Identification of an immunodominant epitope in glycoproteins B and G of herpes simplex viruses (HSVs) using synthetic peptides as antigens in assay of antibodies to HSV in herpes simplex encephalitis patients.

Herpes simplex encephalitis (HSE) is a severe viral infection of the central nervous system (CNS). Assay of antibody response is widely used in diagnostics of HSE. The aim of this study was to identify an immunodominant epitope determining the antibody response to herpes simplex viruses (HSVs) in cerebrospinal fluid (CSF) of HSE patients. The synthetic peptides that resembled type-common as well as type-specific domains of glycoproteins B (gB) and G (gG) of these viruses were evaluated for binding with IgM and IgG antibodies in CSF samples from HSE and non-HSE patients in ELISA. The QLHDLRF peptide, derived from gB of HSV was found to be an immunodominant epitope in the IgM and IgG antibody response. The patients with confirmed and suspected HSE showed in ELISA against this peptide 26% and 23% positivities for IgM, 43% and 37% positivities for IgG and 17% and 15% for both IgM and IgG antibodies, respectively. The total positivities of 86% and 75% for both IgM and IgG antibodies were obtained in the patients with confirmed and suspected HSE, respectively. These results demonstrate that a synthetic peptide-based diagnostics of HSE can be an efficient and easily accessible alternative. This is the first report describing the use of synthetic peptides derived from HSVs in diagnostics of HSE using patientsʹ CSF samples.

Methylene blue used in the treatment of refractory shock resulting from drug poisoning.

BACKGROUND: Methylene blue inhibits the nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathway, decreasing vasodilation and increasing responsiveness to vasopressors. It is reported to improve haemodynamics in distributive shock from various causes including septicaemia and post-cardiac surgery. Reports of use in overdose are limited. We describe the use of methylene blue to treat a case of refractory distributive shock following a mixed drug poisoning. CASE DETAILS: A 41-year-old male presented following reported ingestion of 18 g extended-release quetiapine, 10 g controlled-release carbamazepine, 240 mg fluoxetine, 35 g enteric-coated sodium valproate and 375 mg oxazepam. He was comatose and intubated on presentation. Progressive hypotension developed. Echocardiogram revealed a hyperdynamic left ventricle, suggesting distributive shock. The patient remained hypotensive despite intravenous fluid boluses, escalating vasopressor infusions. Arterial blood gas revealed metabolic acidaemia and high lactate. Methylene blue was administered as loading-dose of 1.5 mg/kg and continuous infusion (1.5 mg/kg/h for 12 h, then 0.75 mg/kg/h for 12 h) resulting in rapid improvement in haemodynamic parameters and weaning of vasopressors. Serum quetiapine concentration was 18600 ng/mL (30-160 ng/mL), collected at the time of peak toxicity. CONCLUSION: Severe quetiapine poisoning produces hypotension primarily from alpha-adrenoreceptor antagonism. Methylene blue may have utility in the treatment of distributive shock resulting from poisoning refractory to standard vasopressor therapy.

Evaluation of BioFM liquid medium for culture of cerebrospinal fluid in tuberculous meningitis to identify Mycobacterium tuberculosis.

The present study was designed to evaluate the sensitivity and specificity of liquid culture medium (BioFM broth) for the diagnosis of tuberculous meningitis (TBM) in cerebrospinal fluid (CSF). CSF samples from 200 patients (TBM group = 150 and non-TBM group = 50) were tested for culture of Mycobacterium tuberculosis in BioFM liquid culture medium. Out of 150 TBM cases, 120 were found to be culture positive, indicating a sensitivity of 80% in BioFM broth within 2-3 weeks of inoculation. Positive cultures were also observed for CSF from 32 (64%) out of 50 non-TBM patients in BioFM liquid culture medium within 4 days of sample inoculation. Therefore, according to our study, BioFM broth system yielded 80% sensitivity [95% confidence interval (CI): 67-93%] and 36% specificity (95% CI: 57-98%) for TBM diagnosis. Our results indicate that although BioFM broth allows the detection of positive cultures within a shorter time, it has a high potential for contamination or for the coexistence of M. tuberculosis and non-tuberculous meningitis (NTM). This coexistence may go undetected or potentially lead to erroneous reporting of results.

Determination of polymerase chain reaction efficiency for diagnosis of tuberculous meningitis in Chelex-100 extracted DNA samples.

SETTING: Polymerase chain reaction (PCR) offers great promise for the rapid, sensitive and specific diagnosis of tuberculous meningitis (TBM). However, the isolation of DNA of high quantity and quality from cerebrospinal fluid (CSF) samples is critical for successful PCR assays. OBJECTIVE: To develop and use a single-tube method for the isolation of PCR-compatible DNA from Mycobacterium tuberculosis using Chelex-100 chelating resin, which does not require organic solvents or detergents. DESIGN: The study focused on the standardisation of a suitable Chelex protocol and its evaluation in 32 CSF samples from TBM and non-TBM subjects. A simultaneous comparison was made with the conventional phenol/chloroform extraction method. RESULT: PCR was found to be more sensitive, more rapid and less technically demanding with the Chelex protocol than the conventional phenol/chloroform extraction method (sensitivity 84.2% vs. 73.6%). CONCLUSION: The single-tube method and the simplicity of the procedure permits early and reliable diagnosis of TBM and makes it an attractive method for routine laboratory assays.

Diagnostic value of early secreted antigenic target-6 for the diagnosis of tuberculous meningitis patients.

BACKGROUND: The early diagnosis of tuberculous meningitis (TBM) is very crucial, since delayed diagnosis can lead to various neurological manifestations. We have previously developed an in-house indirect enzyme-linked immunosorbent assay (ELISA) for TBM diagnosis using the Antigen 85 (Ag 85) complex. It has been suggested that the Ag 85 complex might give false-positive reactions for individuals vaccinated with Bacillus Calmette-Guérin (BCG). OBJECTIVES: In the present study, we describe a prospective evaluation demonstrating that early secreted antigenic target- 6 (ESAT-6), which is absent in Mycobacterium bovis BCG strains, is in the cerebrospinal fluid (CSF) of TBM patients. METHODS: We used an indirect ELISA to detect ESAT-6 antigens in the CSF of TBM patients using polyclonal antibodies against ESAT-6. RESULTS: Using the indirect ELISA method, we demonstrated a sensitivity and specificity of 80% and 94%, respectively, for the diagnosis of TBM. CONCLUSION: The detection of ESAT-6 in the CSF of TBM patients by indirect ELISA is a promising method and can be used to develop an immunodiagnostic assay with increased sensitivity and specificity.

Serodiagnosis of tuberculosis based on the analysis of the 65 kD heat shock protein of Mycobacterium tuberculosis.

OBJECTIVE: To evaluate the 65 kD heat shock protein (hsp) antigen in serum samples of tuberculosis (TB) patients by enzyme-linked immunosorbent assay (ELISA) using monoclonal antibodies (mAbs) specific to the 65 kD hsp antigen for the diagnosis of TB. DESIGN: Serum samples were obtained from 188 different groups of patients (confirmed TB [n = 24], clinically suspected TB [n = 48], non-TB disease controls [n = 74] and healthy individuals [n = 42]) and analysed by ELISA using mAbs to the 65 kD hsp antigen. The Kruskal Wallis test (non-parametric analysis of variance) with the Dunnett post test was used for statistical analysis. RESULTS: The method yielded 82% sensitivity and 89% specificity for the diagnosis of TB. The mean (+ or -SD) absorbance value of the 65 kD hsp antigen in TB patients (1.73 + or - 0.27) was significantly higher than in the non-TB disease control group (1.12 + or - 0.42, P < 0.001) and was also higher than among healthy individuals (1.06 + or - 0.42, P < 0.001). CONCLUSION: The detection of the 65 kD hsp antigen in serum samples from confirmed and suspected TB patients by ELISA using mAb against purified 65 kD antigen gives a reliable diagnosis and could be considered as a diagnostic marker for TB. The absence of the 65 kD hsp antigen in healthy control BCG-vaccinated subjects indicates the diagnostic value of this assay in regions of endemicity.

Demonstration of IgG antibodies to 30 Kd protein antigen in CSF for diagnosis of tuberculous meningitis by antibody-capturing ELISA.

OBJECTIVE: A simple and rapid immunological assay method has been developed to demonstrate the presence of IgG antibodies to 30Kd protein antigen (30Kdpa) and culture filtrate protein (CFP) in the CSF of patients with Tuberculous meningitis (TBM). METHOD: Antibody capturing Enzyme Linked Immunosorbent Assay (ELISA) was standardized with CFP antigen of MTB. The IgG antibodies were assayed in CSF sample from TBM and non-TBM patients against 30 Kdpa. RESULTS: The sensitivity and specificity of IgG antibodies for the diagnosis of suspected patients of TBM using 30 Kdpa was 80% and 91% respectively and the corresponding figures for CFP were 85% and 94% respectively. The sensitivity and specificity in two confirmed cases of TBM was 100%. CONCLUSION: The presence of this 30Kdpa in the CSF of suspected cases of TBM consistently would indicate that the selected protein band carries the candidate protein marker antigen, which is specific to M. tuberculosis and could be considered as a diagnostic marker for TBM.

Study of factors responsible for recurrence of seizures in controlled epileptics for more than 1 years after withdrawal of antiepileptic drugs.

531 epileptic patients, who had achieved remission mostly for 2 years or more were studied. The mean follow up period was 5 years. Recurrence was noted in 103 patients (19%) after gradual withdrawal of AED, over a period of 3-4 months. 424 patients (81%) did not have recurrence. The recurrence rate was influenced adversely by factors like adolescent age and later onset seizures, pre-treatment duration of symptoms more than 3 years, pre-treatment precipitating factors like emotional stress, lack of sleep and meals (however, number in each group is small), positive family history of epilepsy, focal neurodeficit, absence and myoclonic plus grandmal type of clinical seizures, paroxysmal generalized spike and wave discharges and generalized short polyspike and wave discharges in the pretreatment EEG, atrophic changes on CT brain scan (in small numbers), head trauma at birth or later and hereditary factors as etiology of epilepsy, and more than 30 number of seizures before achieving the remission. Factors like, sex, frequency of seizures, period of remission i.e. two years or more and number of drugs used to achieve remission, did not have any significant adverse effect. However, in the last parameter 95% remission was achieved by one or a combination of two drugs (72% and 23% respectively).

Electrocardiographic manifestations following electric injury.

A 23-year-old man admitted with electric injury had intermittent episodes of non-sustained ventricular tachycardia, recurrent ventricular premature complexes, atrial fibrillation, supraventricular tachycardia and ST changes suggestive of pericarditis. The non-sustained ventricular tachycardia and ventricular premature complexes responded to intravenous lidocaine, atrial fibrillation and supraventricular tachycardia to intravenous digoxin.

Clinical and electroneurophysiological assessment of leprosy patients on dapsone monotherapy--a two year follow-up study.

Fifty-three persons with tuberculoid type of leprosy having a thickened nerve on one side and a clinically normal nerve on the contralateral side were studied before, during and after two years of therapy for electrophysiological abnormalities in apparently normal and in obviously thickened nerves. Twenty-seven patients had received treatment with dapsone 100 mg orally and 26 cases had received rifampicin therapy. It was found that there was no extension of anesthesia or diminution of motor power over a period of two years. There was no significant difference between the initial and final recordings of motor and sensory nerve conductions if aggregate figures were taken. However, taking individual cases, deterioration in nerve conduction (increased latency and decreased velocity) was found in two patients, of whom one had received dapsone and the other had received rifampicin.

Hemifacial spasm: early postoperative normalization of blink reflex latency.

Electrophysiological improvement in the blink reflex latency after neurovascular decompression of the facial nerve in patients with hemifacial spasm is believed to be related to remyelination and occurs 2-8 months after surgery. We report a patient with hemifacial spasms for 3 years, in whom the increased blink reflex latency returned to normal within a week after surgery. This suggests that compression without demyelination may be responsible for increased blink reflex latency in some of the patients with hemifacial spasms.

Vibration induced acute carpal tunnel syndrome in a neurosurgeon following use of bone drill.

Exposure of the hand to vibration for many years is known to result in a carpal tunnel syndrome in forest workers. A case is reported of a neurosurgeon who developed features of carpal tunnel syndrome following continuous exposure to vibrations froma bone drill for several hours. The numbness and Paresthesia improved and the median nerve latencies returned to normal after 3 weeks.

Prognostic significance of electrodiagnostic studies in the Guillain-Barré syndrome.

Nerve conduction and needle electromyographic studies were carried out in 50 patients with the Landry-Guillain-Barré syndrome to assess the reliability of this procedure in predicting the prognosis. Two main groups were identified. The first group was characterized by gross abnormalities in nerve conduction and absence of fibrillation potentials during the entire course of the illness. Twenty-five of 31 patients (80.6%) belonging to this group recovered rapidly, and the quality of recovery was good. In 19 patients belonging to the second group, profuse fibrillations were noted within the first four weeks of the illness with or without associated nerve conduction deficits. Recovery in this group was poor (31.5%) and pronounced residual deficits were more common. Electrophysiological studies therefore are of value not only in the diagnosis but also as a reliable prognostic index in this syndrome.