Evaluating the efficacy of a phage cocktail against Pseudomonas fluorescens group strains in raw milk: microbiological, physical, and chemical analyses.

The objective of this study was to investigate the effectiveness of a phage cocktail against Pseudomonas fluorescens group and its effect on the microbial, physical and chemical properties of raw milk during different storage conditions. A phage cocktail consisting of Pseudomonas fluorescens, Pseudomonas tolaasii, and Pseudomonas libanensis phages was prepared. As a result, reductions in fluorescent Pseudomonas counts of up to 3.44 log units for the storage at 4 °C and 2.38 log units for the storage at 25 °C were achieved. Following the phage application, it is found that there was no significant difference in the total mesophilic aerobic bacteria and Enterobacteriaceae counts. However, it was observed that the number of lactic acid bacteria was higher in phage-treated groups. The results also showed that pH values in the phage added groups were lower than the others and the highest titratable acidity was obtained only in the bacteria-inoculated group. As a future perspective, this study suggests that, while keeping the number of target microorganisms under control in the milk with the use of phages during storage, the microbiota and accordingly the quality parameters of the milk can be affected. This work contributes to the development of effective strategies for maintaining the quality and extending the shelf life of milk and dairy products.

Effect of wax separation on macro- and micro-elements, phenolic compounds, pesticide residues, and toxic elements in propolis.

Propolis, a natural product with many biological activities, is a resinous material produced by honeybees. It contains not only valuable components but also some possible contaminants in varying amounts. Hence, this study aimed to examine how the process step of wax separation affects certain elements, pesticide residues, and phenolic compounds in propolis. Total phenolics, elements, and some pesticide residues were analyzed in the crude propolis (CP samples), wax portion (W samples), and remaining propolis fraction (PF samples) after wax separation. Total phenolics of the CP samples were determined in the range of 31.90-45.00 mg GAE g-1 sample, while those of the PF samples were in the range of 54.97-162.09 mg GAE g-1 sample. Loss/reduction values by means of wax separation for phenolics were calculated as 10.88% and 17.89%, respectively. Pb contents of all PF samples were low (0.232-1.520 mg kg-1), but it was also noteworthy that nearly 40% or even more of Cr, As, Cd, and Pb were removed by wax separation. Removal of significant amounts of carbendazim (38.09%-67.35%), metalaxyl (81.57%-72.67%), tebuconazole (65.99%-78.36%), and propargite (88.46%-83.05%) was also achieved. Wax separation enables the removal of toxic substances from crude propolis without causing huge losses in phenolic compounds.

Evaluation of long- and short-term storage conditions and efficiency of a novel microencapsulated Salmonella phage cocktail for controlling Salmonella in chicken meat.

This study aims to assess the stability and activity of using a lyophilization, formulation design and to evaluate their efficiency for controlling Salmonella in chicken meat. The phage-loaded 0.3 M sucrose gelatin mixture at 4 and 25 °C displayed significantly less phage titer loss (p < 0.05) than the other excipients and liquid phage cocktail in 12 months. The results showed that there were significant reductions of Salmonella at the end of the storage in chicken meat for newly prepared phage powder (1.86 log CFU/cm2 and 2.18 log CFU/cm2), lyophilized phage powders stored at 4 °C (1.08 log CFU/cm2 and 1.26 log CFU/cm2) and stored at 25 °C (0.66 log CFU/cm2 and 1.00 log CFU/cm2) for 10 months at MOI 100 and 1000, respectively. The results demonstrated that lyophilized phages in a simple food grade formulation can be successfully stored and might be used in biocontrol of Salmonella in meat.

Estimation of time since deposition of semen stain on different fabric types using ATR-FTIR spectroscopy and chemometrics.

Various body fluids such as blood, semen, vaginal secretions, and saliva are frequently encountered at crime scene. In cases of sexual assault, semen stains are one of the most reliable evidence of biological origin. In this study, our objective was to develop a method for estimating the time since deposition of semen stains on five different fabric types using Attenuated Total Reflectance-Fourier Transform Infrared (ATR-FTIR) Spectroscopy, with a focus on a time frame of up to 8 weeks. Semen samples from six different volunteers were dripped onto five distinct fabric materials, and ATR-FTIR measurements were obtained at 17 different time points. Principal component analysis (PCA) and partial least squares (PLS) methods were employed to differentiate semen stains on various fabric samples and estimate the age of semen stains. Models constructed using PCA and PLSR achieved high R2 values and low root-mean-square error (RMSE). While the performance varies depending on fabric types, it was observed that age estimation of semen stains can be made within following intervals: 0.39-0.76 days for 0-7 day range, 2.59-3.38 days for the 1-8 week range, and 3.98-8.1 days for the 0-56 day range. This study demonstrates the effectiveness of using ATR-FTIR spectroscopy in combination with chemometrics to estimate the age of human semen stains on various fabric types based on time-dependent spectral changes.

Principles, Methods, and Real-Time Applications of Bacteriophage-Based Pathogen Detection.

Bacterial pathogens in water, food, and the environment are spreading diseases around the world. According to a World Health Organization (WHO) report, waterborne pathogens pose the most significant global health risks to living organisms, including humans and animals. Conventional bacterial detection approaches such as colony counting, microscopic analysis, biochemical analysis, and molecular analysis are expensive, time-consuming, less sensitive, and require a pre-enrichment step. However, the bacteriophage-based detection of pathogenic bacteria is a robust approach that utilizes bacteriophages, which are viruses that specifically target and infect bacteria, for rapid and accurate detection of targets. This review shed light on cutting-edge technologies about the novel structure of phages and the immobilization process on the surface of electrodes to detect targeted bacterial cells. Similarly, the purpose of this study was to provide a comprehensive assessment of bacteriophage-based biosensors utilized for pathogen detection, as well as their trends, outcomes, and problems. This review article summaries current phage-based pathogen detection strategies for the development of low-cost lab-on-chip (LOC) and point-of-care (POC) devices using electrochemical and optical methods such as surface-enhanced Raman spectroscopy (SERS).

Isolation, characterization, and application of bacteriophage cocktails for the biocontrol of Pseudomonas fluorescens group strains in whole and skimmed milk.

Pseudomonas fluorescens group strains can lead to spoilage of milk as well as loss of quality in dairy products through their heat-resistant enzymes. Phages are important alternatives for combating spoilage bacteria in food industry and used successfully in many applications. The aim of this study was the isolation and characterization of phages and to assess the efficiency of a phage cocktail in whole and skimmed milk. For this purpose, phages effective against Pseudomonas fluorescens (L23.2), Pseudomonas tolaasii (P22.1), and Pseudomonas rhodesiae (A11.1) were isolated. Their host range was found to be highly specific, and the transmission electron micrographs indicates that they belonged to Tectiviridae family. Their genome sizes were found to be vary between 38.3 and 53.5 kb. The latent periods and burst sizes were determined as 15, 10, 15 min and 91, 20, 80 PFU/infected cell for L23.2, P22.1, and A11.1, respectively. All three phages were found to be sensitive to low pH and high temperature. The effect of the phage cocktail was monitored in milk with different fat contents during storage at 4 °C for 5 days. As a result, bacterial reductions up to 4.09 and 5.29 log-units were observed for the whole and skimmed milk, respectively. Thus, the efficacy of a phage cocktail against a bacterial mixture of different P. fluorescens strains was tested in milk samples with different fat contents in accordance with real-life scenarios for the first time.

Evaluation of antibacterial efficacy of Lawsonia inermis Linn (henna) on periodontal pathogens using agar well diffusion and broth microdilution methods: An in-vitro study.

Background: Although widely explored in medicine, limited evidence exists in the literature regarding the efficacy of Lawsonia inermis Linn (henna) in the dental field. Aim: This study aimed to investigate the antibacterial effect of henna on Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis in vitro. Methods: The agar well diffusion and broth microdilution methods were used to evaluate the antibacterial effect of henna extracts. Dimethyl sulfoxide was used to prepare the ethanol extract of henna, and distilled water was used to prepare the water extract. For both ethanol and water extracts, 4 different concentrations were prepared as 15, 30, 60, and 120 mg/mL. Results: It was determined that the water and ethanol extracts of the henna samples did not show an inhibition zone on P.gingivalis and A.actinomycetemcomitans. As a result of the evaluations made with the broth microdilution method, it was found that the ethanol extract had a higher inhibitory effect on both bacteria, and both extracts had more inhibitory effects against A.actinomycetemcomitans. Conclusion: To understand the effect of henna on periodontal pathogens, more comprehensive in vitro studies should be performed on henna samples at different concentrations and with different bases.

Overcoming obstacles: Analysis of blood and semen stains washed with different chemicals with ATR-FTIR.

INTRODUCTION: Blood and semen stains are the most common biological stains encountered at crime scenes. The washing of biological stains is a common application that perpetrators use to spoil the crime scene. With a structured experiment approach, this study aims to investigate the effects of washing with various chemicals on the ATR-FTIR detection of blood and semen stains on cotton. MATERIALS AND METHODS: On cotton pieces, a total of 78 blood and 78 semen stains were applied, and each group of six stains was immersed or mechanically cleaned in water, 40% methanol, 5% sodium hypochlorite solution, 5% hypochlorous acid solution, 5 g/L soap dissolved pure water, and 5 g/L dishwashing detergent dissolved water. ATR-FTIR spectra gathered from all stains and analyzed with chemometric tools. RESULTS AND DISCUSSION: According to performance parameters of developed models, PLS-DA is a powerful tool for discrimination of washing chemical for both washed blood and semen stains. Results from this study show that FTIR is promising for use in detecting blood and semen stains that have become invisible to the naked eye due to washing of the findings. CONCLUSION: Our approach allows blood and semen to be detected on cotton pieces using FTIR combined with chemometrics, even though it is not visible to the naked eye. Washing chemicals also can be distinguished via FTIR spectra of stains.

Escherichia coli Enumeration in a Capillary-Driven Microfluidic Chip with SERS.

Pathogen detection is still a challenging issue for public health, especially in food products. A selective preconcentration step is also necessary if the target pathogen concentration is very low or if the sample volume is limited in the analysis. Plate counting (24-48 h) methods should be replaced by novel biosensor systems as an alternative reliable pathogen detection technique. The usage of a capillary-driven microfluidic chip is an alternative method for pathogen detection, with the combination of surface-enhanced Raman scattering (SERS) measurements. Here, we constructed microchambers with capillary microchannels to provide nanoparticle-pathogen transportation from one chamber to the other. Escherichia coli (E. coli) was selected as a model pathogen and specific antibody-modified magnetic nanoparticles (MNPs) as a capture probe in a complex milk matrix. MNPs that captured E. coli were transferred in a capillary-driven microfluidic chip consisting of four chambers, and 4-aminothiophenol (4-ATP)-labelled gold nanorods (Au NRs) were used as the Raman probe in the capillary-driven microfluidic chip. The MNPs provided immunomagnetic (IMS) separation and preconcentration of analytes from the sample matrix and then, 4-ATP-labelled Au NRs provided an SERS response by forming sandwich immunoassay structures in the last chamber of the capillary-driven microfluidic chip. The developed SERS-based method could detect 101-107 cfu/mL of E. coli with the total analysis time of less than 60 min. Selectivity of the developed method was also tested by using Salmonella enteritidis (S. enteritidis) and Staphylococcus aureus (S. aureus) as analytes, and very weak signals were observed.

Sorghum Flour Application in Bread: Technological Challenges and Opportunities.

Sorghum has a long history of use in the production of different types of bread. This review paper discusses different types of bread and factors that affect the physicochemical, technological, rheological, sensorial, and nutritional properties of different types of sorghum bread. The main types of bread are unleavened (roti and tortilla), flatbread with a pre-ferment (injera and kisra), gluten-free and sorghum bread with wheat. The quality of sorghum flour, dough, and bread can be improved by the addition of different ingredients and using novel and traditional methods. Furthermore, extrusion, high-pressure treatment, heat treatment, and ozonation, in combination with techniques such as fermentation, have been reported for increasing sorghum functionality.

Laser-induced breakdown spectroscopy as a reliable analytical method for classifying commercial cheese samples based on their cooking/stretching process.

The present work evaluates the possibility of using laser-induced breakdown spectroscopy (LIBS) coupled with chemometric methods to classify cheese samples (namely Kashar cheese and processed cheese) based on their cooking/stretching process. Chemometric analysis of the data provided by LIBS and ICP-OES/AAS analyses made it possible to discriminate between the two cheese types regarding their elemental profiles. The principal component analysis model was able to discriminate the Kashar cheese with an explained variance of 97.02%. Furthermore, the partial least squares discriminant analysis model perfectly classified the Kashar samples with a prediction ability of 100%. Furthermore, calibration and validation models for Mg, Ca, Na, P, Zn, and K elements for both Kashar and processed cheese samples were developed using partial least square regression yielding high correlation coefficients and low root mean square errors. Overall, this study indicates that LIBS with chemometrics can be an easy-to-use and rapid monitoring system for cheese classification.

Surface-enhanced Raman scattering-based detection of plasmin activity by specific peptide substrate.

In this study, a new surface-enhanced Raman scattering (SERS)-based method has been developed for the detection of plasmin activity. Firstly, different peptide sequences, which are specific to plasmin, were examined. Then, SERS substrates were prepared by chosen peptide substrate. Enzyme activity was determined by pursuing the reduction of DTNB band at 1331 cm-1 with Raman spectroscopy. The reduction in SERS intensity was related to the plasmin activity, and changes in SERS intensity vs. plasmin concentration graph was obtained. Limit of detection (LOD) and limit of quantification (LOQ) values were calculated as 2.14 U/mL and 6.42 U/mL, respectively. Intra- and inter-day repeatability results were determined as 1.45% and 1.47% relative standard deviation (RSD). Also, recovery of the method was determined for the plasmin spiked milk samples. The results demonstrated that the proposed method could be successfully used to detect the plasmin activity in milk samples.

Applications of Bacteriophage Cocktails to Reduce Salmonella Contamination in Poultry Farms.

Salmonella contamination is a critical problem in poultry farms, with serious consequences for both animals and food products. The aim of this study is to investigate the use of phage cocktails to reduce Salmonella contamination in poultry farms. Within the scope of the study, Salmonella phages were isolated from chicken stool. After the host range of phages was determined, morphological characterization was performed through transmission electron microscopy analysis. Then, replication parameters and adsorption rates were determined by one-step growth curves. After that, phage cocktail was prepared, and its effectiveness was tested in three environments, which were drinking water, shavings, and plastic surfaces. The results obtained have demonstrated that the phage cocktail can reduce Salmonella count up to 2.80 log10 units in drinking water, up to 2.30 log10 units on shavings, and 2.31 log10 units on plastic surfaces. It has been determined that phage cocktails could be a successful alternative in reducing Salmonella contamination in poultry environment. This work is the first study to investigate the use of phage cocktails for reducing Salmonella contamination in poultry water and on shavings, and it is presumed that the results obtained will contribute to the fight against pathogens by making them applicable to poultry farms.

Investigation of phage and molasses interactions for the biocontrol of E. coli O157:H7.

Resistance to antibiotics is one of the most critical health problems in the world. Therefore, finding new treatment methods to be used as alternatives to antibiotics has become a priority for researchers. Similar to phages, certain products containing antimicrobial components, such as molasses, are widely used to eliminate resistant bacteria. Molasses has a strong antimicrobial effect on bacterial cells, and this effect is thought to be due to the breakdown of the cytoplasmic cell membrane and cell proteins of the polyphenols in molasses. In the present study, phage-molasses interactions were investigated to examine the effects of concomitant use. It was found that molasses samples increased the size of phage plaques by up to 3-fold, and MIC and 1/2 × MIC concentrations of molasses increased the burst size of phages. Although no synergistic effect was found between the phage and molasses, the antimicrobial activities of the components and the effect of molasses on phage activity were demonstrated.

A new and facile route to prepare gold nanoparticle clusters on anodic aluminium oxide as a SERS substrate.

In the present study, a new SERS-active gold nanoparticle clusters having a flower-shape have been prepared easily on nano porous anodic aluminium oxide (AAO) by immersing it in auric chloride solution without any need for complex production steps. In this process, presented for the first time, the metallic aluminum which were released under the influence of chloride ions due to pitting corrosion act as a reducing agent, while gold ions were reduced onto the AAO layer based on the difference in standard reduction potentials between aluminum and gold. Gold nanoparticle clusters on AAO layer formed "hot spots" providing enhanced Raman signal. Optical microscope, SEM, EDX, AFM, and UV-vis spectrophotometer have been used to characterize the substrate. In order to demonstrate applicability of the method, label free SERS measurements of nitrate ion was performed on the proposed sensing platform. A high sensitivity with 1.03 ppm of limit of detection level and the enhancement factor of 2.9 × 105 were obtained for nitrate ion. In addition, remarkable recoveries ranging from 98.4% to 106.8% were obtained for nitrate spiked into drinking water samples. The inter-day and intra-day precisions of the method as relative standard deviation (RSD) were determined as 3.3% and 5.2%, respectively. The sensor platform, developed using a facile method and a low-cost base material (aluminum), can be a good alternative for SERS based sensing applications.

Replacement of antibodies with bacteriophages in lateral flow assay of Salmonella Enteritidis.

In this study, the analytical performance of bacteriophages for Salmonella Enteritidis was investigated using lateral flow assay (LFA) technique. The analytical performance characteristics of bacteriophages were compared with antibodies which are regularly used as analyte-specific agents in the lateral flow immunoassay test strip. Bacteriophages could be an alternative analyte-specific agents to antibodies in lateral flow assay testing of bacteria since they offer comparable sensitivity, specificity, and accuracy. In the present study, Surface Enhanced Raman Spectroscopy (SERS) and colorimetric measurements were combined in one platform and sensitive quantitation of target bacteria was accomplished with a total quantitative analysis time of less than 30 min. The developed Salmonella Enteritidis F5-4 phage-based LFA specifically responds to Salmonella Enteritidis, while lower SERS responses to different bacteria types including Bacillus subtilis, Micrococcus luteus, Escherichia coli, Salmonella Typhimurium were observed. The developed test strips were also applied for the determination of Salmonella Enteritidis in spiked chicken and egg samples.

Determination of milk fat authenticity in ultra-filtered white cheese by using Raman spectroscopy with multivariate data analysis.

Cheese is one of the most widely consumed food products in the world. However, the increasing demand for nutritionally enhanced or functional products by the cheese industry has created new approaches that partially or fully replace milk fat. With this, new methods of adulteration have also been noted, potentially leading to these fully/partially-replaced products being offered as cheese. In this study, Raman spectroscopy was used to determine origins of fats in margarine, corn, and palm oils present in white and ultra-filtered cheese samples. Raman spectra were evaluated with partial least square-discriminant (PLS-DA) and PLS to identify fat/oil origins and adulteration ratios. The coefficients of determination and limits of detection for margarine, and corn and palm oil adulteration were found to be 0.990, 0.993, 0.991 and 3.38%, 3.36% and 3.59%, respectively.

Protein based evaluation of meat species by using laser induced breakdown spectroscopy.

Meat adulteration through partial substitution with cheaper species or mislabeling causes significant problems in terms of health, religious beliefs, economy, and product quality. Therefore, identification of meat species is crucial for monitoring and prevention of meat fraud. In the present study, protein based laser induced breakdown spectroscopy method was developed for the first time to identify three meat species (beef, chicken and pork) by using bulk proteins and protein fractions, namely actin and myosin. LIBS spectra were evaluated with principal component analysis for clustering pattern of meat species, and partial least square analysis was performed to determine adulteration ratio. In PLS analysis, limit of detection (LOD) values for beef adulteration with chicken and pork meat were calculated as 2.84% and 3.89% by using bulk proteins, respectively.

Multiplex enumeration of Escherichia coli and Salmonella enteritidis in a passive capillary microfluidic chip.

Multiplex detection and quantification of bacteria in water by using portable devices are particularly essential in low and middle-income countries where access to clean drinking water is limited. Addressing this crucial problem, we report a highly sensitive immunoassay sensor system utilizing the fluorescence technique with magnetic nanoparticles (MNPs) to separate target bacteria and two different types of quantum dots (CdTe and Ni doped CdTe QDs) incorporated into a passive microfluidic chip to transport and to form sandwich complexes for the detection of two target bacteria, namely Escherichia coli (E. coli) and Salmonella enteritidis (S. enteritidis) in less than 60 min. The assay is carried out on a capillary driven microfluidic chip that can be operated by merely pipetting the samples and reagents, and fluorescence measurements are done by using a handheld fluorescence spectrophotometer, which renders the system portable. The linear range of the method was found to be 101 to 105 cfu mL-1 for both E. coli and S. enteritidis. The limit of detection (LOD) was calculated to be 5 and 3 cfu mL-1 for E. coli and S. enteritidis, respectively. The selectivity of the method was examined by testing Enterobacter dissolvens (E. dissolvens) and Staphylococcus aureus (S. aureus) samples, and no significant interference was observed. The method was also demonstrated to detect bacteria in tap water and lake water samples spiked with target bacteria.

Immunomagnetic separation and Listeriamonocytogenes detection with surface-enhanced Raman scattering

Background/aim: We aimed to develop a rapid method to enumerate Listeria monocytogenes (L. monocytogenes) utilizing magnetic nanoparticle based preconcentration and surface-enhanced Raman spectroscopy measurements. Materials and methods: Biological activities of magnetic Au-nanoparticles have been observed to have the high biocompatibility, and a sample immunosensor model has been designed to use avidin attached Au-nanoparticles for L. monocytogenes detection. Staphylococcus aureus (S. aureus) and Salmonella typhimurium (S. typhimurium) bacteria cultures were chosen for control studies. Antimicrobial activity studies have been done to identify bio-compatibility and bio-characterization of the Au-nanoparticles in our previous study and capturing efficiencies to bacterial surfaces have been also investigated. Results: We constructed the calibration graphs in various population density of L. monocytogenes as 2.2 × 101 to 2.2 × 106 cfu/mL and the capture efficiency was found to be 75%. After the optimization procedures, population density of L. monocytogenes and Raman signal intensity showed a good linear correlation (R2 = 0.991) between 102 to 106 cfu/mL L. monocytogenes. The presented sandwich assay provides low detection limits and limit of quantification as 12 cfu/mL and 37 cfu/mL, respectively. We also compared the experimental results with reference plate-counting methods and the practical utility of the proposed assay is demonstrated using milk samples. Conclusion: It is focused on the enumeration of L. monocytogenes in milk samples and the comparision of results of milk analysis obtained by the proposed SERS method and by plate counting method stay in food agreement. In the present study, all parameters were optimized to select SERS-based immunoassay method for L. monocytogenes bacteria to ensure LOD, selectivity, precision and repeatablity.