Mycobacteria mobility shift assay: a method for the rapid identification of Mycobacterium tuberculosis and nontuberculous mycobacteria

The identification of mycobacteria is essential because tuberculosis (TB) and mycobacteriosis are clinically indistinguishable and require different therapeutic regimens. The traditional phenotypic method is time consuming and may last up to 60 days. Indeed, rapid, affordable, specific and easy-to-perform identification methods are needed. We have previously described a polymerase chain reaction-based method called a mycobacteria mobility shift assay (MMSA) that was designed for Mycobacterium tuberculosis complex (MTC) and nontuberculous mycobacteria (NTM) species identification. The aim of this study was to assess the MMSA for the identification of MTC and NTM clinical isolates and to compare its performance with that of the PRA-hsp65 method. A total of 204 clinical isolates (102 NTM and 102 MTC) were identified by the MMSA and PRA-hsp65. For isolates for which these methods gave discordant results, definitive species identification was obtained by sequencing fragments of the 16S rRNA and hsp65 genes. Both methods correctly identified all MTC isolates. Among the NTM isolates, the MMSA alone assigned 94 (92.2%) to a complex or species, whereas the PRA-hsp65 method assigned 100% to a species. A 91.5% agreement was observed for the 94 NTM isolates identified by both methods. The MMSA provided correct identification for 96.8% of the NTM isolates compared with 94.7% for PRA-hsp65. The MMSA is a suitable auxiliary method for routine use for the rapid identification of mycobacteria.     

Biologically meaningful expression profiling across species using heterologous hybridization to a cDNA microarray

Background  

Unravelling the path from genotype to phenotype, as it is influenced by an organism's environment, is one of the central goals in biology. Gene expression profiling by means of microarrays has become very prominent in this endeavour, although resources exist only for relatively few model systems. As genomics has matured into a comparative research program, expression profiling now also provides a powerful tool for non-traditional model systems to elucidate the molecular basis of complex traits.     

Phylogeny of the genus trichoderma based on sequence analysis of the internal transcribed spacer region 1 of the rDNA cluster

Sequences of the internal transcribed spacer region 1 (ITS1) of the ribosomal DNA were used to determine the phylogenetic relationships of species of Trichoderma sect. Pachybasium. To this end, 85 strains-including all the available ex-type strains-were analyzed. Parsimony analysis demonstrated that the section is nonmonophyletic, distributing the 85 strains among three main groups that were supported by bootstrap values. Group A comprises two clades (A1 and A2), with A1 including T. polysporum, T. piluliferum, and T. minutisporum, while A2 included T. hamatum, T. pubescens, and T. strigosum in addition to species previously included in sect. Trichoderma (i.e., T. viride, T. atroviride, and T. koningii). The ex-type strain of T. fasciculatum formed a separate branch basal to clade A. Clade B contained the sect. Pachybasium members T. harzianum, T. fertile, T. croceum, T. longipile, T. strictipile, T. tomentosum, T. oblongisporum, T. flavofuscum, T. spirale, and the anamorphs of Hypocrea semiorbis and H. cf. gelatinosa. Sequence differences among clades A1, A2, and B were in the same order of magnitude as between each of them and T. longibrachiatum, which was used as an outgroup in these analyses. Sequence differences within clades A1, A2, and B were considerably smaller: in some cases (i.e., T. virens and T. flavofuscum; T. strictipile and H. cf. gelatinosa), the ITS1-sequences were identical, suggesting conspecifity. In other cases (e.g., T. crassum and T. longipile; T. harzianum, T. inhamatum, T. croceum, T. fertile, and H. semiorbis; T. hamatum and T. pubescens; and T. viride, T. atroviride, and T. koningii) differences were in the range of 1-3 nt only, suggesting a very close phylogenetic relationship. The sequence of a previously described aggressive mushroom competitor group of T. harzianum strains (Th2) was strikingly different from that of the ex-type strain of T. harzianum and closely related species and is likely to be a separate species. Copyright 1998 Academic Press.     

Patterns of inflammation and the use of reversibility testing in smokers with airway complaints

Background

Although both smoking and respiratory complaints are very common, tools to improve diagnostic accuracy are scarce in primary care. This study aimed to reveal what inflammatory patterns prevail in clinically established diagnosis groups, and what factors are associated with eosinophilia.

Method

Induced sputum and blood plasma of 59 primary care patients with COPD (n = 17), asthma (n = 11), chronic bronchitis (CB, n = 14) and smokers with no respiratory complaints ('healthy smokers', n = 17) were collected, as well as lung function, smoking history and clinical work-up. Patterns of inflammatory markers per clinical diagnosis and factors associated with eosinophilia were analyzed by multiple regression analyses, the differences expressed in odds ratios (OR) with 95% confidence intervals.

Results

Multivariately, COPD was significantly associated with raised plasma-LBP (OR 1.2 [1.04–1.37]) and sTNF-R55 in sputum (OR 1.01 [1.001–1.01]), while HS expressed significantly lowered plasma-LBP (OR 0.8 [0.72–0.95]). Asthma was characterized by higher sputum eosinophilic counts (OR 1.3 [1.05–1.54]), while CB showed a significantly higher proportion of sputum lymphocytic counts (OR 1.5 [1.12–1.9]). Sputum eosinophilia was significantly associated with reversibility after adjusting for smoking, lung function, age, gender and allergy.

Conclusion

Patterns of inflammatory markers in a panel of blood plasma and sputum cells and mediators were discernable in clinical diagnosis groups of respiratory disease. COPD and so-called healthy smokers showed consistent opposite associations with plasma LBP, while chronic bronchitics showed relatively predominant lymphocytic inflammation compared to other diagnosis groups. Only sputum eosinophilia remained significantly associated with reversibility across the spectrum of respiratory disease in smokers with airway complaints.     

Impact of Ramadan intermittent fasting on cognitive function in trained cyclists: a pilot study

This study assessed selected measures of cognitive function in trained cyclists who observed daylight fasting during Ramadan. Eleven cyclists volunteered to participate (age: 21.6±4.8 years, VO₂max: 57.7±5.6 ml kg⁻¹·min⁻¹) and were followed for 2 months. Cognitive function (Cambridge Neuropsychological Test Automated Battery (CANTAB), Reaction Time index (RTI) and Rapid Visual Information Processing (RVP) tests) and sleep architecture (ambulatory EEG) were assessed: before Ramadan (BR), in the 1st week (RA1) and 4th week of Ramadan (RA4), and 2 weeks post-Ramadan (PR). Both cognitive tests were performed twice per day: before and after Ramadan at 8-10 a.m. and 4-6 p.m., and during Ramadan at 4-6 p.m. and 0-2 a.m., respectively. Training load (TL) by the rating of perceived exertion (RPE) method and wellness (Hooper index) were measured daily. If the TL increased over the study period, this variable was stable during Ramadan. The perceived fatigue and delayed onset muscle soreness (DOMS) increased at RA4. Sleep patterns and architecture showed clear disturbances, with significant increases in the number of awakenings and light sleep durations during Ramadan (RA1 and RA4), together with decreased durations of deep and REM sleep stages at PR. RTI (simple and multiple reaction index) reaction and movement times did not vary over the study period. The RVP test showed reduced false alarms during Ramadan, suggesting reduced impulsivity. Overall accuracy significantly increased at RA1, RA4 and PR compared to baseline. At RA4, the accuracy was higher at 0-2 a.m. compared to 4-6 p.m. Despite the observed disturbances in sleep architecture, Ramadan fasting did not negatively impact the cognitive performance of trained cyclists from the Middle East.     

Inter-specific hybridization between freshwater catfish Mystus cavasius (Ham & Buch) and M. seenghala (Sykes) by artificial fertilization

By employing the technique of induced ovulation for artificial fertilization, inter-specific hybrids between the threatened catfish species (Mystus cavasius x M. seenghala) were produced. Fertilization, hatching and survival were significantly different between control and hybrids. The hatching time of the hybrid was significantly lower than that of the control. The average performance viz., hatching time and viability of larvae of the control fish, was significantly better than that of the hybrids. In the hybrid cross, hatchlings were mostly deformed and abnormal and after yolk absorption ultimately succumbed.