Recent methods for discovering novel bioactive metabolites, specifically antimicrobial agents, from marine-associated micro-organisms.

Marine micro-organisms are a promising source for novel natural compounds with many medical and biotechnological applications. Here, we demonstrate limitations and recent strategies for investigating the marine microbial community for novel bioactive metabolites, specifically those of antimicrobial potential. These strategies include culture-dependent methods such as modifying the standard culture media, including changing the gelling agent, dissolving vehicle, media supplementation and preparation to access a broader range of bacterial diversity from marine samples. Furthermore, we discuss strategies such as in situ cultivation, dilution-to-extinction cultivation and long-term incubation. We are presenting recent applications of culture-independent methods such as genome mining, proteomics profiling and the application of metagenomics as a novel strategy for structure confirmation in the discovery of the marine micro-organism for novel antimicrobial metabolites. We present this review as a simple guide and a helpful resource for those who seek to enter the challenging field of applied marine microbiology.

Analysis of the beak and feather disease viral genome indicates evidence of multiple introduction events into Saudi Arabia.

Psittacine beak and feather disease (PBFD), caused by beak and feather disease virus (BFDV) is a highly contagious disease in wild and captive psittacine populations and has an almost global presence. However, the BFDV infection in Saudi Arabia remains largely unknown. In the present study, we report the full genome sequence of BFDV strains from Saudi Arabia and its genetic diversity. The complete genome sequences were analyzed for 14 BFDV-infected birds representing 6 psittacine species. The complete genome sequence of BFDV strains was compared with 201 previously reported sequences to evaluate their diversity and possible recombination events, if any. Our analysis revealed that newly sequenced BFDV genomes from Saudi Arabia belonged to six different strains. Phylogenetic analysis suggested that the isolated BFDV genomes were highly recombinant with a high degree of diversity. It is evident from the study that psittacine species in Saudi Arabia are at risk from the spread of BFDV. As per the CITES trade database, about 190,000 parrots have been imported to Saudi Arabia since 1975 over a thousand instances. Presumably, during any of these trade events or unregulated trade of birds has predisposed the introduction of BFDV to Saudi Arabia. Understanding the epidemiology of BFDV is necessitated to address the threat posed by the virus to the psittacine population of Saudi Arabia.

Morphologic and biochemical characteristics of transplantable neurogenic tumors induced by N-ethyl-N-nitrosourea in inbred BD IX rats.

Brain and nerve tumors were induced transplacentally in inbred BD IX rats by systemic application of N-ethyl-N-nitrosourea. Because primary gliomas and neurinomas produced in this way are composed of heterogeneous cell populations, changes in tumor morphology were expected to occur during serial transplantation in syngeneic hosts. In this study such changes in morphology were correlated with the expression of two biochemical nervous system markers, S-100 protein and 2',3'-cyclic nucleotide 3'-phosphohydrolase. Several changes were observed during serial transplantation, including increased growth rate (even after one passage), preferential growth of anaplastic versus differentiated glial and Schwann's cells, varying degrees of fibrosarcomatous changes after prolonged serial transplantation, and reduced levels of S-100 protein. In contrast, tumors derived from biochemically differentiated clonal cell lines retained their morphologic and biochemical characteristics to a much greater extent, even after prolonged periods of sc transplantation.

Preparation and characterization of alginate/silver/nicotinamide nanocomposites for treating diabetic wounds.

Silver/Alginate/Nicotinamide nanoparticles composite (Ag/ALG/Nic) was prepared and used for the first time to fabricate wound dressing material. Sodium alginate (ALG) was used as reducing and stabilizing agents for preparation of silver nanoparticles (Ag-NPs). Effect of concentrations of alginate (ALG) on the particle size of silver were studied and confirmed by different techniques like UV/vis spectroscopy, transmission electron microscope (TEM) and dynamic light scattering (DLS). Nonwoven viscous fabrics were used as a carrier for silver/alginate/nanoparticles composite by impregnated the nonwoven fabrics as per the padding-curing technique. Nicotinamide (Nic) as anti-inflammatory drug was entrapped into Ag-NPS/ALG/nonwoven fabrics. Scanning electron microscope and energy dispersive x-ray (SEM-EDX) were used to evaluate the presence of Ag/ALG/Nic nanoparticles composite anchored the nonwoven fabrics. The antibacterial activity of the Ag/ALG/Nic wound dressing material was evaluated against Escherichia coli (E. coli) and Staphylococcus Aureus (St. Aureus). The wound healing and histological studied were evaluated by using burn diabetic rat animals.

Pycnodysostosis: clinical, radiologic, and endocrine evaluation and linear growth after growth hormone therapy.

Pycnodysostosis is a rare hereditary bone abnormality with an autosomal recessive mode of inheritance. We report the clinical, radiologic, and endocrine status of 8 children with this rare disease. All patients had the characteristic phenotype of the disorder including short stature (8 of 8), increased bone density (7 of 8), separated cranial sutures (8 of 8), large fontanel with delayed closure (8 of 8), obtuse mandibular angle (8 of 8), delayed teeth eruption (8 of 8), enamel hypoplasia (7 of 8), dysplastic acromial ends of the clavicles (6 of 8), frontal bossing (6 of 8), ocular proptosis (8 of 8), and dysplastic nails (8 of 8). Developmental evaluation according to the revised Denever developmental screening showed normal motor, fine motor-adaptive language, and personal social abilities in all the children. All had normal hepatic and renal functions. Serum calcium and phosphorus concentrations were normal. Two children had low serum alkaline phosphatase concentration. Short stature is a characteristic feature of pycnodysostosis. Seven of the 8 children were born short (length standard deviation score [SDS] = -3 to -1.5). Deceleration of linear growth was significant during the first 3 years of life. All the children had height SDS below -3 at the end of their third year of life. Although short stature is a feature of this genetic disorder, defective growth hormone (GH) secretion in response to provocation with clonidine and glucagon was found in 4 of the 8 patients. These 4 patients had pituitary hypoplasia on the magnetic resonance imaging (MRI) of their brain. In addition, 3 of these 4 patients had demyelination of the cerebrum. Patients with pycnodysostosis (n = 8) had low circulating concentrations of insulin-like growth factor-1 (IGF-1) compared with normal age-matched short children with constitutional short stature (CSS). IGF-I increased significantly after injecting GH for 3 days in these patients. Physiologic replacement with GH (18 U/m(2)/week) divided in daily evening doses subcutaneously increased IGF-1 concentration and improved linear growth velocity and height standard deviation scores (HtSDS) in the 4 children with GH deficiency. These data ruled out GH resistance and proved the usefulness of GH therapy in the management of short stature in these patients. In summary, some patients with pycnodysostosis have partial GH deficiency and low IGF-1 concentration. GH therapy markedly increases IGF-I secretion and improves their linear growth. MRI study of the brain including the hypothalamic-pituitary area is recommended in these children because of the high incidence of pituitary hypoplasia and cerebral demyelination.

Postantibiotic effect of roxithromycin on streptolysin O production, hydrophobicity, and bactericidal activity of PMNL by Streptococcus pyogenes.

Exposure of Streptococcus pyogenes to 5 x minimum inhibitory concentration of roxithromycin for 1 h produced a significant postantibiotic effect. More than 2.5 h was necessary for roxithromycin-treated bacteria to increase by 1 log10 in colony-forming units after drug removal, compared with the unexposed cells. After exposure to and removal of the drug, treated cells failed to exhibit normal hemolytic activity for at least 4 h. The inhibitory effect persisted for 20 h after drug removal, although the extent of growth for treated and untreated cells was almost the same. Hydrophobicity of treated cells, studied throughout the logarithmic growth phase with a water-hexadecan two-phase system, was markedly decreased by 40%, compared with untreated cells 4 h after drug removal. Cells that had been treated with roxithromycin became more susceptible to the bactericidal activity of human PMNL than untreated bacteria. The data indicate that some of the metabolic activity that contributes to the virulence of S. pyogenes is affected by postexposure to roxithromycin, and its minimum inhibitory concentration and serum level might not be the best indicators of efficacy in this class of drugs.

Post-antibiotic effect of azithromycin and erythromycin on streptococcal susceptibility to phagocytosis.

The effect of azithromycin and erythromycin on growth, cell surface hydrophobicity and the susceptibility to the bactericidal activity of human polymorphonuclear leucocytes (PMNL) was examined in four Streptococcus species. Exposure to either 10 x MIC azithromycin or erythromycin induced a post-antibiotic effect (PAE) of between 2.4 and 4.3 h. Erythromycin caused a longer PAE for S. sanguis than azithromycin under the same conditions. The cell surface charge (hydrophobic or hydrophilic) of the streptococci was altered significantly during PAE; loss of hydrophobicity was induced by both macrolides, and this effect was variable amongst the species. The decrease in hydrophobicity was not related to inhibition of growth. The effect of each drug during PAE on the interaction of opsonised suspensions of the streptococci with human PMNL revealed that erythromycin, and to a lesser extent azithromycin, increased susceptibility to the bactericidal activity of human PMNL; this effect was abolished following PAE. The present study clearly showed that PAE should not only be considered as delayed bacterial growth, but also as modulation of bacterial susceptibility to phagocytosis which may influence the outcome of the host-parasite relationship.

Comparative efficacy of successive exposure of Pseudomonas aeruginosa to gentamicin and ceftazidime.

Aminoglycoside and beta-lactam antibiotics, when used in combination, are usually given simultaneously, however, successive administration may be more efficient. The killing capacity was used to assess the effect of time intervals between low and high concentrations (2-8xMICs) of gentamicin and/or ceftazidime on Pseudomonas aeruginosa and to determine which drug is better to be administered first. The killing capacity after exposure to the antibiotic for 1 h were compared: (i) cells treated with gentamicin alone; (ii) cells treated with ceftazidime alone; and (iii) ceftazidime was added to (i) or (iv) gentamicin was added to (ii) at 0, 1 and 3 h of antibiotic removal. The bactericidal activity of gentamicin was potentiated and the viable cells decreased up to 6 h after antibiotic removal when the ceftazidime was added at O and at 1 h but the extent of bactericidal activity was reduced, when it was added at 3 h after gentamicin removal. Alternatively, treating the cells first with ceftazidime and then gentamicin was added after drug removal at O and at 1 h resulted in a marked decline in the viable cells, while addition of gentamicin after 3 h from ceftazidime removal, the extent of bactericidal activity was reduced. The non-treated cells with gentamicin started to grow heavily within 6 h of ceftazidime removal. No viable cells were detected after overnight incubation in cultures treated first with 6 or 8xMIC of gentamicin for 0.5 or 1 h. This in vitro study suggests that the optimum interval between gentamicin and ceftazidime doses, which gave the maximum bactericidal effect and the time before re-growth, appeared to be 1-2 h.

Minor phenolics from Crinum bulbispermum bulbs.

From the bulbs of Crinum bulbispermum Milne, four new minor compounds were isolated viz. 4-hydroxy-2',4'-dimethoxydihydrochalcone (1), 4,5-methylenedioxy-4'-hydroxy-2-aldehyde[1,1'-biphenyl] (4), hippacine (6), and 4'-hydroxy-7-methoxyflavan-3-ol (7). In addition, four known compounds were isolated and identified as 2(S),3',4'-dihydroxy-7-methoxy flavan (2), isolarrien (3), isoliquiritigenin (5) and liquiritigenin (8). The structures of the isolated compounds were established by spectral evidence.

Effect of surface geometry and morphic features on the flow characteristics of microsphere suspensions.

In this paper we present a study on the effect of surface ruggedness of microspheres on the rheological behavior of their suspensions. For this purpose, different types of ragweed pollen grains were selected as models of natural microspheres. A computer-image processing system based on Fourier and fractal analysis of the contour was used to quantitate the micromorphology and surface roughness. The viscosity of suspensions, prepared by the dispersion of the different types of microspheres in heavy liquid paraffin, was determined. It was found that an increase in surface roughness causes an increase in the viscosity of the suspension. Additional resistance to flow could be attributed to internal friction within the suspension due to an increase in the area of contact (during collision or aggregation) among the microspheres, and between microspheres and liquid environment. These findings suggest that the surface geometry of solid particles (e.g., microcapsules, beads, and microspheres) could have a significant effect on the performance of these microparticles in suspension.

Differential inhibition by clindamycin on slime formation, adherence to teflon catheters and hemolysin production by Staphylococcus epidermidis.

Slime formation was detected in Staphylococcus epidermidis strains isolated from either infected patients or healthy individuals. Cells of S. epidermidis, that either formed slime or not, adhered to teflon catheters. There was no correlation between adherence of bacteria to teflon catheters and slime formation. Clindamycin at subinhibitory concentration significantly inhibited slime formation without inhibiting bacterial growth. Adherence of S. epidermidis to teflon catheters was affected by the presence of clindamycin whether slime was produced or not. Clindamycin at subinhibitory concentrations markedly inhibited hemolysin production by S. epidermidis without appreciably altering the cell density, and cells grown in the presence of the drug showed very low hemolytic activity upon disruption. These results suggest that clindamycin at low concentration alters S. epidermidis virulence properties, apart from inhibiting growth.

Effect of beta-lactamase inhibitors on normal immune capabilities and their interactions with staphylococcal pathogenicity.

The effects of the beta-lactamase inhibitors, clavulanic acid, sulbactam and tazobactam on normal immune responses were investigated. These agents did not interfere with either humoral or cell-mediated immune responses as measured by the hemolytic plaque assay and delayed type hypersensitivity reaction assay respectively. In addition, human polymorphonuclear leukocyte phagocytic activity was not altered by these agents. When these agents were tested for their effect on Staphylococcus aureas adherence to buccal epithelial cells we found that all inhibitors suppressed staphylococcal adherence at therapeutic serum concentrations. Among the inhibitors investigated, sulbactam was found to significantly inhibit the hemolysin production of S. aureus. These data suggest that beta-lactamase inhibitors do not exhibit immunomodulating activity, but they interfere with some of the virulence factors of S. aureus. These findings suggest the advantage of preparations containing these inhibitors.

Preparation, characterization and antibacterial activity of chitosan-g-poly acrylonitrile/silver nanocomposite.

Chitosan-grafted-poly acrylonitrile silver nanocomposites (Cs-g-PAN/Ag) were prepared via in-situ chemical reduction of Ag ions in graft copolymerization of acrylonitrile onto chitosan. Graft copolymerization process was provided by FTIR and gravimetric methods. UV spectra and TEM images show silver nanoparticles with average 15-20nm dispersed homogeneously in CS-g-PAN/Ag nanocomposite-ray and TGA evident the change in crystallography and thermal stability in consequence of presence Ag nanoparticles. Cs-g-PAN/Ag nanocomposite showed excellent antimicrobial performance towards bacteria such as Escherichia coli and Staphylococcus aureus.