Comparison of real time polymerase chain reaction with microscopy and antigen detection assay for the diagnosis of malaria.

OBJECTIVE: To determine the sensitivity of a real time polymerase chain reaction (PCR) for malaria diagnosis and to compare its accuracy with microscopy and an antigen based rapid diagnostic test (OptiMal). STUDY DESIGN: Cross-sectional analytical study. PLACE AND DURATION OF STUDY: Military Hospital, Armed Forces Institute of Transfusion and Armed Forces Institute of Pathology, Rawalpindi, from July to December 2011. METHODOLOGY: Venous blood samples of 300 clinically suspected patients of malaria were tested for malaria parasite by microscopy and OptiMal; and malaria parasite index was calculated for the positive samples. Plasmodium genus specific real time PCR was performed on all specimens, targeting small subunit rRNA gene. Diagnostic accuracy of three tests was compared and cost analysis was done. RESULTS: Out of 300 patients, malaria parasite was detected in 110, 106 and 123 patients by microscopy, OptiMAL and PCR respectively. Real time PCR was 100% sensitive while microscopy and OptiMal had sensitivity of 89.4% and 86.2% respectively. All methods were 100% specific. The cost per test was calculated to be 0.2, 2.75 and 3.30 US$ by microscopy, OptiMal and PCR respectively, excluding the once capital cost on PCR equipment. CONCLUSION: Genus specific real time PCR for the diagnosis of malaria was successfully established as a highly sensitive and affordable technology that should be incorporated in the diagnostic algorithm in this country.

Serum leptin values in the healthy obese and non-obese subjects of Rawalpindi.

OBJECTIVE: To determine serum leptin concentrations from a sample of Rawalpindi population in relation to body mass index, age and gender. METHODS: The observational, comparative study was conducted at the Armed Forces Institute of Pathology, Rawalpindi, and Benazir Bhutto Hospital, Rawalpindi from August 2008 to December 2008. Subjects were 100 including healthy obese, overweight and non-obese of both genders aged between 20-50 years. Sampling was done by non-probability convenience method. Body Mass Index was calculated by formula BMI = weight in kg/height in m2: non-obese subjects were defined as 18.5-23.0 kg/m2; overweight 23.1-27.4 kg/m2; and obese 27.5-40 kg/m2. Serum glucose was measured using Glucose oxidase-phenol amino phenazone (GOD-PAP) method and serum leptin by sandwich enzyme-linked immunosorbent assay method. RESULTS: Serum leptin concentrations were higher in obese subjects (mean 52.8 +/- 24.6 ng/mL; range 28.2-77.4 ng/mL; P < 0.001) than in non-obese subjects (mean 12.7 +/- 6.1 ng/mL, range 6.6-18.8ng/mL). Mean Body Mass Index in obese group was 31.7 +/- 3.1 kg/m2 (range 28.6-34.8 kg/m2) while it was 21.2 +/- 1.5 kg/m2 (range 19.7-22.7 kg/m2) in the nonobese group. Body Mass Index was strongly positively correlated with serum leptin concentration (r = 0.59, p < 0.001) in the obese group. The mean serum leptin concentration was much higher in the healthy obese and non-obese women (64.4 ng/mL and 8.7 ng/mL respectively) than in men of both categories (40.4 ng/mL and 5.5 ng/mL respectively). Age had no significant relation with serum leptin level (p = 0.416). CONCLUSIONS: In the study sample, serum leptin concentration was positively correlated with Body Mass Index in healthy obese and non-obese subjects of both genders. The levels were higher in women than in men. Age had no significant relation with serum leptin level in this age group.

Frequency of conversion to overt hypothyroidism in patients with and without subclinical hypothyroidism.

BACKGROUND: Sub-clinical hypothyroidism (SCO) poses diagnostic and management difficulties for pathologists and clinicians. Cases of SCO are now diagnosed with increasing frequencies worldwide mainly because of availability of more sensitive methods for serum TSH, total T3 and free T4 assays. The rate of progression of SCO to overt hypothyroidism varies in different populations. Sub-clinical hypothyroidism has been suggested as a risk factor for hypothyroid complications. There is a need to identify and treat patients with SCO before they convert to overt hypothyroidism and develop complications. Objective was to compare the development of overt hypothyroidism in a cohort of patients of sub-clinical hypothyroidism and in subjects with normal thyroid function tests. METHODS: It was Cohort study conducted at Department of Chemical Pathology and Endocrinology, Armed Forces Institute of Pathology, Rawalpindi. Hundred patients of SCO and almost equal number of age and sex-matched subjects with normal Thyroid function test (TFT) were enrolled as healthy controls. Subclinical hypothyroid patients and controls were followed for a period of one year on six monthly bases. The patients were examined for signs and symptoms of hypothyroidism and serum TSH, total T3 and free T4 were estimated. The clinical history, physical examination and thyroid function tests were recorded on the pre-designed Performa. RESULTS: Fourteen (14%) out of 100 cases of the SCO patients developed overt hypothyroidism, SCO had 2.8 times more risk for conversion to overt hypothyroidism as compared to healthy controls. Moreover female gender and initial levels of serum TSH were the most important predictors for conversion of SCO to overt hypothyroidism. CONCLUSION: The frequency of conversion to overt hypothyroidism in SCO patients approaches 14% and was more commonly observed in female patients.

Pro/anti-inflammatory cytokines in the pathogenesis of premature coronary artery disease.

Proinflammatory interleukin-18 (IL-18), high-sensitivity C-reactive protein (hS-CRP), tumor necrosis factor-alpha (TNF-alpha), and anti-inflammatory IL-10 are involved in the pathogenesis of atherosclerosis. The aim of this study was to determine the imbalance between pro- and anti-inflammatory cytokines in premature coronary artery disease (PCAD) and their association with the degree of angiographic atherosclerotic blockade. A case-control study was carried out at the National Institute of Heart Diseases, Rawalpindi, Pakistan. Three hundred eighty-five stable coronary artery disease patients aged <45 years were screened. A total of 172 subjects participated in this study, comprising 98 PCAD patients and 74 angionegative controls. Serum IL-10, IL-18, and TNF-alpha were measured using enzyme-linked immunosorbent assay and hS-CRP was analyzed using Immulite 1000. The mean age of the patients was 40 ± 4.23 years (69 men and 9 women). Serum IL-18, TNF-alpha, hS-CRP, and IL-18/IL-10 ratio were significantly raised in PCAD patients (P<0.01), whereas IL-10 was moderately increased (P<0.05) when compared with controls. Serum IL-18 and hS-CRP were significantly raised in patients with 86%-100% stenosis (n=57) when compared with patients with 71%-85% stenosis (n=41) (P<0.05). Serum IL-18, TNF-alpha, IL-18/IL-10 ratio, and hS-CRP significantly correlated (P<0.01) with the degree of angiographic blockade. Pro/anti-inflammatory cytokines play a vital role in the pathogenesis of PCAD and have potential to identify the degree of atherosclerosis.

Impact of CYP2C9 genetic polymorphism on warfarin dose requirements in Pakistani population.

Variations of cytochrome-P450 enzyme system (CYP2CP) are associated with impaired metabolism of warfarin. The objective of our study was to estimate the frequency of genetic and allelic variants of CYP2C9 in Punjabi population of Pakistan and their effects on warfarin dose requirement. One hundred and twenty unrelated Pakistani subjects belong to Punjab province, were randomly included from the registry of National Institute of Heart Disease Rawalpindi, Pakistan. The patients had stable international normalized ratio (INR) of 2 to 3 for last 3 months with warfarin therapy after heart valves replacement. The detection of CYP2C9 variant was done on polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay. Total 120 patients (73 males; 47 females) of mean age of 37 years participated in the study. Nine patients had mutant allele CYP2C9*3 (7.5%), one CYP2C9*2 (0.8%) and 110 patients exhibited wild type CYP2C9*1 (91.7%). The frequency of CYP2C9 genotype was *1/*1 (0.858) ; *1/*3 (0.117) ; 2/*20 (0.08 ) and *3/*3 (0.017) in our study population. A high dose of warfarin (42.2+9.56) mg/week is required for patients with *1/*1 genotype as compared to patients with *2/*2 (17.5+1.9) and *1/*3 (16.6+2.3) allele (p<0.001). Individuals with CYP2C9*3/3* need lowest (8.75±1.76 mg/week) daily warfarin dose. In conclusion, the genetic variations in the CYP2C9 occur in 14% of Punjabi ethnic group in Pakistan. Presence of CYP2C9*2 or *3 variants is an independent predictor of low warfarin dose requirement in our patients. CYP2C9 variants assay may be used in high risk groups for appropriate dose adjustment to avoid complications on long term basis.

Risk assessment of pesticide exposure on health of Pakistani tobacco farmers.

Tobacco is an important cash crop of Pakistan. Pesticides are commonly used to increase the crop yield, but their health impact has not been studied yet. The objectives of the study were to determine the frequency of pesticide poisoning and to explore the knowledge, attitudes and practices (KAP) towards safety measures among the tobacco farmers in Swabi, Pakistan. One hundred and five tobacco farmers involved in pesticide application were randomly selected from two villages of district Swabi. A structured questionnaire was used for clinical and KAP information. Plasma cholinesterase (PChE) levels were measured by Ellman's method by using GD Italy kits. All tobacco farmers were males with a mean (SD) age of 26 (9) years. The majority of the farmers reported multiple symptoms headache, dizziness, vomiting, shortness of breath, muscle weakness and skin rash correlate with the clinically significant depression of PChE levels. Out of 105 pesticide applicators, 58 (55%) had post-exposure reduction in PChE levels <20% from baseline, 35 (33%) had mild poisoning (20-40% reduction) and 12 (11%) had moderate poisoning (>40% reduction). Most of the farmers did not use any personal protective equipment during pesticide handling. Only a few used shoes (31%), masks (14%) and gloves (9%) during pesticide spray. In conclusion, the tobacco farmers had mild to moderate pesticide poisoning, which was correlated with depression in PChE levels. Moreover, most farmers had little knowledge about the safety measures, casual attitude and unsatisfactory safety practices with regard to the use of basic protective equipments during pesticide applications on the tobacco crop.

Psychological autopsy of suicide-a cross-sectional study.

BACKGROUND: Psychological autopsy is the reconstruction of events leading to death. There are few studies on psychological autopsy. AIM: To understand the profile of suicide completers and find out ways of dealing with it. METHODS: Fifty suicide cases were analysed. Using a semi-structured, self-designed questionnaire, the family, friends and relatives of the deceased were interviewed. RESULTS: The presence of some type of psychiatric disorder and stressful life events are two important reasons for committing suicide. CONCLUSION: Psychological autopsy is a very important tool for assessing the causes and precipitants of suicide. More and more studies in this field are required with a larger sample size for the evaluation of suicides.