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The razor clam Sinonovacula constricta is the first marine mollusk demonstrated to possess the complete long-chain polyunsaturated fatty acids (LC-PUFA) biosynthetic pathway. This study explored the impact of different microalgae on growth, Fad and Elovl expressions, and fatty acid (FA) profiles in juvenile S. constricta. Results revealed that juveniles fed with Isochrysis galbana (rich in DHA) or Chaetoceros calcitrans (rich in EPA) consistently exhibited higher growth than those fed Chlorella sp. (rich in LA and ALA), underscoring the importance of dietary LC-PUFA in S. constricta's development. Expression of most Fad and Elovl in C. calcitrans and I. galbana-fed juveniles were initially up-regulated, then down-regulated, suggesting LC-PUFA demand for faster growth. Although Chlorella sp.-fed juveniles exhibited decreased mRNA levels for most genes, levels were notably higher lately compared to those fed C. calcitrans or I. galbana, hinting at potential LC-PUFA biosynthesis induction. FA profiles in S. constricta generally mirrored those in ingested microalgae, implying direct FA accumulation from diets. Some microalgal FA were absent in farmed S. constricta, while others emerged, indicating S. constricta's ability to selectively accumulate and synthesize FA. This study enhances the understanding of dietary FA metabolism in S. constricta, valuable for selecting appropriate microalgae in its farming practices. [ABSTRACT FROM AUTHOR]

Meleagrin and oxaline, which belong to the roquefortine alkaloids with a unique dihydroindole spiroamide framework, have significant bioactivities, especially tumor cell inhibitory activity. In order to discover the requefortine alkaloids, Penicillium sp. OUCMDZ-1435 was fished and identified from marine fungi using molecular probe technology. Meleagrin (1) and oxaline (2) were isolated from it. In addition, we first reported that compounds 1 and 2 could effectively inhibit the proliferation and metastasis of the human HepG2 cell and induce HepG2 cell apoptosis and cell cycle arrest in the G2/M phase. Additionally, the fermentation of Meleagrin (1) was optimized to increase its yield to 335 mg/L. These results provided bioactive inspiration and fungus resources for roquefortine alkaloid development. [ABSTRACT FROM AUTHOR]

Sichuan taimen (Hucho bleekeri Salmonidae) populations are declining, and the status of remnant populations in the Qinling Mountains is poorly known. In this study, eDNA and creel netting were used to estimate the distributions of Sichuan taimen in two upper tributaries of the Hanjiang River. A qPCR assay was applied to detect the eDNA of H. bleekeri, and the presence of Sichuan taimen in the Taibai River, but not in the Xushui River, was confirmed. Sampling during summer achieved a relatively higher detection rate. The utility of eDNA techniques for detecting low-density Sichuan taimen was demonstrated in intricate stream ecosystems. The eDNA method can increase the efficiency of the management of endangered freshwater species, such as Sichuan taimen, by providing reliable distribution data. [ABSTRACT FROM AUTHOR]

Engineering light-controlled K+ pumps from Na+-pumping rhodopsins (NaR) greatly expands the scope of optogenetic applications. However, the limited knowledge regarding the kinetic and selective mechanism of K+ uptake has significantly impeded the modification and design of light-controlled K+ pumps, as well as their practical applications in various fields, including neuroscience. In this study, we presented K+-dependent photocycle kinetics and photocurrent of a light-driven Na+ pump called Nonlabens dokdonensis rhodopsin 2 (NdR2). As the concentration of K+ increased, we observed the accelerated decay of M intermediate in the wild type (WT) through flash photolysis. In 100 mM KCl, the lifetime of the M decay was approximately 1.0 s, which shortened to around 0.6 s in 1 M KCl. Additionally, the K+-dependent M decay kinetics were also observed in the G263W/N61P mutant, which transports K+. In 100 mM KCl, the lifetime of the M decay was approximately 2.5 s, which shortened to around 0.2 s in 1 M KCl. According to the competitive model, in high KCl, K+ may be taken up from the cytoplasmic surface, competing with Na+ or H+ during M decay. This was further confirmed by the K+-dependent photocurrent of WT liposome. As the concentration of K+ increased to 500 mM, the amplitude of peak current significantly dropped to approximately ~60%. Titration experiments revealed that the ratio of the rate constant of H+ uptake (kH) to that of K+ uptake (kK) is >108. Compared to the WT, the G263W/N61P mutant exhibited a decrease of approximately 40-fold in kH/kK. Previous studies focused on transforming NaR into K+ pumps have primarily targeted the intracellular ion uptake region of Krokinobacter eikastus rhodopsin 2 (KR2) to enhance K+ uptake. However, our results demonstrate that the naturally occurring WT NdR2 is capable of intracellular K+ uptake without requiring structural modifications on the intracellular region. This discovery provides diverse options for future K+ pump designs. Furthermore, we propose a novel photocurrent-based approach to evaluate K+ uptake, which can serve as a reference for similar studies on other ion pumps. In conclusion, our research not only provides new insights into the mechanism of K+ uptake but also offers a valuable point of reference for the development of optogenetic tools and other applications in this field. [ABSTRACT FROM AUTHOR]

Environmental DNA (eDNA) techniques have emerged as a cost-effective and non-invasive strategy for monitoring the distribution of endangered aquatic species. Despite their numerous advantages, operational uncertainty at each step of the process represents a significant challenge that can impact the reliability of results. Hence, the optimization of the eDNA analytic protocol is of utmost importance. In this study, the rare and endangered fish species Sichuan taimen was chosen as the target species to evaluate the concordance between different approaches (tissue, pond, and field water samples) for eDNA detection. The results showed that membrane filtration, collection of a 2-L water sample, and utilization of the DNeasy Tissue and Blood DNA extraction kit were suitable for the analysis of DNA in water samples. Furthermore, the mtDNA D-loop region demonstrated potential as a specific molecular marker for the precise identification of Sichuan taimen. Our results indicated that TaqMan fluorescence PCR can serve as the optimal detection method for reliable identification of target DNA even at extremely low concentrations in the field. This study established an optimized eDNA analysis protocol for Sichuan taimen detection, which will provide the basis for future resource investigations aimed at protection of this endangered aquatic species. [ABSTRACT FROM AUTHOR]

The Chinese giant salamander (Andrias davidianus) is the biggest extant and endangered amphibian, which has an incongruous sex ratio during the artificial breeding process and is sex indistinguishable in appearance. The sex development mechanism of this rare species thus needs to be explored. We therefore conducted this study to explore the molecular mechanisms of sex hallmarks or gonad maintenance genes for CGS. In this study, we sequenced and analyzed the sexually mature male and female gonads of Chinese giant salamanders (CGSs) using RNA-seq technology and found a total of 15,063 differentially expressed genes (DEGs) in the testicular and ovarian tissues, of which 7973 (52.93%) were expressed in testicular and 7090 (47.07%) in ovarian tissues. Thirteen gonad DEGs were selected based on gene expression abundance and quantified in the muscle and skin tissues of both male and female CGSs. The results showed that these 13 DEGS, out of the 20 analyzed DEGs, were expressed in both the muscle and skin tissues and only two of them (Bmp15 and ZP3) were consistent in the muscle and skin compared to the gonads of CGSs of the two sexes. Overall, our study should provide an important reference for the sex-characteristics-related genes in this type of species, shedding new light on future research on this topic. [ABSTRACT FROM AUTHOR]

The targeted delivery of drugs to tumor cells and prevention of premature release before reaching the target is one of the key challenges to developing nanomedicines. In this paper, galactose decorated trimethyl chitosan (GT)–camptothecin (CPT) prodrug nanoparticles (GT-ss-CPT NPs) were prepared from GT-CPT conjugates linked by dithiodipropionic acid. The obtained GT-ss-CPT NPs were spherical with a particle size of 184.1 nm. GT-ss-CPT NPs displayed low drug release under physiological conditions, whereas efficient drug release was triggered by high GSH concentration. GT-ss-CPT NPs exhibited a higher antitumor effect both in vitro and in vivo than the free drug counterpart. More importantly, GT-ss-CPT NPs reduced the high systematic toxicity of CPT to tumor-bearing mice. In summary, GT-ss-CPT NPs can not only inhibit the premature release of CPT but also have a great potential for targeted hepatocellular carcinoma chemotherapy. [ABSTRACT FROM AUTHOR]

Heat shock proteins (HSPs) play a key role in anti-stress and immune processes and are associated with autoimmune diseases. In order to explore the immunological role of HSPs from Schizothorax prenanti (S. prenanti), SpHSP60 was cloned for the first time in this study, and the gene expressions of SpHSP27, SpHSP60, SpHSP70 and SpHSP90 in the hepatopancreas, head kidney, hindgut and spleen were analyzed by quantitative real-time PCR (qPCR) after treatment with lipopolysaccharide (LPS). The open reading frame of the SpHSP60 gene (GenBank accession number ON245159) is 1728 bp. It encodes a protein of 575 amino acids. Its C-terminus is a highly conserved and repeated glycine sequence, which is an important cofactor in ATP binding. Compared with the control group, most of the SpHSPs were significantly upregulated in the tissues examined at 12 or 24 h after LPS challenge. The most abundant expression of SpHSP70 was found in the head kidney at 24 h after LPS injection, followed by SpHSP27 in the spleen at 24 h; both of these SpHSPs displayed strong expression under the LPS stresses, about 20–70 fold more than that of SpHSP60 and SpHSP90. The temporal expression patterns of the four SpHSP genes were different in the four tissues examined. Taken together, the results suggest that SpHSP27, SpHSP60, SpHSP70 and SpHSP90 participate in innate immunity stimulated by LPS, and the response intensity of the SpHSPs was organ-specific, indicating they could provide early warning information against bacterial infection. The findings in our study will contribute to better understanding the biological processes and important roles of SpHSPs involved in defending against pathogenic bacterial challenge. [ABSTRACT FROM AUTHOR]

Simple Summary: Due to environmental pollution, habitat loss, and overutilization, the wild population of Chinese giant salamanders (Andrias davidianus) is decreasing continuously. Therefore, artificial proliferation and release are effective ways to restore the wild population of this species. However, the sex of Chinese giant salamanders (CGSs) needs to be confirmed before reintroduction or rejuvenation into the wild. The sex identification of amphibians is difficult and is usually conducted through blood hormone testing, ultrasound, or even the observation of gonads and other invasive sampling methods, and the operation is cumbersome and expensive. As one of the main aspects of animal welfare, non-invasive or minimally invasive sampling is increasingly important in wildlife research. As one of the raw materials of non-invasive sampling, urine has been considered by more and more researchers engaged in wildlife physiology and biochemistry. Our objective was to identify the sex of CGSs by detecting the ratio of hormone testosterone (T) to estrone-3-glucuronide (E1G) in urine collected by minimally invasive sampling. Perhaps this method can be used for the determination of sex in other animals, especially amphibians. Minimally invasive sampling was used to determine the sex of Chinese giant salamanders (Andrias davidianus). Urine samples (n = 25) were collected from 6 adults in the breeding season and from 19 individuals (7 adults and 12 juveniles) in the non-breeding season. The hormone testosterone (T) and estrone-3-glucuronide (E1G) in urine were collected from Chinese giant salamanders (CGSs), and the hormone extracts were analyzed by enzyme immunoassays (EIA). The data demonstrated that the urine T concentration of the male CGSs was significantly higher than that of the females during the breeding season (p < 0.05) and even more pronounced during the non-breeding season (p < 0.01). The urine E1G concentration of the males was less pronounced than that of the females during the breeding season (p < 0.01) and significantly lower during the non-breeding season (p < 0.05). The urine T/E1G values of all the male salamanders were significantly higher than those of the females (p < 0.01) during both the breeding season and the non-breeding season. An interesting pattern was found in this study: the value of urine log10(T/E1G) of the male CGSs was higher than 1, whereas the value for the females was lower than 1, during both the breeding and non-breeding seasons, and in the adult and sub-adult age groups of CGSs. There were 25 salamanders in this study and the accuracy rate reached 100% by using a log10(T/E1G) value of 1. The results of the log10(T/E1G) value provide new insight into the future development of the sex identification of CGSs and also lay the foundation for accurate sex identification in the preparation for artificial release. This is the first study to show that the T/E1G ratio in urinary hormones is reliable for the sex identification of CGSs. Additionally, urinary hormone T/E1G measures are promising sex identification tools for amphibian or monomorphic species and for those whose secondary sex characteristics are visible only during the breeding season. [ABSTRACT FROM AUTHOR]

Simple Summary: The Taibai stream salamander (Batrachuperus taibaiensis) is a protected species endemic to the Zhouzhi Heihe River. Few traditional studies have been conducted on the distribution of B. taibaiensis, leading to irregular discoveries. Environmental DNA (eDNA) is an ideal species detection technique that can increase accuracy and decrease the cost of population surveys. Here, we have established an optimal method for obtaining the eDNA of B. taibaiensis from water samples, which provides a theoretical basis and reference for resource investigation and protection of B. taibaiensis. Moreover, this study highlights the detection of a rare species in a river for use in further research. The Taibai stream salamander (Batrachuperus taibaiensis) is a recently described species of the genus Batrachuperus that occurs in the Zhouzhi Heihe River and is endangered in its native range. Here, we have established a method for water environmental DNA (eDNA) analysis of Batrachuperus using a series of optimizations. We have designed a specific set of primers for the genus Batrachuperus to amplify a 160 bp fragment of Cytb. The sequences were obtained from nested PCR on eDNA extracted from water samples, after which DNA barcoding was performed according to sequence analysis to determine the presence of the target species in the water. The method was validated using water from the Zhouzhi Heihe River with known B. taibaiensis populations and found that B. taibaiensis eDNA can move at least 150 m downstream from its point of origin. This study is the first to establish an optimal method for obtaining the eDNA of Batrachuperus from water samples, which provides a theoretical basis for resource investigation and the protection of B. taibaiensis in future research. It is also an example of the eDNA extraction of other species that live in similar waters and are less genetically diverse between species. [ABSTRACT FROM AUTHOR]