Thidiazuron (TDZ) induced organogenesis and clonal fidelity studies in <Emphasis Type="Italic">Haloxylon persicum</Emphasis> (Bunge ex Boiss &amp; Buhse): an endangered desert tree species

Haloxylon persicum (Bunge ex Boiss & Buhse), is one of the hardy woody desert shrubs, which is now endangered and/or nearing extinction. Urban landscape development and overgrazing are the major threats for the erosion of this important plant species. For conserving the species, it is critical to develop an efficient in vitro regeneration protocol for rapid multiplication of large number of regenerants. Leaf explants, cultured on Murashige and Skoog (MS) medium supplemented with different concentrations of thidiazuron (TDZ) (0.5, 1, 2 µM), showed significant difference in bud sprouting and adventitious shoot induction. The highest shoot bud formation was recorded on MS medium supplemented with 0.5 µM TDZ. Shoot tip necrosis (STN), observed after first subculture of shoot buds in same medium, increased in severity with subculture time. Application of calcium (4 mM) and boron (0.1 mM) in combination with kinetin (10 µM) in the subculture medium significantly reduced the intensity of STN. On an average eight shoots/explant were produced by alleviating this problem. ISSR marker analysis revealed monomorphic banding pattern between progenies and parents, indicating the true to type nature of the clones and its parents.     

Synthesis of sulfadiazinyl acyl/aryl thiourea derivatives as calf intestinal alkaline phosphatase inhibitors,pharmacokinetic properties,lead optimization,Lineweaver-Burk plot evaluation and binding analysis

To seek the new medicinal potential of sulfadiazine drug, the free amino group of sulfadiazine was exploited to obtain acyl/aryl thioureas using simple and straightforward protocol. Acyl/aryl thioureas are well recognized bioactive pharmacophore containing moieties. A new series (4a–4j) of sulfadiazine derived acyl/aryl thioureas was synthesized and characterized through spectroscopic and elemental analysis. The synthesized derivatives 4a–4j were subjected to calf intestinal alkaline phosphatase (CIAP) activity. The derivative 4a–4j showed better inhibition potential compared to standard monopotassium phosphate (MKP). The compound 4c exhibited higher potential in the series with IC₅₀ 0.251?±?0.012?µM (standard KH₂PO₄ 4.317?±?0.201?µM). Lineweaver-Burk plots revealed that most potent derivative 4c inhibition CIAP via mixed type pathway. Pharmacological investigations showed that synthesized compounds 4a–4j obey Lipinsk’s rule. ADMET parameters evaluation predicted that these molecule show significant lead like properties with minimum possible toxicity and can serve as templates in drug designing. The synthetic compounds show none mutagenic and irritant behavior. Molecular docking analysis showed that compound 4c interacts with Asp273, His317 and Arg166 amino acid residues.     

A genetics‐based approach confirms immune associations with life history across multiple populations of an aquatic vertebrate (Gasterosteus aculeatus)

Understanding how wild immune variation covaries with other traits can reveal how costs and trade‐offs shape immune evolution in the wild. Divergent life history strategies may increase or alleviate immune costs, helping shape immune variation in a consistent, testable way. Contrasting hypotheses suggest that shorter life histories may alleviate costs by offsetting them against increased mortality, or increase the effect of costs if immune responses are traded off against development or reproduction. We investigated the evolutionary relationship between life history and immune responses within an island radiation of three‐spined stickleback, with discrete populations of varying life histories and parasitism. We sampled two short‐lived, two long‐lived and an anadromous population using qPCR to quantify current immune profile and RAD‐seq data to study the distribution of immune variants within our assay genes and across the genome. Short‐lived populations exhibited significantly increased expression of all assay genes, which was accompanied by a strong association with population‐level variation in local alleles and divergence in a gene that may be involved in complement pathways. In addition, divergence around the eda gene in anadromous fish is likely associated with increased inflammation. A wider analysis of 15 populations across the island revealed that immune genes across the genome show evidence of having diverged alongside life history strategies. Parasitism and reproductive investment were also important sources of variation for expression, highlighting the caution required when assaying immune responses in the wild. These results provide strong, gene‐based support for current hypotheses linking life history and immune variation across multiple populations of a vertebrate model.     

On-farm monitoring of mouse-invasive Salmonella enterica serovar enteritidis and a model for its association with the production of contaminated eggs.

Mice (Mus musculus) captured in henhouses were assessed for the presence of salmonellae in spleens. Of 621 and 526 spleens cultured during the first and second years of collection, 25.0 and 17.9%, respectively, were positive for Salmonella enterica serovar Enteritidis. Contaminated eggs were cultured from nine houses during the first year of sampling, and for eight of these houses, serovar Enteritidis was recovered from the spleens of mice. Rank sum statistical analysis of positive mouse spleens indicated that three overlapping bacterial populations were present. This pattern of infection was repeated when lipopolysaccharide (LPS) variants were used to infect chicks, and the worst infections were associated with isolates producing high-molecular-weight (HMW) LPS. Mouse isolates were capable of producing unprecedented amounts of HMW LPS as indicated by compositional analysis of six isolates that swarmed across 2% agar, which is a type of bacterial migration dependent upon production of HMW LPS. It is suggested that serovar Enteritidis cultured from the spleens of mice caught on farms will detect strains that are enhanced in their ability to contaminate eggs, in part because they are able to produce HMW LPS.     

Gene cloning, sequencing and enzymatic properties of glutamate synthase from the hyperthermophilic archaeon Pyrococcus sp. KOD1

The gltA gene encoding a glutamate synthase (GOGAT) from the hyperthermophilic archaeon Pyrococcus sp. KOD1 was cloned as a 6.6 kb HindIII-BamHI fragment. Sequence analysis indicates that gltA encodes a 481- amino acid protein (53 269 Da). The deduced amino acid sequence of KOD1-GltA includes conserved regions that are found in the small subunits of bacterial GOGAT: two cysteine clusters, an adenylate-binding consensus sequence and an FAD-binding consensus sequence. However, no sequences homologous to the large subunit of bacterial GOGAT were found in the upstream or downstream regions. In order to examine whether GltA alone can act as a functional GOGAT, GltA was overexpressed in Escherichia coli BL21 (DE3) cells using an expression plasmid. GltA was purified to homogeneity and shown to be functional as a homotetramer of approximately 205 kDa, which is equivalent to the molecular weight of the native GOGAT from KOD1, thus indicating that KOD1-GOGAT is the smallest known active GOGAT. GltA is capable of both glutamine-dependent and ammonia-dependent synthesis of glutamate. Synthesis of glutamate by KOD1-GltA required NADPH, indicating that this enzyme is an NADPH-GOGAT (EC 1.4.1.13). The optimum pH for both activities was 6.5. However, GltA exhibited different optimum temperatures for activity depending on the reaction assayed (glutamine-dependent reaction, 80° C; ammonia-dependent reaction, 90° C). Received: 30 October 1996 / Accepted: 13 January 1997     

A PCR-based method for mating type determination inCochliobolus heterostrophus

A rapid and accurate method based on multiplex PCR using three different primers was developed for determining mating type inCochliobolus heterostrophus. The primers of MAT113 and MAT123 were uniquely derived fromMAT1-1 andMAT1-2, respectively, and the primer MATcon5 is conserved in the flanking regions of the idiomorphs. The amplification product was estimated to be 702 bp (MAT1-1) or 547 bp (MAT1-2). Crossing experiments confirmed the accuracy of this method, which requires less time than the conventional determination of mating type.     

Thermostabilization of carboxymethylcellulase from Aspergillus niger by carboxyl group modification

The carboxyl groups of purified carboxymethylcellulase (CMCase) from Aspergillus niger NIAB280 were modified by 1-ethyl-3(3-dimethylaminopropyl) carbodiimide (EDC) in the presence of glycinamide for 15 min (GAM15) and glycinamide plus cellobiose for 75 min (GAM75). The half-lives of GAM15 at different temperatures were significantly enhanced whereas those of GAM75 were reduced as compared with the native CMCase. The activation energies of denaturation of native, GAM15 and GAM75 were 40, 35 and 59kJ mol respectively. Native CMCase and GAM15 showed no compensation effect, whereas native and GAM75 gave temperature of compensation of 44¡C. Gibb's free energy of activation for denaturation (DG*) of GAM15 was increased as compared with native CMCase. Surprisingly the entropies (DS*) of activation for denaturation were negative for native and GAM75 and decreased further for GAM15 between the temperature range of 45 to 65¡C. A possible explanation for the thermal inactivation of native and increased thermal stability of GAM15 is also discussed.