Trichosporon asahii urinary tract infection in immunocompetent patients.

Trichosporonosis is an emerging infection predominantly caused by Trichosporon asahii which is a ubiquitous and exclusively anamorphic yeast. T. asahii urinary tract infection is rare and remains scantily reported. T. asahii was isolated from urine of two immunocompetent patients who were receiving in-patient treatment for multiple comorbidities. T. asahii was identified phenotypically by a combination of manual and automated systems. Antifungal susceptibility done by E-test revealed multiresistance with preserved susceptibility to voriconazole. The ubiquity and biofilm formationposes difficulty in establishing pathogenicity and delineating environmental or nosocomial infections. Risk factors such as prolonged multiple antimicrobials, indwelling catheter and comorbidities such as anemia and hypoalbuminemia may be contributory to the establishment of a nosocomial opportunistic T. asahii infection. Dedicated efforts targeted at infection control are needed to optimize management and control of Trichosporon infections.

Complete genome sequencing and evolutionary phylogeography analysis of Indian isolates of Dengue virus type 1.

Dengue is now hyper-endemic in most parts of south and southeast Asia including India. The northern India particularly national capital New Delhi witnessed major Dengue outbreaks with Dengue virus type 1 (DENV-1) as the dominant serotype since last five years. This study was initiated to decipher the complete genome information of recently circulating DENV-1 (2009-2011) along with the prototype Indian DENV-1, isolated in 1956. Further extensive ML phylogenetic and Bayesian phylogeography analysis was carried out to investigate the evolution of this virus and understand its spatiotemporal diffusion across the globe. The complete genome analysis revealed deletion of a unique 21-nucleotide stretch in the 3' un-translated region of recent Indian DENV-1. The north Indian DENV-1 revealed up to 5.2% nucleotide sequence difference compared to recent isolates from southern India. Selection pressure analysis revealed positive selection in few amino acid sites of both structural and non-structural proteins. The molecular phylogeny classified the Indian DENV-1 into genotype III, which is also known as cosmopolitan genotype. The northern and southern Indian DENV-1 were grouped into distinct clades. The molecular clock analysis estimated a mean evolutionary rate of 7.08×10(-4) substitutions/site/year for cosmopolitan genotype. The phylogeography analysis revealed that the cosmopolitan genotype DENV-1 originated ∼1938 in India and subsequently spread globally. The diffusion of virus from India to Caribbean and South America was confirmed through SPREAD analysis. This study also confirmed the temporal displacement of different clades of DENV-1 in India over last five decades.

Emerging organisms in a tertiary healthcare set up.

BACKGROUND: One-tenth of all infectious diseases are attributable to emerging organisms. As emerging organisms sporadically affect a relatively small percentage of population they are not studied at large. This study was aimed at studying the characteristics of emerging organisms encountered from various clinical samples in an apex tertiary care multispeciality teaching and research hospital. METHODS: 16,918 positive isolates obtained from 66,323 culture samples processed in the clinical microbiology lab of an apex multispeciality hospital during 2011-2012 were included after a pilot study. Both manual and automated systems were used for identification and antimicrobial susceptibility. The frequency of isolation, sources, referring centers, resistance and susceptibility profiles, phenotypic characteristics and number of reports in PubMed were studied. RESULTS: Out of 16,918 isolates, 13,498 (79.78%) were Gram negative bacteria, 3254 (19.23%) were Gram positive bacteria and 166 (0.98%) were yeasts. A total of 483 (2.85%, 95% CI 2.6%-3.1%) emerging organisms including 116 (0.69%, 95% CI 0.57%-0.81%) emerging species were identified comprising 54 genera. CONCLUSION: Emerging organisms are likely to evade routine identification or be disregarded as non-contributory. Astute efforts directed at identification of emerging isolates, decisions by clinical microbiologists and treating physicians and containment of infection are required.

Pleuropulmonary paragonimiasis: mimicker of tuberculosis.

Infection caused by the lung fluke is endemic in north eastern parts of India. Paragonimus westermani and Paragonimus heterotremus are known to be endemic in eastern Indian states of Manipur and Nagaland. The infection is related to eating habits of the locals and is acquired by ingestion of raw, inadequately cooked crabs or crayfish containing encysted metacercariae which act as second intermediate hosts during the life cycle of the lung fluke. Diagnosis is generally delayed due to lack of suspicion and presentation similar to tuberculosis which is endemic in the population. We report pleuropulmonary paragonimiasis in a soldier from eastern India who presented with chest pain, haemoptysis, and eosinophilia. He gave history of consumption of raw crabs while on leave at his native village in Nagaland. Ova morphologically resembling Paragonimus heterotremus were detected in sputum and bronchoalveolar lavage specimen. Symptoms resolved with praziquantel treatment.

Reverse transcription loop-mediated isothermal amplification (RT-LAMP) for diagnosis of dengue.

BACKGROUND: Dengue is an emerging public health problem causing serious morbidity and mortality in tropical developing countries. Early, sensitive and specific diagnosis is paramount for clinical decision making. Currently available diagnostic tests are limited in scope and utility. This study highlights applicability of RT-LAMP in dengue diagnosis. METHODS: 100 dengue confirmed cases, 100 dengue negative cases and 79 healthy negative controls from dengue epidemic between Sep 2009 to Jul 2011 were included. Dengue cases were profiled using WHO guidelines 2006, haematological and biochemical parameters evaluated and diagnosed using NS1 antigen, IgM and IgG enzyme immunoassay, RT-PCR and RT-LAMP. Positive cases were serotyped, genotyped and various tests were compared. RESULTS: Mean haematocrit, PT, PTT, platelet count, activated lymphocytes, serum fibrinogen, transaminases, bilirubin, lactate dehydrogenase, protein and sodium were significantly elevated in DHF/DSS as compared to DF. NS1 antigen, RT-PCR and RT-LAMP were sensitive during 1-3 days while µ-capture IgM EIA was specific after 5-7 days of initial infection. DEN-1 genotype III was predominant. CONCLUSION: Deranged haematocrit and liver function tests are indicators of the severity of the disease. RT-LAMP is rapid, cost effective, highly sensitive and specific qualitative and quantitative technique which can detect dengue infection in both early and intermediary stages when NS1 antigen titres are not in the detectable range and the IgM antibody titres have just started to rise. Its superiority over existing techniques, amenability for automation and promising utility in low resource healthcare setups and field conditions raise it as the new gold standard for dengue diagnosis.

Production and characterization of monoclonal antibody specific to recombinant dengue multi-epitope protein.

Monoclonal antibodies against novel dengue recombinant protein were produced following immunization of Balb/c mice with recombinant dengue multi-epitope protein (r-DMEP) expressed in Escherichia coli vector and purified in a single-step chromatography system. Antigenicity of r-DMEP was evaluated by dot enzyme immunoassay. Mice were immunized intraperitoneally with five doses each of 100 microg of this novel antigen at 1-week intervals and a final intravenous booster dose prior to the fusion. Hybridomas resulted from fusion of myeloma cells and splenocytes using PEG-1500 as an additive. Selection of the hybrids was done using HAT medium, and the hybrids thus selected were finally screened qualitatively and quantitatively by dot and plate immunoassays, respectively. Five antibody secretory hybrid clones exhibited specific reactivity against r-DMEP by dot-ELISA, whereas a lone clone was found to be cross-reactive with Japanese encephalitis virus (JEV). Monoclonal antibodies (MAbs) specific to r-DME protein recognized the envelope and non-structural epitopes by Western blot analysis. These MAbs were further checked for their diagnostic efficacy using dengue suspected clinical samples and found overall sensitivity and specificity for DRDE dipstick ELISA. MAb-based dipstick ELISA results were 85%, 75% and 85%, 90%, respectively.