Cytomegalovirus (CMV) Identification and Quantification from the Saliva of Human Immunodeficiency Virus (HIV) Seropositive and Seronegative Patients using Real Time Polymerase Chain Reaction

Divya, Uppala (2011) Cytomegalovirus (CMV) Identification and Quantification from the Saliva of Human Immunodeficiency Virus (HIV) Seropositive and Seronegative Patients using Real Time Polymerase Chain Reaction. Masters thesis, Ragas Dental College & Hospital, Chennai.


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INTRODUCTION: Oral lesions are a common manifestation of human immunodeficiency Virus (HIV) infection, with up to 90% of HIV infected patients developing an oral lesion during the course of their HIV disease. These oral lesions are often an early finding in HIV infection and are useful markers of disease progression and immune suppression. Common HIV related oral conditions include candidiasis, gingivitis, intraoral pigmentation, periodontitis, oral hairy leukoplakia, ulcers, Kaposi`s sarcoma, Non Hodgkin`s lymphoma, salivary gland disease including xerostomia and sialadenitis. Opportunistic infections play an important role in immunocompromised patients. Among the viral opportunistic pathogens, the Human Herpes Group (HHV) has been implicated in various oral lesions5. In the HHV group, Cytomegalovirus (CMV) of the Herpesvirinae subfamily has been studied the least. Although most CMV infections are asymptomatic, certain patient groups are at increased risk in developing serious illness. This virus remains the leading cause of congenital viral infection and a significant cause of transfusion-acquired infections in patients who are immunocompromised. It is a frequent contributor to morbidity and mortality among organ transplant recipients as well as subjects infected with HIV. CMV infection is also believed to accelerate the course of HIV disease to the Acquired Immunodeficiency. Syndrome (AIDS). CMV disease typically occurs when latent virus reactivates in AIDS patients with CD4 cells less than 10028. CMV is a well known pathogen causing various systemic disorders. Delayed diagnosis can lead to complications including CMV retinitis, pneumonia, hepatitis, encephalitis and leucopenia of which CMV retinitis is the most commonly manifested one. Studies from our centre have reported a significant percentage of ulcers among immunocompromised persons, some being herpetic and the others being apthous and the rest being non specific in origin. It has been postulated that CMV plays an important role in the pathogenesis of ulcerations of the mucocutaneous and the gastrointestinal tract and causes salivary gland dysfunction. Few studies have linked the role of CMV to its intra oral manifestations, the most common being the formation of non specific oral ulcers. Clinically, CMV-associated ulcers are non-specific and involve either the keratinized or non keratinized tissues. Microscopically, at certain times, typical cytomegalic inclusion bodies can be seen. While there are a very few studies where CMV was detected in ulcers of the mucocutaneous region, there are no documented evidences linking the viral load to non specific ulcers in the oral cavity. Thus, this study attempts to ascertain the efficacy of the normally used techniques such as Polymerase Chain Reaction (PCR) as a diagnostic tool. As CMV commonly manifests itself as CMV retinitis, this criteria has been used to select this study group. To further aid in the accuracy and sensitivity of the study, Real time polymerase chain Reaction (PCR) was been used as a diagnostic tool as it has shown to be rapid and effective in the diagnosis of CMV related ocular diseases. Efforts of correlation between non specific intra oral ulcers, the time lag till CMV retinitis develops and detecting the viral load by Real Time PCR in compromised patients can aid in diagnosing and starting proper antiviral drugs at the right time thereby preventing the disease to reach advanced stages which often have fatal consequences. AIM AND OBJECTIVES: 1. To assess and quantify the Cytomegalovirus (CMV) carriage in HIV seropositive and seronegative patients by Real Time PCR for CMV morphologically transforming region (mtr II) sequence. 2. To correlate its presence with oral findings. STUDY DESIGN: A cross sectional study was done to detect and quantify CMV in unstimulated saliva from HIV seropositive and seronegative individuals using quantitative Real Time PCR technique for CMV morphologically transforming region (mtr II) sequence. STUDY GROUPS: Group I: Study group (n = 5). HIV seropositive patients diagnosed with CMV retinitis and /or nonspecific oral ulcers. 1. Human Immunodeficiencey Virus seropositivity confirmed by Western Blot/ ELISA. 2. CMV retinitis diagnosed as per the recommended diagnostic criteria*, of Sankara Netralaya. *Indirect Ophthalmoscopy and Slit Lamp Technique for Typical Cases. Group II: Study group (n = 5). HIV seronegative patients clinically diagnosed with retinitis/nonspecific oral ulcers. EXCLUSION CRITERIA: • Patients on antiviral drugs to treat CMV infection were not included in either Group I or Group II*. • Ulcers caused by trauma (mechanical, chemical or thermal) or as a result of herpetic stomatitis were not considered. STUDY SETTING: Saliva samples were collected from the patients attending the outpatient wing of RAGAS - YRG Care, VHS, and Sankara Netralaya, Vision Research Foundation (VRF), Chennai. Demographic details of the patient, including the name, age, gender, habits, route of HIV transmission, past medical history, routine blood count and list of current medications taken by the patient were recorded. Ethical clearance was obtained from the institutional review board of Ragas Dental College and YRG-CARE, Chennai. An informed consent formatted for both seropositive and seronegative patients was obtained. A thorough oral examination was done by a trained dental surgeon and the findings were recorded in a pre-structured case sheet. Saliva samples were collected from patients, and stored at - 70° Celsius in the Department of Oral & Maxillofacial Pathology at Ragas Dental College & Hospital. DNA extraction and Real Time PCR was conducted in the Sankara Netralaya, VRF, Chennai. This study was done to detect and quantify CMV in the saliva of HIV seropositive and seronegative patients with active CMV retinitis. • 5 HIV seropositive and 5 HIV seronegative patients were included in the study. • All the study participants were males. • Mean age of the patients in the study group was 35.12 ± 2 years and in control group, it was 28.7 ± 5 years. • Oral ulcers were present in two patients, one in Group I and one in Group II. • DNA extraction was done from all the samples and quantified for cytomegalovirus • CMV DNA was quantified in the saliva in 2 samples in group I and 1 sample in group II above the threshold limit using the Real Time – PCR. • Saliva can be used as an diagnostic tool to assess the CMV infection. In our study, patients with CMV retinitis who had intraoral non-specific ulcers and xerostomia had significant CMV viral load as detected by Real Time PCR. We hypothesize that non specific ulcers not responding to regular treatment, in immunocompromised patients could be indicative of CMV retinitis and associated morbidity. We also conclude that saliva could be recommended as a diagnostic tool for quantifying CMV viral load especially in those patients who present with xerostomia and oral ulcers. More studies are necessary in order to enhance the methods used to detect CMV DNA and the sample collection techniques, so as to shed more light on the CMV infection and its prevalence in the oral cavity of HIV seropositive patients.

Item Type: Thesis (Masters)
Uncontrolled Keywords: Cytomegalovirus (CMV) Identification ; Quantification ; Saliva ; Human Immunodeficiency Virus (HIV) ; Seropositive and Seronegative Patients ; Real Time Polymerase Chain Reaction.
Subjects: DENTAL > Oral Pathology and Microbiology
Depositing User: Kambaraman B
Date Deposited: 12 May 2018 05:14
Last Modified: 12 May 2018 05:14

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