Yeasts and yeast-like fungi in tap water and groundwater,and their transmission to household appliances

In the present study we describe the occurrence of fungi in 100 tap water and 16 groundwater samples from Slovenia. We used culture-dependent and culture-independent techniques. 28 fungal species belonging to 16 genera were isolated with selected culturing conditions, targeting human opportunistic yeasts and yeast-like fungi. Of special concern was the detection of Aureobasidium melanogenum, Exophiala dermatitidis, Rhinocladiella similis, Candida parapsilosis and Rhodotorula mucilaginosa. The DGGE analysis of ITS1 rDNA revealed from 6 to 16 bands hypothetically corresponding to different taxa, while pyrosequencing showed the presence of Aspergillus and Exophiala. According to the statistic machine learning methodology, the profile of fungi in water is determined by the concentration of calcium and magnesium ions and the presence of nitrate. Exophiala spp., C. parapsilosis and R. mucilaginosa are known as dominant contaminants of household appliances. It appears that they are transferred with water to dishwashers and washing machines, where they subsequently proliferate.     

Wheat-yellow pumpkin composite flour: Physico-functional,rheological, antioxidant potential and quality properties of pan and flat bread

The impact of yellow pumpkin powder (YP) substitution (5, 10, and 15%) on wheat flour's physico-functional, pasting, gel texture, and dough rheology was studied. Moreover, the nutritional, organoleptic properties and bioactivity of the composite pan and pita bread were evaluated. An improved water holding capacity was noticed for the blended flour than for control. The pasting parameters were declined significantly (p < 0.05) with increasing YP. Composite flours presented a softer gel texture in the presence of YP. Reduced water absorption, increased dough development time, and lower stability for combined flour dough. The bread with YP depicted increased protein, fat, fiber, and mineral contents, while a reduced volume and specific volume were noticed for pan bread. YP incorporated 5% and did not compromise pan and pita bread's color and overall acceptability. Additionally, composite bread depicted higher total phenolics with enhanced antioxidant activities at the higher substitution of YP.     

沙冬青AmNAC3转录因子基因在抗旱性和抗寒性中的功能鉴定

为了研究强抗逆植物沙冬青Am NAC3转录因子基因在抗旱性和抗寒性中的功能,首先利用半定量RT-PCR方法对该基因进行了表达分析。结果表明,在室内培养的沙冬青幼苗中,Am NAC3有一定量的基础表达,在干旱胁迫下其转录水平明显上调,而在低温胁迫下其表达上调较弱。然后利用5'RACE技术获得该基因的5'端序列及全长cDNA序列,并利用RT-PCR方法克隆到其全长编码区(846bp)。将编码区片段构建到植物表达载体上,利用农杆菌介导法获得转基因拟南芥。进一步分析表明,转基因拟南芥对于干旱和低温胁迫的抗性表型与野生型无明显差异,但其离体叶片的失水率和气孔开度均大于野生型。此外,转基因幼苗中气孔开闭相关基因ABI1和ABI2的表达量降低。这些结果表明,Am NAC3可能主要在响应干旱胁迫和调节气孔开闭及叶片保水性中发挥功能,而在抵抗低温胁迫中无明显作用。     

iQPR法处理水对SD鼠生物安全性的研究

iQPR技术处理污水是一项新型尖端的技术,此技术可以成功降低污水乃至受到污染的地下水中的各种污染指标。但是,iQPR技术处理污水尤其是地下水是否存在潜在的生物安全性问题有待于进一步研究。因此,为评估iQPR技术对生物安全性的影响,本研究首先分析了三种不同iQPR法处理水的水质成分;其次系统研究了iQPR水对SD鼠在个体水平、组织水平和病理形态学损伤的研究。研究表明:iQPR处理的水质成分较对照组普通饮用水好,在个体组织水平检测未见异常,尽管其中一组iQPR处理水造成了SD鼠的脾小体增大,但是可能的原因是水处理环节存在微生物污染现象,因此,初步认定此技术未造成SD大鼠的个体损伤。本研究为揭示iQPR处理的水对生物体的安全性评价提供一个理论依据。     

Have the cake and eat it: Optimizing nondestructive DNA metabarcoding of macroinvertebrate samples for freshwater biomonitoring

DNA metabarcoding can contribute to improving cost‐effectiveness and accuracy of biological assessments of aquatic ecosystems, but significant optimization and standardization efforts are still required to mainstream its application into biomonitoring programmes. In assessments based on freshwater macroinvertebrates, a key challenge is that DNA is often extracted from cleaned, sorted and homogenized bulk samples, which is time‐consuming and may be incompatible with sample preservation requirements of regulatory agencies. Here, we optimize and evaluate metabarcoding procedures based on DNA recovered from 96% ethanol used to preserve field samples and thus including potential PCR inhibitors and nontarget organisms. We sampled macroinvertebrates at five sites and subsampled the preservative ethanol at 1 to 14 days thereafter. DNA was extracted using column‐based enzymatic (TISSUE) or mechanic (SOIL) protocols, or with a new magnetic‐based enzymatic protocol (BEAD), and a 313‐bp COI fragment was amplified. Metabarcoding detected at least 200 macroinvertebrate taxa, including most taxa detected through morphology and for which there was a reference barcode. Better results were obtained with BEAD than SOIL or TISSUE, and with subsamples taken 7–14 than 1–7 days after sampling, in terms of DNA concentration and integrity, taxa diversity and matching between metabarcoding and morphology. Most variation in community composition was explained by differences among sites, with small but significant contributions of subsampling day and extraction method, and negligible contributions of extraction and PCR replication. Our methods enhance reliability of preservative ethanol as a potential source of DNA for macroinvertebrate metabarcoding, with a strong potential application in freshwater biomonitoring.     

The impact of binding site waters on the activity/selectivity trade-off of Janus kinase 2 (JAK2) inhibitors

Structure based optimization of B39, an indazole-based low micromolar JAK2 virtual screening hit is reported. Analysing the effect of certain modifications on the activity and selectivity of the analogues suggested that these parameters are influenced by water molecules available in the binding site. Simulation of water networks in combination with docking enabled us to identify the key waters and to optimize our primary hit into a low nanomolar JAK2 lead with promising selectivity over JAK1.     

Influence of age on the bioaccumulation of heavy metals in Apodemus sylvaticus at Merja Zerga lagoon,Morocco

The influence of age and sex on the bioaccumulation of heavy metals in Apodemus sylvaticus was studied in Merja Zerga lagoon in northern Morocco. Five trace metal elements (Zn, Pb, Cr, Cu and Fe) were quantitatively analyzed by Varian AA 240 atomic absorption spectroscopy with graphite furnace in three organs (Liver, Kidney and Heart) from animals of different age and sex. The maximum metal level of the analyzed samples was recorded in adults and was limited to 46.62 μg/g for Pb and 35.1 μg/g for Cu, while it reached 22.69 μg/g, 7.59 μg/g and 6.78 μg/g for Cr, Zn and Fe, respectively. Highly significant differences were found for bioaccumulation of heavy metals according to animal ages and no significant differences were observed between the two sexes among the studied animals. Our results revealed also the existence of a strong correlation (r > 0.65) between the majority of biometric parameters and the trace element concentrations. In general, we found that age is a critical factor in estimating the level of heavy metal pollution. Other characteristics such as habitat, feeding habits and anti-predator behavior of the species need to be studied.     

Spatiotemporal evaluation of trace elements in river water using multivariate methods

The multivariate statistical methods (cluster analysis and discriminant analysis [DA]) are used to evaluate the quality of water spatiotemporally in the Brahmani River basin. A total of 15 water quality indicators (pH, Chloramines, Sodium, Sulfate, Phosphate, Potassium, Boron, Fluoride, Cadmium, Copper, Lead, Nickel, Zinc, Iron, and Total Chromium) are collected for 13 years in two seasons (the dry and the wet season) from nine sampling stations. These stations are grouped into four clusters (Considerable [Co], Low Pollution [LP], Moderate Pollution [MP], and Extreme Pollution [(EP]) by hierarchical agglomerative cluster analysis based on the similarities of water quality characteristics. The nine important influencing discriminating water quality indicators are extracted by stepwise method of temporal DA,where as eight important influencing indicators for dry season and eight indicators (Potassium, Phosphate, Fluoride, Copper, Lead, Nickel, Iron, and Total Chromium) for wet season are found out by the spatial DA. The pollution status in the river basin was observed from overall trend in (Considerable) Co to (Extreme Pollution) EP regions. The study could be very useful in prioritizing the important sites for water resources and pollution management.     

Front Cover Image,Volume 116, Number 11, November 2019

CRISPR/Cas9-guided cytidine deaminase enables C:G to T:A base editing in bacterial genome without introduction of lethal double-stranded DNA break, supplement of foreign DNA template, or dependence on inefficient homologous recombination. However, limited by genome-targeting scope, editing window, and base transition capability, the application of base editing in metabolic engineering has not been explored. Herein, four Cas9 variants accepting different protospacer adjacent motif (PAM) sequences were used to increase the genome-targeting scope of bacterial base editing. After a comprehensive evaluation, we demonstrated that PAM requirement of bacterial base editing can be relaxed from NGG to NG using the Cas9 variants, providing 3.9-fold more target loci for gene inactivation in Corynebacterium glutamicum. Truncated or extended guide RNAs were employed to expand the canonical 5-bp editing window to 7-bp. Bacterial adenine base editing was also achieved with Cas9 fused to adenosine deaminase. With these updates, base editing can serve as an enabling tool for fast metabolic engineering. To demonstrate its potential, base editing was used to deregulate feedback inhibition of aspartokinase via amino acid substitution for lysine overproduction. Finally, a user-friendly online tool named gBIG was provided for designing guide RNAs for base editing-mediated inactivation of given genes in any given sequenced genome ( www.ibiodesign.net/gBIG ).