Impact of diet and bacterial supplementation regimes on Orius strigicollis microbiota and life history performance.

Given the growing interest in manipulating microbiota to enhance the fitness of mass-reared insects for biological control, this study investigated the impact of an artificial diet on the microbiota composition and performance of Orius strigicollis. We compared the microbiota of O. strigicollis fed on an artificial diet and moth eggs via culturing and 16S rRNA gene amplicon sequencing. Subsequently, we assessed life history traits and immune gene expression of O. strigicollis fed on the artificial diet supplemented with Pantoea dispersa OS1. Results showed that microbial diversity remained largely unaffected by the artificial diet, with similar microbiota compositions in both diet groups. OS1, a minor member of the microbiota but significantly enriched in bugs fed on the artificial diet, improved nymphal survival rates and shifted adult longevity-reproduction life history in females. Additionally, OS1 supplementation elevated the transcription of antimicrobial peptide diptericin. According to population parameters, the group receiving OS1 only during the nymphal stage showed higher population growth potential compared to the group supplemented across all life stages. These findings reveal the resilience of O. strigicollis microbiota under distinct dietary conditions and highlight the potential of using natural symbionts and specific supplementation regimes to improve Orius rearing for future biocontrol programs.

Monitoring the Season-Prevalence Relationship of Vairimorpha ceranae in Honey Bees (Apis mellifera) over One Year and the Primary Assessment of Probiotic Treatment in Taichung, Taiwan.

Microsporidiosis, which is caused by the pathogen Vairimorpha ceranae, is a prevalent disease in the honey bee (Apis mellifera) and might lead to significant adult honey bee mortality. In this study, we conducted an annual survey of the mature spore load of V. ceranae in the guts of nurse bees and forager bees in the apiary of National Chung Hsing University (NCHU) in Taiwan. The results indicated that, on average, honey bees hosted approximately 2.13 × 106 mature spore counts (MSCs)/bee in their guts throughout the entire year. The highest number of MSCs was 6.28 × 106 MSCs/bee, which occurred in April 2020, and the lowest number of MSCs was 5.08 × 105 MSCs/bee, which occurred in November 2020. Furthermore, the guts of forager bees had significantly higher (>58%) MSCs than those of nurse bees. To evaluate the potential of the probiotic to treat microsporidiosis, the lactic acid bacterium Leuconostoc mesenteroides TBE-8 was applied to honey bee colonies. A significant reduction (>53%) in MSCs following probiotic treatment was observed, indicating the potential of probiotic treatment for managing microsporidiosis. This research provided information on V. ceranae MSCs in the honey bee gut at NCHU in Taiwan and the MSCs' correlation with the annual season. Furthermore, a potential probiotic treatment for microsporidiosis was assessed for future management.

Radiographical presentations of inadvertent subdural placement of an epidural catheter for successful labor analgesia: A case report.

RATIONALE: Lumbar epidural analgesia is the gold standard for labor pain control. However, misplacement of epidural catheters into the subdural space may inadvertently happen. Unrecognized subdural administration of local anesthetics could result in serious consequences, including high spinal and brainstem blocks. This case report describes a case where subdural epidural catheter placement was recognized early but labor pain was adequately managed by dosage titration of subdural analgesia. PATIENT CONCERNS: This case report describes a 29-year-old primiparous pregnant woman who was admitted to our obstetric unit for labor induction at the gestational age of 38 weeks. An epidural catheter was inserted via the L2-3 intervertebral space using the standard loss of resistance to air technique. DIAGNOSES: The parturient experienced weakness in the lower extremities and numbness in the upper extremities within 15 minutes after administration of 5 mL of 2% v/v lidocaine as a loading dose and systolic blood pressure also dropped by 25%. INTERVENTIONS: The dose regimen (a mixture of 0.1% ropivacaine and 4 µg/mL fentanyl) for patient-controlled analgesia was given with bolus doses of 0.1 mL per demand and lockout intervals of 20 minutes. The analgesic effects were adequately maintained below the T8 dermatome for more than 12 hours without hypotensive episodes or obvious signs of neurological deficits. Computed tomographic myelography was performed by instillation of a nonionic iodinated contrast medium via the epidural catheter on postpartum day 2 for imaging confirmation of catheter placement in the extradural space. LESSONS: Early recognition that epidural catheters for neuraxial analgesia have been inserted into the subdural space is important for the prevention of high spinal blocks. Subdural analgesia could still be achieved by careful clinical assessment and titration of low analgesic doses. This report also presents important and clear serial computed tomographic images of catheter placement in the thoracic-lumbar subdural spaces and the extent of volume spread in the subdural space following administration of contrast medium.

Exploring the Functional Properties of Propolis, Geopropolis, and Cerumen, with a Special Emphasis on Their Antimicrobial Effects.

Bee propolis has been touted as a natural antimicrobial agent with the potential to replace antibiotics. Numerous reports and reviews have highlighted the functionalities and applications of the natural compound. Despite much clamor for the downstream application of propolis, there remain many grounds to cover, especially in the upstream production, and factors affecting the quality of the propolis. Moreover, geopropolis and cerumen, akin to propolis, hold promise for diverse human applications, yet their benefits and intricate manufacturing processes remain subjects of intensive research. Specialized cement bees are pivotal in gathering and transporting plant resins from suitable sources to their nests. Contrary to common belief, these resins are directly applied within the hive, smoothed out by cement bees, and blended with beeswax and trace components to create raw propolis. Beekeepers subsequently harvest and perform the extraction of the raw propolis to form the final propolis extract that is sold on the market. As a result of the production process, intrinsic and extrinsic factors, such as botanical origins, bee species, and the extraction process, have a direct impact on the quality of the final propolis extract. Towards the end of this paper, a section is dedicated to highlighting the antimicrobial potency of propolis extract.

Transcriptome of Nosema ceranae and Upregulated Microsporidia Genes during Its Infection of Western Honey Bee (Apis mellifera).

Nosema ceranae is one of the fungal parasites of Apis mellifera. It causes physical and behavioral effects in honey bees. However, only a few studies have reported on gene expression profiling during A. mellifera infection. In this study, the transcriptome profile of mature spores at each time point of infection (5, 10, and 20 days post-infection, d.p.i.) were investigated. Based on the transcriptome and expression profile analysis, a total of 878, 952, and 981 differentially expressed genes (DEGs) (fold change ≥ 2 or ≤ -2) were identified in N. ceranae spores (NcSp) at 5 d.p.i., 10 d.p.i., and 20 d.p.i., respectively. Moreover, 70 upregulated genes and 340 downregulated genes among common DEGs (so-called common DEGs) and 166 stage-specific genes at each stage of infection were identified. The Gene Ontology (GO) analysis indicated that the DEGs and corresponding common DEGs are involved in the functions of cytosol (GO:0005829), cytoplasm (GO:0005737), and ATP binding (GO:0005524). Furthermore, the pathway analysis found that the DEGs and common DEGs are involved in metabolism, environmental information processing, and organismal systems. Four upregulated common DEGs with higher fold-change values, highly associated with spore proteins and transcription factors, were selected for validation. In addition, the stage-specific genes are highly involved in the mechanism of pre-mRNA splicing according to GO enrichment analysis; thus, three of them showed high expression at each d.p.i. and were also subjected to validation. The relative gene expression levels showed a similar tendency as the transcriptome predictions at different d.p.i., revealing that the gene expression of N. ceranae during infection may be related to the mechanism of gene transcription, protein synthesis, and structural proteins. Our data suggest that the gene expression profiling of N. ceranae at the transcriptomic level could be a reference for the monitoring of nosemosis at the genetic level.

Effect of harvest time span on physicochemical properties, antioxidant, antimicrobial, and anti-inflammatory activities of Meliponinae honey.

BACKGROUND: The maturity of honey has a great impact on its quality and contents. Additionally, stingless bee honey contains high moisture, which allows microorganisms to survive and ferment, contributing to honey's variable flavor and physicochemical properties. Therefore, there is a need for better quality control of the honey process, especially the harvest time of honey. RESULTS: We gathered honey from the nest of stingless bees Heterotrigona itama and Tetrigona binghami over different time periods, i.e. 15, 30, and 45 days. The results show harvest time considerably affects the physicochemical properties, antioxidant activity, and antimicrobial activity of honey. Good antioxidant activity and antimicrobial activity can be found in honey produced from a longer harvest time. Compared with 15-day harvest time, at 30- or 45-day harvest time water, trehalulose, and protein content and total acidity increased, and the content of reducing sugars, fructose and glucose, and pH values, decreased in both types of honey. Moreover, compared with 15-day harvest time, the sum of six organic acids in the 45-day honey of H. itama fluctuated between 2.78 to 4.12 g 100 g-1 and in the 45-day honey of T. binghami increased from 1.66 to 3.61 g 100 g-1 , respectively. CONCLUSION: Honey harvest time had a significant effect on the physicochemical properties, antioxidant activity, and antimicrobial activity of stingless bee honey (H. itama or T. binghami). This study provides a reference for beekeepers to adjust harvest time to obtain honey with suitable physicochemical parameters. © 2022 Society of Chemical Industry.

A Generic Nomogram Predicting the Stage of Liver Fibrosis Based on Serum Biochemical Indicators Among Chronic Hepatitis B Patients.

Introduction: Liver fibrosis staging is of great importance for reducing unnecessary injuries and prompting treatment in chronic viral hepatitis B patients. Liver biopsy is not suitable to act a screening method although it is a gold standard because of various shortcomings. This study aimed to establish a predictive nomogram as a convenient tool to effectively identify potential patients with different stages of liver fibrosis for patients with chronic hepatitis B. Methods: A nomogram for multinomial model was developed in a training set to calculate the probability for each stage of fibrosis and tested in a validation set. Fibrosis stages were subgrouped as followed: severe fibrosis/cirrhosis (F3-F4), moderate fibrosis (F2), and nil-mild fibrosis (F0-F1). The indicators were demographic characteristics and biochemical indicators of patients. Continuous indicators were divided into several groups according to the optimal candidate value generated by the decision tree. Results: This study recruited 964 HBV patients undergoing percutaneous liver biopsy. The multinomial model with 10 indicators was transformed into the final nomogram. The calibration plot showed a good agreement between nomogram-predicted and observed probability of different fibrosis stages. Areas under the receiver operating characteristics (AUROCs) for severe fibrosis/cirrhosis were 0.809 for training set and 0.879 for validation set. For moderate fibrosis, the AUROCs were 0.75 and 0.781. For nil-mild fibrosis, the AUROCs were 0.792 and 0.843. All the results above showed great predictive performance in predicting the stage of fibrosis by our nomogram. Conclusion: Our model demonstrated good discrimination and extensibility in internal and external validation. The proposed nomogram in this study resulted in great reliability and it can be widely used as a convenient and efficient way.

A potential probiotic Leuconostoc mesenteroides TBE-8 for honey bee.

An isolated bacterium TBE-8, was identified as Leuconostoc mesenteroides according to the sequences of 16S rDNA and the 16S-23S rDNA intergenic spacer region. The probiotic properties of the L. mesenteroides TBE-8 strain were characterized and revealed that TBE-8 could utilize various carbohydrates, exhibited high tolerance to sucrose's osmotic pressure and acidic conditions, and could mitigate the impact of the bee pathogen Paenibacillus larvae. In addition, we found that the TBE-8 broth increased the expression of the nutrition-related genes major royal jelly protein 1 and vitellogenin in bees by approximately 1400- and 20-fold, respectively. The expression of genes encoding two antibacterial peptides, hymenoptaecin and apidaecin, in the bee abdomen was significantly increased by 17- and 7-fold in bees fed with the TBE-8 fermented broth. Furthermore, we fed four-frame bee colonies with 50% sucrose syrup containing TBE-8 and can detect the presence of approximately 2 × 106 16S rDNA copies of TBE-8 in the guts of all bees in 24 h, and the retention of TBE-8 in the bee gut for at least 5 days. These findings indicate that the L. mesenteroides TBE-8 has high potential as a bee probiotic and could enhance the health of bee colonies.

Nutritive Value of 11 Bee Pollen Samples from Major Floral Sources in Taiwan.

Bee pollen is a nutrient-rich food that meets the nutritional requirements of honey bees and supports human health. This study aimed to provide nutritive composition data for 11 popular bee pollen samples (Brassica napus (Bn), Bidens pilosa var. radiata (Bp), Camellia sinensis (Cs), Fraxinus griffithii (Fg), Prunus mume (Pm), Rhus chinensis var. roxburghii (Rc), Bombax ceiba (Bc), Hylocereus costaricensis (Hc), Liquidambar formosana (Lf), Nelumbo nucifera (Nn), and Zea mays (Zm)) in Taiwan for the global bee pollen database. Macronutrients, such as carbohydrates, proteins, and lipids, were analyzed, which revealed that Bp had the highest carbohydrate content of 78.8 g/100 g dry mass, Bc had the highest protein content of 32.2 g/100 g dry mass, and Hc had the highest lipid content of 8.8 g/100 g dry mass. Only the bee pollen Hc completely met the minimum requirements of essential amino acids for bees and humans, and the other bee pollen samples contained at least 1-3 different limiting essential amino acids, i.e., methionine, tryptophan, histidine, valine, and isoleucine. Regarding the fatty acid profile of bee pollen samples, palmitic acid (C16:0), stearic acid (C18:0), oleic acid (C18:1), linoleic acid (C18:2), and linolenic acid (C18:3) were predominant fatty acids that accounted for 66.0-97.4% of total fatty acids. These data serve as an indicator of the nutritional quality and value of the 11 bee pollen samples.

A Potential Fungal Probiotic Aureobasidium melanogenum CK-CsC for the Western Honey Bee, Apis mellifera.

Aureobasidium melanogenum has been used as an animal feed additive for improving thehealth of pets, however, it has not yet been applied in honey bees. Here, a fungal strain CK-CsC isolated from bee bread pollen, was identified as A. melanogenum. Following characterizing CK-CsC fermentation broth, the 4-days fermentation broth (SYM medium or bee pollen) of the CK-CsC was used to feed newly emerged adult honey bees in cages under laboratory-controlled conditions for analysis of survival, gene expression of nutrient and antibacterial peptide, and gut microbiota of honey bees. It was found that the CK-CsC fermentation broth (SYM medium or bee pollen) is nontoxic to honey bees, and can regularly increase nutrient gene expression of honey bees. However, significant mortality of bees was observed after bees were fed on the supernatant liquid of the fermentation broth. Notably, this mortality can be lowered by the simultaneous consumption of bee pollen. The honey bees that were fed bee pollen exhibited more γ-Proteobacteria, Bacteriodetes, and Actinobacteria in their gut flora than did the honey bees fed only crude supernatant liquid extract. These findings indicate that A. melanogenum CK-CsC has high potential as a bee probiotic when it was fermented with bee pollen.

Honey proteomic signatures for the identification of honey adulterated with syrup, producing country, and nectar source using SWATH-MS approach.

Honey is widely consumed by humans, due to its multiple applications as a food constituent and its therapeutic effects. This study reports on the discrimination of honey products from different geographical and botanical sources, as well as honey products containing distinct forms of syrup used in honey adulteration. Sequential window acquisition of all theoretical fragment ion spectra mass spectrometry (SWATH-MS)-based proteomic analysis combined with chemometrics was successfully applied in identifying characteristic proteins that can be used as biomarkers of the original source of honey. Honey samples from different producing regions (Tainan, Changhua, and Taichung), countries (Taiwan and Thailand), and distinct botanical sources (longan and litchi) were clearly distinguished by the developed orthogonal projections to latent structures discriminant analysis (OPLS-DA) model with good fitness and prediction ability. Furthermore, we successfully discriminated the adulteration of honey with syrup in different proportions (even with honey content as low as 20%) with this proteomic SWATH-MS platform.

A nationwide survey of intraoperative management for one-lung ventilation in Taiwan: time to accountable for diversity in protective lung ventilation.

BACKGROUND: There is a major paradigm shift for intraoperative mechanical ventilator support by the introduction of lung protective ventilation strategies to reduce postoperative pulmonary complications and improve overall clinical outcomes in non-thoracic surgeries. However, there is currently a lack of standardized practice guideline for lung protection during thoracic surgeries that require one-lung ventilation (OLV). This study aimed to collect the expert opinions of the thoracic anesthesiologists in perioperative care for OLV surgery in Taiwan. METHODS: This prospective cross-sectional study was undertaken in 16 tertiary hospitals in Taiwan from January to February 2019. A structured survey form was distributed across the participating hospitals and the thoracic anesthesiologists were invited to complete the form voluntarily. The survey form consisted of three parts, including the basic information of the institutional anesthesia care standards, ventilatory settings for a proposed patient receiving OLV surgery and expert opinions on OLV. RESULTS: A total of 71 thoracic anesthesiologists responded to the survey. Double-lumen tubes are the most commonly used (93.8%) airway devices for OLV. The most commonly recommended ventilator setting during OLV is a tidal volume of 6-7 ml/kg PBW (67.6%) and a PEEP level of 4-6 cmH2O (73.5%). Dual controlled ventilator modes are used by 44.1% of the anesthesiologists. During OLV, high oxygen fraction (FiO2 > 0.8) is more commonly supplemented to achieve an oxygen saturation higher than 94%. The consensus of anesthesiologists on the indices for lung protection in thoracic surgery is considerably low. Large majority of the anesthesiologists (91.5%) highly recommend that an international clinical practice guideline on the protective lung ventilation strategy for thoracic anesthesia should be established. CONCLUSIONS: This study found that the thoracic anesthesiologists in Taiwan share certain common practices in ventilator support during OLV. However, they are concerned about the lack of fundamental clinical evidences to support the beneficial outcomes of the current lung protective strategies applicable to OLV. Large-scale trials are needed to form an evidence-based clinical practice guideline for thoracic anesthesia.

Activities of daily living, life orientation, and health-related quality of life among older people in nursing homes: a national cross-sectional study in China.

PURPOSE: This study aimed to explore the current status of activities of daily living (ADLs), life orientation, and health-related quality of life (HRQoL) among older people in nursing homes and to further examine the mediating role of life orientation in the impact of ADLs on HRQoL. METHODS: A national cross-sectional study was conducted among older people aged 60 and above in nursing homes by the randomly stratified cluster sampling method. The status of ADLs, life orientation and HRQoL were measured using an ADL scale, a life orientation scale and the SF-12v2 scale, respectively. Multiple linear regression models were used to identify explanatory factors associated with ADLs, life orientation, and HRQoL. The potential mediating role of life orientation in the relationship between ADL and HRQoL was explored by mediation analysis. RESULTS: The overall prevalence of ADL disability was 52.67%, and 84.37% of older people in nursing homes had a negative life orientation. The mean scores of physical health and mental health among older people in nursing homes were 45.44 ± 6.46 and 42.67 ± 8.48, respectively. Some sociodemographic characteristics were associated with poor physical health and mental health. After adjustments were made for covariates, the life orientation score mediated 13.81% of the total effect of the ADL score on physical component score of HRQoL and mediated 45.33% of the mental component score of HRQoL. CONCLUSION: A sizeable proportion of older people had ADL disability and negative life orientation, and HRQoL was poor among older Chinese people in nursing homes. Life orientation partially mediates the relationship between ADLs and HRQoL.

Potential probiotic of Lactobacillus strains isolated from the intestinal tracts of pigs and feces of dogs with antibacterial activity against multidrug-resistant pathogenic bacteria.

The occurrence of multidrug-resistant pathogenic bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA), multidrug-resistant Acinetobacter baumannii (MDRAB), extended-spectrum ß-lactamase (ESBL) Escherichia coli, and Pseudomonas aeruginosa, has become a serious problem in animals and public. The objective of this study was to identify and isolate lactic acid bacterial (LAB) strains from the intestinal tracts of pigs and feces of dogs and then characterize them as potential probiotics with antimicrobial activity against multidrug-resistant pathogenic bacteria. In a preliminary isolation screening, 45 of 1167 isolated LAB strains were found to have anti-S. aureus ATCC 27,735 activity. Using 16S rDNA and 16S-23S rDNA intergenic spacer region (ISR) sequences, five of these isolates were further identified as Lactobacillus animalis 30a-2, Lactobacillus reuteri 4-12E, Weissella cibaria C34, Lactococcus lactis 5-12H, and Lactococcus lactis 6-3H. Antimicrobial substance assays suggest that the L. lactis 5-12H, L. lactis 6-3H, L. animalis 30a-2, L. reuteri 4-12E, and W. cibaria C34 strains might produce bacteriocins and hydrogen peroxide (H2O2) as antimicrobial substances. The L. animalis 30a-2 and W. cibaria C34 strains were further characterized for probiotic properties and shown to have high acid and bile salt tolerance. Additionally, they have broad antimicrobial spectra, and can significantly repress the growth of all of the tested strains of MRSA isolates, some MDRAB, ESBL E. coli, and P. aeruginosa isolates, along with food-borne pathogenic bacteria such as Bacillus cereus ATCC 11778, Listeria monocytogens ATCC 19111, Salmonella spp., Shigella spp., and Yersinia enterocolitica BCRC 12986. This is the first report of H2O2-producing L. animalis 30a-2 and W. cibaria C34 isolated from the intestinal tracts of pigs and feces of dogs that have good antimicrobial activity against multidrug-resistant and food-borne pathogenic bacteria and have excellent probiotic properties.

The ability of different imputation methods for missing values in mental measurement questionnaires.

BACKGROUND: Incomplete data are of particular important influence in mental measurement questionnaires. Most experts, however, mostly focus on clinical trials and cohort studies and generally pay less attention to this deficiency. We aim is to compare the accuracy of four common methods for handling items missing from different psychology questionnaires according to the items non-response rates. METHOD: All data were drawn from the previous studies including the self-acceptance scale (SAQ), the activities of daily living scale (ADL) and self-esteem scale (RSES). SAQ and ADL dataset, simulation group, were used to compare and assess the ability of four imputation methods which are direct deletion, mode imputation, Hot-deck (HD) imputation and multiple imputation (MI) by absolute deviation, the root mean square error and average relative error in missing proportions of 5, 10, 15 and 20%. RSES dataset, validation group, was used to test the application of imputation methods. All analyses were finished by SAS 9.4. RESULTS: The biases obtained by MI are the smallest under various missing proportions. HD imputation approach performed the lowest absolute deviation of standard deviation values. But they got the similar results and the performances of them are obviously better than direct deletion and mode imputation. In a real world situation, the respondents' average score in complete data set was 28.22 ± 4.63, which are not much different from imputed datasets. The direction of the influence of the five factors on self-esteem was consistent, although there were some differences in the size and range of OR values in logistic regression model. CONCLUSION: MI shows the best performance while it demands slightly more data analytic capacity and skills of programming. And HD could be considered to impute missing values in psychological investigation when MI cannot be performed due to limited circumstances.

MiRNA Polymorphisms and Hepatocellular Carcinoma Susceptibility: A Systematic Review and Network Meta-Analysis.

BACKGROUND: Hepatocellular carcinoma (HCC) is an intractable public health threat worldwide, representing the second leading cause of cancer-related mortality, with limited early detection and therapeutic options. Recent findings have revealed that the susceptibility of HCC is closely related to microRNA (miRNA). We performed this systematic review with a network meta-analysis to investigated four single nucleotide polymorphisms (SNPs) that most regularly reported in miRNAs, exploring their involvement in HCC susceptibility and interaction with hepatitis B virus (HBV). METHODS: Databases were reviewed for related studies published up to May 2019 to identify all studies that compared genotypes of miR-146a rs2910164, miR-149 rs2292832, miR-196a2 rs11614913, and miR-499 rs3746444 with no language and date restrictions. A pairwise meta-analysis was performed to estimate pooled odds ratios and 95% confidence intervals incorporating heterogeneity to assess the relationship between four miRNA polymorphisms and HCC. To further clarify the effect of polymorphisms on HCC, a Bayesian network meta-analysis was conducted to combine the effective sizes of direct and indirect comparisons. Calculations were performed by R version 3.6.1 and STATA 14.0. All steps were performed according to PRISMA guidelines. RESULTS: A total of 20 studies were enrolled in this network meta-analysis, providing 5,337 hepatocellular carcinoma cases and 6,585 controls. All included studies had an acceptable quality. Pairwise meta-analysis demonstrated that miR-196a2 rs11614913 was significantly associated with the susceptibility of HCC, while the other three SNPs were not found to have a significant association. In the analysis of HCC patients under different HBV infection status, only miR-196a2 revealed correlation of threefold risk. The network results showed no significant difference in the distribution of genotype frequencies except for miR-196a2, which appeared to have the highest superiority index when comparing and ranking four SNPs. CONCLUSION: MiR-196a2 rs11614913 was significantly associated with the susceptibility of HCC, especially for HBV- related HCC, and that individuals with TC/CC were more susceptible. No significant association was found in the other three miRNA genes. MiR-196a2 could serve as the best predictor of susceptibility in HCC.

PCR-RFLP of mitochondrial DNA reveals two origins of Apis mellifera in Taiwan.

Beekeeping has been a highly valued industry in Taiwan. As a result, many subspecies of Apis mellifera have been introduced to Taiwan since 1911, leading to the hybridization of different subspecies. In order to know the matrilineal origins of Taiwan A. mellifera, a total of 280 samples collected from 33 apiaries throughout the island were examined. Using PCR-RFLP of four mitochondrial gene fragments, i.e., the non-coding region between tRNAleu and cytochrome c oxidase subunit II (intergenic tRNAleu-COII), cytochrome b (Cyt b), large subunit rRNA (Ls rRNA) and cytochrome c oxidase subunit I (COI), we only found two haplotypes exist in 280 samples. Haplotypes ababa and bbbaa account for 87% of these Western bees belonged to the Eastern European (C) lineage and 13% belonged to the Middle East (Z) lineage, respectively, with the latter being totally absent in northern Taiwan. African (A) and Mellifera (M) lineages, officially imported once in 1990s and 1930s respectively, were not detected. The identification of subspecies of A. mellifera and survey of their distribution on the island are expected to facilitate efficient breeding programs and establish a more booming beekeeping industry.

Gene expression changes in honey bees induced by sublethal imidacloprid exposure during the larval stage.

Honey bee larvae exposed to sublethal doses of imidacloprid show behavioural abnormalities as adult insects. Previous studies have demonstrated that this phenomenon originates from abnormal neural development in response to imidacloprid exposure. Here, we further investigated the global gene expression changes in the heads of newly emerged adults and observed that 578 genes showed more than 2-fold changes in gene expression after imidacloprid exposure. This information might aid in understanding the effects of pesticides on the health of pollinators. For example, the genes encoding major royal jelly proteins (MRJPs), a group of multifunctional proteins with significant roles in the sustainable development of bee colonies, were strongly downregulated. These downregulation patterns were further confirmed through analyses using quantitative reverse transcription-polymerase chain reaction on the heads of 6-day-old nurse bees. To our knowledge, this study is the first to demonstrate that sublethal doses of imidacloprid affect mrjp expression and likely weaken bee colonies.

Rational Re-engineering of a Transcriptional Silencing PreQ1 Riboswitch.

Re-engineered riboswitches that no longer respond to cellular metabolites, but that instead can be controlled by synthetic molecules, are potentially useful gene regulatory tools for use in synthetic biology and biotechnology fields. Previously, extensive genetic selection and screening approaches were employed to re-engineer a natural adenine riboswitch to create orthogonal ON-switches, enabling translational control of target gene expression in response to synthetic ligands. Here, we describe how a rational targeted approach was used to re-engineer the PreQ1 riboswitch from Bacillus subtilis into an orthogonal OFF-switch. In this case, the evaluation of just six synthetic compounds with seven riboswitch mutants led to the identification of an orthogonal riboswitch-ligand pairing that effectively repressed the transcription of selected genes in B. subtilis. The streamlining of the re-engineering approach, and its extension to a second class of riboswitches, provides a methodological platform for the creation of new orthogonal regulatory components for biotechnological applications including gene functional analysis and antimicrobial target validation and screening.

Engineered biosynthesis of enduracidin lipoglycopeptide antibiotics using the ramoplanin mannosyltransferase Ram29.

The lipopeptides ramoplanin from Actinoplanes sp. ATCC 33076 and enduracidin produced by Streptomyces fungicidicus are effective antibiotics against a number of drug-resistant Gram-positive pathogens. While these two antibiotics share a similar cyclic peptide structure, comprising 17 amino acids with an N-terminal fatty acid side chain, ramoplanin has a di-mannose moiety that enduracidin lacks. The mannosyl substituents of ramoplanin enhance aqueous solubility, which was important in the development of ramoplanin as a potential treatment for Clostridium difficile infections. In this study we have determined the function of the putative mannosyltransferase encoded by ram29 from the ramoplanin biosynthetic gene cluster. Bioinformatics revealed that Ram29 is an integral membrane protein with a putative DxD motif that is suggested to bind to, and activate, a polyprenyl phosphomannose donor and an extracytoplasmic C-terminal domain that is predicted to bind the ramoplanin aglycone acceptor. The ram29 gene was cloned into the tetracycline inducible plasmid pMS17 and integrated into the genome of the enduracidin producer S. fungicidicus. Induction of ram29 expression in S. fungicidicus resulted in the production of monomannosylated enduracidin derivatives, which are not present in the WT strain. Tandem MS analysis showed that mannosylation occurs on the Hpg11 residue of enduracidin. In addition to confirming the function of Ram29, these findings demonstrate how the less common, membrane-associated, polyprenyl phosphosugar-dependent glycosyltransferases can be used in natural product glycodiversification. Such a strategy may be valuable in future biosynthetic engineering approaches aimed at improving the physico-chemical and biological properties of bioactive secondary metabolites including antibiotics.