Real-time multiplex polymerase chain reaction with high-resolution melting-curve analysis for the diagnosis of enteric infections associated with diarrheagenic Escherichia coli.

INTRODUCTION: Although diarrheagenic Escherichia coli (DEC) strains are important bacterial causative agents of diarrhoea, they are not routinely sought as stool pathogens in clinical laboratories as conventional microbiological testing are unable to distinguish between normal flora and pathogenic strains of E. coli. This study was undertaken to determine the prevalence of DEC pathotypes amongst children with and without diarrhoea and to detect specific virulent genes present in different DEC pathotypes, using real-time multiplex polymerase chain reaction (PCR) with high-resolution melting (HRM) technology. MATERIALS AND METHODS: Stool samples were obtained from cases and controls. Using a set of conventional biochemical tests, E. coli strains were identified. Further, these isolates were subjected to multiplex PCR system for the detection of virulence genes of different pathotypes of DEC. Real-time multiplex PCR was performed for the detection of specific virulent genes of DEC pathotypes, using Rotor-Gene Q instrument (Qiagen) having High-resolution Melt analyser using Type-it HRM PCR kit (Qiagen) containing EvaGreen fluorescent intercalating dye. RESULTS: In this study, we had successfully standardised two multiplex PCR assays which were found to be effective for direct detection of enteropathogenic E. coli (EPEC), enteroaggregative E. coli (EAEC), enterotoxigenic E. coli (ETEC) and enteroinvasive E. coli (EIEC). A total of 42 DEC strains were detected at an overall rate of 19.3% (n = 42), from the total 217 E. coli isolates recovered from the cases (n = 39, 17.9%) and control (n = 3, 3.8%) groups. Amongst the 42 DEC pathotypes (39 from cases and 3 from controls), EPEC (10%), EAEC (8.82%), ETEC (2.94%) and EIEC (1.18%) were found in children with diarrhoea (cases) and in children without diarrhoea (control) only EAEC (2.13%) and EPEC (4.26%) were detected. Age distribution, gender variation, seasonal variation and clinical features were also analysed. CONCLUSION: This study helped evaluate the prevalence of DEC amongst children (<18 years of age) with and without diarrhoea using multiplex real-time PCR with HRM analysis.

Pottels Equation for Estimation of Glomerular Filtration Rate.

The retrospective study was carried out to examine performance of Pottels height- independent equation compared to Schwartzs height-dependent equation to estimate glomerular filtration rate in 115 children in Indian setting. The Pottels equation performed well compared to updated Schwartz equation (R2=0.94, mean bias 0.25, 95% LOA=20.4, -19.9). The precision was better at lower range of estimated GFR.

Effective ultrafiltration with acute peritoneal dialysis in a child with diuretic-resistant nephrotic edema.

Edema is a cardinal feature of the nephrotic syndrome and sometimes merits independent treatment. The use of diuretics is usually sufficient in the treatment of edema. Ultrafiltration (UF) may sometimes be needed in diuretic-resistant states. The use of UF for steroid-resistant nephrotic edema is scarce in children. We report a child with steroid-resistant nephrotic syndrome with diuretic-resistant nephrotic edema treated successfully using acute peritoneal dialysis as a means of UF.

Invasive meningococcal infection: analysis of 110 cases from a tertiary care centre in North East India.

OBJECTIVES: To report an outbreak of invasive meningococcal disease from Meghalaya, in the north east India, from January 2008 through June 2009. METHODS: Retrospective review of case sheets was done. One hundred ten patients with invasive meningococcal disease were included for the study. RESULTS: Of the total patients, 61.8 % were boys and 38.2 % were girls (boy to girl ratio = 1.62:1). The average age of presentation was 8.48 ± 5.09 y. Meningococcal meningitis was seen in 61.8 % of cases, meningococcemia in 20 % and 18.2 % had both. Fever was the most common manifestation (100 %) followed by meningeal signs (78.2 %), headache (56.4 %), vomiting (53.6 %), shock (38.2 %), low Glasgow coma scale (GCS) (25.5 %), purpura and rashes (23.6 %), seizures (9.1 %), abdominal symptoms (4.5 %), irritability and excessive crying (4.5 %) and bulging anterior fontanalle (23 %) in those below 18 mo of age. Raised intracranial pressure (ICP) was the most common complication (28.2 %) followed by coagulopathy (16.4 %), hepatopathy (10 %), herpes labialis (9.1 %), syndrome of inappropriate ADH secretion (SIADH) (8 %), pneumonia (7 %), arthritis (6 %), purpura fulminans, respiratory failure, sixth nerve palsy and diabetes insipidus in 4.5 % each, subdural empyema, optic neuritis, ARDS and ARF in 1.8 % each, cerebral salt wasting syndrome, third nerve palsy, cerebritis and hearing impairment in 0.9 % each. Culture was positive in 35.5 %. Patients were treated initially with ceftriaxone and dexamethasone but later on with chloramphenicol due to clinical drug resistance. Mortality was 6.4 %. CONCLUSIONS: This is the first epidemic report of invasive meningococcal disease from the north east India. Chloramphenicol acts well in areas with penicillin or cephalosporin resistance. Mortality reduces significantly with early diagnosis and prompt intervention.

Characteristics of pediatric scrub typhus during an outbreak in the North Eastern region of India: peculiarities in clinical presentation, laboratory findings and complications.

OBJECTIVE: To identify and report a recent outbreak of scrub typhus cases recorded from October 2009 to January 2010 in the state of Meghalaya, India. METHODS: The case sheets of all the children were retrieved and reviewed retrospectively. Twenty four (24) patients, who were both clinically and serologically confirmed as scrub typhus cases were included in the study. RESULTS: Fever for more than 1 wk duration was the most common manifestation (100%) followed by splenomegaly (45.8%), eschar (41.7%), cough (37.5%), hepatomegaly (33.3%), headache and body ache (25%), pain abdomen (25%), vomiting (20.8%), altered sensorium (16.6%), seizures (12.5%) lymphadenopathy (12.5%), and loose stools (8.3%). Meningoencephalitis was the most common complication (29.2%) followed by pneumonia (16.3%) and subconjunctival hemorrhage (8.3%).Cortical blindness, septic shock, peritonitis, myocarditis with CCF, pancytopenia, acute renal failure, coagulopathy, prolonged oxygen dependency and urinary tract infection (UTI) were found in one of each case. Hyponatremia (66.7%), elevated liver enzymes without significant rise of bilirubin (58.3%), hypoalbuminemia (52.2%) and thrombocytopenia (26%) were the other significant laboratory findings. Patients were treated with chloramphenicol, doxycycline and azithromycin. There was no mortality. CONCLUSIONS: This is the first outbreak report from the north eastern region of India with varied clinical presentations, laboratory investigations and complications. Weil Felix test still remains fruitful for diagnosing this disease in a resource limited set up.

Empyema thoracis: analysis of 150 cases from a tertiary care centre in North East India.

OBJECTIVES: To analyze the clinical characteristics, microbiological profile, management, complications and outcome of cases with empyema thoracis. METHODS: All cases diagnosed as 'empyema thoracis' over a period from January 2006 through June 2010 were identified from the inpatient records and discharge summaries. Of the 160 cases identified, 150 cases were taken up for analysis and the rest 10 cases, of which two had significant predisposing co-morbidity and eight cases diagnosed as tubercular empyema thoracis were excluded from the analysis. RESULTS: Mean age of presentation was 4.74 ± 3.53 years and two thirds of the children were under 5 years with male to female ratio of 1.42:1. Pus culture was positive in 32% (48 cases) with Streptococcus pneumoniae being the commonest organism isolated (31 cases, 64.6%) followed by Staphylococcus aureus (11 cases, 22.9%), Klebsiella pneumoniae (3 cases, 6.3%), Haemophilus influenzae type b (2 cases, 4.2%) and Enterococcus (1 case, 2%). Clustering was seen in the hot and humid months from April to July (63.3%). Fever was the commonest presentation (96.7%) followed by cough (90%), breathing difficulty (66.7%), chest pain (26.7%) and pain abdomen (10.7%). Ampicillin and cloxacillin was used as the first line antibiotic in 57.3% cases. Average duration of intercostal water seal drainage (ICWSD) in situ was 13.5 ± 8.05 days and 59 patients (39.3%) received fibrinolytic therapy. The commonest complications were collapse (18%), thickened pleura (16.7%), pericardial effusion (8%), cardiac tamponade (3.3%) and bronchopleural fistula (3.3%). Surgical procedures involved in this case series were decortication (14 cases, 9.3%), pericardiocentesis (5.3%), pericardiostomy (2.7%) and pericardiectomy (1.6%). Mortality was 3.3%. CONCLUSIONS: This is the first report of empyema thoracis from the north eastern region of India. Streptococcus pneumoniae was found to be the leading cause of empyema thoracis in this case series. Conservative management with ICWSD and antibiotics or early use of fibrinolytic therapy if indicated are effective modalities of treatment.

Macrophage activation syndrome in malaria.

Macrophage activation syndrome (MAS) is a clinical syndrome caused by an excessive proliferation of T lymphocytes and well-differentiated macrophages; an entity distinct from malignant histiocytosis. Although rheumatologic conditions are the common cause of MAS, a wide range of infections are also seen to cause MAS. We report an adolescent with severe Plasmodium falciparum malaria and MAS. He fulfilled six out of eight criteria required to diagnose hemophagocytic lymphohistiocytosis.