Comparative real-time PCR quantification of cytomegalovirus in severe early childhood caries and caries-free children.

PURPOSE: The present study was undertaken to evaluate cytomegalovirus (CMV) load in carious lesions in children with severe early childhood caries (S-ECC) and compare it with caries-free (CF), healthy children, by quantitative real-time polymerase chain reaction (PCR). MATERIALS AND METHODS: Dental plaque samples were collected from 57 subjects including S-ECC (n = 33) and CF (n = 24) patients. The two groups were matched in terms of age, gender, and socioeconomic status. After deoxyribonucleic acid (DNA) extraction, SYBR Green real-time PCR was used to quantify CMV DNA. RESULTS: The CMV DNA was detected in seven patients in the S-ECC group. The mean of decayed, missing, and filled tooth surfaces (dmfs) was 11.90 ± 5.95 in the S-ECC group. There was no significant difference in gender between the two groups. There was a significant relationship between CMV positivity in children and the S-ECC group (p = 0.043). CONCLUSION: Results of this study suggest a relationship between the presence of CMV in the oral cavity and S-ECC. We can design clinical trials to confirm these results and improve children's dental care.

Molecular Characterization of Environmental Non-Tuberculous Mycobacteria Using PCR- RFLP Analysis of 441 Bp Heat Shock Protein 65 Fragments.

BACKGROUND: Non- Tuberculous Mycobacteria are environmental opportunistic pathogens that can be found in various terrestrial and aquatic habitats. There are an epidemiological links between species isolated in tap water and those isolated from patients. hsp65 gene has more variability in its sequences, compared to the some more conserved genes in NTM, for identification of mycobacteria to species level. In this study, the prevalence of NTM in Isfahan City water samples was determined using culture, biochemical tests and PCR-RFLP analyses of hsp65 gene. METHODS: Eighty-five water samples were collected and cultured. The mycobacterial isolates were identified by conventional biochemical tests. A 441 bp fragment of hsp65 genes was amplified and digested by two restriction enzymes, BstEII and HaeII. Digested products were analyzed using polyacrilamid gel electrophoresis (PAGE). RESULTS: 25.9% of the water samples contained different species of NTM. Dominant isolates were M. fortuitum (26.7%), M. chelonae like organism (13.3%) and M. mucogenicum (13.3%). Nineteen isolates of Mycobacteria were differentiated using hsp65 genes PCR-RFLP. Three isolates could not be identified at the species level because their RFLP patterns were different from other known PCR-RFLP profiles. There were different hsp65 gene PCR-RFLP profiles produced by digestion with BstEII and HaeIII. CONCLUSION: This study showed that PCR-RFLP of hsp65 gene in mycobacteria is more reliable method for identification of NTM at the specie level than conventional phenotypic methods (P<0.05). In comparing of RFLP patterns of this study to other investigation, some minor differences were negligible.

Periodontopathic bacteria and herpesviruses in chronic periodontitis.

Periodontal disease involves complex interactions of microorganisms and host defenses. This work investigated the associations between putative bacterial pathogens, herpesviruses and chronic periodontitis. Subgingival samples were collected from 40 periodontally healthy individuals and from 40 patients with chronic periodontitis with probing depths of < or =3 mm or > or =6 mm. Multiplex and nested polymerase chain reactions were used to identify bacterial pathogens and herpesviruses. Porphyromonas gingivalis, Tannerella forsythia, Epstein-Barr virus (EBV) type 1, cytomegalovirus (CMV), Aggregatibacter actinomycetemcomitans and EBV type 2 were detected in, respectively, 95, 75, 72.5, 50, 12.5 and 10% of sites with probing depths > or =6 mm. P. gingivalis, T. forsythia, EBV-1 and CMV were statistically associated with probing depths > or =6 mm. A. actinomycetemcomitans and EBV-2 showed no association with periodontitis sites, and no significant associations were found for any of the test infectious agents and probing depths < or =3 mm. Our results confirm an association between P. gingivalis, T. forsythia, EBV-1 and CMV, and chronic periodontitis. These infectious agents may play an important synergistic role in the pathogenesis of chronic periodontitis.

Quantification of ergovaline using HPLC and mass spectrometry in Iranian Neotyphodium infected tall fescue.

Ergovaline, the main ergopeptine alkaloid produced in tall fescue (Fescue arundinacea Schreb.) infected with endophyte (Neotyphodium coenophialum Morgan- Jones & Gams), is known to cause tall fescue toxicosis. This study was conducted to examine the presence of fungal endophytes in five populations of tall fescue collected from various regions of Iran. The existence of Neotyphodium mycelia in the tissues of the samples was confirmed by microscopic examination, and the isolation was performed from leaf tissues of the hosts on potato dextrose agar. All isolates were confirmed as the Neotyphodium species by PCR, using specific primers. Mass detection and determination of ergovaline were performed by HPLC at three plant growth stages. Ergovaline was detected in all isolates, with the mean concentrations of 0.24 to 3.48 µg/g dry matter of different populations for the whole three plant growth stages. The differences in ergovaline content between plant populations and sampling time were statistically significant. This is the first report of ergovaline content in endophyte infected Fescue arundinacea from natural grasslands in Iran.

Prevalence of Epstein-Barr virus type 1 in patients with chronic periodontitis by nested-PCR.

The present study evaluated the subgingival presence of Epstein-Barr virus type 1 (EBV-1) in patients with chronic periodontitis with nested-PCR. Subgingival plaque samples from 61 patients with chronic periodontitis with Probing Depth (PD) > or = 6 and 40 healthy controls were collected by sterile curette. DNA was extracted. A nested Polymerase Chain Reaction (PCR) method determined presence of EBV-1. The study included 61 patients (34 women, 27 men; 24-69 years of age; mean 43) and 40 periodontally health controls (22 Women, 18 men, 21-69 years in age; mean 41.35%). EBV type 1 was detected in 37 samples (60.7%) and 1 samples (2.5%) of chronic periodontitis patients and healthy subjects, respectively. This study demonstrated that EBV-1 infection is associated with the activity of chronic periodontitis.