Exploitation of strong constitutive and stress-driven promoters from Acetobacter pasteurianus for improving acetic acid tolerance.

Acetobacter pasteurianus is an excellent cell factory for production of highly-strength acetic acid, and attracts an increasing attention in metabolic engineering. However, the available well-characterized constitutive and inducible promoters are rather limited to adjust metabolic fluxes in A. pasteurianus. In this study, we screened a panel of constitutive and acid stress-driven promoters based on time-series of RNA-seq data and characterized in A. pasteurianus and Escherichia coli. Nine constitutive promoters ranged in strength from 1.7-fold to 100-fold that of the well-known strong promoter Padh under non-acetic acid environment. Subsequently, an acetic acid-stable red fluorescent visual reporting system was established and applied to evaluate acid stress-driven promoter in A. pasteurianus during highly-acidic fermentation environment. PgroES was identified as acid stress-driven strong promoters, with expression outputs varied from 100% to 200% when acetic acid treatment. To assess their application potential, ultra-strong constitutive promoter Ptuf and acid stress-driven strong promoter PgroES were selected to overexpress acetyl-CoA synthase and greatly improved acetic acid tolerance. Notably, the acid stress-driven promoter displayed more favorable for regulating strain robustness against acid stress by overexpressing tolerance gene. In summary, this is the first well-characterized constitutive and acid stress-driven promoter library from A. pasteurianus, which could be used as a promising toolbox for metabolic engineering in acetic acid bacteria and other gram-negative bacteria.

Pectin mediated synthesis of nano hydroxyapatite-decorated poly(lactic acid) honeycomb membranes for tissue engineering.

The fabrication of porous films with bioactive nanoparticles has been arousing interest in nanobiotechnology. The biocompatible nanocomposite membrane could mimic the functions of basement membrane besides the augmented cell functions including adhesion, spreading, proliferation, and differentiation. We have reported the green template synthesis of nano hydroxyapatite (nHAp) using pectin from bitter gourd fruits followed by the fabrication of nHAp (2 wt%, 4 wt%, and 6 wt%) incorporated honeycomb-like poly(lactic acid) (PLA) films to evaluate the effect of nHAp on the surface patterning of PLA films. The use of naturally available bitter gourd with pectin content could efficiently stabilize or envelop the nanoparticles after nucleation process, resulting in reduced particle size. The cell viability over normal cells and cancer cells were evaluated for H-PLA/nHAp films. The present work recommends the biofriendly surface engineering of PLA films with the assistance of nHAp particles for bone tissue engineering.

Influência do ácido lático e da embalagem em atmosfera modificada sobre a validade comercial da linguiça frescal de frango / Influence of lactic acid and modified atmosphere packing in shelf life of fresh poultry sausage

O objetivo deste trabalho foi caracterizar a influência da embalagem em atmosfera modificada (EAM) e do ácido lático (AL) navalidade comercial da linguiça frescal de frango. Para tanto, foram processadas duas massas de linguiça, sendo uma delasadicionado ácido lático (0,15%). As linguiças foram então embaladas nas seguintes atmosferas: 100%ar atmosférico, 100%N2, 100% CO2, 80% CO2/20% N2, 40% CO2/60% N2 e 20% CO2/80% N2, e armazenadas durante 16 dias a 4±2oC. Foramrealizadas, nos dias 0, 1, 9 e 16, análises bacteriológicas: contagem em placa de bactérias heterotróficas aeróbias mesófilas,bactérias heterotróficas aeróbias psicrotróficas, bactérias ácido-láticas, enterobactérias e Pseudomonas; e determinaçãodo pH. A EAM a 80/20 CO2/N2 mostrou-se como método mais eficaz, sob o ponto de vista microbiológico, em termos deconservação da linguiça frescal de frango, atingindo ao final do experimento valores de 6,0; 5,3 e 5,0 Log UFC/g (bactériasmesófilas, psicrotróficas e ácido-láticas, respectivamente). Em relação à adição do AL, tal tratamento foi eficaz sob o aspectode inibição microbiológica em quase todas as atmosferas, com exceção da 80/20 CO2/N2. Durante o experimento, não foiobservado crescimento de enterobactérias e Pseudomonas. Em relação ao pH, pôde-se observar que a adição de 0,15% deAL provocou uma queda do pH de 5,89 para ~5,50, suficiente para acarretar uma inibição significativa da taxa de multiplicaçãomicrobiana nas diferentes atmosferas, exceto na 80/20 CO2/N2. Pôde se concluir que a EAM com 80/20 CO2/N2, mostrou-secomo o método de conservação mais eficaz quanto ao aumento da vida útil da linguiça de frango e a adição de 0,15% de ALdemonstrou ser uma alternativa eficaz para conservação deste produto.

The present work has an objective to observe and to characterize, through microbiological analyses and pH determination, the influence of the Modified Atmosphere Packaging (MAP) and the lactic acid addictive in shelf life of fresh poultry sausage. Two sausage batches were made under laboratory control. One of them was added of lactic acid (0.15%). The samples were processed, and packaged in plastic bags (four sausages per bag). Finally, the bags were filled with different atmospheres: 100% air, 100% N2, 100% CO2, 80/20 CO2/N2, 40/60 CO2/N2 and 20/80 CO2/N2. Samples were stored in walk?in cold rooms at 4±2°C. Samples were taken at different days of storage (zero, 1, 9 and 16). Both added and not added with lactic acid samples were subjected to total viable aerobic counts (mesophylic and psycrophylic), lactic acid bacteria, Enterobacteriaceae, and Pseudomonas sp. in specific media plates. Also, it was determinate the pH of all the samples. The results were arranged in tables and graphs for the descriptive statistical analyses. The MAP 80/20 CO2/N2 was the more effective method to conserve fresh poultry sausages, reaching at the end of the experiment values of 6.0; 5,3 and 5.0 Log UFC/g (mesophylic, psycrophylic and acid-lactic bacteria, respectively), comparing to the conventional package (100% air) and 100% N2 that reached at the end of the experiment maximum values of 8.8 and 9.4 Log UFC/g, respectively. The addition of the acid lactic was effective about the aspect of microbiological inhibition, reaching inferior values in almost all the atmospheres comparing to the samples without addition of the acid, except to 80/20 CO2/N2, that the values of the microbial count in both treatments (with and without acid) they were very close. During the experiment, it was not observed the growth of Enterobacteriaceae and Pseudomonas. Analyzing the pH, it can be observed that the addition of 0.15% lactic acid caused a fall of the pH of 5.89 for ~5.50, enough to inhibit the microbial multiplication in the different atmospheres, except in 80/20 CO2/N2. After analyzed this results, it was concluded that MAP to 80/20% of CO2/N2 was the more effective method to increase the shelf life of the fresh poultry sausage, added or not added with lactic acid; the addition of 0.15% of lactic acid was an effective alternative for conservation of fresh poultry sausage chicken, wrapped or not in modified atmosphere