Extensive use of indwelling devices in modern medicine has revoked higher incidence of device associated infections and most of these devices provide an ideal surface for microbial attachment to form strong biofilms. These obnoxious biofilms are responsible for persistent infections, longer hospitalization and high mortality rate. Gene regulations in bacteria play a significant role in survival, colonization and pathogenesis. Operons being a part of gene regulatory network favour cell colonization and biofilm formation in various pathogens. This review explains the functional role of various operons in biofilm expression and regulation observed in device-associated pathogens such as Staphylococcus aureus, Staphylococcus epidermidis and Pseudomonas aeruginosa.

Purpose : The objective of this study was to develop a sensitive, specific and rapid approach to diagnose hand foot and mouth disease (HFMD) for an early treatment by using loop-mediated isothermal amplification (LAMP) technique. Materials and Methods : A reverse-transcription loop-mediated isothermal amplification (RT-LAMP) for detecting EV71 virus was developed, the specificity and sensitivity of RT-LAMP was tested, and the clinical specimens was assayed by the RT-LAMP comparing with conventional reverse-transcription polymerase chain reaction (RT-PCR) and real-time PCR. Results : A total of 116 clinical specimens from the suspected HFMD individual were detected with the RT-LAMP. The detection rate for EV71 was 56.89% by RT-LAMP, 41.38% by real-time PCR and 34.48% by RT-PCR. The minimum detection limit of RT-LAMP was 0.01 PFU, both of RT-PCR and real-time PCR was 0.1PFU. Non-cross-reactive amplification with other enteroviruses was detected in the survey reports. Conclusions : The effectiveness of RT-LAMP is higher than RT-PCR and real-time PCR. The protocol is easy to operate and time saving. It was not an expensive instrument, which was needed; it is an applicable method for rapid diagnosis of the disease, especially in resource-poor countries or in developing countries.

Purpose: Rapid and specific detection of viral nucleic acid is increasingly important in the diagnosis of infectious diseases. The objective was to develop a rapid, efficient process of nucleic acid based detection of hepatitis C virus (HCV) infection for its diagnosis and treatment follow-up. Materials and Methods: A two-step nested reverse-transcriptase polymerase chain reaction (RT-PCR) has been standardised on a sample set of 125 individuals from different liver clinics in Kolkata. The method utilises a novel fast nested RT-PCR for HCV detection and genotyping from HCV infected patient plasma with high processivity. Results: The overall time required from ribonucleic acid (RNA) isolation to nested PCR amplified product detection is reduced to 42% when compared with conventional nested RT-PCR amplification. The method is sensitive as conventional PCR and detected all HCV RNA positive samples. Sequencing, phylogenetic analysis of the PCR amplified product by this method showed concordant genotypes with conventional PCR. Conclusion: Though being a two-step process, this method is fast, cost-efficient, reliable and feasible for regular HCV RNA screening and apt even in resource limited settings. This method could be translated to regular nucleic acid screening for other infectious diseases as regular PCR regimen.

Purpose: The management of burn patients is always challenging for the clinician due to high risk of bacterial sepsis, multi-organ failure and death. Our objective was to study complement activation, C3 and interleukin-6 (IL-6) levels in burn patients and evaluate their role as prognostic markers. Materials and Methods: A total of 63 burn patients and 60 healthy controls were included in this study. Blood was collected from patients within 24 h and at 7 ^th day of injury. Complement activation was determined by crossed electrophoresis and counter-current immunoelectrophoresis. C3 levels were measured using a single radial immunodiffusion. IL-6 was detected by ELISA. Results: All patients showed initial complement activation. Mean C3 levels showed an inverse correlation with the severity of burn. Patients with ≥20% burns had lower C3 than the controls (P < 0.001) and those with <20% burns (P < 0.001). Patients with ≥40% burns had activated complement and low C3 in 2 ^nd week; they subsequently developed infection. Complement was inactive and C3 levels recovered in patients with <40% burns. The non-survivors showed significantly lower C3 than the survivors (P < 0.05) in 2 ^nd samples. Patients who developed infection had C3 significantly lower than those who remained free of infection (P < 0.05). All patients showed initial elevation in IL-6 levels. Patients with ≥60% burns had significantly higher IL-6 than controls (P < 0.001) and those with <60% burns (P < 0.001). Non-survivors had higher IL-6 than survivors in both samples (P < 0.001). Patients who developed infection showed significantly higher IL-6 in 2 ^nd samples than those without infection (P < 0.001). Conclusions: Complement activation, C3 and IL-6 levels correlated well with the severity of injury and development of infection in burn patients. These parameters can be used to predict the onset of infection, septicaemia and mortality in burn patients.

Background: The genus Acinetobacter is a diverse group of Gram-negative bacteria involve at least 33 species using the molecular methods. Although the genus Acinetobacter comprises a number of definite bacterial species, some of these species are of clinical importance. Therefore, it is of vital importance to use a method which is able to reliably and efficiently differentiate the numerous Acinetobacter species. Objectives: This study aims to identify Acinetobacter of clinical isolates from Assir region to the species level by 16S-23S intergenic spacers internal transcribed spacer (ITS) of ribosomal ribonucleic acid (rRNA). Materials and Methods: Deoxyribonucleic acid extraction, polymerase chain reaction amplification of 16S-23S intergenic spacer sequences (ITS) was performed using the bacterium-specific universal primers. Results: Based on the 16S-23S intergenic spacers (ITS) of rRNA sequences, all isolates tested were identified as Acinetobacter baumannii. The isolates shared a common ancestral lineage with the prototypes A. baumannii U60279 and U60280 with 99% sequence similarities. Conclusion: These findings confirmed 16S-23S rRNA ITS for the identification of A. baumannii of different genotypes among patients.

Purpose: Enteric fever is endemic in India with Salmonella Typhi being the major causative agent. Antibiotic therapy constitutes the mainstay of management. The present study was undertaken to find the susceptibility profile of Salmonella enterica var Typhi (S. Typhi) blood isolates in a tertiary care hospital between January 2001 and December 2012. Materials and Methods: A retrospective analysis of laboratory records was carried out. Conventional blood culture method was used until 2009; from January 2010 onwards BACTEC 9240 system has been in use. Salmonella were confirmed by serotyping using group and type specific antisera. Antibiotic susceptibility was performed using the disk diffusion method. In addition 116 isolates were subjected to minimum inhibitory concentration testing for chloramphenicol, ciprofloxacin, amoxicillin and nalidixic acid (NA) using agar dilution and for ceftriaxone and azithromycin using E-strips (Biomerieux). Result: A total of 1016 typhoidal salmonellae were obtained. The predominant serotype obtained was S. Typhi (852, 83.8%) followed by Salmonella enterica var Paratyphi A (164, 16.2%). We observed a re-emergence of susceptibility to first line antibiotics and a notable decline in multidrug resistant (MDR) strains. We also found all recent isolates resistant to NA and susceptible to third generation cephalosporins and 84.5% of isolates having decreasing ciprofloxacin susceptibility using revised criteria as per Clinical and Laboratory Standards Institute 2012 guidelines. Conclusion: There has been re-emergence of susceptibility to first line antibiotics and a notable decline in MDR strains of S. Typhi. We have a very high resistance to NA and decreasing susceptibility to ciprofloxacin. Third generation cephalosporins and azithromycin seem to be effective therapeutic options. Judicious use of these antibiotics is mandatory to prevent emergence of resistant strains.

Background: Sputum smear microscopy is the main-stay in the diagnosis of pulmonary tuberculosis in many developing countries. To overcome the drop outs, same day diagnosis is ideal. Materials and Methods: In the current study, two spot sputum samples (SS ₂ approach) are collected within a gap of one hour (same day sputum smear microscopy) in addition to the standard spot morning (SM) approach. The smears were stained with standard Ziehl Neelsen (ZN) and modified ZN staining techniques. Results: Out of 1537 patients, sputum smear positivity (SSP) was 9.43% (146 patients) in SM approach with standard ZN staining. Smear positivity was increased to 9.8% (151 patients) with modified ZN staining. For SS ₂ approach, SSP was 9.37% (144 patients) and 9.8% (151 patients) with standard and modified ZN staining procedures, respectively. Conclusions: Diagnosis of lung tuberculosis is possible with two spot sputum samples with modified ZN staining.

Acute cholangitis is inflammation of biliary ductal system from infection with an associated biliary obstruction. This retrospective study was done to determine the factors responsible for cholangitis and the microbiological profile of the bile in patients with cholangitis. In the study involving 348 patients, 36.4% had associated malignancy. A total of 54% of the bile samples were positive for aerobic culture. Nearly 66-73% of the Escherichia coli and Klebsiella isolates were Extended spectrum beta lactamases (ESBL) producers. Two isolates of Candida spps were also obtained. Polymicrobial infection was seen in 31.5% of the culture positive cases. Ideal antibiotics in case of cholangitis would be those which are excreted in the bile such as third-generation cephalosporins, ureidopenicillins, carbapenems and fluoroquinolones to combat resistance and polymicrobial aetiology. Anti-fungal drugs may also be necessary if the patient is not responding to biliary decompression and antibacterial agents to prevent fungaemia.

The rise of multidrug resistant strains of Salmonella Typhi in the last decade of the previous century led to the use of fluoroquinolones as the drug of choice. However, over the past few years fluoroquinolone resistance has been increasingly reported. In accordance with the revised Clinical and Laboratory Standards Institute (CLSI) breakpoints, only 3% of the isolates were susceptible to ciprofloxacin in comparison to 95% as per the earlier guidelines when 488 isolates collected between 2010 and 2012 were re-interpreted. Interestingly, re-emergence of strains susceptible to chloramphenicol, ampicillin and cotrimoxazole is being seen. Amidst the changing susceptibility profile, azithromycin remains a promising alternative.

The conventional method of transfection of suspension cells by chemical has proven to be very difficult. We present a new transfection protocol, wherein, low-speed centrifugation of cell culture plates immediately after adding the lipid: DNA complex significantly enhances the transfection efficiency. Peripheral blood mononuclear cells (PBMCs) were transfected with BLOCK-iT™ Fluorescent Oligo (scrambled siRNA) and lipofectamine complex using conventional and low-speed centrifugation modified transfection protocols. The efficiency of transfection was determined using flowcytometer and cell viability was checked using MTT assay. Incorporation of low-speed centrifugation significantly enhances the transfection efficiency of BLOCK-iT™ in the suspension culture of PBMCs as compared to conventional transfection method (99.8% vs 28.3%; P < 0.0001), even at a low concentration of 40 picomoles without affecting the cell viability. Centrifugation enhanced transfection (CET) technique is simple, time-saving and novel application without compromising the cell viability in the context of recently popular RNA interference in suspension cultures of PBMCs. This undemanding modification might be applicable to a wide variety of cell lines and solve crucial problem of researchers working with RNA interference in suspension cultures.

A total of 37 street vended food samples were examined for bacterial and the colony forming units counts ranged from 4.5 × 10 ⁵ to 1.12 × 10 ⁶ . The isolates were identified as Escherichia coli (37.5%), Pseudomonas aeruginosa (3.57%), Staphylococcus aureus (14.20%), Salmonella sp. (5.36%), Klebsiella sp. (10.71%), Shigella sp. (19.64%) and Enterobacter sp. (8.93%) respectively. All the 56 isolates were susceptible to ciprofloxacin while their susceptibility to the other drugs varied. These findings demonstrated that the ready-to-eat foods vended in Silchar city constitute an important potential hazard to human health and provision of health education to the vendors would improve quality of street foods.

We investigated the faecal carriage of extended spectrum β-lactamases (ESBL) producing Escherichia coli in different groups of human subjects and in the environment. A total of 363 E. coli strains were isolated from stool samples of patients (n = 77), healthy subjects (n = 170) and from different environmental samples (n = 116). A total of 124 ESBL producing E. coli strains were isolated in this study. The frequency of ESBL producing E. coli was found to be highest (60.3%) among the strains isolated from patients, followed by healthy individuals (38%) and the environment (10.5%). The environment was observed to have a very low number of ESBL producing E. coli.

The study was carried out to detect the adhesive genes pap (pyelonephritis associated pili), sfa (S fimbrial adhesin) and afa (afimbrial adhesin) from Escherichia coli strains isolated in patients diagnosed with urinary tract infection (UTI). A total of 23% of the isolates were positive for pap, sfa and afa genes with a prevalence of 60.87% (14/23), 39.1% (9/23) and 39.1% (9/23), respectively. Prevalence of multiple adhesive genes was 8.7% (2/23) for pap and afa, 30.43% (7/23) for pap and sfa. Significant numbers of isolates were positive for Congo red binding (80%) and haemolysin production 60%. The prevalence of multiple adhesive genes indicate the potential to adhere and subsequently cause a systemic infection among UTI patients.

A bacteriologically proven case of brain abscess, due to Streptococcus oralis is being reported in a 12-year-old girl who is a known case of congenital heart disease. The patient presented with fever, headache and vomiting. Pus cultures yielded S. oralis.

Hemophagocytic lymphohistiocytosis (HLH) triggered by Parvovirus B19 and Epstein-Barr virus co-infection is rare and unknown in infants. A 2-month-old male infant with fever, rash, bicytopenia and hepato-splenomegaly died owing to diagnostic dilemmas. Hence simply testing for hyperferritinaemia and hypertriglyceridemia/hypofibrinogenemia could diagnose HLH early while robust treatment be life-saving.

In this paper we describe a clinical scenario of high grade fever, jaundice, hypotension and multi-organ dysfunction syndrome in a 30-year-old homeless male without radiological biliary obstruction. He was brought to our hospital by the emergency medical service of Saudi Arabia (Red Crescent) from the street. After an initial resuscitation he proved to have Borrelia recurrentis. The clinical course during his hospital stay was not a smooth one, but the patient finally improved. After 20 days of hospitalisation, he was discharged in a stable condition. The present case underlines the need for high clinical vigilance, even in a non-endemic area.

Cysticercosis is a disease caused by larval form of tapeworm. Cysticercus cellulose primarily develops in tissues of pigs. Infection of human tissues is unusual and affliction of the oral cavity is rare. We, herein, present a case of lingual cysticercosis without involvement of any other site. A 5-year-old male, non-vegetarian child presented with a painless, pearly white, solitary nodular swelling on posterior dorsal side of tongue since 3 months. Excision biopsy was done. Histopathology revealed cysticercous cellulose in tongue muscles.

Cerebral phaeohyphomycosis is a rare and frequently fatal disease. We report a case of cerebral phaeohyphomycosis caused by Exophiala dermatitidis in a young immuno competent male presenting to a tertiary care hospital in Jaipur.

A multidrug-resistant clinical isolate of Ralstonia pickettii from a woman was analysed. Modified Hodge test was positive for carbapenemase production. Conjugation experiment revealed the presence of conjugative plasmid of >140 Kb size typed as IncN type. This is the first report of emergence bla_VIM-2 in R. pickettii in India.

Primary amoebic meningoencephalitis is a rare fatal meningitis caused by free living amoeba Naegleria fowleri, found in freshwater ponds and lakes. It infects children and young adults with exposure due to swimming or diving. We report a case of N. fowleri meningitis in a 6-year-old boy who presented with signs and symptoms of acute bacterial meningitis. No history of travelling or swimming was present. However, the boy frequently played with water stored from a "kuhl" (diversion channels of water). Wet mount of cerebrospinal fluid (CSF) revealed amoeboid and actively motile flagellate forms of trophozoites. CSF culture done on 1.5% non-nutrient agar plates with a lawn culture of Escherichia coli kept at 37°C for 15 days did not reveal any growth. The test of flagellation on passing CSF in distilled water was however positive in 3 h. Water of the "kuhl" from the stored tank also showed actively motile trophozoites similar to the forms obtained from the CSF. Based on our reports, the boy was immediately treated with amphotericin B, rifampicin and fluconazole for 21 days. Repeat CSF examination after 14 days did not reveal any trophozoites in wet mount and patient was discharged after 3 weeks of successful treatment.