Rajiv Gandhi University of Health Sciences s54

RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES, BANGALORE, KARNATAKA.

ANNEXURE II

PROFORMA FOR REGISTRATION OF SUBJECT FOR

DISSERTATION

1. NAME OF THE CANDIDATE AND ADDRESS:

DR. MOHAMMED IQBAL

P.M.N.M. DENTAL COLLEGE AND HOSPITAL

BAGALKOT-587101

KARNATAKA

2. NAME OF THE INSTITUTION:

BAGALKOT-587101

KARNATAKA

3. COURSE OF STUDY AND SUBJECT:

M.D.S. (MASTER OF DENTAL SURGERY)

ORAL MEDICINE AND DIAGNOSTIC RADIOLOGY

4. DATE OF ADMISSION TO COURSE:

31th MAY 2010 5. TITLE OF THE TOPIC:

“ SALIVARY GLUCOSE LEVELS AND ORAL CANDIDAL CARRIAGE IN

TYPE 1 AND TYPE 2 DIABETES MELLITUS PATIENTS”

6. BRIEF RESUME OF THE INTENDED WORK:

6.1 NEED FOR THE STUDY:

Diabetes mellitus (DM) is a complex multisystemic metabolic disorder characterized by a relative or absolute insufficiency of insulin secretion and / or concomitant resistance to metabolic action of insulin on target tissues. It is estimated that there will be over 230 million people with DM by the year 2010 and half of this population will be in Asia.

Diabetes has profound and variable effects on the oral tissues. Patients with poor glycemic control are particularly prone to severe and/ or recurrent oral bacterial or fungal infections. Oral candidiasis is reported to be more prevalent among diabetics than in normal individuals1, 2.

The health of the oral tissues is known to be related to saliva and both the composition and flow of saliva may be altered in diabetics. The in vitro study in the past and recent in vivo studies have provided the evidence that the saliva of DM patients encouraged the candidal growth. Higher salivary glucose levels (SGL) are shown to be associated with increased candidal colonization in such patients3, 4, 5.

Numerous studies have reported higher SGL levels in diabetics than in controls with significant correlation with blood glucose levels (BGL) 6. Despite implications in the literature that possible higher glucose levels in saliva of diabetic patients could predispose to oral candidal infections, this hypothesis to our knowledge has not been widely explored in the Indian population. This project is undertaken to explore the relationship between the salivary glucose levels and candida colonization in DM patients and healthy subjects visiting the dental and medical OPD’s of Bagalkot town of North Karnataka, India.

6.2 REVIEW OF LITERATURE:

Group of authors conducted the study to determine whether hyperglycemia in IDDM could interfere with salivary glucose levels and salivary microbial counts (yeast count and lactobacillus count). Salivary factors like salivary flow rate, SGL and salivary microbial counts were performed on two study groups. Group1

(n=14) consisted of newly diagnosed IDDM cases with diagnosed cases of IDDM (n=50) forming the Group2. Glycosylated haemoglobin (HbA1) values were determined for both groups. Higher salivary glucose levels, lower salivary flow rates, higher mean blood glucose levels in the newly diagnosed IDDM cases and higher Hb1A values in the long- term IDDM cases were observed with increasing yeast and lactobacilli counts.

The study supported the view that the high glucose levels (BGL & SGL) and hyperglycaemic status (HbA1) favoured the yeast adherence and colonization in diabetes mellitus patients7.

Four hundred and twelve patients (218 males, 194 females) were studied after collecting venous blood and saliva for assessing the relationship between glycemic control and candida carriage rate. Glycemic control was assessed by random blood glucose (hexokinase method) and glycosylated haemoglobin (HbA1) as estimated by gel electrophoresis method (normal range 5.8%) The oral rinse technique and Sabourauds dextrose agar was used for candida isolation and culture .The yeast isolates were identified by germ tube production and sugar assimilation.

Candida was isolated in 210 diabetics (51%) with higher carriage rate observed in diabetic denture wearers than dentate diabetic patients Although, no association was observed between carriage rate and glycemic control, the authors concluded that in certain sub groups the carriage rates were higher and involved some uncommon candidal species8

The glucose concentration in unstimulated mixed saliva and serum was assayed and correlated with oral candida colonization in 41 diabetics (52±16 yrs) and 34 healthy controls (52±18 yrs) by group of authors. The concentrated oral rinse technique was used for quantitative oral candildal isolation. An enzymatic ultraviolet detection method was used for salivary glucose estimation. In diabetic patients, salivary glucose concentration was significantly higher than in controls (P<0.02). Diabetic patients who harboured candida species intra-orally (56.1%) had higher SGL than those in whom candida could not be isolated, although the difference in frequency and quantity of oral candidal isolation failed to reach the significance between two groups. The median number of candida isolated in DM patients was 1300 cfu/ml (range 140-5280 cfu/ml) whereas in non-diabetic subjects (41%) the median number was 200 cfu/ml with the range of 40-460 cfu/ml. The findings of this study suggest that high SGL levels appear to encourage the growth of candida species in DM patients5.

The group of authors conducted the study to evaluate the relationship between SGL and salivary candidal carriage and to determine if SGL could be used as a noninvasive tool to monitor glycemic control in diabetics. A total of 150 adults (100 with type 2 DM and 50 controls, aged 40-60 yrs) participated in the study.

Both unstimulated and stimulated saliva were collected and investigated for salivary glucose levels (glucose oxidase method) and colony forming units (CFU) of candida by using oral rinse technique. Random non-fasting plasma glucose (RNFPG) levels and glycosylated haemoglobin (HbA1c) were determined for the study groups.

Salivary glucose levels were significantly higher in diabetic subjects than in non- diabetic subjects. Both unstimulated and stimulated salivary glucose (USSG,

SSG) showed significant positive correlation with blood glucose and CFUs of candida.

Candidal CFUs were significantly higher in diabetic subjects than in non-diabetic subjects. The median range of CFU was significantly higher in Group 2 DM (RNFPG values>200mg/dL,uncontrolled diabetes) than in Group 1 DM(RNFPG values<200mg/dL, controlled diabetic). CFU of candida showed a positive correlation with USSG (r=0.519:P<0.000) and SSG (r=0.611:P=0.000) in the study population . The authors concluded that increased candida reflects increased salivary glucose levels.

Increased salivary glucose has potential for increased susceptibility for oral candidiasis9

6.3 OBJECTIVES OF THE STUDY:

1. To study the salivary glucose levels and oral candidal carriage in diabetic subjects

in comparision with healthy controls.

2. To investigate the prevalence of different candidal strains.

3. To investigate the relationship between salivary glucose concentrations with oral

candidal carriage in a group of diabetic subjects and matched controls.

7. MATERIALS AND METHODS:

7.1 SOURCE OF DATA:

The study design includes 60 DM subjects attending Diabetic OPD Clinic at

S.N. MEDICAL COLLEGE AND HOSPITAL, BAGALKOT and 30 non-diabetic healthy dental patients attending the OPD of P.M.N.M DENTAL COLLEGE AND

HOSPITAL, BAGALKOT for routine dental care like oral prophylaxis and fillings.

. a. Inclusion criteria: -

Confirmed diagnosed cases of Type 1(n=30) and Type 2 DM (n=30) for more than 3-5 yrs of duration and are being treated for diabetes are included in the study.

The subjects are grouped as:-

Group1:- 30 patients with Type1 diabetes mellitus

Group2:- 30 patients with Type2 diabetes mellitus

Group3:- 30 age, sex matched healthy non-diabetic individuals b. Exclusion criteria: -

Patients with history of salivary gland surgeries, diagnosed with malignancy, under long term local and systemic drug therapy (antibiotics, steroids etc),

HIV positive individuals, heavy smokers, pregnant women and individuals with systemic illnesses other than diabetes mellitus and any oral mucosal lesions are excluded from the study.

Details of history (duration, type, glycemic control, medications), clinical and oral examination data, biochemical values obtained are recorded on the specially prepared pro-forma.

7.2 METHOD OF COLLECTION OF DATA:

Collection of samples: -

1) Saliva: - Approximately 2 ml of unstimulated whole saliva is collected from study and control groups in a sterile graduated tube by ‘Spitting method’ over a period of 5 minutes.

Saliva is collected in resting position after rinsing the mouth with distilled water between

8.00a.m and 10.30 a.m. Patient is asked not to eat or drink for 2 hours prior to collection.

Saliva thus obtained is stored at -20 degree Celsius, for no more than 2 hours or sent to the laboratory immediately. It is centrifuged at 2000 rpm for 5 minutes and subjected to analysis.

2) Serum: - Under aseptic conditions 2 ml of patients intravenous blood is obtained from median cubital vein of forearm, centrifuged at 2000 rpm, serum thus obtained is analyzed. 3) Serum Glucose and Salivary Glucose determination: - Serum and salivary glucose are assayed by use of Glucose Oxidase Peroxidase method- GOD- POD- Enzymatic

Colorimetric method using test kit- UV- 1601 VISIBLE SPECTROPHOTOMETER,

SHIMADZU CORPORATION, KYOTO, JAPAN.

4) Salivary sampling and assessment of yeast count: - The salivary samples are collected using Oral Rinse Technique. The patients are asked to thoroughly rinse the oral cavity using 10ml of sterile phosphate-buffered saline solution (0.1M, pH-7.4) for 60 sec.

The expectorate is collected in a sterile universal container and stored at -4 degree

Centigrade till the transport to microbiology laboratory for further analysis. Candida isolation, quantification and strain characterization are done using Sabouraud dextrose agar (SDA), chromogenic agar culture (CHROMagar Candida) and appropriate biochemical tests.

All the samples are assayed on the same day of saliva collection. Analysis and interpretation of test values will be done in S.N. Medical College, Department of

Microbiology and Biochemistry and Pharmacology department, Bagalkot. Results obtained will be statistically analyzed appropriately & interpreted. 7.3 DOES YOUR STUDY REQUIRE ANY INVESTIGATION

INTERVENTIONS TO BE CONDUCTED ON PATIENTS OR

OTHER HUMANS OR ANIMALS?

7.4 HAS ETHICAL CLEARANCE BEEN OBTAINED FROM

YOUR INSTITUTION IN CASE OF 7.3?

8. LIST OF REFERENCES:

1. Manfredi M, McCullough MJ, Vescovi P, Al-kaaarawi ZM, Porter SR. Update on

diabetes mellitus and related oral diseases. Oral Dis 2004; 10:187-200.

2. Soysa NS,Samaranayake LP,Ellepola ANB. Diabetes mellitus as a contributory

factor in oral candidosis. Diabetic Med 2005:23; 455-9.

3. Scully C and Kabir M EL, Samaranayake LP. Candida and Candidosis: A

Review. Cri Rev Oral Bio Med 1994;5(2):125-157 4. Knight L, Fletcher J. Growth of candida albicans in saliva stimulated by glucose

associated with antibiotics, corticosteroid and diabetes mellitus, J Infect Dis

1971;123:371-7

5. Darwazeh AMG, MacFarlane TW, McCuish A, Lamey P.J. Mixed salivary

glucose levels and candidal carriage in patients with diabetes mellitus. J Oral

Pathol Med 1991;20:280-283

6. Amer S,Yousuf M,,Siddiqui PQ, Alam J. Salivary glucose concentrations in

patients with diabetes mellitus- a minimally invasive technique for monitoring

blood glucose levels. Pak J Pharm Sci; 2001;14:33-37.

7. Karjalainen KM, Knuuttila ML, Kaar ML. Salivary factors in children and

adolescents with insulin-dependent diabetes mellitus. Paediatric Dentistry

1996;18;4;306-11

8. Fisher BM, Lamey P-J, Samaranayake LP, Macfarlane TW, Frier BM.

Carriage of candida species in the oral cavity in diabetic patients: - relationship to

glycemic control. J Oral Pathol 1987; 16: 282-284.

9. Sashikumar R, Kannan R. Salivary glucose levels and candidal carriage in type 2

diabetics .Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2010; 109:706-711.

9. SIGNATURE OF THE CANDIDATE:

10. REMARKS OF THE GUIDE: SATISFACTORY 11.1 NAME AND DESIGNATION OF THE GUIDE:

DR. SUREKHA R.PURANIK M. D. S

PROFESSOR

DEPARTMENT OF ORAL MEDICINE, RADIOLOGY AND DIAGNOSIS

BAGALKOT-587101

KARNATAKA

11.2 SIGNATURE OF THE GUIDE:

11.3 HEAD OF THE DEPARTMENT:

DR. MANGALA METI M. D. S

PROFESSOR AND HEAD

DEPARTMENT OF ORAL MEDICINE, RADIOLOGY AND DIAGNOSIS

BAGALKOT-587101

KARNATAKA 11.4 SIGNATURE OF THE HEAD OF THE DEPARTMENT:

12.1 REMARKS OF THE PRINCIPAL:

12.2 SIGNATURE OF THE PRINCIPAL: