Molecular evidence for nitrite-dependent anaerobic methane-oxidising bacteria in the Jiaojiang Estuary of the East Sea (China).

Nitrite-dependent anaerobic methane oxidation (N-DAMO) is a recently discovered process linking the global carbon and nitrogen cycles. This process was reported to be mediated by "Candidatus Methylomirabilis oxyfera". To date, M. oxyfera-like bacteria have been detected in a limited number of freshwater habitats, but whether these bacteria occur in estuarine habitats is currently unknown. In this study, the distribution, diversity and abundance of M. oxyfera-like bacteria were studied in the sediment of the Jiaojiang Estuary of the East Sea (China). Both the 16S ribosomal RNA (rRNA) and pmoA genes confirmed the occurrence of M. oxyfera-like bacteria in the examined estuary. The recovered 16S rRNA gene sequences showed 91.5-97.2 % identity to the 16S rRNA gene of M. oxyfera, and the recovered pmoA gene sequences showed 85.1-95.4 % identity to the pmoA gene of M. oxyfera. Quantitative PCR further confirmed the occurrence of M. oxyfera-like bacteria in this estuary, with the abundance varying from 5.80 ± 0.28 × 10(4) to 8.35 ± 0.52 × 10(7) copies g (dry weight)(-1). Correlation analysis indicated that the sediment organic content was the most important factor affecting the distribution of M. oxyfera-like bacterial communities in the examined sediments among the environmental factors investigated. This study demonstrated for the first time the existence of M. oxyfera-like bacteria in an estuarine environment and showed the correlations between the distribution of these bacteria and the estuarine environmental conditions.

Optimizing heat pump unit selection for recirculating aquaculture workshops through computational fluid dynamics.

The selection of water temperature regulation equipment plays a crucial role in the design of workshops. At present, the choice of water temperature control equipment is usually based on the volume of the fish pond and thermal parameter calculation, combined with aquaculture experience. Empirical formulas only work in specific conditions due to factors like the environment, climate, and fish types,resulting in inaccurate equipment selection outcomes. Recognizing this limitation, this paper proposes to apply CFD simulation of the temperature field to accurately calculate the heat exchange value between indoor air and water, thereby predicting the heat exchange values during aquaculture activities in the aquaculture workshop. providing a new approach for equipment selection. This paper selects a puffer fish breeding workshop in Dalian as the simulation object, establishing a 3D unsteady-state Computational Fluid Dynamics model. The model considers outdoor temperature, solar radiation, and phase-change heat transfer in water. Comparison with experimental data reveals a root mean square error of 0.46°C for the simulated results. During summer, the highest cooling load occurs at 16:00, reaching 94.6 kW. It is recommended to employ the Daikin GCHP-40MAH ground source heat pump as the water temperature control equipment. CFD simulation validates its effectiveness in shaping the indoor temperature field post-installation. the investment in water temperature control equipment can be reduced to a certain degree. This provides a reference value for the selection of water temperature equipment in aquaculture workshops.

Molecular characterization of a phenylalanine ammonia-lyase gene (BoPAL1) from Bambusa oldhamii.

Phenylalanine ammonia-lyase is the first enzyme of general phenylpropanoid pathway. A PAL gene, designated as BoPAL1, was cloned from a Bambusa oldhamii cDNA library. The open reading frame of BoPAL1 was 2,139 bp in size and predicted to encode a 712-amino acid polypeptide. BoPAL1 was the first intronless PAL gene found in angiosperm plant. Several putative cis-acting elements such as P box, GT-1motif, and SOLIPs involved in light responsiveness were found in the 5'-flanking sequence of BoPAL1 which was obtained by TAIL-PCR method. Recombinant BoPAL1 protein expressed in Pichia pastoris was active. The optimum temperature and pH for BoPAL1 activity was 50°C and 9.0, respectively. The molecular mass of recombinant BoPAL1 was estimated as 323 kDa using gel filtration chromatography and the molecular mass of full-length BoPAL was about 80 kDa, indicating that BoPAL1 presents as a homotetramer. The Km and kcat values of BoPAL1 for L-Phe were 1.01 mM and 10.11 s(-1), respectively. The recombinant protein had similar biochemical properties with PALs reported in other plants.

Cloning and expression of a phenylalanine ammonia-lyase gene (BoPAL2) from Bambusa oldhamii in Escherichia coli and Pichia pastoris.

Phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) is the first committed enzyme of phenylpropanoid pathway. A PAL gene, designated as BoPAL2, was cloned from a Bambusa oldhamii cDNA library. The open reading frame of BoPAL2 was 2142bp in size encoding a 713-amino acid polypeptide. BoPAL2 was heterologous expressed in Escherichia coli and Pichia pastoris. The recombinant proteins were exhibited PAL and tyrosine ammonia-lyase activities. The recombinant BoPAL2 had a subunit mass of 80kDa and existed as a homotetramer. The optimum temperature and pH of BoPAL2 were 50-60 degrees C and 8.5-9.0, respectively. The K(m) and k(cat) values of BoPAL2 expressed in E. coli were 250microM and 10.12s(-1). The K(m) and k(cat) values of BoPAL2 expressed in P. pastoris were 331microM and 16.04s(-1). The recombinant proteins had similar biochemical properties and kinetic parameters with PALs reported in other plants.

Trans-splicing into highly abundant albumin transcripts for production of therapeutic proteins in vivo.

Spliceosome-mediated RNA trans-splicing has emerged as an exciting mode of RNA therapy. Here we describe a novel trans-splicing strategy, which targets highly abundant pre-mRNAs, to produce therapeutic proteins in vivo. First, we used a pre-trans-splicing molecule (PTM) that mediated trans-splicing of human apolipoprotein A-I (hapoA-I) into the highly abundant mouse albumin exon 1. Hydrodynamic tail vein injection of the hapoA-I PTM plasmid in mice followed by analysis of the chimeric transcripts and protein, confirmed accurate and efficient trans-splicing into albumin pre-mRNA and production of hapoA-I protein. The versatility of this approach was demonstrated by producing functional human papillomavirus type-16 E7 (HPV16-E7) single-chain antibody in C57BL/6 mice and functional factor VIII (FVIII) and phenotypic correction in hemophilia A mice. Altogether, these studies demonstrate that trans-splicing to highly abundant albumin transcripts can be used as a general platform to produce therapeutic proteins in vivo.

Mg2+ regulation of kinase signaling and immune function.

Mg2+ is required at micromolar concentrations as a cofactor for ATP, enzymatic reactions, and other biological processes. We show that decreased extracellular Mg2+ reduced intracellular Mg2+ levels and impaired the Ca2+ flux, activation marker up-regulation, and proliferation after T cell receptor (TCR) stimulation. Reduced Mg2+ specifically impairs TCR signal transduction by IL-2-inducible T cell kinase (ITK) due to a requirement for a regulatory Mg2+ in the catalytic pocket of ITK. We also show that altered catalytic efficiency by millimolar changes in free basal Mg2+ is an unrecognized but conserved feature of other serine/threonine and tyrosine kinases, suggesting a Mg2+ regulatory paradigm of kinase function. Finally, a reduced serum Mg2+ concentration in mice causes an impaired CD8+ T cell response to influenza A virus infection, reduces T cell activation, and exacerbates morbidity. Thus, Mg2+ directly regulates the active site of specific kinases during T cell responses, and maintaining a high serum Mg2+ concentration is important for antiviral immunity in otherwise healthy animals.

[Survey of prevention knowledge and behavior of echinococcosis and influencing factors in residents in agricultural and pastoral areas of Maerkang City].

OBJECTIVE: To master the prevention knowledge and behavior of echinococcosis and the influencing factors in residents in agricultural and pastoral areas of Maerkang City, so as to provide the reference for formulating the policy and strategy of echinococcosis prevention and control. METHODS: In April, 2017, 796 residents in agricultural and pastoral areas of Maerkang City were selected by the multi-stage stratified random sampling method, and surveyed with questionnaires. The main questions included the prevention knowledge and related behaviors of echinococcosis. RESULTS: The awareness rate of prevention knowledge of echinococcosis was 70.1% (558/796). The awareness rate was the lowest in the group aged 15 to 25 years. The difference of knowledge awareness rate was statistically significant among the different age groups (χ2 = 16.408, P < 0.01). The knowledge awareness rate of the people with middle school or above educational level was higher than that of the people with primary school or below educational level (χ2 = 4.869, P = 0.027). The knowledge awareness rate of the dog owners was lower than that of the people without a dog (χ2 = 11.384, P = 0.001). The knowledge awareness rate in the people who received the knowledge training of three times or more was higher than that in the people who received the knowledge training of two times or less (χ2 = 11.439, P = 0.003). The logistic regression showed that the age, educational level, training times and dog raising were the main influencing factors. The related behavior survey showed that 32.7% (260/796) of residents did not drink unboiled water, 37.0% (61/165) of dog owners tied the dog, 43.6% (72/165) of dog owners fed the dogs with anthelmintic regularly, and only 10.3% (17/165) of dog owners buried the dog's dung. CONCLUSIONS: The awareness rate of echinococcosis prevention knowledge in Maerkang City should be improved. The health education should be focused on the young people, people with low educational level, and people who have dogs. The behaviors of drinking unboiled water, tying dogs, feeding dogs with anthelmintic regularly, and burying dog' s dung should be advocated.

Primary care reform in the Peoples' Republic of China: implications for training family physicians for the world's largest country.

Just as China has struggled with bringing an expanding economy to a country with large areas of economic underdevelopment, the Chinese health care system is experiencing tension between tradition and modernization. Because of this tension, health care in the Peoples' Republic of China has been undergoing significant reform since the beginning of the 1980s. Experiments in market-based health systems have been unsuccessful and have exacerbated disparities. New reforms, announced in 2006, stress the role of family physicians in leading the health care system. This paper discusses the history of the developments that led up to the new reforms and the educational challenges of training sufficient numbers of family physicians to meet the requirements of the new system.

Socioeconomic status is a critical risk factor for human rabies post-exposure prophylaxis.

The socioeconomic status of the patients is the important factor for post-exposure prophylaxis (PEP). However, few investigations were designed to study the correlation between the socioeconomic status and PEP. This study set out to determine the importance of socioeconomic status for PEP. All of the 11,670 at-risk populations of rabies in the public health centre of San Sheng County in Chengdu from January 2002 to December 2009 were reviewed retrospectively. We identified 11,350 patients on vaccination and 550 patients with rabies immunoglobulin. RIG was administered to 4.85% bite victims attending the rabies prevention clinics, while 61.36% had a category III exposure. The incidence of receiving RIG in the population of the high level of income (49.38%) was much higher than the groups of the medium level (8.08%) and the low level of income (1.46%) (P<0.05). The incidence of receiving RIG with above high school (23.08%) was much higher than the groups of the primary school (3.01%), the junior school (12.56%) and the illiteracy (2.08%) (P<0.05). In the logistic regression analysis by stepwise approach, the socioeconomic status was the most important factor for PEP (95% CI 1.20-2.04). Vaccination and immunoglobulin proved to be the most prominent two factors for PEP but whether receiving Vaccination and immunoglobulin treatment or not is determined by the socioeconomic status. So, the socioeconomic status was the most important factor for PEP.

Cloning, expression, site-directed mutagenesis and immunolocalization of phenylalanine ammonia-lyase in Bambusa oldhamii.

Phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) from green bamboo was isolated and cloned from the shell of Bambusa oldhamii. The K(m) of bamboo shell PAL for L-Phe was 476 µM, and the molecular mass of native PAL was estimated as 275 kDa and the molecular mass of a subunit was about 76 kDa, indicating that PAL from bamboo also exists as a tetramer. The optimum temperature for PAL activity was 50°C and the optimal pH 9.0. The identity of the purified bamboo shell PAL was confirmed using Q-TOF tandem MS/MS de novo sequencing. Four PAL genes, designated as BoPAL1 to BoPAL4, were cloned from B. oldhamii. The open reading frames of BoPAL3 and BoPAL4 were 2142 and 2106 bp in size, respectively: BoPAL2-4 contained one intron and two exons, but no intron was found in BoPAL1. BoPAL4 expressed in Escherichia coli possessed both PAL and tyrosine ammonia-lyase activities. While recombinant wild-type PAL proteins had similar biochemical properties to the native bamboo shell PAL, both site-directed mutagenesis of BoPAL1 F133H and BoPAL2 F134H, respectively, showed decreased k(cat)/K(m) values toward L-Phe, whereas BoPAL2 F134H showed a slightly increased k(cat)/K(m) value toward L-Tyr. These data suggest other residues largely control Phe/Tyr substrate specificity. An antibody raised against the purified shell PAL was generated for histochemical studies. In bamboo shell and branch shoots, PAL was localized primarily in sclerenchyma cells.