A Clinical Study on the Circadian Rhythm of Salivary Cortisol on Aggressive Periodontitis and Its Correlation with Clinical Parameters using Electrochemiluminescence Immunoassay Method.

AIM: Periodontal pathologies are gaining importance as there is a clear indication of bi-way control on general homeostasis of an individual. The study of HPA axis in various diseases has proved that there is evident vulnerability existing for any organism when the Cortisol diurnal rhythm is altered. The aim was to compare the diurnal rhythm of salivary cortisol in aggressive periodontitis with control patients. This study also compared various parameters like body mass index (BMI), waist circumference, Hamilton anxiety scale, OHI-S, clinical attachment loss in aggressive periodontitis. MATERIALS AND METHODS: 30 control patients were compared against 30 aggressive periodontitis patients in Salivary cortisol diurnal rhythm. It was estimated using the electrochemiluminescence (ECL) method on a 3 point analysis-Soon after waking up, 30 minutes after waking up, 1 hour before sleep to see the diurnal variation in aggressive periodontitis patients. The samples were transferred to CABRI labs to be frozen to -20°C. The analysis was done using Cobas e-411 autoanalyzer by Roche, USA. RESULTS: The average cortisol in aggressive patients was found to be higher compared to control patients and was found to be statistically significant with a p value of 0.012. Control group is moderately skewed left (negative skewness graph) while the aggressive p periodontitis patients showed moderately skewed right (+ve skewness graph). CONCLUSION: The cortisol awakening response seen in control patients is not observed in aggressive periodontitis. Instead of giving a surge, the cortisol showed a dip in the first 30 minutes followed by a gradual increase in aggressive periodontitis instead of decline as observed in normal patients. CLINICAL SIGNIFICANCE: The study will focus on the importance of cortisol circadian rhythm on periodontal health allowing the microorganism to create an environment of dysbiosis.

Allelic variants of ADH, ALDH and the five factor model of personality in alcohol dependence syndrome.

BACKGROUND: The etiology of alcohol dependence is a complex interplay of biopsychosocial factors. The genes for alcohol-metabolizing enzymes: Alcohol dehydrogenase (ADH2 and ADH3) and aldehyde dehydrogenase (ALDH2) exhibit functional polymorphisms. Vulnerability of alcohol dependence may also be in part due to heritable personality traits. AIM: To determine whether any association exists between polymorphisms of ADH2, ADH3 and ALDH2 and alcohol dependence syndrome in a group of Asian Indians. In addition, the personality of these patients was assessed to identify traits predisposing to alcoholism. MATERIALS AND METHODS: In this study, 100 consecutive males with alcohol dependence syndrome attending the psychiatric outpatient department of a tertiary care service hospital and an equal number of matched healthy controls were included with their consent. Blood samples of all the study cases and controls were collected and genotyped for the ADH2, ADH3 and ALDH2 loci. Personality was evaluated using the neuroticism, extraversion, openness (NEO) personality inventory and sensation seeking scale. RESULTS: Allele frequencies of ADH2*2 (0.50), ADH3*1 (0.67) and ALSH2*2 (0.09) were significantly low in the alcohol dependent subjects. Personality traits of NEO personality inventory and sensation seeking were significantly higher when compared to controls. CONCLUSIONS: The functional polymorphisms of genes coding for alcohol metabolizing enzymes and personality traits of NEO and sensation seeking may affect the propensity to develop dependence.

Polymerase chain reaction in rapid diagnosis of neonatal sepsis.

In a prospective study a total of hundred neonates who fulfilled the American College of Obstetrics and Gynecology's (ACOG) criteria for probable sepsis admitted to NICU of tertiary care armed forces hospital were investigated for evidence of sepsis. The investigation protocol included sepsis screen, blood culture and 1 mL of venous blood for molecular analysis by polymerase chain reaction (PCR) for bacterial DNA component encoding 16 s RNA in all cases. 100 newborns with probable sepsis were studied to evaluate the molecular diagnosis of sepsis using PCR amplification of 16 S RNA in newborns with risk factors for sepsis or those who have clinical evidence of sepsis. We compared the results of PCR with blood culture and other markers of sepsis screen (total leucocyte count (TLC), absolute neutrophil count (ANC), immature/total neutrophil count ratio (I/T ratio), peripheral blood smear, micro ESR and C reactive protein (CRP). Controls consisted of 30 normal healthy newborns with no overt evidence of sepsis. Sepsis screen was positive in 24 (24%) of cases in study group with sensitivity and specificity of 100% and 83.5% respectively. Blood culture was positive in 09(9%t) with sensitivity of 69.2% and specificity of 100%. PCR was positive in 13(13%) of cases (9% are both blood culture and sepsis screen positive and 4% are positive by sepsis screen); the sensitivity of PCR was 100% and specificity was 95.6%. Blood culture is the most reliable method for diagnosis of neonatal sepsis. Polymerase chain reaction is useful and superior to blood culture for early diagnosis of sepsis in neonates.

Outbreak of Multi-drug Resistant Salmonella Typhi Enteric Fever in Mumbai Garrison.

BACKGROUND: There was an epidemic of enteric fever in Mumbai garrison during Nov-Dec 2000 with more than 150 cases admitted to a tertiary care service hospital. METHODS: All the cases presented with fever and some had splenomegaly, bradycardia, abdominal pain and diarrhoea. The epidemic was investigated by the station health organization (SHO) and the case and bacteriological study was carried out in pathology laboratory of the service hospital. The serological study was carried out at Armed Forces Medical College (AFMC), Pune and the Phage typing was carried out at Lady Harding Medical College, New Delhi. RESULTS: Blood cultures were positive in 92(63%) for Salmonella typhi and Widal test was positive in 83(55%). All strains were resistant to four primary drugs i.e. ampicillin, chloramphenicol, co-trimoxazole and tetracycline. All but two were treated successfully with ceftriaxone. The Salmonella typhi belonged to phage group E1 and biotype I. CONCLUSION: Extensive epidemiological investigation of cases and water sources of cantonment area pointed to a common source of the epidemic i.e. the well near 'Gurudwara'.

Phylogenetic conservation of the stem-loop III structure of the 5' untranslated region of Hepatitis C virus RNA among natural variants in samples collected from Southern India.

The stem-loop III (SLIII) structure within the 5' untranslated region has been shown to be critical for internal initiation of translation of Hepatitis C virus (HCV). Using 'Single Strand Conformation Polymorphism (SSCP)' of the SLIII region we have investigated for natural mutations and demonstrated presence of some non-covariant changes in certain sub-domains. However, overall SLIII-RNA structure was found to be phylogenetically conserved. Additionally, by SSCP analysis we have determined the genotype of 50 HCV isolates collected from Southern India, 25 random samples were confirmed by DNA sequencing. Results showed the prevalence of genotype 1 in this part of India.

Random Amplification of Polymorphic DNA Based Typing of Pseudomonas Aeruginosa.

Pseudomonas aeruginosa was isolated from various sources during the course of an epidemic outbreak of bacterial endophthalmitis following an eye camp at Sangli, Maharashtra. 15 distinct isolates were obtained from clinical samples. Typing of the 15 isolates was performed by random amplified polymorphic DNA (RAPD) analysis, pyocin typing and antibiogram. RAPD typing was rapid, labour friendly and could be done within six hours. RAPD analysis produced reproducible electrophoretic band patterns on the basis of which three distinct amplification patterns could be visualised. The conventional typing methods were labour intensive and took about 48 hours. However, the results of RAPD typing, pyocin typing and antibiogram did not correlate with each other. This study suggests that RAPD typing could be an additional rapid typing method for studying the epidemiology of infectious disease outbreaks due to P aeruginosa.

HIV - 1 SUBTYPES, ITS IMPLICATIONS AND VIRAL DYNAMICS.

Subtyping of HIV has important implications for developing candidate vaccine and understanding the biological behaviour and dynamics of HIV transmission in various populations. The third variable region (V3) in the envelope gene of HIV-1 has been shown to be a major determinant influencing a number of biological characteristics of the virus. HIV-1 evolves by rapid mutation and by recombination, both processes actively contributing to its genetic diversity. Most of the multiple genetic subtypes and intersubtype recombination of HIV-1 that comprise the global pandemic have not been characterized by full genome sequencing. The development of an effective human immunodeficiency virus type-1 (HIV-1) vaccine is likely to depend on knowledge of circulating variants of genes other than the commonly sequenced gag and env genes.

A STRATEGY FOR RAPID IDENTIFICATION OF METHICILLIN RESISTANT STAPHYLOCOCCUS AUREUS NASAL CARRIER STATUS.

Methicillin Resistant Staphylococcus aureus (MRSA) is a multi drug resistant organism responsible for severe outbreaks of life threatening infections in hospitals which are difficult to treat They are spread by nasal carriage among the hospitalised patients, staff and visitors. Mannitol cloxacillin salt agar (MCSA) is a single tube method to identify MRSA. However, tubes showing growth and change in colour on biochemical characterisation often do not prove to be MRSA. In this study we have combined two strategies for the rapid identification and isolation of MRSA by culture in MCSA and multiplex PCR for mecA and femB genes. Anterior nasal swabs obtained from nursing staff and patients admitted to a large referral hospital, were inoculated into MCSA. Of the 100 tubes inoculated, 8 tubes showed change in colour and growth. On conventional testing 4 were MRSA, 3 were methicillin sensitive S aureus (MSSA) and 1 was Methicillin Sensitive Coagulase Negative S aureus (MSCNS). Genotyping by multiplex PCR revealed 5 MRSA, 2 MSSA and 1 MRCNS. The Multiplex PCR technique to rapidly identify presence of mecA and femB genes showed presence of both mecA and femB bands in all MRSA. The methicillin sensitive organisms showed absence of mecA gene while coagulase negative organisms showed absence of the fern B gene. Combining MSCA with multiplex PCR for mec A and fem B genes made the test both rapid and specific. Use of this strategy would enable rapid screening of nasal carriers and early implementation of hospital infection control measures.

Plasmid mediated antibiotic resistance in Klebsiella pneumoniae.

Klebsiella pneumoniae species (108) isolated from tertiary care hospitalized patients were investigated for antibiotic resistance patter. 74% isolates were from urine, 13.5% from pus, 4.5% from blood and 8% from sputum. The resistance pattern of the organisms to various antibiotics were as follows: ampicillin 93.3%, cefotaxime 70%, ceftazidime 81%, gentamicin 68.5%, amikacin 63.75%, netilmicin 74%, norfloxacin 55%, ofloxacin 53%, chloramphenicol 82%, tetracycline 85%, sulphamethoxazole 96%, trimethoprim 94%. 2-4 drug resistance was found in 34.5%, 5-8 drug resistance was in 25.1% and 9-12 drug resistance was found in 40.4% of strains. Out of 65 strains, 25 strains could be successfully conjugated. Common drugs which were transferred to transconjugants were sulphamethoxazole, trimethoprim, gentamicin, netilmicin, amikacin, chloramphenicol, tetracycline, ampicillin, cefotaxime and ceftazidime. A large plasmid of 98.7% kb could be demonstrated in these strains by alkali denaturation method and agarose gel electrophoresis.

COMPARISON OF RAPID METHOD OF DNA EXTRACTION USING MICROWAVE IRRADIATION WITH CONVENTIONAL PHENOL CHLOROFORM TECHNIQUE FOR USE IN MULTIPLEX PCR FOR mec A AND fem B GENES TO IDENTIFY GENOTYPES OF MRSA FROM CULTURES.

Methicillin Resistant Staphylococcus aureus (MRSA) infection is a major cause of morbidity and mortality in hospitalised patients and requires vancomycin for effective therapy. Rapid identification of MRSA is vital to control MRSA outbreaks in hospitals. Identification of MRSA is a time consuming process requiring more than 48 hours and is labour intensive involving culture, biochemical tests and antimicrobial susceptibility testing. In this study we have used microwave irradiation of the bacterium obtained from cultures which was then directly subjected to a multiplex PCR technique to accurately and rapidly identify the presence of mec A and fem B genes which characterise MRSA. This has been compared with the standard method of lysing the bacterium and DNA extraction using phenol chloroform method followed by multiplex PCR. The microwave lysis method followed by direct PCR has been found to be less time consuming, 5 hours, as compared to 9 hours by conventional technique. Use of this strategy would enable early identification and early implementation of control measures.

RECENT ADVANCES IN TUBERCULOSIS DIAGNOSTIC TECHNIQUES.

Tuberculosis is re-emerging as an important cause of morbidity and mortality in man. This article outlines current strategies available for the diagnosis of tuberculosis, and its applicability. Fluorescent staming, modified culture methods, antigen detection, ELISA based assays against various antigen preparation and recent advances in molecular techniques have been outlined. Present strategies being developed at Armed Forces Medical College for the early diagnosis, speciation, antibiotic sensitivity testing and epidemiologic testing have also been alluded to.

SURGEON AND HIV INFECTION: Post Exposure Prophylaxis: Need of the hour.

HIV infection in HCW is an occupational hazard requiring policy decisions by the health care administrators. In this article we have outlined a post exposure prophylaxis protocol following HIV exposure in HCWs. By determining the HIV status code of the source and the HIV exposure code of the HCWs, recommendations for PEP are forwarded. Occupational exposures should be considered urgent medical and surgical concerns to ensure timely administration of PEP. At the end, an algorithm is provided to guide exposed health-care workers in deciding when to consider PEP.

POLYMERASE CHAIN REACTION AND ADVANCES IN INFECTIOUS DISEASE DIAGNOSIS.

The use of Polymerase Chain Reaction (PCR) in infectious disease diagnosis, has resulted in an ability to diagnose early and treat appropriately diseases due to fastidious pathogens, determine the antimicrobial susceptibility of slow growing organisms, and ascertain the quantum of infection. This article outlines the PCR, some of its modifications and their application in infectious disease diagnosis.

T HELPER SUBSETS AND INTERLEUKIN 12: ITS ROLE AS A POTENT IMMUNOMODULATOR OF CELL MEDIATED IMMUNITY.

This article reviews Interleukin (IL) IL-12 a recently described cytokine secreted by monocyte/macrophages and its interaction with T cells. IL 12 is important in the immune response by providing a link between natural resistance mediated by phagocytic cells and NK cells and adaptive immunity mediated by T4, T8 and B cells. The article also evaluates the effect of IL12 in the evolution of T helper cell subsets, its action as a bridge between innate immunity and acquired immunity and the important role it plays in modulating the cellular and humoral immune response mechanisms. Its possible role as an immunomodulator of disease is postulated.

CLINICO-PATHOLOGICAL CORRELATION IN NEONATAL AUTOPSIES.

Of the 253 neonates admitted to a neonate intensive care unit during the period Jan 91 to Sep 93, 43 neonates died. Autopsy was done in 23 of these (53%). The mean duration of stay of the neonates in the intensive care unit prior to death was 5.6 days (range 2 hours to 10 days). Antemortem diagnoses included asphyxia neonatorum (4), meconium aspiration syndrome (2), septicemia (5), prematurity (3), birth trauma (2), congenital anomalies (2), hypoxic ischemic encephalopathy (1), and non-specific diagnosis (4). There were 6 major autopsy findings that, if known prior to death, would have altered clinical management and might have resulted in cure or prolonged survival. There were 8 additional major findings that, if known prior to death, would not have altered management There were 14 minor findings related to major diagnoses but unrelated to the primary cause of death.

SCANNING ELECTRON MICROSCOPIC STUDY OF HeLa CELL INVASION BY SALMONELLA TYPHI.

Invasion of cultured non-pliagoeytic cells by salmonella is illustrated by a scanning electron microscopie study of HeLa cell invasion by S. typhi. This study shows that after bacterial adherence the HeLa cell cytoplasmic membrane shows ruffling and formation of filopodia which gradually engulf the bacterium and draw it into the cytoplasm. The available literature is reviewed and the probable mechanism underlying phagocytosis is hypothesized.

Postal follow up of patients with epilepsy.

BACKGROUND: Epilepsy services in India are mostly located in urban areas and are often overcrowded. It is difficult, therefore, to organize long term management programmes. We report our experience at a tertiary referral centre on follow up of patients with epilepsy through regular postal review. METHODS: One hundred consecutive patients with epilepsy (63 men, 37 women, mean age 17 years) who had only seizures were followed up by post using a questionnaire, instead of reviewing them in a clinic. The safety, utility and efficiency of this system were evaluated. RESULTS: Sixty patients had generalized seizures, 30 had complex partial seizures and 10 had other types of seizures. The indication for shifting to postal review was good control of seizures in 87 cases and economic reasons in the remaining. Postal review constituted 60% of the total follow up period in 55 cases. Sixty-six patients could be maintained on postal review which was suspended or discontinued in 34 patients. Of these 34, 16 were returned to it after being seen in the clinic on a further occasion. Poor control of seizures, fresh medical or social problems, lack of confidence or a combination of these were the reasons for discontinuing the postal review. The economic benefit to a patient by way of savings in travel, incidental expenses and lost wages was estimated to be Rs 750 per annum. The work load in the epilepsy clinic was decreased by 40%. No serious medical problems or mortality were reported in the study population. CONCLUSION: Systematic postal review is a cost-effective alternative to clinic review in the long term follow up of a certain group of patients with epilepsy.