Opportunistic Pathogenic Microorganisms Research Articles

Antagonistic and antibacterial activity of <i>Lactobacillus rhamnosus</i> HN001 and <i>Lactobacillus acidophilus</i> La-14 included in the oral probiotic

BACKGROUND: A microbial imbalance in the vaginal biotope leads to the development of vaginal infections and is associated with urinary tract infections. The ineffectiveness of the therapy and the frequent recurrence of these infections are important reasons for searching for alternative treatment strategies. A progressive solution to this issue has been the use of oral probiotics with a targeted effect on the intestinal and vaginal microflora. AIM: The aim of this study was to evaluate the antagonistic and antibacterial activity of the probiotic strains Lactobacillus rhamnosus HN001 and Lactobacillus acidophilus La-14, which are the components of the oral probiotic, against opportunistic microorganisms of intestinal origin, while assessing the sensitivity of these strains to antimicrobial drugs. MATERIALS AND METHODS: The probiotic strains Lactobacillus rhamnosus HN001 and Lactobacillus acidophilus La-14 included in the oral probiotic Duogynal® were cultivated and the grown colonies were identified using the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry method (Bruker Daltonics GmbH Co., Germany). To study the antagonism of the probiotic strains of lactobacilli and opportunistic pathogenic microorganisms, the following 11 clinical isolates were collected: Escherichia coli (two isolates), Klebsiella pneumoniae, Klebsiella oxytoca, Enterobacter asburiae, Proteus mirabilis, Citrobacter freundii, Staphylococcus aureus, Enterococcus faecalis, Streptococcus agalactiae, and Candida albicans. To assess the sensitivity of the probiotic strains to antimicrobial drugs, the Gram Positive AST panel (Autobio Diagnostics Co., Ltd, China) was used. RESULTS: When the microorganisms were co-cultivated with the probiotic lactobacilli (Lactobacillus rhamnosus HN001 and Lactobacillus acidophilus La-14), we noted a decrease in pH and the death of opportunistic microorganisms such as Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca, Enterobacter asburiae, Proteus mirabilis, Citrobacter freundii, Staphylococcus aureus, Enterococcus faecalis, and Streptococcus agalactiae, as well as yeast-like fungi Candida albicans. When studying antibiotic resistance, it was found that the both strains of lactobacilli were resistant to ampicillin, vancomycin, daptomycin, clindamycin, linezolid, moxifloxacin, nitrofurantoin, oxacillin, oritavancin, penicillin, rifampin, teicoplanin, tigecycline, trimethoprim/sulfamethoxazole, cefoxitin, ceftarolin, and ciprofloxacin. CONCLUSIONS: The oral probiotic, which contains a combination of two lactobacilli (Lactobacillus rhamnosus HN001 and Lactobacillus acidophilus La-14), has pronounced antagonistic activity against opportunistic microorganisms. These strains are resistant to most antibacterial drugs used in widespread clinical practice, which may be the basis for recommending the use of this probiotic drug together with prescribed antibiotics for prevention of vaginal or intestinal dysbiosis.

Влияние антибактериального рассола на качество и микробиологическую безопасность продуктов из мяса

Nomad livestock farming is an extra raw material resource for regions with harsh climate. Foods based on meat of indigenous animals expand the range of products with high consumer properties. The quality and safety of meat products usually depend on the brine. The present research featured the effect of brine with Island moss (Cetrária islándica (L.) Ach.) on the sensory profile and microbiological safety of meat products.
 The study involved meat of two-year-old indigenous cows, as well as multicomponent antibacterial and standard extrusion brines. The brine was injected with a syringe-injector with a hollow perforated needle at a 20 mm pitch. The tenderization took place in a universal express tenderizer with a rotation speed of 16 rpm and lasted for 10 h. The shelf-life was determined based the on microbiological, chemical, and sensory indicators.
 By the end of the reserve shelf-life, the total microbial count was 1.2×103 CFU/g in the control and 1.0×103 CFU/g in the experimental sample. No opportunistic pathogenic bacteria or microorganisms were detected. No opportunistic pathogenic microorganisms were revealed when the microbiological parameters were studied in standard quantities and in quantities increased by an order of magnitude.
 The changes in acid and peroxide numbers during storage indicated a stable oxidation of the fat component in the experimental sample. Its sensory evaluation exceeded the control sample by 0.9 points in terms color, texture, and smell. The meat marinated with C. islándica compiled with the quality standard and had a shelf-life which exceeded that of the traditionally-marinated meat by 5 days.

Microbiological analysis of broiler chicken slaughter products using “Subtiform” probiotic and establishment of safety and technological process criteria

Feeding broiler chickens a probiotic biopreparation containing bacteria of the genus Bacillus subtilis and Bacillus licheniformis leads to improved feed digestion, increased productivity, and immune status, as well as reduced insemination by opportunistic and pathogenic microorganisms of slaughter products. It enhances the level of prevention and treatment of various poultry diseases. The purpose of the work is to carry out a microbiological analysis of the slaughter products of broiler chickens after drinking the probiotic biological preparation “Subtiform” in doses of 0.5 g, 2.0, and 4.0 g per 10 dm3 of water. The material (major pectoral muscle and internal organs) was examined by bacteriological methods. It was established that the content of MAFAM in the control and experimental groups 1, 2, and 3 of the chilled slaughter products of broiler chickens for 1, 3, and 5 days of storage at a temperature of 0–4 °С was within the limits of standards (no more than 1.0×104 CFU/d). It was established that when broiler chickens were given 4.0 g/10 dm3 of water, the MAFAM content for one day of poultry meat storage decreased by 10.4 % (Р < 0.05); on the third day of storage, it decreased by 9.6 % (Р < 0.05); at the beginning of the fifth day of storage, the decrease was, respectively, by 11.3 % (Р < 0.01) compared to the indicators of the control group. The content of MAFAM in offal, in particular in the heart of broiler chickens, was reduced in experimental group 3 by 36.9 % (Р < 0.001); in the liver – by 33.6% (Р < 0.001); in the muscular part of the stomach – by 27.2 % (Р < 0.001); in the spleen and lungs, there was also a decrease in the content of MAFAM in experimental group 3, respectively, by 20.2 % (Р < 0.01) and 23.6 % (Р < 0.01) compared to the indicators of the control group of broiler chickens. It was established that opportunistic pathogenic microorganisms, in particular BGCP, bacteria of the genus Proteus, and pathogenic microorganisms, in specific bacteria of the genus Salmonella, bacteria of the species Staphylococcus aureus, Listeria monocytogenes in 25 grams of the large pectoral muscle of broiler chickens and internal organs (heart, spleen, liver, the muscular part of the stomach and lungs) were not detected. According to the results of the conducted research, an adverse effect of the probiotic biopreparation at a dose of 4.0 g/10 dm3 of water on the development of microorganisms in the large pectoral muscle and internal organs of broiler chickens was established. Therefore, the probiotic “Subtiform” at a dose of 4.0 g/10 dm3 of water during the drinking of broiler chickens can be recommended to increase productivity and obtain safe slaughter products. The practical value of the work consisted of establishing the microbiological indicators of poultry slaughter products after drinking probiotics, as well as establishing safety criteria and the technological process.

Influence of bee products on intestinal microbiota formation in healthy birds and birds with candidiasis

Candidamycosis causes significant damage to the poultry industry. Mortality rate in young birds reaches 95100 %. The aim of the study was to optimize formation and intestinal colonization resistance under the influence of biologically active bee products (BABP) and the features of recovery in birds infected with candidamycosis of digestive tract (CDT). Studies were carried out on Japanese meat 10-day-old to 90-day-old quails. All the studied beekeeping products extracts of wax moth, drone homogenate and propolis contribute to the stabilization of the hidden genetically laid down mechanisms of natural intestinal microbiocenosis in healthy quails bred in captivity, without causing damage to the body, restoring the balance of normoflora and opportunistic pathogenic microorganisms to physiological values. However, a constant complex of stressors encountered during bird breeding, associated with the conditions of keeping, feeding, veterinary and zootechnical measures, lead to a significant activation of Candida albicans in large intestine, with the subsequent development of candidiasis of digestive tract, deep dysbacterioses, characterized by increased reproduction and increase in the content of opportunistic pathogenic microorganisms: Candida albicans by 8.34 times, Staphylococcus aureus by 4.37 times, Pseudomonas spp. by 3.29 times; inhibition of reproduction and decrease in the level of normoflora: Lactobacillus spp. by 6.0 times, Bifidobacterium spp. by 7.25 times. The use of extracts of wax moth, drone homogenate and propolis in candidamycosis-i nfected birds contributed to restoration of quail intestinal microbiocenosis, which was manifested by: a) decrease in the level of opportunistic pathogenic Candida albicans by 3.3; 4.61 and 3.97 times; Staphylococcus aureus by 4.0; 7.78 and 4.5 times; Pseudomonas spp. by 3.05; 5.32 and 3.96 times; b) activation and increase of normoflora: Lactobacillus spp. by 6.38; 10.0 and 8.84 times; Bifidobacterium spp. by 5.36; 8.42 and 7.5 times.

D-histidine combated biofilm formation and enhanced the effect of amikacin against Pseudomonas aeruginosa in vitro

Pseudomonas aeruginosa is an opportunistic gram-negative pathogenic microorganism that poses a significant challenge in clinical treatment. Antibiotics exhibit limited efficacy against mature biofilm, culminating in an increase in the number of antibiotic-resistant strains. Therefore, novel strategies are essential to enhance the effectiveness of antibiotics against Pseudomonas aeruginosa biofilms. D-histidine has been previously identified as a prospective anti-biofilm agent. However, limited attention has been directed towards its impact on Pseudomonas aeruginosa. Therefore, this study was undertaken to explore the effect of D-histidine on Pseudomonas aeruginosa in vitro. Our results demonstrated that D-histidine downregulated the mRNA expression of virulence and quorum sensing (QS)-associated genes in Pseudomonas aeruginosa PAO1 without affecting bacterial growth. Swarming and swimming motility tests revealed that D-histidine significantly reduced the motility and pathogenicity of PAO1. Moreover, crystal violet staining and confocal laser scanning microscopy demonstrated that D-histidine inhibited biofilm formation and triggered the disassembly of mature biofilms. Notably, D-histidine increased the susceptibility of PAO1 to amikacin compared to that in the amikacin-alone group. These findings underscore the efficacy of D-histidine in combating Pseudomonas aeruginosa by reducing biofilm formation and increasing biofilm disassembly. Moreover, the combination of amikacin and D-histidine induced a synergistic effect against Pseudomonas aeruginosa biofilms, suggesting the potential utility of D-histidine as a preventive strategy against biofilm-associated infections caused by Pseudomonas aeruginosa.

An improved recombinase polymerase amplification assay for the visual detection of Staphylococcus epidermidis with lateral flow strips

Staphylococcus epidermidis is an opportunistic pathogenic microorganism that is an important cause of cross-infection in hospitals. The development of rapid and effective detection techniques is important for its control. The application of traditional identification and PCR-based methods is limited by their requirements for both laboratory instrumentation and trained personnel. To overcome this issue, we developed a fast detection approach for S. epidermidis that was based on recombinase polymerase amplification (RPA) and lateral flow strips (LFS). First, five pairs of primers were designed for molecular diagnosis using the sesB gene as the target, and were screened for their amplification performance and the formation of primer dimers. Specific probes were then designed based on the best primer pairs screened, which were susceptible to primer-dependent artifacts and generated false-positive signals when used for LFS detection. This weakness of the LFS assay was overcome by modifying the sequences of the primers and probes. The efficacy of these measures was rigorously tested, and improved the RPA-LFS system. Standardized systems completed the amplification process within 25 min at a constant temperature of 37 °C, followed by visualization of the LFS within 3 min. The approach was very sensitive (with a detection limit of 8.91 CFU/μL), with very good interspecies specificity. In the analysis of clinical samples, the approach produced results consistent with PCR and 97.78% consistent with the culture-biochemical method, with a kappa index of 0.938. Our method was rapid, accurate, and less dependent on equipment and trained personnel than traditional methods, and provided information for the timely development of rational antimicrobial treatment plans. It has high potential utility in clinical settings, particularly in resource-constrained locations.

New Perspective on Comparative Chemometric and Molecular Modeling of Antifungal Activity and Herbicidal Potential of Alkyl and Cycloalkyl s-Triazine Derivatives

The contamination of the environment by pesticides is becoming a burning issue in many countries in the World. Development, design, and synthesis of new eco-friendly pesticides and modification of existing ones in order to improve their efficacy with the lowest impact on the environment are two main future possibilities in crop protection and the provision of sufficient food for the growing world population. The present study is focused on the comparative analysis of a series of eight symmetrical triazine derivatives, as potential herbicide candidates with acyclic (alkyl) and cyclic (cycloalkyl) substituents, in terms of their antifungal activity towards Aspergillus flavus as an opportunistic fungal pathogenic microorganism responsible for frequent contaminations of crops with aflatoxin, and in terms of their potential application as herbicides in maize, common wheat, barley, and rice crops. The applied methods include the chemometric pattern recognition method (hierarchical cluster analysis), experimental microbiological analysis of antifungal activity (agar well-diffusion method), and molecular docking of the triazines in the corresponding enzymes. The main findings of the conducted study indicate the significant antifungal activity of the studied triazine derivatives towards A. flavus, particularly the compounds with acyclic substituents; five out of eight studied triazines could be applied as systematic herbicides, while the other three triazines could be used as contact herbicides; the compounds with acyclic substituents could be more suitable for application for various crops protection than triazines with cyclic substituents.

The Role of Iron in Staphylococcus aureus Infection and Human Disease: A Metal Tug of War at the Host-Microbe Interface.

Iron deficiency anemia can be treated with oral or intravenous Fe supplementation. Such supplementation has considerable effects on the human microbiome, and on opportunistic pathogenic micro-organisms. Molecular understanding of the control and regulation of Fe availability at the host-microbe interface is crucial to interpreting the side effects of Fe supplementation. Here, we provide a concise overview of the regulation of Fe by the opportunistic pathogen Staphylococcus aureus. Ferric uptake regulator (Fur) plays a central role in controlling Fe uptake, utilization and storage in order to maintain a required value. The micro-organism has a strong preference for heme iron as an Fe source, which is enabled by the Iron-regulated surface determinant (Isd) system. The strategies it employs to overcome Fe restriction imposed by the host include: hijacking host proteins, replacing metal cofactors, and replacing functions by non-metal dependent enzymes. We propose that integrated omics approaches, which include metalloproteomics, are necessary to provide a comprehensive understanding of the metal tug of war at the host-microbe interface down to the molecular level.

Extraction possibilities of lipid fraction and authentication assessment of chaga (Inonotus obliquus)

Chaga (Inonotus obliquus) (Fr.) Pilát is a black perennial fungus that grows on adult birch trunks and has been traditionally used as a health promoting remedy in different countries. The lipophilic compounds of chaga were isolated applying conventional (Soxhlet), intensive (ultrasound assisted, accelerated solvent extraction) and environmentally friendly (supercritical fluid) extraction methods utilizing both polar and non-polar solvents. Soxhlet extraction showed the highest inotodiol yield (231 mg/100 g chaga) using cyclohexane. For authentication of chaga samples, isotope ratio mass spectrometry (IRMS) and multi-elemental analysis was performed. The extraction yield and profile of lipids, sterols and triterpenoids depend on the chaga origin. The amount of lipids in chaga samples varied in the range from 0.5 to 1.0%. The antimicrobial activity of chaga extracts against pathogenic and opportunistic pathogenic microorganisms was demonstrated. The results showed that extraction using supercritical CO2 delivered yields comparable to those obtained by conventional and intensive extraction techniques. Amount of target compounds (sterols and triterpenoids) obtained using supercritical CO2 extraction could represent the future for sustainable industrial-scale production of biologically active ingredients with antimicrobial activity. Authentication using IRMS allowed to differentiate between natural (fruiting bodies) and cultivated (mycelium) chaga.

Influence of grain physical characteristics on functional value of poultry feed

The purpose of this investigation was to analyze the scientific literature concerning the influence of grain physical characteristics on functional value of poultry feed. The review considers the role of grain feed particle size and its shape in ensuring optimal development and functioning of digestive tract and increasing productivity of birds. The results showed that compared to the feeding with smaller particles, the use of coarse feed increased stomach weight in broilers, enhanced enzymatic degradation of feed in digestive tract and improved nutrient bioavailability. The study found that increase in the level of coarse corn in the diet increased the number of Lactobacillus spp. and Bifidobacteria spp. beneficial bacteria in digestive tract. Meanwhile, number of opportunistic pathogenic microorganisms decreased. In birds which were given granular feed we found insufficient development of stomach, mainly due to the lack of mechanical stimulation. We assume that in order to improve stomach function and health of birds, it is necessary to apply a source of dietary fiber to their diet. The results of studies suggest that adding 2-3% of insoluble source of fiber to the routine diets of young chickens based on high-protein soy flour and corn can improve their digestive tract development and growth characteristics.

Effect of hypo– and hypersaline stress conditions on physiological, metabolic, and immune responses in the oyster Crassostrea corteziensis (Bivalvia: Ostreidae)

Salinity in the oceans is changing due to climate change and global warming. Intense rainfalls and freshwater runoff decrease salinity along the coastal areas. In contrast, intense drought seasons and river damming have certainly increased salinity in lagoons and estuaries. Few studies have focused on aspects of the biology and culture of oyster Crassostrea corteziensis, but until now, physiological and immunological responses in this species have not been assessed under acute hypo– and hypersaline stress conditions. Oysters obtained from a local farm were acclimated for three weeks in laboratory conditions. To avoid closure of oyster valves during salinity induced–stress conditions, a notch was done on each organism shell not only to facilitate oyster tissue exposure to rearing water but also for sampling hemolymph. Oysters (N = 180) were abruptly exposed to three salinity treatments: (HO) hypo–, (C) control, and (HP) hypersaline stress conditions (10, 35, and 50 PSU, respectively). Four oysters per treatment were sampled at 1, 2, 3, 6, 12, 24, and 48 h after exposure. Hemolymph osmolality, water content and total protein concentration in tissues, metabolic and immune responses were assessed for each organism. Oyster survival was not different among treatments and was maintained above 96% at the end of the experimental trial. Hemolymph osmolality reached the value of rearing water at 6 and 48 h of exposure to HP and HO stress conditions, where oysters exposed to salinity increase showed less resilience than those to decrease. Higher glucose levels in plasma and lower ones of hemocyanin were assessed in the oysters exposed to HP compared to HO conditions, suggesting more stressful conditions or susceptibility of oysters during salinity increase. Total hemocyte (THC), hyalinocyte (HC), and granulocyte (GC) counts decreased in oysters exposed to HP condition, while total and differential hemocyte counts were similar among oysters exposed to HO and control conditions. Despite hemocyte phagocytosis was not different among treatments, viability decreased in those exposed to HP condition. Contrastingly, superoxide anion (SOA) production (oxidative capacity) increased in oysters exposed to both induced salinity–stress conditions, which suggest susceptibility increase in oysters, particularly during salinity increase. The results show that HP condition is particularly stressful for C. corteziensis. In turn, this condition could increase both their vulnerability to other environmental stressors, such as temperature and/or acidification or susceptibility to opportunistic pathogenic microorganisms that cause the most common oyster diseases.

Колонізаційна резистентність слизової оболонки товстої кишки білих щурів з експериментальним гіпотиреозом

Актуальність. Мікроекосистема організму людини тісно взаємодіє з імунною та ендокринною системами, і власне мікробіом приепітеліальної біоплівки слизової оболонки товстої кишки є одним з найчутливіших біотопів, здатних зазнавати змін за різних захворювань. Мета. Встановлення мікробіологічної структури (видовий склад і популяційний рівень мікрофлори) приепітеліальної біологічної плівки слизової оболонки товстої кишки білих щурів з експериментальним гіпотиреозом. Матеріали та методи. Проведено експерименти на 25 статевозрілих самцях білих щурів масою 220–240 г, з яких 15 тварин були віднесені до контрольної групи (інтактні тварини), 10 щурів належали до основної групи. Експериментальний гіпотиреоз моделювали шляхом внутрішньошлункового введення мерказолілу впродовж 14 днів. У стерильних умовах проводили лапаротомію, брали відрізок (до 3 см) товстої кишки з її вмістом. Відмиту частину кишки гомогенізували зі стерильним 0,9% розчином хлориду натрію. Готували серію десятикратних розведень з концентрацією суміші від 10–2 до 10–7. З кожної пробірки 0,01 мл висівали на тверді оптимальні живильні середовища з подальшим виділенням та ідентифікацією мікробів за морфологічними, тинкторіальними, культуральними та біохімічними властивостями. Результати. Колонізаційна резистентність приепітеліальної біоплівки слизової оболонки товстої кишки в експерименті на дослідних тваринах при моделюванні гіпотиреозу зазнає змін: знижується популяційний рівень лактобактерій на 51,58 %, біфідобактерій — на 37,35 %, бактероїдів — на 22,82 %, пептострептококів — на 6,5 %. Приепітеліальна біоплівка контамінується кишковою паличкою та іншими ентеробактеріями (Proteus, Klebsiella), P.niger, стафілококами. Встановлено зростання популяційного рівня кишкових паличок на 10,31 % і клостридій на 8,2 %. Детерміновано дисбіотичні порушення І–III ступеня, що потребує корекції і лікування за допомогою пробіотиків. Висновки. В експерименті на білих щурах при мерказоліл-модельованому гіпотиреозі встановлено елімінацію бактерій роду Bifidobacterium, Lactobacillus, контамінацію і колонізацію біотопу умовно-патогенними мікроорганізмами родів Proteus, Klebsiella, Staphylococcus, Peptococcus на тлі зростання популяційного рівня кишкової палички. Наявні порушення мікробіоценозу у вигляді дисбіозу I–III ступеня.

Nutmeg nutraceutical constituents: In vitro and in vivo pharmacological potential

Spices consumptions are in direct relation with the healthy survivability of consumers throughout life as they are loaded with lists of phytoconstituents of human values. Nutmeg owes to its essential, volatile, and fixed oil is in prime most interest by researchers to explore for its benefits through food utilization. Besides being loaded with these phytochemical-based benefit traits it is also rich on the mineral side preferably with potassium, magnesium, and phosphorous. As for as the oil variation is concern, nutmeg essential oil composition behaves as per the geographical and varietal factors. Its beneficial aspect highlighted wherein oil of nutmeg contains Myristicin (phenylpropene organic compound) can exhibit anti-inflammatory response and have the cytotoxic activity against human colorectal carcinoma (HCT-116) and human breast carcinoma Michigan Cancer Foundation (MCF-7). Associated benefits of being on antiangiogenic compounds such as myristicin in Myristica fragrans essential oil resulted in antiangiogenic activity with an IC50 of 77.64 g/ml. Practical applications Nutmeg has its wider acceptability among medicinal and therapeutic uses in today's world. Indeed, many would say that nutmeg use has increased. Nutmeg's traditional applications in treating stomach conditions, curing rheumatic pain, soothing skin wounds and illnesses, and acting as a relaxing agent have resulted in extensive research into its various extracts and fractions. One of the most commonly observed habits is the influence of nutmeg essential oil and extracts on the central nervous system. Nutmeg acetone extract's antioxidant and antimicrobial properties can help to prevent or delay oxidative stress-related diseases and infections caused by opportunistic pathogenic microorganisms. Nutmeg requires further work on several fronts, including the advancement of analytical approaches, reproducible separation procedures, and successful synthetic methods for promising secondary metabolites, as a source of complex secondary metabolites with substantial potential as prototype agents for drug testing.

Plastics in our water: Fish microbiomes at risk?

Water contaminated with plastic debris and leached plasticizers can be ingested or taken up by aquatic invertebrates and vertebrates alike, exerting adverse effects on multiple tissues including the gastrointestinal tract. As such, gut microbiomes of aquatic animals are susceptible targets for toxicity. Recent studies conducted in teleost fishes report that microplastics and plasticizers (e.g., phthalates, bisphenol A) induce gastrointestinal dysbiosis and alter microbial diversity in the gastrointestinal system. Here we synthesize the current state of the science regarding plastics, plasticizers, and their effects on microbiomes of fish. Literature suggests that microplastics and plasticizers increase the abundance of opportunistic pathogenic microorganisms (e.g. Actinobacillus, Mycoplasma and Stenotrophomonas) in fish and reveal that gamma-proteobacteria are sensitive to microplastics. Recommendations moving forward for the research field include (1) environmentally relevant exposures to improve understanding of the long-term impacts of microplastic and plasticizer contamination on the fish gastrointestinal microbiome; (2) investigation into the potential impacts of understudied polymers such as polypropylene, polyamide and polyester, and (3) studies with elastomers such as rubbers that are components of tire materials, as these chemicals often dominate plastic debris. Focus on both microplastics and the gut microbiota is intensifying in environmental toxicology, and herein lies an opportunity to improve evaluation of global ecological impacts associated with plastic contamination. This is important as the microbiota is intimately tied to an individual's health and fragmentation of microbial community networks and gut dysbiosis can result in disease susceptibility and early mortality events.

Effect of Cocultivation on Lactobacillus plantarum Strains Growth and Antagonistic Activity

The use of bacterial starters for the production of fermented foods has several advantages over traditional spontaneous fermentation, as it provides a rapid and controlled decrease of pH, improves the microbiological quality of the product, and prolongs the shelf-life. Fermented foods are typically produced using mixed cultures of lactic acid bacteria (LAB) due to the synergism between their constituent bacterial cultures. So, the compatibility of the LAB strains decides the efficacy of a multi-strain starter. The purpose of this study was to investigate the effect of the cocultivation of Lactobacillus plantarum strains on the growth, acidification, and antagonistic activity to determine suitable strain combinations for fermented vegetable production. Methods. The effect of cocultivation on growth characteristics of four L. plantarum strains was determined in MRS medium and cabbage-based medium with 2.5% NaCl. After 8 h of cultivation at 30°C and 37°C, the number of viable cells (CFU/ml) and the pH of the medium were determined. The antagonistic activity of monocultures of L. plantarum and their six compositions against opportunistic pathogenic microorganisms was determined by the method of delayed antagonism. Results. During growth in MRS broth at 30°C cocultivation of L. plantarum 47SM with L. plantarum 691T or L. plantarum 1047K strains led to enhanced rates of growth compared to the monocultures, suggesting some degree of symbiosis between these strains. Viable cell counts of L. plantarum 47SM, 1047K and 691T strains and ΔpH values of L. plantarum 952K, 1047K, and 691T strains were higher after 8 h growth in the cabbage-based medium at 30°C compared to MRS broth. Despite the intensive growth of L. plantarum monocultures in cabbage-based medium, a significant decrease of viable cell counts and ΔpH values during cocultivation at 30°C were found. Cocultivation did not affect the average size of the growth inhibition zones of most of the indicator strains used. However, growth inhibition zones of Shigella flexneri, Escherichia coli, and Proteus vulgaris decreased in some L. plantarum mixed cultures compared to monocultures. Thus, the growth inhibition zones of E. coli and S. flexneri by mixed culture L. plantarum 47 SM+1047K were significantly smaller compared to the growth inhibition zones of L. plantarum monocultures. Conclusions. Thus, based on the data obtained in present work, we can assume that some of these L. plantarum strains used in the work may be bactericinogenic. Although the four L. plantarum strains studied are compatible when cocultivated in a standard rich MRS medium, the results of cocultivation in a cabbage-based medium with 2.5% NaCl does not allow to recommend the use of these L. plantarum strains simultaneously in the starter for vegetable fermentation. Further investigation of bacteriocinogenic properties and mechanisms of growth inhibition under cocultivation in vegetable-like conditions are needed, which will allow combining of some of these L. plantarum strains with LAB strains of other species or genera to create multi-starters for vegetable fermentation.

Biotyping and Antimicrobial Susceptibility of Enterococcus faecalis and E. faecium Isolated from Urine and Stool Samples

Background: Enterococci are one of the opportunistic pathogenic microorganisms that can cause significant problems for human and animal health. Enterococcus faecium seems to be more resistant to antibiotics than E. faecalis. It is thought that pathogenic E. faecium can develop antibiotic resistance very quickly, and the ability to transfer this feature is considered to be an important health risk. Objectives: This study aimed to determine the prevalence, biotypes, and in vitro antimicrobial susceptibility of E. faecalis and E. faecium strains isolated from 267 routine urine and stool samples that were brought to the microbiology laboratory of Regional Training and Research Hospital of Van, with permission of the patients. Methods: In the present study, enterococci using species-specific primers to examine E. faecalis and E. faecium multiplex PCR technique was applied. Biotyping of the isolates was used to identify them as E. faecalis and E. faecium by molecular techniques, and antibiotic susceptibility of all samples was examined, as well. Results: The isolates were identified by multiplex PCR using species-specific primers for E. faecalis and E. faecium. Biotyping based on 13 biochemical tests showed that 72.5%, 12.5%, and 15% of E. faecalis strains were of biotypes I, II, and III, respectively, whereas E. faecium strains could be divided into biotype I (10%), biotype II (12.5%), biotype III (27.5%), and biotype IV (50%). Additionally, all E. faecalis strains were found to be susceptible to penicillin G and imipenem. On the other hand, 95% of the E. faecalis strains were found to be resistant to clindamycin, 77.5% to tetracycline and trimethoprim/sulfamethoxazole, 42.5% to erythromycin, 32.5% to gentamicin, and 17.5% to ciprofloxacin. Of E. faecium strains, 37.5% were found to be resistant to clindamycin, 32.5% to penicillin G, 27.5% to erythromycin and imipenem, 20% to ciprofloxacin, 17.5% to tetracycline and trimethoprim/sulfamethoxazole, 15% to gentamicin, and 5% to vancomycin. Conclusions: In conclusion, the identification of E. faecalis and E. faecium strains by PCR is reliable and faster than biochemical tests. Additionally, the results of antimicrobial susceptibility tests may provide important contributions to the clinical approach.

Serratia fajok jellemzése, valamint Serratia marcescens kvalitatív kimutatása nyers és pasztőrözött tejből polimeráz láncreakción alapuló vizsgálati módszerrel

Serratia species are opportunistic pathogenic microorganisms primarily known as nosocomial infectious agents, which can also cause food quality problems. The appearance of the extracellular pigment-producing Serratia marcescens in cow’s milk causes its red discoloration, posing a challenge to the dairy industry and food certification laboratories. The detection of the bacterium by conventional procedures based on microbiological methods is time-consuming and labor-intensive, and in many cases does not lead to satisfactory results due to the competitive inhibitory effect of the accompanying microflora. Following the analysis of the relevant literature, the published endpoint PCR methods and the primers used for the detection of S. marcescens were evaluated in in silico and in vitro assays, and then the procedure was tested on farm milk samples. Using the method, a total of 60 raw and pasteurized milk samples were analyzed, more than half of which (i.e., 32) were identified as S. marcescens positive. The significance of our work is mainly represented by the application of the published test methods in food industry practice. Our results highlight to the importance of detecting this bacterial species.

Characteristics of bactericidal activity of healthy donors' peripheral blood against Escherichia coli

Recently, the role of opportunistic pathogenic microorganisms in the development of inflammatory diseases of various localization has been relatively widely discussed. It is known that Escherichia coli are the etiological agent of such diseases, especially extraintestinal localization, a distinctive feature of which is considered to be a wide range of clinical manifestations. The aim of investigation was to assess the contribution of some components to the total microbicidal activity of healthy donors peripheral blood. Materials and methods. The bactericidal capacity of peripheral blood was assessed by the number of viable E. coli after incubation with blood samples by plating on Endo medium. The antibiofilm activity of whole blood and blood plasma was assessed by direct contact with the biofilm and its subsequent visualization using O'Toole technique. Phagocytic activity of neutrophils and monocytes was assessed separately in a microscopic version of the test. The production of reactive oxygen species was assessed using the stimulated and spontaneous luminol-dependent chemiluminescence reaction. An immunoglobulin G was used to opsonize E. coli. Results and discussion. It was shown that the peripheral blood of healthy volunteers has pronounced bactericidal properties against E. coli, reducing the number of viable bacteria by 2 times. Opsonization of E. coli reduces the number of bacteria by 4 times. Among leukocytes, the predominant place is occupied by neutrophils, which activate their absorption capacity and production of radicals faster than monocytes. The number of actively phagocytic neutrophils after E. coli treatment with an immunoglobulin G increased to 4.1±1.0% (without opsonization - 1.7±0.3%; p<0.05). Conclusion. In general, the microbicidal activity of the blood of healthy people is a multi-component system, in which a special place should be given to the factors of opsonization.

Grapevine extracts and their effect on selected gut-associated microbiota: In vitro study

The biological activity of polyphenol substances contained in food supplements prepared from Vitis vinifera can affect the microorganisms present in the digestive tract in terms of their representation and activity of the individual species. This study deals with resveratrol and two polyphenol-rich extracts (extract from V. vinifera canes and the commercial product Regrapex-R-forte) and their effect on selected gut microbiota (Bifidobacterium animalis subsp. lactis Bb-12, Lactobacillus acidophilus LA-5, Lactobacillus casei Lafti L-26, Citrobacter freundii DBM 3127, Escherichia coli DBM 3125). The effect of the studied agents on planktonic and biofilm growth of the microorganisms was determined as minimum inhibitory concentration (MIC80) and minimum biofilm inhibitory concentration (MBIC80), respectively. The extracts induced metabolic activity as well as total biofilm biomass production in probiotic strain L. acidophilus LA-5 while successfully inhibiting the growth of opportunistic pathogenic microorganisms C. freundii DBM 3127 and E. coli DBM 3125.

Virulence Characteristics of mecA-Positive Multidrug-Resistant Clinical Coagulase-Negative Staphylococci.

Coagulase-negative staphylococci (CoNS) are an important group of opportunistic pathogenic microorganisms that cause infections in hospital settings and are generally resistant to many antimicrobial agents. We report on phenotypic and genotypic virulence characteristics of a select group of clinical, mecA-positive (encoding penicillin-binding protein 2a) CoNS isolates. All CoNS were resistant to two or more antimicrobials with S. epidermidis strain 214EP, showing resistance to fifteen of the sixteen antimicrobial agents tested. Aminoglycoside-resistance genes were the ones most commonly detected. The presence of megaplasmids containing both horizontal gene transfer and antimicrobial resistance genetic determinants indicates that CoNS may disseminate antibiotic resistance to other bacteria. Staphylococcus sciuri species produced six virulence enzymes, including a DNase, gelatinase, lipase, phosphatase, and protease that are suspected to degrade tissues into nutrients for bacterial growth and contribute to the pathogenicity of CoNS. The PCR assay for the detection of biofilm-associated genes found the eno (encoding laminin-binding protein) gene in all isolates. Measurement of their biofilm-forming ability and Spearman’s rank correlation coefficient analyses revealed that the results of crystal violet (CV) and extracellular polymeric substances (EPS) assays were significantly correlated (ρ = 0.9153, P = 3.612e-12). The presence of virulence factors, biofilm-formation capability, extracellular enzymes, multidrug resistance, and gene transfer markers in mecA-positive CoNS clinical strains used in this study makes them powerful opportunistic pathogens. The study also warrants a careful evaluation of nosocomial infections caused by CoNS and may be useful in studying the mechanism of virulence and factors associated with their pathogenicity in vivo and developing effective strategies for mitigation.