Identification and characterization of inulinases by bioinformatics analysis of bacterial glycoside hydrolases family 32 (GH32)

The glycoside hydrolase family contains enzymes that break the glycosidic bonds of carbohydrates by hydrolysis. Inulinase is one of the most important industrial enzymes in the family of Glycoside Hydrolases 32 (GH32). In this study, to identify and classify bacterial inulinases initially, 16,002 protein sequences belonging to the GH32 family were obtained using various databases. The inulin-effective enzymes (endoinulinase and exoinulinase) were identified. Eight endoinulinases (EC 3.2.1.7) and 4318 exoinulinases (EC 3.2.1.80) were found. Then, the localization of endoinulinase and exoinulinase enzymes in the cell was predicted. Among them, two extracellular endoinulinases and 1232 extracellular exoinulinases were found. The biochemical properties of 363 enzymes of the genus Arthrobacter, Bacillus, and Streptomyces (most abundant) showed that exoinulinases have an acid isoelectric point up to the neutral range due to their amino acid length. That is, the smaller the protein (336 aa), the more acidic the pI (4.39), and the larger the protein (1207 aa), the pI is in the neutral range (8.84). Also, a negative gravitational index indicates the hydrophilicity of exoinulinases. Finally, considering the biochemical properties affecting protein stability and post-translational changes studies, one enzyme for endoinulinase and 40 enzymes with desirable characteristics were selected to identify their enzyme production sources. To screen and isolate enzyme-containing strains, now with the expansion of databases and the development of bioinformatics tools, it is possible to classify, review and analyze a lot of data related to different enzyme-producing strains. Although, in laboratory studies, a maximum of 20 to 30 strains can be examined. Therefore, when more strains are examined, finally, strains with more stable and efficient enzymes were selected and introduced for laboratory activities. The findings of this study can help researchers to select the appropriate gene source from introduced strains for cloning and expression heterologous inulinase, or to extract native inulinase from introduced strains.     

Isolation and identification of Streptomyces sp. Act4Zk,a good producer of Staurosporine and some derivatives

In this study, strain Streptomyces sp. Act4Zk was isolated based on a method developed for the isolation of myxobacteria. Due to the low efficiency of the majority of conventional DNA extraction techniques, for molecular identification of the strain Streptomyces sp. Act4Zk, a new technique for DNA extraction of Actinobacteria was developed. In order to explore potential bioactivities of the strain, extracts of the fermented broth culture were prepared by an organic solvent (i.e. ethyl acetate) extraction method using. These ethyl acetate extracts were subjected to HPLC fractionation against standard micro-organisms, followed by LC/MS analysis. Based on morphological, physiological, biochemical and 16S rRNA gene sequence data, strain Streptomyces sp. Act4Zk is likely to be a new species of Streptomyces, close to Streptomyces genecies and Streptomyces roseolilacinus. Antimicrobial assay indicated high antifungal activity as well as antibacterial activity against Mycobacterium smegmatis and Gram-positive bacteria for the new strain. HPLC and LC/MS analyses of the extracts led to the identification of three different compounds and confirmed our hypothesis that the interesting species of the genus Streptomyces being a good producer of staurosporine and some derivatives.     

The assessment of the rice cultivars/lines resistance to blast disease in Mazandaran province, Iran

Blast, caused by Magnaporthe grisea, is one of the most important diseases in rice production regions of the world including Iran. To determine progress of rice blast disease on the selective cultivars and lines also to assay some components of partial resistance, a set of Iranian rice cultivars (Local and breeding) along with near-isogenic lines (NILs) and breeding lines from International Rice Research Institute (IRRI) were tested with some field races of the fungus in blast nursery and five of selective races in greenhouse. These experiments were conducted in a Randomized complete Block Design (RCBD) with three replications (except greenhouse experiment on the leaves). Traits in this study consisted of Infection Neck Number (INN), Neck Lesion Size (NLS), Infection Type (IT), percent Diseased Leaf Area (DLA) and Area Under Disease Progress Curve (AUDPC); also IT, Sporulation Lesion Number (SLN), Sporulating Region Diameter (SRD) and percent DLA were measured in leaf blast in greenhouse (one replication). The Iranian local cultivars and NILs i.e. Co-39 and C104-PKT located as susceptible group for AUDPC, IT, INN and NLS. Iranian breeding cultivars, breeding lines from IRRI and NILs (except Co-39 and C104-PKT) were resistant or indicated hypersensivity reaction (HR). Some cultivars (Fujiminori, Onda, and Hassan Saraii) were semi susceptible to leaf blast in nursery. The main point is correlation in 1% (a = 0.0001) between the traits in greenhouse and blast nursery. Neck node infection of Haraz cultivar in greenhouse experiment to IA-89 race is very important, because Haraz is a resistant cultivar to blast disease in Iran.     

Onychomycosis in Tehran,Iran: Prevailing fungi and treatment with itraconazole

A total of 187 Patients with suspected onychomycosis were examined for causative fungal agents between 1996 and 1997. Laboratory examination confirmed onychomycosis in 115 patients, of which 97 cases were presented with positive microscopic and cultural examinations, and they were selected for itraconazole pulse therapy. From an etiological point of view, 48.4% of the nail infections, mainly toenail infections, were caused by dermatophytes, 43.3% were infected with Candida spp, specially infected fingernails, and 8.2% by non-dermatophytic molds. Trichophyton mentagrophytes var. interdigital and T. violaceum were the most prevalent species. Candida albicans and C. parapsilosis were the predominant species of the Genus Candida. Scopolariopsis brevicaulis was the most common non-dermatophyte molds observed. Female affected more frequently than male and in both sexes, those who were 30–49 years old, more infected. Toenails were affected more frequently than fingernails. In this study, itraconazole pulse therapy (400 mg daily) gave during the first week of per month for 3 months. The study included 51 patients with toenail onychomychosis (group 1) and 46 patients with fingernail infections (group 2). Patients were followed up for 9 months after the last treatment. Clinical response rates were 83% in the group 1, 95% in the group 2 at month 12; the corresponding mycological cure rates were 71 and 87%, respectively. This revised version was published online in June 2006 with corrections to the Cover Date.     

Variation within Mycobacterium scrofulaceum

The purpose of this study was to identify Mycobacterium scrofulaceum reliably and rapidly and investigate diversity within the species. Fifty-four cultures were identified as Myco. scrofulaceum by preliminary cultural and biochemical tests, thin-layer chromatography and double diffusion. These strains were examined by PCR based on the 65 kDa heat stress protein gene, followed by restriction enzyme analysis of the product with Bst EII and Hae III. This produced seven groups, most with fewer fragments than had been reported previously. The technique was a rapid and reliable method for studying variation within Myco. scrofulaceum but alone, was unable to discriminate between some of these variants and other genetically similar species. When PCR-RFLP results were combined with biochemical tests, the major groups appeared to relate to different disease situations and thus, may have some epidemiological value.     

Cyclic Strain Alters the Expression and Release of Angiogenic Factors by Human Tendon Cells

Angiogenesis is associated with the tissue changes underlying chronic overuse tendinopathy. We hypothesized that repetitive, cyclic loading of human tendon cells would lead to increased expression and activity of angiogenic factors. We subjected isolated human tendon cells to overuse tensile loading using an in vitro model (1 Hz, 10% equibiaxial strain). We found that mechanically stimulated human tendon cells released factors that promoted in vitro proliferation and tube formation by human umbilical vein endothelial cells (HUVEC). In response to cyclic strain, there was a transient increase in the expression of several angiogenic genes including ANGPTL4, FGF-2, COX-2, SPHK1, TGF-alpha, VEGF-A and VEGF-C, with no change in anti-angiogenic genes (BAI1, SERPINF1, THBS1 and 2, TIMP1-3). Cyclic strain also resulted in the extracellular release of ANGPTL4 protein by tendon cells. Our study is the first report demonstrating the induction of ANGPTL4 mRNA and release of ANGPTL4 protein in response to cyclic strain. Tenocytes may contribute to the upregulation of angiogenesis during the development of overuse tendinopathy.     

Effect of nutritional supplements on bio-plastics (PHB) production utilizing sugar refinery waste with potential application in food packaging

Abstract

Polyhydroxyalkanoates (PHAs) are intracellular carbon and energy storage reserve material stored by gram-negative bacteria under nutrient limitation. PHAs are best alternative biodegradable plastics (bio-plastics) due to their resemblance to conventional synthetic plastic. The present study investigated the synergistic effect of nutritional supplements (amino acid and vitamin) on the PHA production by Alcaligenes sp. NCIM 5085 utilizing a sugar refinery waste (cane molasses) under submerged fermentation process. Initially, the effect of individual factor on PHA yield was studied by supplementing amino acids (cysteine, isoleucine, and methionine), vitamin (thiamin), and cane molasses at varying concentration in the production medium. Further, the cultivation medium was optimized by varying the levels of cane molasses, methionine and thiamin using response surface methodology to enhance the PHA yield. The maximum PHA yield of 70.89% was obtained under the optimized condition, which was then scaled up on 7.5?L-bioreactor. Batch cultivation in 7.5?L-bioreactor under the optimized condition gave a maximum PHA yield and productivity of 79.26% and 0.312 gL^?1 h^?1, respectively. The PHA produced was subsequently characterized as PHB by FTIR. PHB extracted was of relatively high molecular weight and crystallinity index. DSC analysis gave T_g, T_m, and X_c of 4.2, 179?°C and 66%, respectively. TGA analysis showed thermal stability with maximized degradation occurring at 302?°C, which is above the melting temperature (179?°C) of the purified polymer. The extracted polymer, therefore, possessed desirable material properties to be used in food packaging.     

Cerumen as a potential risk for transmission of Hepatitis B virus

Hepatitis B virus (HBV) transmission via blood and other body fluids from infected individuals to healthy people has been largely demonstrated. However, in the current literature, there is little information available on the potential role of cerumen in HBV transmission. Cerumen and blood were collected from 70 patients infected with HBV and 70 volunteer healthy people were selected as the control group, and the samples were evaluated by ELISA and Real-time PCR. All the patients proved positive for HBsAg and anti HBc total. Sixty-one of the 70 cerumen samples of cases (82.1%) and 5 (7%) of controls were positive for HBV DNA with ranges from 1.53 × 102 to 2.9 × 108 and 1.3 × 102-2.6 × 105/ml, respectively. In three patients, the level of HBV DNA in cerumen was higher than that in the serums. The patients who were positive for HBeAg showed a higher rate of HBVDNA in the serum and cerumen.The results of this study showed the level of HBV DNA as a probably indicator of high risk transmission factor, which was present in the cerumen of chronic hepatitis B patients in west of Iran.     

Application of Probiotics in Folate Bio-Fortification of Yoghurt

Folate deficiency is a public health concern affecting all age groups worldwide. The available evidence reveals that adding probiotic bacteria to the yoghurt starter cultures during yoghurt production process under fermentation conditions increases the folate content of yoghurt. The present study was conducted to measure two folate derivatives, i.e., 5-methyltetrahydrofolate and 5-formyltetrahydrofolate, in bio-fortified yoghurt samples including (1) yoghurt containing Streptococcus thermophilus and Lactobacillus bulgaricus, (2) probiotic yoghurt containing Lactobacillus acidophilus LA-5 and Bifidobacterium lactis BB-12, (3) probiotic yoghurt containing native strains of Lactobacillus plantarum 15HN, (4) probiotic yoghurt containing native strains of Lactococcus lactis 44Lac, and (5) probiotic yoghurt containing commercial strains of Lactobacillus plantarum LAT BY PL. During storage at 4 °C for 21 days, the highest levels of 5-methyltetrahydrofolate and 5-formyltetrahydrofolate, which were statistically significant, were detected in the yoghurt made using Lact. plantarum 15HN. Moreover, the highest total folate concentration (1487 ± 96.42 μg/L) was specified in the yoghurt containing Lact. plantarum 15HN on the 7th day. It can be conjectured that this product can be suggested as a proper alternative to synthetic folic acid and may not have the side effects of using synthetic folic acid overdoses.

     

Genetic structure and ecological niche segregation of Indian gray mongoose (Urva edwardsii) in Iran

Combining genetic data with ecological niche models is an effective approach for exploring climatic and nonclimatic environmental variables affecting spatial patterns of intraspecific genetic variation. Here, we adopted this combined approach to evaluate genetic structure and ecological niche of the Indian gray mongoose (Urva edwardsii) in Iran, as the most western part of the species range. Using mtDNA, we confirmed the presence of two highly differentiated clades. Then, we incorporated ensemble of small models (ESMs) using climatic and nonclimatic variables with genetic data to assess whether genetic differentiation among clades was coupled with their ecological niche. Climate niche divergence was also examined based on a principal component analysis on climatic factors only. The relative habitat suitability values predicted by the ESMs for both clades revealed their niche separation. Between‐clade climate only niche comparison revealed that climate space occupied by clades is similar to some extent, but the niches that they utilize differ between the distribution ranges of clades. We found that in the absence of evidence for recent genetic exchanges, distribution models suggest the species occurs in different niches and that there are apparent areas of disconnection across the species range. The estimated divergence time between the two Iranian clades (4.9 Mya) coincides with the uplifting of the Zagros Mountains during the Early Pliocene. The Zagros mountain‐building event seems to have prevented the distribution of U. edwardsii populations between the western and eastern parts of the mountains as a result of vicariance events. Our findings indicated that the two U. edwardsii genetic clades in Iran can be considered as two conservation units and can be utilized to develop habitat‐specific and climate change‐integrated management strategies.