Microbiological profile of lower respiratory tract infections in neurological intensive care unit of a tertiary care center from Central India.

BACKGROUND: Lower respiratory tract infections (LRTI's) are the most frequent infections among patients in intensive care units. The consequences of increased drug resistance are far reaching since bacterial infection of the lower respiratory tract (LRT) is a major cause of death from infectious disease. OBJECTIVE: The study was conducted with the aim of determining the bacterial etiology of LRTI in the neuro intensive care unit (NICU) as well as to update the clinicians with the various antimicrobial alternatives available in the treatment of LRTI. SUBJECTS AND METHODS: The study was conducted for the period of 3 years from January 2010 to December 2012 in the Microbiology Department of a Teaching Tertiary Care Hospital. The LRT specimens from 230 patients admitted in a NICU during the study period were processed. Following culture, the isolated organisms were identified and antimicrobial sensitivity was performed by standard methods. RESULTS: Out of the 230 LRT specimens evaluated, 198 (86.08%) were culture positive. A total of 254 pathogens were recovered with a predominance of Gram-negative isolates (n = 243; 96.05%) Pseudomonas aeruginosa was the most dominant pathogen followed by Klebsiella pneumoniae. Alarmingly high percentage of extended spectrum beta-lactamase and methicillin resistant Staphylococcus aureus isolates were detected. The resistance to cephalosporins, aminoglycosides and carbapenem were remarkable. CONCLUSIONS: Therefore, we can conclude that for effective management of LRTI's, an ultimate and detailed bacteriological diagnosis and susceptible testing is required to overcome global problem of antibiotic resistance.

Surface disinfection by exposure to germicidal UV light.

The present study was aimed to design a simple model to check efficacy of germicidal UV tube, to standardise the position, distance and time for UV light and also to find out its efficacy against medically important bacteria, the bacterial spores and fungi. The microbial cultures tested included gram positive and gram negative bacteria, bacterial spores and fungal spores. The microbes streaked on solid media were exposed to UV light. The inactivation of the order of four logs was observed for bacteria. UV light can have efficient inactivation of bacteria up to a distance of eight feet on either side and exposure time of 30 minutes is adequate.

In vitro susceptibility to meropenem and other antimicrobial agents among gram-negative bacilli isolated from hospitalized patients in central India.

BACKGROUND: Growing multiple drug resistance among gram-negative bacilli among hospitalized patients is a serious therapeutic problem, and the aim of the study was to assess the situation in our hospital. METHODS: Antimicrobial susceptibility testing with the disk method was carried out on 1,533 isolates of gram-negative bacilli from urine, pus, body fluid and blood from hospitalized patients. RESULTS: Seventeen percent of isolates were susceptible only to meropenem and either to piperacillin + tazobactam, to cefoperazone + sulbactam or to both. Eleven percent of isolates were susceptible only to meropenem and 6% were resistant to all antimicrobial agents including meropenem. CONCLUSION: Growing multiple drug resistance among gram-negative bacilli in hospital practice demands a rigid antibiotic policy and strict infection control measures.

Biomedical waste in laboratory medicine: audit and management.

Pathology, microbiology, blood bank and other diagnostic laboratories generate sizable amount of biomedical waste (BMW). The audit of the BMW is required for planning proper strategies. The audit in our laboratory revealed 8 kgs anatomical waste, 600 kgs microbiology waste, 220 kgs waste sharps, 15 kgs soiled waste, 111 kgs solid waste, 480 litres liquid waste along with 33,000 litres per month liquid waste generated from labware washing and laboratory cleaning and 162 litres of chemical waste per month. Section wise details are described in the text. Needle sharps are collected in puncture proof containers and the needles autoclaved before sending to needle pit. The glass forms the major sharp category and is disinfected with hypochlorite before washing/recycling. All microbiology waste along with containers/plates/tubes are autoclaved before recycling/disposal. The problem of formalin fixed anatomical waste as histology specimens is pointed out. The formalin containing tissues cannot be sent for incineration for the fear of toxic gas release and the guidelines by the Biomedical waste rule makers need to be amended for the issue. The discarded/infected blood units in blood bank need to be autoclaved before disposal since chemical treatments are difficult or inefficient. The liquid waste management needs more attention and effluent treatment facility needs to be viewed seriously for hospital in general. The segregation of waste at source is the key step and reduction, reuse and recycling should be considered in proper perspectives.

High-energy particle acceleration in the shell of a supernova remnant.

A significant fraction of the energy density of the interstellar medium is in the form of high-energy charged particles (cosmic rays). The origin of these particles remains uncertain. Although it is generally accepted that the only sources capable of supplying the energy required to accelerate the bulk of Galactic cosmic rays are supernova explosions, and even though the mechanism of particle acceleration in expanding supernova remnant (SNR) shocks is thought to be well understood theoretically, unequivocal evidence for the production of high-energy particles in supernova shells has proven remarkably hard to find. Here we report on observations of the SNR RX J1713.7 - 3946 (G347.3 - 0.5), which was discovered by ROSAT in the X-ray spectrum and later claimed as a source of high-energy gamma-rays of TeV energies (1 TeV = 10(12) eV). We present a TeV gamma-ray image of the SNR: the spatially resolved remnant has a shell morphology similar to that seen in X-rays, which demonstrates that very-high-energy particles are accelerated there. The energy spectrum indicates efficient acceleration of charged particles to energies beyond 100 TeV, consistent with current ideas of particle acceleration in young SNR shocks.

Bacterial population changes in hospital effluent treatment plant in central India.

Hospital effluent with its high content of multidrug resistant (MDR) enterobacteria and the presence of enteric pathogens could pose a grave problem for the community. It was planned at our tertiary care hospital in central India to study the population changes at various steps of effluent treatment plant (ETP) like collection, aeration, clarification, liquid sludge, dried sludge, high-pressure filter and treated wastewater. The study included viable bacterial counts, coliform counts, staphylococcal, enterococcal, Pseudomonas and multiple drug resistant (MDR) gram negative bacterial counts in the different stages of ETP. In order to study the distribution of bacteria as free floating in liquid and adherent to suspended particles, enumeration of the bacteria in the filterate and the sediment was also carried out. The effluent input showed 55% of the 8.6 x 10(6)/ml bacteria as coliforms and E. coli which was a typical of fecal flora. The prevalence of MDR coliforms was 0.26%. The substantial reduction (> 3log) was seen for the effluent coming from the clarifier. The bulk of the bacteria in the hospital effluent remains firmly adhered to solid particles; aeration and clarification removes bulk of the bacteria by physical processes like flocculation. The treated liquid effluent still contains sizeable loads of MDR bacteria and inactivation by procedure such as chlorination is required. The bacteria get concentrated in sludge and a greater concentration of chlorine is required for decontamination.

Solar disinfection of infectious biomedical waste: a new approach for developing countries.

Poor developing countries cannot afford expensive technologies such as incineration for management of infectious biomedical waste. We assessed solar heating as an alternative technology. We immersed simulated infectious waste with added challenge bacteria in water in a box-type solar cooker, which was left in the sun for 6 h. In 24 sets of observations, the amount of viable bacteria was reduced by about 7 log. We also tested infectious medical waste with a heavy load of bacteria (10(8)-10(9)/g) from our hospital's burn unit for solar heat disinfection in 20 experiments. Our results showed a similar 7 log reduction in the amount of viable bacteria. Solar heating thus seems to be a cheap method to disinfect infectious medical waste in less economically developed countries.

Current status of drug resistance among gram-negative bacilli isolated from admitted cases in a tertiary care centre.

AIMS: To study the prevalence of drug resistance among gram-negative bacilli and susceptibility to the Cefoperazone + Sulbactam and Piperacillin + Tazobactam combination among the hospital isolates resistant to all other antimicrobial agents. METHODS: A total of 367 gram-negative bacilli isolated from various pathological samples were tested for susceptibility to antimicrobial agent by the disc diffusion technique and 300 isolates of gram-negative bacilli which were resistant to all the other antibiotics were checked for sensitivity to piperacillin + tazobactam and cefoperazone + sulbactam combination. RESULTS: Resistance to ampicillin and amoxicillin + clavulanic acid ranged from (72.1-83.8%) which was highest. Resistance to cotrimoxazole, gentamicin, netilmycin, quinolones and all the generation of cephalsporins was often above 50%. piperacillin + tazobactam combination had the lowest incidence of resistance (14 to 20.5%). CONCLUSION: Drug resistance for most of the antimicrobial agents was greater than 50% including the various generations of cephalosporins. Inefficiency of amoxicillin + clavulanic acid (resistance among 72-79.5% isolates) suggests the presence of extended spectrum beta-lactamases among the isolates. Addition of sulbactam to cefoperazone showed increased activity (resistance 27 to 34%) but piperacillin + tazobactam was the most effective combination in vitro.

Treatment of discarded blood units: disinfection with hypochlorite / formalin versus steam sterilization.

Blood bank regulations and bio medical waste rules of India advocate disinfection of contaminated blood units. Incineration is not recommended due to poly-vinyl chloride (PVC) content of blood bags. This study was designed to evaluate the efficacy of chemical disinfection of blood units deliberately contaminated with Staphylococcus aureus and E. coli with 1 and 6 % hypochlorite, 10% formalin and 33% formaldehyde and autoclaving of blood units contaminated with the above mentioned vegetative forms and B. stearothermophilus spores. Only 33 % formaldehyde could bring about 5 log reduction of bacteria but it is highly irritating and toxic. Autoclaving at 15 lbs pressure for 2 hours uniformly inactivated the vegetative forms and B. stearothermophilus spores. Thus, autoclaving of PVC blood bags is a safer and reliable method compared to chemical disinfection.

Hypochlorite (1%) is inefficient in decontaminating blood containing hypodermic needles.

Infectious biomedical waste and sharps have a potential hazard of transmission of pathogens. Among sharps, used needles form a major share and disinfection by 1 % hypochlorite is recommended in biomedical waste management rules of India. The aim of the present study was to evaluate the efficacy of hypochlorite for the decontamination of needles. Needles (16 g) filled with suspensions of standard strains and clinical isolates of gram positive and gram negative bacteria in plain normal saline and in human blood containing anticoagulant, were exposed to 1% hypochlorite and the surviving bacteria were subjected to viable counts. The observations indicated that 85 - 90 % of the needles filled with bacterial suspensions in saline are disinfected to a level of > 5 log bacterial reduction (standard disinfection) on exposure to hypochlorite but only 15 to 30% needles contaminated with the challenge bacteria suspended in blood showed > 5 log reduction in viable counts. Thus, hypochlorite treatment is inadequate for disinfecting needles contaminated with pathogenic bacteria in presence of blood and should not be recommended as an option for disinfection of the needles.

Patterns of CD8 T cell clonal dominance in response to change in antiretroviral therapy in HIV-infected children.

OBJECTIVE: To examine the influence of change in antiretroviral therapy (ART) on patterns of CD8 T cell clonal dominance in HIV-infected children. DESIGN: Seventeen HIV-infected children with plasma virus loads between 3.1 and 5.7 log10 were investigated before and after changes in ART. METHODS: CDR3 spectratyping was performed in 22 T cell receptor (TCR) Vbeta subfamilies by multiplex polymerase chain reaction (PCR) in purified peripheral blood CD8 T cells in conjunction with CD4 cell counts, plasma HIV-RNA copies and lymphoproliferative assays (LPA). RESULTS: CD8 T cell clonal dominance in two or more Vbeta families was present in eight out of 17 children. After a change in therapy, 13 patients (76%) acquired new clones whereas three patients (17.6%) showed a loss in CD8 cell clones. An increase in the numbers of dominant clones correlated with an increase in percentage CD4 cell counts (P < 0.001) and with improved LPA responses to tetanus (P < 0.05) and alloantigens (P < 0.01). CD4 cell increase was associated with an initial mean gain of 3.1+/-2.1 CD8 cell clones, independent of a virological response. A loss of CD8 cell clones or failure to achieve CD4 T cell increase was associated with failure to achieve virological suppression. CONCLUSION: Children with chronic HIV infection manifest CD8 T cell clonal dominance, which appears to be dependent upon the adequacy of the CD4 cells. With optimization of therapy, a gain in clonal dominance is the predominant response, except in situations of failure to contain viral replication.

Growing problem of methicillin resistant staphylococci--Indian scenario.

In the present study MRSA prevalence increased from 12% in 1992 to 80.83% in 1999. Indian literature shows that MRSA incidence was as low as 6.9% in 1988 and reached to 24% and 32.6% in Vellore and Lucknow in 1994 and was of the same order in Mumbai, Delhi and Bangalore in 1996 and in Rohtak and Mangalore in 1999. However, in some of the centres it was as high as 87%. All the MRSA isolates in India including in the present study were sensitive to vancomycin and resistance to netilmycin appears to be low among MRSA isolates in India. All the MRSA isolates were also found to be sensitive to teicoplanin in the present study. Like in other Indian studies, resistance to cotrimoxazole, erythromycin, gentamicin, other penicillins and cephalosporins appeared to be a common feature for MRSA isolates in the present study.

Clonal dominance patterns of CD8 T cells in relation to disease progression in HIV-infected children.

CD8 T cells are important mediators of cellular immune responses as evidenced by clonal expansions in the CD8 TCR V beta repertoire during primary HIV infection in adults. This study investigated the CD8 TCR V beta repertoire by complementarity-determining region 3 length analysis using multiplex PCR in purified peripheral blood CD8 T cells of 22 HIV-infected children (age range was 0.75-15 yr, mean was 8.2 +/- 4.1 yr). Evidence of clonal dominance in one or more V beta families was obtained in 15 of 22 children. The patterns of clonal dominance were designated as major, minor, single, and none to indicate the involvement of three or more, two, one, or no V beta families, respectively. A pattern of major or minor clonal dominance was observed in 12 children (group 1), whereas 10 children had single or no clonal dominance (group 2). In comparison with group 2, the children in group 1 had a higher percentage of CD4 cells (28.3 +/- 11.6 vs 8.6 +/- 4.8, p < 0.001); a higher stimulation index in lymphoproliferative responses to Candida (92.0 +/- 59.5 vs 12.3 +/- 14.4, p = 0.002), tetanus (76.3 +/- 51.2 vs 11.2 +/- 12.7, p = 0.002), and alloantigens (178.3 +/- 298.9 vs 32.9 +/- 35.2, p < 0.001); and a lower percentage of CD8+HLA-DR+CD38+ cells (37.4 +/- 13.1 vs 54.6 +/- 14.2, p < 0.01). The number of dominant CD8 T cell clones was significantly correlated with the percentage of CD4 T cells (r = 0.669, p < 0.001) but not with plasma HIV RNA. Compared with group 1, patients in group 2 had a 4.8 times greater probability of having < 15% CD4 cells. These findings indicate that CD8 clonal dominance in HIV-infected children reflects robustness of immune responses, regardless of time since infection and virus load.

Characterization of a murine monoclonal antibody against human esophageal squamous cell carcinoma-associated antigen.

A murine monoclonal antibody (MAb) 2G3 of the IgG1 type was raised using the human esophageal squamous cell carcinoma (SCC) cell line TE-2. Immunoblotting with 2G3 indicated that the antigen recognized by 2G3 has a molecular weight of 34 kD. Its activity was evaluated by immunoperoxidase and immunofluorescence on frozen and paraffin sections of various normal tissues, normal and benign tumors as well as various established cell lines. The pattern of reactivity revealed that the antigen recognized by 2G3 was expressed mainly by esophageal SCC. The only exception was represented by malignant breast tumors, where it reacted weakly. Scatchard analysis using 125I-labelled 2G3 showed that TE-2 has approximately 7.5 times more binding sites than the MCF-7 breast cancer cell line. The use of this new MAb is therefore proposed for the histopathological diagnosis of esophageal SCC.

Topography of the photosystem I core proteins of the cyanobacterium Synechocystis sp. PCC 6803.

PsaA and PsaB are homologous integral membrane proteins that form the heterodimeric core of photosystem I. Domain-specific antibodies were generated to examine the topography of PsaA and PsaB. The purified photosystem I complexes from the wild type strain of Synechocystis sp. PCC 6803 were treated with eight proteases to study the accessibility of cleavage sites in PsaA and PsaB. Proteolytic fragments were identified using the information from N-terminal amino acid sequencing, reactivity to antibodies, apparent mass, and specificity of proteases. The extramembrane loops of PsaA and PsaB differed in their accessibility to proteases, which indicated the folded structure of the loops or their shielding by the small subunits of photosystem I. NaI-treated and mutant photosystem I complexes were used to identify the extramembrane loops that were exposed in the absence of specific small subunits. The absence of PsaD exposed additional proteolytic sites in PsaB, whereas the absence of PsaE exposed sites in PsaA. These studies distinguish PsaA and PsaB in the structural model for photosystem I that has been proposed on the basis of x-ray diffraction studies (Krauss, N., Schubert, W.-D., Klukas, O., Fromme, P., Witt, H. T., and Saenger, W. (1996) Nat. Struct. Biol. 3, 965-973). Using osmotically shocked cells for protease treatments, the N terminus of PsaA was determined to be on the n side of the photosynthetic membranes. Based on these data and available published information, we propose a topological model for PsaA and PsaB.

The PsaD subunit of photosystem I. Mutations in the basic domain reduce the level of PsaD in the membranes.

The PsaD subunit of photosystem I (PSI) is a peripheral protein that provides a docking site for ferredoxin and interacts with the PsaB, PsaC, and PsaL subunits of PSI. We used site-directed mutagenesis to determine the function of a basic region in PsaD of the cyanobacterium Synechocystis sp. PCC 6803. We generated five mutant strains in which one or more charged residues were altered. Western blotting showed that replacement of lysine (Lys)-74 with glutamine or glutamic acid led to a substantial decrease in the level of PsaD in the membranes. The mutant PSI complexes showed reduced NADP+ photoreduction activity mediated by ferredoxin; the decrease in activity correlated with the reduced level of PsaD. Using protein synthesis inhibitors we showed that the degradation rates of the mutant and wild-type PsaD were similar, indicating a defect in the assembly of the mutant protein. Treatment of the mutant PSI complexes with a different concentration of NaI showed that the mutations decreased affinity between PsaD and the transmembrane components of PSI. With glutaraldehyde, the mutant and wild-type PsaD proteins could be cross-linked with PsaC, but the PsaD-PsaL cross-linked product was reduced drastically when arginine-72, Lys-74, and Lys-76 were mutated simultaneously. These studies demonstrate that the basic residues in the central region of PsaD, especially Lys-74, are crucial in the assembly of PsaD into the PSI complex.