Rapid detection of antibiotic resistance genes in lactic acid bacteria using PMMA-based microreactor arrays.

The emergence of lactic acid bacteria (LABs) resistant to existing antimicrobial drugs is a growing health crisis. To decrease the overuse of antibiotics, molecular diagnostic systems that can rapidly determine the presence of antibiotic resistance (AR) genes in LABs from yogurt samples are needed. This paper describes a fully integrated, miniaturized plastic chip and closed-tube detection chemistry that performs multiplex nucleic acid amplification. High-throughput identification of AR genes was achieved through this approach, and six AR genes were analyzed simultaneously in < 2 h. This time-to-result included the time required for the extraction of DNA. The detection limit of the chip was 103 CFU mL-1, which was consistent with that of tube LAMP. We detected and identified multiple DNAs, including streptomycin, tetracycline, and vancomycin resistance-associated genes, with complete concordance to the Kirby-Bauer disk diffusion method.Key Points• A miniaturized chip was presented, and multiplex nucleic acid amplification was performed.• The device can be integrated with LAMP for rapid detection of antibiotic resistance genes.• The approach had a high throughput of AR gene analysis in lactic acid bacteria.

Differential effects of major inhibitory compounds from sugarcane-based lignocellulosic hydrolysates on the physiology of yeast strains and lactic acid bacteria.

OBJECTIVES: Major lignocellulosic inhibitory compounds found in sugarcane-based industrial hydrolysate samples were tested in laboratory and industrial yeast strains, as well as in lactic acid bacteria, in order to verify their effects on important physiological parameters. RESULTS: Saccharomyces cereviaise SA-1, an industrial strain, stood out as compared to the remaining strains for virtually all inhibitors investigated. This strain presented the highest growth rate and the lowest lag-phase in the presence of acetic acid, levulinic acid, p-coumaric acid, ferulic acid, and HMF, when compared to the other strains. In sugarcane-based hydrolysate fermentations, both SA-1 and CEN.PK113-7D presented similar fermentation performances. Industrial isolates of contaminating lactic acid bacteria were evaluated in the presence of an inhibitory cocktail, containing a mixture of 76.6 mM acetic acid, 1.3 mM HMF, 7.1 mM furfural, and 1.9 mM p-coumaric acid. Whilst all yeast strains were unable to grow under such conditions, bacteria had an average inhibition of roughly 50% on their growth rates. CONCLUSIONS: Overall, industrial strain SA-1 might be a promising microbial chassis for second generation ethanol production and for future metabolic and evolutionary engineering strategies, and for strain robustness understanding.

Exploring the Behavior and Metabolic Transformations of SeNPs in Exposed Lactic Acid Bacteria. Effect of Nanoparticles Coating Agent.

The behavior and transformation of selenium nanoparticles (SeNPs) in living systems such as microorganisms is largely unknown. To address this knowledge gap, we examined the effect of three types of SeNP suspensions toward Lactobacillus delbrueckii subsp. bulgaricus LB-12 using a variety of techniques. SeNPs were synthesized using three types of coating agents (chitosan (CS-SeNPs), hydroxyethyl cellulose (HEC-SeNPs) and a non-ionic surfactant, surfynol (ethoxylated-SeNPs)). Morphologies of SeNPs were all spherical. Transmission electron microscopy (TEM) was used to locate SeNPs in the bacteria. High performance liquid chromatography (HPLC) on line coupled to inductively coupled plasma mass spectrometry (ICP-MS) was applied to evaluate SeNP transformation by bacteria. Finally, flow cytometry employing the live/dead test and optical density measurements at 600 nm (OD600) were used for evaluating the percentages of bacteria viability when supplementing with SeNPs. Negligible damage was detected by flow cytometry when bacteria were exposed to HEC-SeNPs or CS-SeNPs at a level of 10 µg Se mL-1. In contrast, ethoxylated-SeNPs were found to be the most harmful nanoparticles toward bacteria. CS-SeNPs passed through the membrane without causing damage. Once inside, SeNPs were metabolically transformed to organic selenium compounds. Results evidenced the importance of capping agents when establishing the true behavior of NPs.

Effects of nitrite incorporated active films on quality of pork.

Nitrite (0.25-2%) incorporated thermoplastic starch (TPS) and linear low-density polyethylene (LLDPE) blend films were produced by the conventional blown-film extrusion process. Films were characterized and determined for efficacy as active packaging for pork steak. Scanning electron micrographs showed enhanced starch granule disruption due to melting. Increased nitrite contents enhanced the starch network and improved mechanical properties. Water vapor and barrier properties of blend films were improved by nitrite incorporation, with increased compatibility between TPS and LLDPE networks. Films containing 0.5% nitrite effectively improved redness and corresponded to 0.06 ppm residual nitrite and 75 ppm nitrosomyoglobin in packaged pork. Nitrite addition modified protein secondary conformation involving CO stretching bonds via H-bonding, while films with 1% and 2% nitrite significantly inhibited growth of lactic acid bacteria, yeast and mold, and retained softer texture during storage. Nitrite incorporated films were demonstrated as efficient active packaging to improve the quality of red meat products.

Physicochemical responses and microbiological changes of bream (Megalobrama ambycephala) to pectin based coatings enriched with clove essential oil during refrigeration.

The effectiveness of pectin coatings enriched with clove essential oil (CEO), as new edible coatings were investigated to preserve bream (Megalobrama ambycephala) fillets during refrigeration over a period of 15 days. All samples were analyzed for physicochemical (pH, PV, TBA and TVB-N), microbiological (Total viable count, Psychrophilic bacteria, Lactic acid bacteria, Enterobacteriaceae, Pseudomonas spp., H2S producing bacteria) and organoleptic attributes. The results revealed that the CEO incorporation reduced the extent of lipid oxidation, as judged by PV, TBA and TVB-N, thus extending the shelf life of bream fillets by at least 15 days. Moreover, the application of pectin coatings with CEO improved the weight loss, water holding capacity, textural and color attributes of the bream samples significantly compared to untreated sample. Pectin coating along with CEO was effective in inhibiting bacterial growth especially in gram-negative bacteria, while the growth of lactic acid bacteria remained constant for most of the storage period. The effect on the microorganisms during storage was in accordance with biochemical indexes of the quality, representing the viability of these coatings for bream preservation. Thus, the coatings developed in present study could inhibit the development of lipid oxidation during cold storage, representing an option as a seafood preservative.