Computational optimization of in vitro parameters for di-(2-ethylhexyl) phthalate production from Anabaena circinalis

Di-(2-ethylhexyl) phthalate (DEHP) is a ubiquitous environmental toxicant, and finds extensive commercial application as a plasticizer to reduce the rigidity of polyvinyl chloride. Besides numerous negative impacts on environment and public health, the compound exhibits acute bioactivity against microbes and has therapeutic value too. Considering this biochemical significance, searching of its new biogenic sources has become an active area of research. Here, DEHP is identified from the biomass of a toxic strain of Anabaena circinalis, which is quite unobvious, and simultaneously, the in vitro physical conditions are optimized by using a swarm-based multiparameter optimization technique. A purified fraction collected from column chromatography is subjected to gas chromatography (GC) to fetch the compound peak and then subsequent mass analysis for its identification. The mass spectrum describes the molecular weight along with the structure of DEHP with 99.9% experimental accuracy. An experimental observation table has been used to frame a fitness function using the curve fitting approach when temperature (T), light and dark period (LD), and duration of subculture cycle (DSC) are considered as the target parameters to be optimized. The optimum values obtained for T, LD, and DSC are 20°C, 14:10 hour light and dark ratio approximately, and 40 days, respectively. This experimental finding of A. circinalis FSS 124 as a novel source of DEHP and subsequent optimization using soft computing tools might be a benchmark for process optimization in biological research.     

Unique multiple paternity in the endangered big‐headed turtle (Platysternon megacephalum) in an ex situ population in South China

Understanding the mating system and reproductive strategies of an endangered species is critical to the success of captive breeding. The big‐headed turtle (Platysternon megacephalum) is one of the most threatened turtle species in the world. Captive breeding and reintroduction are necessary to re‐establish wild populations of P. megacephalum in some of its historical ranges in China, where the original populations have been extirpated. However, the captive breeding of P. megacephalum is very difficult and this may be due to its mysterious reproductive strategies and special behavior (e.g., aggressive temperament and territoriality). In this study, we achieved successful captive breeding of P. megacephalum by creating a habitat that mimics natural conditions and then investigated its mating system using microsatellite makers. A total of 16 clutches containing 79 eggs of P. megacephalum were collected, and 52 were hatched successfully over two breeding seasons. Of the 15 effective clutches, 6 clutches (40%) exhibited multiple paternity. There was no significant correlation between clutch size and multiple paternity, and no significant difference in hatching success between multiple‐sired and single‐sired clutches. However, there was significant correlation between male body size and the number of offspring, with higher‐ranked males contributing to more clutches. Our results provide the first evidence of multiple paternity and male hierarchy in P. megacephalum. These findings suggest that multiple paternity and male hierarchy should be considered in captive breeding programs for P. megacephalum, and creating a habitat that mimics natural conditions is an effctive way to achieve successful captive breeding and investigate the mating systems of this species.     

Morphological characterization and phylogenetic relationships of Indochinese box turtles—The Cuora galbinifrons complex

The members of the Indochinese box turtle complex, namely Cuora galbinifrons, Cuora bourreti, and Cuora picturata, rank the most critically endangered turtle species on earth after more than three decades of over‐harvesting for food, traditional Chinese medicine, and pet markets. Despite advances in molecular biology, species boundaries and phylogenetic relationships, the status of the C. galbinifrons complex remains unresolved due to the small number of specimens observed and collected in the field. In this study, we present analyses of morphologic characters as well as mitochondrial and nuclear DNA data to reconstruct the species boundaries and systematic relationships within the C. galbinifrons complex. Based on principal component analysis (PCA) and statistical analysis, we found that phenotypic traits partially overlapped among galbinifrons, bourreti, and picturata, and that galbinifrons and bourreti might be only subspecifically distinct. Moreover, we used the mitochondrial genome, COI, and nuclear gene Rag1 under the maximum likelihood criteria and Bayesian inference criteria to elucidate whether C. galbinifrons could be divided into three separate species or subspecies. We found strong support for a sister relationship between picturata and the other two species, and consequently, we recommend maintaining picturata as a full species, and classifying bourreti and galbinifrons as subspecies of C. galbinifrons. These findings provide evidence for a better understanding of the evolutionary histories of these critically endangered turtles.     

The red bayberry genome and genetic basis of sex determination

Morella rubra, red bayberry, is an economically important fruit tree in south China. Here, we assembled the first high‐quality genome for both a female and a male individual of red bayberry. The genome size was 313‐Mb, and 90% sequences were assembled into eight pseudo chromosome molecules, with 32 493 predicted genes. By whole‐genome comparison between the female and male and association analysis with sequences of bulked and individual DNA samples from female and male, a 59‐Kb region determining female was identified and located on distal end of pseudochromosome 8, which contains abundant transposable element and seven putative genes, four of them are related to sex floral development. This 59‐Kb female‐specific region was likely to be derived from duplication and rearrangement of paralogous genes and retained non‐recombinant in the female‐specific region. Sex‐specific molecular markers developed from candidate genes co‐segregated with sex in a genetically diverse female and male germplasm. We propose sex determination follow the ZW model of female heterogamety. The genome sequence of red bayberry provides a valuable resource for plant sex chromosome evolution and also provides important insights for molecular biology, genetics and modern breeding in Myricaceae family.     

Genetic diversity of Indian garlic core germplasm using agro-biochemical traits and SRAP markers

The characterization of garlic germplasm improves its utility, despite the fact that garlic hasn't been used much in the past. Garlic has an untapped genetic pool of immense economic and medicinal value in India. Hence, using heuristic core collection approach, a core set of 46 accessions were selected from 625 Indian garlic accessions based on 13 quantitative and five qualitative traits. The statistical measures (CV per cent, CR per cent, VR per cent) were used to sort the core set using Shannon-Wiener diversity index and the Nei diversity index. In addition, the variation within the core set was tested for 18 agro-morphological and six biochemical characteristics (allicin, phenol content, pyruvic acid, protein, allyl methyl thiosulfinate (AMTHS), and methyl allyl thiosulfinate (MATHS)). Further study of the core set's molecular diversity was performed using sequence related amplified polymorphism (SRAP) markers, which revealed a wide range of diversity among the core set's accessions, with an average polymorphism efficiency (PE) of 80.59 percent, polymorphism information content (PIC) of 0.29, effective multiplex ratio (EMR) of 3.51, and marker index (MI) of 0.99. The findings of this study will be useful in identifying high-yielding, elite garlic germplasm lines with the trait of interest. Since this core set is indicative of total germplasm, these selected breeding lines will be used for genetic improvement of garlic in the future.     

Lipase member H is a novel secreted protein selectively upregulated in human lung adenocarcinomas and bronchioloalveolar carcinomas

Lung cancer is one of the most frequent causes of cancer-related death worldwide. However, molecular markers for lung cancer have not been well established. To identify novel genes related to lung cancer development, we surveyed publicly available DNA microarray data on lung cancer tissues. We identified lipase member H (LIPH, also known as mPA-PLA1) as one of the significantly upregulated genes in lung adenocarcinoma. LIPH was expressed in several adenocarcinoma cell lines when they were analyzed by quantitative real-time polymerase chain reaction (qPCR), western blotting, and sandwich enzyme-linked immunosorbent assay (ELISA). Immunohistochemical analysis detected LIPH expression in most of the adenocarcinomas and bronchioloalveolar carcinomas tissue sections obtained from lung cancer patients. LIPH expression was also observed less frequently in the squamous lung cancer tissue samples. Furthermore, LIPH protein was upregulated in the serum of early- and late-phase lung cancer patients when they were analyzed by ELISA. Interestingly, high serum level of LIPH was correlated with better survival in early phase lung cancer patients after surgery. Thus, LIPH may be a novel molecular biomarker for lung cancer, especially for adenocarcinoma and bronchioloalveolar carcinoma.     

Distinction among North Atlantic cod Gadus morhua stocks by tissue fatty acid profiles

The fatty acid (FA) profiles of the white muscle and heart tissues of cod Gadus morhua from five locations, Faroe Bank, Faroe Plateau, North‐West Iceland, Norway–Barents Sea and Denmark–Skagerrak, were population dependent. The interregional differences of FAs were significantly dissimilar (P < 0·01) in most cases. By way of a rapid and simple analytical method, the stock dependence and harvest location of individual G. morhua were chemometrically determined by multivariate principal component analysis. The difference among the stocks was correlated with the average water temperature at the harvest locations. It thus appears that the tissue FA profile is a phenotypic trait that is partly temperature driven.     

Technical guide for genetic advancement of underdeveloped and intractable Clostridium

In recent years, the genus Clostridium has risen to the forefront of both medical biotechnology and industrial biotechnology owing to its potential in applications as diverse as anticancer therapy and production of commodity chemicals and biofuels. The prevalence of hyper-virulent strains of C. difficile within medical institutions has also led to a global epidemic that demands a more thorough understanding of clostridial genetics, physiology, and pathogenicity. Unfortunately, Clostridium suffers from a lack of sophisticated genetic tools and techniques which has hindered the biotechnological exploitation of this important bacterial genus. This review provides a comprehensive summary of biotechnological progress made in clostridial genetic tool development, while also aiming to serve as a technical guide for the advancement of underdeveloped clostridial strains, including recalcitrant species, novel environmental samples, and non-type strains. Relevant strain engineering techniques, from genome sequencing and establishment of a gene transfer methodology through to deployment of advanced genome editing procedures, are discussed in detail to provide a blueprint for future clostridial strain construction endeavors. It is expected that a more thorough and rounded-out genetic toolkit available for use in the clostridia will bring about the construction of superior bioprocessing strains and a more complete understanding of clostridial genetics, physiology, and pathogenicity.     

Comparing simple root phenotyping methods on a core set of rice genotypes

Interest in belowground plant growth is increasing, especially in relation to arguments that shallow‐rooted cultivars are efficient at exploiting soil phosphorus while deep‐rooted ones will access water at depth. However, methods for assessing roots in large numbers of plants are diverse and direct comparisons of methods are rare. Three methods for measuring root growth traits were evaluated for utility in discriminating rice cultivars: soil‐filled rhizotrons, hydroponics and soil‐filled pots whose bottom was sealed with a non‐woven fabric (a potential method for assessing root penetration ability). A set of 38 rice genotypes including the OryzaSNP set of 20 cultivars, additional parents of mapping populations and products of marker‐assisted selection for root QTLs were assessed. A novel method of image analysis for assessing rooting angles from rhizotron photographs was employed. The non‐woven fabric was the easiest yet least discriminatory method, while the rhizotron was highly discriminatory and allowed the most traits to be measured but required more than three times the labour of the other methods. The hydroponics was both easy and discriminatory, allowed temporal measurements, but is most likely to suffer from artefacts. Image analysis of rhizotrons compared favourably to manual methods for discriminating between cultivars. Previous observations that cultivars from the indica subpopulation have shallower rooting angles than aus or japonica cultivars were confirmed in the rhizotrons, and indica and temperate japonicas had lower maximum root lengths in rhizotrons and hydroponics. It is concluded that rhizotrons are the preferred method for root screening, particularly since root angles can be assessed.