Abundance, diversity, and host assignment of total, intracellular, and extracellular antibiotic resistance genes in riverbed sediments.

Human health risk assessment for environmental antibiotic resistant microbes requires not only quantifying the abundance of antibiotic resistance genes (ARGs) in environmental matrices, but also understanding their hosts and genetic context. Further, differentiating ARGs in intracellular and extracellular DNA (iDNA and eDNA) fractions may help refine our understanding of ARG transferability. The objectives of this study were to understand the (O1) abundance and diversity of extracellular, intracellular, and total ARGs along a land use gradient and (O2) impact of bioinformatics pipeline on the assignment of putative hosts for the ARGs observed in the different DNA fractions. Sediment samples were collected along a land use gradient in the Raritan River, New Jersey, USA. DNA was extracted to separate eDNA and iDNA and qPCR was performed for select ARGs and the 16S rRNA gene. Shotgun metagenomic sequencing was performed on DNA extracts for the different DNA fractions. ARG hosts were assigned via two different bioinformatic pipelines: network analysis of raw reads versus assembly. Results of the two pipelines were compared to evaluate their performance in terms of number and diversity of linkages and accuracy of in silico matrix spike host assignments. No differences were observed in the 16S rRNA gene normalized sul1 concentrations between the DNA fractions. The overall microbial community structure was more similar for iDNA and total DNA compared to eDNA and generally clustered by sampling site. ARGs associated with mobile genetic elements increased in iDNA for the downstream sites. Regarding host assignment, the raw reads pipeline via network analysis identified 247 ARG hosts as compared to 53 hosts identified by assembly pipeline. Other comparisons between the pipelines were made including ARG assignment to taxa containing waterborne pathogens and practical considerations regarding processing time.

Nitrogen-doped graphene-like carbon nanosheets from commercial glue: morphology, phase evolution and Li-ion battery performance.

We report a two-step process to synthesize nitrogen-doped graphene-like carbon nanosheets (N-CNS), using commercially available ethyl cyanoacrylate based super glue as a carbon precursor. In this process, super glue is polymerized in aqueous NaCl solution, followed by carbonization at 1000 °C. Scanning electron microscopy (SEM) and high resolution transmission electron microscopy (HRTEM) studies show that the resultant material consists of micron-sized carbon nanosheets with a wrinkled morphology. HRTEM, X-ray diffraction (XRD), XPS and Raman spectroscopic studies confirm the formation of nanocrystalline and graphitic, nitrogen-doped carbon nanosheets. A detailed FTIR analysis of the degradation products of the polymeric precursor (polyethyl cyanoacrylate) at various heat treatment temperatures under an inert atmosphere reveals that the polymer undergoes a cyclization process similar to polyacrylonitrile (PAN) during carbonization to yield the N-CNS. The N-CNS used as an anode for a lithium-ion battery shows stable reversible capacities of 480 mA h g-1 for 100 cycles, which indicates that N-CNS are promising materials for lithium-ion battery applications. In a broader perspective, a unique chemical transformation of polyethyl cyanoacrylate to graphitic carbon may be useful to design new nanostructured carbons for a plethora of applications.

Evaluation of various laboratory assays in detection of heparin-induced thrombocytopenia in an adult general ICU.

INTRODUCTION: Heparin-induced thrombocytopenia (HIT) is an immune-mediated complication of heparin therapy. Our objective was (i) to compare various laboratory assays for HIT against clinical probability (4-T score) and (14) C-serotonin release assay (SRA), which was the composite gold standard and (ii) to determine the incidence of HIT in the ICU. METHODS: The study group (n = 217) consisted of consecutive ICU patients with heparin exposure followed by thrombocytopenia. The clinical probability (4-T score) was applied to the study group. Enzyme-linked immunosorbent assay (ELISA), particle gel immunoassay (PGIA), SRA, and platelet aggregation assay (PAA) were performed. RESULTS: The 4-T score showed that 1/217 patients had high probability, 48 had intermediate probability, and 168 had low probability for HIT. One patient was positive by SRA, three by PGIA, and 33 by ELISA. The incidence based on a combination of clinical features and laboratory findings was 1.8%. CONCLUSIONS: A greater number of false positives were observed by ELISA than by PGIA when compared to a composite gold standard of SRA and clinical probability. The incidence of SRA-positive HIT was 0.46% (1/217).

Stability indicating HPLC method for simultaneous determination of mephenesin and diclofenac diethylamine.

A simple, specific, accurate and stability-indicating reversed phase high performance liquid chromatographic method was developed for the simultaneous determination of mephenesin and diclofenac diethylamine, using a Spheri-5-RP-18 column and a mobile phase composed of methanol: water (70:30, v/v), pH 3.0 adjusted with o-phosphoric acid. The retention times of mephenesin and diclofenac diethylamine were found to be 3.9 min and 14.5 min, respectively. Linearity was established for mephenesin and diclofenac diethylamine in the range of 50-300 mug/ml and 10-60 mug/ml, respectively. The percentage recoveries of mephenesin and diclofenac diethylamine were found to be in the range of 99.06-100.60% and 98.95-99.98%, respectively. Both the drugs were subjected to acid, alkali and neutral hydrolysis, oxidation, dry heat, photolytic and UV degradation. The degradation studies indicated, mephenesin to be susceptible to neutral hydrolysis, while diclofenac diethylamine showed degradation in acid, H(2)O(2), photolytic and in presence of UV radiation. The degradation products of diclofenac diethylamine in acidic and photolytic conditions were well resolved from the pure drug with significant differences in their retention time values. This method can be successfully employed for simultaneous quantitative analysis of mephenesin and diclofenac diethylamine in bulk drugs and formulations.

Semi-automated imaging system to quantitate estrogen and progesterone receptor immunoreactivity in human breast cancer.

A semi-automated imaging system is described to quantitate estrogen and progesterone receptor immunoreactivity in human breast cancer. The system works for any conventional method of image acquisition using microscopic slides that have been processed for immunohistochemical analysis of the estrogen receptor and progesterone receptor. Estrogen receptor and progesterone receptor immunohistochemical staining produce colorimetric differences in nuclear staining that conventionally have been interpreted manually by pathologists and expressed as percentage of positive tumoral nuclei. The estrogen receptor and progesterone receptor status of human breast cancer represent important prognostic and predictive markers of human breast cancer that dictate therapeutic decisions but their subjective interpretation result in interobserver, intraobserver and fatigue variability. Subjective measurements are traditionally limited to a determination of percentage of tumoral nuclei that show positive immunoreactivity. To address these limitations, imaging algorithms utilizing both colorimetric (RGB) as well as intensity (gray scale) determinations were used to analyze pixels of the acquired image. Image acquisition utilized either scanner or microscope with attached digital or analogue camera capable of producing images with a resolution of 20 pixels /10 mu. Areas of each image were screened and the area of interest richest in tumour cells manually selected for image processing. Images were processed initially by JPG conversion of SVS scanned virtual slides or direct JPG photomicrograph capture. Following image acquisition, images were screened for quality, enhanced and processed. The algorithm-based values for estrogen receptor and progesterone receptor percentage nuclear positivity both strongly correlated with the subjective measurements (intraclass correlation: 0.77; 95% confidence interval: 0.59, 0.95) yet exhibited no interobserver, intraobserver or fatigue variability. In addition the algorithms provided measurements of nuclear estrogen receptor and progesterone receptor staining intensity (mean, mode and median staining intensity of positive staining nuclei), parameters that subjective review could not assess. Other semi-automated image analysis systems have been used to measure estrogen receptor and progesterone receptor immunoreactivity but these either have required proprietary hardware or have been based on luminosity differences alone. By contrast our algorithms were independent of proprietary hardware and were based on not just luminosity and colour but also many other imaging features including epithelial pattern recognition and nuclear morphology. These features provide a more accurate, versatile and robust imaging analysis platform that can be fully automated in the near future. Because of all these properties, our semi-automated imaging system 'adds value' as a means of measuring these important nuclear biomarkers of human breast cancer.

Evaluation of efficacy of nizer versus nimesulide tablets in otitis media.

An evaluation of clinical efficacy of nizer versus nimesulide tablets was undertaken in 118 patients suffering from otitis media. Nizer demonstrated rapid and powerful analgesic and antipyretic effects compared to plain nimesulide, without additional side effects.

Fetal hydantoin syndrome with rheumatic valvular heart disease.

Research has shown that anticonvulsants are teratogens and pose a risk for fetal malformations. Though Fetal Hydantoin Syndrome (FHS) was first reported by Langhman and others, wide phenotypic variability of this syndrome has lead many clinicians to question its very existence. We report a twelve year old girl with FHS with rheumatic valvular heart disease.

Asian paralysis syndrome.

We report 20 children admitted to the paediatric ward of a public general hospital for acute flaccid paralysis, which was bilaterally symmetrical in all cases and was associated with bulbar involvement in eight of them. Recovery was partial. Nerve conduction studies showed motor axonal neuropathy. This new disease, variously termed as non-inflammatory neuropathy/Chinese paralysis syndrome must be differentiated from Guillain-Barré syndrome (GBS) and poliomyelitis. Both GBS and Asian paralysis syndrome have bilaterally symmetrical flaccid paralysis but GBS tends to have sensory involvement, full recovery occurs in 90% of cases and nerve conduction shows demyelinating neuropathy. Asian paralysis syndrome and poliomyelitis are pure motor lesions without sensory changes and partial recovery, but poliomyelitis differs in that paralysis is asymmetrical and unequal, muscle spasm is always present in the initial stage and there are prodromal symptoms. Nerve conduction studies show anterior horn cell disease. This new entity, common in Asian populations, assumes public health importance when it mimics poliomyelitis in a country that has tried to eliminate poliomyelitis by universal immunization. To the best of our knowledge, this is the first report of Asian paralysis syndrome in children in our area.

The phenyl propanoid pathway enzymes in Solanum tuberosum exist as a multienzyme complex.

The elution profile of the core sequence enzymes of the phenyl propanoid pathway, namely phenyl alanine ammonia lyase, t-cinnamic acid 4-hydroxylase and p-coumaryl CoA ligase, on AcA 34 column suggested the existence of a high molecular form (P1) and a low molecular form (P2) for all the three enzymes. All the P1 forms eluted together in same fractions, while the P2 forms eluted out according to their respective molecular mass. Rechromatography of P1 form under identical conditions showed a similar elution profile (Q1 and Q2 forms). Further, the Q1 form did not show any significant increase in specific activity when compared to the P1 form. These results suggested the possibility of these enzymes existing as a protein cluster. Further confirmation was obtained on repeated column chromatography of the Q1 form in presence of 0.1 M KCl which did not result in complete dissociation of the complex to its individual enzyme components. The identification of the subunit polypeptide of the individual enzyme components in the multi enzyme complex and the in vitro demonstration of the phenyl propanoid core pathway reaction sequence using phenylalanine alone as a substrate supplementing the required cofactors for appropriate reactions substantiated that at least the core enzymes of the phenyl propanoid sequence existed as a multi enzyme complex.

Identification of a non-microsomal cinnamic acid 4-hydroxylase from potato tuber (S. tuberosum) and its partial purification.

The cytoplasmic localisation of cinnamic acid 4-hydroxylase (CA4H) has been shown by isolation and subcellular fractionation of the enzyme in Hepes buffer. The enzyme was purified by ammonium sulphate fractionation followed by AcA-34 molecular sieve chromatography. The enzyme existed as a high molecular mass which dissociated to a lower form on dilution on the column. The pH optimum, sulphydryl requirement, molecular and preliminary kinetic characteristics were investigated.

Studies on the induction of cinnamic acid 4-hydroxylase in potato tuber.

The change in activity of cinnamic acid 4-hydroxylase (CA4H) in potato parenchyma tissue exposed to various conditions has been examined. Maximum induction of CA4H activity was obtained at 18 hr of incubation. Though CA4H induction can occur in dark, over 100% increase in enzyme activity was obtained on exposure of the tissue to light. Actinomycin D and cycloheximide inhibited the induction process. Mn2+, though known to cause an induction of CA4H in Jerusalem Artichoke, strongly inhibited potato CA4H induction. Dithiothreitol enhanced the CA4H activity due to either activation or protection of the enzyme. CA4H induction was significantly regulated at very low concentrations of trans-cinnamate and paracoumarate.

A platform for biological sequence comparison on parallel computers.

We have written two programs for searching biological sequence databases that run on Intel hypercube computers. PSCANLIB compares a single sequence against a sequence library, and PCOMPLIB compares all the entries in one sequence library against a second library. The programs provide a general framework for similarity searching; they include functions for reading in query sequences, search parameters and library entries, and reporting the results of a search. We have isolated the code for the specific function that calculates the similarity score between the query and library sequence; alternative searching algorithms can be implemented by editing two files. We have implemented the rapid FASTA sequence comparison algorithm and the more rigorous Smith-Waterman algorithm within this framework. The PSCANLIB program on a 16 node iPSC/2 80386-based hypercube can compare a 229 amino acid protein sequence with a 3.4 million residue sequence library in approximately 16 s with the FASTA algorithm. Using the Smith-Waterman algorithm, the same search takes 35 min. The PCOMPLIB program can compare a 0.8 million amino acid protein sequence library with itself in 5.3 min with FASTA on a third-generation 32 node Intel iPSC/860 hypercube.

Serum Zinc levels in pregnant women, under oral contraceptive therapy and in normal (ovulating) women.

PIP: Serum zinc levels were estimated by atomic absorption spectrophotometer in 15 women with normal menstrual cycles, in 15 women taking oral contraceptives (OCs), and in 25 women with normal pregnancies in the 3rd trimester. In normally menstruating women, a significant increase in serum zinc levels was observed at the time of ovulation; i.e., from 0.586 mcgm/ml-0.840 mcgm/ml. In women taking combined OCs (Orlest), zinc levels were lower and a midcycle peak was missing. The serum zinc levels were decreased during the 3rd trimester of pregnancy to 0.289 mcgm/ml.^ieng