Highly efficient soluble expression and purification of recombinant human basic fibroblast growth factor (hbFGF) by fusion with a new collagen-like protein (Scl2) in Escherichia coli

Abstract

Human basic fibroblast growth factor (hbFGF) is involved in a wide range of biological activities that affect the growth, differentiation, and migration. Due to its wound healing effects and therapy, hbFGF has the potential as therapeutic agent. Therefore, large-scale production of biologically active recombinant hbFGF with low cost is highly desirable. However, the complex structure of hbFGF hinders its high-level expression as the soluble and functional form. In the present study, an efficient, cost-effective, and scalable method for producing recombinant hbFGF was developed. The modified collagen-like protein (Scl2-M) from Streptococcus pyogenes was used as the fusion tag for producing recombinant hbFGF for the first time. After optimization, the expression level of Scl2-M-hbFGF reached approximately 0.85?g/L in the shake flask and 7.7?g/L in a high cell-density fermenter using glycerol as a carbon source. Then, the recombinant Scl2-M-hbFGF was readily purified using one-step acid precipitation and the purified Scl2-M-hbFGF was digested with enterokinase. The digested mixture was further subject to ion-exchange chromatography, and the final high-purity (96%) hbFGF product was prepared by freeze-drying. The recovery rate of the whole purification process attained 55.0%. In addition, the biological activity of recombinant hbFGF was confirmed by using L929 and BALB/c3T3 fibroblasts. Overall, this method has the potential for large scale production of recombinant hbFGF.     

Microbial diversity from the continental shelf regions of the Eastern Arabian Sea: A metagenomic approach

The marine microbiome is a complex and least-understood habitat, which play a significant role in global biogeochemical cycles. The present study reported the culture-independent assessment of microbial diversity from the Arabian Sea (AS) sediments (from Gujarat to Malabar; at 30 m depth) by using metagenome sequence analysis. Our results elucidated that bacterial communities in the Malabar coastal region are highly diverse than the Gujarat coast. Moreover, Statistical analysis (Spearman rank correlation) showed a significant correlation co-efficient value (r = P < 0.001) between microbial communities and physicochemical parameters (salinity and dissolved oxygen) in the water column. A total of 39 bacterial phyla were recorded from the eastern side of AS, of which six phyla Proteobacteria, Bacteroidetes, Actinobacteria, Cyanobacteria, Firmicutes, and Planctomycetes were found to be the most dominant group. The most dominant genus from Valapad region (Malabar Coast) was found to be Halomonas sp., while other regions were dominated with Psychrobacter pulmonis. The subsequent Principal Coordinate Analysis (PCoA) showed 99.53% variance, which suggests that, highly distinct microbial communities at Valapad (Malabar Coast) sampling location than other sites. Moreover, the microbial metabolic activity analysis revealed the important functions of microbial communities in the AS are hydrocarbon degradation, polymer degradation, nutrient oxidation and sulphate reduction (biodegradation process). Further extended studies are needed to be carried out for better understanding the functional diversity of microbial communities from the marine sediments.     

The hybrid bacterial foraging algorithm based on many-objective optimizer

A new multi-objective optimized bacterial foraging algorithm - Hybrid Multi-Objective Optimized Bacterial Foraging Algorithm (HMOBFA) is presented in this article. The proposed algorithm combines the crossover-archives strategy and the life-cycle optimization strategy, look for the best method through research area. The crossover-archive strategy with an external archive and internal archive is assigned to different selection principles to focus on diversity and convergence separately. Additionally, according to the local landscape to satisfy population diversity and variability as well as avoiding redundant local searches, individuals can switch their states periodically throughout the colony lifecycle with the life-cycle optimization strategy. all of which may perform significantly well. The performance of the algorithm was examined with several standard criterion functions and compared with other classical multi-objective majorization methods. The examiner results show that the HMOBFA algorithm can achieve a significant enhancement in performance compare with other method and handles many-objective issues with solid complexity, convergence as well as diversity. The HMOBFA algorithm has been proven to be an excellent alternative to past methods for solving the improvement of many-objective problems.     

BacStalk: A comprehensive and interactive image analysis software tool for bacterial cell biology

Prokaryotic cells display a striking subcellular organization. Studies of the underlying mechanisms in different species have greatly enhanced our understanding of the morphological and physiological adaptation of bacteria to different environmental niches. The image analysis software tool BacStalk is designed to extract comprehensive quantitative information from the images of morphologically complex bacteria with stalks, flagella, or other appendages. The resulting data can be visualized in interactive demographs, kymographs, cell lineage plots, and scatter plots to enable fast and thorough data analysis and representation. Notably, BacStalk can generate demographs and kymographs that display fluorescence signals within the two-dimensional cellular outlines, to accurately represent their subcellular location. Beyond organisms with visible appendages, BacStalk is also suitable for established, non-stalked model organisms with common or uncommon cell shapes. BacStalk, therefore, contributes to the advancement of prokaryotic cell biology and physiology, as it widens the spectrum of easily accessible model organisms and enables highly intuitive and interactive data analysis and visualization.     

Root-Associated Endophytic Bacterial Community Composition of Asparagus officinalis of Three Different Varieties

Asparagus (Asparagus officinalis L) is an economically important crop, rich in nutrients, and is also conducive to solving ecological and environmental problems. Plants may acquire benefits from root-associated endophytic bacteria. However, the composition of the endophytic bacterial community associated with the roots of asparagus is poorly elucidated. In this study, the nine root samples of asparagus from three different varieties including Asparagus officinalis var. Grande (GLD), A. officinalis var. Jinglvlu3 (JL3) and A. officinalis var. Jingzilu2 (JZL) were investigated by high-throughput sequencing technology of the 16S rDNA V5-V7 hypervariable region of endophytic bacteria. A total of 16 phyla, 29 classes, 90 orders, 171 families, and 312 genera were identified. Endophytic bacteria diversity and bacteria structure was different among the three varieties and was influenced by rhizosphere soil properties and varieties. In the GLD variety, the main phyla were Proteobacteria, Actinobacteria, and Firmicutes. The main phylum in JL3 and JZL varieties was Proteobacteria. The observations showed that GLD had the highest diversity of endophytes as indicated by the Shannon index (GLD > JZL > JL3). The order of the endophytes richness was GLD > JL3 > JZL. The PCA and PCoA analysis revealed the microbial communities were different between three different asparagus varieties, and the microbial composition of GLD and JZL was more similar. This report provides an important reference for the study of endophytic microorganisms of asparagus. Supplementary informationThe online version contains supplementary material available at (10.1007/s12088-021-00926-6) contains supplementary material, which is available to authorized users.     

利用细菌人工染色体技术构建整合F基因的重组MDV疫苗株*

目的:预防马立克氏病病毒(MDV)和新城疫病毒(NDV)混合感染鸡引起的疾病,构建表达NDV F蛋白的MDV疫苗株CVI988 BAC重组载体,并包装成重组病毒,为疫苗免疫提供更多的重组疫苗选择。方法:首先利用PCR扩增带有卡那霉素(Kanamycin,Kana)抗性基因片段的F基因,采用同源重组的方法将其整合到CVI988 BAC上,进一步诱导I-SceI表达敲除Kana基因而获得重组质粒CVI988 BAC-F。通过磷酸钙法转染鸡胚成纤维细胞获得重组病毒。结果:Western blot和间接免疫荧光实验证实重组病毒能够表达F蛋白。病毒生长曲线和蚀斑大小测定结果表明,F基因的插入不影响病毒的体外增殖。结论:利用BAC技术成功构建了整合F基因的重组MDV病毒CVI988 BAC-F,为MDV重组疫苗研发,防控NDV与MDV共感染奠定了基础。     

阴道加德纳菌检出率及唾液酸酶A基因携带与细菌性阴道病的关系

目的探究阴道加德纳菌(Gardnerella vaginalis)检出率及唾液酸酶A基因携带与细菌性阴道病(BV)的关系。方法选择2017年1月至2019年8月确诊的BV患者82例作为BV组,并随机选择同时期健康女性82例作为健康组,比较2组人群G. vaginalis检出率和唾液酸酶A基因携带情况,相关统计学资料分析其对BV发生的影响。结果BV组人群G. vaginalis阳性检出率高于健康组(χ²=11.511,P<0.05)。BV组人群共检出G. vaginalis 1、2、3、4、5、6和7型,其中2型占比最高;BV组人群G. vaginalis 2、3、4型占比率高于健康组(χ²=4.148,17.009,9.973,均P<0.05)。BV组人群唾液酸酶A基因携带率高于健康组(χ²=39.234,P<0.05)。较健康组人群,BV组PCR DGGE宽度更窄,肠道菌群条带数少(t=9.217,P<0.05)。结论G. vaginalis检出率和唾液酸酶A基因携带情况与BV发生相关,有待成为相关生物学治疗靶点。     

Population dynamics,hunting nature on insect pests and existence of symbiotic bacterial microbes among leading transgenic cotton spiders

The genetically modified cotton holding Bt proteins, is noxious to bollworms larvae but very little is known about its impact on population of non-target spiders within the field. Studies conducted with the aim to identify spiders along with their abundance, prevalence pattern, preying habit on insect pests in Bt cotton (L280, Cry2Ab4) and commonness of symbiotic bacteria in these spiders. Spiders collections were made via vial tapping/jarring, from May to end of October (2018). In total, 13,342 spiders were recorded during the entire cropping season. Least relative abundance was in May (0.28%), highest in August (30.39%) while ahead decline was observed, reached to 12% in October. The 27 species were verified, least richness was in May (3 species), high in August (19 species), later in the end, reduced to 7 species. High abundance was of Hylyphantes graminicola (68.56%) and Neoscona theisi (19.98%). Existence of H. graminicola and Pardosa astrigera observed during the whole cropping season. Presence of seven guilds among spiders had a diverse nature of hunting insect pests and on number of species existed in the crop, maximum was from direct hunting habit. Composition and community structure of microbiota varied as of spider species. Proteobacteria was the most abundant bacterial phylum and topmost genus was Wolbachia in observed spiders. Five groups of bacteria distinguished across 4 clusters of spiders depending on core community of symbiotic bacterial genera. This effort is an initial step to get knowhow about spiders in Bt cotton, which will facilitate future research studies concerning spiders.     

Phytochemical screening and anti-oxidant activity of Sargassum wightii enhances the anti-bacterial activity against Pseudomonas aeruginosa

In this study, the phytochemical, phenolic, flavonoid and bioactive compounds were successfully screened from crude extract of Sargassum wightii by LC-MS analysis after NIST interpretation. Bacterial growth inhibition study result was shown with 24 mm zone inhibition at 200 µg/mL concentration against P. aeruginosa. The increased phenolic content was much closed to gallic acid and the range was observed at 250 μg/mL concentration. In addition, flavonoid contents of the algae extract was indicated more significant with rutin at 200 μg/mL. In result, both the phenolic and flavonoid contents of the extract were more correlated with gallic acid and rutin. Further, the total anti-oxidant and DPPH radical scavenging activities were shown increased activity at 200 μg/mL concentrations. Furthermore, the excellent anti-bacterial alteration result was observed at 200 μg/mL concentration by minimum inhibition concentration. Therefore, the result was revealed that the marine algae Sargassum wightii has excellent phytochemical and anti-oxidant activities, and it has improved anti-bacterial activity against P. aeruginosa.     

Honest signals and sexual conflict: Female lizards carry undesirable indicators of quality

Sex differences in animal coloration often result from sex‐dependent regulatory mechanisms. Still, some species exhibit incomplete sexual dimorphism as females carry a rudimentary version of a costly male trait, leading to intralocus sexual conflict. The underlying physiology and condition dependence of these traits can inform why such conflicts remain unresolved. In eastern fence lizards (Sceloporus undulatus), blue iridophore badges are found in males and females, but melanin pigmentation underneath and surrounding badges is male‐exclusive. We track color saturation and area of badges across sexual maturity, and their relationship to individual quality (body condition and immunocompetence) and relevant hormones (testosterone and corticosterone). Saturation and testosterone were positively correlated in both sexes, but hormone and trait had little overlap between males and females. Saturation was correlated with body condition and immunocompetence in males but not in females. Co‐regulation by androgens may have released females from resource allocation costs of color saturation, even when in high condition. Badge area was independent of testosterone, but associated with low corticosterone in females, indicating that a nonsex hormone underlies incomplete sexual dimorphism. Given the evidence in this species for female reproductive costs associated with ornamentation, this sex‐nonspecific regulation of an honest signal may underlie intralocus sexual conflict.