Enumeration Of Lactic Acid Bacteria Research Articles

Development of fermented beverage with citrus fruit extract using probiotics: impact on antioxidant activity and in vitro digestibility

Redhyang (Citrus hybrid ‘Kanpei’,CHK) is a subtropical citrus species introduced in Korea due to climate change. To enhance the nutritional value and usability of CHK as a processed food product, CHK extract was fermented with four types of commercial starters (YoFlex Harmony 1.0 (YFH), ABY-3 (ABY), YC-X11 (YXC), and YC-180 (YC)), and their antioxidant activities and changes in chemical properties during fermentation were investigated. The consumer acceptance of probiotic beverages containing fermented CHK extracts and their viability and antioxidant activity through in vitro digestion were also elucidated. The enumeration of lactic acid bacteria (LAB) in all samples after fermentation was above 7.60 log colony-forming units (CFU)/mL, with YC exhibiting the highest number after 24 h. Fermented CHK extracts containing higher levels of organic acids, total polyphenols, and flavonoids tended to exhibit higher antioxidant activities. YFH, ABY, and YC showed maximum antioxidant activity at 24 h, whereas YXC showed differences in the types of LAB at 12 h. After in vitro digestion, YXC showed higher antioxidant activity and LAB viability than the control. This result indicates that CHK extract fermented with YXC can increase antioxidant activity, bioactive ingredients, and sensory preference and positively impact the production of probiotic beverages.

Test of Antioxidants and Alpha-glucosidase Inhibitors from Lactic Acid Bacteria in Fermentation of Soy Milk (Soygurt)

Soygurt is an innovative product from yogurt which is fermented from soy milk. This research is an experimental study performed by an in vivo approach. The disc diffusion method evaluated the antibacterial activity of lactic acid bacteria isolates from Soygurt. Meanwhile, the antioxidant activity and inhibition of α-glucosidase enzymes were evaluated from cell-free supernatants of lactic acid bacteria isolated by spectroscopic methods. The results of the enumeration of lactic acid bacteria in Soygurt with 5% Lactobacillus acidophilic yogurt starter were 8.00 x 104 CFU/ml, and the calculation of lactic acid bacteria in Soygurt with Yakult starter was 5.00 x 107 CFU/ml. Lactic acid bacteria isolates from Soygurt with 5% Lactobacillus acidophilic starter showed no antibacterial activities against Staphylococcus aureus bacteria by the disc diffusion method. However, this lactic acid bacteria isolate had antioxidant activity and inhibited the α-glucosidase enzyme. It was shown by the percentage of DPPH inhibition, which ranges from 27.06-58.63%. Meanwhile, the moderate inhibitory activity of the α-glucosidase enzyme was 50.55%. Therefore, it can be concluded that lactic acid bacteria isolated from Soygurt with 5% Lactobacillus acidophilic yogurt starter have antioxidant and antidiabetic activity.

Sustainability of food side streams: a case study of fermented blends made with sour whey and sunflower press cake powder using the back-slopping technique

The exploitation of by-products is a key factor to increase the sustainability of the agri-food chain and fermentation is a simple and eco-friendly process for achieving safe and suitable food materials. In this study, we investigated the possibility to manage a spontaneous fermentation of blends made with different proportions of two food side streams (bovine acid whey and sunflower press cake powder) through the application of a back-slopping technique of the mixed material incubated at 26°C in static conditions. A full-factorial 2-factor 3-level design of experiment was applied to infer the effect of the percent (w/w) of press cake powder in the mixture (20, 25, and 30%) and the rate of back-slopping inoculum (15, 30, and 45%). The pH value, titratable acidity, content of sugars, organic acids, and phenolic acids, enumeration of lactic acid bacteria, yeasts and molds, bacterial contaminants, presumptive Bacillus cereus, and Escherichia coli were measured for each fermentation step at 0, 24, 48, and 72 h. On the same samples, a metataxonomics analysis, targeted on bacterial 16S rRNA gene and fungal ITS region, was performed by using the Illumina MiSeq platform. Acidification of the blends (on average, starting pH = 5.45 ± 011, final pH = 4.61 ± 0.11; starting acidity =13.68 ± 1.02 °SH/50 mL, final acidity = 28.17 ± 2.92°SH/50 mL) and high LAB counts (on average, 9.39 log CFU/g ± 0.25) were observed at the end of each refreshment. In all fermented mixtures, B. cereus, E. coli, and molds counts were lower than the detection limit (<2 log CFU/g), whereas bacterial contaminants, overall spore-formers, were always present (3.74 log CFU/g ± 0.27). After 72 h, the dropping of pH value was maximum, yielding significant differences compared to previous fermentation steps (p < 0.01); particularly, the lowest pH (4.45 ± 0.06) was achieved in the central points of DoE (25% of press cake powder and 30% of back-slopping rate), representing the most suitable condition. Results from both culture-dependent and -independent techniques were consistent; although Lactococcus lactis, continuously deriving from the acid whey, was the main LAB, Pediococcus pentosaceus appeared and, in some cases, became the dominant species. Finally, a long-term trial (about 1 month), using the best condition previously pointed out, was performed with an extension of the incubation time to 84 h for each refreshment. The increase in acidity forced the natural selection toward acid-tolerant microbial strains confirming the former results. Although preliminary, these findings can be useful for developing innovative operations to manage these two relevant side streams implementing the circularity of food resources.

Inoculation with heterofermentative strains <i>Lentilactobacillus buchneri</i> CNCM 40788 and <i>Lentilactobacillus hilgardii</i> CNCM I-4785 either alone or combined improves fermentation and aerobic stability of ensiled triticale (X-triticosecale)

<abstract> <p>Triticale, a small-grain forage, was harvested for an ensiling experiment evaluating different silage inoculants. Fresh material (mean dry matter 404.1 g kg<sup>−1</sup>) was wilted, chopped, and assigned to one of four treatment groups including water only (Control), heterofermentative strains <italic>Lentilactobacillus buchneri</italic> (LB), <italic>Lentilactobacillus hilgardii</italic> (LH), or combination (Combo) of both. Inoculants were applied at 4.0 x 10<sup>5</sup> CFU per g of fresh forage, and the Combo contained both strains at 2.0 x 10<sup>5</sup> CFU per g. Treated forage was packed into 7.57 L mini-silos for openings after 15, 30, and 130 d. Samples were collected at each opening for microbial enumeration of lactic acid bacteria, enterobacteria, yeasts, and moulds. Additional samples were collected for analysis of fermentation profiles, and nutritional analysis following dry matter determination and grinding. Aerobic stability was also evaluated at each opening through a 10-d period. Fermentation profiles were notably affected, including increases in acetic acid (g/kg DM) in LB and Combo treated silage after 15 d which resulted in reduced pH. Aerobic stability was vastly improved through inoculation by prolonging the time to reach 2˚ C above ambient, which was likely a result of decreased yeast counts. Our research validates the use of combined heterofermentative lactic acid bacteria strains on an ensiled small-grain specie, and further agrees with previous findings of prolonged aerobic stability through increases in lactate and acetate in response to co-inoculation.</p> </abstract>

Microbiological Biodiversity of Regional Cow, Goat and Ewe Milk Cheeses Produced in Poland and Antibiotic Resistance of Lactic Acid Bacteria Isolated from Them.

(1) Unique sensory values of traditional and regional dairy products made them more and more popular among consumers. Lactic acid bacteria naturally occurring in these products can express antibiotic resistance and be a reservoir of antibiotic resistance genes (ARG) in the environment. The aim of the study was to characterize the microbial diversity of twenty regional cheeses produced from non-pasteurized cow, goat and ewe milk, and investigate the phenotypic and genotypic antibiotic resistance (AR) of lactic acid bacteria isolated from these products. (2) Conventional microbiological methods were applied for the enumeration of lactic acid bacteria (lactobacilli and lactococci) and their isolation, and for the enumeration of Enterococcus, Staphylococcus, Enterobacteriaceae and spores. The disc diffusion method was applied for phenotypic AR. The PCR-based methods were used for strain identification, microbiological diversity of cheeses (PCR-DGGE), and for AR gene detection. (3) Among 79 LAB isolates the most frequent species were L. plantarum (n = 18), Leuc. lactis (n = 17), Lc. lactis (n = 11), Leuc. mesenteroides (n = 9) and L. pentosus (n = 8). Additionally, by using the PCR-DGGE method, DNA of L. casei was found in nine products. Lactobacilli (5.63-8.46 log cfu/g) and lactococci (6.15-8.41 log cfu/g) predominated over Enterococcus (max. 4.89 log cfu/g), Staphylococcus (max. 4.18 log cfu/g), and Enterobacteriaceae (mostly up to 4.88 log cfu/g). Analysis of phenotypic resistance to tetracycline (30 µg), erythromycin (15 µg), and chloramphenicol (30 µg) showed that 29% of LAB isolates were resistant to one antibiotic, 8%-to two, and 12%-to all tested antibiotics. Antibiotic resistance genes (AGR) for tetracycline (tet(M), tet(L), tet(W)), erythromycin (erm(B)) and chloramphenicol (cat-TC) were detected in 30 (38%), 29 (36.7%) and 33 (43.4%) LAB isolates, respectively. Among 31 LAB isolates phenotypically susceptible to all tested antibiotics, only 5 (16%) had no ARGs. (4) The results obtained in our work shed light on the potential threat posed by the widespread presence of ARGs in LAB present in regional cheeses.

Development of a selective medium for the enumeration of lactic acid bacteria and bifidobacteria in food products.

The online version contains supplementary material available at 10.1007/s10068-022-01202-z.

Survival of some food-borne bacteria in kefir produced by microbial levan and pullulan

Kefir is a fermented milk product produced by kefir grains traditionally or lyophilized starter culture. Some quality characteristics of kefir produced at 22 °C and 30 °C using kefir grains after addition of microbial levan and pullulan were investigated for 14 days. Additionally, the survival of different Gram-positive and Gram-negative bacteria strains which were inoculated before and after fermentation were investigated during storage. The pH and titratable acidity were in the ranges of 4.67–4.39 and 1.10–0.87%, respectively. The highest exopolysaccharide values (458.3–465.3 mg/kg) were found in the samples produced with the addition of 1% (w/v) of microbial levan at 30 °C. Yeast counts were in the ranges of 6.73–5.11 log CFU/mL while the lactobacilli and lactic streptococci counts were in the ranges of 9.64–7.91 and 9.64–8.69 log CFU/mL, respectively. The microbial polysaccharide addition did not show any significant differences in the enumeration of lactic acid bacteria in kefir (p > 0.05). All Gram-positive and Gram-negative bacteria grew/survived during fermentation however Listeria monocytogenes was the most susceptible test bacterium to metabolites during storage. The highest reduction in the counts of L. monocytogenes was determined as 2.39 log units after 24 h of fermentation at 30 °C in the samples which were inoculated before fermentation and produced with the addition of pullulan. Kefir samples produced at 30 °C without polysaccharide addition were considered as the best with respect to appearance, taste, consistency and overall acceptability.

Simple and rapid method for selective enumeration of lactic acid bacteria in commercially prepared yogurt by image analysis and K-means clustering.

We developed a simple and rapid method based on the combination of image analysis and k-means clustering to selectively enumerate cocci and bacilli from among lactic acid bacteria (LAB) in commercially prepared yogurt. We used our previously reported method for recovering only LAB without non-microbial substances from commercial yogurt, and found that the shape and light intensity of LAB cell images taken by optical microscopy were factors that could distinguish cocci and bacilli, allowing the selective enumeration of LAB. Also, k-means clustering was executed on a dataset of the mean light intensity and aspect ratio of each LAB obtained by image analysis, and each LAB in the image could be automatically assigned to either the cocci or bacilli group. The results obtained by this automated method were in good agreement with those obtained by manually counting the LAB under a microscope, with an overall error within 10%. In addition, this method could provide results within a few hours, which is approximately 1/32 of the time required for the conventional colony-counting method.

Effect of Using Probiotic Bacteria as Adjunct Culture in Domiati Cheese Making

The composition and quality of probiotic Domiati cheese were studied during 90 days of ripening in brine. Six cheese treatments were made using different types of cultures, which are: C (Control): Lactococcus lactis subsp. lactis and Lactococcus lactis subsp. cremoris, T1: as control + Lactobacillus casei, T2: as control +Lactobacillus acidophilus, T3: as control + Lactococcus lactis biovar diacetylactis, T4:Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus and T5:Lactobacillus acidophilus, Bifidobacterium bifidum and Streptococcus thermophilus. A gradual decrease in moisture content was observed as pickling proceeded. Titratable acidity, salt and fat content continuously increased as storage proceeded in all treatments. The soluble nitrogen and amino nitrogen were higher in the probiotic cheese, compared with the control. Enumeration of lactic acid bacteria (LAB) increased up to 45 days of storage and started to decrease with prolonging the storage period. The highest numbers of total bacteria and LAB were detected in T1 and T4, respectively, at the end of the storage, while the lowest was found in T5and T3, respectively. All fresh cheese treatments were found completely free of yeasts and moulds, while it started to appear after 15 days of storage. Absence of coliform bacteria was detected in all treatments. Higher overall scores were achieved in all treatments with added probiotic bacteria compared with control at the end of the storage period. Using probiotic bacteria as adjuncts enhanced the quality and improved the organoleptic properties of the resultant cheese.

Isolation and Characterization of Lactic Acid Bacteria and Yeasts from Typical Bulgarian Sourdoughs

Traditional sourdoughs in Bulgaria were almost extinct during the centralized food production system. However, a rapidly developing trend of sourdough revival in the country is setting the demand for increased production and use of commercial starter cultures. The selection of strains for such cultures is based on geographical specificity and beneficial technological properties. In this connection, the aim of this study was to isolate, identify and characterize lactic acid bacteria (LAB) and yeasts from typical Bulgarian sourdoughs for the selection of strains for commercial sourdough starter cultures. Twelve samples of typical Bulgarian sourdoughs were collected from different geographical locations. All samples were analyzed for pH, total titratable acidity and dry matter content. Enumeration of LAB and yeast was also carried out. Molecular identification by 16S rDNA sequence analysis was performed for 167 LAB isolates, and 106 yeast strains were identified by ITS1-5.8S-ITS2 rRNA gene partial sequence analysis. The LAB strains were characterized according to their amylolytic and proteolytic activity and acidification capacity, and 11 strains were selected for further testing of their antimicrobial properties. The strains with the most pronounced antibacterial and antifungal activity are listed as recommended candidates for the development of starter cultures for sourdoughs or other food products.

Evaluation of the Estimation Capability of Response Surface Methodology and Artificial Neural Network for the Optimization of Bacteriocin-Like Inhibitory Substances Production by Lactococcus lactis Gh1.

Bacteriocin-like inhibitory substances (BLIS) produced by Lactococcus lactis Gh1 had shown antimicrobial activity against Listeria monocytogenes ATCC 15313. Brain Heart Infusion (BHI) broth is used for the cultivation and enumeration of lactic acid bacteria, but there is a need to improve the current medium composition for enhancement of BLIS production, and one of the approaches is to model the optimization process and identify the most appropriate medium formulation. Response surface methodology (RSM) and artificial neural network (ANN) models were employed in this study. In medium optimization, ANN (R2 = 0.98) methodology provided better estimation point and data fitting as compared to RSM (R2 = 0.79). In ANN, the optimal medium consisted of 35.38 g/L soytone, 16 g/L fructose, 3.25 g/L sodium chloride (NaCl) and 5.40 g/L disodium phosphate (Na2HPO4). BLIS production in optimal medium (717.13 ± 0.76 AU/mL) was about 1.40-fold higher than that obtained in nonoptimised (520.56 ± 3.37 AU/mL) medium. BLIS production was further improved by about 1.18 times higher in 2 L stirred tank bioreactor (787.40 ± 1.30 AU/mL) as compared to that obtained in 250 mL shake flask (665.28 ± 14.22 AU/mL) using the optimised medium.

Effect of Fermentation Media on the Quality of Arabica Wine Coffee

The fermentation of arabica wine coffee from the cherry is one of the methods in processing the coffee which has the wine taste. The selection of media of fermentation is also one of the factors influencing the taste of the arabica wine coffee. The difference in the fermentation process of the coffee bean affects the taste and quality of the coffee. The fermentation process can take place in the aerobic, anaerobic and semi-aerobic state. The process is a fermentation using a glass bottle, can, and a plastic sack as the media and kept in the storage for seven days. After the fermentation, the coffee was dried for 21 days until the moisture content reaches 10% and the total bacteria were enumerated. The enumeration of Lactic Acid Bacteria (LAB) which was conducted before and after fermentation with a dilution factor of 10-1 to 10-6 shows that the total LAB tends to decrease along with the higher factor of dilution. This result comes from the dilution factor of 10-4 to 10-6 with the total bacteria of 2 to 8 colonies depends on the number of dilutions. The sample analysis used was cupping test which gives the result that the coffee aroma of ages, woody, strawy, fermented, clean and fresh, which is the characteristics of wine taste. The best container obtained in this research is the coffee which fermented in a glass bottle and can. Meanwhile, the coffee fermented in a plastic sack was highly contaminated when the fermentation occurs.

The variation of ethanol concentration and kombucha characterization on several incubation periods

Kombucha is a type of fermented tea that recently popular in Indonesia. It gives so many health benefits for human body, as prevention and therapeutic cure for the disease. Some of the benefits are for improving digestion and reducing the risk of cancer and cardiovascular disease. It also has significant potential as an antioxidant agent. The heterofermentative fermentation process in kombucha causes this product to produce organic acid and ethanol as an alcohol compound. However, ethanol concentration in fermented food has become a major concern in Indonesian Halal products studies. One cause of ethanol concentration differences is incubation times. In this study, we measured alcohol concentration at several incubation times: 5, 6, 7 and 8 days incubation. We also assessed characteristics of kombucha by testing its antioxidant activity, lactic acid content, pH, and enumeration of lactic acid bacteria. The result shows that the incubation time, from 5 to 8 days, did not produce alcohol more than 0,5% as restricted by MUI Fatwa 2018. It has been shown that the various time of incubations did not give a significant difference in the quality of the antioxidant activity and lactic acid levels, and pH (p>0,05).

3M Petrifilm Lactic Acid Bacteria Count Plate Is a Reliable Tool for Enumerating Lactic Acid Bacteria in Bacon

This study aimed to evaluate the behavior of Petrifilm Lactic Acid Bacteria Count Plates (PLAB) as an alternative methodology to enumerate lactic acid bacteria (LAB) in bacon. Bacon samples (n = 40) were obtained from retail sale, 10-fold diluted with buffered peptone water (BPW, 0.2% [w/v]) and Letheen broth, and subjected to LAB enumeration according to four protocols: (i) de Man Rogosa Sharpe (MRS) agar, pH 5.7, 30°C; (ii) MRS, pH 5.7, 30°C, anaerobiosis; (iii) all-purpose Tween agar (APT), 25°C; and (iv) PLAB, 30°C. Colonies were enumerated at 24, 48, and 72 h, and the results expressed as log CFU per gram for comparison by analysis of variance and regression (P < 0.05). Furthermore, colonies were randomly selected and characterized as LAB (Gram staining and catalase). Mean LAB counts from MRS and PLAB did not present significant differences independently of incubation time or diluent (P > 0.05), whereas counts in APT with BPW after 24 h were significantly lower (P < 0.05). PLAB counts with BPW (24, 48, and 72 h) presented significant correlation with MRS (r ranging from 0.87 to 0.89; in anaerobiosis, r ranging from 0.94 to 0.95) and APT (r ranging from 0.84 to 0.86). With Letheen broth, PLAB (24, 48, and 72 h) presented significant correlation with MRS (r ranging from 0.92 to 0.94; in anaerobiosis, r ranging from 0.93 to 0.96) and APT (r ranging from 0.77 to 0.79). In total, 1,032 colonies (97%) from 1,063 colonies were characterized as LAB. Thus, PLAB can be considered as an alternative tool for enumerating LAB in bacon, with reliable results even after 24 h of incubation.

Microbiological and Chemical Quality of Portuguese Lettuce-Results of a Case Study.

In addition to environmental pollution issues, social concerns about the sustainability, safety, and quality of conventionally grown fruits and vegetables have been increasing. In order to evaluate if there were any microbiological differences between samples of organic and conventional lettuce, a wide range of parameters were tested, including pathogens and indicator organisms: the enumeration of Escherichia coli; the detection of Salmonella spp.; the detection/enumeration of Listeria monocytogenes; the enumeration of lactic acid bacteria, Pseudomonas spp. yeasts and molds, and Enterobacteriaceae. This study also evaluated the chemical safety of the lettuce samples, quantifying the nitrate concentration and 20 pesticides (14 organochlorine and 6 organophosphorus pesticides). Significant differences (p < 0.05) between the conventional and organic samples were only detected for the counts of total microorganisms at 30 °C. Pathogens were absent in all the samples. The analytical method, using the quick, easy, cheap, effective, rugged, and safe (QuEChERS) approach for pesticide extraction, was suitable for detecting the targeted analytes; the limit of quantification (LOQ) was between 0.6 and 1.8 µg/kg (lower than the Maximum Residue Levels (MRLs) established by EU legislation). In three organic lettuce samples, one organochlorine pesticide (α-HCH) was observed below the MRLs. For the samples analyzed and for the parameters investigated, except for the total mesophilic counts, the organic and conventional lettuces were not different.

The complex microbiota of artisanal cheeses interferes in the performance of enumeration protocols for lactic acid bacteria and staphylococci

We assessed the interference of a complex food microbiota, present in Minas artisanal cheese (MAC) produced in the Serro region, on the performance of enumeration protocols for lactic acid bacteria (LAB) and staphylococci. Initially, 140 isolates were obtained from 6 Serro MAC samples and plated for enumeration of LAB (3 variations of MRS, M17 and Petrifilm™ LAB) and staphylococci (Baird–Parker and Petrifilm™ STX); some target isolates did not grow or present atypical morphologies in their selective protocols. Then, Serro MAC samples (n = 55) were obtained and plated for LAB and staphylococci based on the same protocols: 2569 isolates were obtained from LAB and 1164 from staphylococci protocols: Petrifim™ LAB presented the highest frequencies of LAB-confirmed colonies, as Petrifilm™ STX for staphylococci (p < 0.05).

Isolation and identification of lactic acid bacteria from the intestinal tracts of honey bees, Apis mellifera L., in Egypt

The objective of this study was enumeration and identification of lactic acid bacteria (LAB) from honey bee (Apis mellifera L) and testing their antibacterial activity against different types of bacteria. Dilution method and cultivation in selective media were used for enumeration of LAB, the isolates were identified by their morphological and biochemical characteristics and by genotype method. The antibacterial activity was performed using the agar well diffusion method. The results showed that five isolates of LAB were isolated from Egyptian honey bees intestinal tract. The isolates were identified as Enterococcus faecalis MG890204, Enterococcus faecalis KX073783, Enterococcus faecalis EU594564, Lactobacillus brevis MH191230 and Lactobacillus casei KT273339. The results of antibacterial activity showed that four of LABs had antibacterial activity against clinical pathogenic strains tested. The mechanisms of action are partly shown by elucidating the production of some active compounds like organic acids in different amounts, Lactic, Oxalic, Glutaric, and acetic acids were produced by all LABs. The findings revealed that honey bee has the potential to be a source of new bacteria with probiotic activities in honey bee or as natural food preservatives for the food industry.

Enumeration of lactic acid bacteria: lacuna and improvement areas highlighted by proficiency testing

During the 2011 to 2018 food microbiology proficiency testing (PT) schemes organized by REQUASUD, a systematic analytical issue was detected. For the enumeration of the lactic acid bacteria (LAB), laboratories carrying out the same analytical method (ISO 15214) yielded completely incoherent results while analysing naturally contaminated samples (raw milk, smoked salmon and ham sausage), leading to an atypically large distribution of the PT results. A comprehensive study of these atypical PT results provided insight into the source of systematic errors affecting the LAB enumeration. The main sources of analytical variability turned out to be linked to the ISO 15214:1998 standard. To better harmonize the analytical results, practical improvements can be proposed for the revision of this standard, mainly: the addition of precise criteria for the colony count, a restriction of the allowed incubation conditions and the use of a more selective culture medium (e.g. supplemented with sorbate) to limit the growth of interfering colonies. PT is a privileged observation post to highlight such analytical issues, which would otherwise remain unnoticed. Whenever an unusual distribution of PT results is observed (e.g. high dispersion or non-normal distribution of the values), it is the responsibility of the PT provider to try to identify the causes of non-normality, in a perspective of method improvement. The statistical analysis of atypical PT results requires specific attention: ignoring the non-normality of PT results when it occurs can lead to set an erroneous assigned value, unfairly penalizing participants.

Identification and characterization of lactic acid bacteria isolated from some medicinal and/or edible Philippine plants

Probiotics research on lactic acid bacteria (LAB) continues to be paramount in the development of nutraceutical or functional foods. In this study, 47 selected Philippine plants having nutritional (edible such as vegetables) and/or medicinal values (therapeutic), were collected, from which selected fruit and leaves were subjected to LAB enumeration. Among these, 46.7% plant leaves reported to have strong antimicrobial property resulted in non-isolation of LAB while edible plant leaves with less or no antimicrobial properties generally gave numerous LAB isolates. Isolates coming from ripened guava (Psidium guajava L.), lobo-lobohan or cape gooseberry (Physalis peruviana L.) fruit, parsley (Petroselinum crispum), pandan (Pandanus amaryllifolius), spinach (Spinacea oleracea), leek (Allium ampeloprasum var. porrum) and niyog-niyogan (Quisqualis indica L.) leaves were identified through partial 16S rRNA analysis and tested for probiotic properties. Overall, Streptococcus luteciae Lb17 from ripe gooseberry fruit exhibited the highest antimicrobial activities against Staphylococcus aureus BIOTECH 1526, Escherichia coli O157: H7 BIOTECH 10311 and Bacillus cereus BIOTECH 1509. On the other hand, Enterococcus hirae (H and S63) from stevia were susceptible to streptomycin at minimum inhibitory concentration or MIC of 128 ug/mL). Lactobacillus plantarum F39 and all the other strains tested, meanwhile, was susceptible to ampicillin at MIC of 2 ug/mL and 0.125 ug/mL. Results were lower or equal to the established cut off value indicating the absence of antibiotic resistance genes among the identified strains, except for Pediococcus (Par5 and NN39) which showed resistance against streptomycin. Further investigation is needed to rule out the possibility of transfer of antibiotic resistance to pathogens present in the gut. All isolates tested were able to survive at artificial gastric juice (pH 2), revived at the simulated intestinal fluid (pH 8), and exhibited minimal titratable acidity and diacetyl production. For genetic screening of plantaricin genes, F39 possesses both plantaricin EF and plantaricin J, while Lactobacillus fermentum F36 has plantaricin EF. Both isolates were subjected to DNA fingerprinting. Such findings on the local isolates’ probiotic properties suggest the possibility of incorporating them into different plant-based probiotic foods.

Microbial analysis of commercially available US Queso Fresco

Queso Fresco (QF), a fresh Hispanic-style cheese, is often associated with Listeria monocytogenes outbreaks and recalls. Queso Fresco's susceptibility to bacterial contamination is partially due to its high pH and moisture content as well as Listeria's tolerance for the salt content typical for QF. Nine different brands of US QF, 2 packages from 4 different lots (to account for temporal variability), were sampled. The pH, salt content, and moisture content were analyzed in addition to microbial testing including yeasts and molds, coliforms, lactic acid bacteria enumeration, and L. monocytogenes counts. The cheeses were also inoculated with a cocktail of 5 food and human isolates of food-borne outbreak-associated Listeria monocytogenes strains to evaluate how the differences between brands influenced Listeria growth. Three of the cheeses underwent additional genus-level microbial analysis using extracted 16S rDNA, allowing for phylogenetic analysis between bacterial taxa including diversity and relative abundance. We found little variation between the sampled QF pH (range = 6.62-6.86), salt content (1.53-2.01%), and moisture content (43.90-54.50%). Yeasts and molds were below the detection limit of enumeration in all of the cheeses and coliforms were below the detection limit across the first 3 lots, but were detected at varying levels in the fourth lot (>3.0 most probable number/g) for 3 of the brands. Listeria monocytogenes was not isolated after enrichment in any of the samples. All cheeses tested positive for the presence of lactic acid bacteria, with only 1 of the cheeses being labeled as produced with added cultures having substantial counts. Fourteen days after inoculation with L. monocytogenes, at least 2.5 log10 cfu/g of growth was found for all QF brands stored at 4°C. Microbial genus analysis showed that, among the 3 brands, the microbial community was more similar within brand than when compared with the other 2 brands. Thermus, Anoxybacillus, and Streptococcus accounted for the dominant genera of brands A, B, and C, respectively. These variations within the microbial community may account for sensory differences and help manufacturers determine quality control consistency more readily than culture-based methods.