Taxonomic and molecular characterization of lactic acid bacteria and yeasts in nunu,a Ghanaian fermented milk product

Produced from raw unpasteurized milk, nunu is a spontaneously fermented yoghurt-like product made in Ghana and other parts of West Africa. Despite the importance of nunu in the diet of many Africans, there is currently only limited information available on the microorganisms associated with nunu processing. With the aim of obtaining a deeper understanding of the process and as a first step towards developing starter cultures with desired technological properties for nunu production, a microbiological characterization of nunu processing in three different towns in the Upper East region of Ghana, namely Bolgatanga, Paga and Navrongo, was carried out. Lactic acid bacteria (LAB) and yeasts associated with nunu processing were isolated and identified using a combination of pheno- and genotypic methods including morphological and carbohydrate fermentation tests, (GTG)₅-based rep-PCR, multiplex PCR, and 16S and 26S rRNA gene sequencing. The LAB counts during nunu processing increased from 4.5 ± 0.4 log cfu/ml at 0 h to 8.7 ± 1.8 log cfu/ml at 24 h of fermentation while yeasts counts increased from 2.8 ± 1.2 log cfu/ml at 0 h to 5.8 ± 0.5 log cfu/ml by the end of fermentation. Lactobacillus fermentum was the dominant LAB throughout the fermentations with Lactobacillus plantarum and Leuconostoc mesenteroides playing prominent roles during the first 6–8 h of fermentation as well. Less frequently isolated LAB included Lactobacillus helveticus, Enterococcus faecium, Enterococcus italicus, Weissella confusa and a putatively novel Lactococcus spp. The yeasts involved were identified as Candida parapsilosis, Candida rugosa, Candida tropicalis, Galactomyces geotrichum, Pichia kudriavzevii and Saccharomyces cerevisiae with P. kudriavzevii and S. cerevisiae being the dominant yeast species.     

Evaluating the individual effects of temperature and salt on table olive related microorganisms

Statistical modelling techniques were used in the present study to assess the individual effects of temperature and NaCl concentration on the growth of 10 lactic acid bacteria and 6 yeast strains mostly isolated from different forms of table olive processing and belonging to the species Lactobacillus pentosus, Lactobacillus plantarum, Saccharomyces cerevisiae, Wickerhamomyces anomalus and Candida boidinii. The mathematical models obtained in synthetic laboratory media show that yeasts, except for C. boidinii, were more resistant to a high salt concentration than lactic acid bacteria, with an MIC value ranging from 163.5 (S. cerevisiae) to 166.9 g/L (W. anomalus); while for L. pentosus and L. plantarum this parameter ranged from 110.6 to 117.6 g/L, respectively. With regards to temperature, lactic acid bacteria showed a slight trend towards supporting higher temperature values than yeasts, with the exception of S. cerevisiae. The maximum temperatures for growth of L. pentosus and L. plantarum were 41.9 and 43.0 °C, respectively; while for W. anomalus and C. boidinii they were 38.2 and 36.5 °C. The optimum temperatures for growth were also higher for L. pentosus and L. plantarum (35.5 and 32.9 °C), compared to W. anomalus and C. boidinii (29.3 and 26.9 °C, respectively). Additional experiments carried out in natural olive brines confirmed previous results, showing that high NaCl concentrations clearly favoured yeast growth and that at high temperatures LAB slightly overcame yeasts. Results obtained in this paper could be useful for industry for a better control of both table olive fermentation and packaging.     

Lactic acid bacteria and yeasts isolated from the starter doughs for Chinese steamed buns in Thailand

Isolation of lactic acid bacteria (LAB) and yeasts from the starter dough of Chinese steamed buns from four different commercial sources in Thailand was carried out. Thirty-one lactic acid bacteria and eight yeast strains were isolated. Total counts of LAB were from 1.8 × 10⁴ to 10⁸ colonies/g sample, whilst yeasts were very low from 10 to 2.3 × 10² colonies/g sample. The pH values of all starter doughs ranged from 3.36 to 3.52 and the Total Titratable Acidity (TTA) varied from 11.1 to 17.0 ml of 0.1 N NaOH/10 g dough. All LAB isolates were identified as Lactobacillus. The phenotypic characteristics were used to cluster all the LAB isolates into two major groups (Group A and Group B), with the B group subdivided into four groups. Phylogenetic analysis, based upon partial 16S rRNA gene sequences, showed that isolates of Group A, which all contained meso-diaminopimelic acid in their cell wall and produced dl-lactic acid, were closely related to Lactobacillus plantarum, whilst the strains of Group B that produced l-lactic acid were closely related to Lactobacillus casei. For yeasts, eight isolates based on the D1/D2 domain sequences of 26S rRNA were identified as Candida tropicalis, Pichia stipitis, Candida parapsilosis, Issatchenkia orientalis and Saccharomyces cerevisiae.     

Lactic acid bacteria population dynamics during minced beef storage under aerobic or modified atmosphere packaging conditions

A total of 266 lactic acid bacteria (LAB) have been isolated from minced beef stored at 0, 5, 10 and 15 °C aerobically and under modified atmosphere packaging consisting of 40% CO₂–30% O₂–30% N₂ in the presence MAP (+) and absence MAP (−) of oregano essential oil. Sequencing of their 16S rRNA gene along with presence of the katA gene demonstrated dominance of the LAB microbiota by Leuconostoc spp. during aerobic storage at 5, 10 and 15 °C, as well as during MAP (−) and MAP (+) storage at 10 and 15 °C; Lactobacillus sakei prevailed during aerobic storage at 0 °C, as well as at MAP (−) and MAP (+) storage at 0 and 5 °C. The sporadic presence of other species such as Leuconostoc mesenteroides, Weisella viridescens, Lactobacillus casei and Lactobacillus curvatus has also been determined. Pulsed-Field Gel Electrophoresis of high molecular weight genomic DNA revealed the dynamics of the isolated LAB strains. Prevalence of Leuconostoc spp. was attributed to one strain only. On the other hand, packaging conditions affected Lb. sakei strain spoilage dynamics.     

FTIR-based polyphasic identification of lactic acid bacteria isolated from traditional Greek Graviera cheese

This study used a combination of phenotypic, physical (Fourier Transformed Infra-Red [FTIR] spectroscopy) and molecular (RFLP and SSCP analysis of 16S rRNA genes) methods to identify the lactic acid bacteria (LAB) flora present in traditional Greek Graviera cheese after five weeks of ripening. A total of 300 isolates collected from high dilution plates of TSAYE (incubated at 30 °C), M-17 (22 °C) and M-17 (42 °C) agar media were clustered by FTIR and then representative strains of each cluster were cross-identified blindly by all methods. Based on their FTIR spectra, 282 isolates were LAB grouped in 28 clusters. The LAB species identified and their prevalence in the cheese samples were: Lactobacillus casei/paracasei (68.8%), Lactobacillus plantarum (19.5%), Streptococcus thermophilus (8.9%), Enterococcus faecium (2.1%), and Lactococcus lactis (0.7%). Also, Staphylococcus equorum (11 isolates), Corynebacterium sp. (5 isolates) and Brevibacterium sp. (1 isolate) were recovered from TSAYE. Comparative identification results showed that phenotypic and molecular methods were in mutual agreement as regards the LAB species identified. The present polyphasic identification approach based on rapid FTIR screening of 10-fold more isolates than a previous classical identification approach allowed or improved detection of few sub-dominant species; however the predominant LAB species in the cheese samples were the same with both approaches.     

Spontaneous organic cocoa bean box fermentations in Brazil are characterized by a restricted species diversity of lactic acid bacteria and acetic acid bacteria

Spontaneous organic cocoa bean box fermentations were carried out on two different farms in Brazil. Physical parameters, microbial growth, bacterial species diversity [mainly lactic acid bacteria (LAB) and acetic acid bacteria (AAB)], and metabolite kinetics were monitored, and chocolates were produced from the fermented dry cocoa beans. The main end-products of the catabolism of the pulp substrates (glucose, fructose, and citric acid) by yeasts, LAB, and AAB were ethanol, lactic acid, mannitol, and/or acetic acid. Lactobacillus fermentum and Acetobacter pasteurianus were the predominating bacterial species of the fermentations as revealed through (GTG)₅-PCR fingerprinting of isolates and PCR-DGGE of 16S rRNA gene PCR amplicons of DNA directly extracted from fermentation samples. Fructobacillus pseudoficulneus, Lactobacillus plantarum, and Acetobacter senegalensis were among the prevailing species during the initial phase of the fermentations. Also, three novel LAB species were found. This study emphasized the possible participation of Enterobacteriaceae in the cocoa bean fermentation process. Tatumella ptyseos and Tatumella citrea were the prevailing enterobacterial species in the beginning of the fermentations as revealed by 16S rRNA gene-PCR-DGGE. Finally, it turned out that control over a restricted bacterial species diversity during fermentation through an ideal post-harvest handling of the cocoa beans will allow the production of high-quality cocoa and chocolates produced thereof, independent of the fermentation method or farm.     

Diversity of bacteria and yeast in the naturally fermented cotton seed and rice beverage produced by Brazilian Amerindians

Microorganisms associated with the fermentation of cotton seed and rice were studied using a combination of culture-dependent and -independent methods. Samples of the cotton seed and rice beverage were collected every 8 h during the fermentation process for analysis of the microbiota present over 48 h. The lactic acid bacteria (LAB) population reached values of approximately 8.0 log cfu/mL. A total of 162 bacteria and 81 yeast isolates were identified using polyphasic methods. LAB (Lactobacillus plantarum, Lactobacillus vermiforme, Lactobacillus paracasei) were the most frequently isolated bacteria. Bacillus subtilis was present from 16 h until the end of the fermentation process. A decrease in pH value from 6.92 (0 h) to 4.76 (48 h) was observed, and the concentration of lactic acid reached 24 g/L at the end of the fermentation process. DGGE (denaturing gradient gel electrophoresis) was performed to determine the dynamics of the communities of bacteria and yeast, and the analysis revealed a predominance of LAB throughout the fermentation process. No changes were observed in the yeast community. The yeast species detected were Candida parapsilosis, Candida orthopsilosis, Clavispora lusitaniae and Rhodotorula mucilaginosa. Our studies indicate that the DGGE technique combined with a culture-dependent method is required to discern the dynamics in the fermentation of cotton seed and rice.     

Enhancement of survival of probiotic and non-probiotic lactic acid bacteria by yeasts in fermented milk under non-refrigerated conditions

The effects of yeasts on the survival of probiotic and non-probiotic lactic acid bacteria (LAB) were studied in fermented milk under non-refrigerated conditions (30 °C) with a view to develop ambient-stable fermented milk with live LAB. Five yeasts tested (Saccharomyces bayanus, Williopsis saturnus var. saturnus, Yarrowia lipolytica, Candida kefyr and Kluyveromyces marxianus) enhanced the survival of Lactobacillus bulgaricus (but not Streptococcus thermophilus) in a mixed yoghurt culture in yoghurt by ~ 10² to 10⁵-fold. Seven yeasts examined (Candida krusei, Geotrichum candidum, Pichia subpelliculosa, Kloeckera apiculata, Pichia membranifaciens, Schizosaccharomyces pombe and Y. lipolytica) improved the survival of Lactobacillus rhamnosus in fermented milk by ~ 10³ to 10⁶-fold. W. saturnus var. saturnus enhanced the survival of Lactobacillus acidophilus, L. rhamnosus (probiotic) and Lactobacillus reuteri by up to 10⁶-fold, but the same yeast failed to improve the survival of Lactobacillus johnsonii (probiotic), S. thermophilus and L. bulgaricus in fermented milk. These results provide definitive evidence that yeasts possess stability-enhancing effects on LAB and that the specific effects of yeasts on LAB stability vary with yeasts as well as with LAB. However, the molecular mechanism of such interaction of yeasts with LAB remains to be found.     

Microbiological and physicochemical characterisation of green table olives of Halkidiki and Conservolea varieties processed by the Spanish method on industrial scale

The microbiological and physicochemical changes of industrially fermented Halkidiki and Conservolea green table olives were determined. Samples were analysed to monitor the population of lactic acid bacteria (LAB), yeasts and Enterobacteriaceae, together with changes in pH, acidity, salinity, colour, lactic acid, acetic acid and ethanol. LAB and yeast species diversity was evaluated at the beginning (1 day), middle (75 days) and final (135 days) stages of fermentation by RAPD-PCR genomic fingerprinting. Results revealed vigorous lactic acid processes as indicated by the dominance of LAB over yeasts. No Enterobacteriaceae could be detected after 30 days. Lactiplantibacillus plantarum (formerly Lactobacillus plantarum) dominated in the beginning of fermentation in both varieties. In the end, Lactiplantibacillus pentosus (formerly Lactobacillus pentosus) and Pediococcus ethanolidurans prevailed in Halkidiki and Conservolea varieties, respectively. As for yeasts, Kluyveromyces lactis/marxianus and Pichia manshurica prevailed at the onset of fermentation in Halkidiki and Conservolea varieties, whereas in the end Pichia membranifaciens dominated in both varieties.     

Effects of two different sourdoughs on the characteristics of Pandoro, a typical Italian sweet leavened baked good

Pandoro is a traditional Italian sweet-leavened baked product, usually consumed at Christmas. It is manufactured according to specific procedures and preparation starts from a sourdough called ‘‘madre’’ (mother sponge) continuously refreshed. This sourdough is the result of a complex microbial association including yeasts and lactic acid bacteria (LAB) and its use can improve sensory quality and shelf-life of the resulting products. The aim of this work was to evaluate the effect of the use of two different sourdoughs matured at different temperature (13 and 19 °C) on some metabolites, which can affect the organoleptic characteristics of Pandoro. Different samples, taken throughout the 19 h of process, were analysed for the determination of yeast and LAB counts, pH, a_w, carbohydrates, organic acids content and volatile profile. The results showed that, at the end of fermentation process, the sourdough propagated at 19 °C reached lower pH values than that at 13 °C. This was probably due to higher LAB counts (1 log unit higher), resulting also in higher lactic acid concentration and faster sugars depletion. On the contrary, temperature of dough during maturation did not affected yeast concentrations. Different production processes strongly affected also the volatile profile, both of sourdough at the end of fermentation and of the final products.     

Use of a starter culture of lactic acid bacteria in plaa-som, a Thai fermented fish

Plaa-som is a Thai fermented fish prepared from freshwater fish and various ingredients. In this study, two strains of lactic acid bacteria (LAB) isolated from natural plaa-som fermentation were used as starter cultures: Lactobacillus plantarum IFRPD P15 and Lactobacillus reuteri IFRPD P17. These strains were used as a mixed starter culture for plaa-som using an air-drying method (laminar airflow) with sterilized rice grains as the filler. This method produced a suitable starter culture, which was maintained at 4 °C for more than 20 weeks. LAB were the dominant bacteria in the starter culture and produced high acidity from 24 h until the end of fermentation. This resulted in decreased pH in plaa-som. L. plantarum IFRPD P15 was dominant as an acidity producer, whereas L. reuteri IFRPD P17 showed an ability to suppress and eliminate pathogenic bacteria such as Escherichia coli within 24 h. The use of a single starter culture for plaa-som resulted in incomplete suppression of pathogenic bacteria and elimination of E. coli. Thus, L. plantarum IFRPD P15 and L. reuteri IFRPD P17 have great potential for use as a mixed starter culture in plaa-som fermentation and may possibly help to reduce fermentation time.     

Impact of ecological factors on the stability of microbial associations in sourdough fermentation

The limits for the stability of the microbial association 1 (Lactobacillus sanfranciscensis and Candida humilis) and association 2 (Lactobacillus reuteri, Lactobacillus johnsonii and Issatchenkia orientalis) during sourdough fermentation were evaluated by investigating the effects of the ecological factors substrate, refreshment time, temperature, amount of backslopping and competing species in different combinations on their growth. Sourdoughs were fermented in 28 batches under different conditions using the associations and possible competing strains as starters. The dominating microbiota was characterized by bacteriological culture, rRNA gene sequence analysis and RAPD-PCR. Association 1 was found to be competitive in doughs with rye and wheat flour at temperatures between 20 and 30 °C, refreshment times of 12 and 24 h, amounts of backslopping dough from 5 to 20% and against all competing lactic acid bacteria and yeasts. The processing parameters for the competitiveness of the association 2 were temperatures of 35-40 °C, refreshment times of 12-24 h and the substrates rye bran, wheat and rye flour, but not in every case. Issatchenkia orientalis could only grow when enough oxygen was available. Its cell counts fell rapidly under the limit of detection when using high amounts of doughs (small ratio of surface to volume) and refreshment times of 12 h. In conclusion, the results demonstrated that the two associations were remarkably stable under most of the investigated process conditions.     

Prevalence and impact of single-strain starter cultures of lactic acid bacteria on metabolite formation in sourdough

Flavour of type II sourdoughs is influenced by the ingredients, processing conditions, and starter culture composition. It is, however, not fully clear to what extent different sourdough lactic acid bacteria (LAB) contribute to flavour. Therefore, two types of flour (rye and wheat) and different LAB starter culture strains were used to prepare sourdoughs, thereby leaving the yeast microbiota uncontrolled. All LAB starter culture strains tested were shown to be prevalent and to acidify the flour/water mixture to pH values between 3.1 and 3.9 after 24 h of fermentation. Multiple aldehydes, alcohols, ketones, and carboxylic acids were produced by the sourdough-associated microbiota throughout the fermentation period. Based on the organoleptic evaluation of breads produced with these sourdoughs, five LAB strains were selected to perform prolonged wheat and rye fermentations as to their capacity to result in an acidic (Lactobacillus fermentum IMDO 130101, Lactobacillus plantarum IMDO 130201, and Lactobacillus crustorum LMG 23699), buttermilk-like (Lactobacillus amylovorus DCE 471), or fruity flavour (Lactobacillus sakei CG1). Upon prolonged fermentation, higher metabolite concentrations were produced. For instance, L. sakei CG1 produced the highest amounts of 3-methyl-1-butanol, which was further converted into 3-methylbutyl acetate. The latter compound resulted in a fruity banana flavour after 48 h of fermentation, probably due to yeast interference. Rye fermentations resulted in sourdoughs richer in volatiles than wheat, including 3-methyl-1-butanol, 2-phenylethanol, and ethyl acetate.     

Identification of the lactic acid bacteria in Kimchi according to initial and over-ripened fermentation using PCR and 16S rRNA gene sequence analysis

This paper aimed to identify the lactic acid bacteria species involved in kimchi fermentation at different fermentation periods by using culture-independent 16S rRNA gene clone libraries, and develop polymerase chain reaction for the detection of lactic acid bacteria (LAB). We investigated 6 commercially produced kimchi samples, including kimchi at an initial stage of fermentation and kimchi that was fermented to an over-ripened stage. The results of our study show that the analysis with cultureindependent 16S rRNA gene clone libraries could successfully identify 134 clones, 11 species, including Weissella, Lactobacillus, Pediococcus, and Leuconostoc from the 6 commercial kimchi samples. Weisella koreensis and Lactobacillus brevis were the predominant LAB in the initial stage of kimchi fermentation at 4°C (pH 4.96–5.27 and acidity 0.81–0.88), and Leuconostoc gelidum and Lactobacillus sakei subsp. sakei may play an important role in kimchi fermentation at the over-ripened stage (pH 3.61–3.91 and acidity 1.70–1.79).     

Effect of lactic acid fermentation on antioxidant, texture, color and sensory properties of red and green smoothies

Weissella cibaria, Lactobacillus plantarum, Lactobacillus sp. and Lactobacillus pentosus were variously identified from blackberries, prunes, kiwifruits, papaya and fennels by partial 16S rRNA gene sequence. Representative isolates from each plant species were screened based on the kinetics of growth on fruit juices. A protocol for processing and storage of red and green smoothies (RS and GS) was set up, which included fermentation by selected lactic acid bacteria starters and exo-polysaccharide producing strains. Starters grew and remained viable at ca. 9.0 log cfu g⁻¹ during 30 days of storage at 4 °C. No contaminating Enterobacteriaceae and yeast were found throughout storage. Values of soluble solids, total titratable acidity and viscosity distinguished started RS and GS compared to spontaneously (unstarted) fermented smoothies. Color difference dE∗_ab and browning index were positively affected by lactic acid fermentation. Consumption of carbohydrates by lactic acid bacteria was limited as well as it was the lactic fermentation. Consumption of malic acid was evident throughout storage. Polyphenolic compounds and, especially, ascorbic acid were better preserved in started RS and GS compared to unstarted samples. This reflected on the free radical scavenging activity. A statistical correlation was only found between the level of ascorbic acid and free radical scavenging activity. As shown by a ?rst-order equation, the rate of degradation of ascorbic acid through storage were found to be higher in the unstarted compared to started RS and GS. Fermentation by lactic acid bacteria clearly improved the sensory attributes of RS and GS.     

Adaptability of lactic acid bacteria and yeasts to sourdoughs prepared from cereals, pseudocereals and cassava and use of competitive strains as starters

The adaptability of lactic acid bacteria (LAB) and yeasts to sourdoughs prepared from cereals, pseudocereals and cassava was investigated using PCR-DGGE and bacteriological culture combined with rRNA gene sequence analysis. Sourdoughs were prepared either from flours of the cereals wheat, rye, oat, barley, rice, maize, and millet, or from the pseudocereals amaranth, quinoa, and buckwheat, or from cassava, using a starter consisting of various species of LAB and yeasts. Doughs were propagated until a stable microbiota was established. The dominant LAB and yeast species were Lactobacillus fermentum, Lactobacillus helveticus, Lactobacillus paralimentarius, Lactobacillus plantarum, Lactobacillus pontis, Lactobacillus spicheri, Issatchenkia orientalis and Saccharomyces cerevisiae. The proportion of the species within the microbiota varied. L. paralimentarius dominated in the pseudocereal sourdoughs, L. fermentum, L. plantarum and L. spicheri in the cassava sourdough, and L. fermentum, L. helveticus and L. pontis in the cereal sourdoughs. S. cerevisiae constituted the dominating yeast, except for quinoa sourdough, where I. orientalis also reached similar counts, and buckwheat and oat sourdoughs, where no yeasts could be detected. To assess the usefulness of competitive LAB and yeasts as starters, the fermentations were repeated using flours from rice, maize, millet and the pseudocereals, and by starting the dough fermentation with selected dominant strains. At the end of fermentation, most of starter strains belonged to the dominating microbiota. For the rice, millet and quinoa sourdoughs the species composition was similar to that of the prior fermentation, whereas in the other sourdoughs, the composition differed.     

Culture-independent study of the diversity of microbial populations in brines during fermentation of naturally-fermented Aloreña green table olives

Aloreña table olives are naturally fermented traditional green olives with a denomination of protection (DOP). The present study focused on Aloreña table olives manufactured by small and medium enterprises (SMEs) from Valle del Guadalhorce (Southern Spain) under three different conditions (cold storage, and ambient temperature fermentations in small vats and in large fermentation tanks). The microbial load of brines during fermentation was studied by plate counting, and the microbial diversity was determined by a culture-independent approach based on PCR-DGGE analysis. The viable microbial populations (total mesophilic counts, yeasts and molds, and lactic acid bacteria — LAB) changed in cell numbers during the course of fermentation. Great differences were also observed between cold, vat and tank fermentations and also from one SME to another. Yeasts seemed to be the predominant populations in cold-fermented olives, while LAB counts increased towards the end of vat and tank fermentations at ambient temperature. According to PCR-DGGE analysis, microbial populations in cold-fermented olives were composed mostly by Gordonia sp./Pseudomonas sp. and Sphingomonas sp./Sphingobium sp./Sphingopyxis sp. together with halophilic archaea (mainly by haloarchaeon/Halosarcina pallida and uncultured archaeon/uncultured haloarchaeon/Halorubrum orientalis) and yeasts (Saccharomyces cerevisiae and Candida cf. apicola). Vat-fermented olives stored at ambient temperature included a more diverse bacterial population: Gordonia sp./Pseudomonas sp., Sphingomonas sp./Sphingobium sp./Sphingopyxis sp. and Thalassomonas agarivorans together with halophilic archaea and yeasts (mainly S. cerevisiae and C. cf. apicola, but also Pichia sp., and Pichia manshurica/Pichia galeiformis). Some LAB were detected towards the end of vat fermentations, including Lactobacillus pentosus/Lactobacillus plantarum and Lactobacillus vaccinostercus/Lactobacillus suebicus. Only the tank fermentation showed a clear predominance of LAB populations (Lactobacillus sp., Lactobacillus paracollinoides, and Pediococcus sp.) together with some halophilic archaea and a more selected yeast population (P. manshurica/P. galeiformis). The present study illustrates the complexity of the microbial populations in naturally-fermented Aloreña table olives.     

Diversity of lactic acid bacteria from modified atmosphere packaged sliced cooked meat products at sell-by date assessed by PCR-denaturing gradient gel electrophoresis

The predominant lactic acid bacteria (LAB) microbiota associated with three types of modified atmosphere packaged (MAP) sliced cooked meat products (i.e. ham, turkey and chicken) was analyzed at sell-by date using a combination of culturing and molecular population fingerprinting. Likewise routine analyses during industrial MAP production, meat samples were plated on the general heterotrophic Plate Count Agar (PCA) and on the LAB-specific de Man, Rogosa, Sharpe (MRS) agar under different temperature and atmosphere conditions. Subsequently, community DNA extracts were prepared from culturable bacterial fractions harvested from both media and used for PCR targeting the V3 hyper-variable region of the 16S rRNA gene followed by denaturing gradient gel electrophoresis (DGGE) of PCR amplicons (PCR-DGGE). Irrespective of aerobic or anaerobic incubation conditions, V3-16S rDNA DGGE fingerprints of culturable fractions from PCA and MRS medium displayed a high level of similarity indicating that LAB constituted the most dominant group in the culturable bacterial community. Comparison of DGGE profiles of fractions grown at 20, 28 or 37 °C indicated that part of the culturable community consisted of psychrotrophs. Four DGGE bands were common among cooked ham, turkey and chicken products, suggesting that these represent the microbiota circulating in the plant where all three MAP product types were sliced and packaged. Based on band sequencing and band position analysis using LAB reference strains, these four bands could be assigned to Lactobacillus sakei and/or the closely related Lactobacillus fuchuensis, Lactobacillus curvatus, Carnobacterium divergens and Leuconostoc carnosum. In conclusion, the PCR-DGGE approach described in this study allows to discriminate, identify and monitor core and occasional LAB microbiota of MAP sliced cooked meat products and provides valuable complementary information to the current plating procedures routinely used in industrial plants.     

Identification of lactic acid bacteria isolated during traditional fura processing in Ghana

Fura is a millet-based spontaneously fermented dumpling produced and consumed in parts of West Africa, particularly Nigeria, Burkina Faso and Ghana. From eight traditional fura production sites in northern Ghana, 862 lactic acid bacteria were isolated and identified to species level using a combination of genotypic and phenotypic methods including (GTG)₅-based PCR fingerprinting and 16S rRNA gene sequencing, multiplex PCR by means of recA gene sequence comparison, conventional morphological characteristics and carbohydrate fermentation profiling. During millet dough fermentation, pH decreased from 5.6–6.4 to 4.1–3.7 and total lactic acid bacteria (LAB) counts increased from 4.4–5.3 to 7.9–9.2 log₁₀ (cfu/g). The initial stages of the fermentation were characterized by co-dominance of homo- and heterofermentative species of Pediococcus acidilactici, Weisella confusa, Lactobacillus fermentum, Lactobacillus reuteri, Lactobacillus salivarius, and Lactobacillus paraplantarum whereas L. fermentum was dominating at the end of the fermentation. L. fermentum was predominant in all fermentations (p < 0.05) and a high uniformity was observed among production sites regarding the dominance of L. fermentum. L. fermentum and W. confusa were isolated in all production sites and almost at all fermentation stages indicating that they are indigenous to traditional fura processing. The other LAB bacteria species which comprised a minor proportion of the total LAB occurred occasionally and in an irregular pattern among the production sites.     

Lactic acid bacteria isolated from ethnic preserved meat products of the Western Himalayas

We used culture- and molecular-biology-based methods to investigate the diversity of lactic acid bacteria (LAB) in the ethnic chevon (goat) meat products chartayshya, jamma and arjia of the Western Himalayas. In six chartayshya, six jamma and four arjia samples, LAB were the predominant microbial component involved in the fermentation of these samples, and the total LAB population in arjia (7.8 ± 0.1 log cfu g⁻¹; mean ± SD) was significantly higher (P < 0.05) than in chartayshya (6.9 ± 0.1 log cfu g⁻¹) and jamma (7.5 ± 0.1 log cfu g⁻¹). We identified 53 LAB samples by 16S rRNA and phenylalanyl-tRNA synthase (pheS) genes sequencing. The LAB isolates were identified as Enterococcus durans, Enterococcus faecalis, Enterococcus faecium, Enterococcus hirae, Leuconostoc citreum, Leuconostoc mesenteroides, Pediococcus pentosaceus, and Weissella cibaria. These results revealed that there is a high level of diversity of LAB in the Himalayan ethnic preserved meat products.