Phylotype-Level Characterization of Complex Communities of Lactobacilli Using a High-Throughput, High-Resolution Phenylalanyl-tRNA Synthetase (pheS) Gene Amplicon Sequencing Approach.

The lactobacilli identified to date encompass more than 270 closely related species that were recently reclassified into 26 genera. Because of their relevance to industry, there is a need to distinguish between closely related and yet metabolically and regulatory distinct species, e.g., during monitoring of biotechnological processes or screening of samples of unknown composition. Current available methods, such as shotgun metagenomics or rRNA gene-based amplicon sequencing, have significant limitations (high cost, low resolution, etc.). Here, we generated a phylogeny of lactobacilli based on phenylalanyl-tRNA synthetase (pheS) genes and, from it, developed a high-resolution taxonomic framework which allows for comprehensive and confident characterization of the community diversity and structure of lactobacilli at the species level. This framework is based on a total of 445 pheS gene sequences, including sequences of 276 validly described species and subspecies (of a total of 282, including the proposed L. timonensis species and the reproposed L. zeae species; coverage of 98%), and allows differentiation between 265 species-level clades of lactobacilli and the subspecies of L. sakei The methodology was validated through next-generation sequencing of mock communities. At a sequencing depth of ∼30,000 sequences, the minimum level of detection was approximately 0.02 pg per µl DNA (equaling approximately 10 genome copies per µl template DNA). The pheS approach, along with parallel sequencing of partial 16S rRNA genes, revealed considerable diversity of lactobacilli and distinct community structures across a broad range of samples from different environmental niches. This novel complementary approach may be applicable to industry and academia alike.IMPORTANCE Species formerly classified within the genera Lactobacillus and Pediococcus have been studied extensively at the genomic level. To accommodate their exceptional functional diversity, the over 270 species were recently reclassified into 26 distinct genera. Despite their relevance to both academia and industry, methods that allow detailed exploration of their ecology are still limited by low resolution, high cost, or copy number variations. The approach described here makes use of a single-copy marker gene which outperforms other markers with regard to species-level resolution and availability of reference sequences (98% coverage). The tool was validated against a mock community and used to address diversity of lactobacilli and community structure in various environmental matrices. Such analyses can now be performed at a broader scale to assess and monitor the assembly, structure, and function of communities of lactobacilli at the species level (and, in some cases, even at the subspecies level) across a wide range of academic and commercial applications.

Composition of lactic acid bacteria during spontaneous curly kale (Brassica oleracea var. sabellica) fermentation.

The present work is the first report on spontaneous fermentation of curly kale and characteristics of autochthonous lactic acid bacteria (LAB). Our results indicate that curly kale fermentation is the new possibility of the technological use of this vegetable. Bacteria representing ten different species were isolated from three phases of curly kale fermentation and identified by MALDI-TOF mass spectrometry and 16S rRNA gene sequencing. Among them, four species were identified as Lactobacillus spp. (Lb. plantarum 332, Lb. paraplantarum G2114, Lb. brevis R413, Lb. curvatus 154), two as Weissella spp. (W. hellenica 152, W. cibaria G44), two as Pediococcus spp. (P. pentosaceus 45AN, P. acidilactici 2211), one as Leuconostoc mesenteroides 153, and one as Lactococcus lactis 37BN. The functional properties of isolates, i.e. acid, NaCl and bile salt tolerance, enzyme activities, adhesion to hydrocarbons, and antibiotic resistance, were examined. Among the tested strains, Lb. plantarum 332, Lb. paraplantarum G2114, P. pentosaceus 2211, and Lb. brevis R413 exhibited the best hydrophobicity value and high tolerance to bile salts, NaCl, and low pH.

Sourdough-type propagation of faba bean flour: Dynamics of microbial consortia and biochemical implications.

The microbial ecology of faba bean sourdoughs obtained from an Italian (Ita) and a Finnish (Fi) cultivar, belonging respectively to Vicia faba major and V. faba minor groups, was described by 16S rRNA gene pyrosequencing and culture-dependent analysis. The sourdoughs were propagated with traditional backslopping procedure throughout 14days. Higher microbial diversity was found in the sourdough deriving from V. faba minor (Fi), still containing residual hulls after the milling procedure. After 2days of propagation, the microbial profile of Ita sourdough was characterized by the dominance of the genera Pediococcus, Leuconostoc and Weissella, while the genera Lactococcus, Lactobacillus and Escherichia, as well as Enterobacteriaceae were present in Fi sourdoughs. Yeasts were in very low cell density until the second backslopping and were not anymore found after this time by plate count or pyrosequencing analysis. Among the lactic acid bacteria isolates, Pediococcus pentosaceus, Leuconostoc mesenteroides and Weissella koreensis had the highest frequency of occurrence in both the sourdoughs. Lactobacillus sakei was the only lactobacillus isolated from the first to the last propagation day in Fi sourdough. According to microbiological and acidification properties, the maturity of the sourdoughs was reached after 5days. The presence of hulls and the different microbial composition reflected on biochemical characteristics of Fi sourdoughs, including acidification and phenolic compounds. Moreover, proteolysis in Fi sourdough was more intense compared to Ita. The microbial dynamic of the faba bean sourdoughs showed some differences with the most studied cereal sourdoughs.

Rapid and accurate identification of species of the genus Pediococcus isolated from Korean fermented foods by matrix-assisted laser desorption/ionization time-of-flight MS with local database extension.

Pediococci are halophilic lactic acid bacteria, within the family Lactobacillaceae, which are involved in the fermentation of various salted and fermented foods, such as kimchi and jeotgal. In this study, a matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS method was developed for the rapid identification of species of the genus Pediococcus. Of the 130 Pediococcus spectra aligned with the Biotyper taxonomy database, 122 isolates (93.9 %) yielded log scores <1.7, which means they were not identifiable. After registering the spectra of 11 reference strains of the genus Pediococcus, all of the isolates were correctly identified, of which 84 (64.6 %) and 46 (35.4 %) were identified at the species and genus level, respectively. In comparing food origins, no relationship was found between the bacterial characteristics and food environment. We were able to produce a Biotyper system for identification of members of the genus Pediococcus with locally extended Pediococcus reference strains. The MALDI-TOF MS method is fast, simple and reliable for discriminating between species in the genus Pediococcus and therefore will be useful for quality control in determining the spoilage of alcoholic beverages or in the production of fermented food.

Isolation of lactic acid-forming bacteria from biogas plants.

Direct molecular approaches provide hints that lactic acid bacteria play an important role in the degradation process of organic material to methanogenetic substrates in biogas plants. However, their diversity in biogas fermenter samples has not been analyzed in detail yet. For that reason, five different biogas fermenters, which were fed mainly with maize silage and manure from cattle or pigs, were examined for the occurrence of lactic acid-forming bacteria. A total of 197 lactic acid-forming bacterial strains were isolated, which we assigned to 21 species, belonging to the genera Bacillus, Clostridium, Lactobacillus, Pediococcus, Streptococcus and Pseudoramibacter-related. A qualitative multiplex system and a real-time quantitative PCR could be developed for most isolates, realized by the selection of specific primers. Their role in biogas plants was discussed on the basis of the quantitative results and on physiological data of the isolates.

Biodiversity and technological potential of lactic acid bacteria isolated from spontaneously fermented amaranth sourdough.

UNLABELLED: Spontaneous fermented sourdoughs prepared from amaranth flour were investigated for the presence of autochthonous lactic acid bacteria (LAB) predominating microbiota. The doughs were fermented with daily backslopping on a laboratory scale at 30°C for 10 days. LAB counts ranged from 2·60 to 8·54 log CFU g(-1) with a pH declined from 6·2 to 3·8 throughout fermentation. The combined use of randomly amplified polymorphic DNA (RAPD)-PCR analysis and sequence analysis of 16S rRNA was applied for LAB intraspecies differentiation and taxonomic identification, respectively. Enterococcus, Pediococcus and Lactobacillus species were present in amaranth sourdoughs (AS). After the first refreshment step, Lactobacillus plantarum dominated AS until the end of fermentation. In coincidence, when DGGE analysis was performed, the occurrence of a progressive change in bacterial communities allowed the selection of Lact. plantarum as a dominant species. Moreover, technological, functional and safety characteristics of representative RAPD-biotypes were investigated. Lact. plantarum CRL1898 was selected as a potential candidate for gluten-free amaranth sourdough starter. SIGNIFICANCE AND IMPACT OF THE STUDY: Nowadays, there is an increasing interest in ancient noncereal gluten-free (GF) crops such as amaranth, due to their reported nutritional and health benefits. However, the use of these grains is still limited to traditional foods and bread making processes that are not yet well standardized. Results on the dynamics of autochthonous lactic acid bacteria (LAB) microbiota during laboratory spontaneous amaranth sourdoughs (AS) fermentation will contribute to overcome challenges for GF-fermented products development. In addition, knowledge about LAB diversity involving Enterococcus, Pediococcus and Lactobacillus species, with Lactobacillus plantarum predominating during AS fermentation, and their technological and functional properties provides the basis for the selection of autochthonous strains as starters cultures for novel gluten-free bakery products with enhanced nutritional, sensory and/or safety quality.

Effects of probiotics Pediococcus acidilactici strain MA18/5M and Saccharomyces cerevisiae subsp. boulardii strain SB-CNCM I-1079 on fecal and intestinal microbiota of nursing and weanling piglets.

In this study, the influence of (PA) and subsp. (SCB) on fecal and intestinal microbiota of piglets during lactation and after weaning was monitored. Forty sows and their litters were used and allocated to the following dietary treatments: 1) PA, 2) SCB, 3) a mixture of the 2 probiotics (PA+SCB), 4) antibiotics (ATB), and 5) control (CTRL). Four weeks before parturition, probiotic-treated sows started receiving a daily probiotic dose of at least 2.5 × 10 cfu mixed in 500 g of feed until the end of lactation. The other groups were fed a diet without probiotics and ATB. Two days after birth, piglets received, daily, 1 × 10 cfu of the same probiotics as their mother. At weaning (d 21), these piglets were fed a basal diet enriched with the same probiotics whereas piglets from untreated litters were fed the basal diet with or without ATB. Two piglets per litter were randomly chosen to evaluate the influence of treatments on fecal microbial composition (d 10 and 28) and on ileum and colon microbiota at d 37. The microbiota was characterized by culture on selective media and by 16S rRNA gene diversity assessment using the terminal RFLP technique and clone library analysis to evaluate diversity index and phylum affiliation. Terminal RFLP profiles were also analyzed to determine differences in microbial composition between animals receiving different treatments and to identify diet-specific terminal restriction fragments (TRF) using pairwise multiresponse permutation procedures (MRPP) and indicator species analysis. Before weaning, administration of probiotics to sows and piglets had minor effect on fecal microbiota of piglets. Most modulatory effects of probiotics on ileum and colon microbiota were observed on d 37. Results revealed that PA or ATB treatments reduced ileal microbiota diversity compared with the CTRL ( < 0.05) and promoted the establishment of Firmicutes whereas SCB consumption positively influenced the establishment of the Porphyromonadaceae and Ruminococcaceae bacterial families in the colon. Moreover, pairwise MRPP analysis indicated that ileum bacterial communities of pigs treated with PA or ATB differed from those of CTRL pigs ( < 0.05). In conclusion, PA and SCB supplements, respectively, influenced, in a strain-dependent manner, the ileum and colon microbiota of weaned piglets. Results also suggest that PA and SCB have the potential as feed additives to modulate bacterial populations associated with gut health.

A Genomic View of Lactobacilli and Pediococci Demonstrates that Phylogeny Matches Ecology and Physiology.

Lactobacilli are used widely in food, feed, and health applications. The taxonomy of the genus Lactobacillus, however, is confounded by the apparent lack of physiological markers for phylogenetic groups of lactobacilli and the unclear relationships between the diverse phylogenetic groups. This study used the core and pan-genomes of 174 type strains of Lactobacillus and Pediococcus to establish phylogenetic relationships and to identify metabolic properties differentiating phylogenetic groups. The core genome phylogenetic tree separated homofermentative lactobacilli and pediococci from heterofermentative lactobacilli. Aldolase and phosphofructokinase were generally present in homofermentative but not in heterofermentative lactobacilli; a two-domain alcohol dehydrogenase and mannitol dehydrogenase were present in most heterofermentative lactobacilli but absent in most homofermentative organisms. Other genes were predominantly present in homofermentative lactobacilli (pyruvate formate lyase) or heterofermentative lactobacilli (lactaldehyde dehydrogenase and glycerol dehydratase). Cluster analysis of the phylogenomic tree and the average nucleotide identity grouped the genus Lactobacillus sensu lato into 24 phylogenetic groups, including pediococci, with stable intra- and intergroup relationships. Individual groups may be differentiated by characteristic metabolic properties. The link between phylogeny and physiology that is proposed in this study facilitates future studies on the ecology, physiology, and industrial applications of lactobacilli.

Antioxidant potential of a unique LAB culture isolated from Harbin dry sausage: In vitro and in a sausage model.

The lactic acid bacteria Pediococcus pentosaceus, Lactobacillus curvatus, Lactobacillus brevis, and Lactobacillus fermentum isolated from Harbin dry sausage were evaluated for their potential antioxidant activity. The in vitro results showed that P. pentosaceus had the strongest H2O2 resistance, radical scavenging activity, reducing power, and inhibition of lipid peroxidation (P<0.05). Additionally, superoxide dismutase and glutathione peroxidase activities in P. pentosaceus were higher than those observed in three other strains (P<0.05). The inoculation of P. pentosaceus in dry sausage significantly decreased the quantities of thiobarbituric acid-reactive substance and carbonyl formation, while it also reduced the sulfhydryl loss in sausages (P<0.05). Furthermore, the lower content of volatile compounds that are related to lipid-oxidation, such as aldehydes, ketones and hydrocarbons, was observed in the P. pentosaceus inoculated sausage (P<0.05). These results demonstrate that P. pentosaceus has the potential to be employed as an antioxidant starter culture in fermented meat products.

Identification and characterization of wild lactobacilli and pediococci from spontaneously fermented Mountain cheese.

The Traditional Mountain Malga (TMM) cheese is made from raw cow's milk by spontaneously fermentation in small farms called "Malga" located in Trentino region. This study was designed to characterize the lactic acid bacteria (LAB) growing on MRS medium, of TMM-cheese at the end of the ripening. Ninety-five LAB were isolated and genotypically characterized by Randomly Amplified Polymorphic DNA-Polymerase Chain Reaction (RAPD-PCR) with two primers, species-specific PCR and partial sequencing of 16S rRNA gene. The 95 LAB clustered in 70 biotypes. Pediococcus pentosaceus and Lactobacillus paracasei were the dominant species. Isolates were tested for their growth properties, carbohydrate metabolism, acidifying ability, proteolytic and lipolytic activities, acetoin production, amino-peptidase (AP) activity, biogenic amines production, bile salts hydrolysis, conjugated linoleic acid and γ-aminobutyric acid production. Lb. paracasei isolates resulted to be well adapted to Malga environment and to show the best AP activity and acetoin production. TMM-cheese related LAB showed also interesting health promoting properties and produced bioactive substances. In particular, one Lb. brevis biotype produced a GABA mean value of 129 mg/L that is considered a high concentration. The results confirmed that TMM-cheese resident LAB could be exploited for dairy production.

Probiotic Potential of Pediococcus pentosaceus CRAG3: A New Isolate from Fermented Cucumber.

A novel strain of lactic acid bacterium isolated from fermented cucumber was selected due to its high glucansucrase activity. It was identified on the basis of 16S rRNA sequence analysis as Pediococcus pentosaceus CRAG3 (GenBank accession number JX679020). The isolate was round shaped, Gram positive, and catalase negative displaying typical features of lactic acid bacterium. It produced 145 ± 3.27 mg lactic acid per ml of cell-free supernatant. It showed ability to ferment carbohydrates such as sucrose, dextrose, and arabinose; showed resistance to antibiotics such as ciprofloxacin, kanamycin, vancomycin; displayed acid production in triple sugar iron agar test and non-motile nature. Interestingly, the isolate also displayed potential probiotic properties such as hydrophobicity, autoaggregation, coaggregation, and in vitro cell adhesion ability. It exhibited resistance against lysozyme and simulated gastric juice at pH 3.0 with 75 and 58% survival, respectively. It also showed tolerance toward 0.3%, w/v bile salts with 73% survival and ability to deconjugate bile salts. The isolate exhibited antibacterial activity and ability to utilize prebiotics such as inulin and raffinose. These results indicate both probiotic property and glucansucrase-producing ability of P. pentosaceus CRAG3.

Putrescine production from different amino acid precursors by lactic acid bacteria from wine and cider.

The aim of this work was to study the production of biogenic amines and particularly putrescine in lactic acid bacteria (LAB) related to wine and cider. We applied an analytical protocol that involves the use of PCR and TLC techniques to determine the production of putrescine from different precursors. Moreover, we also studied the ability of the Lactobacillus and Pediococcus tested to produce histamine and tyramine. The results showed that the majority of the Lactobacillus brevis analyzed harbour both AgDI and tdc genes and are tyramine and putrescine producers. Conversely, among the other LAB tested, only one Lactobacillus hilgardii and one Pediococcus pentosaceus produced putrescine. The AgDI gene was also detected in two other LAB (Lactobacillus mali and Pediococcus parvulus), but no putrescine production was observed. Finally, hdc gene and histamine production were found in strains (L. hilgardii 5211, isolated from wine, and Lactobacillus casei 18, isolated from cider) that were not putrescine producers.

A description of the lactic acid bacteria microbiota associated with the production of traditional fermented vegetables in Vietnam.

An important part of the daily nourishment in Vietnam constitutes of fermented vegetables. Bacteria and especially lactic acid bacteria play a central role in the production of many fermented vegetables. The current study was conducted to investigate the diversity of native lactic acid bacteria (LAB) populations in 'dua muoi' (mustard and beet fermentation) and 'ca muoi' (eggplant fermentation), three types of popular traditional fermented vegetables of Vietnamese origin. To this end a polyphasic approach combining matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and pheS gene sequence analysis was used. In addition, denaturing gradient gel electrophoresis was performed as a culture-independent method to complement the observed culturable diversity data. A total of 881 LAB isolates were recovered from 21 different samples. Predominant LAB associated with 'dua muoi' and 'ca muoi' were identified as Lactobacillus fermentum (56.6%), Lactobacillus pentosus (24.4%) and Lactobacillus plantarum (17.1%). Less abundant species were Pediococcus pentosaceus (1.0%) and Lactobacillus brevis (0.5%). Species present less than 0.1% included Lactobacillus paracasei, Lactobacillus pantheris and Pediococcus acidilactici. In contrast to fermented mustard and beet with the highest prevalence of L. fermentum, the species most recovered from fermented eggplant samples was L. pentosus. In addition, an important degree of genetic variability within the different predominant species was observed and strain dependency correlating with the type of fermented vegetable or location of production could be demonstrated using multivariate statistics. This research gives an extensive and detailed inventory of the LAB diversity associated with the production of diverse Vietnamese fermented vegetables and demonstrates the influence of type of raw material and/or production location and conditions on this diversity.

Bacterial population dynamics during the ensiling of Medicago sativa (alfalfa) and subsequent exposure to air.

AIMS: To describe, at high resolution, the bacterial population dynamics and chemical transformations during the ensiling of alfalfa and subsequent exposure to air. METHODS AND RESULTS: Samples of alfalfa, ensiled alfalfa and silage exposed to air were collected and their bacterial population structures compared using 16S rRNA gene libraries containing approximately 1900 sequences each. Cultural and chemical analyses were also performed to complement the 16S gene sequence data. Sequence analysis revealed significant differences (P < 0·05) in the bacterial populations at each time point. The alfalfa-derived library contained mostly sequences associated with the Gammaproteobacteria (including the genera: Enterobacter, Erwinia and Pantoea); the ensiled material contained mostly sequences associated with the lactic acid bacteria (LAB) (including the genera: Lactobacillus, Pediococcus and Lactococcus). Exposure to air resulted in even greater percentages of LAB, especially among the genus Lactobacillus, and a significant drop in bacterial diversity. CONCLUSIONS: In-depth 16S rRNA gene sequence analysis revealed significant bacterial population structure changes during ensiling and again during exposure to air. SIGNIFICANCE AND IMPACT OF THE STUDY: This in-depth description of the bacterial population dynamics that occurred during ensiling and simulated feed out expands our knowledge of these processes.

Genotypic and phenotypic diversity of Pediococcus pentosaceus strains isolated from food matrices and characterisation of the penocin operon.

Lactic acid bacteria (LAB) are widely used in the food industry. Pediococcus spp. belong to the LAB group and include several species that are essential for the quality of fermented food. Pediococcus pentosaceus is the species that is most frequently isolated from fermented food and beverages but its uncontrolled growth during food fermentation processes can contribute to undesired flavours. Hence, the characterisation of these bacteria at the strain level is of great importance for the quality of fermented products. Despite their importance, misidentification at the species level is common for members of the genus Pediococcus. To clarify the taxonomic relationships among strains, a multilocus sequencing approach was developed for the characterisation of a collection of 29 field strains, 1 type strain and 1 reference strain of P. pentosaceus isolated from food. These strains were also tested for several phenotypic properties of technological interest and for the production of bacteriocins. The chromosomal operon involved in the synthesis of the bacteriocin penocin was also investigated. The present study enabled a good genomic characterisation, identifying 17 sequence types, with an overview of phenotypic characteristics related to different technological abilities, and also provides a thorough characterisation of the operon involved in penocin production.

Characterisation of the bacterial microbiota of the vagina of dairy cows and isolation of pediocin-producing Pediococcus acidilactici.

BACKGROUND: Uterine infections in dairy cows lower profitability of dairy operations. Infections of the reproductive tract are related to the overgrowth of pathogenic bacteria during the first three weeks after parturition. However, alterations in the vaginal microbiota composition in the first weeks after parturition remain poorly documented. RESULTS: In this study, bacteria isolated from the vagina of healthy pregnant, and infected postpartum cows were characterised by random amplification of polymorphic DNA (RAPD) analysis and partial 16S ribosomal RNA (rDNA) gene sequencing. Populations of bacilli and lactic acid bacteria of the genera Enterococcus, Lactobacillus, and Pediococcus were present in both healthy and infected cows. Infected cows had a significant increase in the vaginal enteric bacteria population which consisted mainly of Escherichia coli. Three E. coli isolates harboured the gene coding for Shiga-like-toxin (SLT) I or II. Several isolates of the Pediococcus acidilactici were found to produce the bacteriocin pediocin AcH/PA-1. Quantitative PCR analyses of vaginal mucus samples collected from ten metritic cows before and after parturition confirmed the presence of the Lactobacillus group (Lactobacillus spp., Pediococcus spp., Leuconostoc spp., and Weissella spp.); Enterobacteriaceae, E. coli, and bacilli. The presence of the pediocin AcH/PA-1 structural gene and SLT genes were also confirmed with qPCR. CONCLUSIONS: In conclusion, overgrowth of pathogenic bacteria, particularly E. coli, after parturition likely contributes to the development of metritis. Our microbiota analysis extends the information related to the composition of commensal bacteria in the bovine female reproductive tract and may facilitate the development of novel intervention strategies for prevention of uterine infections in dairy cows.

Broad-spectrum antifungal-producing lactic acid bacteria and their application in fruit models.

A large-scale screen of some 7,000 presumptive lactic acid bacteria (LAB), isolated from animal, human, or plant origin, identified 1,149 isolates with inhibitory activity against the food-spoilage mould Penicillium expansum. In excess of 500 LAB isolates were subsequently identified to produce a broad spectrum of activity against P. expansum, Penicillium digitatum, Penicillium notatum, Penicillium roqueforti, Rhizopus stolonifer, Fusarium culmorum, Aspergillus fumigatus and Rhodotorula mucilaginosa. Partial 16S rRNA sequencing of 94 broad spectrum isolates revealed that the majority of antifungal producers were strains of Lactobacillus plantarum. The remaining population was composed of Weissella confusa and Pediococcus pentosaceous isolates. Characterization of six selected broad-spectrum antifungal LAB isolates revealed that antifungal activity is maximal at a temperature of 30 °C, a pH of 4.0 and is stable across a variety of salt concentrations. The antifungal compound(s) was shown to be neither proteinaceous nor volatile in nature. P. pentosaceous 54 was shown to have protective properties against P. expansum spoilage when applied in pear, plum and grape models, therefore representing an excellent candidate for food-related applications.

A novel thermotolerant Pediococcus acidilactici B-25 strain for color, COD, and BOD reduction of distillery effluent for end use applications.

The present study was aimed to characterize physico-chemical and microbial population of distillery effluent and isolate a novel thermotolerant bacterium for color, COD, and BOD reduction of spentwash. The level of alkalinity, TSS, DO, COD, BOD, TN, ammonical nitrogen, nitrate nitrogen, phosphorous, potassium, chloride, and calcium of spentwash (SW), bioreactor effluent (BE), and secondary treated effluent (STE) were well above the permissible limits. The level of color, TS, and TDS were under the permissible limits for STE but not for SW and BE. The microbial population was higher in BE. The results revealed that effluent was highly polluted and require suitable treatment before discharge. A novel thermotolerant bacterium, identified as Pediococcus acidilactici, was isolated which exhibited maximum 79 % decolorization, 85 % COD, and 94 % BOD reduction at 45 °C using 0.1 %, glucose; 0.1 %, peptone; 0.05 %, MgSO4; 0.05 %, K2HPO4; pH 6.0 within 24 h under static condition. The ability of this strain to decolorize melanoidin at minimum carbon and nitrogen supplementation warrants its possible application for effluent treatment at industrial level. In addition, it is first instance when melanoidin decolorization was reported by P. acidilactici. This study could be an approach towards control of environmental pollution and health hazards of people in and around the effluent distillery unit.

Pediococcus lolii DSM 19927T and JCM 15055T are strains of Pediococcus acidilactici.

Strain NGRI 0510Q(T), isolated from ryegrass silage, was recently classified as a representative of a novel Pediococcus species, Pediococcus lolii Doi et al. 2009. It was deposited in the DSMZ and JCM culture collections as DSM 19927(T) and JCM 15055(T), respectively. A polyphasic taxonomic study, including matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, pheS and 16S rRNA gene sequence analysis, fluorescent amplified fragment length polymorphism and DNA-DNA hybridization, was used to prove that both subcultures of the type, and only, strain of this species are strains of Pediococcus acidilactici.

Isolation of Pediococcus acidilactici Kp10 with ability to secrete bacteriocin-like inhibitory substance from milk products for applications in food industry.

BACKGROUND: Lactic acid bacteria (LAB) can be isolated from traditional milk products. LAB that secrete substances that inhibit pathogenic bacteria and are resistant to acid, bile, and pepsin but not vancomycin may have potential in food applications. RESULTS: LAB isolated from a range of traditional fermented products were screened for the production of bacteriocin-like inhibitory substances. A total of 222 LAB strains were isolated from fermented milk products in the form of fresh curds, dried curds, and ghara (a traditional flavor enhancer prepared from whey), and fermented cocoa bean. Eleven LAB isolates that produced antimicrobial substances were identified as Lactococcus lactis, Lactobacillus plantarum, and Pediococcus acidilactici strains by biochemical methods and 16S rDNA gene sequencing. Of these, the cell-free supernatant of Kp10 (P. acidilactici) most strongly inhibited Listeria monocytogenes. Further analysis identified the antimicrobial substance produced by Kp10 as proteinaceous in nature and active over a wide pH range. Kp10 (P. acidilactici) was found to be catalase-negative, able to produce ß-galactosidase, resistant to bile salts (0.3%) and acidic conditions (pH 3), and susceptible to most antibiotics. CONCLUSION: Traditionally prepared fermented milk products are good sources of LAB with characteristics suitable for industrial applications. The isolate Kp10 (P. acidilactici) shows potential for the production of probiotic and functional foods.