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The Sip1Aa protein from Bacillus thuringiensis is highly toxic to Colaphellus bowringi Baly. In order to obtain mutant proteins with higher insecticidal activity, a random recombinant library of Sip1Aa protein was constructed using error-prone PCR. A total number of 100 positive transformants were randomly selected for sequence determination, and 25 mutants (M1 to M25) were selected and expressed the respective Sip1Aa mutants. These Sip1Aa variants had a total of 29 base mutations, with an average of 1.2 base mutations per mutant. Compared with that of the wild-type Sip1Aa protein, the insecticidal activity of the mutants M1 (A31G, Y118C, D227E), M5 (K168R) and M21 (I307T) was significantly decreased, with and LC50 values 4 to 6 times higher than the Sip1Aa protein. The mutant M8 (R174S) showed increase in the insecticidal activity against the Colaphellus bowringi Baly was obtained, with an LC50 value 4-fold less than the Sip1Aa protein. The results of this study provide reference for the molecular modification of Sip1Aa protein and the study of key sites of its insecticidal activity.

Prometryne is a widely used herbicide in China to control annual grasses and broadleaf weeds. However, the stability of prometryne makes it difficult to be degraded, which poses a threat to human health. This study presents a bacterial strain isolated from soil samples with a prometryne application history, designated strain DY-1. Strain DY-1, identified as Pseudomonas sp., is capable of utilizing prometryne as a sole carbon source for growth and degrading 100% of prometryne within 48 h from an initial concentration of 50 mg L−1. To further optimize the degradation of prometryne, the prometryne concentration, temperature, pH, and salt concentration were examined. The optimal conditions for degradation of prometryne by strain DY-1 were an initial prometryne concentration of 50 mg L−1, 30 °C, pH 7–8, and NaCl concentration of 200 mg L−1. The same strain also degraded other s-triazine herbicides, including simetryne, ametryne, desmetryne, and metribuzin, under the same conditions. The biodegradation pathway of prometryne was established by isolating sulfoxide prometryne as the first metabolite and by the identification of sulfone prometryne and 2-hydroxy prometryne by liquid chromatography-mass spectrometry (LC–MS/MS). The results illustrated that strain DY-1 achieved the removal of prometryne by gradually oxidizing and hydrolyzing the methylthio groups. A bioremediation trial with contaminated soil and pot experiments showed that after treating the prometryne-contaminated soil with strain DY-1, the content of prometryne was significantly reduced (P

The search for therapeutic targets for Parkinson’s disease (PD) is hindered by the incomplete understanding of the pathophysiology of the disease. Mitochondrial dysfunction is an area with high potential. The neurobiological signaling connections between the gut microbiome and the central nervous system are incompletely understood. Multiple lines of evidence suggest that the gut microbiota participates in the pathogenesis of PD. Gut microbial dysbiosis may contribute to the loss of dopaminergic neurons through mitochondrial dysfunction. The intervention of gut microbial metabolites via the microbiota-gut-brain axis may serve as a promising therapeutic strategy for PD. In this narrative review, we summarize the potential roles of gut microbial dysbiosis in PD, with emphasis on microbial metabolites and mitochondrial function. We then review the possible ways in which microbial metabolites affect the central nervous system, as well as the impact of microbial metabolites on mitochondrial dysfunction. We finally discuss the possibility of gut microbiota as a therapeutic target for PD.

Understanding how gut flora influences gut-brain communications has been the subject of significant research over the past decade. The broadening of the term “microbiota-gut-brain axis” from “gut-brain axis” underscores a bidirectional communication system between the gut and the brain. The microbiota-gut-brain axis involves metabolic, endocrine, neural, and immune pathways which are crucial for the maintenance of brain homeostasis. Alterations in the composition of gut microbiota are associated with multiple neuropsychiatric disorders. Although a causal relationship between gut dysbiosis and neural dysfunction remains elusive, emerging evidence indicates that gut dysbiosis may promote amyloid-beta aggregation, neuroinflammation, oxidative stress, and insulin resistance in the pathogenesis of Alzheimer’s disease (AD). Illustration of the mechanisms underlying the regulation by gut microbiota may pave the way for developing novel therapeutic strategies for AD. In this narrative review, we provide an overview of gut microbiota and their dysregulation in the pathogenesis of AD. Novel insights into the modification of gut microbiota composition as a preventive or therapeutic approach for AD are highlighted.

The fall armyworm Spodoptera frugiperda and cotton bollworm Helicoverpa armigera are major insect pests of corn and cotton worldwide. Genetically engineered crops producing Vip3Aa, a potent endotoxin, from the bacterium Bacillus thuringiensis (Bt) are effective in controlling these two harmful pests. However, Vip3Aa efficacy is relatively weak compared to that of other Bt proteins such as Cry1A and Cry1F. This study sought to modify Vip3Aa for increased insecticidal activity and determine the cause of elevated activity.The two triple Vip3Aa mutants in domains IV and V (Vip3Aa-S543N/I544L/E627A and Vip3Aa-S543N/I544L/S686R) exhibited 7.3-fold and 2.8-fold increased toxicity against S. frugiperda, respectively, compared with the wild type while the toxicity of Vip3Aa-S543N/I544L/S686R was 3.2 times that of wild-type protein in H. armigera. The mutants had enhanced stability in midgut juice and 2.6-5.1 times higher binding affinity against S. frugiperda and H. armigera compared with wild type protein.The enhanced toxicity of Vip3Aa mutants was due to increased stability and binding affinity during infection. The amino acids S543 and I544 combined with E627 or S686 in domains IV and V of Vip3Aa are important for maintaining structural stability and receptor binding. The results match insecticidal activity (LC

The secretory insecticidal protein Sip1Ab and crystal protein Cry8Ca from

Vip3Aa11 and Vip3Aa39 were isolated from Bacillus thuringiensis strains C9 and T03B001, respectively. Despite that there is 95.06% similarity of amino acid sequences between these two prote...

Sip1Aa is an insecticidal protein of Bacillus thuringiensis at the secretory stage. It has a strong toxic effect on the members of order Coleoptera. To date, there are few available studies on Sip1Aa protein and the inclusion body problem is serious, and this raises the importance to conduct further studies on Sip1Aa protein. Disulfide bonds, as the only covalent bond on protein side chains, play an important role in the stability and function of the proteins. The tertiary structure of Sip1Aa protein was analyzed by homologous modeling and other bioinformatics methods to predict the conserved domain of Sip1Aa protein. Cysteine used to replace these amino acids by site-directed mutation. Consequently, we were able to successfully construct Sip149-251, Sip153-248, Sip158-243, and Sip178-314. These were exposed to ultraviolet radiation, and we found that Sip153-248 and Sip158-243 were the most stable, followed by Sip149-251 and Sip178-314 when compared with Sip1Aa. After the mutant strain was transferred into Escherichia coli BL21, sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used to detect the inducible expression products. Approximately 37.6 kDa of proteins that were highly expressed in E. coli. We found no significant change in the insecticidal activity.

The microbiota-gut-brain axis has emerged as a focal point of biomedical research. Alterations of gut microbiota are involved in not only various immune/inflammatory disorders but also neurological disorders including Alzheimer's disease (AD). The initial stage of the involvement of gut microbiota in the pathogenesis of AD may be the dysfunction of the blood-brain barrier (BBB). Gut microbiota-derived products in the circulation can worsen the BBB integrity, easily cross the disrupted BBB and enter the brain to promote pathological changes in AD. In this review, we first summarize the current evidence of the associations among gut microbiota, AD, and BBB integrity. We then discuss the mechanism of gut microbiota on BBB dysfunction with a focus on bacteria-derived lipopolysaccharide and exosomal high-mobility group box 1. Novel insights into the modification of the BBB as an intervention approach for AD are highlighted as well.

In this study, based on the pUC19 vector and using overlapping PCR technology, the researchers constructed an expression vector of the Sip1Aa protein guided by a cry1Ac promoter. The expression situation, insecticidal activity and solubility were researched, and the expression of the Sip1Aa protein, as guided by a T7 promoter, was compared. Additionally, the fermentation conditions were explored on a preliminary basis and a histidine label was added to the recombinant plasmid via reverse PCR for subsequent purification of recombinant proteins. The results showed that both the cry1Ac and T7 promoters could guide Sip1Aa to express as a soluble protein of 37.6 kDa, and there was no significant difference in insecticidal activity against Colaphellus bowringi Baly, with LC50 values of 1.637 mg/mL and 1.683 μg/mL, respectively. The soluble component of the Sip1Aa protein, when guided by the cry1Ac promoter, was significantly higher than when it was guided by the T7 promoter. The expression of the cry1Ac guider-promoted Sip1Aa protein was more suitable at 37℃ and 16 h. The recombinant protein was purified after an exogenous histidine sequence was added. This provides a new research method and idea to solve the problem of the Sip1Aa protein usually producing a large number of inclusion bodies when expressed in E. coli, and provides new ideas for the study of sip gene rapid expression, functional verification and insecticidal mechanisms.

Background Investigations have revealed the association between inflammation and post-stroke depression (PSD). However, whether the C-reactive protein (CRP) level, a biomarker of inflammation, would affect the development of PSD is still controversial. Methods A systematic search of databases was performed for eligible studies. Standardized Mean Difference (SMD) with 95 % Confidence Interval (CI) was used to assess the association between the CRP level in the acute phase of stroke and the risk of PSD. Results 13 cohort studies that involved 3536 participants were included. Combined results showed that compared with non-PSD patients, the CRP level of PSD patients was significantly higher on admission (SMD = 0.19, 95 % CI: 0.12−0.27). A subgroup analysis by classifying the assessment time of depression showed obvious differences of the CRP levels between the PSD patients who were diagnosed more than 1 month after stroke and the non-PSD (1−3 months: SMD = 0.16, 95 % CI: 0.06−0.25; >3months: SMD = 0.34, 95 % CI: 0.18−0.51). Conclusion A higher level of CRP in the acute phase of stroke suggests an increased risk for PSD.

Immunoglobulin G4 (IgG4)-related disease is a systemic disease characterized by sclerosing lesions and an increased serum IgG4 level. This condition can involve any organ, but IgG4-related spinal pachymeningitis is relatively rare. In the current study, we report a case of spinal cord compression caused by IgG4-related spinal pachymeningitis. A 39-year-old man presented to us with a 15-day history of back pain and a 3-day history of dysuresia, exacerbated by weakness in the lower extremities for 2 days. Cervical magnetic resonance imaging (MRI) showed strip-shaped abnormal signals along the anterior and posterior borders of the spinal cord at the C5-T4 levels. The IgG level in cerebrospinal fluid was 718.0 mg/L. Thoracic MRI revealed strip-shaped abnormal signals with remarkable enhancement along the anterior and posterior borders of the dural sac at the T1-T6 levels. Histopathological examination confirmed IgG4-related spinal pachymeningitis. The symptoms worsened rapidly, and surgical resection of the space-occupying lesion in the vertebral canal was performed for spinal decompression. Corticosteroid therapy was administered, and the patient's motor functions were mildly improved. IgG4-related disease can manifest as spinal pachymeningitis and cause spinal cord compression. Clinicians should be aware of this rare condition, and early diagnosis, timely surgical decompression, and appropriate corticosteroid therapy should be highlighted.

Objective To explore the short-term efficacy and tolerability of paroxetine in the treatment of panic disorder in adults. Methods Multiple electronic databases were searched to find randomized controlled trials (RCTs) on paroxetine and panic disorder. The primary efficacy outcomes were: the mean change compared to the baseline in the total number of full panic attacks, Clinical Global Impression-Severity of Illness (CGI-S) score, and the proportion of participants with zero full panic attacks and with a 50% or greater reduction in the number of full panic attacks. The tolerability outcomes included withdrawal rate and the incidence of adverse events (AEs). Results 13RCTs were included. The pooled analyses showed patients who received paroxetine experienced greater improvements in the number of full panic attacks (total: MD=-1.96, 95%CI -3.45 to -0.47, P=0.010; ≥50% reduction: OR=1.66, 95%CI 1.08 to 2.55, P=0.02; zero full panic attacks: OR=1.70, 95%CI 1.42 to 2.03, P < 0.00001) and CGI-S (MD=-0.37, 95%CI -0.74 to -0.01, P=0.05) than placebo. There was no evident difference in the total withdrawal rate (OR=0.91, 95%CI 0.76 to 1.08, P=0.26) and withdrawal rate due to AEs (OR=1.29, 95%CI 0.97 to 1.72, P=0.07) between the two groups. Withdrawal rate due to lack of efficacy or relapse (OR=0.44, 95%CI 0.31 to 0.63, P < 0.00001) and the incidence of serious AEs (OR=0.42, 95%CI 0.23 to 0.79, P=0.007) in the paroxetine group was lower than the placebo group. Meanwhile, the incidence of any treatment-emergent adverse events (TEAEs) (OR=1.32, 95%CI 1.05 to 1.64, P=0.02) in the paroxetine group was higher in comparison with the placebo. Conclusions Paroxetine is an effective and well-tolerated short-term treatment for adults with panic disorder.

Transgenic plants expressing insecticidal proteins originating from Bacillus thuringiensis (Bt) have successfully been used to control lepidopteran and coleopteran pests with chewing mouthparts. However, only a handful of Bt proteins have been identified that have bioactivity against sap sucking pests (Hemiptera), including aphids, whiteflies, plant bugs and planthoppers. A novel Bt insecticidal protein with significant toxicity against a hemipteran insect pest is described here. The gene encoding the 359 amino acid, 40.7 kDa protein was cloned from strain C9F1. After expression and purification of the toxin, its median lethal concentration (LC) values against Laodelphax striatellus and Nilaparvata lugens were determined as 6.89 μg/mL and 15.78 μg/mL respectively. Analysis of the toxin sequence revealed the presence of both Toxin_10 and Ricin_B_Lectin domains.

The vegetative insecticidal proteins (VIPs) of Bacillus thuringiensis (Bt) have a broad-spectrum insecticidal activity against Lepidopteran pests and no cross-resistance with the insecticidal crystal protein Cry protein. So there are great potentials for the control of agricultural pests and the resolution of resistance problems. The structural information of Vip3Aa protein and the predicted key amino acid sites on the C-terminal domain of Vip3Aa were analyzed with the methods of bioinformatics such as homology modeling and molecular docking. Site-directed mutagenesis was used to replace these amino acids with alanine, and there was difference in the activities of the mutant protein and Vip3Aa protein. Y619A had improved insecticidal activity against Helicoverpa armigera, but the toxicity of W552A and E627A to Helicoverpa armigera was significantly reduced. The mutants of W552A and E627A had reduced insecticidal activity against Spodoptera exigua. This study demonstrated that the C-terminal domain played an important role in the function of Vip3Aa protein toxin, and the deletion of the side chain of key residues had a significant effect on the activity of the insecticidal protein. This study provides the theoretical basis for revealing the relationship between the structure and function of Vip3Aa protein.

In this study, we collected 540 soil samples from northeast China and isolated the wild-type strain of Bacillus thuringiensis (Bt) by identifying and cloning 9 Bt strains that expressed the secreted insecticidal protein (Sip) gene. We selected the strain QZL38 for further study. The sip gene was identified from the Bt strain QZL38 using polymerase chain reaction (PCR). We sequenced a 1095-base pair fragment of DNA that encodes 364 amino acid residues of a 41.18 kDa pro-toxin and compared it with the registered Sip1Ab protein amino acid residue sequence. The sequence was submitted to GenBank with the accession no. KP231523, and the gene was named sip1Ab. The Sip1Ab protein expressed in Escherichia coli showed insecticidal activity against Colaphellus bowringi Baly, with an LC50 of 1.051 μg mL−1. To identify the active fragment of the Sip1Ab toxin, four pairs of primers with different truncation positions were designed, and the recombinant proteins were expressed in E. coli. The truncated Sip protei...

Bacillus thuringiensis (Bt) has been used globally as a biopesticide for effective and environmentally friendly pest control. Research has intensified following the development of resistance by lepidopteran species to Bt insecticidal crystal proteins. Discovering new Bt strains with novel toxin properties which can overcome resistance is one of the strategies to improve pesticide sustainability. The genome of the Bacillus thuringiensis LTS290 strain was sequenced and assembled in 252 contigs containing a total of 6,391,328 bp. The novel cry79Aa1 gene from this strain was identified and cloned. Cry79Aa1 contains 729 amino acid residues and a molecular mass of 84.8 kDa by SDS-PAGE analysis. Cry79Aa1 was found to be active against the lepidopteran larvae of Spodoptera exigua, Helicoverpa armigera, and Plutella xylostella with LC50 values of 13.627 µg/mL, 42.8 µg/mL, and 38.086 µg/mL, respectively. However, Cry79Aa1 protein showed almost no insecticidal activity against Leguminivora glycinivorella, although some degree of growth retardation was observed.

The genus Pseudomonas is widely recognized for its potential for environmental remediation and plant growth promotion. Pseudomonas sp. DY-1 was isolated from the agricultural soil contaminated five years by prometryne, it manifested an outstanding prometryne degradation efficiency and an untapped potential for plant resistance improvement. Thus, it is meaningful to comprehend the genetic background for strain DY-1. The whole genome sequence of this strain revealed a series of environment adaptive and plant beneficial genes which involved in environmental stress response, heavy metal or metalloid resistance, nitrate dissimilatory reduction, riboflavin synthesis, and iron acquisition. Detailed analyses presented the potential of strain DY-1 for degrading various organic compounds via a homogenized pathway or the protocatechuate and catechol branches of the β-ketoadipate pathway. In addition, heterologous expression, and high efficiency liquid chromatography (HPLC) confirmed that prometryne could be oxidized by a Baeyer-Villiger monooxygenase (BVMO) encoded by a gene in the chromosome of strain DY-1. The result of gene knock-out suggested that the sulfate starvation-induced (SSI) genes in this strain might also involve in the process of prometryne degradation. These results would provide the molecular basis for the application of strain DY-1 in various fields and would contribute to the study of prometryne biodegradation mechanism as well.

The insecticidal activities and specificities of the Vip3Aa proteins derived from different Bt strains are very different, although the similarities between these proteins are higher than 95%. In t...

Epilepsy is a brain and neurological disorder with high prevalence. It was reported that more than 70% of epileptic seizures were controlled by anti-epileptic medications, whereas the lack of evidence with respect to head-to-head comparisons motivated researchers to seek alternative approaches that are able to provide deep insights into the profile of anti-epileptic medications. In this study, we performed a network meta-analysis (NMA) to evaluate the efficacy and safety of anti-epileptic medications for partial seizures of epilepsy. Publications were retrieved from PubMed, Embase, and Cochrane Library. Then, studies were screened and selected based on the inclusion criteria. Data were extracted and a NMA was performed to combine both direct and indirect evidence. Surface under the cumulative ranking curve (SUCRA) was obtained for ranking purposes. Consistency between direct and indirect evidence was assessed by using the node-splitting method. Seventeen anti-epileptic medications from 90 publications were enrolled. Fifty percent responder and state of seizure freedom were studied as outcomes for efficacy; treatment emergent adverse effect (TEAE), including dizziness, somnolence, headache, fatigue, and nausea were evaluated as safety outcomes. Topiramate, levetiracetam, pregabalin, and oxcarbazepine were recommended for their relatively high efficacy and low-risk of adverse events for partial seizures. Rufinamide was the least preferable medication due to its low efficacy and high-risk of adverse effects. J. Cell. Biochem. 118: 2850-2864, 2017. © 2017 Wiley Periodicals, Inc.

Cerebral small vessel diseases (CSVDs) are common causes of cognitive impairments and mood disorders. In recent years, event-related potential P300 has received increasing attention as a biomarker of cognitive impairments or mood disorders. Previous studies on P300 mainly focused on anxiety, depression or cognitive impairments, and few results have been reported on P300 in CSVD patients. The present study aimed to explore the relationship between neuropsychological test scores and P300 in patients with CSVDs.The clinical data of 52 patients with CSVDs admitted to the Neurology ward of the First Hospital of Jilin University from June 2016 to October 2017 were collected. All patients who met the inclusion criteria were assessed by both cognitive tests and mood scales within 1 week after enrollment, followed by measurement of P300. Accordingly, patients were assigned to the following four groups: cognitive impairment, non-cognitive impairment, mood disorder, and non-mood disorder.The amplitude and latency values of P300 were measured from the Pz, Fz, Fpz, C3, C4 and Cz electrode sites. In addition, correlations of P300 responses and neuropsychological test scores were analyzed.Significant differences were found in the P300 latency values between the cognitive impairment group and non-cognitive impairment group (P0.05). P300 latency values were more significantly prolonged in the mood disorder group at the Fz, C3 and Cz electrode sites than in the non-mood disorder group. Positive correlations were found between Hamilton Depression Scale scores and C3, Fz and Cz latencies. Females tended to have a statistically higher risk of emotional impairment than did males (p0.05).P300 latency values can be used as a objective indicator of cognitive impairments and mood disorders in CSVD patients.

Vip insecticidal proteins are produced by Bacillus thuringiensis (Bt) during its vegetative growth phase. In the present study, Vip3Aa11 and Vip3Aa39 proteins were investigated. These two proteins present 39 amino acid differential sites and they shared 95.06% amino acid sequence similarity. They are effective against some Lepidoptera insect larvae. In a previous study, using artificial diet bioassays, we estimated the LC50 of Vip3Aa11 and Vip3Aa39 strains against Agrotis ipsilon larvae were 73.41 μg/mL (with 95% confidence interval of 2.34–11.19) and 5.43 μg/mL (with 95% confidence interval of 43.20–115.03), respectively. To investigate the response of Agrotis ipsilon transcriptome in defending against Vip3Aa11 and Vip3Aa39 toxins, we performed high-throughput RNA-sequencing on cDNA generated from the midguts of Agrotis ipsilon larvae that consumed a control diet (CK-M-A), Vip3Aa11 (Vip3Aa11-M-A) and Vip3Aa39 (Vip3Aa39-M-A) proteins. We generated about 98.87 Gb bases in total on BGISEQ-500 sequencing platform. After assembling all samples together and filtering the abundance, we got 51,887 unigenes, the total length, average length, N50 and GC content of unigenes are 64,523,651 bp, 1243 bp, 2330 bp and 41.81% respectively. We revealed 558 midgut genes differential expressed in Vip3Aa11-M-A and 65 midgut genes differentially expressed in Vip3Aa39-M-A. The differentially expressed genes were enriched for serine proteases and potential Bt Vip toxin midgut receptor genes. Eleven serine proteases related genes and 13 Bt toxin potential receptor genes with differential expression were found. Based on transcriptome profiling, we focused on validation the sensitivity of these two Vip3Aa proteins to trypsin and their binding properties to Agrotis ipsilon midgut BBMV (Brush Border Membrane Vesicles). The results show that the sensitivity of the two proteins to trypsin is similar. Binding experiments revealed that both proteins can bind to Agrotis ipsilon midgut BBMV, and there is a competitive binding between them. This transcriptome dataset provided a comprehensive sequence resource of Agrotis ipsilon and provides a foundation for comparative studies with other species of insects.

The original version of this article unfortunately contained a mistake. The missing acknowledgement is provided below.

In the context of the deep integration of Internet technology and teaching, the Internet has been increasingly applied in practice teaching. At present, although the national policy supports the network practice teaching of ideological and political theory courses in colleges and universities, it is still in the exploratory stage in terms of practical operation, and some problems in its operation process deserve to be paid attention to, followed up and deeply analyzed.

UNLABELLED Atrazine has been used worldwide for over 50 years as a chemical herbicide. A strain of bacteria, YJY4 which utilizes atrazine as its sole nitrogen source for growth was isolated from agricultural black soil in northeastern China. 16S rDNA sequencing identified YJY4 as a Shewanella sp. PCR analysis and sequencing confirmed that YJY4 contained atrazine-degrading atzA, atzB and atzC genes. These genes revealed high similarity with those in Pseudomonas sp. ADP and Arthrobacter sp. TC1. The strain YJY4 was observed to degrade atrazine (100 mg l(-1) ) to cyanuric acid completely after 36 h. To the best of our knowledge, YJY4 was the first reported Shewanella sp. to grow in pure culture with atrazine serving as a sole source of nitrogen. Therefore, YJY4 may help with atrazine biodegradation and may become an abundant resource of atrazine degradation strains. SIGNIFICANCE AND IMPACT OF STUDY A new isolated strain, YJY4 showed high atrazine-degrading ability, being able to degrade 100 mg l(-1) atrazine completely in 36 h. Strain YJY4 was identified as Shewanella sp. and contained atrazine-degrading atzA, atzB and atzC genes. This study examined the degradation mechanism and metabolic ability of this strain and for the bioremediation of contaminated environments, provides more strain selection and determines the strain of atrazine bioremediation potential.

Bacillus thuringiensis vegetative insecticidal proteins (VIPs) are not homologous to other known Cry proteins, and they act against lepidopteran larvae via a unique process. All reported studies on the mode of action of Vip3 proteins have been performed on the Vip3A family, mostly on the Vip3Aa subfamily. Vip3Aa proteins are activated by midgut proteases, and they cross the peritrophic membrane and bind specific proteins in apical membrane epithelial midgut cells, which results in pore formation and, eventually, death to the insects. Some studies of trypsin-activated protein (core fragment) and the full-length protein show differences in mortality on the same insect species. The N-terminus of Vip3A proteins is responsible for the translocation of the protein across the cell membrane. To determine whether the N-terminus of Vip3Aa11 proteins contribute to insecticidal activity, we exchanged Vip3Aa11 residues with Vip3Aa39 no-core fragment residues using site-directed mutagenesis. Bioassays showed that the toxicity of S9N, S193T, and S194L mutants displayed approximately one- and twofold increases in toxicity against Helicoverpa armigera. Mutant protein R115H demonstrated a threefold decrease in toxicity. This work serves as a guideline for the study of the Vip3Aa11 no-core fragment protein insecticidal mechanism.

Tilletia controversa Kühn (TCK) is an important quarantine pathogen that causes wheat dwarf bunt and results in devastating damage to wheat production. The fungus is difficult to be distinguished from T. caries and T. laevis, which cause wheat common bunt, based on morphological, physiological and symptomatological characteristics of the pathogens. The traditional detection of the fungus can be a long and tedious process with poor accuracy. The inter-simple sequence repeat (ISSR) technique has been used for identifying molecular markers for detection of TCK. Of 28 ISSR primers screened, ISSR-859 amplified a specific 678 bp DNA fragment from all TCK isolates but not from any isolates of the common bunt fungi or other pathogenic fungi tested. Based on the fragment sequence, a pair of sequence characterized amplified region (SCAR) primers was designed, which amplified a 372 bp DNA fragment specifically in TCK. The SCAR marker was detected using as low as 1 ng template DNA of TCK, and was also detected using broken teliospores and DNA from asymptomatic wheat samples. We developed the SYBR Green I and TaqMan Green I and TaqMan real-time polymorphism chain reaction methods to detect TCK with the detection limit of 0.1 fg with asymptomatic wheat samples. Further work is needed to develop a rapid test kit for this pathogenic fungus using the designed specific primers.

Bacillus thuringiensis (Bt) is the most widely used insecticidal microbe due to its specific toxicity and safe use with respect to animals and the environment. In this study, we isolated Bt strain Q52-7 from a soil sample collected in the Qian Shan District, Liao Ning Province, China. We observed that the Q52-7 strain produced spherical crystals. The Bt Q52-7 strain had high toxicity against Asian Cockchafer (Holotrichia parallela), exhibiting an LC50 of 3.80 × 10(9) cfu/g, but is not toxic for Anomala corpulenta Motschulsky and Holotrichia oblita. Using general cry8 primers, we amplified a 1.3 kb fragment with the polymerase chain reaction. Specific primers were designed for the amplified fragment to clone the full-length coding region. A novel gene, cry8Na1, had 69 % sequence similarity with cry8Ca1. cry8Na1 gene was successfully expressed in the HD-73(-) acrystalliferous mutant of Bt subsp. Kurstaki HD-73. Bioassays demonstrated that the Cry8Na1 protein is highly toxic for the H. parallela, with a 50 % lethal concentration of 8.18 × 10(10) colony forming units per gram.

Bacillus thuringiensis (Bt) is the most widely applied type of microbial pesticide due to its high specificity and environmental safety. The activity of Bt is largely attributed to the insecticidal crystal protein encoded by the cry genes. Different insecticidal crystal proteins of Bt have different bioactivity against distinct agricultural insect pests, and combination of these proteins not only increases insecticidal activity, but also overcomes and delays development of resistance. A Bt strain, S185, was isolated from a soil sample collected in Songfeng Shan district, Heilongjiang Province, China. Bt S185 has highly specific insecticidal activity against Coleoptera, and was determined to contain cry8 -type genes by peptide mess fingerprint (PMF) analysis. Application of polymerase chain reaction-restricted fragment length polymorphisms (PCR-RFLP) analysis further determined the genotype due to the high homology of cry8Ea1 and cry8Fa1 genes. Through the full-length primers design, two insecticidal crystal protein genes cry8Ca and cry8Ea were obtained. Using prokaryotic cloning vectors, the recombinant plasmids pEB-cry8Ca and pEB-cry8Ea were transferred into expression host strain Escherichia coli Rosetta, thus the two genes were successfully expressed in heterologous bacteria. Keywords: Bacillus thuringiensis , peptide mess fingerprint, identification, clone, insecticidal crystal protein. African Journal of Biotechnology , Vol 13(25) 2484-2490

Epilepsy is a brain and neurological disorder with high prevalence. It was reported that more than 70% of epileptic seizures were controlled by anti-epileptic medications, whereas the lack of evidence with respect to head-to-head comparisons motivated researchers to seek alternative approaches that are able to provide deep insights into the profile of anti-epileptic medications. In this study, we performed a network meta-analysis (NMA) to evaluate the efficacy and safety of anti-epileptic medications for partial seizures of epilepsy. Publications were retrieved from PubMed, Embase, and Cochrane Library. Then, studies were screened and selected based on the inclusion criteria. Data were extracted and a NMA was performed to combine both direct and indirect evidence. Surface under the cumulative ranking curve (SUCRA) was obtained for ranking purposes. Consistency between direct and indirect evidence was assessed by using the node-splitting method. Seventeen anti-epileptic medications from 90 publications were enrolled. Fifty percent responder and state of seizure freedom were studied as outcomes for efficacy; treatment emergent adverse effect (TEAE), including dizziness, somnolence, headache, fatigue, and nausea were evaluated as safety outcomes. Topiramate, levetiracetam, pregabalin, and oxcarbazepine were recommended for their relatively high efficacy and low-risk of adverse events for partial seizures. Rufinamide was the least preferable medication due to its low efficacy and high-risk of adverse effects. J. Cell. Biochem. 118: 2850-2864, 2017. © 2017 Wiley Periodicals, Inc.

The cry1 -type genes of Bacillus thuringiensis represent the largest cry gene family, which contains 50 distinct holotypes. It is becoming more and more difficult to identify cry1 -type genes using current methods because of the increasing number of cry1 -type genes. In the present study, an improved PCR-restriction fragment length polymorphism (PCR-RFLP) method which can distinguish 41 holotypes of cry1 -type genes was developed. This improved method was used to identify cry1 -type genes in 20 B. thuringiensis strains that are toxic to lepidoptera. The results showed that the improved method can efficiently identify single and clustered cry1 -type genes and can be used to evaluate cry1 -type genes in novel strain collections of B. thuringiensis . Among the detected cry1 -type genes, we identified four novel genes, cry1Ai , cry1Bb , cry1Ja , and cry1La . The bioassay results from the expressed products of the four novel cry genes showed that Cry1Ai2, Cry1Bb2, and Cry1Ja2 were highly toxic against Plutella xylostella , whereas Cry1La2 exhibited no activity. Moreover, Cry1Ai2 had good lethal activity against Ostrinia furnacalis , Hyphantria cunea , Chilo suppressalis , and Bombyx mori larvae and considerable weight loss activity against Helicoverpa armigera .

We analyzed the transcriptional regulation of aco gene cluster and the phenotype of acoR mutant, to determine the effect of acoR deletion on sporulation efficiency and Cry protein production.Sequence of aco gene cluster in Bacillus thuringiensis was analyzed by sequence alignment. RT-PCR was carried out to reveal the transcriptional units of the aco gene cluster. acoR insertion mutant was constructed by homologous recombination. Transcriptional activity was analyzed by promoter fusions with lacZ gene. Comparison of the Cry1Ac protein production was determined by protein quantitation.The aco gene cluster was composed of four genes. The acoABCL formed one transcriptional unit. The transcriptional activity of acoA promoter sharply decreased in sigL and acoR mutants, respectively. Deletion of acoR had no effect on growth and Cry protein production, but decreased the motility of cells and sporulation efficiency.The aco gene cluster is controlled by Sigma 54 and activated by AcoR. Deletion of acoR has no effect on Cry protein production, but decreased the motility of the cells.

In this study, vegetative insecticidal proteins vip3 genes from Bacillus thuringiensis strains were detected based on polymerase chain reaction-high resolution melt (PCR-HRM) analysis. A pair of primers was designed according to the conservative sequences in 150 bp region of the known vip3 subfamily. The 150 bp regions of difference vip3 genes have only a few nucleotide difference vip3 genes were detected in 8 of 11 standard B. thuringiensis strains, and vip3Aa genes, vip3Af genes and vip3Ba gene can be distinguished as different melting curves by this method. The results demonstrate the utility of the HRM assay for mutant screening using vip3 gene. The PCR-HRM method may be a valuable and reliable tool for specific detection and identification of vip3 genes.

16S rDNA and ERIC (Enterobacteia Repetitive Intergenic Consensus Sequences) based on PCR method were tested for the effectiveness of the differentiation of B. thuringiensis and B. cereus. 16S rDNA-PCR primers were designed based on the sequence difference in variable regions of B. cereus 16S rDNA and B. thuringiensis 16S rDNA. 16S rDNA-PCR showed no obvious difference between B. cereus and B. thuringiensis. The only difference was that one 1600-bp amplificon could be obtained from all the three B. Cereus strains, and none amplificon from any B. thuringiensis strains. ERIC was optimized based on previous reports. The genomic DNA was used for the template of ERIC-PCR, and the following DNA fingerprints were analyzed by the agarose gel electrophoresis. The results showed that DNA fingerprint of three B. thuringiensis strains had a unique amplicon less than 100-bp, while DNA fingerprint of three B. cereus strains had none. Moreover, DNA fingerprint of B. cereus showed a 700-bp amplicon, but didn't have any DNA fingerprints of B. thuringiensis genome. Therefore, ERIC-PCR technique should be able to be used for the differentiation of B. thuringiensis and B. cereus.

Angiostrongylus cantonensis is the most common cause of eosinophilic meningitis in humans. It is usually caused by ingestion of raw or inadequately cooked intermediate hosts or food contaminated with infective third-stage larvae. We describe a case of eosinophilic meningitis caused by A. cantonensis in a male Chinese patient. The patient had a history of eating raw fish and snail. We describe the clinical features of the patient, the diagnostic process and treatments. We also provide a brief update for physicians on the characteristics, diagnosis and treatment of eosinophilic meningitis caused by A. cantonensis, with particular emphasis on the update of prevalence and treatment of the disease in China.

Angiostrongylus cantonensis is the most common cause of eosinophilic meningitis in humans. It is usually caused by ingestion of raw or inadequately cooked intermediate hosts or food contaminated with infective third-stage larvae. We describe a case of eosinophilic meningitis caused by A. cantonensis in a male Chinese patient. The patient had a history of eating raw fish and snail. We describe the clinical features of the patient, the diagnostic process and treatments. We also provide a brief update for physicians on the characteristics, diagnosis and treatment of eosinophilic meningitis caused by A. cantonensis, with particular emphasis on the update of prevalence and treatment of the disease in China.

The effects of milbemycin A(4) (MB A(4)), milbemycin oxime A(4) (MBO A(4)) and milbemycin beta(1) (MB beta(1)) on reversing multidrug resistance (MDR) of tumor cells were firstly conducted according to the following research, including MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] assay, the accumulation of adriamycin, the accumulation and efflux of rhodamine 123 (Rh123), the regulations of MDR1 gene, and expression of P-gp. The three milbemycins (5muM) showed strong potency to increase adriamycin cytotoxicity toward adriamycin-resistant human breast carcinoma cells MCF-7/adr with reversal fold (RF) of 21.42, 19.06 and 14.89, respectively. In addition, the mechanisms of milbemycins on P-glycoprotein (P-gp)-mediated MDR demonstrated that the milbemycins significantly increased the intracellular accumulations of adriamycin and Rh123 via inhibiting P-gp transport function. Based on the analysis of the P-gp and MDR1 gene expression using flow cytometry and RT-PCR, the results revealed that milbemycin compounds, particularly MB A(4), could regulate down the expression of the P-gp and MDR1 gene. These findings suggest that the milbemycins probably represent promising agents for overcoming MDR in cancer therapy, and especially MB A(4) is better modulator with the lowest toxicity.

A new Bacillus thuringiensis strain, Bt185, was isolated from HeBei soil samples in China. Observations after transmission electron microscopy found that the strain produced spherical parasporal inclusions similar to that of the B. thuringiensis subsp. japonensis Buibui strain, which showed toxicity to both Anomala corpulenta and Popillia japonica. The plasmid profile seen on an agarose gel revealed that Bt185 contained six large bands of 191 kb, 161 kb, 104 kb, 84 kb, 56 kb, and 37 kb. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed one major band with an estimated molecular mass of 130 kDa. Polymerase chain reaction-restriction fragment length polymorphism results showed that a novel cry8-type gene sequence was found in the Bt185 strain. When we screened for this novel gene sequence, an additional novel cry8-type gene was isolated, having a partial sequence of 2340 bp and encoding a protein of 780 amino acids. Bioassay results showed that Bt185 had no toxicity against several Coleopteran and Lepidopteran pests. However, Bt185 exhibited toxicity against larvae of the Asian cockchafer, Holotrichia parallela. This is the first report of the occurrence of a Bacillus strain that has insecticidal activity against Holotrichia parallela larvae.