gyrB as a phylogenetic discriminator for members of the Bacillus anthracis-cereus-thuringiensis group

Bacillus anthracis, the causative agent of the human disease anthrax, Bacillus cereus, a food-borne pathogen capable of causing human illness, and Bacillus thuringiensis, a well-characterized insecticidal toxin producer, all cluster together within a very tight clade (B. cereus group) phylogenetically and are indistinguishable from one another via 16S rDNA sequence analysis. As new pathogens are continually emerging, it is imperative to devise a system capable of rapidly and accurately differentiating closely related, yet phenotypically distinct species. Although the gyrB gene has proven useful in discriminating closely related species, its sequence analysis has not yet been validated by DNA:DNA hybridization, the taxonomically accepted "gold standard". We phylogenetically characterized the gyrB sequences of various species and serotypes encompassed in the "B. cereus group," including lab strains and environmental isolates. Results were compared to those obtained from analyses of phenotypic characteristics, 16S rDNA sequence, DNA:DNA hybridization, and virulence factors. The gyrB gene proved more highly differential than 16S, while, at the same time, as analytical as costly and laborious DNA:DNA hybridization techniques in differentiating species within the B. cereus group.     

Proteome analysis to assess physiological changes in Escherichia coli grown under glucose-limited fed-batch conditions

Proteome analysis was used to compare global protein expression changes in Escherichia coli fermentation between exponential and glucose-limited fed-batch phase. Two-dimensional gel electrophoresis and MALDI-TOF mass spectrometry were used to separate and identify 49 proteins showing >2-fold difference in expression. Proteins upregulated during exponential phase include ribonucleotide biosynthesis enzymes and ribosomal recycling factor. Proteins upregulated during fed-batch phase include those involved in high-affinity glucose uptake, transport and degradation of alternate carbon sources and TCA cycle, suggesting an enhanced role of the cycle under glucose- and energy-limited conditions. We report the upregulation of several putative proteins (ytfQ, ygiS, ynaF, yggX, yfeX), not identified in any previous study under carbon-limited conditions.     

Cytologic diagnosis of pulmonary nocardiosis: a report of 3 cases

BACKGROUND: Nocardiosis is an uncommon infection and presents as an opportunistic infection in an immunocompromised host. Pulmonary infection by Nocardia may be difficult to diagnose based on clinical and radiologic features, as these are not specific. Sputum examination, bronchoalveolar lavage and transthoracic ultrasound/computed tomography-guided fine needle aspiration cytology offer a simple means of procuring material for diagnostic evaluation. Very few articles have described the morphologic appearance of this uncommon pathogen in cytologic material. CASES: Three cases occurred in patients with an underlying immunocompromised state. Patient 1 was on steroid therapy for nephrotic syndrome, patient 2 was on immunosuppressant therapy after renal transplantation, and patient 3 was HIV positive. A diagnosis of pulmonary nocardiosis was suspected on Papanicolaou stain. Modified Ziehl-Neelsen stain and silver methanamine stains were useful in confirming the diagnosis. CONCLUSION: A high index of suspicion for nocardiosis must be maintained while assessing cytologic material in immunosuppressed individuals as it may be masked by the intense inflammatory exudate associated with this infection. A meticulous search may reveal the presence of delicate, thin, faintly stained, branching filaments of Nocardia on routine Papanicolaou stain. Special stains and culture studies are useful in confirming the diagnosis.     

Survival of spacecraft-associated microorganisms under simulated martian UV irradiation

Spore-forming microbes recovered from spacecraft surfaces and assembly facilities were exposed to simulated Martian UV irradiation. The effects of UVA (315 to 400 nm), UVA+B (280 to 400 nm), and the full UV spectrum (200 to 400 nm) on the survival of microorganisms were studied at UV intensities expected to strike the surfaces of Mars. Microbial species isolated from the surfaces of several spacecraft, including Mars Odyssey, X-2000 (avionics), and the International Space Station, and their assembly facilities were identified using 16S rRNA gene sequencing. Forty-three Bacillus spore lines were screened, and 19 isolates showed resistance to UVC irradiation (200 to 280 nm) after exposure to 1,000 J m(-2) of UVC irradiation at 254 nm using a low-pressure mercury lamp. Spores of Bacillus species isolated from spacecraft-associated surfaces were more resistant than a standard dosimetric strain, Bacillus subtilis 168. In addition, the exposure time required for UVA+B irradiation to reduce the viable spore numbers by 90% was 35-fold longer than the exposure time required for the full UV spectrum to do this, confirming that UVC is the primary biocidal bandwidth. Among the Bacillus species tested, spores of a Bacillus pumilus strain showed the greatest resistance to all three UV bandwidths, as well as the total spectrum. The resistance to simulated Mars UV irradiation was strain specific; B. pumilus SAFR-032 exhibited greater resistance than all other strains tested. The isolation of organisms like B. pumilus SAFR-032 and the greater survival of this organism (sixfold) than of the standard dosimetric strains should be considered when the sanitation capabilities of UV irradiation are determined.     

Tree population changes in a tropical dry evergreen forest of south India over a decade (1992–2002)

Changes in species composition and density of trees >10 cm gbh in a tropical dry evergreen forest in Puthupet, south India are interpreted for the period between 1992 and 2002. A 1-ha plot was inventoried in 1992 and was recensused in 2002. During the 10-year interval tree taxa diversity as well as stand density increased, but the basal area value decreased. Tree species richness increased by 21% (from 24 to 29 species) by an addition of eight species and local extinction of three species. The tree density increased just by eight individuals (from 1330 stems ha⁻¹ in 1992 to 1338 ha⁻¹ in 2002), but the basal area decreased by 8% (from 37.5 to 34.5 m² ha⁻¹). Many species (11 numbers) have increased in abundance rather than decreased. Many surviving species seem to have considerable stability in abundance at the local scale. The density of smaller stems (10 29 cm gbh) increased by 15.3%, while that of the larger trees decreased drastically (81.6%). Ninety percent of the missing stems were from the middlestorey of the forest. Tree density changes among the three ecological guilds revealed a decrease in stem density and an increase in basal area in the lowerstorey; while the middlestorey exhibited a reverse trend. Family-wise, tree density changes revealed that the majority of families (67%) showed an increase in stem density. Long-term studies on tree population changes are essential to estimate tree mortality and recruitment rates, which will provide a greater insight in tropical forest dynamics.     

The role of sinking particles in the overwintering process of Vibrio parahaemolyticus in a marine environment

Abstract The behavioral pattern of Vibrio parahaemolytics during the winter season (December 1988 to March 1989) in the water column, sediment, plankton and sinking particles was determined in a eutrophic marine environment. A total of 15 environmental parameters and seven microbial characteristics were examined. This halophile was isolated sporadically from bottom water and plankton materials, whereas it was undetectable in the surface water and sediment samples. However, V. parahaemolyticus was isolated from the sinking particles continuously throughout the sampling period with highest counts during February 1989. Out of 195 strains identified, 10 Vibrio spp. and 3 Listonella spp. were observed of which V. alginolyticus was predominantly isolated irrespective of the samples tested. Simple correlation and multiple regression analyses show that the occurrence of V. parahaemolyticus is not governed by any single biotic or abiotic factor of the environment. Possibly, the cumulative effect of all these environmental parameters on the distribution of V. parahaemolyticus is conceivable.     

De Novo Mutations in Moderate or Severe Intellectual Disability

Genetics is believed to have an important role in intellectual disability (ID). Recent studies have emphasized the involvement of de novo mutations (DNMs) in ID but the extent to which they contribute to its pathogenesis and the identity of the corresponding genes remain largely unknown. Here, we report a screen for DNMs in subjects with moderate or severe ID. We sequenced the exomes of 41 probands and their parents, and confirmed 81 DNMs affecting the coding sequence or consensus splice sites (1.98 DNMs/proband). We observed a significant excess of de novo single nucleotide substitutions and loss-of-function mutations in these cases compared to control subjects, suggesting that at least a subset of these variations are pathogenic. A total of 12 likely pathogenic DNMs were identified in genes previously associated with ID (ARID1B, CHD2, FOXG1, GABRB3, GATAD2B, GRIN2B, MBD5, MED13L, SETBP1, TBR1, TCF4, WDR45), resulting in a diagnostic yield of ∼29%. We also identified 12 possibly pathogenic DNMs in genes (HNRNPU, WAC, RYR2, SET, EGR1, MYH10, EIF2C1, COL4A3BP, CHMP2A, PPP1CB, VPS4A, PPP2R2B) that have not previously been causally linked to ID. Interestingly, no case was explained by inherited mutations. Protein network analysis indicated that the products of many of these known and candidate genes interact with each other or with products of other ID-associated genes further supporting their involvement in ID. We conclude that DNMs represent a major cause of moderate or severe ID.     

Development of β-Carotene Rich Maize Hybrids through Marker-Assisted Introgression of β-carotene hydroxylase Allele

Development of vitamin A-rich cereals can help in alleviating the widespread problem of vitamin A deficiency. We report here significant enhancement of kernel β-carotene in elite maize genotypes through accelerated marker-assisted backcross breeding. A favourable allele (543 bp) of the β-carotene hydroxylase (crtRB1) gene was introgressed in the seven elite inbred parents, which were low (1.4 µg/g) in kernel β-carotene, by using a crtRB1-specific DNA marker for foreground selection. About 90% of the recurrent parent genome was recovered in the selected progenies within two backcross generations. Concentration of β-carotene among the crtRB1-introgressed inbreds varied from 8.6 to 17.5 µg/g - a maximum increase up to 12.6-fold over recurrent parent. The reconstituted hybrids developed from improved parental inbreds also showed enhanced kernel β-carotene as high as 21.7 µg/g, compared to 2.6 µg/g in the original hybrid. The reconstituted hybrids evaluated at two locations possessed similar grain yield to that of original hybrids. These β-carotene enriched high yielding hybrids can be effectively utilized in the maize biofortification programs across the globe.     

The neuroprotective role of melatonin against amyloid β peptide injected mice

Widespread cerebral deposition of a 40–42 amino acid peptide called amyloid β peptide (Aβ) in the form of amyloid fibrils is one of the most prominent neuropathologic features of Alzheimer's disease (AD). The clinical study provides evidence that accumulation of protofibrils due to the Arctic mutation (E22G) causes early AD onset. Melatonin showed beneficial effects in an AD mouse model. Mice were divided into four different groups (n=8 per group): (i) control group, (ii) scrambled Aβ-injected group, (iii) Aβ protofibril-injected group and (iv) melatonin-treated group. A single dose of (5 µg) Aβ protofibril was administered to the Aβ protofibril-injected and melatonin-treated groups via intracerebroventricular injections. The results demonstrate that melatonin treatment significantly reduces Aβ protofibril-induced reactive oxygen species (ROS) production, intracellular calcium levels and acetylcholinesterase activity in the neocortex and hippocampus regions. Based on these findings it is suggested that melatonin therapy might be a useful treatment for AD patients.     

TRIAL: a tool for finding distant structural similarities

Finding structural similarities in distantly related proteins can reveal functional relationships that can not be identified using sequence comparison. Given two proteins A and B and threshold ε ?, we develop an algorithm, TRiplet-based Iterative ALignment (TRIAL) for computing the transformation of B that maximizes the number of aligned residues such that the root mean square deviation (RMSD) of the alignment is at most ε ?. Our algorithm is designed with the specific goal of effectively handling proteins with low similarity in primary structure, where existing algorithms perform particularly poorly. Experiments show that our method outperforms existing methods. TRIAL alignment brings the secondary structures of distantly related proteins to similar orientations. It also finds larger number of secondary structure matches at lower RMSD values and increased overall alignment lengths. Its classification accuracy is up to 63 percent better than other methods, including CE and DALI. TRIAL successfully aligns 83 percent of the residues from the smaller protein in reasonable time while other methods align only 29 to 65 percent of the residues for the same set of proteins.