DNA In Saliva Research Articles

Persistent Epstein--Barr virus infection in the common marmoset (Callithrix jacchus)

Epstein-Barr virus (EBV) infection of the common marmoset causes long-term infection, with production of antibodies to virus-induced antigens, without clinical illness. Attempts to show the presence of EBV DNA in saliva of infected animals by PCR were initially unsuccessful, although slot-blot hybridization analysis demonstrated that viral DNA was present. Further investigations showed that most samples of pilocarpine-induced saliva, and 33% of the samples of whole mouth fluids (WMF) tested, were inhibitory to PCR. Similar results were found using human WMF. A method of assessing samples of marmoset WMF for the presence of EBV, by PCR using an EBV BamHI W probe, and removing inhibition with Chelex 100, is described. A total of 202 samples from 21 EBV infected, and seven non-infected animals was tested. Five seropositive animals shed virus on every occasion, and 15 intermittently. Two marmosets, infected as neonates, showed progressively increasing humoral responses to viral antigens, and shed virus on every occasion tested over 3 years. When mated with uninfected animals, the latter seroconverted 4 and 6 weeks later, respectively, and later shed virus into their WMF. The naturally infected animals were paired with naive marmosets, and were able to pass on infection. These results establish that long-term, permissive EBV infection occurs in the common marmoset, and demonstrate again the similarities in the response to EBV between marmoset and man.

Polymerase chain reaction reveals herpes simplex virus DNA in saliva of patients with Behçet's disease.

The etiology of Behçet's disease is unclear, but viral infection is thought to be one etiologic factor. The aims of this study were to detect herpes simplex virus (HSV) DNA in the saliva of patients with Behçet's disease and of healthy persons, to determine whether the presence of HSV in saliva is associated with the presence of intraoral ulcer, and to investigate the relationship between HSV and Behçet's disease. The polymerase chain reaction (PCR) was used to detect HSV DNA sequences in DNA extracted from the saliva of patients with Behçet's disease and of healthy control subjects. Of 66 patients with Behçet's disease diagnosed clinically, 19 were diagnosed as complete type, 29 as incomplete type and 18 as suspected type. Of 66 DNA preparations from the saliva of the patients, 26 (39.4%) showed the 289-bp band. This contrasts with 12 of 87 preparations (13.8%) from healthy controls (P < 0.01). There were no significant differences among the three patient groups. All the 289-bp bands analyzed by restriction endonuclease digestion yielded the expected 158-bp and 131-bp fragments when digested with PstI.HSV DNA was detected in 12 of 33 Behçet's disease patients (36.4%) with oral ulceration and 14 of 33 patients (42.4%) without oral ulceration at the time of testing. There was no statistically significant correlation in the PCR results between the two groups.

Detection of proviral human T-cell lymphotrophic virus type I DNA in mouthwash samples of HAM/TSP patients and HTLV-I carriers

Human T-cell lymphotrophic virus type I (HTLV-I), is a member of the oncogenic retroviruses family endemic in several parts of the world and also recently identified in the Jewish Mashhadi population who immigrated from Iran to Israel. The virus is the causative agent of adult T-cell leukemia (ATL) and a chronic myelopathy known both as tropical spastic paraparesis (TSP) or HTLV-I associated myelopathy (HAM). The known modes of HTLV-I transmission are by sexual intercourse, from mother to child in breast milk, via blood transfusion, and by sharing of needles by parenteral drug users. In the present study we examined the presence of HTLV-I provirus genomic DNA by nested polymerase chain reaction (PCR) and by DNA hybridization in mouthwash samples obtained from 13 Mashhadi-born Iranian Jews with spastic paraparesis associated with HTLV-I, 4 Mashhadi-born Iranian Jews asymptomatic carriers for HTLV-I and 21 healthy controls. Proviral HTLV-I DNA was detected by mouthwash PCR in 12 of 17 HTLV-I infected subjects (71%) but in none of 21 controls. Proviral DNA was also detected in mouthwash samples using HTLV-I probe by dot blot hybridization assay. The presence of HTLV-I proviral DNA in whole saliva may suggest a possible transmission of the virus via saliva and explain the increased rate of infection in elderly Mashhadi-Jewish population.

Epstein-Barr Virus (EBV) DNA in saliva and EBV serology of HIV-1-infected persons with and without hairy leukoplakia

Secretion of Epstein-Barr virus (EBV) in saliva, as well as serum antibody titres against various EBV antigens, were analyzed in respect of (1) 15 HIV-1-infected patients with oral hairy leukoplakia proven to contain EBV by in situ hybridization, (2) 45 HIV-1 infected patients without hairy leukoplakia, (3) 10 HIV-1 infected patients treated with acyclovir or foscarnet and (4) 21 healthy controls. The numbers of CD4+ cells in the peripheral blood were also recorded. The HIV-1 infected patients were at various stages of HIV-1-associated disease. Excretion of EBV DNA in the saliva was determined by means of the polymerase chain reaction (PCR) while the amount of EBV DNA in positive samples was estimated by repeated titrations. The frequency of shedding of EBV DNA increased from 33% in healthy controls to 78% in asymptomatic HIV-1 infected persons, but did not increase significantly with progression of HIV-1-associated disease. The titres of EBV DNA in saliva correlated inversely and significantly with the number of CD4+ cells in the peripheral blood. All patients with hairy leukoplakia shed by EBV DNA in their saliva but the titres were not significantly higher than those of other HIV-1 infected persons. The serum titres of antibodies against EBV nuclear antigen 1 (EBNA-1) correlated positively and significantly with the CD4+ cell count in the peripheral blood. EBNA-1 IgG antibody in the serum was also significantly lower in symptomatic than in asymptomatic HIV-1 infected persons. There were, however, no significant differences in serum antibodies to various EBV antigens between patients with and without hairy leukoplakia.

Detection of H. pylori DNA in saliva as a means of monitoring eradication therapy : C. Li 1, T. Ha 1, DA. Ferguson, Jr 2., RG. Zhao, DS. Chi 1, and [formula omitted]1,3 Departments of Internal Medicine 1 and Microbiology 2, James H. Quillen College of Medicine, East Tennessee State University and VA Medical Center 3, Johnson City, TN 37684

Detection of H. pylori DNA in saliva as a means of monitoring eradication therapy : C. Li 1, T. Ha 1, DA. Ferguson, Jr 2., RG. Zhao, DS. Chi 1, and [formula omitted]1,3 Departments of Internal Medicine 1 and Microbiology 2, James H. Quillen College of Medicine, East Tennessee State University and VA Medical Center 3, Johnson City, TN 37684

Infection Of Oral Mucosal Cells By Human Immunodeficiency Virus Type 1 In Seropositive Persons

Molecular studies have revealed significant amounts of human immunodeficiency virus type 1 (HIV-1) provirus DNA in saliva of HIV-infected persons. However, cellular localization has not been determined. In situ polymerase chain reaction (IS-PCR) was done on saliva-associated cells for localization of HIV-1 provirus DNA. Results indicate its presence in the nuclei of saliva-associated epithelial cells in 29 (83%) of 35 HIV-1-seropositive subjects. In 24 (83%) of the 29 IS-PCR-positive samples, 0.1%-4.0% of the mucosal epithelial cells exhibited nuclear localization of HIV-1 DNA. In addition, HIV-1 provirus DNA was detected in monocytes or lymphocytes of all salivary samples from the 35 subjects. The localization of HIV-1 provirus DNA indicates that epithelial cells are another cell type infected by HIV-1 in vivo. These findings suggest epithelial cells in other body sites might also be infected with HIV-1.

Human Herpesvirus 6 DNA in Blood Cells of Human Immunodeficiency Virus-Infected Men: Correlation of High Levels with High CD4 Cell Counts

The relationship between human herpesvirus 6 (HHV-6) and the progression of human immunodeficiency virus type 1 (HIV-1) infection to AIDS was evaluated using the polymerase chain reaction to quantitate HHV-6 genome equivalents in peripheral blood mononuclear cells (PBMC) and saliva of 32 HIV-1-seropositive men. Unlike other herpesviruses, HHV-6 was found with higher frequency and in higher copy numbers of HIV-infected men with high rather than low CD4 cell counts. Among subjects with > 400 CD4 cells/mL, 100% had detectable HHV-6 genomes in PBMC compared with 57.9% of subjects with < 400 CD4 cells/mL (P < .05). There was no significant correlation between HHV-6 antibody titer or number of amplifiable copies of HHV-6 DNA in saliva and stage of HIV disease or CD4 cell number.

Human herpesvirus 6 DNA in peripheral blood cells and saliva from immunocompetent individuals

Human herpesvirus 6 (HHV-6) genome equivalents were quantitated in peripheral blood mononuclear cells (PBMCs) and saliva from 20 healthy individuals by the polymerase chain reaction (PCR). Nineteen of 20 subjects (95%) harbored HHV-6 DNA: 18 (90%) had HHV-6 in their PBMCs and 18 had HHV-6 in their saliva. Quantitative PCR revealed HHV-6 DNA levels ranging from negative to 4,000 HHV-6 genome equivalents per 10(6) PBMCs and from negative to 200,000 HHV-6 genome equivalents per ml of saliva. Longitudinal saliva samples from 15 HHV-6-seropositive subjects revealed salivary HHV-6 DNA persistence in 13 subjects. HHV-6 antibodies were detected in 17 of 19 subjects, with titers ranging from 1:400 to 1:51,200 (geometric mean titer, 1:2,500). Antibody titers did not correlate with HHV-6 DNA levels in PBMCs or saliva (P = 0.27 and P = 0.44, respectively). One subject with persistent HHV-6 DNA lacked detectable HHV-6 antibodies. The high prevalence of HHV-6 DNA in PBMCs and saliva supports the concept that HHV-6 exists at these sites in normal individuals.

Correlation of HBeAg/anti‐HBe, ALT levels, and HBV DNA PCR results in HBsAg‐positive patients

A highly sensitive polymerase chain reaction (PCR) was used to analyse serum samples from HBsAg-positive patients in Sweden. Forty-two chronic carriers were tested, five of whom were of Swedish origin. Of the total, there were 13 HBeAg-positive and 27 anti-HBe-positive patients, while 1 patient was negative for both HBeAg and anti-HBe and one was positive for both markers. Nine of the 13 HBeAg-positive carriers and only 7 of the 27 anti-HBe-positive carriers had elevated alanine transaminase (ALT) levels (P = 0.01). Two PCR tests of marginally different sensitivity were used on all patient samples. All 13 HBeAg-positive patients and the patients with and lacking both HBeAg and anti-HBe markers, respectively, were positive in both PCR tests. One HBeAg-positive patient was shown to shed hepatitis B virus (HBV) DNA in both saliva and urine. Twelve of the 27 anti-HBe-positive carriers, 6 of whom had elevated ALT levels, were PCR positive. The remaining 15 had no evidence of HBV DNA and all but 1 had normal ALT levels. A positive PCR result was more common in those anti-HBe-positive patients with elevated ALT levels (P < 0.02). The precore gene from 18 samples was sequenced and, with a few exceptions, showed a high degree of conservation. We suggest that in the absence of optimal tests for infectivity of serum from HBsAg-positive patients, and until PCR becomes more widely available, all anti-HBe-positive patients with elevated ALT levels be considered highly infectious.

Detection of human herpesvirus‐6 DNA in peripheral blood and saliva

Saliva and peripheral blood samples from 20 healthy adults were examined for the presence of human herpesvirus-6 (HHV-6) DNA sequences using the polymerase chain reaction. Eighteen out of 20 whole saliva samples contained detectable HHV-6 genomes. The majority of peripheral blood samples were also positive; however, the results suggest that only a rare cell in the peripheral blood is infected. Serological studies did not reveal any correlation between HHV-6 antibody titre and the ability to detect HHV-6 DNA. The data indicate that HHV-6 genomes persist in the peripheral blood and oropharynx or salivary glands of most healthy individuals following primary infection.

Quantitative analysis of hepatitis B virus DNA in saliva and semen of chronically infected homosexual men.

Serum, saliva, and semen from fifteen homosexual men who were chronic surface antigen (HBsAg) carriers were quantitatively assayed for hepatitis B virus (HBV) DNA by Southern analysis. HBV DNA (2 pg/ml-42,000 pg/ml) was detected in 11 carriers and correlated with endogenous DNA polymerase activity and e antigen (HBeAg) titer greater than or equal to 1:640. Viral DNA was detected in the saliva, at concentrations indicating a virus titer of 10(5)-10(7)/ml, of eight carriers and in semen from three patients (10(6)-10(7) virions/ml). HBV DNA in saliva and semen had electrophoretic mobilities similar to those in serum HBV DNA, but HBV DNA was present at much lower concentrations (10(-2)-10(-4) that of serum DNA). Results of isopycnic centrifugation suggested that DNA in saliva and semen was virion associated. These data provide an indication of the quantity of virus present in the semen and saliva of HBV carriers and the relative frequency with which shedding of high titers of virus (greater than 10(5) virions/ml) occurs in these secretions.

Detection of hepatitis B virus DNA in spermatozoa, urine, saliva and leucocytes, of chronic HBsAg carriers: A lack of relationship with serum markers of replication

Urine, saliva and semen from 18 chronic HBsAg carriers, material that might be concerned in the transmission of hepatitis B virus infection, was examined for the presence of HBV-DNA using molecular hybridisation. HBV-DNA was detected in samples from 14 patients: semen was positive in all of these, saliva in 12 and urine in 10. Autoradiographic studies indicated that HBV-DNA was present in the free form in all cases and in 10, higher molecular weight bands were also observed. While the latter suggested integration of HBV-DNA into host chromosome, resolution to 3.2 Kb following digestion with Eco RI in every case was more consistent with the HBV-DNA being free. In a further group of patients peripheral blood leucocytes were also found to contain HBV-DNA and were the likely source of HBV-DNA in saliva and urine, while in semen both spermatozoa and mononuclear cells contained HBV-DNA. Moreover, free HBV-DNA was identified in the semen of patients without markers of viral replication in serum indicating that sexual transmission could still occur and this is of importance in relation to current vaccination policies.