Number Of Bacterial Cells Research Articles

Nanocomposite complex ZnO in combination with a triazoloazepinium derivative for inhibition of microbial steel corrosion

We have established the possibility of using ZnO nanoparticles to inhibit microbial corrosion with simultaneous inhibition of growth and sulfate-reducing activity of bacteria of the corrosive active group. The search for ways to increase the antibacterial and anticorrosive effect of ZnO nanoparticles makes it possible to combine them with substances, in particular, inhibitors-biocides of microbial steel corrosion. The nanocomposite complex of ZnO nanoparticles and cationic heterocyclic compound (triazoloazepinium derivative) was studied as a biocide and inhibitor of microbial corrosion on steel. The component concentrations are 3 mg/mL and 1 mg/mL accordingly. The experiments were carried out by using a microbiological and corrosion control methods. It has been established that the proposed nanocomposite complex affects the number of bacteria in the plankton and biofilm. In its presence, there is a complete inhibition of the growth of sulfate-reducing bacteria (the most aggressive component of the microbial community) in plankton. Also, the number of denitrifying bacteria and iron-reducing bacteria in plankton decreases by 3 and 4 orders, respectively. The biofilm formed in the presence of the nanocomposite in the culture medium is less dense in terms of the number of bacterial cells of the sulfidogenic community.

Effects of Phenolic Plant Extracts on Biofilm Formation by Klebsiella pneumoniae Isolated from Urinary Tract Infections

Ten isolates of Klebsiella pneumoniae, seven isolates of Pseudomonas aeruginosa and nine isolates of Staphylococcus aureus, were obtained from 100 urine samples collected from Baghdad hospitals. All isolates were identified biochemically and confirmed by using VITEK 2 and were then tested for their susceptibility towards 6 antibiotics and for phenolic extracts of Thymus vulgaris and Cinnamomum cassia. All bacteria were greatly affected by T. vulgaris, especially K. pneumoniae. Viable count was performed, it was noted that the number of bacterial cells reduced from 1×108 CFU to 1.2× 103, 2×105 and 1.8×106CFU of K. pneumoniae, P. aeruginosa and S. aureus respectively. While C. cassiahad a slight effect on them. K. pneumoniae isolates which were affected by phenolic extract more than the other bacteria under study and at the same time were resistant to more than one type of tested antibiotics. These isolates were taken to detect their ability to form biofilm by using Congo red as screening method for it. The results showed that all isolates produced biofilms. Also, by using microtiter plate method, the results confirmed that all isolates produced biofilm where 7 isolates were strong biofilm producers and 3 were moderate. The strongest isolate was taken to study the effect of T. vulgaris and C. cassia phenolic extract on its biofilm formation by using microtiter plate method with two concentrations (20 and 40 ml/L). The results showed that biofilm reduction was 45% and 73% for T. vulgaris and that for C. cassiait was 15% and 20% after using 20 and 40 ml/L respectively.

Modification of Antibacterial Copolymers on the Surface of PVA-Based Microfibers via Thermal Cross-Linking and Their Antibacterial Properties.

Bacterial infections on material surfaces are a serious public health concern worldwide. Although poly(vinyl alcohol) (PVA)-based materials have great potential as medical devices, they lack antibacterial properties on their surfaces and pose bacterial infection risks during implantation surgery. Copolymers containing antibacterial [2-(methacryloyloxy)ethyl]trimethylammonium chloride (METAC) units were used to modify the surfaces of chemically cross-linked water-insoluble PVA-based microfibers. The copolymers also had carboxy units that were used to react with the hydroxy group of the PVA-based microfibers via a simple thermal treatment at 135 °C. PVA-based materials containing METAC units exhibit significant swelling due to electrostatic repulsions. Because the copolymers were modified on the extreme surface of the microfibers, no difference in the diameters between unmodified microfibers (PM-fiber) and copolymers with METAC unit-modified microfibers (PM-METAC-fiber), in both the dry and swollen states, was observed. The viable bacterial cell numbers, which were evaluated by colony counting, decreased by exposure to the poly(METAC-co-methacrylic acid (MAAc)) aqueous solution or PM-METAC-fibers. The value of CFU/mL decreased to 0.1% (against B. subtilis) and 3.9% (against E. coli) after contact with the PM-METAC-fibers compared to the PM-fibers. The percentage of hemolysis against rabbit red blood cells was equivalent to that of the negative control, suggesting that PM-METAC-fibers can selectively exhibit antibacterial properties. This modification method can be applied to various PVA-based materials if hydroxy groups are present on their surface. This study provides a facile, cost-effective, and promising strategy to impart antibacterial properties to the surface of PVA-based materials without significantly affecting their physicochemical properties.

Calcium sorption and isotope fractionation in Bacillus subtilis and Pseudomonas aeruginosa

Bacteria are a key component of the critical zone, because of their role in the nutrient availability for the vegetation. There is still little knowledge on the direct role of bacteria on Ca storage/leaching in soils while it is an essential macronutrient for vegetation growth. In recent years, Ca stable isotopes have shown their potential in understanding the Ca biogeochemical cycle. Preliminary studies highlighted that in the presence of soil bacteria, the plant uptake of nutrients is increased due to the mineral bioweathering. Moreover, Ca isotope signatures of nutrient media also showed differences between growth experiments in batch in the presence and absence of bacteria. In this study, the focus is to verify if Ca adsorption and incorporation into/onto bacterial strains induce such isotopic fractionation. Batch experiments were carried out on Pseudomonas aeruginosa (a Gram-negative bacterium) and on the vegetative and sporulated forms of Bacillus subtilis (a Gram-positive bacterium). These experimentations showed that: (i) no observable isotopic fractionations were induced during calcium/bacteria contact for all experimental parameters (pH, kinetic, bacterial cell number, interaction time, dead/alive bacteria); (ii) Ca was mainly stored in the bacterial cell wall compartments. On the other hand, significant Ca isotopic differences between the spores and the sporulation medium (Δ44/40Caspores–sporulation medium ranging from − 0.53 to − 1.15‰), suggest isotopic fractionation during the sporulation process, likely occurring during the attachment of Ca to carboxyl acid groups as calcium chelates with dipicolinic acid. The absence of Ca isotope fractionation during Ca sorption on vegetative and sporulated bacteria via passive channels indicates that the tested bacteria’s contribution to the Ca biogeochemical cycle is indirect primary enhancing bioweathering and Ca bioavailability for vegetation. If confirmed by further studies, only the sporulation mechanisms itself may directly impact the Ca biogeochemical cycle.

Role of the SaeRS Two-Component Regulatory System in Group B Streptococcus Biofilm Formation on Human Fibrinogen.

Streptococcus agalactiae, also known as Group B Streptococcus or GBS, is a commensal colonizer of human vaginal and gastrointestinal tracts that can also be a deadly pathogen for newborns, pregnant women, and the elderly. The SaeRS two-component regulatory system (TCS) positively regulates the expression of two GBS adhesins genes, but its role in the formation of biofilm, an important step in pathogenesis, has not been investigated. In the present study, we set up a novel model of GBS biofilm formation using surfaces coated with human fibrinogen (hFg). Biofilm mass and structure were analyzed by crystal violet staining and three-dimensional fluorescence microscopy, respectively. GBS growth on hFg resulted in the formation of a mature and abundant biofilm composed of bacterial cells and an extracellular matrix containing polysaccharides, proteins, and extracellular DNA (eDNA). Enzymatic and genetic analysis showed that GBS biofilm formation on hFg is dependent on proteins and eDNA in the extracellular matrix and on the presence of covalently linked cell wall proteins on the bacterial surface but not on the type-specific capsular polysaccharide. In the absence of the SaeR regulator of the SaeRS TCS, there was a significant reduction in biomass formation, with reduced numbers of bacterial cells, reduced eDNA content, and disruption of the biofilm architecture. Overall, our data suggest that GBS binding to hFg contributes to biofilm formation and that the SaeRS TCS plays an important role in this process.

Serine affects engulfment during the sporulation process in Clostridium perfringens strain SM101

ObjectivesAlthough Clostridium perfringens sporulation is a key event in the pathogenesis of food-borne illness, the molecules and underlying mechanisms responsible for regulating sporulation are incompletely understood. The present study sought to identify amino acids that affect sporulation in C. perfringens strain SM101. MethodsA C. perfringens strain was cultured in the chemically defined medium deficient in an amino acid. The bacterial growth was determined by spectrophotometrically measuring culture turbidity and by calculating colony-forming unit. Morphological characteristics were assessed by phase-contrast microscopy with fluorescent staining and by electron microscopy. ResultsThe amino acids Arg, Cys, Gly, His, Ile, Leu, Met, Phe, Thr, Trp, Tyr, and Val were important for sporulation, and furthermore, Ser reduced sporulation. The mechanism underlying Ser-induced prevention of sporulation was assessed morphologically. The numbers of bacterial cells in sporulation stage II were significantly higher in the presence than in the absence of Ser. In the presence of Ser, almost all cells were in stage II−III, characterized by polar septation−early engulfment, and did not proceed to late engulfment. ConclusionsThese results suggest that Ser accelerated the early stage of sporulation of C. perfringens strain SM101, but disturbed the engulfment process, resulting in reduction of sporulation. To the best of our knowledge, this is the first study reporting that an amino acid affects engulfment during the C. perfringens sporulation process.

Lag Plot: A Novel Method to Determine Viable Bacterial Cell Number via a Single OD Measurement.

Bactericidal activity is a valuable parameter which is considered and measured for antimicrobial compounds. The available standard protocol to evaluate bactericidal activity is based on the direct colony count. Colony counting requires serial dilution, plating, overnight incubation, and direct counting, which is time-and labor-intensive. In regard to eliminate direct plate count, novel techniques were developed based on the real-time growth monitoring which can come with some limitations and drawbacks. These drawbacks encourage us to develop a novel technique with simple procedure to determine viable bacterial cell count. In this procedure, real-time growth monitoring is not required. In fact, after an incubation time, the number of viable bacteria can be determined with a single OD measurement and through an equation. In this regard, four standard bacterial strains, including Staphylococcus aureus (ATCC 25923), Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC 27853), and Bacillus subtilis (ATCC 23857), were cultured with descending inoculum densities (1.5 × 107 to 15CFU/mL) and growth curves were drawn. As expected, growth in the samples with lower inoculum densities recorded with longer lag phase. Also, a direct relation was observed between the recorded turbidities and initial cell counts. A logarithmic curve (lag plot) was obtained by plotting the OD, after 12-18h incubation, against initial cell counts. In all examined strains, R2 was calculated in the range of 0.96-0.99 which is acceptable value for coefficient of determination. Equation corresponding to the lag plot was obtained and called lag equation. This equation is applicable to calculate the number of viable cells in unknown samples simply by inoculation, incubation, and single OD measurement. Developed technique can be introduced as a simple substitution for labor- and time-intensive direct colony counting.

Carbon-carbon composite material as a potential basis for orthopedic implants

Aim. To determine the cytocompatibility of carbon-carbon composite materials (CCCM) and assess their ability to be impregnated with vancomycin.Materials and Methods. The study included samples of carbon-carbon composite materials (CCCM). The cytocompatibility of CCCM blocks was evaluated using a culture of eukaryotic cells (Vero cell line). Biofilms of S. aureus ATCC 29213 (MSSA), S. aureus ATCC 43300 (MRSA), S. epidermidis ATCC 12228 (MSSE), and S. epidermidis ATCC 29887 (MRSE) were formed by immersing sterile test samples of CCCM into a nutrient medium which contained bacteria. After 24-hour incubation, the samples were washed, placed in an ultrasonic bath, and sonication fluid was inoculated using the sector method. To saturate the CCCM blocks with antibiotics, they were placed into a vancomycin solution and then lyophilized under negative pressure with gradual heating. The antimicrobial activity of the resulting blocks was studied using the cup plate method against the same reference cultures of staphylococci. The dynamics of vancomycin elution from CCCM was investigated using high-performance liquid chromatography.Results. Vero cells maintained their viability in the presence of the tested material. Considering the highly porous structure of CCCM and variable diameter of the pores, we suggested a good osteointegration potential of this material. On the samples without an impregnation with an antibacterial drug, reference strains of staphylococci were able to form a biofilm with a sufficient number of bacterial cells to initiate an infectious process. The duration of antimicrobial activity of the antibioticim-pregnated samples against the reference staphylococcal strains was up to 3 days. The majority of the antibiotic eluted from the CCCM into the incubation medium during the first two days.Conclusion. The cytocompatibility and porosity of CCCM in combination with a vancomycin impregnation makes this material promising for the fabrication of implants with antimicrobial activity as well as tissue engineering constructs.

Immobilized consortium of heterotrophic nitrifying-aerobic denitrifying bacteria on various matrices for nitrogen removal from synthetic wastewater

Environmental contamination by nitrogen compounds such as ammonium and nitrate has increased extensively in the recent past, which necessitates the development of eco-friendly remediation technologies. In this study, three matrix types including pumice, aquarium ceramic filter, and calcium alginate beads were used to facilitate nitrogen removal with an immobilized heterotrophic nitrifying-aerobic denitrifying (HNAD) bacterial consortium. The HNAD bacterial consortium was made of Pseudomonas monteilii Nht, Pseudomonas mendocina AquaN, Rhodococcus erythropolis R1, and Acinetobacter calcoaceticus SCC2. The quality parameters for immobilization, such as the number of immobilized cells and their viability, were assessed. The highest number of bacterial cells (3.4 × 10 9) was immobilized on the aquarium ceramic filter, with 53% cell viability at 30°Ⅽ for two months. Pumice, aquarium ceramic filter, and calcium alginate achieved NH4+-N removal efficiencies of 85.3 ± 1.7%, 87.3 ± 2.2%, and 77.5 ± 3.99% within 24 h, respectively, and removed NO3−-N by 88.23 ± 0.36%, 93.95 ± 0.00%, and 71.29 ± 6.49% over 60 h. Additionally, immobilized cells on pumice and ceramic filter retained up to 84% of NH4+-N removal efficiency after 14 reuse cycles. These findings indicate that the immobilized HNAD bacterial consortium on the aquarium ceramic filter can be used as a suitable biofilter for treatment of high nitrogen wastewater.

Nascently generated microplastics in freshwater stream are colonized by bacterial communities from stream and riparian sources.

The purpose of this study was to examine bacterial colonization of different types of microplastics through time in a freshwater ecosystem. Microplastics are persistent pollutants in aquatic ecosystems. Bacteria readily colonize microplastic surfaces and may contribute to their degradation, but the taxa involved, and their degradative abilities, differ based on factors such as microplastic chemistry, plastic age, and specific ecosystem types. Four different common types of newly manufactured microplastics, high-density polyethylene, low-density polyethylene, polypropylene, and polystyrene, were incubated for 7 weeks in a freshwater stream and sampled. Sample collection was timed to examine the development of early and late bacterial biofilm communities. Microplastics were analyzed for changes to buoyancy, weight, contact angles (an indicator of surface roughness), bacterial community composition, and the number of bacterial cells. Time was the only significant contributing factor in the development of bacterial biofilm communities on microplastic disks over the 7-week study. Notably, the Comamonadaceae were abundant early in the study and decreased in abundance with time, while the Methylococcaceae demonstrated the opposite trend. Different physicochemical properties among the various types of microplastics had only a minor effect on bacterial community compositions of biofilms growing on the microplastics. Additionally, the surfaces of all microplastic disks became rougher over time in the stream. Collectively, our results show that microplastic surfaces undergo surface modification and community succession as time progresses, regardless of microplastic type, in a freshwater stream ecosystem.

Photocatalytic Degradation of Paracetamol and Antibacterial Activity of La-Modified TiO2 Obtained by Non-Hydrolytic Sol–Gel Route

The current study aims to synthesize and analyze both pure and La-doped TiO2, and evaluate the photocatalytic and antibacterial activity of as-prepared samples. Doped and undoped samples were prepared by the non-hydrolytic sol–gel method from titanium(IV) chloride, benzyl alcohol, and lanthanum(III) nitrate followed by thermal treatment. Lanthanum content in synthesized samples was 0.4, 1, and 5 mol%. The resulting nanopowders’ structure and morphology were described using XRD, IR, and UV–Vis analysis. The average particle sizes of pure and doped TiO2 were about 6–15 nm and anatase was found to be a dominant crystalline phase in the samples. It was observed that particle sizes decreased on increasing La content. The photocatalytic activity of the pure and La-doped sol–gel powders was estimated in the decomposition of paracetamol in distilled water using ultraviolet light illumination. Doping with lanthanum ions has been shown to increase the photocatalytic properties on the degradation of paracetamol. Furthermore, the annealed catalysts (pure and La3+ doped) showed increased photocatalytic activity and degradation of the analgesic in comparison with non-annealed materials. In both cases, the highest photocatalytic efficiency is observed at the optimal La3+ (1 mol%) concentration. The antimicrobial activity of 1 mol% La/TiO2 was tested against a reference strain E. coli in the presence of ultraviolet light and in dark conditions. The number of viable bacterial cells was determined by a spread plate method, and kill curves were performed. The results showed that photoactivated 1 mol% La/TiO2 exhibited a strong bactericidal effect, and in concentration, 1 mg/mL efficiently killed bacteria at an initial cell density of about 105 colony forming units in 1 mL within 15 min.

IN VIVO STUDY EVALUATES EFFECTIVENESS OF ADDING CIPROFLOXACIN TO CALCIUM HYDROXIDE VERSUS SAUSSUREA COSTUS TO CALCIUM HYDROXIDE MEDICATION AGAINST BACTERIAL BIOFILM IN ROOT CANAL TREATMENT

Background and objectives: Elimination of root canal bacteria is a perquisite for a successful root canal treatment. This research aims to determine the effectiveness of adding ciprofloxacin versus Saussurea costus extract to calcium hydroxide medicament against bacterial biofilm in root canal treatment in vivo study. Materials and methods: In vivo study on 70 single-rooted human teeth at Sana'a University dental clinics used Saussurea costus extract preparation, including washing, drying, crushing, mixing, centrifugation, drying, and homogenization.The intracanal drug concentration was standardized by mixing 1 g of calcium hydroxide powder with 1 ml of ciprofloxacin or 1 ml of Saussurea costus extract in both groups. Three bacteriological samples were taken before root canal treatment, before Medicaments were placed, and 7 days after, and CFU and bacterial isolation were performed using standard methods. Results: An intragroup analysis showed that all drug groups had a significant decrease in the number of intracanal bacterial cells from S1 to S2 and from S2 to S3 (p<0.0001). When comparing quantitative S1 or S2 data, there was no significant difference between the groups; however, at Sample 3, the Saussurea costus extract calcium hydroxide (34.05±21.8) had considerably higher bacterial counts than the calcium hydroxide + Ciprofloxacin group (11.9±29 CFU). The most isolated bacteria in S1 and S2 were Staphylococcus aureus (58.6%, 51.4%), followed by Klebsiella species (27.1%, 18.6%), Escherichia coli (25.7%, 17.1%), Enterococcus faecalis (22.9%, 15.7%) and Pseudomonas aeruginosa (21.4%, 12.9%). Conclusions: The addition of Ciprofloxacin to calcium hydroxide and Saussurea costus- calcium hydroxide provided further antibacterial effectiveness when used as an intracanal medicament in vivo during root canal treatment. Peer Review History: Received 27 March 2024; Revised 5 May 2024; Accepted 28 June; Available online 15 July 2024 Academic Editor: Essam Mohamed Eissa, Beni-Suef University, Egypt, dressamceutics@yahoo.com Average Peer review marks at initial stage: 6.5/10 Average Peer review marks at publication stage: 7.0/10 Reviewers: Dr. Kingsley C Anukam, University of Benin, Nigeria, kanukam@gmail.com Dr. Liliya Logoyda, Horbachevsky Ternopil State Medical University, Ukraine, logojda@tdmu.edu.ua

Enhancing heavy metal phytoremediation in landfill soil by Chrysopogon zizanioides (L.) roberty through the application of bacterial-biochar pellets

This work investigated the performance of bacterial-biochar pellets (BBP) on promoting heavy metal phytoremediation by Chrysopogon zizanioides (L.) Roberty planted in landfill soil. Micrococcus sp. MU1, an indole-3-acetic acid (IAA)-producing bacterium, was incorporated into biochar pellets (MU1-BBP). MU1-BBP contained a high number of viable bacterial cells with 90 % viability after storage at 4°C for two months. In addition, it had a greater survival capability in heavy metals-contaminated landfill soil than MU1-free cells. The application of MU1-BBP significantly promoted the growth performance and chlorophyll synthesis of C. zizanioides planted in landfill soil. Cd, Cu, Pb and Zn were mostly retained in the roots relative to the shoots of C. zizanioides. The accumulation of Cd, Cu, Pb, and Zn in the roots of C. zizanioides were significantly boosted by MU1-BBP inoculation. Plants with MU1-BBP inoculation enhanced the accumulation coefficient (AC) of all four heavy metals. The values of AC values of Cd, Cu, and Zn in C. zizanioides with MU1-BBP inoculation were greater than 1.0. However, translocation factors of all four metals were low. The findings suggest C. zizanioides may be a good heavy metal phytostabilizer. The findings support the potential of bacterial-biochar pellets to enhance the efficiency of heavy metal phytostabilization of C. zizanioides in landfill sites.

Preliminary Study: Antimicrobial Permeation and Release Rates of Biocellulose Wound Dressing Produced Using Acetobacter Xylinum 0416 in Pineapple Peel Waste Culture

Modern wound dressings are employed to hasten the healing of damaged skin. Alginate, foam, hydrogel, hydrocolloid and film are contemporary wound dressings. These contemporary wound dressings are effective for treating wounds because they keep moisture in the wound to help the skin of the damaged tissue heal. Another application for biocellulose (BC) is as a material for treating wounds. This is because biocellulose can absorb exudates from skin wounds and has a considerable water-holding capacity. Because pineapple peel waste has a high concentration of glucose and fructose, it can be used as a fermentation medium to make biocellulose. The manufacture of biocellulose also makes use of the bacterium Acetobacter xylinum 0416. The biocellulose fermentation process lasts five days at 30 °C in an incubator with static conditions. Utilising antimicrobial agents such as silver nanoparticle compounds, Moringa oleifera, or Melastoma malabathricum leaf extract will improve the antibacterial capabilities of biocellulose. The growth of A. xylinum 0416 indicates an exponential phase at 48 hours, where the number of bacterial cells increases, and the process of transferring 20% of the inoculum to the fermentation medium is carried out. BC fermentation in 6-well plates with 3 ml of pineapple peel extract volume at 30°C and pH 5.15 recorded the highest BC wet weight reading at 1.740g and an average of 1.393g. Low concentrations of silver nanoparticles and Moringa leaf extract at 0.02mg/ml, 5mg/l and 0.0125mg/l showed a good effect against the pathogen. The water absorption capacity of the raw BC shows a good absorption value of 144.84.

Selection versus transmission: Quantitative and organismic biology in antibiotic resistance

We aimed to determine the importance of selection (mostly dependent on the anthropogenic use of antimicrobials) and transmission (mostly dependent on hygiene and sanitation) as drivers of the spread of antibiotic-resistant bacterial populations. The first obstacle to estimating the relative weight of both independent variables is the lack of detailed quantitative data concerning the number of bacterial cells, potentially either pathogenic or harmless, and bacterial species exposed to antimicrobial action in the microbiotas of specific environments. The second obstacle is the difficulty of considering the relative importance of the transmission and selection exerting their combined effects on antibiotic resistance across eco-biological levels. As a consequence, advances are urgently required in quantitative biology and organismic biology of antimicrobial resistance. The absolute number of humans exposed to antibiotics and the absolute number of potentially pathogenic and commensal bacteria in their microbiomes should influence both the selection and transmission of resistant bacterial populations. The “whole Earth” microbiome, with astonishingly high numbers of bacterial cells and species, which are also exposed to anthropogenic antimicrobials in various biogeographical spaces, shapes the antibiotic resistance landscape. These biogeographical spaces influence various intensities of selection and transmission of potentially pathogenic bacteria. While waiting for more precise data, biostatistics analysis and mathematical or computational modeling can provide proxies to compare the influence of selection and transmission in resistant bacteria. In European countries with lower sanitation levels, antibiotic consumption plays a major role in increasing antibiotic resistance; however, this is not the case in countries with high sanitation levels. Although both independent variables are linked, their relative influence on the level of antibiotic resistance varies according to the particular location. Therefore, interventions directed to decrease antibiotic resistance should be designed “a la carte” for specific locations with particular ecological conditions, including sanitation facilities.

Quantitative analysis of the effects of brushing, flossing, and mouthrinsing on supragingival and subgingival plaque microbiota: 12-week clinical trial

BackgroundTranslational microbiome research using next-generation DNA sequencing is challenging due to the semi-qualitative nature of relative abundance data. A novel method for quantitative analysis was applied in this 12-week clinical trial to understand the mechanical vs. chemotherapeutic actions of brushing, flossing, and mouthrinsing against the supragingival dental plaque microbiome. Enumeration of viable bacteria using vPCR was also applied on supragingival plaque for validation and on subgingival plaque to evaluate interventional effects below the gingival margin.MethodsSubjects with gingivitis were enrolled in a single center, examiner-blind, virtually supervised, parallel group controlled clinical trial. Subjects with gingivitis were randomized into brushing only (B); brushing and flossing (BF); brushing and rinsing with Listerine® Cool Mint® Antiseptic (BA); brushing and rinsing with Listerine® Cool Mint® Zero (BZ); or brushing, flossing, and rinsing with Listerine® Cool Mint® Zero (BFZ). All subjects brushed twice daily for 1 min with a sodium monofluorophosphate toothpaste and a soft-bristled toothbrush. Subjects who flossed used unflavored waxed dental floss once daily. Subjects assigned to mouthrinses rinsed twice daily. Plaque specimens were collected at the baseline visit and after 4 and 12 weeks of intervention. Bacterial cell number quantification was achieved by adding reference amounts of DNA controls to plaque samples prior to DNA extraction, followed by shallow shotgun metagenome sequencing.Results286 subjects completed the trial. The metagenomic data for supragingival plaque showed significant reductions in Shannon-Weaver diversity, species richness, and total and categorical bacterial abundances (commensal, gingivitis, and malodor) after 4 and 12 weeks for the BA, BZ, and BFZ groups compared to the B group, while no significant differences were observed between the B and BF groups. Supragingival plaque vPCR further validated these results, and subgingival plaque vPCR demonstrated significant efficacy for the BFZ intervention only.ConclusionsThis publication reports on a successful application of a quantitative method of microbiome analysis in a clinical trial demonstrating the sustained and superior efficacy of essential oil mouthrinses at controlling dental plaque compared to mechanical methods. The quantitative microbiological data in this trial also reinforce the safety and mechanism of action of EO mouthrinses against plaque microbial ecology and highlights the importance of elevating EO mouthrinsing as an integral part of an oral hygiene regimen.Trial registrationThe trial was registered on ClinicalTrials.gov on 31/10/2022. The registration number is NCT05600231.

The use of stem cells in the treatment of mastitis in dairy cows

Mastitis is a multifactorial inflammatory disease. The increase in antibiotic resistance of bacteria that cause mastitis means that cattle breeders would prefer to reduce the use of antibiotics. Recently, therapies using mesenchymal stem cells (MSCs) from various sources have gained significant interest in the development of regenerative medicine in humans and animals, due to their extraordinary range of properties and functions. The aim of this study was to analyze the effectiveness of an allogeneic stem cells derived from bone marrow (BMSC) and adipose tissue (ADSC) in treating mastitis in dairy cattle. The research material consisted of milk and blood samples collected from 39 Polish Holstein-Friesian cows, 36 of which were classified as having mastitis, based on cytological evaluation of their milk. The experimental group was divided into subgroups according to the method of MSC administration: intravenous, intramammary, and intravenous + intramammary, and according to the allogeneic stem cells administered: BMSC and ADSC. The research material was collected at several time intervals: before the administration of stem cells, after 24 and 72 h, and after 7 days. Blood samples were collected to assess hematological parameters and the level of pro-inflammatory cytokines, while the milk samples were used for microbiological assessment and to determine the somatic cells count (SCC). The administration of allogeneic MSCs resulted in a reduction in the total number of bacterial cells, Staphylococcus aureus, bacteria from the Enterobacteriaceae group, and a systematic decrease in SCC in milk. The therapeutic effect was achieved via intravenous + intramammary or intramammary administration.

Effects of Bifidobacterium animalis subsp. lactis BB-12 and yogurt on mice during oral antibiotic administration

Probiotics have the potential to prevent disruptions to normal gastrointestinal function caused by oral antibiotic use. In this study, we examined the capacity of Bifidobacterium animalis subspecies lactis BB-12 (BB-12) and yogurt, separately and combined, to mitigate the effects of the antibiotic amoxicillin-clavulanate (AMC) on the gut microbiota and metabolomes of C57BL/6 J mice. Male and female mice were administered either BB-12, yogurt, BB-12 in yogurt, or saline for 10 days concurrent with the inclusion of AMC in the drinking water. Male mice exposed to AMC exhibited significant reductions (p<0.05) in body weight over the course of the study compared to sham (no AMC) controls whereas no such effects were observed for female mice. AMC administration resulted in rapid alterations to the intestinal microbiota in both sexes irrespective of BB-12 or yogurt treatment, including significant (p<0.05) losses in bacterial cell numbers and changes in microbial alpha-diversity and beta-diversity in the feces and cecal contents. The effects of AMC on the gut microbiota were observed within one day of administration and the bacterial contents continued to change over time, showing a succession marked by rapid reductions in Muribaculaceae and Lachnospiraceae and temporal increases in proportions of Acholeplasmataceae (day 1) and Streptococcaceae and Leuconostocaceae (day 5). By day 10 of AMC intake, high proportions of Gammaproteobacteria assigned as Erwiniaceae or Enterobacteriaceae (average of 63 %), were contained in the stools and were similarly enriched in the cecum. The cecal contents of mice given AMC harbored significantly reduced concentrations of (branched) short-chain fatty acids (SCFA), aspartate, and other compounds, whereas numerous metabolites, including formate, lactate, and several amino acids and amino acid derivatives were significantly enriched. Despite the extensive impact of AMC, starting at day 7 of the study, the body weights of male mice given yogurt or BB-12 (in saline) with AMC were similar to the healthy controls. BB-12 (in saline) and yogurt intake was associated with increased Streptococcaceae and both yogurt and BB-12 resulted in lower proportions of Erwiniaceae in the fecal and cecal contents. The cecal contents of mice fed BB-12 in yogurt contained levels of formate, glycine, and glutamine that were equivalent to the sham controls. These findings highlight the potential of BB-12 and yogurt to mitigate antibiotic-induced gut dysbiosis.

IM156, a new AMPK activator, ameliorates polymicrobial sepsis by regulating inflammatory responses

Abstract IM156, a biguanide with a higher potency of AMP-activated protein kinase (AMPK) activation than metformin, exhibits inhibitory activity against angiogenesis and cancer. In this study, we explored the anti-inflammatory effects of IM156 against polymicrobial sepsis. Administration of IM156 significantly improved the survival rate and tissue damage in cecal ligation and puncture (CLP)-induced sepsis. Mechanistically, IM156 markedly reduced the number of bacterial cells in the peritoneal fluid and peripheral blood in a CLP-induced sepsis model. IM156 also inhibited splenocyte apoptosis and the production of inflammatory cytokines, including IL-1β, IL-6, and IL-10, in CLP mice. Furthermore, IM156 strongly inhibited the reactive oxygen species (ROS) generation by enhancing antioxidative effects, which can lead to reduced neutrophil extracellular trap (NET) formation in response to lipopolysaccharide. In conclusion, these findings collectively indicate that IM156 can suppress the inflammatory response and provide protection against polymicrobial sepsis. These results suggest the potential of IM156 as a novel therapeutic agent for sepsis control.

Biofilm Production and Porin Permeability Activity in Clinical Isolates of Ciprofloxacin-Resistant Klebsiella pneumoniae in the Tertiary Hospital in Purwokerto, Indonesia

World Health Organization reports that the current use of ciprofloxacin as a broad-spectrum antibiotic shows a pattern of high bacterial resistance in many countries. Klebsiella pneumoniae is known as a Gram-negative bacterium that most often causes infection and is resistant to antibiotics. This study aimed to find patterns of resistance mechanisms based on biofilm production and porin permeability activity in K. pneumoniae clinical specimens from patients at RSUD Prof. Dr Margono Soekardjo Purwokerto, Indonesia. Several isolates of ciprofloxacin-resistant K. pneumoniae were isolated from clinical specimens of blood, sputum, urine, pus, stool and pleural fluid from August to October 2022. Identification and sensitivity testing of K. pneumoniae to ciprofloxacin were performed using Vitek® 2 Compact. A test for biofilm production is carried out by measuring optical density (OD) with a microplate at a wavelength of 630 nm. The porin permeability activity was determined using the value of the minimum inhibitory concentration (MIC) by the LC-MS spectrophotometry method at a wavelength of 630 nm. The results showed that 72 (47 %) isolates were resistant to ciprofloxacin. Among them, 41.3 % (24/58) of ciprofloxacin-resistant K. pneumoniae were able to produce strong biofilm. Porin permeability is indicated by MIC values of 1, 2 and 4 mg/L with bacterial cell counts of 32×106, 19×106 and 34×106 CFU/mL, respectively. This number decreased from the initial control with a bacterial cell count of 10.15×107 CFU/mL. Analysis of the correlation between the significance of biofilm production and the number of bacterial cells indicated that biofilm formation was related to the number of bacterial cells (R = 0.628, p ≤ 0.05). In conclusion, the resistance mechanism of ciprofloxacin-resistant K. pneumoniae involves biofilm production and decreased porin permeability activity. HIGHLIGHTS Major resistance mechanisms in pneumoniae decreased cell permeability from porin activity and biofilm formation Bacterial outer membrane permeability to antimicrobial substances influences inherent resistance. Bacteria produce biofilm to increase their resistance to antibiotics. Porins are non-specific diffusion transport proteins that play a part in the antibiotic resistance process The evolutionary development in the case of antibiotic resistance has an impact on the emergence of understanding in new perspective patterns on how to overcome health problems related to antibiotic resistance GRAPHICAL ABSTRACT