Riesgo potencial de parásitos zoonóticos presentes en heces caninas en Puerto Escondido, Oaxaca / Potential hazard of zoonotic parasites present in canine feces in Puerto Escondido, Oaxaca

Objetivo. Estimar la prevalencia de parásitos potencialmente zoonóticos en heces caninas de Puerto Escondido. Material y métodos. La ciudad se dividió en diez zonas de estudio y éstas se categorizaron en hábitats natural, urbano y suburbano. Se colectaron muestras fecales caninas del piso. Se recuperaron los parásitos por medio de técnicas coproparasitológicas de flotación y frotis directo para su observación microscópica y posterior identificación. Se estimó la prevalencia parasitaria en las heces caninas. Resultados. Todas las zonas presentaron fecalismo canino. La prevalencia parasitaria fue de 73.33%. Los parásitos con mayor prevalencia fueron Toxocara canis (47.78%), Ancylostoma caninum (17.88%) y Dipylidium caninum (13.89%). Conclusión. El fecalismo canino proviene de perros errantes y con dueño. Del total de parásitos encontrados, 66.66% son zoonóticos. Los factores que favorecen la problemática son el hábitat suburbano, el manejo indeseable de la basura y la tenencia irresponsable de los cánidos.

Objective. To estimate the zoonotic parasites prevalence in feral dog feces in Puerto Escondido. Material and methods. The fecalism frecuency was estimated in ten zones. To identify the parasites parasitological flotation and direct smear methods were used. The parasitic prevalence was estimated in the canine feces. Results. All the zones presented canine fecalism. The parasitic prevalence in the feces was 73.33%. The parasites with the highest prevalence were Toxocara canis (47.78%), Ancylostoma caninum (17.88%), and Dipylidium caninum (13.89%). Conclusion. Canine fecalism comes from strayed and owned dogs. 66.66% of the parasites found in the dog feces are zoonotics. The factors associated to this problem are the suburban habitat, waste mishandling and nil tenure of stray dogs.

Tutorial for writing systematic reviews for the Brazilian Journal of Physical Therapy (BJPT) / Tutorial para elaboração de revisões sistemáticas para o Brazilian Journal of Physical Therapy (BJPT)

Systematic reviews aim to summarize all evidence using very rigorous methods in order to address a specific research question with less bias as possible. Systematic reviews are widely used in the field of physical therapy, however not all reviews have good quality. This tutorial aims to guide authors of the Brazilian Journal of Physical Therapy on how systematic reviews should be conducted and reported in order to be accepted for publication. It is expected that this tutorial will help authors of systematic reviews as well as journal editors and reviewers on how to conduct, report, critically appraise and interpret this type of study design. .

Revisões sistemáticas têm como objetivo sumarizar toda a evidência disponível, através de métodos rigorosos, para responder a uma pergunta de pesquisa específica com o mínimo de viés possível. Revisões sistemáticas são amplamente utilizadas na fisioterapia, porém nem todas as revisões possuem boa qualidade. Esse tutorial tem como objetivo guiar os autores do Brazilian Journal of Physical Therapy sobre como revisões sistemáticas deveriam ser conduzidas e descritas para que sejam aceitas para publicação. Espera-se que esse tutorial irá auxiliar autores de revisões sistemáticas, assim como editores e revisores de periódicos em como conduzir, descrever, fazer análise crítica e interpretar esse tipo de delineamento de pesquisa.

Prejuízos nutricionais e distúrbios no padrão de sono de trabalhadores da Enfermagem / Nutritional damages and disturbances in the sleep pattern of nursing workers / Prejuicios nutricionales y alteraciones en los patrones de sueño de los trabajadores de Enfermería

Este artigo apresenta uma revisão integrativa das publicações científicas da última década, que investigaram os hábitos de sono, a ingestão alimentar e o estado nutricional de profissionais de enfermagem. Foram analisados artigos publicados em periódicos nacionais e internacionais no período de 2002 a 2014, disponibilizados na base de dados PubMed/MEDLINE (USA National Library of Medicine), Lilacs / SciELO (Scientific Eletronic Library Online) e Google Acadêmico. Trinta e um artigos preencheram os critérios estabelecidos. Na análise destes estudos foi identificada elevada prevalência de sobrepeso e obesidade, além de uma modificação negativa nos hábitos alimentares, bem como prejuízos na dinâmica do sono dos profissionais da área de enfermagem.

This article presents an integrative review of national and international scientific publications that investigate the sleep habits, the feed intake and nutritional status of nursing professionals. It was analyzed articles published in national and international journals in the period 2002 to 2014 and made available in the database PubMed / MEDLINE (USA National Library of Medicine), Lilacs / SciELO (Scientific Eletronic Library Online) and Google Scholar. Thirty one articles met the criteria. In the analysis of these studies it has been found a high prevalence of overweight and obesity, a negative change in the eating habits, as well as losses in the sleep patterns of nursing professionals.

En este artículo se presenta una revisión integradora de las publicaciones científicas nacionales e internacionales que investigan los hábitos de sueño, el consumo de alimento y el estado nutricional de los profesionales de enfermería. Se analizaron los artículos publicados en revistas nacionales e internacionales en el período de 2002 a 2014, disponibles en la base de datos PubMed / MEDLINE (USA Biblioteca Nacional de Medicina), Lilacs / SciELO (Scientific Eletronic Library Online) y Google Scholar. Treinta y uno artículos cumplieron con los criterios de inclusión. En el análisis de estos estudios se encontró una alta prevalencia de sobrepeso y obesidad, un cambio negativo en los hábitos alimenticios, así como prejuicios en la dinámica del sueño de los profesionales de enfermería.

Diffusional restrictions in glyoxyl-agarose immobilized penicillin G acylase of different particle size and protein loading / Restricciones para la difusión de penicilina G acilasa inmovilizada en glicosil agarosa

Particle size and enzyme protein loading are design parameters of enzyme immobilization affecting biocatalyst performance that can be varied within broad margins. Their effect on mass transfer limitations at different bulk penicillin G concentrations has been studied with glyoxyl agarose immobilized penicillin G acylase biocatalysts of average particle size of 5·10-5m and 10·10-4m at protein loadings from 15 to 130 mg/g gel. Internal diffusional restrictions were evaluated for such biocatalysts: Thiele modulus varied from 1.17 for the small particles at the lower protein load to 5.84 for the large particles at the higher protein load. Effectiveness factors at different bulk substrate concentrations were determined for all biocatalysts, values ranging from 0.78 for small particle size at 25 mM penicillin G to 0.15 for large particle size at 2 mM penicillin G. Enzyme protein loading had a strong impact on the effectiveness factors of immobilized penicillin G acylase, being it more pronounced in the case of large particle size biocatalysts. At conditions in which 6-aminopenicillanic acid is industrially produced, all biocatalysts tested were mass-transfer limited, being this information valuable for reactor design and performance evaluation.

Phase transfer catalyzed bioconversion of penicillin G to 6-APA by immobilized penicillin acylase in recyclable aqueous two-phase systems with light/pH sensitive copolymers / 南方医科大学学报

Immobilized penicillin acylase was used for bioconversion of penicillin PG into 6-APA in aqueous two-phase systems consisting of a light-sensitive polymer PNBC and a pH-sensitive polymer PADB. Partition coefficients of 6-APA was found to be about 5.78 in the presence of 1% NaCl. Enzyme kinetics showed that the reaction reached equilibrium at roughly 7 h. The 6-APA mole yields were 85.3% (pH 7.8, 20 degrees C), with about 20% increment as compared with the reaction of single aqueous phase buffer. The partition coefficient of PG (Na) varied scarcely, while that of the product, 6-APA and phenylacetic acid (PA) significantly varied due to Donnan effect of the phase systems and hydrophobicity of the products. The variation of the partition coefficients of the products also affected the bioconversion yield of the products. In the aqueous two-phase systems, the substrate, PG, the products of 6-APA and PA were biased in the top phase, while immobilized penicillin acylase at completely partitioned at the bottom. The substrate and PG entered the bottom phase, where it was catalyzed into 6-APA and PA and entered the top phase. Inhibition of the substrate and products was removed to result in improvement of the product yield, and the immobilized enzyme showed higher efficiency than the immobilized cells and occupied smaller volume. Compared with the free enzyme, immobilized enzyme had greater stability, longer life-time, and was completely partitioned in the bottom phase and recycle. Bioconversion in two-phase systems using immobilized penicillin acylase showed outstanding advantage. The light-sensitive copolymer forming aqueous two-phase systems could be recovered by laser radiation at 488 nm or filtered 450 nm light, while pH-sensitive polymer PADB could be recovered at the isoelectric point (pH 4.1). The recovery of the two copolymers was between 95% and 99%.

Antibacterial and toxicological evaluation of beta-lactams synthesized by immobilized beta-lactamase-free penicillin amidase produced by Alcaligenes sp.

Search for anti-beta-lactamase and synthesis of newer penicillin were suggested to overcome resistance to penicillin in chemotherapy. It was found that clavulanic acid, an ant-beta-lactamase was ineffective due to its structural modification by bacteria. Thus, there is a need for the synthesis of newer pencillins. Retro-synthesis was inspired by the success of forward reaction i.e.conversion of penicillin G to 6-aminopenicillanic acid (6-APA) by biological process. In the present study a better enzymatic method of synthesis of newer pencillin by a beta-lactamase-free penicillin amidase produced by Alcaligenes sp. is attempted. Antibacterial and toxicological evaluation of the enzymatically synthesized beta-lactams are reported. Condensation of 6-APA with acyl donor was found to be effective when the reaction is run in dimethyl formamide (DMF 50% v/v) in acetate buffer (25 mM pH 5.0) at 37 degrees C. Periplasm entrapped in calcium alginate exihibited the highest yield (approximately 34%) in synthesis. The minimum inhibitory concentration of the synthetic products against Staphylococcus aureus and Salmonella typhi varied between 20-80 microg/ml. Some of the products exhibited antibacterial activity against enteric pathogens. It was interesting to note that product A was potent like penicillin G. LD50 value of three products (product A, B and C) was more than 12 mg/kg. Furthermore, these synthetic beta-lactams did not exihibit any adverse effect on house keeping enzymes viz., serum glutamate oxalacetate-trans-aminase, serum glutamate pyruvate -trans-aminase, acid phosphatase, alkaline phosphatase of the test animals. The hematological profile (RBC and WBC) of the test animals also remained unaffected.

Dual promoters enhance heterologous enzyme production from bacterial phage based recombinant Bacillus subtilis / 生物工程学报

The effect of dual promoters on recombinant protein production from bacterial phage based Bacillus subtilis expression system was investigated. Alpha amylase (from Bacillus amyloliquefaciens) and penicillin acylase (from Bacillus megaterium) were selected as the indicating enzymes. Both the promoterless genes and the promoter-bearing genes were isolated through PCR amplification with properly designed primers, and were inserted into plasmid pSG703 that contains the lacZ-cat expression cartridge. The lysogenic B. subtilis (phi105 MU331) was transformed with the resultant recombinant plasmids, and the heterologous genes were thereby integrated into the chromosommal DNA of B. subtilis via homologous recombination. The transformants were designated as B. subtilis AMY1, B. subtilis AMY2, B. subtilis PA1, and B. subtilis PA2, respectively. In the recombinant B. subtilis strains, the inserted sequences were located down stream of a strong phage promoter that could be activated by thermal induction. In B. subtilis AMY1 and B. subtilis PA1, transcription of the heterologous genes was only initiated by the phage promoter after heat shock, whereas in B. subtilis AMY2 and B. subtilis PA2, transcription of the heterologous genes was initiated by dual promoters, the phage promoter and the native promoter. The application of dual promoters increased the productivity of both enzymes, with 133% enhancement for alpha-amylase production and 113% enhancement for penicillin acylase production.

Resolution of DL-Phenylglycine by Penicillin G acylase.

The parameters for complete hydrolysis of L-phenyl acetyl phenylglycine (L-PAPG) using immobilized penicillin G acylase (IMEPGA) were investigated. IMEPGA exhibited maximum activity at pH 8.5 and 50 degrees C. The apparent Km value observed was 10 mM. Quantitative hydrolysis (>97%) of the L-PAPG was achieved within 45 min, at pH 7.8 and 37 degrees C, when 0.5% (w/v) of DL-PAPG was used and the concentration of IMEPGA was 133 IU/gm of DL-PAPG. The IMEPGA was used for 50 cycles.

Anticancer activity of N-(phenylacetyl) doxorubicin combined with folate-targeted PGA / 药学学报

<p><b>AIM</b>To demonstrate the specific killing of folate receptor (FR)-positive tumor cells can be achieved by folate-targeted penicillin-G amidase (PGA) combined with its prodrug substrate N-(phenylacetyl) doxorubicin (DOXP).</p><p><b>METHODS</b>Folic acid was covalently linked to PGA and folate content value was determined by quantitative UV spectrophotometry. The ability of folate conjugated PGA to hydrolyze DOXP was measured by RP-HPLC. Visual demonstration of uptake by FR (+) HeLa and SKOV3 cells was detected by using FITC labeled folate-PGA and a fluorescence microscopy. The cytotoxicity of DOXP towards the cells in the presence or absence of folate-PGA was assayed by using MTT method.</p><p><b>RESULTS</b>The folate-PGA has a specific activity of 29. 8 U x mg(-1) (protein). FR selectivity was confirmed by fluorescence microscopy. The combination of DOXP prodrug with folate-PGA generated higher cytotoxicity towards the FR (+) cells than free doxorubicin. The IC50 was 0.72 micromol x L(-1) for HeLa cells and 0.75 micromol x L(-1) for SKOV3 cells, respectively. Further, the enhanced cytotoxicity reduced greatly with the addition of free folic acid.</p><p><b>CONCLUSION</b>Folate conjugated PGA did not significantly compromise PGA catalytic activity and enabled binding prodrug-activating enzyme PGA to folate receptor expressing cells, and increased the sensitivity of the cells to doxorubicin followed by administration of its prodrug substrate.</p>

Optimization of enzymatic synthesis of ampicillin using cross-linked aggregates of penicillin G acylase

Penicillin G acylase from E. coli TA1 was immobilized by Cross-Linked Enzyme Aggregates [CLEA], a new method for immobilization. This biocatalyst and commercial immobilized penicillin G acylase [PGA-450] were used to study the effect of pH, temperature and substrate concentration on the synthesis of ampicillin from phenyl glycine methyl ester [PGME] and 6-aminopenicillanic acid [6-APA]. Compared with PGA-450, this immobilized enzyme showed a high synthesis activity. The optimum conditions for synthetic activity was at pH 6, 25°C and 2:6 [6-APA:PGME] substrate ratio

Constitutive expression and purification of Alcaligenes faecalis penicillin G acylase in Escherichia coli / 生物工程学报

Considering Alcaligenes faecalis pencillin G acylase(AfPGA), which possesses the attractive characteristics for beta-lactam antibiotics conversions, the gene of PGA was cloned into an expressing vector pKKFPGA. The recombinant plasmid contained multicopy replicon(COLE 1), trc promoter, AfPGA gene, rrnB transcript terminator and ampicillin marker transformed Escherichia coli DH5alpha. As both the recombinant plasmid and the host DH5alpha had no laclq gene, the trc promoter was always active and the AfPGA could be constitutively expressed without IPTG induction in the host DH5alpha. In the shaking flask, the recombinant cell was inoculated into the fermentation medium (tryptone 10g/L, yeast extract 5g/L, MgSO4 x 7 H2O 1g, KH2 PO4 2g/L, K2HPO4 x 3H2O 5g/L, Na2HPO4 x 12H2O 7g/L, (NH4)2SO4 1.2g/L, NH4Cl 0.2 g/L, NaCl 0.1g/L, dextrin 30g/L) and cultured at 28 degrees C for 20h. The production of AfPGA reached 2,590u/L(NIPAB method), with a cell-density-specific activity of more than 300(u/L)/A600, this yield increased 432 fold higher than the native expression of Alcaligenes faecalis . Without ammonium sulphate fractionation and dialysis, the supernatant of crude extract was directly loaded on DEAE-Sepharose CL 6B column equilibrated by phosphate buffer (50mmol/L, pH7.8), and the enzyme fraction was not absorbed on the column but impurities were absorbed. Subsequently the effluent was added ammonium sulphate to 1mol/L and loaded on Butyl-Sepharose CL 4B column equilibrated by 50mmol/L phosphate buffer pH7.8-1mol/L ammonium sulphate. The enzyme was eluted as concentration of ammonium sulphate in phosphate buffer decreased to 0, PGA was eluted. After these two column chromatography, the enzyme was enriched 20 times with a 91% activity recovery. The purified enzyme had a specific activity of 68.6u/mg protein. However, the overproduction of PGA was often limited by translocation and/or periplasmic processing steps, subsequently resulted in intracellular accumulation of various types of PGA precursors and then formed inclusion bodies in the cytoplasm and/or periplasm. In this study, 5% PGA precursors formed as inclusion bodies in the cytoplasm while no inclusion bodies formed in the periplasm. It suggested most PGA precursors were transported to the periplasm and matured to active PGA and also explained why PGA gene was highly expressed in the host DH5alpha. On the other hand, inclusion bodies in the cytoplasm indicated that the maturation of PGA in the host DHSalpha was limited by the translocation step.

Nucleotide sequence and protein sequence analysis of GL-7-ACA acylase from Pseudomonas sp. 130 / 生物工程学报

The nucleotide sequence and N-, C-terminal amino acid sequences of alpha,beta-subunit of glutaryl 7-ACA acylase C130 from Pseudomonas sp. 130 were determined. The alignment of the acylase C130 with the other acylases shows that it has high homology with the acylases from Pseudomonas sp. GK16 and C427, but low homology with the others. There is large difference in the N-terminal of alpha-subunit, while the N-terminal of beta-subunit has significant conservation.

Penicillin acylase catalyzed synthesis of penicillin-G from substrates anchored in cyclodextrins.

Penicillin acylase (EC 3.5.1.11) catalyses the condensation of phenylacetic acid (PAA) and 6-aminopenicillanic acid (6-APA) to form benzylpenicillin (BP). Both PAA and 6-APA were found to form host-guest complexes with beta-methylcyclodextrin (beta m-CD) and gamma-cyclodextrin (gamma-CD) respectively. The rate of the reaction catalyzed by the enzyme remained unaffected if one of the substrates used was in the cyclodextrin complexed form. However, in this case, the reaction lasted longer and yielded about 20 per cent more products compared to the condensation reaction involving only uncomplexed substrates. There was distinct increase in the rate of formation of the antibiotic, if both substrates used are in CD-complexed form.

Thermal stability of penicillin G acylase : effect of stabilizing agents.

The role of sugars, polyhydroxy compounds, phenylacetic acid and 6-aminopenicillanic acid in stabilization of immobilized penicillin G acylase (IMPGA) was studied. The loss in the activity of IMPGA at 50 degrees C, 2 h, after incorporation of sucrose and mannitol at 0.1 M concentration was 16 and 18% respectively; the loss in the activity of the enzyme under these conditions in the absence of stabilizing agents was 40%.

Expresión del gen de la penicilino acilasa de la cepa recombinante escherichia coli JM101/pPACII en un sistema de cultivo exponencial de lote alimentado / Expression of the acylasa penicillin gene of the recombinant strain escherichia coli JM101/pPACII in a exponentially culture system of the feed part

La penicilino acilasa es una enzima inducible que cataliza la hidrólisis de las penicilinas generando, entre otros, el intermediario más importante en la producción de las penicilinas semisintéticas de nueva generación, el ácido 6-amino penicilánico (6-Apa). Esta enzima es codificada por el gen estructural pac, el cual, al igual que su producto polipeptídico, se encuentra modulado por diferentes elementos de regulación. En la cepa recombinante Escherichia coli JM101, transformada con el plásmido pPACII, se incluyó este gen con la región reguladora exógena con IPTG. Tradicionalmente, los estudios para evaluar los efectos de los factores ambientales y condiciones de crecimiento sobre la expresión y estabilidad de plásmidos en cepas recombinantes se han realizado en sistemas de escalaminetos por lote o por cultivo contínuo, dificultándose, en primer caso, el control de las condiciones ambientales y, en el segundo, la inherente complejidad operativa del sistema; por lo tanto, en este trabajo se evaluaron los efectos de la concentración del inductor y de la tasa de crecimiento sobre la expresión y estabilidad de la cepa recombinante JM101/pPACII, utilizándose un medio de cultivo balanceado y el sistema de cultivo exponencial de lote alimentado de volumen variable, iniciando la alimentación con un volumen correspondiente al 20 por ciento del volumen final. Un incremento en la expresión del plásmido pPACII para la producción de penicilino acilasa fue observado a medida que disminuyó la tasa de crecimiento y el tiempo de inducción. Se determinó en cultivos por lote, un comportamiento tipo saturación en la expresión del plásmido pPACII a concentraciones superiores de 50 My del indutor, reflejado a través de la actividad máxima de la penicilino acilasa. Por otra parte se observó un incremento en la actividad máxima de la enzima, así como una disminución no significativa en la cinética de crecimiento, a medida que el tiempo de inducción era cercano a la inoculación. Estos resultados demostraron que el cultivo exponencial de lote alimentado de volumen variable puede ser utilizado como una alternativa simple para el estudio de los microorganismos recombinantes

Isolation and purification of penicillin G acylase obtained from Escherichia coli (NCIM-2400) and immobilisation on Eupergit C for the production of 6 amino penicillanic acid.

Penicillin G-Acylase is produced by submerged cultivation of E. Coli (NCIM-2400) and extracted from the harvested fermented broth, purified (affinity chromatography) and immobilised on Eupergit C (Synthetic polymer in bead form). The immobilised penicillin G acylase properties are studied and compared with soluble penicillin G-acylase. The control parameters for conversion of penicillin G-K to 6 APA are optimised [e.g. substrate (Pen G-K) concentration ratio to immobilised penicillin G-acylase, temperature, pH etc.] in a stirred tank reactor. Our findings suggest that immobilised penicillin G-acylase can be used commercially and the productivity of 1 kg. of immobilised enzyme is around 400 kg of 6 APA under given desired stipulated conditions.

Purification and characterization of extracellular penicillin V acylase from Fusarium sp. SKF 235.

Penicillin V acylase from Fusarium sp. SKF 235 culture filtrate was purified to homogeneity. The enzyme was a glycoprotein and composed of single polypeptide chain with molecular weight of 83,200 Daltons. The pH and temperature optima were 6.5 and 55 degrees C, respectively. The KM for penicillin V was 10 mM but the enzyme was inhibited by penicillin V at concentrations above 50 mM. Products of reaction, 6-aminopenicillanic acid and phenoxyacetic acid inhibited the enzyme competitively and noncompetitively with Ki values of 18 mM and 45 mM, respectively. The enzyme specifically hydrolyzed penicillin V, cephalosporanic acid V and penicillin V sulphoxide. Other phenoxy acetyl amides studied were not hydrolysed. It is proposed that phenoxyacetyl moiety alone is not recognized by the penicillin V acylase and in addition, the beta-lactam structure contributes in formation of enzyme-substrate complex.

Cephalosporin acylases: enzyme production, structure and application in the production of 7-ACA.

Cephalosporin acylases have application in the production of 7-aminocephalosporanic acid which forms a key raw material for the preparation of semisynthetic injectable cephalosporins. The enzymes are of industrial importance and hyperproducing genetically engineered strains have been constructed. Different aspects of these enzymes such as subunit structure, post translational modification, primary structure, substrate specificity and their importance in pharmaceutical industry are discussed.

Evaluation of penicillin acylase production by two strains of Bacillus megaterium

Penicillin acylase is a key enzyme for the production of semisynthetic beta-lactam antibiotics. The intracellular enzyme from Escherichia coli has been thoroughly studied and characterized. The extracellular enzyme from Bacillus megaterium, despite its potential advantages, has received less attention in the recent scientific literature. A comparative study is presented for the production of penicillin acylase with two strains of Bacillus megaterium in batch fermentation in previously optimized complex and defined media. The enzyme produced by the selected strain has been recovered, partially purified and its kinetic behaviour determined

Immobilization of penicillin G acylase onto ion exchange and hydrophobic resins.

Penicillin G acylase was immobilized on numerous cation exchange resins and hydrophobic adsorbents. Amberlite XAD-7 was the matrix of choice among the matrices studied for the immobilization of enzyme. Binding of 96.8% and expression of 82.6% of the penicillin G acylase was achieved on XAD-7. Penicillin G acylase immobilised on XAD-7 was used for 80 cycles for the production of 6-PA in a stirred tank reactor.