Determination of growth rate of microorganisms in broth from oxygen-sensitive fluorescence plate reader measurements

A novel method utilizing the BD Oxygen Biosensor System has been developed to rapidly, simply, and accurately determine the growth rate of microorganisms in broth, with no needfor plate counts, standardized inocula, or technically difficult manipulations. The BD Oxygen Biosensor System incorporates an oxygen-sensitive material into the wells of standard Falcon microplates. The time response of this sensor monitored in afluorescence plate reader can be used to quantitate microbe growth. The method entails seeding a dilution series of microorganism onto the plate and reading at regular intervals for 3-10 h. As the organisms grow and consume oxygen, thefluorescence intensities increase over time to form a family of sigmoidal growth curves. A simple mathematical analysis of the time intervals between the curves yields the doubling time, which is independent of the initial concentration of organism. The method is ideally suited as a screening tool for assessing the impact of culture conditions, media composition, or added compounds on growth kinetics.     

Genetic basis of plant height and its degree of indetermination in mungbean (Vigna radiata (L.) Wilczek)

The genetic basis of plant height at various growth stages and the degree of indetermination of plant height in six mungbean genotypes (NM 92, 6601, NM 89, VC 1560D and VC 3902A) were assessed through half diallel cross. Cultivars, 6601 and NM 92, were the best general combiner for pre-flowering dry matter accumulation and minimum increase in plant height from first flower to 90% pods maturity, respectively. For these traits, the combination NM 92 x NM 89 was the best specific combiner of all the crosses. Both additive and dominant gene effects controlled the inheritance of plant height at first pod and to 90% pods maturity, degree of indetermination of plant height (DDh) from first flower to first pod maturity (DDh1), DDh from first flower to 90% pods maturity (DDh2) and DDh from first pod maturity to 90% pods maturity (DDh3). Plant height at first flower was additively inherited. The additive gene action was predominant as compared to dominant gene action for all the traits examined. High narrow and broad sense heritability estimates for DDh2 showed that better response to selection is possible for the development of mungbean genotypes with minimum increase in plant height during post-flowering development.     

Conformation of an immunoreactive undecapeptide fragment (10–20) of Asp f 1 by NMR and molecular modeling

Allergic bronchopulmonary aspergillosis (ABPA), caused by Aspergillus fumigatus, is a complication of allergic asthma. Asp f 1 secreted by A. fumigatus is reported to be a major allergen/antigen involved in pathogenesis of aspergillosis. A 11-mer immunodominant epitope (Leu-Asn-Pro-Lys-Thr⁵-Asn-Lys-Trp-Glu-Asp¹⁰-Lys) of Asp f 1 has shown immunoreactivity with specific IgG and IgE antibodies in the sera of patients with ABPA in ELISA inhibitionassay. Various studies have suggested that the peptide has a potential use in the development of ELISA based diagnostic kit for early diagnosis of infections caused by A. fumigatus.In view of these interesting properties of the undecapeptide wehave embarked on an investigation of its conformation to understand the relationship between structure and immunoreactivity. NMR and molecular modeling studies of the peptide suggest a structure with a -turn spanning residuesAsn⁶ – Glu⁹ in water at pH 4.0, a -pleated sheet in DMSO and a -helix in 40% HFA.     

In vitro propagation of pummelo (Citrus grandis L. osbeck)

Summary Several experiments were carried out to develop protocols for the in vitro propagation of pummelo (Citrus grandis L. Osbeck) using shoot-tip explants from seedlings. Murashige and Skoog (MS) medium supplemented with various concentrations of 6-benzylaminopurine (BA) and thidiazuron (TDZ), singly or in combination with α-naphthaleneacetic acid (NAA), was used to determine the rate of shoot proliferation. The response of explants to all concentrations of TDZ was very poor. After 6 wk culture, the most adventitious shoots per explant (average 5.2) were obtained on medium supplemented with 1.8 μM BA. NAA with cytokinin in the medium did not improve the rate of shoot multiplication significantly. Addition of 5.8 μM gibberellic acid in shoot-proliferation medium during the second subculture improved shoot elongation significantly. Shoot multiplication increased 3.5-fold in each successive subculture. NAA was superior to indolebutyric acid for in vitro root induction. Over 75% of the shoots developed roots when transferred to half-strength MS medium with 1.3, 2.7, or 5.4 μM NAA.     

Singular value decomposition of 3-D DNA melting curves reveals complexity in the melting process

The thermal denaturation of synthetic deoxypolynucleotides of defined sequence was studied by a three dimensional melting technique in which complete UV absorbance spectra were recorded as a function of temperature. The results of such an experiment defined a surface bounded by absorbance, wavelength, and temperature. A matrix of the experimental data was built, and analyzed by the method of singular value decomposition (SVD). SVD provides a rigorous, model-free analytical tool for evaluating the number of significant spectral species required to account for the changes in UV absorbance accompany-ing the duplex – to – single strand transition. For all of the polynucleotides studied (Poly dA – Poly dT; [Poly (dAdT)]₂; Poly dG – Poly dC; [Poly(dGdC)]₂), SVD indicated the existence of at least 4 – 5 significant spectral species. The DNA melting transition for even these simple repeating sequences cannot, therefore, be a simple two-state process. The basis spectra obtained by SVD analysis were found to be unique for each polynucleotide studied. Differential scanning calorimetry was used to obtain model free estimates for the enthalpy of melting for the polynucleotides studied, with results in good agreement with previously published values. Received: 16 April 1997 / Accepted: 9 July 1997     

Role of ACE and PAI-1 Polymorphisms in the Development and Progression of Diabetic Retinopathy

In the present study we determined the association of angiotensin converting enzyme (ACE) and plasminogen activator inhibitor-1 (PAI-1) gene polymorphisms with diabetic retinopathy (DR) and its sub-clinical classes in Pakistani type 2 diabetic patients. A total of 353 diabetic subjects including 160 DR and 193 diabetic non retinopathy (DNR) as well as 198 healthy controls were genotyped by allele specific polymerase chain reaction (PCR) for ACE Insertion/Deletion (ID) polymorphism, rs4646994 in intron 16 and PAI-1 4G/5G (deletion/insertion) polymorphism, rs1799768 in promoter region of the gene. To statistically assess the genotype-phenotype association, multivariate logistic regression analysis was applied to the genotype data of DR, DNR and control individuals as well as the subtypes of DR. The ACE genotype ID was found to be significantly associated with DR (p = 0.009, odds ratio (OR) 1.870 [95% confidence interval (CI) = 1.04–3.36]) and its sub-clinical class non-proliferative DR (NPDR) (p = 0.006, OR 2.250 [95% CI = 1.098–4.620]), while PAI polymorphism did not show any association with DR in the current cohort. In conclusion in Pakistani population the ACE ID polymorphism was observed to be significantly associated with DR and NPDR, but not with the severe form of the disease i.e. proliferative DR (PDR).     

Stereoselective Synthesis of (R)‐3‐Methylthalidomide by Piperidin‐2‐one Ring Assembly Approach

A simple and stereoselective synthesis of 3‐methylthalidomide, a configurationally stable thalidomide analog, is presented. Herein we describe the synthesis of (R)‐3‐methylthalidomide starting from (S)‐alanine by piperidin‐2‐one ring assembly approach in high yield and enantiomeric purity without using a chiral auxiliary or reagent. Starting from (R)‐alanine, the corresponding (S)‐3‐methylthalidomide can be prepared using the same methodology. Chirality 27:619–624, 2015. © 2015 Wiley Periodicals, Inc.     

Predictors of Two Months Culture Conversion in Multidrug-Resistant Tuberculosis: Findings from a Retrospective Cohort Study

Background

Various studies have reported culture conversion at two months as a predictor of successful treatment outcome in multidrug-resistant tuberculosis (MDR-TB).

Objectives

The present study was conducted with the aim to evaluate the rate and predictors of culture conversion at two months in MDR-TB patients.

Methods

All confirmed pulmonary MDR-TB patients enrolled for treatment at Lady Reading Hospital Peshawar, Pakistan from 1 January to 31 December 2012 and met the inclusion criteria were reviewed retrospectively. Rate and predictors of culture conversion at two months were evaluated.

Results

Eighty seven (53.4%) out of 163 patients achieved culture conversion at two months. In a multivariate analysis lung cavitation at baseline chest X-ray (P = 0.006, OR = 0.349), resistance to ofloxacin (P = 0.041, OR = 0.193) and streptomycin (P = 0.017, OR = 0.295) had statistically significant (P<0.05) negative association with culture conversion at two months.

Conclusion

A reasonable proportion of patients achieved culture conversion at two months. Factors negatively associated with culture conversion at two months can be easily identified either before diagnosis or early in the course of MDR-TB treatment. This may help in better care of individual patients by identifying them early and treating them vigorously.