Adolfo Contreras

Comparison of nested polymerase chain reaction (PCR), real‐time PCR and viral culture for the detection of cytomegalovirus in subgingival samples

INTRODUCTION The purpose of this study was to compare nested polymerase chain reaction (PCR), real-time PCR, and shell vial for the detection of human cytomegalovirus (HCMV) in subgingival samples in periodontitis patients. METHODS A group of 44 patients and 24 individuals without periodontitis were included in the study. A full periodontal examination was conducted in each subject. Gingival crevicular fluid (GCF) was collected by pocket lavage and used for viral culture (shell vial). Additional subgingival samples were obtained with paper points and used for molecular analysis. Nested PCR and real-time PCR were used to detect and quantify HCMV. Students t-test and chi-squared test were used to compare groups. The sensitivity and specificity for the tests were calculated on 2 x 2 tables considering the nested PCR as the gold standard. RESULTS The detection of HCMV was greater using nested PCR than with either real-time PCR or shell vial (P < 0.0001). However, the frequency detection of both molecular techniques was higher than in viral culture (P < 0.0001). Only one case of chronic periodontitis was positive by viral culture. Agreement between nested PCR and real-time PCR was observed 47.7% and 4.1% of the time in the periodontitis and control groups, respectively. The sensitivity of real-time PCR was 60%, compared with 2.8% for the shell vial technique. CONCLUSIONS In conclusion, this study confirmed that active HCMV infection occurs in human periodontitis; however, its frequency seems to be low. In contrast, latent periodontal HCMV infection seems to be a more frequent event.

Biology and pathogenesis of cytomegalovirus in periodontal disease

Abstract Human periodontitis is associated with a wide range of bacteria and viruses and with complex innate and adaptive immune responses. Porphyromonas gingivalis, Tannerella forsythia, Aggregatibacter actinomycetemcomitans, Treponema denticola, cytomegalovirus and other herpesviruses are major suspected pathogens of periodontitis, and a combined herpesvirus–bacterial periodontal infection can potentially explain major clinical features of the disease. Cytomegalovirus infects periodontal macrophages and T‐cells and elicits a release of interleukin‐1β and tumor necrosis factor‐α. These proinflammatory cytokines play an important role in the host defense against the virus, but they also have the potential to induce alveolar bone resorption and loss of periodontal ligament. Gingival fibroblasts infected with cytomegalovirus also exhibit diminished collagen production and release of an increased level of matrix metalloproteinases. This article reviews innate and adaptive immunity to cytomegalovirus and suggests that immune responses towards cytomegalovirus can play roles in controlling, as well as in exacerbating, destructive periodontal disease.

Periodontal disease in children and adolescents of Latin America

Periodontal diseases are a group of infectious diseases that mainly include gingivitis and periodontitis. Gingivitis is the most prevalent form of periodontal disease in subjects of all ages, including children and adolescents. Less frequent types of periodontal disease include aggressive periodontitis, acute necrotizing ulcerative gingivitis and various diseases of herpesviral and fungal origin. This review aimed to retrieve relevant information from Latin America on the prevalence of periodontal diseases among children and adolescents of the region. Gingivitis was detected in 35% of young Latin American subjects and showed the highest frequencies in Colombia (77%) and Bolivia (73%) and the lowest frequency in Mexico (23%). The frequency of gingivitis in subjects from other Latin American countries was between 31% and 56%. Periodontitis may affect <10% of the young population in Latin America, but the data are based on only a few studies. A more precise assessment of the distribution and severity of periodontal disease in children and adolescents of Latin America may help policy makers and dentists to institute more effective public health measures to prevent and treat the disease at an early age to avoid major damage to the permanent dentition.

Effects of cytomegalovirus infection on the mRNA expression of collagens and matrix metalloproteinases in gingival fibroblasts

BACKGROUND AND OBJECTIVE The purpose of this in vitro investigation was to study the effects of human cytomegalovirus infection on the mRNA expression for collagens I and III and for matrix metalloproteinases 1 and 2 in gingival fibroblasts. MATERIAL AND METHODS Gingival fibroblasts were experimentally infected with the Towne strain of human cytomegalovirus and the kinetics of expression of mRNA for collagens I and III and for matrix metalloproteinases 1 and 2 was studied at different time-points. Total RNA was isolated at the indicated time, and the reverse transcription-polymerase chain reaction was used to analyze the level of mRNA expression. In addition, gingival specimens were obtained from 14 periodontitis and from three non-periodontitis subjects and mRNA analysis for collagens and metalloproteinases was carried out. Nested polymerase chain reaction was used to determine the presence or absence of human cytomegalovirus in subgingival samples from each subject. RESULTS The infection of gingival fibroblasts with human cytomegalovirus during a 0-72-h period resulted in progressive reduction in the expression of mRNA for collagens I and III (p < 0.05). A higher concentration of human cytomegalovirus resulted in varying degrees of mRNA reduction, suggesting a virally mediated effect. Biopsies from human cytomegalovirus-positive individuals with periodontitis had a higher expression of mRNA for collagens I and III than biopsies from human cytomegalovirus-negative individuals. An up-regulation in the mRNA expression of matrix metalloproteinases 1 and 2 over time was observed (p < 0.05). Analysis of mRNA expression in gingival biopsies demonstrated higher expression of matrix metalloproteinase-1 in human cytomegalovirus-positive periodontitis specimens compared with human cytomegalovirus-negative periodontitis specimens. CONCLUSION Altered expression of mRNA for collagens and metalloproteinases in human cytomegalovirus-infected gingival fibroblasts should be considered as possible modifying mechanisms in periodontitis-infected sites.