Xylanase gene mutation by error-prone pcr and expression in Pichia pastoris / Mutação do gene da xilanase por pcr propensa a erros e expressão em Pichia pastoris

Xylanase can hydrolyze xylan for reducing its anti-nutritional impact and improving nutrient availability, so obtaining suitable xylanase to degrade xylan is essential. Error-prone PCR and gene transformation were used in this study to obtain the ideal xylanase for degrading xylan effectively. The result showed that one mutant xylanase gene with high xylanase expression was obtained. After the mutant xylanase gene was connected with pGAPZA and transformed into Pichia pastoris (P. pastoris), the recombinant P. pastoris with mutant gene was found to produce higher xylanase activity (0.1480 U/mL) than that with the native xylanase gene (0.1360 U/mL) after 12 h incubation (p<0.05). The optimal temperature and pH of xylanase expressed by native and mutant genes were the same, i.e. 40°C and 5.50 (p<0.05). In addition, adding 0.2% Tween 80 during recombinant P. pastoris incubation could significantly increase xylanase yield by about 30-35% (p<0.05). The mutant xylanase could significantly increase xylose yield from wheat meal more than the native xylanase (p<0.05).

A xilanase pode hidrolisar o xilano para reduzir seu impacto antinutricional e melhorar a disponibilidade de nutrientes, portanto, obter xilanase adequada para degradar o xilano é essencial. A PCR propensa a erros e a transformação genética foram utilizadas neste estudo para obter a xilanase ideal para degradar eficazmente a xilana. O resultado mostrou que um gene mutante de xilanase com alta expressão de xilanase foi obtido. Depois que o gene mutante da xilanase foi conectado ao pGAPZA e transformado em Pichia pastoris (P. pastoris), o recombinante P. pastoris com o gene mutante produziu maior atividade de xilanase (0,1480 U / mL) do que com o gene nativo da xilanase (0,1360 U / mL) após 12 h de incubação (p <0,05). A temperatura e o pH ótimos da xilanase expressa pelos genes nativos e mutantes foram os mesmos, ou seja, 40 ºC e 5,50 (p <0,05). Além disso, a adição de Tween 80 a 0,2% durante a incubação de P. pastoris recombinante poderia aumentar significativamente o rendimento de xilanase em cerca de 30-35% (p <0,05). A xilanase mutante poderia aumentar significativamente o rendimento de xilose da farinha de trigo mais do que a xilanase nativa (p <0,05).

Modelo epidémico para el estudio regional de la chikungunya / Epidemic Model for the regional study of chikungunya

En el presente estudio planteamos un modelo matemático como marco cuantitativo para describir el brote epidémico de la enfermedad de chikungunya ocurrido en Guatemala durante las últimas semanas del 2014. El modelo pertenece a la familia de modelos compartimentales, introducidos inicialmente para el estudio epidemiológico de la malaria. Contiene los ingredientes básicos para describir la dinámica de transmisión del virus entre las poblaciones de humanos y mosquitos. El trabajo inicia con un análisis teórico cuyo objetivo es mostrar la estabilidad del modelo y explorar las posibilidades de equilibrio endémico y equilibrio libre de la infección en una población dada. Luego del análisis de estabilidad, se interpretó la incidencia semanal de humanos infectados predicha por el modelo como el valor esperado de las observaciones. Para ello se tomaron en cuenta los datos oficiales reportados durante el 2014 en las memorias epidemiológicas del Ministerio de Salud Pública y Asistencia Social de Guatemala (MSPAS), correspondientes a cuatro municipios del departamento de Escuintla. En cada caso, se realizó una simulación numérica, utilizando métodos estadísticos que permiten obtener el mejor ajuste de parámetros biológicos y poblacionales del modelo frente a los datos reportados por el MSPAS. La simulación permitió caracterizar los registros observados en cada localidad a la luz del modelo en discusión. Los métodos presentados en este trabajo podrían ser útiles como herramienta cuantitativa de apoyo para prever escenarios futuros sobre epidemias regionales de enfermedades ocasionadas por el Aedes aegypti, tales como el dengue, el zika y la chikungunya.

In the present study we propose a quantitative framework to describe the epidemic outbreak of chikungunya disease occurred in Guatemala during the last weeks of 2014. The model belongs to the family of compartmental models, initially introduced for the epidemiological study of malaria. It contains the basic ingredients to describe the dynamics of virus transmission between human and mosquito populations. The study begins with a theoretical analysis aimed at showing the stability of the model in order to explore the possibilities of endemic balance and free balance of the infection in a given population. Once the stable equilibrium points were identified, the weekly incidence of infected humans estimated by the model was considered as the expected value of the observations. To this end, the official data reported during 2014 by the Ministry of Public Health and Social Assistance of Guatemala (MSPAS), corresponding to four municipalities in the department of Escuintla were taken into account. In each case, a numerical simulation was performed using statistical methods that allow obtaining the best fit of biological and population parameters of the model compared to the data reported by the MSPAS. The simulation enables to characterize the records observed in each locality on the grounds of the model under discussion. The quantitative methods presented in this paper could be useful for predicting.

Protease gene shuffling and expression in Pichia pastoris

Four kinds of neutral and alkaline protease genes from Aspergillus oryzae and Bacillus subtilis were isolated and shuffled. The shuffled genes were selected, inserted into pGAPZαA plasmid and transformed into Escherichia coli. The gene which could express high-activity protease was selected by screening the sizes of transparent zones around the colonies on casein plates. After an ideal protease gene was selected, it was sequenced and then transformed into Pichia pastoris X33. The result showed that the base in 1022th position of shuffled protease gene was changed from thymine to cytosine, inferring that cysteine was changed to arginine in the mutant protease. After 48 h incubation for the transformed P. pastoris with the mutant or native protease genes, the mutant protease activity was 36.4% higher than the native protease (P<0.05). The optimal pH and temperature of the mutant protease were 6.5-8.0 and 30-70°C, respectively, which indicated better stability than the native protease (P<0.05).

Clinical features of hepatitis associated aplastic anemia for children by eviden ce-based medical analysis / 中华实用儿科临床杂志

Objective To study the clinical features and curative effect of hepatitis associated aplastic anemia (HAAA) in children of China.Methods Patients' records in China Biological Medicine Database (CBM-disc 1980-2011) and Wan Fang Database were reviewed.The clinical data of the children with newly diagnosed HAAA were respectively studied,including clinical manifestations,blood routine,bone marrow examination and viral serology results,as well as the treatment and clinical effect.Results A total of 50 children were confirmed as HAAA.There were 41 boys and 9 girls.The median age was 8.2 years(range 0.8 to 15.0 years) on diagnosis.The causes of hepatitis could not be identified.The median interval between hepatitis occurrence to blood cell reduction was 10 weeks.Twenty-two cases were diagnosed as severe aplastic anemia and 13 cases as very severe aplastic anemia.Fifteen children died within 1 month after diagnosis.In the immunosuppression treatment group,the percentage of overall responders was 69.2％,which was higher than that of the non-immunosuppression treatment group (18.5％) (x2 =9.920,P ＜ 0.01).Conclusions Severe HAAA is very common in school children,especially in boys.The children with HAAA have a higher early death rate.Immunosuppression therapy is effective if combined with androgenic hormone with an earlier diagnosis.

Prognostic scoring system for locoregional control among the patients with nasopharyngeal carcinoma treated by intensity-modulated radiotherapy / 癌症

The prognostic value of T category for locoregional control in patients with nasopharyngeal carcinoma (NPC) has decreased with the extensive use of intensity-modulated radiotherapy (IMRT). We aimed to develop a prognostic scoring system (PSS) that incorporated tumor extension and clinical characteristics for locoregional control in NPC patients treated with IMRT. The magnetic resonance imaging scans and medical records of 717 patients with nonmetastatic NPC treated with IMRT at Sun Yat-sen University Cancer Center between January 2003 and January 2008 were reviewed. Age, pathologic classification, primary tumor extension, primary gross tumor volume (GTV-p), T and N categories, and baseline lactate dehydrogenase (LDH) level were analyzed. Hierarchical cluster analysis as well as univariate and multivariate analyses were used to develop the PSS. Independent prognostic factors for locoregional relapse included N2-3 stage, GTV-p ≥26.8 mL, and involvement of one or more structures within cluster 3. We calculated a risk score derived from the regression coefficient of each factor and classified patients into four groups: low risk (score 0), intermediate risk (score >0 and ≤1), high risk (score >1 and ≤2), and extremely high risk (score >2). The 5-year locoregional control rates for these groups were 97.4%, 93.6%, 85.2%, and 78.6%, respectively (P < 0.001). We have developed a PSS that can help identify NPC patients who are at high risk for locoregional relapse and can guide individualized treatments for NPC patients.

Long-term outcome and late toxicities of simultaneous integrated boost-intensity modulated radiotherapy in pediatric and adolescent nasopharyngeal carcinoma / 癌症

The application of simultaneous integrated boost-intensity modulated radiotherapy (SIB-IMRT) in pediatric and adolescent nasopharyngeal carcinoma (NPC) is underevaluated. This study aimed to evaluate long-term outcome and late toxicities in pediatric and adolescent NPC after SIB-IMRT combined with chemotherapy. Thirty-four patients (aged 8-20 years) with histologically proven, non-disseminated NPC treated with SIB-IMRT were enrolled in this retrospective study. The disease stage distribution was as follows: stage I, 1 (2.9%); stage III, 14 (41.2%); and stage IV, 19 (55.9%). All patients underwent SIB-IMRT and 30 patients also underwent cisplatin-based chemotherapy. The prescribed dose of IMRT was 64-68 Gy in 29-31 fractions to the nasopharyngeal gross target volume. Within the median follow-up of 52 months (range, 9-111 months), 1 patient (2.9%) experienced local recurrence and 4 (11.8%) developed distant metastasis (to the lung in 3 cases and to multiple organs in 1 case). Four patients (11.8%) died due to recurrence or metastasis. The 5-year locoregional relapse-free survival, distant metastasis-free survival, disease-free survival, and overall survival rates were 97.1%, 88.2%, 85.3%, and 88.2%, respectively. The most common acute toxicities were grades 3-4 hematologic toxicities and stomatitis. Of the 24 patients who survived for more than 2 years, 16 (66.7%) and 15 (62.5%) developed grades 1-2 xerostomia and ototoxicity, respectively. Two patients (8.3%) developed grade 3 ototoxicity; no grade 4 toxicities were observed. SIB-IMRT combined with chemotherapy achieves excellent long-term locoregional control in pediatric and adolescent NPC, with mild incidence of late toxicities. Distant metastasis is the predominant mode of failure.

Colonization and plant growth promoting characterization of endophytic Pseudomonas chlororaphis strain Zong1 isolated from Sophora alopecuroides root nodules

The endophytic strain Zong1 isolated from root nodules of the legume Sophora alopecuroides was characterized by conducting physiological and biochemical tests employing gfp-marking, observing their plant growth promoting characteristics (PGPC) and detecting plant growth parameters of inoculation assays under greenhouse conditions. Results showed that strain Zong1 had an effective growth at 28 ºC after placed at 4-60 ºC for 15 min, had a wide range pH tolerance of 6.0-11.0 and salt tolerance up to 5% of NaCl. Zong1 was resistant to the following antibiotics (µg/mL): Phosphonomycin (100), Penicillin (100) and Ampicillin (100). It could grow in the medium supplemented with 1.2 mmol/L Cu, 0.1% (w/v) methylene blue and 0.1-0.2% (w/v) methyl red, respectively. Zong1 is closely related to Pseudomonas chlororaphis based on analysis the sequence of 16S rRNA gene. Its expression of the gfp gene indicated that strain Zong1 may colonize in root or root nodules and verified by microscopic observation. Furthermore, co-inoculation with Zong1 and SQ1 (Mesorhizobium sp.) showed significant effects compared to single inoculation for the following PGPC parameters: siderophore production, phosphate solubilization, organic acid production, IAA production and antifungal activity in vitro. These results suggest strains P. chlororaphi Zong1 and Mesorhizobium sp. SQ1 have better synergistic or addictive effect. It was noteworthy that each growth index of co-inoculated Zong1+SQ1 in growth assays under greenhouse conditions is higher than those of single inoculation, and showed a significant difference (p < 0.05) when compared to a negative control. Therefore, as an endophyte P. chlororaphis Zong1 may play important roles as a potential plantgrowth promoting agent.

Efficacy of bismuth-based quadruple therapy as first-line treatment for Helicobacter pylori infection / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To evaluate the efficacy of bismuth-based quadruple therapy as the first-line treatment for H.pylori infection.</p><p><b>METHODS</b>A total of 136 patients with H.pylori related peptic ulcer or chronic gastritis were randomized into two groups: 67 patients in bismuth-based quadruple group received esomeprazole 20 mg, clarithromycin 0.5 g,amoxicillin 1.0 g,and bismuth potassium citrate 220 mg for 7 d; 69 patients in standard triple group received esomeprazole 20 mg, clarithromycin 0.5 g and amoxicillin 1.0 g for 7 d. Outcome of eradication therapy was assessed by (14)C-UBT. On ITT and PP analysis, calculating the cost-effectiveness ratio (C/E) and the incremental cost-effectiveness ratio (delta C/delta E).</p><p><b>RESULT</b>On ITT and PP analysis, the eradication rates of the quadruple therapy group were 82.09% and 88.71%, and those of the triple therapy group were 66.67% and 73.02% (P<0.05). The cost-effectiveness ratio of two groups was 4.15 and 4.82; The incremental cost-effectiveness ratio of quadruple therapy group was 1.02 as against triple therapy group.</p><p><b>CONCLUSION</b>Compared to the standard triple therapy regimen, the bismuth-containing quadruple therapy regimen has higher eradication rate and cost-effectiveness, which can be recommended as the fist-line treatment for H.pylori infection.</p>

Caracterización de gemelos con esquizofrenia en el municipio de Santiago de Cuba / Characterization of twins with schizophrenia in the municipality of Santiago de Cuba

Se realizó un estudio descriptivo y transversal de 24 pares de gemelos con diagnóstico de esquizofrenia, independientemente de su cigosidad, del municipio de Santiago de Cuba durante el período febrero-abril de 2007, seleccionados mediante un muestreo aleatorio simple, a fin de caracterizarles. Entre las principales observaciones se tuvo en cuenta que la edad promedio de inicio de los síntomas fue más baja en los monocigóticos y los antecedentes prenatales aumentaron el riesgo de la enfermedad. Se constató la existencia de precedentes familiares, con superioridad de los parientes de primer grado. Hubo una mayor frecuencia de concordancia entre gemelos monocigóticos, así como un alto porcentaje en la heredabilidad.

A descriptive and cross-sectional study was conducted in 24 pairs of twins diagnosed with schizophrenia, irrespective of their zygosity, in Santiago de Cuba municipality from February to April, 2007, selected by means of a simple random sampling to characterize them. It was mainly observed that the average age of symptom onset was lower in monozygotic twins and prenatal history increased the risk of disease. The presence of family history was verified, with a predominance of first-degree relatives. There was a higher frequency of concordance between monozygotic twins, as well as a high percentage of heritability.

Detection of luxS gene of quorum sensing signal system in periodontal pathogen / 中华口腔医学杂志

<p><b>OBJECTIVE</b>To detect the presence and distribution of luxS gene in quorum sensing signal system in the periodontal pathogens.</p><p><b>METHODS</b>The total DNA of Porphyromonas gingivalis (Pg), Fusobacterium nucleatum (Fn), Actinobacillus acitinomycetimcomtans (Aa) were extracted. The presence of luxS was detected by polymerase chain reaction (PCR). The products of PCR were detected by electrophoresis, sequenced and identified by a Blast search of the GenBank database.</p><p><b>RESULTS</b>Electrophoresis, sequencing and Blast searching indicated that the PCR products of Pg were highly consistent with the luxS gene in GenBank. The sequencing result of Fn was also identified with the target gene. The PCR product of Aa was the same as reference through electrophoresis.</p><p><b>CONCLUSIONS</b>Pg, Fn, Aa contain luxS gene. Further studies may be required to investigate the functions of luxS in the periodontal pathogens.</p>

Study of adherence of periodontal pathogens to collagen-treated hydroxyapatite / 华西口腔医学杂志

<p><b>OBJECTIVE</b>The ability of oral bacteria to adhere to tooth surface is associated with their pathogenicity. The objective of this study was to compare the ability of 4 strains of periodontal pathogens attaching to collagen-treated hydroxyapatite (C-HA) beads in order to evaluate the ability of the main periodontal pathogens to form the biofilm on root surface.</p><p><b>METHODS</b>The binding amount and binding percentage of 4 strains to C-HA were measured and compared by 3H-labeled binding assay. 4 strains of periodontal pathogens were Fusobacterium nucleatum (F. nucleatum) ATCC 10953, Porphyrin gingivalis (P. gingivalis) ATCC 33277, Prevotella intermedia (P. intermedia) ATCC 25611 and Hemophilic actinomycetemcomitans (H. actinomycetemcomitans) ATCC 29523.</p><p><b>RESULTS</b>The differences of the percentage of relative adherence between F. nucleatum ATCC 10953 and P. gingivalis ATCC 33277, as well as between H. actinomycetemcomitans ATCC 29523 and P. intermedia ATCC 25611 could not be observed. However, the percentage of relative adherence of F. nucleatum ATCC 10953 and P. gingivalis ATCC 33277 was higher than that of P. intermedia ATCC 25611 and H. actinomycetemcomitans ATCC 29523 (P<0.001), no matter cultured 24 h or 48 h. No significant difference of the percentage of the relative adherence of each stain between 24 h and 48 h cultured time could be found.</p><p><b>CONCLUSION</b>F. nucleatum and P. gingivalis exhibited strong binding ability to C-HA. Their adherence to root surface may play an important role in their local aggregation, biofilm formation during the development and recurrence of the periodontitis.</p>

Clinical study on treatment of advanced non-small-cell lung cancer with Chinese herbal medicine in different stages combined with chemotherapy / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To observe the effect of staged Chinese herbal medicinal therapy combined with chemotherapy in treating patients with non-small-cell lung cancer (NSCLC) of stage III or IV.</p><p><b>METHODS</b>Adopting prospective randomized controlled multi-centered method, the 116 patients enrolled were assigned to the treated group (n = 60) and the control group (n = 56). The control group was treated by chemotherapy alone, while the treated group treated by chemotherapy combined with Chinese herbal medicine, i.e. Kangliu Zengxiao Decoction (KLZX) was administered in the chemotherapy stage, followed with Feiyanning Decoction (FYN) in the stage after ending the chemotherapy. The survival time, Karnofsky score, main clinical symptoms and adverse reactions, etc. were observed.</p><p><b>RESULTS</b>The median survival time was 15.57 months in the treated group, which was higher than that in the control group (11.17 months, P< 0.01). The improvements in tumor related symptoms such as fatigue, dyspnea, etc. in the treated group were better than those in the control group (P <0.05). In addition, adverse reactions such as leucopenia, digestive reaction (such as nausea and vomiting) in the treated group were less than those in the control group (P <0.05).</p><p><b>CONCLUSION</b>Chinese herbal medicine combined with chemotherapy shows favorable effect in improving quality of life and prolonging survival time on patients with advanced NSCLC.</p>

Displasia epifisiometafisaria condromatosa múltiple con opacidades corneales / Multiple chonchomatous epiphysealmetaphyseal dysplasia with corneal opacities

Se presentan dos casos de hermanos cuyo cuadro clínico se caracterizaba por facies grotesca, opacidades corneales, deformaciones osteoarticulares y tumoraciones para-articulares. En uno de ellos se encontraba asociada la combinación de una lesión medular y de los nervios periféricos. Los exámenes radiológicos demuestran que las lesiones osteoarticulares tomaban las epífisis y en menor grado las metáfisis de los huesos, con mucha similitud a las alteraciones descritas en las displasias óseas. Además, en el caso de mayor edad se observaron deformidades en las vértebras cervicales y dorsales de la columna vertebral. Sin embargo, la presencia de opacidades corneales, las características de las facies, la corta estatura, los hallazgos del examen histológico de la médula ósea y de las tumoraciones osteoarticulares cuyas células cartllagionosas se teñían con los colorantes para mucopolisacáridos ácidos, sugieren que este trastorno puede tratarse de una alteración del metabolismo de los mucopolisacáridos. Dentro de la clasificación actual de esta última enfermedad, el estudio clínico de estos hermanos difiere de las formas descritas. Otro hallazgo de importancia fue la presencia de condromas y osteocondromas, hecho que por otra parte no se ha encontrado ni en las displasiias óseas ni en las mucopolisacaridosis conocidas