Genetic diversity of human respiratory syncytial virus in children with acute respiratory infections in Chennai, South India

Introduction: Human respiratory syncytial virus (HRSV) an RNA virus belonging to Pneumoviridae family, is an important cause of acute respiratory infections (ARIs) in young children. HRSV circulates as two subgroups A and B, which are further categorised into several genotypes. New genotypes may replace existing ones over successive epidemic seasons and multiple genotypes may cocirculate in the same community rendering it important to monitor them at the molecular level. The present study assessed the circulating genotypes of HRSV in Chennai. Materials and Methods: Two hundred and sixty-seven children with ARI were recruited during the study from April 2016 to March 2018 for detecting HRSV A and B by real-time reverse transcription-polymerase chain reaction. Phylogeny and selection pressure analysis were done. Results: Fifty-seven of the 267 samples (21.3%) were positive for HRSV, of which 7.1% and 14.2% were HRSV A and B, respectively, indicating that HRSV B was the major subgroup circulating in Chennai. Peak activity of HRSV was observed during the monsoon and winter months. Phylogenetic analysis of 2nd hypervariable region (HVR) of attachment glycoprotein gene (G gene) revealed that the HRSV A strains belonged to ON1 and HRSV B strains belonged to BA9 genotypes. Several unique amino acid substitutions were observed among the study strains. The Shannon entropy plot revealed that the HRSV A strains from our study have a high potential for amino acid substitutions in the 2nd HVR of G gene. Conclusion: This study underlines the genetic diversity of HRSV and emphasises the need for continued molecular surveillance for infection management and prevention strategies.

Comparison of nested polymerase chain reaction and real-time polymerase chain reaction targeting 47kda gene for the diagnosis of scrub typhus

Introduction: Scrub typhus is a zoonotic infection caused by Orientia tsutsugamushi which is transmitted by Leptotrombidium mites. The disease manifests as a mild-to-severe illness with non-specific clinical symptoms. Rapid diagnosis and prompt treatment are essential for patient management. Both serological and molecular methods are used for the diagnosis of scrub typhus. The present study assessed the usefulness of detection of the gene encoding the 47kDa outer-membrane protein (OMP) for the laboratory diagnosis of scrub typhus. Materials and Methods: Nested polymerase chain reaction (nPCR) and real-time PCR targeting 47 kDa OMP antigen gene of O. tsutsugamushi were performed on ethylenediaminetetraacetic acid blood samples. Results: Six of the 103 (5.8%) patients showed the presence of 47kDa gene by nPCR. Seventy of 103 (67.9%) cases showed the presence of 47kDa gene by qPCR. Among the 70 positive cases, the majority of them were females (40/70, 57.1%). The highest number of positive cases was observed during October朏ebruary. Conclusion: Real-time PCR targeting O. tsutsugamushi-specific 47-kDa gene is more sensitive than nPCR and may be the assay of choice for the detection of the organism in patients with suspected scrub typhus.

Understanding the viridians group streptococci: Are we there yet.

The viridans group streptococci are a heterogeneous group of organisms which exist as commensals in the oropharynx and the gut. They cause serious infections when they gain entry into sterile sites particularly in patients with predisposing conditions. Classification and species differentiation of these organisms has always been a challenge because of phenotypic differences between strains of the same species. Facklam’s typing scheme based on six metabolic properties has been the most widely used and many commercial identification systems are based on it. Due to the ambiguity in species differentiation based on phenotypic tests, nucleic acid‑based methods have been developed to improve the identification of these organisms. Results using genotypic methods such as 16S rRNA and sodA gene sequencing have been promising. Multilocus sequence analysis of seven house‑keeping genes map, pfl, pyk, ppaC, rpoB, soda and tuf amplified by polymerase chain reaction was found to be an accurate alternative to other methods and could be useful in the characterisation of larger collections of isolates.

Comparison of double disc and three dimensional methods to screen for ESBL producers in a tertiary care hospital.

Extended spectrum beta lactamases (ESBLs) continue to be a major problem in clinical setups world over, conferring resistance to the expanded spectrum cephalosporins. An attempt was made to study ESBL production among Enterobacteriaceae members from a tertiary care center in Chennai. A total of seventy randomly collected isolates of the family Enterobacteriaceae from a tertiary care center was studied for their susceptibility patterns to various antibiotics and detection of ESBL producers by double disc synergy (DDS) test and three dimensional test (TDT). Eighty percent of the isolates were multidrug resistant (MDR) and 20% were ESBL producers. TDT detected 85.7% whereas only 14.2% were detected by DDS. In the present study, a large number of isolates were found to be MDR and ESBL producers. TDTs were found to be better than DDS in the detection of ESBLs. Continued monitoring of drug resistance is necessary in clinical settings for proper disease management.

Effect of Azadirachta indica (neem) on the growth pattern of dermatophytes.

PURPOSE: To determine the minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) for the extracts of the leaves and seeds of the plant Azadirachta indica against various dermatophytes. METHODS: Clinical isolates of dermatophytes(Trichophyton rubrum, Trichophyton mentagrophytes and Microsporum nanum) were treated with extracts of leaves and seeds of the plant Azadirachta indica (neem) for antifungal activity by in vitro tube dilution technique. RESULTS: The MIC of neem seed extracts was 31 microg/mL for all the dermatophytes tested. The neem seed extract at 15 microg/mL concentration (below MIC) was observed to be sufficient for distorting the growth pattern of the organisms tested. CONCLUSIONS: The changes in growth curve of the treated dermatophytes were found to be statistically significant with reference to the untreated fungi.

EMM types of streptococcus pyogenes in Chennai.

The M protein of group A Streptococcus (GAS) is the major virulence factor and is coded by the emm gene. The current serologic M typing methods are now being replaced by alternate means of M type deduction such as emm gene sequencing. This is the first report of emm types of GAS which are prevalent in south India. We found no marked preponderance of any single emm sequence among our clinical isolates with 11 emm sequences being present in 34 isolates.

Enhanced detection of herpes simplex virus from ocular specimens of herpetic keratitis patients.

Corneal scrapings collected from 70 patients were used to assess the diagnostic value of indirect immunofluorescence (indirect IF) procedure in comparison with routine virus culture (RVC) for the diagnosis of Herpes simplex virus induced keratitis (HSK). Virus specific antigen was detected by indirect IF in 22 (31.42%) cases. In contrast, only 20% (14) of the cases had positive viral isolation which sometimes took as long as a week to show a cytopathogenic effect (CPE). It is concluded that antigen detection by indirect IF is a rapid, specific and sensitive technique for demonstrating HSV-1 antigen in corneal scrapings from HSK patients and a useful laboratory tool not only for diagnosing HSK but also for monitoring efficiency of anti HSV treatment for HSK.

Systemic and gastrointestinal candidiasis of infant mice as model for antifungal therapy.

Systemic and gastrointestinal infection was established in infant (15-19 days old) mice after oral-intragastric challenge with Candida albicans. All survivors retained high levels of organisms in the liver, kidney, spleen, stomach and intestine up to the 24th post infection day. These animals with persistent infections were used to study the efficacy of short term antifungal therapy. Drug treatment was initiated on 13th day for a two week period, treatment with fluconazole was compared with amphotericin B, and 5 fluorocytosine. The results suggest that fluconazole is a useful drug in the treatment of gastrointestinal candidiasis.

Kinetics of peripheral blood leucocyte counts in patients following thermal injuries.

Total and differential white cell counts (WCC) were done in 50 patients with burns involving 10% to 40% of the total body surface area (TBSA) and in 32 age and sex matched controls. Polymorphonuclear cell counts were low from 1st to 30th post burn (PB) days, lymphocyte counts were normal throughout the PB period whereas eosinophil counts were high from 1st to 60th PB days. Total leucocyte counts were significantly lower than controls from 8th to 60th PB days.