Plate Count Technique Research Articles

Microbicidal Polymer Nanoparticles Containing Clotrimazole for Treatment of Vulvovaginal Candidiasis

Vulvovaginal candidiasis (VVC) alters the innate cervicovaginal immunity, which provides an important barrier against viruses and other infections. The incidence of this disease has not decreased in the last 30 years, so effective treatments are still needed. Nanoparticles (NPs) of cellulose acetate phthalate (CAP) and clotrimazole (CLZ) were prepared by the emulsification-diffusion method. NPs were characterized using dynamic light scattering, atomic force microscopy and differential scanning calorimetry; their release profile was determined by the dialysis bag technique and mucoadhesion was evaluated with the mucin-particle method. The growth inhibition study of Candida albicans was carried out using the plate counting technique. Finally, accelerated physical stability tests of NPs were carried out, both in water and in SVF. The CAP-CLZ NPs had an average diameter of 273.4 nm, a PDI of 0.284, smooth surfaces and spherical shapes. In vitro release of CLZ from the CAP NPs was categorized with the Weibull model as a matrix system in which initial release was rapid and subsequently sustained. The inhibition of C. albicans growth by the CAP-CLZ NPs was greater than that of free CLZ, and the CAP-only NPs had a microbicidal effect on C. albicans. The NPs showed poor mucoadhesiveness, which could lead to studies of their mucopenetration capacities. An accelerated physical stability test revealed the erosion of CAP in aqueous media. A nanoparticulate system was developed and provided sustained release of CLZ, and it combined an antifungal agent with a microbial polymer that exhibited antifungal activity against C. albicans.Graphical

Study on barrier properties, biodegradability and antimicrobial activity of linear low density polyethylene/starch blends

Purpose The present work aims to prepare biocomposites blend based on linear low density polyethylene/ starch without using harmful chemicals to improve the adhesion between two phases. Also, the efficiency of essential oils as green plasticizers and natural antimicrobial agents were evaluated. Design/methodology/approach Barrier properties and biodegradation behavior of linear low density polyethylene/starch (LLDPE/starch) blends plasticized with different essential oils including moringa oleifera and castor oils wereassessed as a comparison with traditional plasticizer such as glycerol. Biodegradation behavior forLLDPE/starch blends was monitored by soil burial test. The composted samples were recovered then washed followed by drying, and weighting samples after 30, 60, and 90 days to assess the change in weight loss. Also, mechanical properties including retention values of tensile strength and elongation at break were measured before and after composting. Furthermore, scanning electron microscope (SEM) was used to evaluate the change in the morphology of the polymeric blends. In addition to, the antimicrobial activity of plasticized LLDPE/starch blends films was evaluated using a standard plate counting technique. Findings The results illustrate that the water vapor transition rate increases from 2.5 g m−2 24 h−1 for LLDPE/5starch to 4.21 g m−2 24 h−1 and 4.43 g m−2 24 h−1 for castor and moringa oleifera respectively. Also, the retained tensile strength values of all blends decrease gradually with increasing composting period. Unplasticized LLDPE/5starch showed highest tensile strength retention of 91.6% compared to the other blends that were 89.61, 88.49 and 86.91 for the plasticized LLDPE/5starch with glycerol, castor and M. oleifera oils respectively. As well as, the presence of essential oils in LLDPE/ starch blends increase the inhibition growth of escherichia coli, candida albicans and staphylococcus aureus. Originality/value The objective of this work is to develop cost-effective and environmentally-friendly methods for preparing biodegradable polymers suitable for packaging applications.

Evaluation of quantification methods to determine photodynamic action on mono- and dual-species bacterial biofilms.

The effect of photodynamic inactivation (PDI) sensitized by 5,10,15,20-tetra(4-N,N,N-trimethylammoniophenyl)porphyrin (TMAP4+) on different components of mono- and dual-species biofilms of Staphylococcus aureus and Escherichia coli was determined by different methods. First, the plate count technique showed that TMAP4+-PDI was more effective on S. aureus than E. coli biofilm. However, crystal violet staining revealed no significant differences between before and after PDI biofilms of both bacteria. On the other hand, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide method indicated a reduction in viable cells as the light exposure time increases in both, mono- and dual-species biofilms. Furthermore, it was determined that as the irradiation time increases, the amount of extracellular polymeric substances present in the biofilms decreased. This effect was presented in both strains and in the mixed biofilm, being more evident in S. aureus mono-specie biofilm. Finally, scanning electron microscopy analysis showed a decrease in the number of cells forming the biofilm after photosensitization treatments. This information makes it possible to determine whether the photodynamic action is based on damage to metabolic activity, extracellular matrix and/or biomass, which may be useful in establishing a fully effective PDI protocol for the treatment of microorganisms growing as biofilms.

Examining the response of Botrytis cinerea to aqueous ozone: inactivation kinetics, structure, physiology, and growth in different strawberries cultivars

ABSTRACT This work aimed to study the kinetics and mechanisms of Botrytis cinerea conidia inactivation by aqueous ozone treatments (2.5 ± 0.8 mg O3 L−1 and 4.2 ± 0.2 mg O3 L−1, 0.5–16 min), using conventional plate count technique, flow cytometry analysis (FCM), and transmission electron microscopy (TEM). The effect of treatments on the mold incidence on two artificially contaminated strawberries cultivars over refrigerated storage (6 ± 1 °C) was also investigated. The inactivation pattern of B. cinerea conidial suspensions depended on initial level of the microorganism, dissolved ozone concentration and treatment time. Non-linear survival curves were obtained and fitted successfully with the Weibullian model. FCM and TEM analysis revealed alterations in the physiological state and ultrastructure of ozonated conidia that depended on the ozone dose. Ozonation (4.4 mg O3L−1, 5–10 min) reduced B. cinerea incidence throughout storage in strawberries of cv. Fortuna, but no effect was found in cv. Aroma.

Breaking the Mold: Towards Rapid and Cost-Effective Microbial Contamination Detection in Paints and Cosmetics Using ATP-Bioluminescence

Traditional culture-based methods, though a “gold standard” for bacterial detection in various industrial sectors, do often not fulfill today’s high requirements regarding rapidity, on-site applicability, and cost-efficiency both during operation and evaluation. Here, the feasibility of using an adenosine triphosphate (ATP)-based assay for determining microbial contaminations in paints and cosmetics was investigated and compared with standard plate count techniques and dipslides. Therefore, we initially determined the level of sensitivity and assessed the accuracy and concordance among the different methods via spiking tests using a mix of frequently abundant bacterial species to simulate microbial contamination. Bioluminescence intensity was linearly proportional to log colony counts over five orders of magnitude (R2 = 0.99), indicating a high level of sensitivity. Overall, the accuracy varied depending on the test specimen, most probably due to matrix-related quenching effects. Although the degree of conformity was consistently higher at target concentrations ≥ 105 CFU·mL−1, microbial contaminations were detectable down to 103 CFU·mL−1, thus meeting the high requirements of various industries. ATP-based results tended to be within an order of magnitude lower than the reference. However, bearing that in mind, the developed assay serves as a rapid, real-time alternative for routine quality control and hygiene monitoring.

Exonuclease-III Assisted the Target Recycling Coupling with Hybridization Chain Reaction for Sensitive mecAGene Analysis by Using PGM.

In the field of neonatal infections nursing, methicillin-resistant Staphylococcus aureus (MRSA) is a major bacterial pathogen. Here, we present a portable biosensor for MRSA detection that is both highly sensitive and portable, owing to its implementation on the personal glucose meter (PGM) platform. The H probe was fixed on the magnetic bead for mecA gene analysis. A blunt 3' terminus appeared in the MBs-H probe when the mecA gene was present. Exonuclease-III (Exo-III) recognized the blunt terminus and cleaved it, freeing the mecA gene and so facilitating target recycling. In the meantime, the remaining H probe-initiated hybridization chain reaction (HCR) led to the desired signal amplification. Portable quantitative detection of mecA gene is possible because PGM can read the quantity of invertase tagged on HCR product. After optimizing several experimental parameters, such as the concentration of Exo-III and incubation time, the constructed sensor is extremely sensitive, with a detection limit of 2 CFU/mL. The results from this sensitive PGM-based sensor are in agreement with those obtained from plate counting methods, suggesting that it can be used to accurately assess the MRSA content in artificial clinical samples. In addition, the PGM sensor can significantly cut down on time spent compared to plate counting techniques. The manufactured sensor provides a promising option for accurate identification of pathogenic bacteria.

Counting and Identifying Probiotics: From a Systematic Comparison of Three Common Methods to Proposing an Appropriate Method for Identification

Background: The plate count technique had traditionally been used for the determination of viability and counting of probiotic bacteria, which had obvious disadvantages. Efficient tools to identify and count probiotics (alone or in combination) have evolved. Objective: This study aimed to compare two methods of counting and identifying probiotics such as Real-time PCR and flow cytometry, with the culture method and suggest an inexpensive method for the diagnosis of probiotics in dairy products. Methods: Electronic databases such as Scopus, PubMed, and Science Direct were systematically searched, identified, screened, and reviewed from June 2001 to December 2022. Results: This study showed that each technology has its strengths, advantages, and disadvantages, but the Real-time PCR method is more suitable than other methods and can identify and count live cells of probiotics. Conclusion: In conclusion, it should be mentioned that due to the superiority of the Real-time PCR method, we recommend the use of this molecular method, but for more assurance and comparison, several methods can be used to count and correctly identify probiotic strains.

Antioxidant Potential of Jicama (Pachyrhizus erosus) Extract Fermented by Lactobacillus plantarum B1765

Jicama is a source of phenolic compounds that function as antioxidants. However, the presence of polyphenol oxidase and naturally bound phenolic compounds limits its antioxidant potential. Fermentation is one method to enhance the antioxidant potential of jicama. This study investigates the growth of lactic acid bacteria (LAB), pH, total titratable acidity (TTA), total phenolic content (TPC), and antioxidant activity of jicama extract (Pachyrhizus erosus) using L. plantarum B1765 as a starter culture which was fermented for 2, 12, 24, and 36 hours at 37°C. Total LAB is measured using the whole plate count technique, pH using a pH meter, TTA by acid-base titration, TPC using the Folin-Ciocalteu method, and antioxidant activity using 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging method represented by IC50. The fermentation duration of jicama extract increased the growth of LAB to 108 CFU/mL and reached its optimum at 12 hours. However, reducing pH, increasing TTA to 0.223%, increasing TPC, and antioxidant activity still occur up to 36 hours of fermentation. Antioxidant activity was classified as vital. Duration of fermentation increased the growth of total LAB until the defined time, secreted β-glucosidase and inulinase enzymes, and also produced acid as their metabolism product, which hydrolyzed the glycoside bond and reduced pH, both act to free phenolic and increased antioxidant activity. This product has met Indonesian National Standards for beverage-fermented types and could be an antioxidant agent.

The combination of allicin with domiphen is effective against microbial biofilm formation.

Microorganisms in biofilms are particularly difficult to control because of their increased survival and antibiotic resistance. Allicin and domiphen were employed to inhibit the microbial growth and biofilm formation of Staphylococcus aureus, Escherichia coli, and Candida albicans strains. Broth microdilution method and checkerboard assay were conducted to determine the efficacy of allicin combined with domiphen against S. aureus, E. coli, and C. albicans. Microbial biofilm formation was measured using the crystal violet staining method and fluorescence microscopy. And the total viable count of the biofilm cells on material surface after the treatment with antimicrobial reagents was calculated with the plate count technique. The two drugs showed synergistic effects against the pathogens with a fractional bactericidal concentration of less than 0.38. The combination of 64 μg/mL allicin with 1 μg/mL domiphen dispersed over 50% of the biofilm mass of S. aureus, E. coli, and C. albicans. In addition, the drug combination reduced the total viable counts of E. coli and C. albicans biofilm cells on stainless steel and polyethylene surfaces by more than 102 CFU/mL. The combination of allicin and domiphen is an effective strategy for efficiently decreasing biofilms formation on various industrial materials surfaces.

Rationally designed protein A surface molecularly imprinted magnetic nanoparticles for the capture and detection of Staphylococcus aureus.

Staphylococcus aureus (S. aureus), a commensal organism found on the human skin, is commonly associated with nosocomial infections and exhibits virulence mediated by toxins and resistance to antibiotics. The global threat of antibiotic resistance has necessitated antimicrobial stewardship to improve the safe and appropriate use of antimicrobials; hence, there is an urgent demand for the advanced, cost-effective, and rapid detection of specific bacteria. In this regard, we aimed to selectively detect S. aureus using surface molecularly imprinted magnetic nanoparticles templated with a well-known biomarker protein A, specific to S. aureus. Herein, a highly selective surface molecularly imprinted polymeric thin layer was created on ∼250 nm magnetic nanoparticles (MNPs) through the immobilization of protein A to aldehyde functionalized MNPs, followed by monomer polymerization and template washing. This study employs the rational selection of monomers based on their computationally predicted binding affinity to protein A at multiple surface residues. The resulting MIPs from rationally selected monomer combinations demonstrated an imprinting factor as high as ∼5. Selectivity studies revealed MIPs with four-fold higher binding capacity (BC) to protein A than other non-target proteins, such as lysozyme and serum albumin. In addition, it showed significant binding to S. aureus, whereas negligible binding to other non-specific Gram-negative, i.e. Escherichia coli (E. coli), Pseudomonas aeruginosa (P. aeruginosa), and Gram-positive, i.e. Bacillus subtilis (B. subtilis), bacteria. This MIP was employed for the capture and specific detection of fluorescently labeled S. aureus. Quantitative detection was performed using a conventional plate counting technique in a linear detection range of 101-107 bacterial cells. Remarkably, the MIPs also exhibited approximately 100% cell recovery from milk samples spiked with S. aureus (106 CFU mL-1), underscoring its potential as a robust tool for sensitive and accurate bacterial detection in dairy products. The developed MIP exhibiting high affinity and selective binding to protein A finds its potential applications in the magnetic capture and selective detection of protein A as well as S. aureus infections and contaminations.

Comparison of the Structure of Soil Microbial Communities of Different Ecosystems Using the Microbiome Sequencing Approach

In this study, we aimed to compare the functional and taxonomic composition of soil microbial communities in different ecosystems, agricultural, natural grasslands, and old-growth forests, in the context of different environmental conditions. In this research, cultivable microbial quantification was performed by conventional plate-count techniques using different selective media. The taxonomic structure of microbe communities was evaluated using NGS metagenomic sequencing on the Illumina platform NovaSeq. The taxonomic analysis showed that individual land uses are characterized by the specific structure of communities; some taxonomic groups are specific only to agricultural, grassland, or forest ecosystems. After determining the abundance of functional groups of culturable microorganisms by the conventional plate-count method, statistically significant quantitative differences in physiological groups between the individual ecosystems were revealed. The metagenomic analysis revealed that different ecosystems are characterized by specific taxonomic groups of microorganisms and that general alpha diversity varies among individual land-use samples. Since the most unstable soil systems are agricultural, they are likely to suffer the most and will suffer more in the future from climate change than natural ones.

Influence of culture media composition on the rheology of microalgae concentrates on a large scale

The role of microalgae in the production of bioproducts and biofuels, along with their ability to provide a sustainable pathway for wastewater treatment, makes them promising alternatives to conventional processes. Nevertheless, large-scale downstream processing requires an understanding of biomass rheology that needs to be addressed further. This study aimed to characterize microalgal concentrates rheologically in different culture media. The presence of bacteria was quantified by photorespirometry and plate counting techniques. The culture medium was found to significantly influence viscosity, with primary wastewater exhibiting the highest viscosity and seawater plus pig slurry the lowest. The concentration of heterotrophic bacteria was directly related to the viscosity. Extracellular polysaccharides (EPS) in supernatant exhibited an inverse viscosity trend compared to biomass concentrates, with pig slurry cultures having higher concentrations. These findings emphasize the profound influence of culture medium and EPS on the rheology of microalgal biomass, underscoring the need for continued research aimed at facilitating and optimizing large-scale downstream processes within the framework of a circular economy and the attainment of the Sustainable Development Goals (6,8, and 12).

Characteristics of copper oxide and tin oxide nanoparticles produced by using pulsed laser ablation method and their application as an antibacterial agent

The synthesis of copper oxide (CuO) and tin oxide (SnO2) nanoparticles was successfully carried out using the neodymium yttrium aluminum garnet (Nd:YAG) laser ablation method. In this study, a laser beam was focused on the surface of high-purity (99.9%) Cu and Sn plates placed into deionized water (DIW) and 0.2% carboxymethyl cellulose (CMC) liquid media, respectively, to produce colloidal CuO and SnO2 nanoparticles. CuO and SnO2 nanoparticle colloids were successfully produced with greenish and brownish colors, respectively. The colloidal nanoparticles produced in CMC media are more stable. Thus, the antibacterial test on E. coli bacteria uses CuO and SnO2 samples in CMC media. Antibacterial assessments were carried out using disc diffusion and liquid dilution methods equipped with total plate counting (TPC) techniques, with a sample variation of 26 ppm CuO, 78 ppm SnO2, and a CuO-SnO2 mixture of 5:56 ppm and 19:19 ppm samples. The results of the disc diffusion test yielded the diameter of the bacterial inhibition zone. The diameter of the inhibition zone in CuO 26 ppm, SnO2 78 ppm, and CuO-SnO2 5:56 ppm and 19:19 ppm samples, respectively, were 7.43 mm, 6.39 mm, 7.18 mm, and 6.54 mm, while the liquid dilution test equipped with TPC produced the percentage of bacterial degradation, respectively, of 98.93%, 97.60%, 98.47%, and 97.73%. The results show that all variations had the effectiveness of inhibiting and killing bacteria. From the results of various bacterial tests, colloidal CuO nanoparticles are more effective than the SnO2 nanoparticles.

Antibacterial activity of Garcinia kola and Hunteria umbellata extracts on bacterial isolates from consumed sachet water in Edo State, Nigeria

"Water is an essential resource and its purity should not be negotiated when it comes to human consumption. This study investigated the antibacterial activity of Garcinia kola and Hunteria umbellata seed and epicarp on some bacterial isolates from sachet water. Duplicate samples of ten brands of sachet water were purchased from sales outlets around Ugbowo community, Benin City. Plate count techniques, minimum inhibitory and minimum bactericidal concentrations of the extracts were adopted in this investigation. Heterotrophic bacterial counts revealed highest range of 3.72±0.50x102 cfu/mL in EJ water and lowest in IB water (0.00±0.0x102 cfu/mL) while total coliform counts revealed its highest value in EJ water (3.62±0.30x102 cfu/mL) and lowest value (0.00±0.0 x102 cfu/mL), in OL, IB, NOS and UNI water. Aeromonas sp., Bacillus sp., Pseudomonas sp., Staphylococcus aereus and Enterobacter sp. were isolated. The antimicrobial susceptibility profile revealed varying zones of inhibition of 4 mm for Aeromonas sp., for Garcinia Kola and 20 mm recorded against Staphylococcus aureus and Bacillus sp. for Hunteria umbellata epicarp extract. The Minimum Inhibitory Concentration of susceptible bacteria to Hunteria umbellata and Garcinia kola extract were 6.25mg/mL and 12.5mg/mL, respectively, except Aeromonas sp. which had 50 mg/mL. Hunteria umbellata epicarp had a greater bactericidal effect of 6.25mg/mL against Staphylococcus aureus, while Garcinia kola had its greatest bactericidal effect on Enterobacter sp. with a minimum bactericidal concentration of 12.5mg/mL. This study has revealed the potentials of Hunteria umbellata epicarp and Garcinia kola as effective natural therapeutic agents against some harmful bacteria, preventing their pathogenic effect. Keywords: Herbal extracts, Hygiene education, Minimum Bactericidal Concentration, Minimum Inhibitory Concentration, Sachet water."

Degradation of crude oil by the microbial populations of Lagos and Ologe lagoon waters

Introduction: hydrocarbon contamination of surface waters as a result of anthropogenic activity poses threat to ecosystems and counter their beneficial uses. Some indigenous microbial communities have the potential to purify such waters unaided. Aims: To show the biodegradative potential of microbial communities in Lagos and Ologe Lagoons during minimal pollution with crude oil. Materials and Methods: The total heterotrophic bacteria and hydrocarbon utilising bacterial and fungal populations were estimated from Lagos and Ologe lagoon water samples contaminated with 1% crude oil over 42-day incubation period by plate count and vapour-phase transfer techniques. Residual hydrocarbons were determined by Gas chromatography. Results: The predominant bacterial genera identified from the lagoons include Enterobacter, Klebsiella and Proteus, while Escherichia, and Morganella. Aspergillus and Mucor were the predominant fungal genera in both waters. The hydrocarbon degradation rate in the Lagos Island microcosm was 65.391±0.370 mg/l/d, degradation rate constant 0.05±0.01 /d, half-life 9.559±0.093 /d and percentage degradation of 95.315 ± 0.134. Corresponding values in the Ologe water were 61.190± 8.542 mg/l/d, 8.725 ±0.389 /d, 0.055±0.003/d and 96.345±0.488 respectively. There was almost complete disappearance of the various fractions of the oil in the two samples. The microbial communities from both lagoons effectively utilised majority of the hydrocarbon fractions after 42 days where 66.890±1.075 and 100±000 were recorded for benzene, toluene had 100±000 percent degradation, anthracene 96.755±0.119 and 99.726±0.026, and pristane had 91.674±0.222 and 99.943±0.015 while phytane had 96.44±0.058 and 99.670±0.104 respectively. Conclusion: Efficient biodegradation of moderate contamination crude oil could be achieved by indigenous microbial flora present in Lagos and Ologe lagoon waters.

A Label-Free Carbohydrate-Based Electrochemical Sensor to Detect Escherichia coli Pathogenic Bacteria Using D-mannose on a Glassy Carbon Electrode.

Controlling water and food contamination by pathogenic organisms requires quick, simple, and low-cost methods. Using the affinity between mannose and type I fimbriae in the cell wall of Escherichia coli (E. coli) bacteria as evaluation elements compared to the conventional plate counting technique enables a reliable sensing platform for the detection of bacteria. In this study, a simple new sensor was developed based on electrochemical impedance spectroscopy (EIS) for rapid and sensitive detection of E. coli. The biorecogniton layer of the sensor was formed by covalent attachment of p-carboxyphenylamino mannose (PCAM) to gold nanoparticles (AuNPs) electrodeposited on the surface of a glassy carbon electrode (GCE). The resultant structure of PCAM was characterized and confirmed using a Fourier Transform Infrared Spectrometer (FTIR). The developed biosensor demonstrated a linear response with a logarithm of bacterial concentration (R2 = 0.998) in the range of 1.3 × 10 1~1.3 × 106 CFU·mL-1 with the limit of detection of 2 CFU·mL-1 within 60 min. The sensor did not generate any significant signals with two non-target strains, demonstrating the high selectivity of the developed biorecognition chemistry. The selectivity of the sensor and its applicability to analysis of the real samples were investigated in tap water and low-fat milk samples. Overall, the developed sensor showed to be promising for the detection of E. coli pathogens in water and low-fat milk due to its high sensitivity, short detection time, low cost, high specificity, and user-friendliness.

Effect of Pectin and Citric acid on Total Bacterial Load, Fungal Load, and Techno-economic Feasibility from processing to the storage of Guava Jelly

This study was conducted to investigate the Total bacterial load, fungal load, sensory evaluation, and techno-economic feasibility from processing to the storage of guava jelly. Fresh guava (Psidium guajava) jelly were prepared with 0.15%, 0.2%, and 0.25% pectin and with 1.1%, 1.15%, and 1.2% citric acid as a preservative. The bacterial count was determined through the Standard Plate Count (SPC) technique. The fungal analysis was done using a selective Sabouraud Dextrose Agar (SDA) medium. Lastly, the Inventory Theory’s model III was applied for the Techno-economic feasibility of guava jelly production. There was no yeast and mold growth detectable till day 90 of the storage period. However, the total bacterial load in jelly (acceptable limit ≤105cfu/ml) is satisfactory for consumption after 90 days of pectin and citric acid treatment. Furthermore, the production cost was approximately 65 Tk a bottle of 500g, which is considered economical compared to other local brands. Therefore, this formulation could be practiced for escalating the country’s development in the agriculture and food industries.

The effect of land management and organic matter inputs on bacterial population and soil nutrients across different types of agroforestry system

Abstract. Rinady MVP, Nuraini Y, Prayogo C, Arfarita N. 2023. The effect of land management and organic matter inputs on bacterial population and soil nutrients across different types of agroforestry system. Biodiversitas 24: 1333-1345. Different management of agroforestry systems changes crop composition and growth. This will affect organic matter inputs that become the source of food and energy for soil microorganisms. The soil organisms themself play an important role in amplifying soil biogeochemical processes and regulating soil reactions. This process allows the nutrients to be released into the soil and absorbed by the crop. This study aimed to examine the abundance of soil bacterial population and the changes in soil chemical properties (soil pH, total soil organic carbon (C), and total soil N (Nitrogen) under different land management and organic matter inputs. The determination of total organic C and total soil N was used by Walkey and Black and Kjedahl methods. Total Plate Count (TPC) techniques were employed to measure Total Bacteria Population (TBP) and Total Cellulolytic Bacteria (TCB). The Randomized Complete Block Design was used along with the 5% Tukey's test to examine the significant effect of the treatments. These treatments consisted of various agroforestry system as follow: (i) pine-coffee agroforestry system (PK), (ii) pine-banana agroforestry system (PPs), (iii) pine-cardamom agroforestry system (PR), (iv) pine-vegetable agroforestry system (PS), (v) mixed garden (KC), and (vi) citrus (LJ). Each treatment was repeated four times to obtain 24 experimental plots using a size of 20 m x 20 m. The results showed that the highest organic C and N content was obtained at the PK (agroforestry of pine at 41 years and coffee at 11 years old) plot, which was about 6.64% and 0.56%. Those parameters strongly correspond to the greatest soil bacteria population and soil cellulolytic bacteria population at the value of 1,71 x 105 CFU/g and 4,24 x 104 CFU/g, resulting from the greatest quantity of in-situ litter accumulation at PK plot to reach about 201.35 g. The dry weight value of litter in situ at the PK plot is greater than that of the PS and LJ plots by 81% and 87%. A power equation followed the relationship between soil organic C and the total bacteria population. A similar trend has been observed between total soil Nitrogen and total bacteria population. We concluded that changes in different management could affect soil chemical conditions and the changes in the total population of soil bacteria and cellulolytic soil bacteria.

Sugar-Lectin Interactions for Direct and Selective Detection of Escherichia coli Bacteria Using QCM Biosensor.

Pathogenic Escherichia coli (E. coli) remains a safety concern in the preservation and quality of green leafy vegetables. Sugar-lectin interactions provide a reliable, specific, and effective sensing platform for the detection of bacteria as compared to the tedious conventional plate counting technique. Herein, we present the synthesis of 4-(N-mannosyl) benzoic acid (4-NMBA) and 4-thiophenyl-N-mannose (4-TNM) via a two-step reductive amination for the detection of E. coli using a quartz crystal microbalance (QCM) biosensor. The 4-NMBA was synthesized with mannose and para-aminobenzoic (4-PBA), while the 4-TNM was synthesized with mannose and 4-aminophenyl disulfide (4-AHP) using water and acetic acid in a 1:1 ratio. The resultant structure of mannose derivatives (4-NMBA and 4-TNM) was characterized and confirmed using analytical tools, such as Mass Spectrometer, SEM, and FTIR. The choice of ligands (mannose derivatives) is ascribed to the specific recognition of mannose to the FimH lectin of the type 1 pilus of E. coli. Furthermore, the 4-PBA and 4-AHP conjugated to mannose increase the ligand affinity to FimH lectins. The setup of the QCM biosensor was composed of modification of the crystal surface and the covalent attachment of ligands for the detection of E. coli. The piezoelectric effect (frequency shift of the quartz) was proportional to the change in mass added to the gold crystal surface. Both the 4-NMBA- and 4-TNM-coated QCM sensors had a limit of detection of 3.7 CFU/mL and 6.6 CFU/mL with a sensitivity of 2.56 × 103 ng/mL and 8.99 × 10-5 ng/mL, respectively, within the dynamic range of 103 to 106 CFU/mL. This study demonstrates the application of ligand-coated QCM biosensors as a cost-effective, simple, and label-free technology for monitoring pathogenic bacteria via molecular interactions on crystal surfaces.

Insights into the enumeration of mixtures of probiotic bacteria by flow cytometry

The use of flow cytometry to enumerate microorganisms is gaining traction over the traditional plate count technique on the basis of superior accuracy, precision and time-to-result. Here, we assessed the suitability of live/dead flow cytometry for the enumeration of mixed populations of probiotic bacteria (L. acidophilus, L. paracasei, L. plantarum, L. salivarius, B. lactis and B. bifidum) whilst comparing outcomes with plate counting. Using a novel gating strategy designed specifically for the enumeration of mixed populations, the application of flow cytometry resulted in the detection of higher numbers of viable bacteria with a greater level of repeatability than plate counting (RSD of 6.82 and 13.14% respectively). Across all multi-species blends tested, viable cell input was more accurately recovered by flow cytometry (101.8 ± 6.95%) than plate counts (81.37 ± 16.03%). However, when certain probiotic mixtures contained preparations with high numbers of non-viable cells in their total population, flow cytometry had the potential for overestimation of the viable population. Nevertheless, the comparative plate counts of these mixtures were low and variable, thus supporting the use of flow cytometry for the enumeration of viable bacteria in mixed populations.