Generation and exploration of new classes of antitubercular agents: The optimization of oxazolines, oxazoles, thiazolines, thiazoles to imidazo[1,2-a]pyridines and isomeric 5,6-fused scaffolds

Tuberculosis (TB) is a devastating disease resulting in a death every 20s. Thus, new drugs are urgently needed. Herein we report ten classes of compounds-oxazoline, oxazole, thiazoline, thiazole, pyrazole, pyridine, isoxazole, imidazo[1,2-a]pyridine, imidazo[1,2-a]pyrimidine and imidazo[1,2-c]pyrimidine-which have good (micromolar) to excellent (sub-micromolar) antitubercular potency. The 5,6-fused heteroaromatic compounds were the most potent with MIC's as low as <0.195 μM (9 and 11). Overall, the imidazo[1,2-a]pyridine class was determined to be most promising, with potency similar to isoniazid and PA-824 against replicating Mtb H(37)Rv, clinically relevant drug sensitive, multi- and extensively resistant Mtb strains as well as having good in vitro metabolic stability.     

Solid-phase synthesis of 2'-hydroxychalcones. Effects on cell growth inhibition, cell cycle and apoptosis of human tumor cell lines

Thirty-one 2'-hydroxychalcones were prepared via solid-phase synthesis by base-catalyzed aldol condensation of substituted 2'-hydroxyacetophenones and benzaldehydes. Chalcones were tested for their growth inhibitory activity in three human tumor cell lines (MCF-7, NCI-H460 and A375-C5) using the SRB assay. Results revealed that several of the tested compounds caused a pronounced dose-dependent growth inhibitory effect on the tumor cell lines studied in the low micromolar range. To gain further insight on the cellular mechanism of action of this class of compounds, studies of their effect on cell cycle profile as well as on induction of cellular apoptosis were also carried out. Generally, the tested chalcones interfered with the cell cycle profile and increased the percentage of apoptotic MCF-7 cells. The results here presented may help to identify new chalcone-like structures with optimized cell growth inhibitory activity which may be further tested as potential antitumor agents.     

The significance of D-amino acids in soil, fate and utilization by microbes and plants: review and identification of knowledge gaps

Background

D-amino acids are far less abundant in nature than L-amino acids. Both L- and D-amino acids enter soil from different sources including plant, animal and microbial biomass, antibiotics, faeces and synthetic insecticides. Moreover, D-amino acids appear in soil due to abiotic or biotic racemization of L-amino acids. Both L- and D-amino acids occur as bound in soil organic matter and as “free“ amino acids dissolved in soil solution or exchangeably bound to soil colloids. D-amino acids are mineralized at slower rates compared to the corresponding L-enantiomers. Plants have a capacity to directly take up “free“ D-amino acids by their roots but their ability to utilize them is low and thus D-amino acids inhibit plant growth.

Scope

The aim of this work is to review current knowledge on D-amino acids in soil and their utilization by soil microorganisms and plants, and to identify critical knowledge gaps and directions for future research.

Conclusion

Assessment of “free“ D-amino acids in soils is currently complicated due to the lack of appropriate extraction procedures. This information is necessary for consequent experimental determination of their significance for crop production and growth of plants in different types of managed and unmanaged ecosystems. Hypotheses on occurrence of “free“ D-amino acids in soil are presented in this review.     

Relationship between Enterococcal Levels and Sediment Biofilms at Recreational Beaches in South Florida

Enterococci, recommended at the U.S. federal level for monitoring water quality at marine recreational beaches, have been found to reside and grow within beach sands. However, the environmental and ecological factors affecting enterococcal persistence remain poorly understood, making it difficult to determine levels of fecal pollution and assess human health risks. Here we document the presence of enterococci associated with beach sediment biofilms at eight south Florida recreational beaches. Enterococcal levels were highest in supratidal sands, where they displayed a nonlinear, unimodal relationship with extracellular polymeric secretions (EPS), the primary component of biofilms. Enterococcal levels peaked at intermediate levels of EPS, suggesting that biofilms may promote the survival of enterococci but also inhibit enterococci as the biofilm develops within beach sands. Analysis of bacterial community profiles determined by terminal restriction fragment length polymorphisms showed the bacterial communities of supratidal sediments to be significantly different from intertidal and subtidal communities; however, no differences were observed in bacterial community compositions associated with different EPS concentrations. Our results suggest that supratidal sands are a microbiologically unique environment favorable for the incorporation and persistence of enterococci within beach sediment biofilms.     

Intraradical dynamics of two coexisting isolates of the arbuscular mycorrhizal fungus Glomus intraradices sensu lato as estimated by real-time PCR of mitochondrial DNA

Real-time PCR in nuclear ribosomal DNA (nrDNA) is becoming a well-established tool for the quantification of arbuscular mycorrhizal (AM) fungi, but this genomic region does not allow the specific amplification of closely related genotypes. The large subunit of mitochondrial DNA (mtDNA) has a higher-resolution power, but mtDNA-based quantification has not been previously explored in AM fungi. We applied real-time PCR assays targeting the large subunit of mtDNA to monitor the DNA dynamics of two isolates of Glomus intraradices sensu lato coexisting in the roots of medic (Medicago sativa). The mtDNA-based quantification was compared to quantification in nrDNA. The ratio of copy numbers determined by the nrDNA- and mtDNA-based assays consistently differed between the two isolates. Within an isolate, copy numbers of the nuclear and the mitochondrial genes were closely correlated. The two quantification approaches revealed similar trends in the dynamics of both isolates, depending on whether they were inoculated alone or together. After 12 weeks of cultivation, competition between the two isolates was observed as a decrease in the mtDNA copy numbers of one of them. The coexistence of two closely related isolates, which cannot be discriminated by nrDNA-based assays, was thus identified as a factor influencing the dynamics of AM fungal DNA in roots. Taken together, the results of this study show that real-time PCR assays targeted to the large subunit of mtDNA may become useful tools for the study of coexisting AM fungi.     

Synergist effects of n-acetylcysteine and deferoxamine treatment on behavioral and oxidative parameters induced by chronic mild stress in rats

A growing body of evidence has pointed to a relationship between oxidative stress and depression. Thus, the present study was aimed at evaluating the effects of the antioxidants n-acetylcysteine (NAC), deferoxamine (DFX) or their combination on sweet food consumption and oxidative stress parameters in rats submitted to 40 days of exposure to chronic mild stress (CMS). Our results showed that in stressed rats treated with saline, there was a decrease in sweet food intake and treatment with NAC or NAC in combination with DFX reversed this effect. Treatment with NAC and DFX decreased the oxidative damage, which include superoxide and TBARS production in submitochondrial particles, and also thiobarbituric acid reactive substances (TBARS) levels and carbonyl proteins in the prefrontal cortex, amygdala and hippocampus. Treatment with NAC and DFX also increased the activity of the antioxidant enzymes, superoxide dismutase and catalase in the same brain areas. Even so, a combined treatment with NAC and DFX produced a stronger increase of antioxidant activities in the prefrontal cortex, amygdala and hippocampus. The results described here indicate that co-administration may induce a more pronounced antidepressant activity than each treatment alone. In conclusion, these results suggests that treatment with NAC or DFX alone or in combination on oxidative stress parameters could have positive effects against neuronal damage caused by oxidative stress in major depressive disorders.     

Sarcosin (Krp1) in skeletal muscle differentiation: gene expression profiling and knockdown experiments

SARCOSIN, also named Krp1, has been identified as a protein exclusively expressed in striated muscle tissue. Here we report on the role of SARCOSIN in skeletal muscle development and differentiation. We demonstrate, by means of whole-mount in situ hybridization, that Sarcosin mRNA is expressed in the myotome part of the mature somites in mouse embryos from embryonic day 9.5 onwards. Sarcosin is not expressed in the developing heart at these embryonic stages, and in adult tissues the mRNA expression levels are five times lower in the heart than in skeletal muscle. SARCOSIN protein partially co-localizes with the M-band protein myomesin and between and below laterally fusing myofibrils in adult skeletal muscle tissue. RNA interference mediated knock-down of SARCOSIN in the C2C12 myoblast cell line appeared to be stimulatory in the early phase of differentiation, but inhibitory at a later phase of differentiation.     

Transient expression of <Emphasis Type="Italic">Human papillomavirus</Emphasis> type 16 L2 epitope fused to N- and C-terminus of coat protein of <Emphasis Type="Italic">Potato virus X</Emphasis> in plants

Transient expression of foreign genes based on plant viral vectors is a suitable system for the production of relevant immunogens that can be used for the development of a new generation of vaccines against a variety of infectious diseases. In the present study the epitope derived from HPV-16 L2 minor capsid protein (amino acids 108–120) was expressed from Potato virus X (PVX)-based vector pGR106 as N- or C-terminal fusion with the PVX coat protein (PVX CP) in transgenic Nicotiana benthamiana plants. The fusion protein L2_108-120-PVX CP was successfully expressed in plants at a level of 170 mg/kg of fresh leaf tissue. The C-terminal fusion protein PVX CP- L2_108-120 was expressed using mutated vector sequence to avoid homologous recombination at a level of 8 mg/kg of fresh leaf tissue. Immunogenicity of L2_108-120-PVX CP virus-like particles was tested after immunization of mice by subcutaneous injection or tattoo administration. In animal sera the antibodies against the PVX CP and the L2_108-120 epitope were found after both methods of vaccine delivery.     

Ubiquitous urease affects soybean susceptibility to fungi

The soybean ubiquitous urease (encoded by GmEu4) is responsible for recycling metabolically derived urea. Additional biological roles have been demonstrated for plant ureases, notably in toxicity to other organisms. However, urease enzymatic activity is not related to its toxicity. The role of GmEu4 in soybean susceptibility to fungi was investigated in this study. A differential expression pattern of GmEu4 was observed in susceptible and resistant genotypes of soybeans over the course of a Phakopsora pachyrhizi infection, especially 24 h after infection. Twenty-nine adult, transgenic soybean plants, representing six independently transformed lines, were obtained. Although the initial aim of this study was to overexpress GmEu4, the transgenic plants exhibited GmEu4 co-suppression and decreased ureolytic activity. The growth of Rhizoctonia solani, Phomopsis sp., and Penicillium herguei in media containing a crude protein extract from either transgenic or non-transgenic leaves was evaluated. The fungal growth was higher in the protein extracts from transgenic urease-deprived plants than in extracts from non-transgenic controls. When infected by P. pachyrhizi uredospores, detached leaves of urease-deprived plants developed a significantly higher number of lesions, pustules and erupted pustules than leaves of non-transgenic plants containing normal levels of the enzyme. The results of the present work show that the soybean plants were more susceptible to fungi in the absence of urease. It was not possible to overexpress active GmEu4. For future work, overexpression of urease fungitoxic peptides could be attempted as an alternative approach.