Search results for: bacterial 16S rRNA

Authors: Metehan Mutlu, Meltem Elitas

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This study aims to provide an automated method for placing the balloon uterine stents after hysteroscopy adhesiolysis. Currently, there are no automatized tools to place the balloon uterine stent; therefore, surgeons into the endometrial cavity manually fit it. However, it is very hard to pass the balloon stent through the cervical canal, which is roughly 10mm after the surgery. Our method aims to provide an effective and practical way of placing the stent, by automating the procedure through our designed device. Furthermore, our device does the required tasks fast compared to traditional methods, reduces the narcosis time, and decreases the bacterial contamination risks.

Keywords: balloon uterine stent, endometrial cavity, hysteroscopy, motorized-tool

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Authors: Blessing C. Nwachukwu, Michael O. Taiwo, Wasiu A. Abibu, Isaac O. Ayodeji

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Milk is an important source of micro and macronutrients for humans. Soft Cheese (Wara) is an example of a by-product of milk. In addition, water is considered as one of the most vital resources in cattle farms. Due to the high consumption rate of milk and soft cheese and the traditional techniques involved in their production in Nigeria, there was a need for a microbiological assessment which will be of utmost public health importance. The study thus investigated microbial risk assessments associated with consumption of milk and soft cheese (Wara). It also investigated common pathogens present in dairy water in farms and antibiotic sensitivity profiling for implicated pathogens were conducted. Samples were collected from three different Fulani dairy herds in Ido local government, Ibadan, Oyo State, Nigeria and subjected to microbiological evaluation and antimicrobial susceptibility testing. Aspergillus flavus was the only isolated fungal isolate from Wara while Staphylococcus aureus, Vibro cholera, Hafnia alvei, Proteus mirabilis, Escherishia coli, Psuedomonas aeuroginosa, Citrobacter freundii, and Klebsiella pneumonia were the bacteria genera isolated from Wara, dairy milk and dairy drinking water. Bacterial counts from Wara from the three selected farms A, B and C were 3.5×105 CFU/ml, 4.0×105 CFU/ml and 5.3×105 CFU/ml respectively while the fungal count was 3CFU/100µl. The total bacteria count from dairy milk from the three selected farms A, B and C were Farms 2.0 ×105 CFU/ml, 3.5 × 105 CFU/ml and 6.5 × 105 CFU/ml respectively. 1.4×105 CFU/ml, 1.9×105 CFU/ml and 4.9×105 CFU/ml were the recorded bacterial counts from dairy water from farms A, B and C respectively. The highest antimicrobial resistance of 100% was recorded in Wara with Enrofloxacin, Gentamycin, Cefatriaxone and Colistin. The highest antimicrobial susceptibility of 100% was recorded in Raw milk with Enrofloxacin and Gentamicin. Highest antimicrobial intermediate response of 100% was recorded in Raw milk with Streptomycin. The study revealed that most of the cheeses sold at Ido local Government are contaminated with pathogens. Further research is needed on standardizing the production method to prevent pathogens from gaining access. The presence of bacteria in raw milk indicated contamination due to poor handling and unhygienic practices. Thus, drinking unpasteurized milk is hazardous as it increases the risk of zoonoses. Also, the Provision of quality drinking water is crucial for optimum productivity of dairy. Health education programs aiming at increasing awareness of the importance of clean water for animal health will be helpful.

Keywords: dairy, raw milk, soft cheese, Wara

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Authors: Shikha Rani, Neeta Sehgal, Vipin Kumar Verma, Om Prakash

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Introduction: Fish is an important protein source for humans and has great economic value. Fish cultures are affected due to various anthropogenic activities that lead to bacterial and viral infections. Aeromonas hydrophila is a fish pathogenic bacterium that causes several aquaculture outbreaks throughout the world and leads to huge mortalities. In this study, plants of no commercial value were used to investigate their immunostimulatory, antioxidant, anti-inflammatory, anti-bacterial, and disease resistance potential in fish against Aeromonas hydrophila, through fish feed fortification. Methods: The plant was dried at room temperature in the shade, dissolved in methanol, and analysed for biological compounds through GC-MS/MS. DPPH, FRAP, Phenolic, and flavonoids were estimated following standardized protocols. In silico molecular docking was also performed to validate its broad-spectrum activities based on binding affinity with specific proteins. Fish were divided into four groups (n=6; total 30 in a group): Group 1, non-challenged fish (fed on a non-supplemented diet); Group 2, fish challenged with bacteria (fed on a non-supplemented diet); Group 3 and 4, fish challenged with bacteria (A. hydrophila) and fed on plant supplemented feed at 2.5% and 5%. Blood was collected from the fish on 0, 7th, 14th, 21st, and 28th days. Serum was separated for glutamic-oxaloacetic transaminase (SGOT), serum glutamic pyruvic transaminase (SGPT), alkaline phosphatase assay (ALP), lysozyme activity assay, superoxide dismutase assay (SOD), lipid peroxidation assay (LPO) and molecular parameters (including cytokine levels) were estimated through ELISA. The phagocytic activity of macrophages from the spleen and head kidney, along with quantitative analysis of immune-related genes, were analysed in different tissue samples. The digestive enzymes (Pepsin, Trypsin, and Chymotrypsin) were also measured to evaluate the effect of plant-supplemented feed on freshwater fish. Results and Discussion: GC-MS/MS analysis of a methanolic extract of plant validated the presence of key compounds having antioxidant, anti-inflammatory, anti-bacterial, anti-inflammatory, and immunomodulatory activities along with disease resistance properties. From biochemical investigations like ABTS, DPPH, and FRAP, the amount of total flavonoids, phenols, and promising binding affinities towards different proteins in molecular docking analysis helped us to realize the potential of this plant that can be used for investigation in the supplemented feed of fish. Measurement liver function tests, ALPs, oxidation-antioxidant enzyme concentrations, and immunoglobulin concentrations in the experimental groups (3 and 4) showed significant improvement as compared to the positive control group. The histopathological evaluation of the liver, spleen, and head kidney supports the biochemical findings. The isolated macrophages from the group fed on supplemented feed showed a higher percentage of phagocytosis and a phagocytic index, indicating an enhanced cell-mediated immune response. Significant improvements in digestive enzymes were also observed in fish fed on supplemented feed, even after weekly challenges with bacteria. Hence, the plant-fortified feed can be recommended as a regular feed to enhance fish immunity and disease resistance against the Aeromonas hydrophila infection after confirmation from the field trial.

Keywords: immunostimulation, antipathogen, plant fortified feed, macrophages, GC-MS/MS, in silico molecular docking

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Authors: Dmitriy Berillo, Areej K. A. Al-Jwaid, Jonathan L. Caplin, Andrew Cundy, Irina Savina

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Chlorophenols (CPs) are used as a precursor in the production of higher CPs and dyestuffs, and as a preservative. Contamination by CPs of the ground water is located in the range from 0.15-100mg/L. The EU has set maximum concentration limits for pesticides and their degradation products of 0.1μg/L and 0.5μg/L, respectively. People working in industries which produce textiles, leather products, domestic preservatives, and petrochemicals are most heavily exposed to CPs. The International Agency for Research on Cancers categorized CPs as potential human carcinogens. Existing multistep water purification processes for CPs such as hydrogenation, ion exchange, liquid-liquid extraction, adsorption by activated carbon, forward and inverse osmosis, electrolysis, sonochemistry, UV irradiation, and chemical oxidation are not always cost effective and can cause the formation of even more toxic or mutagenic derivatives. Bioremediation of CPs derivatives utilizing microorganisms results in 60 to 100% decontamination efficiency and the process is more environmentally-friendly compared with existing physico-chemical methods. Microorganisms immobilized onto a substrate show many advantages over free bacteria systems, such as higher biomass density, higher metabolic activity, and resistance to toxic chemicals. They also enable continuous operation, avoiding the requirement for biomass-liquid separation. The immobilized bacteria can be reused several times, which opens the opportunity for developing cost-effective processes for wastewater treatment. In this study, we develop a bioremediation system for CPs based on macroporous materials, which can be efficiently used for wastewater treatment. Conditions for the preparation of the macroporous material from specific bacterial strains (Pseudomonas mendocina and Rhodococus koreensis) were optimized. The concentration of bacterial cells was kept constant; the difference was only the type of cross-linking agents used e.g. glutaraldehyde, novel polymers, which were utilized at concentrations of 0.5 to 1.5%. SEM images and rheology analysis of the material indicated a monolithic macroporous structure. Phenol was chosen as a model system to optimize the function of the cryogel material and to estimate its enzymatic activity, since it is relatively less toxic and harmful compared to CPs. Several types of macroporous systems comprising live bacteria were prepared. The viability of the cross-linked bacteria was checked using Live/Dead BacLight kit and Laser Scanning Confocal Microscopy, which revealed the presence of viable bacteria with the novel cross-linkers, whereas the control material cross-linked with glutaraldehyde(GA), contained mostly dead cells. The bioreactors based on bacteria were used for phenol degradation in batch mode at an initial concentration of 50mg/L, pH 7.5 and a temperature of 30°C. Bacterial strains cross-linked with GA showed insignificant ability to degrade phenol and for one week only, but a combination of cross-linking agents illustrated higher stability, viability and the possibility to be reused for at least five weeks. Furthermore, conditions for CPs degradation will be optimized, and the chlorophenol degradation rates will be compared to those for phenol. This is a cutting-edge bioremediation approach, which allows the purification of waste water from sustainable compounds without a separation step to remove free planktonic bacteria. Acknowledgments: Dr. Berillo D. A. is very grateful to Individual Fellowship Marie Curie Program for funding of the research.

Keywords: bioremediation, cross-linking agents, cross-linked microbial cell, chlorophenol degradation

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Authors: Dalal Alghawas, Jetty Lee, Kaisa Poutanen, Hani El-Nezami

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Oat β-glucan a water soluble non starch linear polysaccharide has been approved as a cholesterol lowering agent by various food safety administrations and is commonly used to reduce the risk of heart disease. The molecular weight of oat β-glucan can vary depending on the extraction and fractionation methods. It is not clear whether the molecular weight has a significant impact at reducing the acceleration of atherosclerosis. The aim of this study was to investigate three different oat β-glucan fractionations on the development of atherosclerosis in vivo. With special focus on plaque stability and the intestinal barrier function. To test this, ApoE-/- female mice were fed a high fat diet supplemented with oat bran, high molecular weight (HMW) oat β-glucan fractionate and low molecular weight (LMW) oat β-glucan fractionate for 16 weeks. Atherosclerosis risk markers were measured in the plasma, heart and aortic tree. Plaque size was measured in the aortic root and aortic tree. ICAM-1, VCAM-1, E-Selectin, P-Selectin, protein levels were assessed from the aortic tree to determine plaque stability at 16 weeks. The expression of p22phox at the aortic root was evaluated to study the NADPH oxidase complex involved in nitric oxide bioavailability and vascular elasticity. The tight junction proteins E-cadherin and beta-catenin from western blot analyses were analysed as an intestinal barrier function test. Plasma LPS, intestinal D-lactate levels and hepatic FMO gene expression were carried out to confirm whether the compromised intestinal barrier lead to endotoxemia. The oat bran and HMW oat β-glucan diet groups were more effective than the LMW β-glucan diet group at reducing the plaque size and showed marked improvements in plaque stability. The intestinal barrier was compromised for all the experimental groups however the endotoxemia levels were higher in the LMW β-glucan diet group. The oat bran and HMW oat β-glucan diet groups were more effective at attenuating the development of atherosclerosis. Reasons for this could be due to the LMW oat β-glucan diet group’s low viscosity in the gut and the inability to block the reabsorption of cholesterol. Furthermore the low viscosity may allow more bacterial endotoxin translocation through the impaired intestinal barrier. In future food technologists should carefully consider how to incorporate LMW oat β-glucan as a health promoting food.

Keywords: Atherosclerosis, beta glucan, endotoxemia, intestinal barrier function

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Authors: Delsy Salinas, Andreia Garcês, Augusto Silva, Paula Brilhante Simões

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Urinary tract infection (UTI) is a common disease affecting dogs and cats in both community and hospital environment. Bacteria is the most frequent agent isolated, fewer than 1% of infections are due to parasitic, fungal, or viral agents. Common symptoms and laboratory abnormalities includeabdominal pain, pyrexia, renomegaly, and neutrophilia with left shift. A rapid and precise identification of the bacterial agent is still a challenge in veterinarian laboratories. Therefore, this cross-sectional study aims to describe bacterial colony patterns of urine samples by using chromID™ CPS® EliteAgar (BioMérieux, France) from canine and feline specimens submitted to a veterinary laboratory in Portugal (INNO Veterinary Laboratory, Braga)from January to March2022. All urine samples were cultivated in CPS Elite Agar with calibrated 1 µL inoculating loop and incubated at 37ºC for 18-24h. Color,size, and shape (regular or irregular outline)were recorded for all samples. All colonies were classified as Gram-negative or Gram-positive bacteriausing Gram stain (PREVI® Color BioMérieux, France) and determined if they were pure colonies. Identification of bacteria species was performed using GP and GN cards inVitek 2® Compact(BioMérieux, France). A total of 256/1003 submitted urine samples presented bacterial growth, from which 172 isolates were included in this study. The sample’s population included 111 dogs (n=45 males and n=66 females) and 61 cats (n=35 males and n=26 females). The most frequent isolated bacteria wasEscherichia coli (44,7%), followed by Proteus mirabilis (13,4%). All Escherichia coli isolates presented red to burgundy colonies, a colony diameter between 2 to 6 mm, and regular or irregular outlines. Similarly, 100% of Proteus mirabilis isolates were dark yellow colonies with a diffuse pigment and the same size and shape as Escherichia coli. White and pink pale colonies where Staphylococcus species exclusively and S. pseudintermedius was the most frequent (8,2 %). Cian to blue colonies were mostly Enterococcusspp. (8,2%) and Streptococcus spp. (4,6%). Beige to brown colonies were Pseudomonas aeruginosa (2,9%) and Citrobacter spp. (1,2%).Klebsiella spp.,Serratia spp. and Enterobacter spp were green colonies. All Gram-positive isolates were 1 to 2 mm diameter long and had a regular outline, meanwhile, Gram-negative rods presented variable patterns. This results showed that theprevalence of E coli and P. mirabilis as uropathogenic agents follows the same trends in Europe as previously described in other studies. Both agents presented a particular color pattern in CPS Elite Agar to identify them without needing complementary tests. No other bacteria genus could be correlated strongly to a specific color pattern, and similar results have been observed instudies using human’s samples. Chromogenic media shows a great advantage for common urine bacteria isolation than traditional COS, McConkey, and CLEDAgar mediums in a routine context, especially when mixed fermentative Gram-negative agents grow simultaneously. In addition, CPS Elite Agar is versatile for Artificial Intelligent Reading Plates Systems. Routine veterinarian laboratories could use CPS Elite Agar for a rapid screening for bacteria identification,mainlyE coli and P.mirabilis, saving 6h to 10h of automatized identification.

Keywords: cats, CPS elite agar, dogs, urine pathogens

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Authors: Z. C. Dlamini, R. L. S. Langa, A. I. Okoh, O. A. Aiyegoro

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The indiscriminate usage of antibiotics in swine production have consequential outcomes; such as development of bacterial resistance to prophylactic antibiotics and possibility of antibiotic residues in animal products. The use of probiotics appears to be the most effective procedure with positive metabolic nutritional implications. The aim of this study was to investigate the efficacy of probiotic bacteria (Lactobacillus reuteri ZJ625, Lactobacillus reuteri VB4, Lactobacillus salivarius ZJ614 and Streptococcus salivarius NBRC13956) administered as direct-fed microorganisms in weaned piglets. 45 weaned piglets blocked by weight were dived into 5 treatments groups: diet with antibiotic, diet with no-antibiotic and no probiotic, and diet with probiotic and diet with combination of probiotics. Piglets performance was monitored during the trials. Faecal and Ileum samples were collected for microbial count analysis. Blood samples were collected from pigs at the end of the trial, for analysis of haematological, biochemical and IgG stimulation. The data was analysed by Split-Plot ANOVA using SAS statistically software (SAS 9.3) (2003). The difference was observed between treatments for daily weight and feed conversion ratio. No difference was observed in analysis of faecal samples in regards with bacterial counts, difference was observed in ileums samples with enteric bacteria colony forming unit being lower in P2 treatment group as compared with lactic acid and total bacteria. With exception of globulin and albumin, biochemistry blood parameters were not affected, likewise for haematology, only basophils and segmented neutrophils were differed by having higher concentration in NC treatment group as compared with other treatment groups. Moreover, in IgG stimulation analysis, difference was also observed, with P2 treatment group having high concentration of IgG in P2 treatment group as compared to other groups. The results of this study suggest that probiotics have a beneficial effect on growth performances, blood parameters and IgG stimulation of pigs, most effective when they are administered in synergy form. This means that it is most likely that these probiotics will offer a significant benefit in pig farming by reducing risk of morbidity and mortality and produce quality meat that is more affordable to poorer communities, and thereby enhance South African pig industry’s economy. In addition, these results indicate that there is still more research need to be done on probiotics in regards with, i.e. dosage, shelf life and mechanism of action.

Keywords: antibiotics, biochemistry, haematology, IgG-stimulation, microbial count, probiotics

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Authors: Agnieszka E. Laudy, Karolina Stępien, Sergiusz Lulinski, Krzysztof Durka, Stefan Tyski

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1,3-dihydro-1-hydroxy-2,1-benzoxaborole and its derivatives are a particularly interesting group of synthetic agents and were successfully employed in supramolecular chemistry medicine. The first important compounds, 5-fluoro-1,3-dihydro-1-hydroxy-2,1-benzoxaborole and 5-chloro-1,3-dihydro-1-hydroxy-2,1-benzoxaborole were identified as potent antifungal agents. In contrast, (S)-3-(aminomethyl)-7-(3-hydroxypropoxy)-1-hydroxy-1,3-dihydro-2,1-benzoxaborole hydrochloride is in the second phase of clinical trials as a drug for the treatment of Gram-negative bacterial infections of the Enterobacteriaceae family and Pseudomonas aeruginosa. Equally important and difficult task is to search for compounds active against Gram-negative bacilli, which have multi-drug-resistance efflux pumps actively removing many of the antibiotics from bacterial cells. We have examined whether halogen-substituted benzoxaborole-based derivatives and their analogues possess antibacterial activity and are substrates for multi-drug-resistance efflux pumps. The antibacterial activity of 1,3-dihydro-3-hydroxy-1,1-dimethyl-1,2,3-benzosiloxaborole and 10 halogen-substituted its derivatives, as well as 1,2-phenylenediboronic acid and 3 synthesised fluoro-substituted its analogs, were evaluated. The activity against the reference strains of Gram-positive (n=5) and Gram-negative bacteria (n=10) was screened by the disc-diffusion test (0.4 mg of tested compounds was applied onto paper disc). The minimal inhibitory concentration values and the minimal bactericidal concentration values were estimated according to The Clinical and Laboratory Standards Institute and The European Committee on Antimicrobial Susceptibility Testing recommendations. During the minimal inhibitory concentration values determination with or without phenylalanine-arginine beta-naphthylamide (50 mg/L) efflux pump inhibitor, the concentrations of tested compounds ranged 0.39-400 mg/L in the broth medium supplemented with 1 mM magnesium sulfate. Generally, the studied benzosiloxaboroles and benzoxadiboroles showed a higher activity against Gram-positive cocci than against Gram-negative rods. Moreover, benzosiloxaboroles have the higher activity than benzoxadiboroles compounds. In this study, we demonstrated that substitution (mono-, di- or tetra-) of 1,3-dihydro-3-hydroxy-1,1-dimethyl-1,2,3-benzosiloxaborole with halogen groups resulted in an increase in antimicrobial activity as compared to the parent substance. Interestingly, the 6,7-dichloro-substituted parent substance was found to be the most potent against Gram-positive cocci: Staphylococcus sp. (minimal inhibitory concentration 6.25 mg/L) and Enterococcus sp. (minimal inhibitory concentration 25 mg/L). On the other hand, mono- and dichloro-substituted compounds were the most actively removed by efflux pumps present in Gram-negative bacteria mainly from Enterobacteriaceae family. In the presence of efflux pump inhibitor the minimal inhibitory concentration values of chloro-substituted benzosiloxaboroles decreased from 400 mg/L to 3.12 mg/L. Of note, the highest increase in bacterial susceptibility to tested compounds in the presence of phenylalanine-arginine beta-naphthylamide was observed for 6-chloro-, 6,7-dichloro- and 6,7-difluoro-substituted benzosiloxaboroles. In the case of Escherichia coli, Enterobacter cloacae and P. aeruginosa strains at least a 32-fold decrease in the minimal inhibitory concentration values of these agents were observed. These data demonstrate structure-activity relationships of the tested derivatives and highlight the need for further search for benzoxaboroles and related compounds with significant antimicrobial properties. Moreover, the influence of phenylalanine-arginine beta-naphthylamide on the susceptibility of Gram-negative rods to studied benzosiloxaboroles indicate that some tested agents are substrates for efflux pumps in Gram-negative rods.

Keywords: antibacterial activity, benzosiloxaboroles, efflux pumps, phenylalanine-arginine beta-naphthylamide

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Authors: Reena Aggarwal

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The use of textile chemicals in the various pre and post-textile manufacturing processes has made the textile industry conscious of its negative contribution to environmental pollution. Popular environmentally friendly fibers such as recycled polyester and organic cotton have been now increasingly used by fabrics and apparel manufacturers. However, after these textiles or the finished apparel are manufactured, they have to be dyed in the same chemical dyes that are harmful and toxic to the environment. Dyeing is a major area of concern for the environment as well as for people who have chemical sensitivities as it may cause nausea, breathing difficulties, seizures, etc. Ayurvastra or herbal medical textiles are one step ahead of the organic lifestyle, which supports the core concept of holistic well-being and also eliminates the impact of harmful chemicals and pesticides. There is a wide range of herbs that can be used not only for dyeing but also for providing medicinal properties to the textiles like antibacterial, antifungal, antiseptic, antidepressant and for treating insomnia, skin diseases, etc. The concept of herbal dyeing of fabric is to manifest herbal essence in every aspect of clothing, i.e., from production to end-use, additionally to eliminate the impact of harmful chemical dyes and chemicals which are known to result in problems like skin rashes, headache, trouble concentrating, nausea, diarrhea, fatigue, muscle and joint pain, dizziness, difficulty breathing, irregular heartbeat and seizures. Herbal dyeing or finishing on textiles will give an extra edge to the textiles as it adds an extra function to the fabric. The herbal extracts can be applied to the textiles by a simple process like the pad dry cure method and mainly acts on the human body through the skin for aiding in the treatment of disease or managing the medical condition through its herbal properties. This paper, therefore, delves into producing Ayurvastra, which is a perfect amalgamation of cloth and wellness. The aim of the paper is to design and create herbal disposable and non-disposable medical textile products acting mainly topically (through the skin) for providing medicinal properties/managing medical conditions. Keeping that in mind, a range of antifungal socks and antibacterial napkins treated with turmeric and aloe vera were developed, which are recommended for the treatment of fungal and bacterial infections, respectively. Both Herbal Antifungal socks and Antibacterial napkins have proved to be efficient enough in managing and treating fungal and bacterial infections of the skin, respectively.

Keywords: ayurvastra, ayurveda, herbal, pandemic, sustainable

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Authors: W. Y. Zhu, X. L. Yan, Y. Zhou

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Sulfonamides (SAs) are extensively used antibiotics; photocatalysis is an effective, way to remove the SAs from water driven by solar energy. Here we used WO3 nanoplates and their Ag heterogeneous as photocatalysts to investigate their photodegradation efficiency against sulfanilamide (SAM) which is the precursor of SAs. Results showed that WO3/Ag composites performed much better than pure WO3 where the highest removal rate was 96.2% can be achieved under visible light irradiation. Ag as excellent antibacterial agent also endows certain antibacterial efficiency to WO3, and 100% removal efficiency could be achieved in 2 h under visible light irradiation for all WO3/Ag composites. Generally, WO3/Ag composites are very effective photocatalysts with potentials in practical applications which mainly use cheap, clean and green solar energy as energy source.

Keywords: antibacterial, photocatalysis, semiconductor, sulfanilamide

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Authors: Irina E. Sukovataya, Oleg S. Sutormin, Valentina A. Kratasyuk

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Effect of viscosity of media on kinetic parameters of the coupled enzyme system NADH:FMN-oxidoreductase–luciferase was investigated with addition of organic solvents (glycerol and sucrose), because bioluminescent enzyme systems based on bacterial luciferases offer a unique and general tool for analysis of the many analytes and enzymes in the environment, research, and clinical laboratories and other fields. The possibility of stabilization and increase of activity of the coupled enzyme system NADH:FMN-oxidoreductase–luciferase activity in vicious aqueous-organic mixtures have been shown.

Keywords: coupled enzyme system of bioluminescence bacteria NAD(P)H:FMN-oxidoreductase–luciferase, glycerol, stabilization of enzymes, sucrose

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Authors: Yetti Marlida, Harnentis, Yuliaty Shafan Nur

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Background: Tempoyak is a dish derived from fermented durian fruit. Tempoyak is a food consumed as a side dish when eating rice. Besides being eaten with rice, tempoyak can also be eaten directly. But this is rarely done because many cannot stand the sour taste and aroma of the tempoyak itself. In addition, tempoyak can also be used as a seasoning. The taste of tempoyak is acidic, this occurs because of the fermentation process in durian fruit meat which is the raw material. Tempoyak is already very well known in Indonesia, especially in Padang, Bengkulu, Palembang, Lampung, and Kalimantan. Besides that, this food is also famous in Malaysia. The purpose of this research is to improvement production of γ-aminobutyric acid (GABA) by Lactobacillus plantarum isolated from indigenous fermented durian (tempoyak). Selected Lactic Acid Bacteria (LAB) previously isolated from indigenous fermented durian (tempoyak) that have ability to produce γ-aminobutyric acid (GABA). The study was started with identification of selected LAB by 16 S RNA, followed optimation of GABA production by culture condition using different initial pH, temperature, glutamate concentration, incubation time, carbon and nitrogen sources. Results: The result from indentification used polymerase chain reaction of 16S rRNA gene sequences and phylogenetic analysis was Lactobacillus plantarum (coded as Y3) with a sequenced length of 1400bp. The improvement of Gaba production was found highest at pH: 6.0; temperature: 30 °C; glutamate concentration: 0.4%; incubation time: 60 h; glucose and yeast extract as carbon and nitrogen sources. Conclusions: GABA can be produced with the optimum condition fermentation were 66.06 mM.

Keywords: lactic acid bacteria, γ-amino butyric acid, indigenous fermented durian, PCR

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Authors: Assel E. Molzhigitova, Amankeldi K. Sadanov, Elvira T. Ismailova, Kulyash A. Iskandarova, Olga N. Shemshura, Ainur I. Seitbattalova

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Fire blight is a very harmful for commercial apple and pear production quarantine bacterial disease. To date, several different methods have been proposed for disease control, including the use of copperbased preparations and antibiotics, which are not always reliable or effective. The use of bacteria as biocontrol agents is one of the most promising and eco-friendly alternative methods. Bacteria with protective activity against the causal agent of fire blight are often present among the epiphytic microorganisms of the phyllosphere of host plants. Therefore, the main objective of our study was screening of local epiphytic bacteria as possible antagonists against Erwinia amylovora, the causal agent of fire blight. Samples of infected organs of apple and pear trees (shoots, leaves, fruits) were collected from the industrial horticulture areas in various agro-ecological zones of Kazakhstan. Epiphytic microorganisms were isolated by standard and modified methods on specific nutrient media. The primary screening of selected microorganisms under laboratory conditions to determine the ability to suppress the growth of Erwinia amylovora was performed by agar-diffusion-test. Among 142 bacteria isolated from the fire blight host plants, 5 isolates, belonging to the genera Bacillus, Lactobacillus, Pseudomonas, Paenibacillus and Pantoea showed higher antagonistic activity against the pathogen. The diameters of inhibition zone have been depended on the species and ranged from 10 mm to 48 mm. The maximum diameter of inhibition zone (48 mm) was exhibited by B. amyloliquefaciens. Less inhibitory effect was showed by Pantoea agglomerans PA1 (19 mm). The study of inhibitory effect of Lactobacillus species against E. amylovora showed that among 7 isolates tested only one (Lactobacillus plantarum 17M) demonstrated inhibitory zone (30 mm). In summary, this study was devoted to detect the beneficial epiphytic bacteria from plants organs of pear and apple trees due to fire blight control in Kazakhstan. Results obtained from the in vitro experiments showed that the most efficient bacterial isolates are Lactobacillus plantarum 17M, Bacillus amyloliquefaciens MB40, and Pantoea agglomerans PA1. These antagonists are suitable for development as biocontrol agents for fire blight control. Their efficacies will be evaluated additionally, in biological tests under in vitro and field conditions during our further study.

Keywords: antagonists, epiphytic bacteria, Erwinia amylovora, fire blight

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Authors: Gail Louw, Helena Boshoff, Taeksun Song, Clifton Barry

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Drug resistance in Mycobacterium tuberculosis is primarily attributed to mutations in target genes. These mutations incur a fitness cost and result in bacterial generations that are less fit, which subsequently acquire compensatory mutations to restore fitness. We hypothesize that mutations in specific drug target genes influence bacterial metabolism and cellular function, which affects its ability to develop subsequent resistance to additional agents. We aim to determine whether the sequential acquisition of drug resistance and specific mutations in a well-defined clinical M. tuberculosis strain promotes or limits the development of additional resistance. In vitro mutants resistant to pretomanid, linezolid, moxifloxacin, rifampicin and kanamycin were generated from a pan-susceptible clinical strain from the Beijing lineage. The resistant phenotypes to the anti-TB agents were confirmed by the broth microdilution assay and genetic mutations were identified by targeted gene sequencing. Growth of mono-resistant mutants was done in enriched medium for 14 days to assess in vitro fitness. Double resistant mutants were generated against anti-TB drug combinations at concentrations 5x and 10x the minimum inhibitory concentration. Subsequently, mutation frequencies for these anti-TB drugs in the different mono-resistant backgrounds were determined. The initial level of resistance and the mutation frequencies observed for the mono-resistant mutants were comparable to those previously reported. Targeted gene sequencing revealed the presence of known and clinically relevant mutations in the mutants resistant to linezolid, rifampicin, kanamycin and moxifloxacin. Significant growth defects were observed for mutants grown under in vitro conditions compared to the sensitive progenitor. Mutation frequencies determination in the mono-resistant mutants revealed a significant increase in mutation frequency against rifampicin and kanamycin, but a significant decrease in mutation frequency against linezolid and sutezolid. This suggests that these mono-resistant mutants are more prone to develop resistance to rifampicin and kanamycin, but less prone to develop resistance against linezolid and sutezolid. Even though kanamycin and linezolid both inhibit protein synthesis, these compounds target different subunits of the ribosome, thereby leading to different outcomes in terms of fitness in the mutants with impaired cellular function. These observations showed that oxazolidinone treatment is instrumental in limiting the development of multi-drug resistance in M. tuberculosis in vitro.

Keywords: oxazolidinones, mutations, resistance, tuberculosis

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Authors: Kalaumari Mayoral-Peña, Ana I. Montejano-Montelongo, Josué Reyes-Muñoz, Gonzalo A. Ortiz-Mancilla, Mayrin Rodríguez-Cruz, Víctor Hernández-Villalobos, Jesús A. Guzmán-López, Santiago García-Jacobo, Iván Licona-Vázquez, Grisel Fierros-Romero, Rosario Flores-Vallejo

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The group B-lactamic antibiotics include some of the most frequently used small drug molecules against bacterial infections. Nevertheless, an alarming decrease in their efficacy has been reported due to the emergence of antibiotic-resistant bacteria. Infections caused by bacteria expressing extended Spectrum B-lactamases (ESBLs) are difficult to treat and account for higher morbidity and mortality rates, delayed recovery, and high economic burden. According to the Global Report on Antimicrobial Resistance Surveillance, it is estimated that mortality due to resistant bacteria will ascend to 10 million cases per year worldwide. These facts highlight the importance of developing low-cost and readily accessible detection methods of drug-resistant ESBLs bacteria to prevent their spread and promote accurate and fast diagnosis. Bacterial detection is commonly done using molecular diagnostic techniques, where PCR stands out for its high performance. However, this technique requires specialized equipment not available everywhere, is time-consuming, and has a high cost. Loop-Mediated Isothermal Amplification (LAMP) is an alternative technique that works at a constant temperature, significantly decreasing the equipment cost. It yields double-stranded DNA of several lengths with repetitions of the target DNA sequence as a product. Although positive and negative results from LAMP can be discriminated by colorimetry, fluorescence, and turbidity, there is still a large room for improvement in the point-of-care implementation. DNA origami is a technique that allows the formation of 3D nanometric structures by folding a large single-stranded DNA (scaffold) into a determined shape with the help of short DNA sequences (staples), which hybridize with the scaffold. This research aimed to generate DNA origami structures using LAMP products as scaffolds to improve the sensitivity to detect ESBLs in point-of-care diagnosis. For this study, the coding sequence of the CTM-X-15 ESBL of E. coli was used to generate the LAMP products. The set of LAMP primers were designed using PrimerExplorerV5. As a result, a target sequence of 200 nucleotides from CTM-X-15 ESBL was obtained. Afterward, eight different DNA origami structures were designed using the target sequence in the SDCadnano and analyzed with CanDo to evaluate the stability of the 3D structures. The designs were constructed minimizing the total number of staples to reduce costs and complexity for point-of-care applications. After analyzing the DNA origami designs, two structures were selected. The first one was a zig-zag flat structure, while the second one was a wall-like shape. Given the sequence repetitions in the scaffold sequence, both were able to be assembled with only 6 different staples each one, ranging between 18 to 80 nucleotides. Simulations of both structures were performed using scaffolds of different sizes yielding stable structures in all the cases. The generation of the LAMP products were tested by colorimetry and electrophoresis. The formation of the DNA structures was analyzed using electrophoresis and colorimetry. The modeling of novel detection methods through bioinformatics tools allows reliable control and prediction of results. To our knowledge, this is the first study that uses LAMP products and DNA-origami in combination to delect ESBL-producing bacterial strains, which represent a promising methodology for diagnosis in the point-of-care.

Keywords: beta-lactamases, antibiotic resistance, DNA origami, isothermal amplification, LAMP technique, molecular diagnosis

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Authors: Eduardo Lanzagorta Garcia, Chaitra Venkatesh, Romina Pezzoli, Laura Gabriela Rodriguez Barroso, Declan Devine, Margaret E. Brennan Fournet

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New antimicrobial interventions are urgently required to combat rising global health and medical infection challenges. Here, an innovative antimicrobial technology, providing price competitive alternatives to antibiotics and readily integratable with currently technological systems is presented. Two cutting edge antimicrobial materials, antimicrobial peptides (AMPs) and uncompromised sustained Ag+ action from triangular silver nanoplates (TSNPs) reservoirs, are merged for versatile effective antimicrobial action where current approaches fail. Antimicrobial peptides (AMPs) exist widely in nature and have recently been demonstrated for broad spectrum of activity against bacteria, viruses, and fungi. TSNP’s are highly discrete, homogenous and readily functionisable Ag+ nanoreseviors that have a proven amenability for operation within in a wide range of bio-based settings. In a design for advanced antimicrobial sustainable plastics, antimicrobial TSNPs are formulated for processing within biodegradable biopolymers. Histone H5 AMP was selected for its reported strong antimicrobial action and functionalized with the TSNP (AMP-TSNP) in a similar fashion to previously reported TSNP biofunctionalisation methods. A synergy between the propensity of biopolymers for degradation and Ag+ release combined with AMP activity provides a novel mechanism for the sustained antimicrobial action of biopolymeric thin films. Nanoplates are transferred from aqueous phase to an organic solvent in order to facilitate integration within hydrophobic polymers. Extrusion is used in combination with calendering rolls to create thin polymerc film where the nanoplates are embedded onto the surface. The resultant antibacterial functional films are suitable to be adapted for food packing and biomedical applications. TSNP synthesis were synthesized by adapting a previously reported seed mediated approach. TSNP synthesis was scaled up for litre scale batch production and subsequently concentrated to 43 ppm using thermally controlled H2O removal. Nanoplates were transferred from aqueous phase to an organic solvent in order to facilitate integration within hydrophobic polymers. This was acomplised by functionalizing the TSNP with thiol terminated polyethylene glycol and using centrifugal force to transfer them to chloroform. Polycaprolactone (PCL) and Polylactic acid (PLA) were individually processed through extrusion, TSNP and AMP-TSNP solutions were sprayed onto the polymer immediately after exiting the dye. Calendering rolls were used to disperse and incorporate TSNP and TSNP-AMP onto the surface of the extruded films. Observation of the characteristic blue colour confirms the integrity of the TSNP within the films. Antimicrobial tests were performed by incubating Gram + and Gram – strains with treated and non-treated films, to evaluate if bacterial growth was reduced due to the presence of the TSNP. The resulting films successfully incorporated TSNP and AMP-TSNP. Reduced bacterial growth was observed for both Gram + and Gram – strains for both TSNP and AMP-TSNP compared with untreated films indicating antimicrobial action. The largest growth reduction was observed for AMP-TSNP treated films demonstrating the additional antimicrobial activity due to the presence of the AMPs. The potential of this technology to impede bacterial activity in food industry and medical surfaces will forge new confidence in the battle against antibiotic resistant bacteria, serving to greatly inhibit infections and facilitate patient recovery.

Keywords: antimicrobial, biodegradable, peptide, polymer, nanoparticle

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Authors: Nuha Mansour Alhazmi

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Replacement of petro-based plastics by a biodegradable plastic are vastly growing process. Poly-β-hydroxybutyrate (PHB) is a biodegradable biopolymer, synthesized by some bacterial genera. The objective of the current study is to explore the ability of some fungi to biodegrade PHB. The degradation of (PHB) was detected in Petri dish by the formation of a clear zone around the fungal colonies due to the production of depolymerase enzyme which has an interesting role in the PHB degradation process. Among 10 tested fungi, the most active PHB biodegraded fungi were identified as Trichoderma asperellum using morphological and molecular characters. The highest PHB degradation was at 25°C, pH 7.5 after 7 days of incubation for the tested fungi. Finally, the depolymerase enzyme was isolated, purified using column chromatography and characterized. In conclusion, PHB can be biodegraded in solid and liquid medium using depolymerase enzyme from T. asperellum.

Keywords: degradation, depolymerase enzyme, PHB, Trichoderma asperellum

Procedia PDF Downloads 167

Authors: B.L. España-Sánchez, E. Luna-Hernández, R.A. Mauricio-Sánchez, M.E. Cruz-Soto, F. Padilla-Vaca, R. Muñoz, L. Granados-López, L.R. Ovalle-Flores, J.L. Menchaca-Arredondo, G. Luna-Bárcenas

Abstract:

Treatment of burn injured has been considered an important clinical problem due to the fluid control and the presence of microorganisms during the healing process. Conventional treatment includes antiseptic techniques, topical medication and surgical removal of damaged skin, to avoid bacterial growth. In order to accelerate this process, different alternatives for tissue regeneration have been explored, including artificial skin, polymers, hydrogels and hybrid materials. Some requirements consider a nonreactive organic polymer with high biocompatibility and skin adherence, avoiding bacterial infections. Chitin-derivative biopolymer such as chitosan (CS) has been used in skin regeneration following third-degree burns. The biological interest of CS is associated with the improvement of tissue cell stimulation, biocompatibility and antibacterial properties. In particular, antimicrobial properties of CS can be significantly increased when is blended with nanostructured materials. Silver-based nanocomposites have gained attention in medicine due to their high antibacterial properties against pathogens, related to their high surface area/volume ratio at nanomolar concentrations. Silver nanocomposites can be blended or synthesized with chitin-derivative biopolymers in order to obtain a biodegradable/antimicrobial hybrid with improved physic-mechanical properties. In this study, nanocomposites based on chitosan/silver nanoparticles (CS/nAg) were synthesized by the in situ chemical reduction method, improving their antibacterial properties against pathogenic bacteria and enhancing the healing process in thermal burn injuries produced in an animal model. CS/nAg was prepared in solution by the chemical reduction method, using AgNO₃ as precursor. CS was dissolved in acetic acid and mixed with different molar concentrations of AgNO₃: 0.01, 0.025, 0.05 and 0.1 M. Solutions were stirred at 95°C during 20 hours, in order to promote the nAg formation. CS/nAg solutions were placed in Petri dishes and dried, to obtain films. Structural analyses confirm the synthesis of silver nanoparticles (nAg) by means of UV-Vis and TEM, with an average size of 7.5 nm and spherical morphology. FTIR analyses showed the complex formation by the interaction of hydroxyl and amine groups with metallic nanoparticles, and surface chemical analysis (XPS) shows low concentration of Ag⁰/Ag⁺ species. Topography surface analyses by means of AFM shown that hydrated CS form a mesh with an average diameter of 10 µm. Antibacterial activity against S. aureus and P. aeruginosa was improved in all evaluated conditions, such as nAg loading and interaction time. CS/nAg nanocomposites films did not show Ag⁰/Ag⁺ release in saline buffer and rat serum after exposition during 7 days. Healing process was significantly enhanced by the presence of CS/nAg nanocomposites, inducing the production of myofibloblasts, collagen remodelation, blood vessels neoformation and epidermis regeneration after 7 days of injury treatment, by means of histological and immunohistochemistry assays. The present work suggests that hydrated CS/nAg nanocomposites can be formed a mesh, improving the bacterial penetration and the contact with embedded nAg, producing complete growth inhibition after 1.5 hours. Furthermore, CS/nAg nanocomposites improve the cell tissue regeneration in thermal burn injuries induced in rats. Synthesis of antibacterial, non-toxic, and biocompatible nanocomposites can be an important issue in tissue engineering and health care applications.

Keywords: antibacterial, chitosan, healing process, nanocomposites, silver

Procedia PDF Downloads 276

Authors: C. C. Castro, T. Senechal, D. Lahem, A. L. Hantson

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Formaldehyde is one of the most representative volatile organic compounds present in the indoor air of residential units and workplaces. Increased attention has been given to this toxic compound because of its carcinogenic effect in health. Biological or enzymatic transformation is being explored to degrade this pollutant. Pseudomonas putida is a bacteria able to synthesize formaldehyde dehydrogenase, an enzyme known to use formaldehyde as a substrate and transform it into less toxic compounds. The immobilization of bacterial cells in the surface of different supports through spraying or dip-coating is herein proposed. The determination of the enzymatic activity on the coated surfaces was performed as well as the study of its effect on formaldehyde degradation in an isolated chamber. Results show that the incorporation of microbial cells able to synthesize depolluting enzymes can be an innovative, low-cost, effective and environmentally friendly solution for indoor air depollution.

Keywords: cells encapsulation, formaldehyde, formaldehyde dehydrogenase, indoor air depollution

Procedia PDF Downloads 160

Authors: Tanmaya Pradhan, K. Midhun, M. Joy Thomas

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Increasing the shelf life and improving the quality are important objectives for the success of packaged liquid food industry. One of the methods by which this can be achieved is by deactivating the micro-organisms present in the liquid food through pasteurization. Pasteurization is done by heating, but some serious disadvantages such as the reduction in food quality, flavour, taste, colour, etc. were observed because of heat treatment, which leads to the development of newer methods instead of pasteurization such as treatment using UV radiation, high pressure, nuclear irradiation, pulsed electric field, etc. In recent years the use of the pulsed electric field (PEF) for inactivation of the microbial content in the food is gaining popularity. PEF uses a very high electric field for a short time for the inactivation of microorganisms, for which we require a high voltage pulsed power source. Pulsed power sources used for PEF treatments are usually in the range of 5kV to 50kV. Different pulse shapes are used, such as exponentially decaying and square wave pulses. Exponentially decaying pulses are generated by high power switches with only turn-on capacity and, therefore, discharge the total energy stored in the capacitor bank. These pulses have a sudden onset and, therefore, a high rate of rising but have a very slow decay, which yields extra heat, which is ineffective in microbial inactivation. Square pulses can be produced by an incomplete discharge of a capacitor with the help of a switch having both on/off control or by using a pulse forming network. In this work, a pulsed power-based system is designed with the help of high voltage capacitors and solid-state switches (IGBT) for the inactivation of pathogenic micro-organism in liquid food such as fruit juices. The high voltage generator is based on the Marx generator topology, which can produce variable amplitude, frequency, and pulse width according to the requirements. Liquid food is treated in a chamber where pulsed electric field is produced between stainless steel electrodes using the pulsed output voltage of the supply. Preliminary bacterial inactivation tests were performed by subjecting orange juice inoculated with Escherichia Coli bacteria. With the help of the developed pulsed power source and the chamber, the inoculated orange has been PEF treated. The voltage was varied to get a peak electric field up to 15kV/cm. For a total treatment time of 200µs, a 30% reduction in the bacterial count has been observed. The detailed results and analysis will be presented in the final paper.

Keywords: Escherichia coli bacteria, high voltage generator, microbial inactivation, pulsed electric field, pulsed forming line, solid-state switch

Procedia PDF Downloads 167

Authors: Lina Wu

Abstract:

The excessive discharge of nitrogen in sewage greatly intensifies the eutrophication of water bodies and threatens water quality. Nitrogen pollution control has become a global concern. The concentration of nitrogen in water is reduced by converting ammonia nitrogen, nitrate nitrogen and nitrite nitrogen into nitrogen-containing gas through biological treatment, physicochemical treatment and oxidation technology. However, some wastewater containing high ammonia nitrogen including landfill leachate, is difficult to be treated by traditional nitrification and denitrification because of its high COD content. The core process of denitrification is that denitrifying bacteria convert nitrous acid produced by nitrification into nitrite under anaerobic conditions. Still, its low-carbon nitrogen does not meet the conditions for denitrification. Many studies have shown that the natural autotrophic anammox bacteria can combine nitrous and ammonia nitrogen without a carbon source through functional genes to achieve total nitrogen removal, which is very suitable for removing nitrogen from leachate. In addition, the process also saves a lot of aeration energy consumption than the traditional nitrogen removal process. Therefore, anammox plays an important role in nitrogen conversion and energy saving. The short-range nitrification and denitrification coupled with anaerobic ammoX ensures total nitrogen removal. It improves the removal efficiency, meeting the needs of society for an ecologically friendly and cost-effective nutrient removal treatment technology. In recent years, research has found that the symbiotic system has more water treatment advantages because this process not only helps to improve the efficiency of wastewater treatment but also allows carbon dioxide reduction and resource recovery. Microalgae use carbon dioxide dissolved in water or released through bacterial respiration to produce oxygen for bacteria through photosynthesis under light, and bacteria, in turn, provide metabolites and inorganic carbon sources for the growth of microalgae, which may lead the algal bacteria symbiotic system save most or all of the aeration energy consumption. It has become a trend to make microalgae and light-avoiding anammox bacteria play synergistic roles by adjusting the light-to-dark ratio. Microalgae in the outer layer of light particles block most of the light and provide cofactors and amino acids to promote nitrogen removal. In particular, myxoccota MYX1 can degrade extracellular proteins produced by microalgae, providing amino acids for the entire bacterial community, which helps anammox bacteria save metabolic energy and adapt to light. As a result, initiating and maintaining the process of combining dominant algae and anaerobic denitrifying bacterial communities has great potential in treating landfill leachate. Chlorella has a brilliant removal effect and can withstand extreme environments in terms of high ammonia nitrogen, high salt and low temperature. It is urgent to study whether the algal mud mixture rich in denitrifying bacteria and chlorella can greatly improve the efficiency of landfill leachate treatment under an anaerobic environment where photosynthesis is stopped. The optimal dilution concentration of simulated landfill leachate can be found by determining the treatment effect of the same batch of bacteria and algae mixtures under different initial ammonia nitrogen concentrations and making a comparison. High-throughput sequencing technology was used to analyze the changes in microbial diversity, related functional genera and functional genes under optimal conditions, providing a theoretical and practical basis for the engineering application of novel bacteria-algae symbiosis system in biogas slurry treatment and resource utilization.

Keywords: nutrient removal and recovery, leachate, anammox, Partial nitrification, Algae-bacteria interaction

Procedia PDF Downloads 28

Authors: Sulaiman B. Ali Alharbi, Bassam H. Mashat, Naif Abdullah Al-Harbi, Milton Wainwright, Abeer S. Aloufi, Sulamain Alnaimat

Abstract:

Bismuth salicylate was found to inhibit the growth of a range of bacteria and yeast, Candida albican. In general the growth of bacteria did not result in the increase in bismuth solubilisation, in contrast, bismuth solubilisation increased following the growth of C. albicans. A significant increase in the biomass (dry weight) of Aspergillus niger and Aspergillus oryzae occurred in vitro when these fungi were grown in the presence of bismuth salicylate. Biomass increase occurred over a range of bismuth compound additions, which in the case of A. oryzae was associated with the increase in the solubilisation of the insoluble bismuth compounds.

Keywords: bacterial inhibition, fungal growth stimulation, medical uses of bismuth, yeast inhibition

Procedia PDF Downloads 328

Authors: Yixin Yan, Miao Yan, Irini Angelidaki, Ioannis Fotidis

Abstract:

Ammonia, which derives from the degradation of urea and protein-substrates, is the major toxicant of the commercial anaerobic digestion reactors causing loses of up to 1/3 of their practical biogas production, which reflects directly on the overall revenue of the plants. The current experimental work is aiming to alleviate the ammonia inhibition in anaerobic digestion (AD) process by developing an innovative bioaugmentation method of ammonia tolerant methanogenic consortia. The ammonia tolerant consortia were cultured in batch reactors and immobilized together with biochar in agar (customized inocula). Three continuous stirred-tank reactors (CSTR), fed with the organic fraction of municipal solid waste at a hydraulic retention time of 15 days and operated at thermophilic (55°C) conditions were assessed. After an ammonia shock of 4 g NH4+-N L-1, the customized inocula were bioaugmented into the CSTR reactors to alleviate ammonia toxicity effect on AD process. Recovery rate of methane production and methanogenic activity will be assessed to evaluate the bioaugmentation performance, while 16s rRNA gene sequence will be used to reveal the difference of microbial community changes through bioaugmentation. At the microbial level, the microbial community structures of the four reactors will be analysed to find the mechanism of bioaugmentation. Changes in hydrogen formation potential will be used to predict direct interspecies electron transfer (DIET) between ammonia tolerant methanogens and syntrophic bacteria. This experimental work is expected to create bioaugmentation inocula that will be easy to obtain, transport, handled and bioaugment in AD reactors to efficiently alleviate the ammonia toxicity, without alternating any of the other operational parameters including the ammonia-rich feedstocks.

Keywords: artisanal fishing waste, acidogenesis, volatile fatty acids, pH, inoculum/substrate ratio

Procedia PDF Downloads 111

Authors: Leila Fozouni, Anahita Mazandarani

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Background: Maternal infections are the most common cause of infections in infants, and the level of infection and its severity highly depends on the degree of colonization of the bacteria in the mother; so, the occurrence of aggressive diseases is not unpredictable in mothers with very high colonization. Group B Streptococcus is part of the normal flora of the gastrointestinal and genital tracts in women and is the leading cause of septicemia and meningitis in newborns. Today Zinc oxide nanoparticle is regarded as one of the most commonly used and safest nanoparticles for defeating Gram-positive and Gram-negative bacteria. This study aims to determine the antibacterial effects of Zinc oxide on the growth of drug-resistant group B Streptococcus strains isolated from pregnant women. Materials and Methods: This cross-sectional study was conducted on 150 pregnant women of 28–37 weeks admitted to seven hospitals and maternity wards in Golestan province, northeast of Iran. For bacterial identification, rectovaginal swabs were firstly inoculated to the Todd-Hewitt Broth and cultured in blood agar (containing 5% sheep blood). Then microbiologic and PCR methods were performed to detect group B Streptococci. Disk diffusion and broth microdilution tests were used to determine the bacterial susceptibility to antibiotics according to CLSI M100(2021) criteria. The antibacterial properties of Zinc oxide nanoparticles were evaluated using the agar well-diffusion method. Results: The prevalence of group B Streptococcus was 18% in pregnant women. Out of twenty-seven positive cultures, 62.96% were higher than thirty years old. Ninety percent and 45% of isolates were resistant to clindamycin and erythromycin, respectively, and susceptibility to cefazolin was 71%. In addition, susceptibility to ampicillin and penicillin were 74% and 55%, respectively. The results showed that 82% of erythromycin-resistant, 92% clindamycin-resistant, and 78% of cefazolin-resistant isolates were eliminated by zinc oxide nanoparticles at a concentration of 100 mg/L of the nanoparticle. Furthermore, ZnONPs could inhibit all drug-resistant isolates at a concentration of 200 mg/mL (MIC90 ≥ 200). Conclusion: Since the drug resistance of group B streptococci against various antibiotics is increasing, determining and investigating the drug-resistance pattern of this bacterium to different antibiotics in order to prevent arbitrary consumption of antibiotics by pregnant women and ultimately prevent Infant mortality seems necessary. Generally, ZnONPs showed a high antimicrobial effect, and it was revealed that the bactericide effect increases upon the increase in the concentration of the nanoparticle.

Keywords: group B beta-hemolytic streptococcus, pregnant women, zinc oxide nanoparticles, drug resistance

Procedia PDF Downloads 77

Authors: Marcin Debowski, Marcin Zielinski, Magdalena Zielinska, Paulina Rusanowska

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The problem with digestate management is one of the most important factors influencing on the development and operation of biogas plant. Turbidity and bacterial contamination negatively affect the growth of algae, which can limit the use of the effluent in the production of algae biomass on a large scale. These problems can be overcome by cultivating of algae species resistant to environmental factors, such as Chlorella sp., Scenedesmus sp., or reducing load of organic compounds to prevent bacterial contamination. The effluent requires dilution and/or purification. One of the methods of effluent treatment is the use of a membrane technology such as microfiltration (MF), ultrafiltration (UF), nanofiltration (NF) and reverse osmosis (RO), depending on the membrane pore size and the cut off point. Membranes are a physical barrier to solids and particles larger than the size of the pores. MF membranes have the largest pores and are used to remove turbidity, suspensions, bacteria and some viruses. UF membranes remove also color, odor and organic compounds with high molecular weight. In treatment of wastewater or other waste streams, MF and UF can provide a sufficient degree of purification. NF membranes are used to remove natural organic matter from waters, water disinfection products and sulfates. RO membranes are applied to remove monovalent ions such as Na⁺ or K⁺. The effluent was used in UF for medium to cultivation of two microalgae: Chlorella sp. and Phaeodactylum tricornutum. Growth rates of Chlorella sp. and P. tricornutum were similar: 0.216 d⁻¹ and 0.200 d⁻¹ (Chlorella sp.); 0.128 d⁻¹ and 0.126 d⁻¹ (P. tricornutum), on synthetic medium and permeate from UF, respectively. The final biomass composition was also similar, regardless of the medium. Removal of nitrogen was 92% and 71% by Chlorella sp. and P. tricornutum, respectively. The fermentation effluents after UF and dilution were also used for cultivation of algae Scenedesmus sp. that is resistant to environmental conditions. The authors recommended the development of biorafinery based on the production of algae for the biogas production. There are examples of using a multi-stage membrane system to purify the liquid fraction from digestate. After the initial UF, RO is used to remove ammonium nitrogen and COD. To obtain a permeate with a concentration of ammonium nitrogen allowing to discharge it into the environment, it was necessary to apply three-stage RO. The composition of the permeate after two-stage RO was: COD 50–60 mg/dm³, dry solids 0 mg/dm³, ammonium nitrogen 300–320 mg/dm³, total nitrogen 320–340 mg/dm³, total phosphorus 53 mg/dm³. However compostion of permeate after three-stage RO was: COD < 5 mg/dm³, dry solids 0 mg/dm³, ammonium nitrogen 0 mg/dm³, total nitrogen 3.5 mg/dm³, total phosphorus < 0,05 mg/dm³. Last stage of RO might be replaced by ion exchange process. The negative aspect of membrane filtration systems is the fact that the permeate is about 50% of the introduced volume, the remainder is the retentate. The management of a retentate might involve recirculation to a biogas plant.

Keywords: digestate, membrane filtration, microalgae cultivation, Chlorella sp.

Procedia PDF Downloads 342

Authors: B. T. Naveen Kumar, Anuj Tyagi, Niraj Kumar Singh, Visanu Boonyawiwat, A. H. Shanthanagouda, Orawan Boodde, K. M. Shankar, Prakash Patil, Shubhkaramjeet Kaur

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Early mortality syndrome (EMS)/Acute Hepatopancreatic Necrosis Disease (AHPND) has emerged as a major obstacle for the shrimp farming around the world. It is caused by a strain of Vibrio parahaemolyticus. The possible preventive and control measure is, early and rapid detection of the pathogen in the broodstock, post-larvae and monitoring the shrimp during the culture period. Polymerase chain reaction (PCR) based early detection methods are good, but they are costly, time taking and requires a sophisticated laboratory. The present study was conducted to develop a simple, sensitive and rapid diagnostic farmer level kit for the reliable detection of AHPND in shrimp. A panel of monoclonal antibodies (MAbs) were raised against the recombinant Pir B protein (rPirB). First, an immunodot was developed by using MAbs G3B8 and Mab G3H2 which showed specific reactivity to purified r-PirB protein with no cross-reactivity to other shrimp bacterial pathogens (AHPND free Vibrio parahaemolyticus (Indian strains), V. anguillarum, WSSV, Aeromonas hydrophila, and Aphanomyces invadans). Immunodot developed using Mab G3B8 is more sensitive than that with the Mab G3H2. However, immunodot takes almost 2.5 hours to complete with several hands-on steps. Therefore, the flow-through assay (FTA) was developed by using a plastic cassette containing the nitrocellulose membrane with absorbing pads below. The sample was dotted in the test zone on the nitrocellulose membrane followed by continuos addition of five solutions in the order of i) blocking buffer (BSA) ii) primary antibody (MAb) iii) washing Solution iv) secondary antibody and v) chromogen substrate (TMB) clear purple dots against a white background were considered as positive reactions. The FTA developed using MAbG3B8 is more sensitive than that with MAb G3H2. In FTA the two MAbs showed specific reactivity to purified r-PirB protein and not to other shrimp bacterial pathogens. The FTA is simple to farmer/field level, sensitive and rapid requiring only 8-10 min for completion. Tests can be developed to kits, which will be ideal for use in biosecurity, for the first line of screening (at the port or pond site) and during monitoring and surveillance programmes overall for the good management practices to reduce the risk of the disease.

Keywords: acute hepatopancreatic necrosis disease, AHPND, flow-through assay, FTA, farmer level, immunodot, pond site, shrimp

Procedia PDF Downloads 162

Authors: Maria Lucia G. Lourenco, Keylla H. N. P. Pereira, Vivane Y. Hibaru, Fabiana F. Souza, Joao C. P. Ferreira, Simone B. Chiacchio, Luiz H. A. Machado

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Neonatal sepsis is a systemic response to the acute generalized infection caused by one or more bacterial agents, representing the main infectious cause of neonatal mortality in dogs during the first three weeks of life. This study aims to describe the incidence of sepsis in neonate dogs, as well as the main clinical signs and mortality rates. The study included 735 neonates admitted to the Sao Paulo State University (UNESP) Veterinary Hospital, Botucatu, Sao Paulo, Brazil, between January 2018 and November 2019. Seven hundred thirty-five neonates, 14% (98/703) presented neonatal sepsis. The main sources of infection for the neonates were intrauterine (72.5%, 71/98), lactogenic (13.2%, 13/98), umbilical (5.1%, 5/98) and unidentified sources (9.2%, 9/98). The main non-specific clinical signs observed in the newborns were weakness, depression, impaired or absent reflexes, hypothermia, hypoglycemia, dehydration, reduced muscle tonus and diarrhea. The newborns also manifested clinical signs of severe infection, such as hyperemia in the abdominal and anal regions, omphalitis, hematuria, abdomen and extremities with purplish-blue coloration necrosing injuries in the pads, bradycardia, dyspnea, epistaxis, hypotension and evolution to septic shock. Infections acquired during intrauterine life led to the onset of the clinical signs at the time of birth, with fast evolution during the first hours of life. On the other hand, infections acquired via milk or umbilical cord presented clinical signs later. The total mortality rate was 5.4% (38/703) and the mortality rate among the neonates with sepsis was 38.7% (38/98). The early mortality rate (0 to 2 days) accounted for 86.9% (33/38) and the late mortality rate (3 to 30 days) for 13.1% (5/38) of the deaths among the newborns with sepsis. The main bacterial agents observed were Staphylococcus spp., Streptococcus spp., Proteus spp. Mannheimia spp. and Escherichia coli. Neonatal sepsis evolves quickly and may lead to high mortality in a litter. The prognosis is usually favorable if the diagnosis is reached early and the antibiotic therapy instituted as soon as possible, even before the results of blood cultures and antibiograms. The therapeutic recommendations should meet the special physiological conditions of a neonate in terms of metabolism and excretion of medication. Therefore, it is of utmost importance that the veterinarian is knowledgeable regarding neonatology to provide effective intervention and improve the survival rates of these patients.

Keywords: Neonatal infection , bacteria, puppies, newborn

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Authors: Yasir Arafat, Asma Shah, Hua Shao

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Background and Aims: Legume/cereal intercropping is used worldwide for enhancement in biomass and yield of cereal crops. However, because of intercropping, the belowground biological and chemical interactions and their effect on physiological parameters and yield of crops are limited. Methods: Wheat faba bean (WF) intercropping was designed to understand the underlying changes in the soil's chemical environment, soil microbial communities, and effect on growth and yield parameters. Experimental plots were established as having no root partition (NRP), semi-root partition (SRP), complete root partition (CRP), and their sole cropping (CK). Low molecular weight organic acids (LMWOAs) were determined by GC-MS, and high throughput sequencing of the 16S rRNA gene was carried out to screen microbial structure and composition in different root partitions of the WF intercropping system. Results: We show that intercropping induced a shift in the relative abundance of some genera of plant growth promoting rhizobacteria (PGPR) such as Allorhizobium, Neorhizobium, Pararhizobium, and Rhizobium species and resulted in better growth and yield performance of wheat. Moreover, as the plant's distance of wheat from faba beans decreased, the diversity of microbes increased, and a positive effect was observed on physiological traits and crop yield. Furthermore, an abundance and positive correlations of palmitic acid, arachidic acid, stearic acid, and 9-Octadecenoic with PGPR were recorded in the root zone of WF intercropping, which can play an important role in this facilitative mechanism of enhancing growth and yield of cereals. Conclusion: The two treatments clearly affected soil microbial and chemical composition, which can be reflected in growth and yield enhancement.

Keywords: intercropping, microbial community, LMWOAs, PGPR, soil chemical environment

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Authors: Georgia Saxami, Evangelia N. Kerezoudi, Evdokia K. Mitsou, Marigoula Vlassopoulou, Georgios Zervakis, Adamantini Kyriacou

Abstract:

Autism Spectrum Disorder (ASD) is a complex group of developmental disorders of the brain, characterized by social and communication dysfunctions, stereotypes and repetitive behaviors. The potential interaction between gut microbiota (GM) and autism has not been fully elucidated. Children with autism often suffer gastrointestinal dysfunctions, while alterations or dysbiosis of GM have also been observed. Treatment with dietary components has been postulated to regulate GM and improve gastrointestinal symptoms, but there is a lack of evidence for such approaches in autism, especially for prebiotics. This study assessed the effects of Pleurotus eryngii mushroom (candidate prebiotic) and inulin (known prebiotic compound) on gut microbial composition, using faecal samples from autistic children in an in vitro batch culture fermentation system. Selected members of GM were enumerated at baseline (0 h) and after 24 h fermentation by quantitative PCR. After 24 h fermentation, inulin and P. eryngii mushroom induced a significant increase in total bacteria and Faecalibacterium prausnitzii compared to the negative control (gut microbiota of each autistic donor with no carbohydrate source), whereas both treatments induced a significant increase in levels of total bacteria, Bifidobacterium spp. and Prevotella spp. compared to baseline (t=0h) (p for all <0.05). Furthermore, this study evaluated the impact of fermentation supernatants (FSs), derived from P. eryngii mushroom or inulin, on the expression levels of tight junctions’ genes (zonulin-1, occludin and claudin-1) in Caco-2 cells stimulated by bacterial lipopolysaccharides (LPS). Pre-incubation of Caco-2 cells with FS from P. eryngii mushroom led to a significant increase in the expression levels of zonulin-1, occludin and claudin-1 genes compared to the untreated cells, the cells that were subjected to LPS and the cells that were challenged with FS from negative control (p for all <0.05). In addition, incubation with FS from P. eryngii mushroom led to the highest mean expression values for zonulin-1 and claudin-1 genes, which differed significantly compared to inulin (p for all <0.05). Overall, this research highlighted the beneficial in vitro effects of P. eryngii mushroom on the composition of GM of autistic children after 24 h of fermentation. Also, our data highlighted the potential preventive effect of P. eryngii FSs against dysregulation of the intestinal barrier, through upregulation of tight junctions’ genes associated with the integrity and function of the intestinal barrier. This research has been financed by "Supporting Researchers with Emphasis on Young Researchers - Round B", Operational Program "Human Resource Development, Education and Lifelong Learning."

Keywords: gut microbiota, intestinal barrier, autism spectrum disorders, Pleurotus Eryngii

Procedia PDF Downloads 156

Authors: Suvalux Chaichuchote, Ratchadaporn Thonghem

Abstract:

Pleurotus mushroom is one of popular edible mushrooms in Thailand. It is much favored by consumers due to its delicious taste and high nutrition. It is commonly used as an ingredient in several dishes. The commercially cultivated strain grown in most farms is the Pleurotus sp., Hed Bhutan, that is widely distributed to mushroom farms throughout the country and can be cultivated almost all year round. However, it demands different cultivated strains from mushroom growers, therefore, the improving mushroom strains should be done to their benefits. In this study, we used a di-mon mating method to hybrid production from Hed Bhutan (P-3) as dikaryon material and monokaryotic mycelium were isolated from basidiospores of other three Pleurotus sp. by single spore isolation. The 3 hybrids: P-3XSA-6, P-3XSB-24 and P-3XSE-5 were recognized from the 12 hybridized successfully. They were appropriate hybridized in terms of fruiting body performance in the three time cycles of cultivation such as the number of days until growing, time for pinning, color and shape of fruiting bodies and yield. For genetic study, genomic DNAs of both Hed Bhutan (P-3) and three hybrids were extracted. A couple of primer ITS1 and ITS4 were used to amplify the gene coding for ITS1, ITS2 and 5.8S rRNA. The similarities between these amplified genes and databases of DNA revealed that Hed Bhutan (P-3) was the Pleurotus pulmonarius as well as P-3XSA-6, P-3XSB-24 and P-3XSE-5 hybrids. Furthermore, Hed Bhutan (P3) and three hybrids were distributed to 3 small-scale farms, with mushroom farming experience, in the countryside. To address this, one hundred and twenty mushroom bags of each strain were supplied to them. The findings, by interview, indicated two mushroom farmers were satisfied with P-3XSA-6 hybrid and P-3XSB-24 hybrid, thanks to their simultaneous fruiting time and good yield. While the other was satisfied with P-3XSB-24 hybrid due to its good yield and P-3XSE-5 hybrids thanks to its gradually fruiting body, benefiting in frequent harvest. Overall, farmers adopted all hybrids to grow as commercially cultivated strains as well as Hed Bhutan (P-3) strain.

Keywords: dikaryon, monokaryon, pleurotus, strain improvement

Procedia PDF Downloads 238