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As an intermediate in drug synthesis, uridine has practical applications in the pharmaceutical field. Bacillus subtilis is used as a host to boost uridine yield by manipulating its uridine biosynthesis pathway. In this study, we engineered a high-uridine-producing strain of B. subtilis by modifying its metabolic pathways in vivo. Overexpression of the aspartate ammonia-lyase (ansB) gene increased the relative transcriptional level of ansB in B. subtilis TD320 by 13·18 times and improved uridine production to 15·13 g l -1 after 72-h fermentation. Overexpression of the putative 6-phosphogluconolactonase (ykgB) gene increased uridine production by the derivative strain TD325 to 15·43 g l -1 . Reducing the translation of the amido phosphoribosyl transferase (purF) gene and inducing expression of the subtilisin E (aprE) gene resulted in a 1·99-fold increase in uridine production after 24 h shaking. Finally, uridine production in the optimal strain B. subtilis TD335, which exhibited reduced urease expression, reached 17·9 g l -1 with a yield of 314 mg of uridine g -1 glucose. To our knowledge, this is the first study to obtain high-yield uridine-producing B. subtilis in a medium containing only three components (80 g l -1 glucose, 20 g l -1 yeast powder, and 20 g l -1 urea)., (© 2022 The Society for Applied Microbiology.)

German cockroaches (Blattella germanica L.) harbor and disperse medically important pathogens and are a source of allergens that impact human health and wellbeing. Management of this pest requires an understanding of their distribution and dispersal. In this study, we collected German cockroaches from three apartment buildings in New Jersey, USA. We identified single-nucleotide polymorphisms (SNPs) from DNA extractions using next generation sequencing. We analyzed the SNPs and characterized cockroach population genetic structure using Fst, principal component, phylogenetic, and STRUCTURE analyses. We found significant differences in German cockroach population structure among the buildings. Within buildings, we found variable population structure that may be evidence for multiple colonization events. This study shows that SNPs derived from next generation sequencing provide a powerful tool for analyzing the genetic population structure of these medically important pests., (© The Author(s) 2022. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Riboflavin is a feed additive, food additive and clinical drug, with a significant annual demand of nearly 8000 t. Fermentation using recombinant Bacillus subtilis is currently one of the most important industrial production method for riboflavin. First, a suitable medium was selected and the expression of the ureABC operon was modified. The ykgB gene was overexpressed in B. subtilis RX10, the production of the derivative strain RX20 was increased to 4·61 g l -1 riboflavin, and the yield was increased to 52 mg riboflavin g -1 glucose. The relative transcription level of pyr operon in RX20 was reduced to 71%, the production of the derivative strain RX21 was increased to 5·82 g l -1 riboflavin, and the yield was 76 mg riboflavin g -1 glucose. The start codon of the pyrE gene in RX21 was modified to 'TTG', the production of the derivative strain RX22 was increased to 7·01 g l -1 riboflavin, and the yield was 89 mg riboflavin g -1 glucose. These results indicated that overexpression of the ykgB gene and reduction of the metabolic flux of de novo synthesis of pyrimidine nucleotides were beneficial to the synthesis of riboflavin., (© 2021 The Society for Applied Microbiology.)

Shewanella baltica is one of the most important bacterial species contributing to spoilage of seafood. Principally, RpoS has been recognized as the central regulator of stress resistance in many bacterial species. However, little is known about the role of RpoS in S. baltica. In this study, an rpoS mutant of S. baltica was constructed and analysed for its functions. The results showed that the survival rate of rpoS mutant decreased when treated with heat, ethanol and H 2 O 2, while increased the resistance to NaCl. Moreover RpoS promoted the biofilm formation of S. baltica at 30°C, while declined at 4°C. Interestingly, the rpoS-deficient mutant showed increased swimming motility. Furthermore, the results revealed that the production of quorum-sensing (QS) signals such as cyclo-(l-Pro-l-Leu) and cyclo-(l-Pro-l-Phe) reduced in rpoS mutant. Mainly, rpoS positively regulated QS response regulators, as the expression of all luxR genes in rpoS mutant significantly decreased relative to wild type. This study reveals that RpoS is a major regulator involved in stress responses, biofilm formation and quorum sensing system in S. baltica. The present work provides significant information for the control of microbiological spoilage of seafood., (© 2020 The Society for Applied Microbiology.)

Aim: The aim of this study was to evaluate the ability of Bacillus methylotrophicus M4-1 to protect winter wheat from the harmful effects of soil salinity and alkalinity., Methods and Results: We isolated the halotolerant B. methylotrophicus M4-1. Two representative soils with different salt contents (S1, 213 μs cm -1 ; S2, 786 μs cm -1 ) in the Yellow River delta region of China were selected for experiments. The effects of the M4-1 strain on the typical wheat variety (Jimai 21) in this environment were proven. In S1 soil, the M4-1 strain reduced the wheat rhizosphere soil pH (1·61%) and electrical conductivity (EC) (8·01%) and increased the exchangeable K content (11·14%). The uptake of Mg 2+ (20·73%) by wheat roots and K + (8·84%) by leaves was increased, and the content of Na + (23·62%) in leaves was reduced. In S2 soil, the M4-1 strain was able to reduce soil EC (2·56%) and increase exchangeable K (11·20%) content. The absorption of K + (13·28%) in wheat leaves was increased, and the content of Na + (12·41%) in roots was decreased. Total N and organic matter contents in rhizosphere soil were significantly positively correlated with wheat growth and salt tolerance, whereas EC showed a significant negative correlation., Conclusions: M4-1 attenuates salt stress injury in wheat under both low and high salt stress., Significance and Impact of the Study: We demonstrated the efficacy and value of plant growth-promoting rhizobacteria addition to protect winter wheat against salt stress and improve crop yield. We also elucidated the physicochemical and biochemical interactions among M4-1, the rhizosphere and the host plant., (© 2020 The Society for Applied Microbiology.)

Aims: To investigate the prevalence and characteristics of Salmonella in pork on sale in Wuhan, Central China., Methods and Results: A total of 4744 pork samples were collected from retail or wholesale markets in each season from 2016 to 2018. The samples showed an overall Salmonella prevalence of 19·54% (927/4744), among which the samples collected in 2017 (21·67%, 428/1975) possessed a significantly higher prevalence than those collected in 2016 (18·61%, 209/1123) (P = 0·047) or 2018 (17·51%, 290/1656) (P = 0·002), and the samples collected in winter showed the lowest prevalence (15·86%, 177/1116). The Salmonella prevalence was significantly higher among samples from retail markets (25·68%, 283/1102) than wholesale markets (17·68%, 644/3642) (P = 0·000). Antimicrobial resistance of 922 Salmonella strains was tested by determining the minimal inhibitory concentrations using a broth microdilution method. The strains revealed that 98·92% (912/922) were resistant to at least one of the antimicrobial agents, and 80·04% (738/922) were resistant to three or more antimicrobials (MDR). Resistance to sulfamethoxazole/trimethoprim (89·91%), tetracycline (87·20%) and ampicillin (71·69%) was predominant. The proportion of MDR strains in 2017 (93·62%, 396/423) was significantly higher than that in 2016 (63·16%, 132/209) (P = 0·000) or 2018 (69·66%, 202/290) (P = 0·000). No significant difference was observed in the proportions of MDR strains between wholesale markets (76·07%, 213/280) and retail markets (80·53%, 517/642) (P = 0·075). Multi-locus sequence typing for 554 of the isolates revealed 20 different sequence types (STs), among which ST40 (38·27%, 212/554), ST34 (18·41%, 102/554) and ST469 (14·46%, 79/554) were dominant., Conclusions: A high risk of Salmonella prevalence and antimicrobial resistance was observed in pork in Wuhan. The risk varies between different sampling years, seasons and market types., Significance and Impact of the Study: Providing baseline data on Salmonella contamination in pork on sale in Central China., (© 2019 The Society for Applied Microbiology.)

Despite being the fastest growing sector, the modern aquaculture industry faces serious challenges such as the lack of protein source in feed, the susceptibility to pathogens, and deterioration in quality during culture and storage. Bacterial biomass is considered as a proper protein source for feed, and the beneficial bacterial species protect aquatic animals from infection or reduce spoilage of products. In this review, we summarized the application of beneficial bacteria to aquatic products, focusing mainly on the nutritional, anti-pathogenic, anti-spoilage and immunoregulatory functions of these bacteria. We then discussed the relationship between beneficial bacteria, intestinal microbiota and host immunity, and the recent progress and drawbacks of the technology., (© 2019 The Society for Applied Microbiology.)

Aims: Genotype VII Newcastle disease (ND) is one of the most epidemic and serious infectious diseases in the poultry industry. A novel vaccine targeting VII Newcastle disease virus (NDV) is still proving elusive., Methods and Results: In this study, we constructed regulated delayed lysis Salmonella strains expressing either a fusion protein (F) alone under an eukaryotic CMV promoter or together with chicken IL-18 (chIL-18) as a molecular adjuvant under prokaryotic P trc promoter, named pYL1 and pYL23 respectively. Oral immunization with recombinant strains induced NDV-specific serum IgG antibodies in both pYL1- and pYL23-immunized chickens. The presence of chIL-18 significantly increased lymphocyte proliferation in immunized chickens, as well as the percentages of CD3 + CD4 + and CD3 + CD8 + T cells in serum, even if a statistically significant difference did not exist. After a virulent challenge, pYL23 immunization provided about 80% protection at day 10 postinfection, compared with 60% of protection offered by pYL1 immunization and 100% protection in the inactivated vaccine group, indicating the enhanced immune response provided by chIL-18, which was also confirmed by histochemical analysis., Conclusions: Recombinant lysis Salmonella-vectored DNA vaccine could provide us a novel potential option for controlling NDV infection., Significance and Impact of the Study: This study took use of a regulated delayed lysis Salmonella vector for the design of an orally administrated vaccine against NDV., (© 2019 The Society for Applied Microbiology.)

Aims: We aimed to develop a biological agent that regulates the microbial community structure of the poplar rhizosphere and alleviates the effects of continuous poplar cropping., Methods and Results: Poplar rhizosphere soils were treated with or without Paenibacillus polymyxa CP-S316 microbial fermentation medium. Real-time polymerase chain reaction was performed to measure bacteria and fungi in both groups, and microbial communities were analysed by metabarcoding. In fungi, the operational taxonomic units, abundance-based coverage estimator and Chao index of the CP-S316-treated group were significantly lower than those in the control check (CK) group. In bacteria, the proportions of Bacillus in the CP-S316 and CK groups were 5·20 and 2·38%, respectively, whereas those of Rhizoctonia were 2·20 and 5·82% respectively. The diameter at breast height, culturable bacteria and total bacteria of poplars treated with CP-S316 exceeded those in the CK group., Conclusions: Our data confirmed that CP-S316 could improve the microbial community structure of poplar rhizosphere and promote the growth of poplars., Significance and Impact of the Study: Research aimed at alleviating continuous cropping obstacles and promoting poplar growth via biocontrol agents is uncommon. We analysed the community structures of bacteria and fungi in rhizosphere soil to illustrate the use of CP-S316 for poplar cropping for improving plant health in the continuous cropping of poplar trees., (© 2019 The Society for Applied Microbiology.)

Aim: The aim of this study was to identify efficient plant-beneficial rhizobacterium that has the potential to be developed as biocontrol agent for the control of wheat soil-borne diseases., Methods and Results: Rhizosphere soil samples were collected from a wheat field located in Taian City. Numerous bacteria were isolated and screened for antagonistic activity against soil-borne plant pathogenic fungi by performing dual-culture assays. Among them, XH-9 was selected for its highly antagonistic activity and others growth-promoting characteristics. Subsequently, the strain was identified as Bacillus amyloliquefaciens subsp. plantarum based on phylogenetic analysis of 16S rDNA sequence. Pot experiment indicated that XH-9 has good capacities for wheat, corn, and chili root colonization and considerably increased the biometric parameters of wheat seedlings. Quantitative real-time polymerase chain reaction experiments showed that the amount of Fusarium oxysporum associated with the XH-9 after treatment significantly decreased compared with control group., Conclusions: Bacillus amyloliquefaciens subsp. plantarum XH-9 has the potential as biocontrol agent when applied in local arable land to prevent damage caused by F. oxysporum and other phytopathogens., Significance and Impact of the Study: The development of biocontrol strategies for reducing the damage caused by plant pathogens is fully in accord with the current principles of sustainability., (© 2018 The Society for Applied Microbiology.)

Aim: To investigate the volatile flavour components (VFCs), microbiota and their correlations of three categories of sufu, a Chinese fermented soybean food., Methods and Results: The VFCs were analyzed by headspace solid-phase micro-extraction (HS-SPME) combined with gas chromatography and mass spectrometry. A total of 141 VFCs were tentatively identified, in which 12 esters, eight aldehydes, five alcohols and nine miscellaneous compounds were recognized as the characteristic VFCs through the relative odour activity value. The complex microbial community was revealed by Illumina MiSeq sequencing. Among the total 202 bacteria and 125 fungi species identified, 16 bacteria and 7 fungi species were revealed as dominant community members. Furthermore, the VFC-microbiota correlation was characterized by Spearman's rank correlation coefficients (ρ). Based on the positive VFC-microbiota correlation, two bacterial species and three fungal species were selected as potential flavour-producing microbiota., Conclusions: A variety of VFCs and complicated microbiota were observed in the three categories of sufu. Lactococcus lactis, Sphingobacterium sp., Pichia fermentans, Kodamaea ohmeri and Saccharomyces rouxii were the potential flavour-producing microbiota., Significance and Impact of the Study: The preliminary results would be useful for designing starter cultures to produce sufu with desirable sensory properties, consistent flavour and shorter ripening time. Furthermore, these results will also provide a new insight to improve the flavour quality of traditional fermented soybean food., (© 2018 The Society for Applied Microbiology.)

Aims: This study investigates pinewood nematode's impacts on root mycoflora of Pinus tabulaeformis., Methods and Results: The biomass, colonization rate, community structure and diversity of root-associated fungi were investigated in pinewood nematode-infected and nematode-noninfected P. tabulaeformis. The results indicated that the roots of P. tabulaeformis were colonized highly by root-associated fungi, mainly ectomycorrhizal fungi (ECMF) and dark septate endophytes. Infection of pinewood nematode was associated with a significant (P < 0·05) decrease in root colonization rates by ECMF, dark septate endophytes and total hyphae, as well as in fungal biomass in the roots. Illumina MiSeq sequences of tagged amplicons of 18S rDNA region revealed Basidiomycota (65·70%) and Ascomycota (34·14%) as the dominant root-associated fungi in roots of P. tabulaeformis. Among the detected operational taxonomic units (OTUs), ECMF and dark septate endophytes exhibited a higher relative abundance in trees infected by pinewood nematode compared with noninfected ones., Conclusions: The infection of pinewood nematode altered the composition and OTU abundance of root-associated fungi community in P. tabulaeformis roots with a decrease in the biomass, species richness and diversity of root-associated fungi, as well as in the colonization rates and abundance of ECMF and dark septate endophytes., Significance and Impact of the Study: This study is an important contribution for better understanding the interaction between pine wilt disease and root-associated fungi., (© 2018 The Society for Applied Microbiology.)

Background: Few studies have reported the clinical features, complications and predictors of Kasabach-Merritt phenomenon (KMP) associated with Kaposiform haemangioendothelioma (KHE)., Objectives: To determine the clinical characteristics present at diagnosis and to identify features that may aid clinicians in managing KHE., Methods: We conducted a cohort study of 146 patients diagnosed with KHE., Results: KHE precursors or lesions were present at birth in 52·1% of patients. In 91·8% of patients, lesions developed within the first year of life. The median age at diagnosis of KHE was 2·3 months (interquartile range 1·0-6·0). The extremities were the dominant location, representing 50·7% of all KHEs. Among KHEs in the cohort, 63·0% were mixed lesions (cutaneous lesions with deep infiltration). Approximately 70% of patients showed KMP. A KHE diagnosis was delayed by ≥ 1 month in 65·7% of patients with KMP. Patients with KMP were more likely to have major complications than patients without KMP (P = 0·023). Young age (< 6 months), trunk location, large lesion size (> 5·0 cm) and mixed lesion type were associated with KMP in a univariate analysis. In the multivariate analysis, only age [odds ratio (OR) 11·9, 95% confidence interval (CI) 4·07-34·8; P < 0·001], large lesion size (OR 5·08, 95% CI 2·24-11·5; P < 0·001) and mixed lesion type (OR 2·96, 95% CI 1·23-7·13; P = 0·016) were associated with KMP., Conclusions: Most KHEs appeared before 12 months of age. KHEs are associated with various major complications, which can occur in combination and develop early in the disease process. Young age, large lesion size and mixed lesion type are important predictors of KMP., (© 2018 British Association of Dermatologists.)

Increasing meat consumption by Chinese people has created a focus for improving meat quality for increasing quality of life. Twenty-five Male Oula sheep were selected at 2, 21, 56, 84, and 112 d to investigate the developmental changes associated with age on the intramuscular fat (IMF) content of heart fatty acid-binding protein (H-FABP) mRNA expression in muscle. Longissimus dorsal muscle and biceps femoris muscle were sampled to measure IMF concentrations and total mRNA was extracted to measure H-FABP mRNA expression using real-time polymerase chain reaction (PCR). Growing male Oula sheep demonstrated that the IMF concentrations continuously increased with age and significant differences ( P < 0.05) were detected among the age groups; 2. The IMF concentrations among tissues were different; 3. The development changes in H-FABP mRNA expression in longissimus dorsal muscle and biceps femoris muscle were similar with a decrease from 2 to 21 d, followed by continuously increasing concentrations being significant different ( P < 0.05) among age groups; 4. The H-FABP mRNA expression in the longissimus dorsal muscle tissue was significantly ( P < 0.05) higher compared to the biceps femoris muscle; 5. The muscle H-FABP mRNA expression concentration was positively correlated with IMF concentrations from d 21 to 112; 6. The correlation coefficients were significantly ( P < 0.01)different between H-FABP gene mRNA expression in the longissimus dorsal muscle and IMF concentration of 0.815 compared to the biceps femoris muscle and IMF concentration of 0.787,which indicated that the H-FABP gene may be affecting the IMF concentrations in the early developmental stages of Oula sheep. These results support the hypothesis that H-FABP gene and its expression in muscle tissue is related to the IMF concentration of meat.

Esophageal squamous cell carcinoma (ESCC) is one of the most frequently diagnosed cancers in China, but the etiology and mode of carcinogenesis of this disease remain poorly understood. Phosphatase and tensin homolog deleted on chromosome 10 (PTEN), as a negative regulator of Akt/mTOR pathway, frequently mutates or is inactive in many cancers. Although mTOR has been thought a promising cancer therapeutic target, the sensitivity of tumor cells to rapamycin was still to be revaluated. In this study, we measured the effects of rapamycin on cell proliferation and phosphorylation of Akt in ESCC cells with varying degrees of differentiation. And then, the relationship between PTEN status and the sensitivity of cells to rapamycin was investigated in EC9706 cells with or without wild-type PTEN in vitro and in vivo. The results demonstrated ESCC cells with poor differentiation were insensitive to rapamycin of high concentration and rapamycin obviously promoted the phosphorylation of Akt in these cells, but it had no obvious effects on p-Akt in cells with well differentiation. Also, we showed that wild-type PTEN improved the sensitivity of poor differentiation cells to rapamycin through inhibiting phosphorylation of Akt in vitro and in vivo. This study explored the possible molecular mechanism of some ESCC cells insensitive to rapamycin and provided a measure for treating ESCC patients with PTEN inactivation using mTOR inhibitors., (© 2015 International Society for Diseases of the Esophagus.)

Background: Despite unmet need, 15 years have passed since a topical therapy with a new mechanism of action for atopic dermatitis (AD) has been approved. Janus kinase (JAK) inhibitor treatment effect via topical application in patients with AD is unknown., Objectives: Tofacitinib, a small-molecule JAK inhibitor, was investigated for the topical treatment of AD., Methods: In this 4-week, phase IIa, randomized, double-blind, vehicle-controlled study (NCT02001181), 69 adults with mild-to-moderate AD were randomized 1:1 to 2% tofacitinib or vehicle ointment twice daily. Percentage change from baseline (CFB) in Eczema Area and Severity Index (EASI) score at week 4 was the primary end point. Secondary efficacy end points included percentage CFB in body surface area (BSA), CFB in EASI Clinical Signs Severity Sum Score, proportion of patients with Physician's Global Assessment (PGA) response and CFB in patient-reported pruritus. Safety, local tolerability and pharmacokinetics were monitored., Results: The mean percentage CFB at week 4 in EASI score was significantly greater (P < 0·001) for tofacitinib (-81·7%) vs. vehicle (-29·9%). Patients treated with tofacitinib showed significant (P < 0·001) improvements vs. vehicle across all prespecified efficacy end points and for pruritus at week 4. Significant improvements in EASI, PGA and BSA were observed by week 1 and improvements in pruritus were observed by day 2. Safety/local tolerability were generally similar for both treatments, although more adverse events were observed for vehicle vs. tofacitinib., Conclusions: Tofacitinib ointment showed significantly greater efficacy vs. vehicle across end points, with early onset of effect and comparable safety/local tolerability to vehicle. JAK inhibition through topical delivery is potentially a promising therapeutic target for AD., (© 2016 The Authors. British Journal of Dermatology published by John Wiley & Sons Ltd on behalf of British Association of Dermatologists.)

We conducted a retrospective analysis to assess the toxicity and long-term survival of esophageal squamous cell carcinoma patients treated with three-dimensional conformal radiotherapy (3DCRT) or intensity-modulated radiotherapy (IMRT) versus conventional two-dimensional radiotherapy (2DRT). All data in the present study were based on four prospective clinical trials conducted at our institution from 1996 to 2004 and included 308 esophageal squamous cell carcinoma patients treated with 2DRT or 3DCRT/IMRT. Based on the inclusion and exclusion criteria, 254 patients were included in the analysis. Of these patients, 158 were treated with 2DRT, whereas 96 were treated with 3DCRT/IMRT. The rates of ≥Grade3 acute toxicity of the esophagus and lung were 11.5% versus 28.5% (P = 0.002) and 5.2% versus 10.8% (P = 0.127) in the 3DCRT/IMRT and 2DRT groups, respectively. The incidences of ≥Grade 3 late toxicity of the esophagus and lungs were 3.1% versus 10.7% (P = 0.028) and 3.1% versus 5.7% (P = 0.127) in the 3DCRT/IMRT and 2DRT groups, respectively. The 1-year, 3-year and 5-year estimated overall survival rates were 81%, 38% and 34% in the 3DCRT/IMRT group and 79%, 44% and 31% in the 2DRT group, respectively (P = 0.628). The 1-year, 3-year and 5-year local control rates were 88%, 71% and 66% in the 3DCRT/IMRT group and 84%, 66% and 60% in the 2DRT group, respectively (P = 0.412). Fewer incidences of acute and late toxicities were observed in esophageal squamous cell carcinoma patients treated with 3DCRT/IMRT compared with those treated with 2DRT. No significant survival benefit was observed with the use of 3DCRT/IMRT., (© 2015 International Society for Diseases of the Esophagus.)

The objective of this study was to investigate the prognostic value of peripheral blood monocytes in esophageal squamous cell carcinoma (ESCC) patients who underwent esophagectomy. Records from 218 consecutive patients with histologically diagnosed ESCC who underwent esophagectomy at Qilu Hospital of Shandong University from January 2007 to December 2008 were retrospectively reviewed. The median disease-free survival (DFS) of this cohort was 29.0 months, and the 5-year DFS rate was 34.4%. The median overall survival (OS) was 35.0 months, and the 5-year OS rate was 37.6%. The cut-off value of 0.42 × 10 9 /L for the absolute monocyte count (AMC) was chosen as optimal to discriminate between survival and death by applying receiver operating curve analysis. There were 131 patients (60.1%) who had high AMC (≥0.42 × 10 9 /L) preoperatively. We found that AMC was significantly associated with gender, tumor location, and platelet count. Kaplan-Meier survival analysis of patients with high preoperative AMC had a significant worse prognosis for DFS (high vs. low: 27.5% vs. 39.0%, P = 0.015) and OS (high vs. low: 31.1% vs. 44.8%, P = 0.009) than those with low preoperative AMC. In a multivariate analysis, preoperative AMC was an independent prognostic factor for DFS (P = 0.025, hazard ratio [HR]: 1.469, 95% confidence interval [CI]: 1.050-2.054) and OS (P = 0.015, HR: 1.547, 95% CI: 1.088-2.200). In addition, among 140 patients without both preoperative and postoperative therapy, significantly worse OS (P = 0.012) and marginally reduced DFS (P = 0.079) were found in the high AMC cohort versus the low AMC cohort. A higher preoperative absolute peripheral monocyte count can be considered as a useful prognostic marker of ESCC patients who underwent esophagectomy., (© 2015 International Society for Diseases of the Esophagus.)

Context: Denosumab and zoledronic acid (ZOL) are parenteral treatments for patients with osteoporosis., Objective: The objective of the study was to compare the effect of transitioning from oral bisphosphonates to denosumab or ZOL on bone mineral density (BMD) and bone turnover., Design and Setting: This was an international, multicenter, randomized, double-blind trial., Participants: A total of 643 postmenopausal women with osteoporosis previously treated with oral bisphosphonates participated in the study., Interventions: Subjects were randomized 1:1 to sc denosumab 60 mg every 6 months plus iv placebo once or ZOL 5 mg iv once plus sc placebo every 6 months for 12 months., Main Outcome Measures: Changes in BMD and bone turnover markers were measured., Results: BMD change from baseline at month 12 was significantly greater with denosumab compared with ZOL at the lumbar spine (primary end point; 3.2% vs 1.1%; P < .0001), total hip (1.9% vs 0.6%; P < .0001), femoral neck (1.2% vs -0.1%; P < .0001), and one-third radius (0.6% vs 0.0%; P < .05). The median decrease from baseline was greater with denosumab than ZOL for serum C-telopeptide of type 1 collagen at all time points after day 10 and for serum procollagen type 1 N-terminal propeptide at month 1 and at all time points after month 3 (all P < .05). Median percentage changes from baseline in serum intact PTH were significantly greater at months 3 and 9 with denosumab compared with ZOL (all P < .05). Adverse events were similar between groups. Three events consistent with the definition of atypical femoral fracture were observed (two denosumab and one ZOL)., Conclusions: In postmenopausal women with osteoporosis previously treated with oral bisphosphonates, denosumab was associated with greater BMD increases at all measured skeletal sites and greater inhibition of bone remodeling compared with ZOL.

Background: Dermal microvasculature expansion and angiogenesis are prominent in psoriasis. Our previous microarray study showed that the angiogenesis-related genes EDIL3 (epidermal growth factor-like repeats and discoidin I-like domains 3), AMOT (angiomotin) and ECM1 (extracellular matrix protein 1), had high expression levels in dermal mesenchymal stem cells (DMSCs) from psoriatic skin lesions., Aim: To investigate the mRNA and protein expressions of EDIL3, AMOT and ECM1 in DMSCs derived from psoriatic skin in order to better determine the molecular mechanisms of angiogenesis in the skin., Methods: DMSCs from 12 patients with psoriasis and 14 healthy controls (HCs) were cultured to passage 3, and identified by morphology, immunophenotype and multipotential differentiation. The mRNA and protein expressions of EDIL3, AMOT, and ECM1 in the DMSCs were determined using real-time reverse transcription PCR and western blotting., Results: DMSCs displayed spindle-like morphology and surface protein expression, and were able to differentiate into osteoblasts, chondrocytes and adipocytes. mRNA expression analysis showed that EDIL3, AMOT and ECM1 were expressed at 2.54-fold, 2.11-fold, and 1.90-fold higher levels, respectively, in psoriatic DMSCs compared with HC DMSCs (all P < 0.05). Protein analysis showed significantly (all P < 0.01) higher concentrations of EDIL3, AMOT and ECM1in the psoriasis group than in the HC group., Conclusions: Our data demonstrate for the first time that expression of EDIL3, AMOT and ECM1 is altered in DMSCs in psoriasis, suggesting that EDIL3, AMOT and ECM1 are involved in the excessive angiogenesis and vasodilation observed in psoriasis., (© 2015 British Association of Dermatologists.)

Unlabelled: Syphilis is a multistage sexually transmitted disease that remains a serious public health concern worldwide. The coexistence of Treponema pallidum with other closely related members of spirochaeta, such as Leptospira spp. and Borrelia burgdorferi, has complicated the serodiagnosis due to cross-reactive antigens. In this study, recombinant Tp0821 protein was expressed in Escherichia coli and purified by metal affinity chromatography. Then enzyme-linked immunosorbent assays (ELISAs) based on Tp0821 for the detection of specific antibodies were established. The relative positive rates of the IgM ELISA and the IgG ELISA were found to be 91·0 and 98·3%, respectively, when screening 578 syphilis specimens. The specificities were 94·3 and 100%, respectively, when cross-checking with serum samples obtained from 30 patients with Lyme disease, five patients with leptospirosis, and 52 uninfected controls. In addition, relative positive rates and specificities of Tp0821 for human sera were all 100% in Western blotting. When compared to the syphilis diagnostic tests commonly used in clinical settings, we found that the results of Tp0821-based ELISAs correlated well with the results of the treponemal tests, specifically the T. pallidum particle agglutination (TP-PA) test and the chemiluminescent immunoassay (CIA). Thus, these findings identify Tp0821 as a novel serodiagnostic candidate for syphilis., Significance and Impact of the Study: In this study, we expressed and purified the Treponema pallidum protein Tp0821 and developed Tp0821-based enzyme-linked immunosorbent assays (ELISAs) for the detection of specific antibodies. The serodiagnostic performance of the recombinant protein was then evaluated. When compared to the results of syphilis diagnostic tests commonly used in clinical settings, we found that the reactivities of syphilitic sera with the recombinant antigen correlated well with the results of the treponemal tests, specifically the T. pallidum particle agglutination (TP-PA) test and the chemiluminescent immunoassay (CIA). Thus, the recombinant protein shows promise as a new diagnostic antigen in the ELISAs., (© 2016 The Society for Applied Microbiology.)

Corynespora cassiicola is a plant pathogen associated with leaf-spotting disease. The fungus has been found on diverse substrates: leaves, stems and roots of plants; nematode cysts and human skin. It rarely causes human infections. Here we report one case of subcutaneous phaeohyphomycosis caused by C. cassiicola with prominent tissue necrosis in a woman. All of her clinical features pointed towards a genetic linkage. Hence, whole-exome sequencing and Sanger sequencing were performed on this patient. One mutation of CARD9 was detected., (© 2015 British Association of Dermatologists.)

Background: Previous studies of drug resistance have shown that mutations in the drug resistance-determining region (DRDR) in the Folp1, RpoB and GyrA genes of Mycobacterium leprae are responsible for resistance to dapsone, rifampin and ofloxacin, respectively., Aim: To investigate the prevalence of mutations in genes associated with drug resistance in M. leprae isolates from patients with leprosy in Shandong Province., Methods: The DRDR in the FolP1, RpoB and GyrA genes was analysed by direct sequencing of the PCR product from 85 isolates of M. leprae sampled from patients with leprosy in Shandong, China., Results: Sequencing results were obtained for FolP1, RpoB and GyrA in 67, 57 and 81 of the 85 samples, with mutation rates of 1.5% (1/67), 8.8% 5/57 and 25.9% (21/81). Three multidrug-resistant samples were found among the new cases: one had a mutation in both Folp1 and RpoB, while the other two had a mutation in both RpoB and GyrA., Conclusions: Primary resistance appears to be to either single drugs or combinations of two drugs. The resistance rate to dapsone seems to be low. To our knowledge, this is the first case of multidrug-resistant M. leprae from China., (© 2015 British Association of Dermatologists.)

Unlabelled: A pentachloronitrobenzene (PCNB)-degrading bacterial strain was identified as Arthrobacter nicotianae based on morphological, physiological and biochemical tests, and 16S rRNA gene sequences analysis. The strain could grow on mineral salt medium (MSM) containing PCNB as the sole carbon source. Under optimal conditions (pH 6·85, 30°C and inoculum biomass amount of 1·45 g l(-1) ), more than 90% residues of PCNB were degraded by strain DH19 within 7 days. Strain DH19 could efficiently degrade dichlorodiphenyl trichloroethane, hexachlorocyclohexane, cypermethrin and cyhalothrin. Metabolites from PCNB degradation were identified by using gas chromatography (GC) coupled with triple quadrupole mass spectrometry. A possible degradation pathway for PCNB was deduced. This is the first report of PCNB-degrading strain DH19 isolated from the rhizospheric soil. Therefore, strain DH19 could potentially be employed in bioremediation of PCNB., Significance and Impact of the Study: The degradation of pentachloronitrobenzene (PCNB) by an individual bacterial strain is being reported for the first time. The efficient PCNB-degrading strain DH19 was isolated from ginseng rhizosphere soil and identified as Arthrobacter nicotianae. The strain could utilize PCNB as the sole carbon source for growth and degradation. In addition, strain DH19 could efficiently degrade dichlorodiphenyl trichloroethane, hexachlorocyclohexane, cypermethrin and cyhalothrin. Five metabolites from PCNB degradation were identified, and a possible degradation pathway was deduced. The results suggest that A. nicotianae DH19 has great potential for the bioremediation of PCNB-contaminated environments., (© 2015 The Society for Applied Microbiology.)

The interaction between neutrophils and activation of alternative complement pathway plays a pivotal role in the pathogenesis of anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). ANCAs activate primed neutrophils to release neutrophil extracellular traps (NETs), which have recently gathered increasing attention in the development of AAV. The relationship between NETs and alternative complement pathway has not been elucidated. The current study aimed to investigate the relationship between NETs and alternative complement pathway. Detection of components of alternative complement pathway on NETs in vitro was assessed by immunostain and confocal microscopy. Complement deposition on NETs were detected after incubation with magnesium salt ethyleneglycol tetraacetic acid (Mg-EGTA)-treated human serum. After incubation of serum with supernatants enriched in ANCA-induced NETs, levels of complement components in supernatants were measured by enzyme-linked immunosorbent assay (ELISA). Complement factor B (Bb) and properdin deposited on NETs in vitro. The deposition of C3b and C5b-9 on NETs incubated with heat-inactivated normal human serum (Hi-NHS) or EGTA-treated Hi-NHS (Mg-EGTA-Hi-NHS) were significantly less than that on NETs incubated with NHS or EGTA-treated NHS (Mg-EGTA-NHS). NETs induced by ANCA could activate the alternative complement cascade in the serum. In the presence of EGTA, C3a, C5a and SC5b-9 concentration decreased from 800·42 ± 244·81 ng/ml, 7·68 ± 1·50 ng/ml, 382·15 ± 159·75 ng/ml in the supernatants enriched in ANCA induced NETs to 479·07 ± 156·2 ng/ml, 4·86 ± 1·26 ng/ml, 212·65 ± 44·40 ng/ml in the supernatants of DNase I-degraded NETs (P < 0·001, P = 0·008, P < 0·001, respectively). NETs could activate the alternative complement pathway, and might thus participate in the pathogenesis of AAV., (© 2015 British Society for Immunology.)

Aims: To reveal the effects of fish genotype, feeding habits and serum physiological index on the composition of gastrointestinal microbiota, eight fish species with four different feeding habits were investigated., Methods and Results: The V1 to V3 regions of 16S rRNA gene were analysed by high-throughput sequencing (454 platform) to compare the gut microbiota of different fish species. A total of 551 995 high-quality sequences with an average length of 463 bp were obtained from the 48 samples. No significant difference was observed among the detected sequences obtained from fishes with different feeding habits (One-way anova, F = 1·003, P = 0·400), but the number of OTUs among different feeding habits was significantly different (One-way anova, F = 7·564, P < 0·001). Additionally, significant correlations were detected between the fish genotype and microbial composition (partial Mantel test, all P values = 0·001) in the stomach, foregut and hindgut. Moreover, different core intestinal microbiota was also noticed in the eight fish species with different feeding habits., Conclusions: Feeding habits and genotype clearly affected the gastrointestinal microbiota of fish. Moreover, the evolutionary process shaped the serum physiological indexes of fish., Significance and Impact of the Study: This study provided much important information for developing commercial fish feeds., (© 2014 The Society for Applied Microbiology.)

Unlabelled: In this study, we developed and validated a loop-mediated isothermal amplification (LAMP) assay for Salmonella detection targeting bcfD gene, a conserved fimbrial operon gene existing in Salmonella. The Salmonella LAMP assay we developed successfully amplified 44 Salmonella strains (14 standard strains and 30 clinical isolates), but none of 9 non-Salmonella standard strains (Proteus mirabilis, Listeria monocytogenes, Escherichia coli, Pseudomonas aeruginosa, Shigella flexneri, Shigella sonnei, Klebsiella pneumoniae, Campylobacter jejuni and Vibrio parahemolyticus). The detection limit was 5 CFU of Salmonella pure culture or 200 CFU of artificially spiked faeces per reaction system (equivalent to 5000 CFU g(-1) of faeces), and this method could directly detect Salmonella in chicken faeces free of pre-enrichment in a reaction time of 25 min. Our experiments show that the LAMP method we developed is a rapid, sensitive, specific and practical method for Salmonella detection. The Salmonella LAMP assay can potentially serve as new on-site diagnostics in the food and agricultural industries., Significance and Impact of the Study: A loop-mediated isothermal amplification (LAMP) assay was established to detect Salmonella targeting bcfD gene, a conserved fimbrial operon gene. The detection limit was 5 CFU of Salmonella pure culture or 200 CFU of artificially spiked faeces per reaction system (equivalent to 5000 CFU g(-1) of faeces), and this method could directly detect Salmonella in chicken faeces free of pre-enrichment in a reaction time of 25 min. The Salmonella LAMP assay is a rapid, sensitive, specific and practical method for Salmonella detection and can potentially serve as new on-site diagnostics in the food and agricultural industries., (© 2014 The Society for Applied Microbiology.)

Aims: To evaluate the potential effect of matrine on reducing the growth of hypha and lowering the resistance of a fluconazole-resistant colony of Candida albicans., Methods and Results: Candida albicans SC5314 and a fluconazole-resistant C. albicans 215 were used. As for C. albicans SC5314, minimal inhibitory concentration (MIC(80)) and effective concentration (EC(50)) were determined, 1 mg ml(-1) matrine could inhibit nearly 80% of planktonic growth by inverted microscope, 2 mg ml(-1) matrine suppressed 50% of metabolic activity of biofilm by XTT assay, vanishing hypha could be observed on spider agar containing 2 mg ml(-1) matrine, the expressions of three hypha-related genes, namely ALS 3, SUN 41 and PBS 2, were suppressed by 29, 45 and 61% by 2 mg ml(-1) matrine. Also, matrine could lower the resistance of C. albicans 215, in either the free-floating form or the biofilm phenotype., Conclusions: Matrine had favourable antifungal potential and might be able to reverse the fluconazole resistance of clinical isolates at relatively high concentration. The anti-candidal performance of matrine could be tightly associated with yeast-to-hypha transition proved by spider agar test and qRT-PCR., Significance and Impact of Study: More efforts are needed to find new antifungal agents. Matrine could be a potential candidate to fight against Candida-related infections by regulating yeast-to-hypha transition., (© 2014 The Society for Applied Microbiology.)

Unlabelled: The objective of this study was to evaluate the potential synergistic effect of ethanol on a combination of orthophthalaldehyde (OPA) and didecyldimethylammonium chloride (DDAC) against the spores of Bacillus subtilis var. Niger. The quantitative carrier test for sporicidal testing of high-level disinfectants according to the guideline of China (Technical Standard for Disinfection 2002) was used as method. Considerable synergistic effect was observed after a 30-min treatment at 20°C. There was an augment in mean log reduction as the concentration of DDAC was increased ranging from 0·2 to 3 g l(-1) in combination with 6 g l(-1) OPA. Ten and 20% ethanol in combination with 6 g l(-1) OPA and 2 g l(-1) DDAC caused more than a 3-log reduction while either 6 g l(-1) OPA, 2 g l(-1) DDAC and 20% ethanol alone or a combination of two of the three agents produced less than a 1-log reduction. Further, 40-min exposure time of combination of OPA, DDAC and 20% ethanol led to greater than a 5-log reduction in spores, and no spore growth was observed following 60- and 90-min exposures., Significance and Impact of the Study: Orthophthalaldehyde (OPA) is very effective at concentrations far lower than its recommended in-use concentration of 0·5% (w/v) and is equally effective against both the gram-negative and gram-positive bacteria. However, it shows lower activity against spores. The synergistic sporicidal effect exhibited by ethanol on a combination of OPA and DDAC can be considered to enhance sporicidal activity for using in situations of sterilization, to reduce in-use concentration of OPA used alone, which may minimize its side effect. OPA may be a more satisfactory and the first-choice agent to replace glutaraldehyde (GTA) as a high-level disinfectant for medical devices., (© 2014 The Society for Applied Microbiology.)

Aryl hydrocarbon receptor (AhR) is well known for mediating the toxic effects of dioxin-containing pollutants, but has also been shown to be involved in the natural regulation of the immune response. In this study, we investigated the effect of AhR activation by its endogenous ligands 6-formylindolo[3,2-b]carbazole (FICZ) and 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) on the differentiation, maturation and function of monocyte-derived DCs in Behçet's disease (BD) patients. In this study, we showed that AhR activation by FICZ and ITE down-regulated the expression of co-stimulatory molecules including human leucocyte antigen D-related (HLA-DR), CD80 and CD86, while it had no effect on the expression of CD83 and CD40 on DCs derived from BD patients and normal controls. Lipopolysaccharide (LPS)-treated dendritic cells (DCs) from active BD patients showed a higher level of interleukin (IL)-1β, IL-6, IL-23 and tumour necrosis factor (TNF)-α production. FICZ or ITE significantly inhibited the production of IL-1β, IL-6, IL-23 and TNF-α, but induced IL-10 production by DCs derived from active BD patients and normal controls. FICZ or ITE-treated DCs significantly inhibited the T helper type 17 (Th17) and Th1 cell response. Activation of AhR either by FICZ or ITE inhibits DC differentiation, maturation and function. Further studies are needed to investigate whether manipulation of the AhR pathway may be used to treat BD or other autoimmune diseases., (© 2014 British Society for Immunology.)

Nonalcoholic fatty liver disease is common in developed countries and is associated with obesity, metabolic syndrome, and type 2 diabetes. T deficiency is a risk factor for developing these metabolic deficiencies, but its role in hepatic steatosis has not been well studied. We investigated the effects of T on the pathogenesis of hepatic steatosis in rats fed a high-fat diet (HFD). Adult male rats were randomly placed into four groups and treated for 15 weeks: intact rats on regular chow diet (RCD), intact rats on liquid HFD (I+HFD), castrated rats on HFD (C+HFD), and castrated rats with T replacement on HFD (C+HFD+T). Fat contributed 71% energy to the HFD but only 16% of energy to the RCD. Serum T level was undetectable in castrated rats, and T replacement led to 2-fold higher mean serum T levels than in intact rats. C+HFD rats gained less weight but had higher percentage body fat than C+HFD+T. Severe micro- and macrovesicular fat accumulated in hepatocytes with multiple inflammatory foci in the livers of C+HFD. I+HFD and C+HFD+T hepatocytes demonstrated only mild to moderate microvesicular steatosis. T replacement attenuated HFD-induced hepatocyte apoptosis in castrated rats. Serum glucose and insulin levels were not increased with HFD in any group. Immunoblots showed that insulin-regulated proteins were not changed in any group. This study demonstrates that T deficiency may contribute to the severity of hepatic steatosis and T may play a protective role in hepatic steatosis and nonalcoholic fatty liver disease development without insulin resistance.

Background: This meta-analysis aimed to investigate whether preoperative biliary drainage (PBD) is beneficial to patients with obstructive jaundice., Methods: Data from randomized clinical trials related to safety and effectiveness of PBD versus no PBD were extracted by two independent reviewers. Risk ratios, rate ratios or mean differences were calculated with 95 per cent confidence intervals (c.i.), based on intention-to-treat analysis, whenever possible., Results: Six trials (four using percutaneous transhepatic biliary drainage and two using endoscopic sphincterotomy) including 520 patients with malignant or benign obstructive jaundice comparing PBD (265 patients) with no PBD (255) were included in this review. All trials had a high risk of bias. There was no significant difference in mortality (risk ratio 1.12, 95 per cent c.i. 0.73 to 1.71; P = 0.60) between the two groups. Overall serious morbidity (grade III or IV, Clavien-Dindo classification) was higher in the PBD group (599 complications per 1000 patients) than in the direct surgery group (361 complications per 1000 patients) (rate ratio 1.66, 95 per cent c.i. 1.28 to 2.16; P < 0.001). Quality of life was not reported in any of the trials. There was no significant difference in length of hospital stay between the two groups: mean difference 4.87 (95 per cent c.i. -1.28 to 11.02) days (P = 0.12)., Conclusion: PBD in patients undergoing surgery for obstructive jaundice is associated with similar mortality but increased serious morbidity compared with no PBD. Therefore, PBD should not be used routinely., (© 2013 British Journal of Surgery Society Ltd. Published by John Wiley & Sons Ltd.)

Aims: To understand the effects of Trp residues in linear antimicrobial peptides with α-helical conformations on cell permeation ability and membrane transduction efficacy., Methods and Results: A series of L-K6 analogues were designed and synthesized by replacing Ile or Leu with Trp at different positions on the hydrophobic face of L-K6. The antimicrobial and haemolytic activity and secondary structure of the designed Trp-containing peptides were assessed. In addition, the role of Trp in membrane disruption for these designed peptides was investigated. I1W, I4W and L5W demonstrated stronger activity than the other peptides against both Gram-positive and Gram-negative bacteria. All of the tested peptides preferentially interacted with negatively charged vesicles composed of phosphatidylglycerol (PG)/cardiolipin (CL) or PG/CL/phosphatidylethanolamine, and, to a lesser extent, with zwitterionic vesicles. I1W, I4W and L5W caused calcein release at 2·5 μmol l(-1) ., Conclusions: The position of Trp, rather than the number of Trp residues, in these peptides was an important factor in the antimicrobial activity. Trp residues were deeply inserted into negatively charged membranes but were largely exposed in aqueous buffer solution., Significance and Impact of the Study: These Trp-containing peptides may represent good candidates for new antibiotic agents and for use in new therapeutic approaches., (© 2013 The Society for Applied Microbiology.)

Aims: To examine how storage temperatures influence ensiling fermentation, aerobic stability and microbial communities of total mixed ration (TMR) silage., Methods and Results: Laboratory-scale silos were stored at 5, 15, 25 and 35°C for 10, 30 and 90 days. If silage was stored at 5°C, fermentation was weak until day 30, but acceptable lactic acid production was observed on day 90. The ethanol content was higher than the acetic acid content when stored at 15 and 25°C, whereas the ethanol content was lower when stored at 35 than at 25°C. Aerobic deterioration did not occur when silage was exposed to air at the same temperature at which it was stored. Although 10-day silages stored at 5 and 15°C deteriorated when the aerobic stability test was conducted at 25°C, heating was not observed in silages stored at 25 or 35°C or in any 90-day silages regardless of storage temperature. Denaturing gradient gel electrophoresis demonstrated that bands indicative of Lactobacillus plantarum and Lactobacillus delbrueckii were less prominent, while bands indicative of Lactobacillus panis became more distinct in silages stored at high temperatures. Bands of Kluyveromyces marxianus were seen exclusively in silages that were spoiled at 25°C., Conclusion: High ambient temperature enhances acetic acid production in TMR silage. Lactobacillus panis may be associated with changes in the fermentation products due to differences in storage temperature., Significance and Impact of the Study: The role of Lacto. panis in ensiling fermentation and aerobic stability is worth examining., (© 2013 The Society for Applied Microbiology.)

Context: In the absence of panhypopituitarism and low serum IGF-I levels, the diagnosis of adult GH deficiency (AGHD) requires confirmation with a GH stimulation test. Macimorelin is a novel, orally active ghrelin mimetic that stimulates GH secretion., Objective: The objective of the study was to determine the diagnostic efficacy and safety of macimorelin in AGHD., Design: This was a multicenter open-label study comparing the diagnostic accuracy of oral macimorelin with that of arginine+GHRH in AGHD patients and healthy, matched controls. After 43 AGHD patients and 10 controls were tested, the GHRH analog Geref Diagnostic [GHRH(1-29)NH2] became unavailable in the United States. The study was completed by testing 10 additional AGHD patients and 38 controls with macimorelin alone., Main Outcome Measure: Peak GH area under the receiver operating characteristic curve after macimorelin was measured., Results: Fifty AGHD subjects and 48 controls were evaluated. Peak GH levels in AGHD patients and controls after macimorelin were 2.36 ± 5.69 and 17.71 ± 19.11 ng/mL, respectively (P < .0001). With macimorelin, the receiver operating characteristic analysis yielded an optimal GH cut point of 2.7 ng/mL, with 82% sensitivity, 92% specificity, and 13% misclassification rate. For subjects receiving both tests, macimorelin showed discrimination comparable with arginine+GHRH (area under the receiver operating characteristic curve 0.99 vs 0.94, respectively, P = .29). Obesity (body mass index > 30 kg/m(2)) was present in 58% of subjects, and peak GH levels were inversely associated with body mass index in controls (r = -0.37, P = .01). Using the separate cut points of 6.8 ng/mL for nonobese and 2.7 for obese subjects reduced the misclassification rate to 11%. Only 1 drug-related serious adverse event, an asymptomatic QT interval prolongation on the electrocardiogram, was reported., Conclusion: Oral macimorelin is safe, convenient, and effective in diagnosing AGHD with accuracy comparable with the arginine+GHRH test.

Unlabelled: The object of this study was to develop a reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay for detecting hepatitis D virus (HDV) genotype 1. With an alignment analysis, a highly conserved sequence (nt 820-1020) was chosen as a suitable target to design LAMP primers. The optimal condition of RT-LAMP was a 25-μl reaction volume, which consists of the following components: 1·6 μmol l⁻¹ each of FIP and BIP, 0·2 μmol l⁻¹ each of F3 and B3, 1·5 μmol l⁻¹ dNTPs, 4 mmol l⁻¹ MgSO₄, 8 U Bst DNA polymerase, 2U M-MLV and 2 μl extracted RNA sample. The amplification reaction was carried out at 65°C for 50 min. Compared with conventional qualitative or quantitative real-time reverse transcription polymerase chain reaction, the results of RT-LAMP indicated a 1000-fold increase in sensitivity for detecting HDV. There was no cross-reaction for the RT-LAMP method between HDV 1 and HIV, HAV, HBV, HCV and HEV., Significance and Impact of the Study: The results indicate that RT-LAMP is a simple, rapid, specific, highly sensitive and cost-effective, field-based method for detecting HDV 1. The RT-LAMP assay is an acceptable alternative to diagnose the HDV genotype 1 and to investigate its epidemiology for clinical laboratories lacking specialized equipments., (© 2012 The Society for Applied Microbiology.)

Aims: To understand the structure-activity relationship of chensinin-1, a anti-microbial peptide (AMP) with an unusual structure, and to develop novel AMPs as therapeutic agents., Methods and Results: A series of chensinin-1 analogues were designed and synthesized by one to three replacement of glycines with leucines at the hydrophilic face of chensinin-1 or rearrangement of some of the residues in its sequence. Circular dichroism spectroscopy showed that the analogues adopted α-helical-type conformations in 50% trifluoroethanol/water but adopted β-strand-type conformations in 30 mmol l(-1) sodium dodecyl sulphate. The anti-microbial activities of the peptides against Gram-positive bacteria increased 5- to 30-fold, and these increases paralleled the increases in the peptides' hydrophobicities. Their haemolytic activities also increased. Amphipathicities had little influence on the bactericidal activity of chensinin-1. All peptides caused leakage of calcein entrapped in negatively charged liposomes although with different efficiencies. The peptides did not induce leakage of calcein from uncharged liposomes., Conclusions: Peptide adopted an aperiodic structure can improve the anti-microbial potency by increasing peptide hydrophobicity. Its target is bacteria plasma membrane., Significance and Impact of the Study: Chensinin-1 can act as a new lead molecule for the study of AMPs with atypical structures., (© 2012 The Authors Journal of Applied Microbiology © 2012 The Society for Applied Microbiology.)

Aims: Acetic acid is considered an important preservative in tropical grass ensiling. The objective of the current experiments was to follow the ensiling fermentation of low dry matter (DM) tropical grass as a model to study changes in bacterial communities during acetic acid fermentation., Methods and Results: Direct-cut and wilted guinea grass silage was prepared with and without molasses. A high acetic acid level was observed during the fermentation of direct-cut silage, and long storage increased the butyric acid and ethanol content if molasses was not added. The lactic acid production in wilted silage was greater than the acetic acid production, but prolonged ensiling decreased the lactic to acetic acid ratio regardless of molasses addition. Adding molasses enhanced the lactic acid content in both direct-cut and wilted silage. The bacterial community, identified by denaturing gradient gel electrophoresis, was affected by wilting and molasses addition. Bands for Pantoea sp. and Morganella sp. became faint when acetic acid fermentation was suppressed, and those for Pediococcus pentosaceus and Lactococcus garvieae were detected when lactic acid fermentation was enhanced by wilting and molasses addition. Lactobacillus plantarum and Lactococcus lactis were found throughout the ensiling process in all silage types., Conclusion: Distinct changes occurred in the bacterial community in guinea grass silage because of wilting and molasses addition. These changes could explain how lactic acid fermentation was enhanced but could not help determine which bacteria were associated with enhanced acetic acid fermentation., Significance and Impact of the Study: The study reveals the effects of wilting and molasses during ensiling of low DM tropical grasses and the associated bacteria., (© 2011 The Authors. Letters in Applied Microbiology © 2011 The Society for Applied Microbiology.)

Aims:   The present study was conducted to evaluate the possibility of using cyanobacterial bloom materials as a medium for white rot fungi and the capability of white rot fungi, Trichaptum abietinum 1302BG and Lopharia spadicea to biodegrade dried cyanobacterial bloom material taken from Taihu Lake., Methods and Results:   The results showed T. abietinum 1302BG and L. spadicea could use the cyanobacterial bloom materials taken from Taihu Lake for growth to measure the mycelial plaque and dry-weight mycelial pellicles of fungi. The removal rate of dried cyanobacterial bloom materials incubated with white rot fungi is approximately 100%., Conclusions:   The cyanobacterial bloom material can be used as a glucose substitute in white rot fungi medium. The white rot fungi, T. abietinum 1302BG and L. spadicea, can also directly decrease the biomass of cyanobacterial bloom material taken from Taihu Lake., Significance and Impact of the Study:   Cyanobacterial bloom thrives in eutrophic fresh waters all over the world. Micro-organisms, particularly fungi, have attracted attention as possible agents for the degradation of phytoplankton species. Dealing with cyanobacterial bloom material as a medium for fungi instead of directly discharging them as organic fertilizers is a new, safe and environmentally friendly approach., (© 2011 The Authors. Letters in Applied Microbiology © 2011 The Society for Applied Microbiology.)

Aims: The objective of this study was to estimate UV(254) inactivation constants for four viral pathogens: influenza virus type A, porcine respiratory and reproductive syndrome virus (PRRSV), bovine viral diarrhoea virus (BVDV) and reovirus., Methods and Results: Viruses in culture medium were exposed to one of nine doses of UV(254) and then titrated for infectious virus. Analysis showed that viral inactivation by UV(254) was more accurately described by a two-stage inactivation model vs a standard one-stage inactivation model., Conclusions: The results provided evidence for the existence of two heterogeneous viral subpopulations among the viruses tested, one highly susceptible to UV(254) inactivation and the other more resistant. Importantly, inactivation constants based on the one-stage inactivation model would have underestimated the UV(254) dose required for the inactivation of these viruses under the conditions of the experiment., Significance and Impact of the Study: To improve the accuracy of estimates, it is recommended that research involving the inactivation of micro-organisms evaluates inactivation kinetics using both one-stage and two-stage models. These results will be of interest to persons responsible for microbial agents under laboratory or field conditions., (© 2011 The Authors. Journal of Applied Microbiology © 2011 The Society for Applied Microbiology.)

Aims: To characterize the bacterial communities in commercial total mixed ration (TMR) silage, which is known to have a long bunk life after silo opening., Methods and Results: Samples were collected from four factories that produce TMR silage according to their own recipes. Three factories were sampled three times at 1-month intervals during the summer to characterize the differences between factories; one factory was sampled 12 times, three samples each during the summer, autumn, winter and spring, to determine seasonal changes. Bacterial communities were determined by culture-independent denaturing gradient gel electrophoresis. All silages contained lactic acid as the predominant acid, and the contents appeared stable regardless of factories and product seasons. Acetic acid and 1-propanol contents were different between factories and indicated seasonal changes, with increases in warm seasons compared to cool seasons. Both differences and similarities existed among the bacterial communities from each factory and product season. Lactobacillus parabuchneri was found in the products from three of four factories. Various sourdough lactic acid bacteria (LAB) were identified in commercial TMR silage; Lactobacillus panis, Lactobacillus hammesii, Lactobacillus mindensis, Lactobacillus pontis, Lactobacillus frumenti and Lactobacillus farciminis were detected in many products. Moreover, changes owing to product season were distinctive, and Lact. pontis and Lact. frumenti became detectable in summer products., Conclusion: Sourdough LAB are involved in the ensiling of commercial TMR silage. Silage bacterial communities vary more by season than by factory. The LAB species Lact. parabuchneri was detected in the TMR silage but may not be essential to the product's long bunk life after silo opening., Significance and Impact of the Study: Commercial TMR silage resembles sourdough with respect to bacterial communities and long shelf life. The roles of sourdough LAB in the ensiling process and aerobic stability are worth examining., (© 2010 The Authors. Journal compilation © 2010 The Society for Applied Microbiology.)

Aim: To evaluate a new quantitative reverse transcription-PCR (qRT-PCR) assay for the rapid detection of methicillin-resistant Staphylococcus aureus (MRSA)., Methods and Results: Primers for Staphylococcus-specific regions of 16S rRNA gene, spa gene and mecA gene were newly designed. RNAs extracted from broth-cultured strains were tested by qRT-PCR targeting each primer, and the bacterial counts obtained correlated well with those counted by the plating method with detection limits of 10(0), 10(1) and 10(2) CFU. The qRT-PCR assay targeting the 16S rRNA was 6430-fold or more sensitive than qPCR assay. All Staph. aureus strains tested were detected and none of the other Staphylococcus species and genus strains tested cross-reacted with the assay targeting the spa gene. All MRSAs tested were detected by the assay targeting the mecA gene. Clinical samples, faecal material and bronchial washout solutions were tested by our assay, and MRSAs were detected with a high sensitivity within 6 h., Conclusion: Our qRT-PCR assay targeting three new primers to the target genes is a rapid and sensitive tool for the detection of MRSA directly from clinical samples., Significance and Impact of the Study: Because of its sensitivity and rapidity, our qRT-PCR assay is considered to be a valuable tool for clinical management.

Aims: To investigate the effect of modulation of transcription factors on the fermentation properties of Saccharomyces cerevisiae industrial strains and to evaluate whether overexpression and co-overexpression of transcription factors would result in higher ethanol yield., Methods and Results: A mutant gene spt15 (Phe(177)Ser, Tyr(195)His, Lys(218)Arg) was constructed by polymerase chain reaction mediated site-directed mutagenesis. The fermentation properties of the engineered strains in very high gravity fermentations were investigated. It is found that overexpression of SPT3 can enhance the resistance to ethanol and osmotic stress. On the contrary, overexpression of SPT15 or spt15 cannot obviously improve osmotic and ethanol tolerance of industrial strains. Additionally, simultaneous overexpression of SPT15 and SPT3 can not only distinctly enhance the resistance to ethanol and osmotic stress, but also improve fermentation performance., Conclusions: Simultaneous modulation of the expression level of SPT15 and SPT3 can increase the production of ethanol by improving osmotic tolerance and ethanol tolerance of industrial strains., Significance and Impact of the Study: Modulation of transcription factors provides a route to fermentation phenotypes of industrial yeast strains that are not readily accessible by traditional methods.

Aims: To isolate and characterize an oxalate-degrading Pandoraea sp. OXJ-11., Methods and Results: A new bacterium Pandoraea sp. OXJ-11 was isolated from soil samples, which can grow in the medium with oxalate as the sole carbon and energy source. The isolate OXJ-11 is Gram-negative straight rod. It occurs singly and is motile by means of a double polar flagellum. Catalase is positive and nitrate is not reduced. It grows aerobically and the optimum growth temperature and the optimum pH are at 30 degrees C and pH 6.0, respectively. The polyphasic taxonomic data along with 16S rRNA sequence comparison demonstrate that the isolate OXJ-11 should belong to the genus Pandoraea and represent a new member in this family., Conclusions: Oxalate could be degraded and the oxalate-degrading enzyme activity was detected when the isolate OXJ-11 grew in the medium with oxalate as carbon source., Significance and Impact of the Study: Oxalate-degrading Pandoraea sp. OXJ-11 would be beneficial to the potential application in the control of sclerotinia stem rot in economically important plants caused by fungus Sclerotinia sclerotiorum, and in making plants resistant to the white mold disease by oxalate-degrading enzyme transgene.

Interleukin (IL)-18, a proinflammatory cytokine, has been recognized recently as an important factor in both treated and untreated patients with human immunodeficiency virus type 1 (HIV-1) infection. Consistent with all earlier reports, our quantification of serum IL-18 concentrations in 88 HIV-1 seropositive, North American adolescents (14-18 years old) revealed a positive correlation with cell-free HIV-1 viral load at two separate visits (Spearman's r = 0.31 and 0.50, respectively, P < 0.01 for both), along with a negative correlation with CD4+ T cell counts (r = -0.31 and -0.35, P < 0.01 for both). In additional analyses of 66 adults (21-58 years old) from Zambia, HIV-1 seroconversion was associated uniformly with elevated IL-18 production (P < 0.0001). These epidemiological relationships were independent of other population-related characteristics, including age, gender and ethnicity. In neither study population could serum IL-18 concentrations be associated with the IL-18 gene (IL18) promoter genotypes defined by five major single nucleotide polymorphisms. Collectively, these findings suggest that circulating IL-18 rather than the IL18 genotype may provide a useful biomarker for HIV-1-related events or outcomes.

Chlamydia trachomatis infects epithelial cells at the mucosal surface. While in vitro and animal studies have shown changes in mucosal T(H)1-associated cytokines in the presence of C. trachomatis infection and with its progression to the upper genital tract or clearance, in vivo cytokine responses to chlamydial infection in humans are not well understood. Using a quantitative enzyme-linked immunosorbent assay (ELISA), we examined the endocervical production of two T(H)1-associated cytokines, i.e. interleukin (IL)-2 and IL-12, in relation to C. trachomatis infection in adolescents. At a randomly selected visit for 396 females, median endocervical IL-2 levels were significantly lower (190 versus 283 pg/ml, P = 0.02) and median IL-12 levels significantly higher (307 versus 132 pg/ml, P < 0.001) in subjects testing positive versus negative for C. trachomatis. These divergent T(H)1-associated cytokine responses were: (1) confirmed in paired analyses of 96 individuals before and after infection within 6-month intervals, (2) reversible in 97 patients who cleared infection during consecutive visits, (3) not attributable to sociodemographic factors or other genital infections and (4) independent of common genetic variants at the IL2 and IL12B loci associated previously with differential gene expression. From these findings we infer that increased IL-12 and decreased IL-2, observed commonly during mucosal inflammation, are important features of mucosal immune defence against C. trachomatis infection.