Microbial behaviours inside alternating anaerobic-anoxic environment of a sulfur cycle-driven EBPR system: A metagenomic investigation.

Denitrifying sulfur conversion-assisted enhanced biological phosphorus removal (DS-EBPR) was recently developed for saline wastewater treatment. However, the main functional bacteria and the interrelationship of functional bacteria of the DS-EBPR have not been defined and identified so far. This study used metagenomics and multivariate statistics to deduce the functional microbial community and distribution of functional genes associated with the critical metabolic pathways of carbon (C), nitrogen (N), phosphorus (P) and sulfur (S), particularly regarding how they would behave under the alternating anaerobic-anoxic conditions inside a long-term DS-EBPR system. An analysis of the metagenomics and metabolic functions identified 11 major microbial species which were classifiable into four groups: sulfate reducing bacteria (SRB, 0.8-2.2%), sulfur oxidizing bacteria (SOB, 31.9-37.7%), denitrifying phosphate accumulating organisms (DPAOs, 10.0-15.8%) and glycogen accumulating organisms (GAOs, 3.7-7.7%). The four groups of microorganisms performed their respective metabolisms synergistically. In terms of distribution of functional genes, SRB (Desulfococcus and Desulfobacter) and SOB (Chromatiaceae and Thiobacillus) are not only encoded by the related sulfur conversion genes (sqr, dsrAB, aprAB and sat), but also encoded by the necessary ppx and ppk1 gene for P removal that they can be considered as the potential S-related PAOs. Between the anaerobic and anoxic conditions, the metagenome-based microbial community remained structurally similar, but the functional genes, which encode various key enzymes for the P, N, and S pathways, changed in abundance. This study contributes to our understanding on the interactions and competition between the SRB, SOB, DPAOs, and GAOs in a DS-EBPR system.

A novel online analyzer for accurate and rapid measurement of volatile fatty acids in anaerobic wastewater treatment.

Volatile fatty acids (VFAs), which are largely generated during the anaerobic acidification process, are considered to be reliable indicators of the stable process operation. However, the common methods for monitoring VFAs are offline, and they are typically manual requiring time-consuming, costly and complex instruments. This study aims to develop a novel online analyzer for automatic measuring VFAs, which was based on the 5-pH point titration, embedded with a proportional-integral-derivative (PID) feedback control system. The results show that it can achieve accurate and rapid monitoring of VFAs ranging between 0-400 mg/L (<9 min/sample) but simultaneously faces the problems of overtitration and interference of complex characteristics of wastewater. In order to improve its accuracy and stability, the effects of three general coefficients (KI,KP, and KD) of PID on the titration were investigated, and the optimal values of KI, KP, and KD were found to be 1.5, 1.0, and -1.0~0.5, respectively. Besides, the initial titration speed was set at 0.06 mL/min, equal to the minimum speed of the peristaltic pump, and the dichotomy approach was integrated into the PID feedback controller. Owing to the above improvements, the relative mean deviation and standard deviation of measuring VFAs in both synthetic and real wastewaters were mostly lower than 5.0% and 5.0 mg/L, proving the online analyzer is rapid, accurate and reliable.

Association of serum uric acid with biventricular myocardial dysfunction in patients with type 2 diabetes mellitus.

BACKGROUND AND AIMS: Increased serum uric acid (SUA) is common in type 2 diabetes mellitus (T2DM) and is associated with left ventricular (LV) myocardial dysfunction. Nonetheless the association of SUA with right ventricular (RV) function in T2DM has not been studied. This study aimed to investigate the association of SUA with biventricular myocardial function in patients with T2DM. METHODS AND RESULTS: A total of 560 patients with T2DM were enrolled and divided into four groups according to sex-specific quartiles of SUA. Transthoracic echocardiography was performed and two-dimensional speckle tracking was used to measure biventricular myocardial strain, including LV global longitudinal strain (GLS), circumferential strain (CS), radial strain (RS), and RV free wall longitudinal strain (RV-FWLS). The absolute value of all biventricular strain parameters showed a stepwise decrease across SUA quartiles (all P < 0.01). In particular, LV assessment by GLS, CS and RS demonstrated that those in the 4th quartile were impaired compared with the other quartiles (all P < 0.05). Similarly, RV-FWLS of the 4th quartile was significantly impaired compared with the 1st and 2nd quartiles (all P < 0.05). The same reduction in biventricular strain across SUA quartiles was observed in patients with estimated glomerular filtration rate < or ≥60 ml/min/1.73 m2, and glycated hemoglobin < or ≥7.0% (all P < 0.05). Multivariable linear regression analysis demonstrated that higher quartile of SUA was independently associated with impaired biventricular myocardial strain (all P < 0.05). CONCLUSIONS: SUA was independently associated with biventricular myocardial dysfunction in asymptomatic T2DM patients, regardless of renal function or diabetic control.

Structural Basis for the Broad, Antibody-Mediated Neutralization of H5N1 Influenza Virus.

Human infection with highly pathogenic avian influenza A viruses causes severe disease and fatalities. We previously identified a potent and broadly neutralizing antibody (bnAb), 13D4, against the H5N1 virus. Here, we report the co-crystal structure of 13D4 in complex with the hemagglutinin (HA) of A/Vietnam/1194/2004 (H5N1). We show that heavy-chain complementarity-determining region 3 (HCDR3) of 13D4 confers broad yet specific neutralization against H5N1, undergoing conformational rearrangement to bind to the receptor binding site (RBS). Further, we show that mutating four critical residues within the RBS-Trp153, Lys156, Lys193, and Leu194-disrupts the binding between 13D4 and HA. Viruses bearing Asn193 instead of Lys/Arg can evade 13D4 neutralization, indicating that Lys193 polymorphism might be, at least in part, involved in the antigenicity of recent H5 genotypes (such as H5N6 and H5N8) as distinguished from H5N1. BnAb 13D4 may offers a template for therapeutic RBS inhibitor design and serve as an indicator of antigenic change for current H5 viruses.IMPORTANCE Infection by highly pathogenic avian influenza A virus remains a threat to public health. Our broadly neutralizing antibody, 13D4, is capable of neutralizing all representative H5N1 viruses and protecting mice against lethal challenge. Structural analysis revealed that 13D4 uses heavy-chain complementarity-determining region 3 (HCDR3) to fit the receptor binding site (RBS) via conformational rearrangement. Four conserved residues within the RBS are critical for the broad potency of 13D4. Importantly, polymorphism of Lys193 on the RBS may be associated with the antigenicity shift from H5N1 to other newly emerging viruses, such as H5N6 and H5N8. Our findings may pave the way for highly pathogenic avian influenza virus vaccine development and therapeutic RBS inhibitor design.

Quantitative analysis of mitral valve morphology in mitral valve prolapse with real-time 3-dimensional echocardiography: importance of annular saddle shape in the pathogenesis of mitral regurgitation.

BACKGROUND: Few data exist on the relation of the 3-dimensional morphology of mitral valve and degree of mitral regurgitation (MR) in mitral valve prolapse. METHODS AND RESULTS: Real-time 3-dimensional transesophageal echocardiography of the mitral valve was acquired in 112 subjects, including 36 patients with mitral valve prolapse and significant MR (≥3+; MR+ group), 32 patients with mitral valve prolapse but no or mild MR (≤2+; MR- group), 12 patients with significant MR resulting from nonprolapse pathologies (nonprolapse group), and 32 control subjects. The 3-dimensional geometry of mitral valve apparatus was measured with dedicated quantification software. Compared with the normal and MR- groups, the MR+ group had more dilated mitral annulus (P<0.0001), a reduced annular height to commissural width ratio (AHCWR) (P<0.0001) indicating flattening of annular saddle shape, redundant leaflet surfaces (P<0.0001), greater leaflet billow volume (P<0.0001) and billow height (P<0.0001), longer lengths from papillary muscles to coaptation (P<0.0001), and more frequent chordal rupture (P<0.0001). Prevalence of chordal rupture increased progressively with annulus flattening (7% versus 24% versus 42% for AHCWR >20%, 15%-20%, and <15%, respectively; P=0.004). Leaflet billow volume increased exponentially with decreasing AHCWR in patients without chordal rupture (r(2)=0.66, P<0.0001). MR severity correlated strongly with leaflet billow volume (r(2)=0.74, P<0.0001) and inversely with AHCWR (r(2)=0.44, P<0.0001). In contrast, annulus dilatation but not flattening occurred in nonprolapse MR patients. An AHCWR <15% (odds ratio=7.1; P=0.0004) was strongly associated with significant MR in mitral valve prolapse. CONCLUSION: Flattening of the annular saddle shape is associated with progressive leaflet billowing and increased frequencies of chordal rupture and may be important in the pathogenesis of MR in mitral valve prolapse.

Electrogenerated singlet oxygen and reactive chlorine species enhancing volatile fatty acids production from co-fermentation of waste activated sludge and food waste: The key role of metal oxide coated electrodes.

Electrochemical pretreatment (EPT) has shown to be superior in improving acidogenic co-fermentation (Co-AF) of waste activated sludge (WAS) and food waste (FW) for volatile fatty acids (VFAs). However, the influence of EPT electrode materials on the production of electrogenerated oxidants (such as singlet oxygen (1O2) and reactive chlorine species (RCS)), as well as their effects on properties of electrodes, the microbial community structure and functional enzymes remain unclear. Therefore, this study investigated the effects of various metal oxide coated electrodes (i.e., Ti/PbO2, Ti/Ta2O5-IrO2, Ti/SnO2-RuO2, and Ti/IrO2-RuO2) on EPT and subsequent Co-AF of WAS-FW. The results showed that EPT with Ti/PbO2, Ti/Ta2O5-IrO2, Ti/SnO2-RuO2 and Ti/IrO2-RuO2 electrodes generated 165.3-848.2 mg Cl2/L of RCS and 5.643 × 1011-3.311 × 1012 spins/mm3 of 1O2, which significantly enhanced the solubilization and biodegradability of WAS-FW by 106.4 %-233.6 % and 177.3 %-481.8 %, respectively. Especially with Ti/Ta2O5-IrO2 as the electrode material, an appropriate residual RCS (2.0-10.4 mg Cl2/L) remained in Co-AF step, resulted in hydrolytic and acidogenic bacteria (e.g., Prevotella_7, accounting for 78.9 %) gradually become dominant rather than methanogens (e.g., Methanolinea and Methanothrix) due to their different tolerance to residual RCS. Meanwhile, the functional gene abundances of hydrolytic and acidogenic enzymes increased, while the methanogenic enzymes deceased. Consequently, this reactor produced the highest VFAs up to 545.5 ± 36.0 mg COD/g VS, which was 101.8 % higher than that of the Control (without EPT). Finally, the economic analysis and confirmatory experiments further proved the benefits of WAS-FW Co-AF with EPT.

Thiosulfate/FeCl3 pre-treatment enhances short-chain fatty acid production and mitigates H2S generation during anaerobic fermentation of waste activated sludge: Performance, microbial community and ecological analyses.

Anaerobic fermentation (AF) has been identified as a promising method of transforming waste activated sludge (WAS) into high-value products (e.g., short-chain fatty acids (SCFAs)). This study developed thiosulfate/FeCl3 pre-treatment and investigated the effects of different thiosulfate/FeCl3 ratios (S:Fe = 3:1, 3:2, 1:1, 3:4 and 3:5) on SCFA production and sulfur transformation during the AF of WAS. At a S:Fe ratio of 1:1, the maximal SCFA yield (933.3 mg COD/L) and efficient H2S removal (96.5 %) were obtained. S:Fe ratios ≤ 1:1 not only benefited hydrolysis and acidification but largely mitigated H2S generation. These results were supported by the enriched acidogens and reduced sulfur-reducing bacteria (SRB). Molecular ecological network analysis further revealed that the keystone taxon (g_Saccharimonadales) was found in S:Fe = 1:1, together with reductions in associations among methanogens, acidogens and SRB. This work provides a strategy for enhancing high-value product recovery from WAS and minimising H2S emissions.

Target organ damage in untreated hypertensive patients with primary aldosteronism.

An increased risk of target organ damage (TOD) has been reported in patients with primary aldosteronism (PA). However, there is relatively little related research on the correlation between the degree of TOD and those with and without PA in newly diagnosed hypertensive patients. The aim of this study was to assess the association between PA and TOD among patients with newly diagnosed hypertension. Newly diagnosed hypertensive patients were consecutively recruited from January 2015 to June 2020 at the University of Hong Kong-Shenzhen Hospital. Patients were stratified into those with and without PA. Data for left ventricular mass index (LVMI), carotid intima-media thickness (CIMT) and plaque, and microalbuminuria were systematically collected. A total of 1044 patients with newly diagnosed hypertension were recruited, 57 (5.5%) of whom were diagnosed with PA. Patients with PA had lower blood pressure, serum lipids, body mass index, and plasma renin activity and a higher incidence of hypokalemia than those without PA. In contrast, the prevalence of left ventricular hypertrophy, increased CIMT, and microalbuminuria was higher in patients with PA than in those without PA. Multivariable regression analysis demonstrated that PA was independently associated with increased LVMI, CIMT and microalbuminuria. Among patients with newly diagnosed hypertension, those with PA had more severe TOD, including a higher LVMI, CIMT and microalbuminuria, than those without PA. These findings emphasize the need for screening TOD in newly diagnosed hypertension due to underlying PA.

Enhancing anaerobic fermentation of waste activated sludge by investigating multiple electrochemical pretreatment conditions: Performance, modeling and microbial dynamics.

Electrochemical pretreatment (EPT) is an efficient technology to improve volatile fatty acids (VFAs) production during anaerobic fermentation of waste activated sludge (WAS). This study investigated the co-effects of different current intensities, electrolyte NaCl dosage and pretreatment time for promoting VFAs production. The results showed that it was considerably enhanced by 51.6 % when EPT was performed at 1.0 A, 1.0 g/L and 60 min, and response surface methodology strategy adjusted the optimal EPT conditions to 1.0 A, 1.2 g/L and 66 min. The potential mechanisms were proposed that EPT at optimal conditions greatly enhanced solubilization and hydrolysis of WAS and selectively inactivated methanogens, causing the enrichment of acidogenic bacteria (i.e., Lactobacillus, Saccharimonadales, Tetrasphaera and Prevotella) due to generated reactive chlorine species. Finally, the economic analysis indicated the promising application potential with the profit of EPT at optimal conditions increasing by 36.0 %.

Short-chain fatty acid production and phosphorous recovery from waste activated sludge via anaerobic fermentation: A comparison of in-situ and ex-situ thiosulfate-assisted Fe2+/persulfate pretreatment.

Recently, increasing attention is given on the resource and energy recovery (e.g. short-chain fatty acids (SCFAs) and phosphorus (P)) from waste active sludge (WAS) under the "Dual carbon goals". This study compared four thiosulfate-assisted Fe2+/persulfate (TAFP) pretreatments of WAS, i.e. in-situ TAFP pretreatment (R1), ex-situ TAFP pretreatment (R2), in-situ TAFP pretreatment + pH adjustment (R3) and ex-situ TAFP pretreatment + pH adjustment (R4), followed by anaerobic fermentation over 20 days for SCFA production and P recovery. The results showed that the maximal SCFA yields in R1-4 were 730.2 ± 7.0, 1017.4 ± 13.9, 860.1 ± 40.8, and 1072.0 ± 33.2 mg COD/L, respectively, significantly higher than Control (365.2 ± 17.8 mg COD/L). The findings indicated that TAFP pretreatments (particularly ex-situ TAFP pretreatment) enhanced WAS disintegration and provided more soluble organics and subsequently promoted SCFA production. The P fractionation results showed the non-apatite inorganic P increased from 11.6 ± 0.2 mg P/g TSS in Control to 11.8 ± 0.5 (R1), 12.4 ± 0.3 (R2), 13.2 ± 0.7 (R3) and 12.7 ± 0.7 mg P/g TSS (R4), suggesting TAFP pretreatments improved P bioavailability due to formation of Fe-P mineral (Fe(H2PO4)2·2H2O), which could be recycled through magnetic separators. These findings were further strengthened by the analysis of microbial community and related marker genes that fermentative bacteria containing SCFA biosynthesis genes (e.g. pyk, pdhA, accA and accB) and iron-reducing bacteria containing iron-related proteins (e.g. feoA and feoB) were enriched in R1-4 (dominant in ex-situ pretreatment systems, R2 and R4). Economic evaluation further verified ex-situ TAFP pretreatment was cost-effective and a better strategy over other operations to treat WAS for SCFA production and P recovery.

Optimizing waste activated sludge disintegration by investigating multiple electrochemical pretreatment conditions: Performance, mechanism and modeling.

The complex and rigid floc structure often limits the reutilization of waste activated sludge (WAS). Electrochemical pretreatment (EPT) is one of the most effective technologies that can enhance WAS disintegration. But a comprehensive investigation into how multiple EPT conditions work was rarely reported. The study evaluated the effects of multiple EPT conditions, i.e., different electrolytes (NaCl, Na2SO4, and CaCl2), electrolytes dosage (0 g/L, 0.5 g/L, 1.0 g/L, and 3.0 g/L), EPT current (0 A, 0.5 A, 1.0 A, and 3.0 A) and EPT time (0 min, 30 min, 60 min, and 90 min) on WAS disintegration. The results showed that NaCl was outstanding from other electrolytes in promoting more WAS disintegration. Besides, a relatively higher NaCl dosage, a higher EPT current, and a longer EPT time promoted more reactive chlorine species (RCS), thus enhancing WAS disintegration in terms of extracellular polymeric substances (EPS) structure destruction and biodegradable organic matter release. After EPT for 60 min at NaCl dosage of 1.0 g/L and current of 1.0 A, the EPS multilayer structure destruction, biodegradable organic matters release, and soluble chemical oxygen demand (SCOD) increase in the supernatant were enhanced by 17.2 %, 130.5 %, and 238.7 %, respectively. Then a predictive quadratic model was established and the impact significance of the above EPT factors for enhancing WAS disintegration followed dosage of NaCl > current > EPT time. Furthermore, response surface methodology (RSM) suggested NaCl dosage of 2.75 g/L, current of 2.0 A, and EPT time of 30 min were the optimal EPT conditions, bringing a 42.0 % increase in the net economic benefit of WAS treatment compared to without EPT.

Electrochemical pretreatment enhancing co-fermentation of waste activated sludge and food waste into volatile fatty acids: Performance, microbial community dynamics and metabolism.

Waste activated sludge (WAS) has low biodegradability that restricts acidogenic fermentation (AF), thereby limiting the high-value volatile fatty acids (VFAs) production. This study investigated an alternative electrochemical pretreatment (EPT) approach that can facilitate AF of WAS and food waste (FW) and therefore enhance VFAs production. The results showed through introducing 50 % volatile solid basis of FW (containing massive chloride) into WAS, a 60-min EPT produced reactive chlorine species (RCS), which diffused into WAS-FW inner layers resulting in cell lysis, therefore significantly promoted and accelerated WAS-FW disintegration, contributing to more soluble and biodegradable dissolved organic matter (DOM). Then during the subsequent 15-day acidogenic co-fermentation (Co-AF), the residual RCS (approximate 5 mg Cl2/L) also caused acidogenic bacteria (including Prevotella_7, Lactobacillus and Veillonella) gradually outcompeted methanogens due to their different tolerance to residual RCS. Consequently, the maximum VFAs yield of the WAS-FW Co-AF with EPT was 40.8 % higher than WAS-AF without EPT.

Thiosulfate-assisted Fe2+/persulfate pretreatment effectively alleviating iron dose and enhancing biotransformation of waste activated sludge into high-value liquid products.

Ferrous-based acidogenic fermentation (AF) as a means to treat waste activated sludge (WAS) and produce short-chain fatty acids (SCFAs) has drawn increasing attention, but the massive amount of "iron sludge" that it produces not only significantly increases costs and difficulty of disposal but also poses risks to the environment and human health. This study explored a novel approach to not only reduce the iron dosage required by AF but also to improve its performance by introducing a thiosulfate (TS)-assisted Fe2+/persulfate (TAFP) pretreatment. Effects of the TAFP pretreatment on WAS disintegration and biodegradability, SCFA production, and microbial community structure with different persulfate-Fe2+-thiosulfate molar ratios at 4:4:0 (R1), 4:3:1 (R2), 4:2:2 (R3) and 4:1:3 (R4) were investigated. The results showed that the TAFP pretreatment by a remarkable margin promoted the disintegration of WAS as well as the biodegradability of the organics released, owing to the production of robust free radicals (SO4•- and •OH) triggered by the thiosulfate and Fe2+ cycles. 48-day AF tests further showed maximum SCFA production, ranging roughly between 1283 and 1395 mg COD/L in the TAFP pretreated samples, much higher than Control (<120 mg/L) and R1 (around 593 mg COD/L). At the meantime, the Fe2+ dosage was reduced by 50% in R3 than that of R1. However, a prolonged lag phase of SCFA generation was observed between days 7 and 25, which was ascribable to the acidic conditions (pH < 4.5) closely related to impaired metabolic activities as well as electron transfer efficiencies and limited activities of acidogenic enzymes (i.e., pyruvate-ferredoxin oxidoreductase). Despite such lag phase, the economic and environmental assessment of AF of TAFP-pretreated WAS had a higher net SCFA yield and less "iron sludge" than either without any pretreatment or with Fe2+/persulfate-only pretreatment.

Sex-specific pattern of left ventricular hypertrophy and diastolic function in patients with type 2 diabetes mellitus.

AIMS: Few prospective studies have evaluated sex-specific pattern, natural progression of left ventricular (LV) remodelling, and diastolic dysfunction in patients with type 2 diabetes (T2DM). The aim of this study was to study the sex-specific prevalence, longitudinal changes of LV remodelling, and diastolic dysfunction in patients with T2DM. Further, the prognostic value of diastolic function in women and men was also evaluated. METHODS AND RESULTS: A total of 350 patients with T2DM (mean age 61 ± 11 years; women, 48.3%) was recruited. Detailed echocardiography was performed at baseline and after 25 months. A major adverse cardiovascular event (MACE) was defined as cardiovascular death, heart failure hospitalization, or myocardial infarction. Despite a similar age, prevalence of hypertension and body mass index, women had a higher prevalence of LV hypertrophy and diastolic dysfunction at baseline and follow-up compared with men. A total of 21 patients developed MACE (5 cardiovascular death, 9 hospitalization for heart failure, and 7 myocardial infarction) during a median follow-up of 56 months. Women with diastolic dysfunction had a higher incidence of MACE than those with normal diastolic function but this association was neutral in men. Multivariable Cox-regression analysis indicated that diastolic dysfunction was associated with MACE in women [hazard ratio = 6.30; 95% confidence interval (CI) = 1.06-37.54; P < 0.05] but not men (hazard ratio = 2.29, 95% CI = 0.67-7.89; P = 0.19). CONCLUSION: LV hypertrophy and diastolic dysfunction, both at baseline and follow-up, were more common in women than men. Pre-clinical diastolic dysfunction was independently associated with MACE only in women with T2DM but was neutral in men.

Assessment of left ventricular function and peripheral vascular arterial stiffness in patients with dipper and non-dipper hypertension.

A non-dipper pattern of high blood pressure is associated with increased risk of organ damage and cardiovascular disease in patients with hypertension. The aim of the study was to evaluate the left ventricular (LV) remodeling and function and arterial stiffness in a dipper/non-dipper pattern of high blood pressure in patients with hypertension. A total of 183 hypertensive patients with no history of adverse cardiovascular events were divided into two groups based on 24 hours ambulatory blood pressure monitoring (ABPM): 66 patients with a dipper pattern and 117 patients with non-dipper pattern. Detailed transthoracic echocardiogram was performed and analyzed with advance speckle tracking 3-orthogonal direction strain analysis to assess LV systolic function and tissue Doppler-derived E/E' for LV diastolic function assessment. Cardio ankle vascular index (CAVI) was used to evaluate arterial stiffness. Compared with patients with dipper hypertension, those with non-dipper hypertension had increased LV mass index, higher prevalence of eccentric and concentric LV hypertrophy, more impaired LV diastolic and systolic function and peripheral arterial stiffness. Multivariable analysis revealed that a non-dipper pattern was independently associated with LV systolic dysfunction evaluated by speckle tracking-derived strain analysis. In conclusion, a non-dipper pattern of hypertension is an independent risk factor for LV systolic dysfunction. Treatment that could reverse this non-dipper pattern may reduce cardiac damage in these patients.

Characterization of capsid protein (p495) of hepatitis E virus expressed in Escherichia coli and assembling into particles in vitro.

Hepatitis E virus (HEV) is associated with acute hepatitis disease. Numerous truncated HEV capsid proteins have been successfully expressed using different expression systems. Among these, p495, a protein truncated at its N- and C-termini by 111 and 54 amino acids (aa), respectively (HEV ORF2 aa 112-606) can self-assemble into T = 1 virus-like particles (VLPs) when expressed by insect cells. A shorter p239 (aa 368-606) protein is a particulate antigen that we have previously used in our commercialized HEV vaccine, Hecolin. Here, we sought to express p495 in its soluble form (named Ep495) in E. coli and in baculovirus-infected Tn5 insect cells (named BTp495) as a back-to-back control. Characterization of p495 particles derived from these two expression systems showed similarities in particle size, morphology, and sedimentation coefficient. Antigenicity assays using a panel of anti-HEV monoclonal antibodies also showed similar strong reactivities for Ep495 and BTp495, as well as similar binding profiles that were congruent with p239. Mouse immunization results showed that Ep495 particles had comparable immunogenicity with that of BTp495 VLPs, as well as p239. Overall, our findings suggest that p495 particles produced in E. coli are ideal for the development of next-generation prophylactic vaccines against hepatitis E.

Prevalence, clinical characteristics and echocardiography parameters of non-resistant, resistant and refractory hypertension in Chinese.

OBJECTIVES: Refractory and resistant hypertension is defined as hypertension that is uncontrolled despite the use of multiple antihypertensives. The aim of the present study is to evaluate the prevalence, both in young and elderly, and the pattern of left ventricular hypertrophy (LVH) in South-Eastern Chinese patients with refractory, resistant and non-resistant hypertension. METHODS: A total of 1455 patients (age 60.3 ± 13.9, male 55.7%) with essential hypertension were recruited. Refractory hypertension was defined as uncontrolled blood pressure (systolic/diastolic, ≥140/90 mm Hg) on ≥5 antihypertensive drug classes. Resistant hypertension was defined as uncontrolled blood pressure (systolic/diastolic, ≥140/90 mm Hg) on 3 or <140/90 mm Hg on ≥4 antihypertensive classes. RESULTS: Among the total population, 1273 (87.4%) patients were considered non-resistant hypertension; 170 (11.7%) with resistant hypertension and 12 (0.8%) with refractory hypertension. The prevalence of the three groups of hypertension were similar between patients age <60 or ≥60. Patients with refractory hypertension had the most dilated LV dimension, greatest left ventricular mass index and highest prevalence of diastolic dysfunction than patients with resistant and non-resistant hypertension. In particular, all patients with refractory hypertension had either concentric or eccentric LVH. CONCLUSIONS: In South-Eastern Chinese patients, the prevalence of refractory and resistant hypertension was 0.8% and 11.7%, respectively. Furthermore, no difference was observed in the hypertensive patterns between patients age <60 and ≥60. Importantly, patients with refractory hypertension had the worst LV remodeling with all suffering from either concentric or eccentric hypertrophy.

Burden and contributing factors associated with tricuspid regurgitation: a hospital-based study.

OBJECTIVES: Tricuspid regurgitation (TR) is common in patients referred for cardiac assessment. Nonetheless, current estimates of its prevalence and contributing factors are limited. The aim of the present study was to evaluate the prevalence and demographics of TR in patients referred for echocardiography assessment at two University-affiliated hospitals. METHODS: A total of 6711 consecutive Chinese patients were recruited as part of the Chinese Valvular Heart Disease Study (CVATS). RESULTS: The most common valvular lesion was TR (54.7%), followed by mitral regurgitation (44.7%) and aortic regurgitation (26.5%). Clinically significant (moderate or severe) TR was identified in 8.4% with the proportion increased from 3.9% amongst those aged <51 to 15.9% in those aged ≥81. Multivariable adjustment demonstrated that significant TR was associated with age, congenital heart disease, chronic obstructive pulmonary disease, left-sided valvular heart disease (VHD), impaired left ventricular ejection fraction <50%, atrial fibrillation and pulmonary hypertension. CONCLUSIONS: Among all types of VHD, TR was the most common and was identified in over half of the subjects and clinically significant in 8.4%. These unique data provide contemporary clinical and epidemiological characteristics of TR in a large cohort of patients referred for cardiac assessment and confirm the increased burden of TR in the aged population.

Production of Influenza Virus HA1 Harboring Native-Like Epitopes by Pichia pastoris.

The outbreak of the H5N1 highly pathogenic avian influenza which exhibits high variation had brought a serious threat to the safety of humanity. To overcome this high variation, hemagglutinin-based recombinant subunit vaccine with rational design has been considered as a substitute for traditional virion-based vaccine development. Here, we expressed HA1 part of the hemagglutinin protein using the Pichia pastoris expression system and attained a high yield of about 120 mg/L through the use of fed-batch scalable fermentation. HA1 protein in the culture supernatant was purified using two-step ion-exchange chromatography. The resultant HA1 protein was homogeneous in solution in a glycosylated form, as confirmed by endoglycosidase H treatment. Sedimentation velocity tests, silver staining of protein gels, and immunoblotting were used for verification. The native HA1 reacted well with conformational, cross-genotype, neutralizing monoclonal antibodies, whereas a loss of binding activity was noted with the denatured HA1 form. Moreover, the murine anti-HA1 serum exhibited a virus-capture capability in the hemagglutination inhibition assay, which suggests that HA1 harbors native-like epitopes. In conclusion, soluble HA1 was efficiently expressed and purified in this study. The functional glycosylated protein will be an alternative for the development of recombinant protein-based influenza vaccine.

A shared N-terminal hydrophobic tail for the formation of nanoparticulates.

AIM: Nanoparticulate design is important for the production of nanotechnological materials and passive immunogens. Using lessons from our hepatitis E vaccine, we herein design protein-based nanoparticles through incorporation of an N-terminal hydrophobic tail (NHT, located on HEV ORF2 aa368-460). MATERIALS & METHODS: Flu HA1, HIV gp41/gp120/p24, HBsAg and HPV16 L2 were fused with NHT, expressed in Escherichia coli and subjected to self-assembly in vitro. Nanosized particles were characterized by size-exclusion chromatography and negative electron microscopy. Immunogenicity was assessed in mice. RESULTS: All the NHT-fused proteins spontaneously formed nanoparticulates and presented with immunogenicity approximately 2-log over their nonassembling forms. CONCLUSION: Protein self-assembly provides an attractive means to create nanosized particles that bear specific antigens. Our strategy outlines a novel and shared method for the design of immunogenic nanoparticles.