dentes presente. A doença periodontal foi definida como pelo menos quatro sítios com perda de inserção de >5mm, com um ou mais destes sítios tendo uma bolsa de >4mm. Foram coletadas células de descamação da mucosa oral e a detecção do polimorfismo foi feita através da PCR em tempo real e análise da temperatura de melting. RESULTADOS: Em uma população de diabéticos tipo 2, não houve diferenças estatisticamente significantes nos genótipos do polimorfismo da MBL2 ou freqüência alélica observadas entre os indivíduos com doença periodontal. CONCLUSÃO: Este estudo indicou que o polimorfimos no exon-1 do gene da MBL2 não foi relacionado à doença periodontal em uma população de diabéticos tipo 2.

PURPOSE: To assess the association between the polymorphism in exon-1 of the MBL2 gene and the periodontal disease in type 2 diabetic patients. METHODS: The sample comprised of 100 patients, who were submitted to a clinical periodontal examination that evaluated in six sites per tooth the probing depth (PD), bleeding on probing (BOP), clinical attachment loss (CAL), plaque index (PI) and the number of teeth present. Periodontal disease was defined as at least four sites with loss of attachment of >5 mm, with one or more of those sites having a pocket of > 4 mm. The collection of scaling cells from the oral mucosa was carried out and the detection of MBL2 polymorphism was made by real time PCR and melting temperature curve analysis. RESULTS: In a type 2 diabetic population, no significant statistical differences in MBL2 polymorphisms genotype or allele frequencies were observed among subjects with periodontal disease. CONCLUSION: This study indicates that the polymorphisms in exon-1 of the MBL2 gene are not related to periodontal disease in a type 2 diabetic population.

O advento dos métodos moleculares tem, nos últimos anos, revolucionado a detecção e caracterização dos microorganismos em diversas áreas médicas diagnósticas, tais como virologia, micologia, parasitologia, microbiologia e odontologia. Dentre as técnicas baseadas em biologia molecular, a PCR (Polymerase Chain Reaction) trouxe enormes benefícios e desenvolvimentos científicos, se mostrando como um excelente caminho para a rápida detecção de patógenos, até mesmo aqueles de difícil cultivo. Derivada da PCR convencional, a PCR em Tempo Real se mostra como uma inovação tecnológica e vem conquistando espaço nos diagnósticos clínicos e nos laboratórios de pesquisa por apresentar a capacidade de gerar, além de resultados qualitativos, resultados quantitativos, se mostrando de forma mais rápida e precisa. Este trabalho de revisão tem por objetivo explorar a utilidade clínica da técnica de PCR convencional e em Tempo real nas diversas áreas médicas supracitadas, abrangendo seus principais usos e avanços, direcionando para o cotidiano profissional.

Recent developments in molecular methods have revolutionized the detection and characterization of microorganisms in a broad range of medical diagnostic fields, including virology, mycology, parasitology, microbiology and dentistry. Among these methods, Polymerase Chain Reaction (PCR) has generated great benefits and allowed scientific advancements. PCR is an excellent technique for the rapid detection of pathogens, including those difficult to culture. Along with conventional PCR techniques, Real-Time PCR has emerged as a technological innovation and is playing an ever-increasing role in clinical diagnostics and research laboratories. Due to its capacity to generate both qualitative and quantitative results, Real-Time PCR is considered a fast and accurate platform. The aim of the present literature review is to explore the clinical usefulness and potential of both conventional PCR and Real-Time PCR assays in diverse medical fields, addressing its main uses and advances.

This study compares the detection rates of Chlamydia trachomatis by two techniques, direct immunofluorescence (IMF) and real time polymerase chain reaction (PCR), in patients with and without intra-epithelial cervical lesions (SIL) in Recife. We conducted a transversal study involving 35 women with SIL and 35 without SIL attended at Ambulatório Especializado da Mulher, Recife, Brazil. They were tested for Chlamydia trachomatis using two techniques, direct IMF or real time PCR. The rates of Chlamydia trachomatis detection were compared and the association with intra-epithelial cervical lesions was determined using the chi-square test at a 5% level of significance. Concordance between the tests was evaluated using kappa. The global prevalence of Chlamydia infection was 47.1% by direct IMF and 58.6% by real time PCR. A significant association was observed between Chlamydia diagnosis and presence of intra-epithelial cervical lesions, with about 80% positive results by direct IMF and 77.1% by real time PCR. However, the detected rate of infection with Chlamydia trachomatis was significantly greater in patients without intra-epithelial cervical lesions tested by real time PCR (40%) when compared to direct IMF (14.3%). The concordance between the tests was weak, with a kappa coefficient of 0.4. Both real time PCR and direct IMF detected elevated rates of Chlamydia infection in patients with intra-epithelial cervical lesions (80%) but the tests were discordant when patients without cervical lesions were tested, possibly because sensitivity of real time PCR is greater.