Rapid Detection of Total Viable Count in Intact Beef Dishes Based on NIR Hyperspectral Hybrid Model.

The total viable count (TVC) of bacteria is an important index to evaluate the freshness and safety of dishes. To improve the accuracy and robustness of spectroscopic detection of total viable bacteria count in a complex system, a new method based on a near-infrared (NIR) hyperspectral hybrid model and Support Vector Machine (SVM) algorithms was developed to directly determine the total viable count in intact beef dish samples in this study. Diffuse reflectance data of intact and crushed samples were tested by NIR hyperspectral and processed using Multiplicative Scattering Correction (MSC) and Competitive Adaptive Reweighted Sampling (CARS). Kennard-Stone (KS) and Samples Set Partitioning Based on Joint X-Y Distance (SPXY) algorithms were used to select the optimal number of standard samples transferred by the model combined with root mean square error. The crushed samples were transferred into the complete samples prediction model through the Direct Standardization (DS) algorithm. The spectral hybrid model of crushed samples and full samples was established. The results showed that the Determination Coefficient of Calibration (RP2) value of the total samples prediction set increased from 0.5088 to 0.8068, and the value of the Root Mean Square Error of Prediction (RMSEP) decreased from 0.2454 to 0.1691 log10 CFU/g. After establishing the hybrid model, the RMSEP value decreased by 9.23% more than before, and the values of Relative Percent Deviation (RPD) and Reaction Error Relation (RER) increased by 12.12% and 10.09, respectively. The results of this study showed that TVC instewed beef samples can be non-destructively determined based on the DS model transfer method combined with the hybrid model strategy. This study provided a reference for solving the problem of poor accuracy and reliability of prediction models in heterogeneous samples.

Antimicrobial effect of photodynamic therapy using sinoporphyrin sodium and 390-400 nm light-emitting diode on Porphyromonas gingivalis in vitro.

This study aims to investigate the effect of antimicrobial photodynamic therapy (a-PDT) using a novel combination of sinoporphyrin sodium (DVDMS) and light-emitting diode (LED) with a wavelength of 390-400 nm on Porphyromonas gingivalis in vitro. Absorption spectrum of DVDMS was determined by spectrometer for selecting suitable wavelength light source. The uptake of DVDMS by P. gingivalis was evaluated according to fluorescence intensity detected by a spectrometer. Then effects of DVDMS alone, 390-400 nm LED alone, and photodynamic therapy produced by 10, 20, 40, and 80 µg/mL DVDMS and 390-400 nm LED on the suspension of P. gingivalis were evaluated by counting the number of colony forming units (CFU) after incubation. In the experiment, the LED illumination time was 30, 60, 90, 120, 180, 240, and 360 s, respectively, and the corresponding energy density was 1, 2, 3, 4, 6, 8, and 12 J/cm2, respectively. According to the absorption spectrum of DVDMS, the 390-400-nm light emitted by the LED was selected as the light source. The fluorescence intensity of DVDMS on P. gingivalis increased significantly at 5 min, and with the extension of time, it decreased at 30 min. DVDMS alone did not produce a significant toxicity on P. gingivalis compared with PBS (p = 0.979). While 390-400 nm LED alone had a certain bactericidal effect on P. gingivalis, the bactericidal effect was more obvious as the light dose increased (p < 0.001). The effect of a-PDT produced by 20, 40, and 80 µg/mL DVDMS and 390-400 nm LED were significantly better than that of 390-400 nm LED alone (p < 0.05). Both DVDMS concentration and light dose could enchance the bactericidal effect. The strongest photo-killing effect was generated by 80 µg/mL DVDMS with 360 s illumination (energy density is 12 J/cm2), and the log reduction of bacteria was 5.69 ± 1.70. a-PDT using the combination of DVDMS with 390-400 nm LED shows promise as a new treatment modality for pathogens elimination in periodontal therapy.

Synthetic dye's substitution with chokeberry extract in jelly candies.

Matching the general trend of replacing synthetic additives with healthier natural products, the present research studies the effects of different concentrations of chokeberry extract which substitute carmoisine dye in jelly candies. Also, the colour and antioxidant properties of the aforementioned extract and their changes at various pH and in presence of different mineral salts from foods are analysed. The phenolic content of the extract was determined using HPLC and spectrophotometric methods. A high concentration of polyphenols was found in the chokeberry extract, of which around 97% were flavonoids. Catechin, epicatechin, ferulic acid and its methyl esther, protocatechuic, gallic and para-hydroxybenzoic acids were the major phenolics identified in the extract. The total antioxidant activity decreased in acidic media, while close-to-neutral and alkaline pH values did not exhibit any effect on this parameter. Furthermore, the green/red colour parameter, the chroma and the hue angle were enhanced in the most acidic media (pH 2.3 and 3.5). From the studied salts, CaCl2 and KNO3 had the most significant effects on colour. The chokeberry extract proved to be suitable as replacement of carmoisine dye in jelly candies, as the physicochemical and microbiological properties comply with the regulated requirements. More than that, the extract improved the antioxidant and sensory properties of jellies in all studied concentrations and the best total sensory score was obtained for 1.5% extract. After 5 and 50 days of storage, the microbiological properties were improved in candies prepared with aronia extracts compared to carmoisine, as the total viable count registered important diminutions.

Isothermal microcalorimetry for thermal viable count of microorganisms in pure cultures and stabilized formulations.

BACKGROUND: Quantification of viable microorganisms is an important step in microbiological research as well as in microbial product formulation to develop biological control products or probiotics. Often, the efficiency of the resulting product is dependent on the microbial cell density and their viability, which may decrease over time. Commonly, the number of viable cells is determined by serial dilution and plating techniques or flow cytometry. In 2017, we developed a mathematical model for isothermal microcalorimetry (IMC) data analysis and showed that the new method allows for a more rapid quantification of viable fresh and freeze-dried anaerobic Lactobacillus reuteri cells than traditional viable count methods. RESULTS: This study developed the new method further by applying it to well-known aerophilic plant-beneficial microbial species (Pseudomonas brassicacearum, Bacillus amyloliquefaciens subsp. plantarum and Clonostachys rosea) used in biological control products. We utilized IMC to quantify viable cells in microbial pure cultures as well as when coated onto wheat seeds. The results from this study confirmed that thermal viable count methods are more rapid and sensitive than traditional viable count techniques. Most interestingly, a thermal viable count method was able to quantify microbes coated on seeds despite the presence of the natural microbiota of the seeds. Our results also showed that, in contrast to plating techniques for which clustered cells skew the results, IMC does not require single cells for accurate viable counts. CONCLUSIONS: Thermal viable count methods are novel methods for the rapid quantification of divergent bacterial and fungal species and enhance the speed, sensitivity, and accuracy of routine viable counts of pure cultures and controlled microbiomes such as plant seed coatings.

Heavy metal contamination, microbiological spoilage and biogenic amine content in sushi available on the Polish market.

BACKGROUND: The present study determined the heavy metal contamination (mercury, cadmium, lead, arsenic and nickel) of nori, restaurant-served sushi and ready-to-eat sushi meals available via retail chains. Moreover, both microbiological load and biogenic amine content in ready-to-eat sushi meals were analysed. RESULTS: All of the nori samples contained high levels of Cd (2.122 mg kg-1 ), Ni (0.715 mg kg-1 ), As (34.56 mg kg-1 ) and Pb (0.659 mg kg-1 ). The studied sushi samples contained high levels of Ni and Pb, reaching 0.194 and 0.142 mg kg-1 wet weight, respectively, being potentially hazardous to women during pregnancy and lactation and small children. None of the studied samples contained high levels of Hg. Overall, 37% of ready-to-eat sushi meals exceeded a microbiological load of 106 cfu g-1 . However, biogenic amine content in all of the samples was low, with a highest histamine content of 2.05 mg kg-1 . CONCLUSION: Sushi is not the source of high levels of biogenic amines even with high microbiological loads. Nevertheless, the high microbiological loads at the end of the shelf-life indicate that some processors might have problems with the distribution chain or implement a poor hygienic regime. Moreover as a result of possible risk associated with heavy metal contamination, the present study highlights the need to establish new regulations regarding the contamination of nori and sushi. © 2017 Society of Chemical Industry.

Differential detection of pathogenic Yersinia spp. by fluorescence in situ hybridization.

Yersinia enterocolitica, Y. pseudotuberculosis and Y. pestis are pathogens of major medical importance, which are responsible for a considerable number of infections every year. The detection of these species still relies on cultural methods, which are slow, labour intensive and often hampered by the presence of high amounts of accompanying flora. In this study, fluorescence in situ hybridization (FISH) was used to develop a fast, sensitive and reliable alternative to detect viable bacteria in food. For this purpose, highly specific probes targeting the 16S and 23S ribosomal RNA were employed to differentially detect each of the three species. In order to enable the differentiation of single nucleotide polymorphisms (SNPs), suitable competitor oligonucleotides and locked nucleic acids (LNAs) were used. Starved cells still showed a strong signal and a direct viable count (DVC) approach combined with FISH optimized live/dead discrimination. Sensitivity of the FISH test was high and even a single cell per gram of spiked minced pork meat could be detected within a day, demonstrating the applicability to identify foodborne hazards at an early stage. In conclusion, the established FISH tests proved to be promising tools to compensate existing drawbacks of the conventional cultural detection of these important zoonotic agents.

Antioxidant and Antimicrobial Effects of Pistacia lentiscus L. Extracts in Pork Sausages.

Pistacia lentiscus fruits are ingredients of traditional Cypriot sausages. The objective of this study is to evaluate P. lentiscus extracts as natural additives to the sausages. First, the phenolic content and antioxidant activity of fruit and leaf extracts were determined. Results revealed that leaves are richer source of polyphenolic antioxidants than fruits, with methanol being the better extraction solvent. In the next step, the antioxidant and antimicrobial effects of methanolic extracts (300 mg/kg) in the pork sausage formulation were investigated. Peroxide, acid and thiobarbituric acid-reactive substance values demonstrated that both fruit and leaf extracts reduced the rate of lipid oxidation of sausages at 4 °C. Total viable count revealed significant differences on the fifth day of storage, with better microbial inhibition by leaf extract. No significant differences between the extracts were observed after the tenth day of storage. Overall, the extracts can be used to prevent lipid oxidation and reduce microbial spoilage during the first days of storage of fresh traditional pork sausages.

Performance evaluation of bulk freeze dried starter cultures of dahi and yoghurt along with probiotic strains in standardized milk of cow and buffalo.

Performance of bulk freeze dried (BFD) cultures of dahi (D) and yoghurt (Y) either with or without probiotic cultures (AB -Lactobacillus acidophilus and Bifidobacterium bifidum) in standardized milk of cow and buffalo was evaluated. In buffalo milk, significantly (p < 0.05) low viable count of probiotic culture combination of dahi cultures (DAB) over non probiotic combination (D) was noticed; whereas, difference in counts of yoghurt culture combinations Y and YAB was not significant. The culture activity of D and DAB was similar in both types of milk, however, the volatile acidity (VA) produced by combination D was higher (32.5 ml/50 g sample) in buffalo milk than in cow milk (29.2 ml/50 g sample). Whereas, DAB produced very low amount of VA (16 ml/50 g sample) both in cow and buffalo milk. The diacetyl and tyrosine contents produced by either D or DAB in cow or buffalo milk were in the same order. Although Y and YAB produced slightly more VA in buffalo milk than in cow milk, significant change in the performance of yoghurt cultures (Y or YAB) both in cow and buffalo milk was not noticed. However, the VA and acetaldehyde produced by YAB either in cow and buffalo milk was higher than that by combination Y. Addition of probiotic cultures significantly enhanced the production of acetaldehyde content in both types of milk. Difference in tyrosine content in yoghurt made either with cow or buffalo milk was not significant. Overall, the present study indicated that the BFD cultures can be used to prepare dahi or yoghurt either from cow or buffalo milk, without affecting the biochemical profile of these products.

Isotachophoretic quantification of total viable bacteria on meat and surfaces.

BACKGROUND: The quantification of microbes, particularly live bacteria, is of utmost importance in assessing the quality of meat products. In the context of meat processing facilities, prompt identification and removal of contaminated carcasses or surfaces is crucial to ensuring the continuous production of safe meat for human consumption. The plate count method and other traditional detection methods are not only labour-intensive but also time-consuming taking 24-48 h. RESULTS: In this report, we present a novel isotachophoretic quantification method utilizing two nucleic acid stains, SYTO9 and propionic iodide, for the detection of total viable bacteria. The study employed E. coli M23 bacteria as a model organism, with an analysis time of only 30 min. The method demonstrated a limit of detection (LOD) of 184 CFU mL-1 and 14 cells mL-1 for total viable count and total cell count, respectively. Furthermore, this new approach is capable of detecting the microbial quality standard limits for food contacting surfaces (10 CFU cm-2) and meat (1.99 × 104 CFU cm-2) by swabbing an area of 10 × 10 cm2. SIGNIFICANCE: In contrast to the culture-based methods usually employed in food processing facilities, this isotachophoretic technique enables easy and rapid detection (<30 min) of microorganisms, facilitating crucial decision-making essential for maintaining product quality and safety.

Mathematical modeling of Escherichia coli O157:H7 growth in carrot juice influenced by Thymbra capitata essential oil, heat treatment, and storage temperature.

The aim of this study was to develop a mathematical model describing the survival of Escherichia coli O157:H7 in carrot juice treated with Thymbra capitata essential oil combined with mild heat treatment and stored at different temperatures. The viable count method was used to investigate the effect of the treatment on bacterial survival, and the response surface methodology was used to develop a statistical model fitting the data. The results showed that the variance of bacterial growth is explained by storage temperature (37 %) and heat treatment (35 %), these are followed by Thymbra capitata essential oil (18 %) and their interaction (9 %). Positive multiplicative interaction was obtained for any pair of the studied treatments and cooperative effect synergy was observed over a large domain of these factors. A mathematical model was successfully developed to describe Escherichia coli O157:H7 response to the selected factors, within the study limits, and to estimate the risk of juice contamination and shelf-life. Based on our results, the use of Thymbra capitata essential oil combined with heat treatment may control Escherichia coli O157:H7 growth in carrot juice stored at low temperature.

Effects of cold storage time on the quality and active probiotics of yogurt fermented by Bifidobacterium lactis and commercial bacteria Danisco.

In this study, the commercial bacteria Danisco and Bifidobacterium lactis were used to ferment soy yogurt, and then the quality of yogurt and the number of active probiotics in yogurt during storage were investigated. The results showed that the total number of viable bacteria in soy yogurt increased first and then decreased, but all of them met the standard for the number of viable bacteria in probiotic foods. The content of protein, lipid, and total sugar in soy yogurt decreased gradually with the extension of storage time. The texture, water holding capacity, and rheological properties of soy yogurt were improved within 0-10 days, and there was no significant change after 15 days. However, brightness and whiteness of yogurt were significantly reduced. Based on realizing the reuse of soy whey, this study provided a theoretical basis for the research of the shelf life of soy yogurt. PRACTICAL APPLICATION: This study developed a soy yogurt with good quality and provided a theoretical basis for the study of the shelf life of soy yogurt. In addition, some technical support was provided for the reuse of soy whey.

An improved MTT colorimetric method for rapid viable bacteria counting.

The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay has been employed in the analysis of bacterial growth. In comparison to experiments conducted on mammalian cells, the MTT bacterial assay encounters a greater number of interfering factors and obstacles that impact the accuracy of results. In this study, we have elucidated an improved MTT assay protocol and put forth an equation that establishes a correlation between colony-forming units (CFU) and the amount of formazan converted by the bacteria, drawing upon the fundamental principle of the MTT assay. This equation is represented as CFU=kF. Furthermore, we have explicated a methodology to determine the scale factor "k" by employing S. aureus and E. coli as illustrative examples. The findings indicate that S. aureus and E. coli reduce MTT by a cyclic process, from which the optimal reduction time at room temperature was determined to be approximately 30 mins. Furthermore, individual E. coli exhibits an MTT reduction capacity approximately four times greater than that of S. aureus. HPLC analysis proves to be the most accurate method for mitigating interferences during the dissolution and quantification of formazan. Additionally, this study has identified a new constraint related to the narrow linear range (0-125 µg/mL) of formazan concentration-absorbance and has presented strategies to circumvent this limitation.

Probiotic and postbiotic analytical methods: a perspective of available enumeration techniques.

Probiotics are the largest non-herbal/traditional dietary supplements category worldwide. To be effective, a probiotic strain must be delivered viable at an adequate dose proven to deliver a health benefit. The objective of this article is to provide an overview of the various technologies available for probiotic enumeration, including a general description of each technology, their advantages and limitations, and their potential for the future of the probiotics industry. The current "gold standard" for analytical quantification of probiotics in the probiotic industry is the Plate Count method (PC). PC measures the bacterial cell's ability to proliferate into detectable colonies, thus PC relies on cultivability as a measure of viability. Although viability has widely been measured by cultivability, there has been agreement that the definition of viability is not limited to cultivability. For example, bacterial cells may exist in a state known as viable but not culturable (VBNC) where the cells lose cultivability but can maintain some of the characteristics of viable cells as well as probiotic properties. This led to questioning the association between viability and cultivability and the accuracy of PC in enumerating all the viable cells in probiotic products. PC has always been an estimate of the number of viable cells and not a true cell count. Additionally, newer probiotic categories such as Next Generation Probiotics (NGPs) are difficult to culture in routine laboratories as NGPs are often strict anaerobes with extreme sensitivity to atmospheric oxygen. Thus, accurate quantification using culture-based techniques will be complicated. Another emerging category of biotics is postbiotics, which are inanimate microorganisms, also often referred to as tyndallized or heat-killed bacteria. Obviously, culture dependent methods are not suitable for these products, and alternative methods are needed for their quantification. Different methodologies provide a more complete picture of a heterogeneous bacterial population versus PC focusing exclusively on the eventual multiplication of the cells. Alternative culture-independent techniques including real-time PCR, digital PCR and flow cytometry are discussed. These methods can measure viability beyond cultivability (i.e., by measuring cellular enzymatic activity, membrane integrity or membrane potential), and depending on how they are designed they can achieve strain-specific enumeration.

Periplaneta americana L. a potential source of traditional medicine: chemometric analysis, in vitro and in silico study.

'Mayurbhanj is the ethnic dominant tribal population district in Odisha, India. The triabl's of Mayurbhanj depends on traditional medicines since time immemorial for health-related issues. Due to the imperative ethnic claim of traditional healers, the financial stringency of the patient community and the necessity to produce a better therapeutic effect has led to investigate ethno zoological sources and to find out the biochemical moiety responsible for the healing process. Considering the ethnic communities' acceptability of the zoological source as traditional medicine, the current evidence-based research study is conducted to investigate the biochemical moiety present in Periplaneta americana, responsible for therapeutic activity. The whole powdered Periplaneta americana was extracted using maceration techniques with n-hexane and methanol as solvent. The obtained extracts were subjected to GC-MS analysis to identify the biochemical moiety. To check the potential biological activity, an in-vitro antimicrobial test was carried out in both turbidimetry and a viable count method against E. coli. Moreover, the obtained biochemical molecules were exposed to in silico studies for their binding modes and their affinity using Discovery studio software. The major compounds were found to be hexadecanoic acid, methyl ester, n-hexadecanoic acid, oleic acid, octadecanoic acid along with other minor constituents. The maximum inhibitory activity of n-hexane and methanol extract against S. aureus at a concentration of 400 µg/mL was found to be 89 and 87%, respectively. The binding models of almost all identified compounds confer very good binding affinities with some key and strong non-covalent interactions with various amino acid residues of receptor active site pocket, which predict the compounds to be potent inhibitors of various infectious bacteria. These findings suggested that the hexane extract of P. americana could be exploited as a potential natural source. Communicated by Ramaswamy H. Sarma.

Vitality, fermentation, aroma profile, and digestive tolerance of the newly selected Lactiplantibacillus plantarum and Lacticaseibacillus paracasei in fermented apple juice.

Background: To enrich the probiotic lactic acid bacteria (LAB) strains and expand the commercialization of new fermented juice products, we have identified two LAB strains with excellent potential in fermenting apple juice from pickles. Methods: The two strains were morphologically, physiologically, and genetically characterized. The strains' fermentation performance and alterations in volatile aroma components of apple juice and ability to survive in a simulated gastrointestinal environment were evaluated. Results: Two strains were identified as Lacticaseibacillus paracasei (WFC 414) and Lactiplantibacillus plantarum (WFC 502). The growth of WFC 414 and WFC 502 in apple juice for 48 h reached 8.81 and 9.33 log CFU/mL, respectively. Furthermore, 92% and 95% survival rates were achieved in 2 h simulated gastric juice, and 80.7 and 83.6% survival rates in 4 h simulated intestinal juice. During the fermentation, WFC 414 and WFC 502 reduced the soluble sugars and total polyphenols in apple juice, and consumed malic acid to produce large amounts of lactic acid (3.48 and 5.94 mg/mL). In addition, the esters and aldehydes were reduced, and the production of alcohols, acids and ketones was elevated in the apple juice fermented by both strains. Conclusion: These results show that WFC 414 and WFC 502 have great potential applications in the fermented fruit juice industry.

UV-fluorescence imaging for real-time non-destructive monitoring of pork freshness.

This study aimed to develop a cost-effective fluorescence imaging system to rapidly monitor pork freshness indicators during chilled storage. The system acquired fluorescence images of pork and the color features were extracted from these images to establish partial least squares regression (PLSR) models to predict total volatile basic nitrogen (TVB-N), total viable count (TVC), pH for pork. For TVB-N, TVC and pH values, Rp were 0.92, 0.88 and 0.74, residual predictive deviation (RPD) were 2.24, 2.03, and 1.19, respectively. For TVB-N and TVC indicators showed that the predictive ability of this model was largely comparable to that of fluorescence hyperspectral imaging. However, combining fluorescence and color imaging improved the model's predictive ability. For TVB-N, TVC and pH, Rp were 0.94, 0.93 and 0.85, RPD were 2.62, 2.59, and 1.95, respectively. Therefore, this study developed a system with great potential for detecting the value of most pork quality indicators in real-time.

Extending the Shelf Life of Fresh Khalal Barhi Dates via an Optimized Postharvest Ultrasonic Treatment.

The Barhi date is a high-quality date cultivar whose fruits (dates) are plucked and eaten fresh when they reach the Khalal maturity stage due to their sweetness, crispiness, and yellow skin color. After harvesting, Khalal Barhi fruits rapidly matured to the Rutab stage, where their tissues become soft and their skin color browner. This results in a decrease in their market value and customer demand. This study aims at investigating the effectiveness of the postharvest ultrasonic treatment in conserving the physical, microbial, and nutritional quality of Barhi fruits and extending their shelf life. To achieve the goals of the present work, the response surface methodology (RSM) was used for the optimization of the ultrasonic intensity (50, 100, 150, and 200 W/cm2) and application time (5, 10, 15, and 20 min) to preserve the Barhi dates high quality features for varied storage temperatures (1, 5, 15, and 25 °C) and duration (1, 6, 16, and 21 days). In RSM, a four-factors-mixed-levels central composite rotatable design (CCRD) was applied to optimize the ultrasound treatment and storage environments for better-quality physical [total soluble solids (TSS), firmness, and total color changes (ΔE)], microbial [total viable count (TVC)], nutritional [total phenolic content (TPC), DPPH antiradical activity, glucose, and fructose] features of Barhi dates. The outcomes showed that ultrasound intensity and its application time, storage temperature, and storage period influence the physical, microbial, and nutritional quality attributes in different magnitudes. The ideal settings for lessening the changes in the physical attributes, eliminating the microbial growth, and improving the nutritional quality attributes were 140 W/cm2, 5.2 min, 20.9 °C, and 21 days for ultrasound intensity, ultrasound exposure duration, storage temperature, and storage duration, respectively. In conclusion, this study proved the potential application of ultrasound for persevering the excellence aspects of Barhi dates and identified the ideal ultrasound environments for maintaining the physical, microbial, and nutritional quality features of Barhi dates during extended storing.

Preliminary Results on the Comparative Evaluation of Alkaline Phosphatase Commercial Tests Efficiency in Non-Cow Milk Pasteurization.

The demand for non-cow milk and the products derived from it, is constantly increasing; thus, correct and effective pasteurization becomes necessary. Typical practices for evaluating milk pasteurization are mainly based on the thermal inactivation of an endogenous enzyme, alkaline phosphatase (ALP). The ALP tests, originally designed and applied to pasteurized cow milk, are often used to control pasteurization in non-cow milk, without sufficient data on their suitability; EFSA calls on the scientific world for collecting more information on the subject. In this study, the pertinent details of the ALP assay for non-cow milk products are summarized, and a comparison is performed regarding the evaluation of the adequacy of commercially available tests for the determination of ALP activity in non-cow milk. At the same time, raw and pasteurized non-cow milk was analyzed microbiologically using standard ISO methods and MALDI-TOF MS in order to confirm the thermal effect on common microorganisms. In these preliminary results, various ALP tests do not appear to be fully reliable as indicators for the pasteurization of some types of non-cow milk such as camel and donkey milk or even goat and sheep milk, using the EFSA proposed limits. ALP commercial kits may not be suitable as pasteurization indicators for various types on non-cow milk, and alternatives should be investigated.

Detection of viable Lacticaseibacillus paracasei in fermented milk using propidium monoazide combined with quantitative loop-mediated isothermal amplification.

To quantify viable probiotic Lacticaseibacillus paracasei (L. paracasei) in fermented milk accurately and quickly, propidium monoazide combined with quantitative loop-mediated isothermal amplification (PMA-qLAMP) was applied. The optimal PMA treatment conditions for treating a L. paracasei suspension were determined using an orthogonal test to eliminate the DNA amplification of 108 CFU/mL of dead L. paracasei. Primers were designed based on the species-specific gyrB gene of L. paracasei. A phylogenetic tree based on the gyrB gene showed that L. paracasei clustered on the same branch with 91% support. Compared with the 16 strains commonly found in fermented milk, three strains of L. paracasei showed positive PMA-qLAMP results, and the melting temperature was approximately 82.4°C. There was a linear relationship (R2 = 0.9983) between the Ct values and the logarithm of the concentration of viable bacteria. The PMA-qLAMP detection limit for the L. paracasei artificially added to fermented milk was 7.3 × 102 CFU/mL. There was no significant difference between the logarithm values of the concentration of viable L. paracasei of 50 fermented milk samples within shelf life using the PMA-qLAMP and plate count methods (P > 0.01). PMA-qLAMP is specific and accurate for obtaining reliable results faster than when using plate counts.

Bacteriological quality of raw milk marketed in and around Guwahati city, Assam, India.

BACKGROUND AND AIM: Milk is a highly perishable commodity, which is subjected to various types of contamination right from the farm level to the consumers' table. This study aimed to assess the quality of raw milk sold in and around Guwahati city based on the microbial load. MATERIALS AND METHODS: A total of 200 raw pooled milk samples collected from 25 different locations in and around Guwahati city were subjected to quality evaluation based on the methylene blue reduction test (MBRT), standard plate count, and coliform count as per the standard procedure. RESULTS: Out of the 200 samples evaluated, more than 50% of them were graded as poor to very poor quality based on the MBRT results. None of the samples could be graded as excellent quality and only 14.5% were graded as good quality. The standard plate count and coliform count of all the raw milk samples were found to be significantly higher than the legal standard. A highly significant (p<0.01) difference was observed for standard plate count and coliform count among the different locations in and around Guwahati city. CONCLUSION: From the present study, it could be inferred that raw milk sold in most parts of Guwahati city do not confer to the legal microbiological standard and may pose a high risk of milk-borne illness among consumers of the city, which needs a systematic series of actions to be implemented properly.