Search Results (16)

The domestic cat is the primary reservoir host of three flea-borne Bartonella species, one of which (Bartonella henselae) causes reduced fertility and reproductive failure in experimentally infected cats. Vertical transmission of Bartonella has been documented only in B-cell deficient mice, but not immunocompetent animals. As many free-roaming cats are chronically infected with Bartonella and may be immunocompromised by environmental stress or coinfection, we attempted to isolate Bartonella from the fetal and placental tissues of pregnant queens spayed during trap–neuter–release. Four samples from each tissue (ovary, uterus, fetus, and placenta) were split for direct DNA extraction, liquid culture, and culture on a blood agar plate. Samples from infected queens were inoculated into liquid media and sampled weekly for three weeks for DNA extraction and plating. Bartonella DNA was sequenced directly from 28% (5/18) of the free-roaming queens. For these five queens, liquid enrichment culture was attempted in duplicate for fetal and placental samples. Bartonella clarridgeiae DNA was amplified using qPCR liquid enrichment cultures from the placentas of two cats. These findings suggest that viable Bartonella organisms are present in feline reproductive tissue. Additional studies are needed to assess the transplacental transmission of Bartonella spp. and Bartonella’s influence on fetal development. Full article

The viability and host-seeking behavior of Strongyloides larvae are significantly influenced by soil conditions, emphasizing the critical role of environmental control in disease management. This is particularly relevant given the growing concerns about drug resistance resulting from mass chemotherapy or the use of chemical nematicides. Strongyloides stercoralis was effectively inactivated by exposure to 50 °C for both 12 and 24 h (long-term exposure). Strongyloides ratti was inactivated by 50 °C for 20 min (short-term exposure), 9% saline for 50 min, and a combination of 4% saline and 40 °C for 50 min. The combined treatment successfully inactivated S. ratti in four soil mediums using 5% saline at a central temperature of 40 °C. Thermotaxis responses to noxious heat revealed attraction at 40 °C, increased localized searching at 45 °C, and complete inactivation at 50 °C. Larvae migrating within agar at 45 °C were more readily inactivated. Long-range heat attraction at 5 cm resulted in the inactivation of up to 50% of incoming larvae; however, heat-high concentration saline traps at 3 cm distance proved ineffective. Thermal–saline agar trapping demonstrated promise for larval removal in sand, loam, and laterite soils. This method offers a promising approach to larval removal while minimizing hazards to non-target organisms. Full article

Background and Objectives: Enterococcus faecalis (E. faecalis) is a primary pathogen responsible for dental abscesses, which cause inflammation and pain when trapped between the crown and soft tissues of an erupted tooth. Therefore, this study aims to use specific phages as an alternative method instead of classical treatments based on antibiotics to destroy multidrug-resistant E. faecalis bacteria for treating dental issues. Materials and Methods: In the current study, twenty-five bacterial isolates were obtained from infected dental specimens; only five had the ability to grow on bile esculin agar, and among these five, only two were described to be extensive multidrug-resistant isolates. Results: Two bacterial isolates, Enterococcus faecalis A.R.A.01 [ON797462.1] and Enterococcus faecalis A.R.A.02, were identified biochemically and through 16S rDNA, which were used as hosts for isolating specific phages. Two isolated phages were characterized through TEM imaging, which indicated that E. faecalis_phage-01 had a long and flexible tail, belonging to the family Siphoviridae, while E. faecalis_phage-02 had a contractile tail, belonging to the family Myoviridae. Genetically, two phages were identified through the PCR amplification and sequencing of the RNA ligase of Enterococcus phage vB_EfaS_HEf13, through which our phages shared 97.2% similarity with Enterococcus phage vB-EfaS-HEf13 based on BLAST analysis. Furthermore, through in silico analysis and annotations of the two phages’ genomes, it was determined that a total of 69 open reading frames (ORFs) were found to be involved in various functions related to integration excision, replication recombination, repair, stability, and defense. In phage optimization, the two isolated phages exhibited a high specific host range with Enterococcus faecalis among six different bacterial hosts, where E. faecalis_phage-01 had a latent period of 30 min with 115.76 PFU/mL, while E. faecalis_phage-02 had a latent period of 25 min with 80.6 PFU/mL. They were also characterized with stability at wide ranges of pH (4–11) and temperature (10–60 °C), with a low cytotoxic effect on the oral epithelial cell line at different concentrations (1000–31.25 PFU/mL). Conclusions: The findings highlight the promise of phage therapy in dental medicine, offering a novel approach to combating antibiotic resistance and enhancing patient outcomes. Further research and clinical trials will be essential to fully understand the therapeutic potential and safety profile of these bacteriophages in human populations. Full article

Carnivorous plants capture, digest, and absorb prey via specialized structures such as bladders, pitchers, and other modified leaf traps. Studies have shown that not all carnivorous plants produce digestive enzymes; instead, some species rely on microbes living within their traps to produce the necessary enzymes required for prey digestion. Therefore, this study investigated the microbial community (bacteria and fungi) associated with Genlisea hispidula, a rare carnivorous species. Photosynthetic leaves, rhizophylls, and vesicles were processed after either being cleaned and rinsed in sterile water or after being surface sterilized. Tissues were ground in sterile water, serially diluted, lawn plated onto potato dextrose agar, and incubated in darkness for 24 h at 18–23 °C. Axenic cultures were obtained. Identity was determined via molecular sequence similarity of the full bacterial 16S rDNA gene or fungal ITS barcode regions. In total, 48 bacterial species and 29 fungal species were isolated, with Acidocella facilis and Burkholderia spp. being the most dominant isolated bacteria, and Trichomonascus vanleenenianus and Saitozyma spp. being the most dominant isolated fungi. Microbial diversity was greatest on photosynthetic leaves, while the vesicles had the lowest microbial diversity. This study is important because microbial communities play vital roles in maintaining host health and may be required when considering conservation. Full article

Although wooden cutting boards have been used for centuries, debate remains on how best to coat/seal the boards for microbial protection. A growing body of the literature points to the hygroscopicity of wood—its ability to draw water and bacteria from its surface, deep into the wood, where the bacteria are trapped and die—as the wood attempts to even out its moisture content. There is little research as to how coatings affect this mechanism. In this study, four wood species were coated with either a hardening or nonhardening oil and then, were inoculated with either Listeria or Salmonella, then ‘stamped’ onto sterile agar plates at given time intervals to check microbial growth. The coated woods had significantly more recoverable bacteria on their surface than did the uncoated samples. There was no significant difference in performance between the oils. Remaining bacterial loads did vary significantly by wood species, with European beech having significantly less surface bacteria when inoculated with Salmonella, and the oak species having significantly less surface bacteria with Listeria. Multiple factors appeared to affect the ability of wood to move bacteria from its surface, including porosity of the wood, type of bacteria applied, and time. These results run counter to popular understanding of wood cutting board maintenance and will hopefully impact the way households and professional kitchens use and care for their food-contact wood in the future. Full article

The root extracellular trap (RET) consists of root-associated, cap-derived cells (root AC-DCs) and their mucilaginous secretions, and forms a structure around the root tip that protects against biotic and abiotic stresses. However, there is little information concerning the changes undergone by the RET during droughts, especially for tree species. Morphological and immunocytochemical approaches were used to study the RET of black poplar (Populus nigra L.) seedlings grown in vitro under optimal conditions (on agar-gelled medium) or when polyethylene glycol-mediated (PEG₆₀₀₀—infused agar-gelled medium) was used to mimic drought conditions through osmotic stress. Under optimal conditions, the root cap released three populations of individual AC-DC morphotypes, with a very low proportion of spherical morphotypes, and equivalent proportions of intermediate and elongated morphotypes. Immunolabeling experiments using anti-glycan antibodies specific to cell wall polysaccharide and arabinogalactan protein (AGP) epitopes revealed the presence of homogalacturonan (HG), galactan chains of rhamnogalacturonan-I (RG-I), and AGPs in root AC-DC cell walls. The data also showed the presence of xylogalacturonan (XGA), xylan, AGPs, and low levels of arabinans in the mucilage. The findings also showed that under osmotic stress conditions, both the number of AC-DCs (spherical and intermediate morphotypes) and the total quantity of mucilage per root tip increased, whereas the mucilage was devoid of the epitopes associated with the polysaccharides RG-I, XGA, xylan, and AGPs. Osmotic stress also led to reduced root growth and increased root expression of the P5CS2 gene, which is involved in proline biosynthesis and cellular osmolarity maintenance (or preservation) in aerial parts. Together, our findings show that the RET is a dynamic structure that undergoes pronounced structural and molecular remodeling, which might contribute to the survival of the root tip under osmotic conditions. Full article

The antimicrobial properties of two lichen extracts (LC1 and LC2 solutions extracted in acetone and cyclohexane, respectively) were investigated against both Gram-positive and Gram-negative microorganisms through the agar well diffusion assay. Results displayed that both samples were similarly effective against all the indicator strains. The antimicrobial activity was maintained up to 30 days against Candida albicans ATCC 10231 with an inhibition zone of 38 mm and 37 mm for the LC1 and LC2 solutions extracted, respectively. In order to separate the single chemical components and to associate them with the biological activity, the two extracts were subjected to an activity-guided fractionation followed by a liquid chromatography mass spectrometry (LC–MS) Ion Trap 6310A for the chemical characterization. Chromatogram analysis of each sample that maintained an antimicrobial activity revealed the presence of a significant peak, at a retention time (t_R) of 10.8 min, corresponding to a scabrosin derivative that could likely be associated with the antimicrobial activity. Results obtained in the present investigation, especially against the opportunistic pathogen C. albicans, are encouraging and could represent a preliminary step to a future solution toward a microorganism responsible for fungal infections, mainly occurring in immunocompromised patients and recently caused by drug-resistant strains. Full article

Sclerotium rot causes damping-off and stem rot in seedlings and mature mungbeans, which negatively impacts cultivation. The use of a rhizobacterium to control soil-borne diseases is an alternative method to the excess use of synthetic fungicides; therefore, this study aims to screen rhizosphere actinobacteria with fungicidal activities against Sclerotium rolfsii, the pathogen that causes sclerotium rot in mungbeans. Primary screening showed that the Streptomyces sp. isolate Z1-04-02 displayed the highest effectiveness against S. rolfsii in dual culture plates, with a percentage inhibition of 74.28%. An assay containing enzymes that degrade cell walls, of the cell-free culture filtrate (CF) of Z1-04-02, showed that the activities of chitinase and β-1,3-glucanase were 0.0209 and 1.0210 U/mL, respectively, which was significantly higher than that of the control (media alone). The cell-free CF of Z1-04-02, incubated at 37 °C and 100 °C, using agar well diffusion, effectively inhibited the growth of S. rolfsii with inhibition percentages of 37.78% and 27.78%, respectively. Solid-phase microextraction (SPME) was applied to trap volatiles released from Z1-04-02 and gas chromatography–mass spectrometry (GC/MS); volatile antifungal compounds were tentatively identified as bicyclic monoterpene (1R)-(-)-myrtenal. The application of the cell-free CF, and the spore suspension of Z1-04-02, showed disease severity indexes (DSIs) of 12.5% and 8.25%, respectively, which were significantly lower than those showing inoculation by S. rolfsii alone. The identification of this strain by morphology, biochemistry tests, and 16s rDNA sequences revealed that Z1-04-02 was Streptomyces albulus. This finding revealed that S. albulus Z1-04-02 displayed diverse fungicidal activities against S. rolfsii, and it has the potential to act as a biological control agent in terms of inhibiting sclerotium rot in mungbeans. Full article

Haemonchus contortus (Hc) is a parasite affecting small ruminants worldwide. Arthrobotrys musiformis (Am) is a nematode-trapping fungi that captures, destroys and feeds on nematodes. This study assessed the predatory activity (PA) and nematocidal activity (NA) of liquid culture filtrates (LCF) of Am against Hc infective larvae (L3), and additionally, the mycochemical profile (MP) was performed. Fungal identification was achieved by traditional and molecular procedures. The PA of Am against HcL3 was performed in water agar plates. Means of non-predated larvae were recorded and compared with a control group without fungi. LCF/HcL3 interaction was performed using micro-tittering plates. Two media, Czapek–Dox broth (CDB) and sweet potato dextrose broth (SPDB) and three concentrations, were assessed. Lectures were performed after 48 h interaction. The means of alive and dead larvae were recorded and compared with proper negative controls. The PA assessment revealed 71.54% larval reduction (p < 0.01). The highest NA of LCF was found in CDB: 93.42, 73.02 and 51.61%, at 100, 50 and 25 mg/mL, respectively (p < 0.05). Alkaloids and saponins were identified in both media; meanwhile, coumarins were only identified in CDB. The NA was only found in CDB, but not in SPDB. Coumarins could be responsible for the NA. Full article

Followed by a buildup of its phytochemical profile, Erodium cicutarium is being subjected to antimicrobial investigation guided with its ethnobotanical use. The results of performed in vitro screening on Staphylococcus aureus, Pseudomonas aeruginosa, and Candida albicans strains, show that E. cicutarium has antimicrobial activity, with a particular emphasis on clinical S. aureus strains—both the methicillin sensitive (MSSA) and the methicillin resistant (MRSA) S. aureus. Experimental design consisted of general methods (the serial microdilution broth assay and the agar well diffusion assay), as well as observing bactericidal/bacteriostatic activity through time (the “time-kill” assay), investigating the effect on cell wall integrity and biofilm formation, and modulation of bacterial hemolysis. Observed antibacterial activity from above-described methods led to further activity-guided fractionation of water and methanol extracts using bioautography coupled with UHPLC-LTQ OrbiTrap MS⁴. It was determined that active fractions are predominantly formed by gallic acid derivatives and flavonol glycosides. Among the most active phytochemicals, galloyl-shikimic acid was identified as the most abundant compound. These results point to a direct connection between galloyl-shikimic acid and the observed E. cicutarium antibacterial activity, and open several new research approaches for future investigation. Full article

Chemotaxis in the nematode Caenorhabditis elegans has basically been examined using conventional assay methods. Although these can be problematic, for example, in their use of anesthesia, the method has never been improved. We propose a pond assay for the sensory systems (PASS) of C. elegans as a novel population-based method of behavioral analysis. The test solution is injected into a recess(es) formed on agar and the response of C. elegans to its odor and/or taste is examined. Once C. elegans individuals fall into recesses (ponds) filled with liquid, they cannot return to a solid medium. In this way, the animals are trapped with certainty without the use of anesthesia. The anesthesia used to keep animals in the attractant area in conventional chemotaxis assays is no longer required, allowing pure evaluation of the attractant or repellent response to specific substances. Furthermore, the assay itself can be greatly streamlined because the preparation can be completed simply by providing a recess(es) and filling the liquid. The present paper reports the detailed method and effectiveness of the novel PASS. Full article

Nontuberculous mycobacteria (NTM) are ubiquitous microorganisms naturally resistant to antibiotics and disinfectants that can colonize drinking water supply systems. Information regarding the spread of NTM in specifically South America and Colombia is limited. We aimed to identify and characterize NTM present in tap water samples from Cali, Colombia. Drinking water samples and faucet biofilm swabs were collected in 18 places, including the city’s three main water treatment plants (WTPs). Filter-trapped material and eluates (0.45 μm) from swab washes were plated in 7H11 agar plates. Suspected colonies were evaluated microscopically, and NTM species were identified based on the rpoB gene. Antibiotic susceptibility testing was also performed. Fifty percent (9/18) of sampling points were positive for NTM (including two WTPs), from which 16 different isolates were identified: Mycobacterium mucogenicum (8/16), M. phocaicum (3/16), M. chelonae (2/16), M. mageritense (2/16), and M. fortuitum (1/16), all rapidly growing mycobacteria. A susceptibility profile was obtained from 68.75% (11/16) of the isolates. M. chelonae was the most resistant species. All NTM isolated are potentially responsible for human diseases; our findings might provide a baseline for exploring NTM transmission dynamics and clinical characterization, as well as potential associations between NTM species found in drinking water and isolates from patients. Full article

Fusarium head blight (FHB) is a devastating disease of wheat. Worldwide, Fusarium graminearum is the most dominant FHB-causing species. Its most common toxin, deoxynivalenol (DON), impairs food and feed safety and has an enormous economic impact. Agronomic factors such as crop rotation, soil management and host genotype strongly influence the occurrence of F. graminearum. Infected plant debris from previous crops, on which perithecia and ascospores develop, represent the main source for FHB, and hence, improved cropping systems aim to reduce this inoculum to decrease the infection risk. The best measure to evaluate the disease pressure is spore traps that detect deposited airborne ascospores. Commercial spore traps are expensive and require power sources, thus, they are not suitable for investigations in field experiments with different treatments. In consequence, we developed spore traps containing a Petri dish with Fusarium-selective agar, protected by aluminum dishes and attached on a wooden board. We compared the data of our low-cost trap with those of a commercial high-throughput jet sampler and obtained equivalent results. In field experiments to compare cropping systems, we observed a high correlation between the DON content in wheat grains and the number of colonies from deposited spores. Our spore trap proved to be a highly valuable tool to not only study FHB epidemiology but also to identify innovative cropping systems with a lower risk for FHB and DON contamination. Full article

When screening microbial populations or consortia for interesting cells, their selective retrieval for further study can be of great interest. To this end, traditional fluorescence activated cell sorting (FACS) and optical tweezers (OT) enabled methods have typically been used. However, the former, although allowing cell sorting, fails to track dynamic cell behavior, while the latter has been limited to complex channel-based microfluidic platforms. In this study, digital microfluidics (DMF) was integrated with OT for selective trapping, relocation, and further proliferation of single bacterial cells, while offering continuous imaging of cells to evaluate dynamic cell behavior. To enable this, magnetic beads coated with Salmonella Typhimurium-targeting antibodies were seeded in the microwell array of the DMF platform, and used to capture single cells of a fluorescent S. Typhimurium population. Next, OT were used to select a bead with a bacterium of interest, based on its fluorescent expression, and to relocate this bead to a different microwell on the same or different array. Using an agar patch affixed on top, the relocated bacterium was subsequently allowed to proliferate. Our OT-integrated DMF platform thus successfully enabled selective trapping, retrieval, relocation, and proliferation of bacteria of interest at single-cell level, thereby enabling their downstream analysis. Full article

Hydrogen sulfide (H₂S) is the most recently established gaseous vasodilator, enzymatically produced from cysteine metabolism, involved in a number of pathophysiological processes. However, its accurate detection in vivo is critical due to its volatility and tendency to form sulfane sulfur derivatives, thus limiting the data interpretation of its biological roles. We developed new applications of the simple and rapid method to measure H₂S release in cell culture systems, based on the lead acetate strip test. This test, previously prevalently used in microbiology, was compared with the agar trap method, applied, in parallel, on both cell cultures and cell-free samples. Sulfane sulfur represents the major species derived from intracellular H₂S. Various fluorescent probes are available for quantitation of H₂S derivatives intracellularly. We present here an alternative to the classic imaging method for sulfane sulfur evaluation, running on a flow cytometer, based on SSP4 probe labeling. Flow cytometry turned out to be more direct, fully quantitative and less time-consuming compared to microscopy and more precise with respect to the fluorescence multi-plate reader assay. The new application methods for H₂S determination appear to be fully suitable for the analysis of H₂S release and sulfane sulfur content in biological samples. Full article