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Pseudomonas aeruginosa biofilm enhances tolerance to antimicrobials and immune system defenses. Alginate is an important component of biofilm and a virulence factor of P. aeruginosa. The degradation of alginate by alginate lyases has come to serve as an adjunctive therapeutic strategy against P. aeruginosa biofilm, but poor stability of the enzyme limited this application. Thus, PspAlgL, an alginate lyase, can degrade acetylated alginate but has poor thermostability. The 3D structure of PspAlgL was predicted, and the thermostability of PspAlgL was rationally designed by GRAPE strategy, resulting in two variants with better stability. These variants, PspAlgLS270F/E311P and PspAlgLG291S/E311P, effectively degraded the alginate in biofilm. In addition, compared with PspAlgL, these variants were more efficient in inhibiting biofilm formation and degrading the established biofilm of P. aeruginosa PAO1, and they were also able to destroy the biofilm attached to catheters and to increase the sensitivity of P. aeruginosa to the antibiotic amikacin. This study provides one potential anti-biofilm agent for P. aeruginosa infection.

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Cavefishes represent a taxon that has experienced natural selection pressures. This paper summarizes the results with respect to the taxonomy, diversity, phylogeny, and adaptation aspects of cavefishes research. It showed that: 1) These studies suggest that cavefishes play important roles in the study of geologic history and adaptation to extreme environments, but the mechanisms involved 168 species of cavefishes belonging to 17 genera, four families, and two orders have been recorded in China. Meanwhile, more new species are being discovered recently, and the species diversity of cavefishes are still underestimated, indicating the need to strengthen the survey in field. 2) The biogeography of cavefishes have focused on Sinocyclocheilus and Triplophysa, that have helped understand the geomorphology of karst areas in southern China and the spatial pattern of species diversity. These studies revealed the influences of evolution and geological history in Sinocyclocheilus, but there are still many species that have not been studied accordingly. 3) Some adaptive mechanistic studies have been conducted on cavefishes, primarily focusing on eye and body color degradation and energy metabolism in the genus Sinocyclocheilus to reveal adaptive mechanisms in the dark environment. 4) The IUCN list of protected cavefishes species in China only includes 21 species. The List of Key Protected Wild Animals for 2021 includes all species of Sinocyclocheilus as National Class II.It is necessary to strengthen the research on the biodiversity and adaptation and need consider the conservation actions for cavefishes.

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Pseudomonas aeruginosa can form biofilms at the site of burn wound, leading to infection and the failure of treatment regimens. The previous in vitro study demonstrated that a combination of the quorum-quenching enzyme AidHA147G and the extracellular matrix hydrolase PslG was effective in inhibiting biofilm and promoting antibiotic synergy. The aim of the present study was to evaluate the efficacy of this combination of enzymes in conjunction with tobramycin in treating burn wound infected with P. aeruginosa. The results showed that this treatment was effective in quorum-quenching and biofilm inhibition on infected wounds. Compared with the tobramycin treatment only, simultaneous treatment with the enzymes and antibiotics significantly reduced the severity of tissue damage, decreased the bacterial load, and reduced the expression of the inflammatory indicators myeloperoxidase (MPO) and malondialdehyde (MDA). Topical application of the enzymes also reduced the bacterial load and inflammation to some extent. These results indicate that the combined-enzyme approach is a potentially effective treatment for P. aeruginosa biofilm infections of burn wounds.

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Phthalate esters (PAEs) and parabens are environmental pollutants that can be toxic to human health. Herein, a cold-adapted esterase from the Mao-tofu metagenome named Est1260 was screened for its PAE-hydrolyzing potential in cold temperatures. The results showed that purified Est1260 could degrade a variety of PAEs and parabens at temperatures as low as 0 °C. After careful analysis of the structural information and molecular docking, site-saturation mutation was conducted at the identified hotspots. Protein expression of variant A1B6 doubled, and its thermal stability significantly improved (24 times) without sacrificing activity at low temperatures. In addition, Est1260 and its variants were activated by NaCl and demonstrated resistance to high concentrations of saline (up to 5 M), making it a potential biocatalyst for bioremediation of PAE and paraben-polluted environments.

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The pervasive use of pyrethroids is seriously hazardous to the environment and even human health. Enzymatic bioremediation is potentially a rapid and environmentally friendly technology to combat the pollution of pyrethroid pesticides. The hydrolysis of ester linkages is the initial and critical enzymatic step in microbial degradation pathways. Here, the versatile and thermostable esterase Est816 was cloned and its new function, pyrethroid-hydrolysis activity, was expanded. To further improve its pyrethroid-hydrolysis ability, Est816 was modified by rational design. After two rounds of mutation, the best-performing mutant, Est816A216V/K238N/M97V, was obtained, which could completely degrade 1 mg/L λ-cyhalothrin, cypermethrin, and deltamethrin within 20 min, and efficiently degrade fenvalerate, reaching over 80% conversion. Degradation activity analyses showed that three substitutions (A216V, K238 N and M97V) were beneficial for enhancing the activity of Est816. Enzymatic characterization showed that Est816A216V/K238N/M97V inherited broad substrate specificity and possessed excellent stability and adaptability over wide ranges of temperature and pH, which is essential for bioremediation in frequently changing conditions. Furthermore, Est816A216V/K238N/M97V had the best degradation effect on all four pyrethroid residues in Panax notoginseng root, with more than 87% conversion after 24 h. Pyrethroid residues in tea, cucumber, and soil were reduced by more than 76%, 80%, and 76%, respectively. Taken together, these findings highlight the great potential of Est816A216V/K238N/M97V in the bioremediation of pyrethroid-contaminated soil and agricultural products.

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Pseudomonas aeruginosa is an opportunistic pathogen that forms biofilms during infection, resulting in recalcitrance to antibiotic treatment. Biofilm inhibition is a promising research direction for the treatment of biofilm-associated infections. Here, a combined-enzyme biofilm-targeted strategy was put forward for the first time to simultaneously prevent biofilm formation and break down preformed biofilms. The N-acylhomoserine lactonase AidH was used as a quorum-sensing inhibitor and was modified to enhance the inhibitory effect on biofilms by rational design. Mutant AidHA147G exerted maximum activity at the human body temperature and pH and could reduce the expression of virulence factors as well as biofilm-related genes of P. aeruginosa. Subsequently, the P. aeruginosa self-produced glycosyl hydrolase PslG joined with AidHA147G to disrupt biofilms. Interestingly, under the combined-enzyme intervention for P. aeruginosa wild-type strain PAO1 and clinical strains, no biofilm was observed on the bottom of NEST glass-bottom cell culture dishes. The combination strategy also helped multidrug-resistant clinical strains change from resistant to intermediate or sensitive to many antibiotics commonly used in clinical practice. These results demonstrated that the combined-enzyme approach for inhibiting biofilms is a potential clinical treatment for P. aeruginosa infection.

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Modern accumulations of genetic data offer unprecedented opportunities for understanding the systematic classification and origins of specific groups of organisms. The genus Sinocyclocheilus is among the most cave abundant genera in Cyprinidae, with 76 recognized species, belonging to 4 species groups. Recent phylogenetic studies have shown that the classification of species groups within the genus Sinocyclocheilus remains controversial. In this study, we constructed a sequence supermatrix of 26 species from 4 species groups of the genus Sinocyclocheilus using the mitochondrial genome to reveal phylogenetic relationships, historical biogeography and patterns of species diversification in the genus Sinocyclocheilus. Phylogenetic analysis strongly supports the monophyletic groups of the 3 species groups (S. jii, S. cyphotergous, and S. tingi groups) except the S. angularis group. Phylogenetic analysis showed that S. anshuiensis and S. microphthalmus, which were recognized as numbers of S. angularis group, formed a strongly supported independent clade. Therefore, we propose a new species group, the S. microphthalmus group, which contains S. anshuiensis and S. microphthalmus. Biogeographic reconstruction suggests that the living Sinocyclocheilus may have originated in north-central Guangxi at the late Eocene and dispersed outward after a vicariance at 32.31 Million years ago (Ma). Early diversification is focused on the late Oligocene (ca. 25 Ma), which is related to the second uplift of the Qinghai-Tibetan Plateau and the lateral extrusion of the Indochina at the Oligocene/Miocene boundary. Our results suggest that 2 uplifts of the Qinghai-Tibetan Plateau and climate change in the Miocene may have influenced the diversification of the Sinocyclocheilus lineage.

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The complete mitochondrial DNA genome of Triplophysa nasobarbatula was sequenced and characterized. Triplophysa nasobarbatula revealed that the complete length of its mitochondrial genome was 16,316 bp, composed of A (29.71%), C (24.79%), G (17.22%), T (28.29%), A + T (57.99%), and C + G (42.01%). Its genetic constitution and arrangement were consistent with the taxon of the Teleost, including 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and 2 main non-coding regions, D-loop region and OL region. All genes were encoded by the H-strand, except for 1 protein-coding gene (ND6) and 8 tRNA genes (tRNA-Gln, tRNA-Ala, tRNA-Cys, tRNA-Asn, tRNA-Tyr, tRNA-Ser, tRNA-Glu and tRNA-Pro) are encoded by the L-strand. Our mitochondrial genome data may provide information for taxonomic resolution, taxonomic resolution, and other studies about this genus of Triplophysa.

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Triplophysa lewanggensis revealed that the complete length of its mitochondrial genome was 16,568 bp, composed of A (31.7%), T (27.1%), G (15.9%), C (25.4%), and A + T (58.8%). Its genetic constitution and arrangement were consistent with other Triplophysa, including 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and 1 control area (D-loop). All genes were encoded by the H-strand, except for one protein-coding gene (ND6) and eight tRNA genes (tRNAGln, tRNAAla, tRNACys, tRNAAsn, tRNATyr, tRNASer(UCN), tRNAGlu, tRNAPro) are encoded by the L chain, and the remaining genes are encoded by the H chain. The phylogenetic tree was divided into two main clades: one just including Triplophysa and the other including Oreonectes, Homatula, Schistura, Nemacheilus, Traccatichthys. T. lewanggensis is distributed in Southwest Guizhou Province attached to Yunnan-Guizhou Plateau, T. xiangxiensis is distributed in Hunan Province, they are near relatives, but their kinship with Triplophysa distributed in Tibetan Plateau is farther.

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Oreonectes daqikongensi revealed that the complete length of its mitochondrial genome was 16,578 bp, composed of A (30.7%), T (25.6%), G (16.2%), C (27.5%) and A + T (56.3%). Its genetic constitution and arrangement were consistent with those of other Osteichthyes, including 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes, and 1 control area (D-loop). All genes were encoded by the H-strand, except for 1 protein-coding gene (ND6) and 8 tRNA genes (tRNA-Gln, tRNA-Ala, tRNA-Cys, tRNA-Asn, tRNA-Tyr, tRNA-Ser, tRNA-Glu and tRNA-Pro) are encoded by the L-strand. The mitochondrial genes were arranged very closely. There were four gene overlapping regions, with an overall length of 21 bp and a base covering number range of 1-14 bp, and 23 intergenic regions, with an overall length of 147 bp and an intergenic length range of 1-19 bp. There were 10 gene pairs that were neither overlapping nor intergenic.

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This study aimed to describe a new specimen of cavefish collected from a karst cave in the Daqikong area of Libo County, Guizhou. Twenty-six cavefish specimens were collected and identified as a new species of Balitoridae: Nemacheilinae, and named Oreonectes daqikongensissp. n. A genetic analysis was performed and showed that its genetic distances from Oreonectes shuilongensis and Oreonectes platycephalus are higher than intraspecific distances. Discovery of this species will be helpful to understand the distribution of Oreonectes.