Plasmid profiles of ten strains of Lactobacillus plantarum

Abstract The presence of extrachromosomal DNA elements has been investigated in 10 strains of Lactobacillus plantarum .
8 of the strains contained from 1 to 6 plasmids of different M _r values spanning from 1.35 · 10⁶ to 15.4 · 10⁶.
6 of the strains are commonly used as starter cultures in dry sausage and all these strains contained plasmids. The remaining 4 strains were obtained from the American Type Culture Collection and only 2 of these strains were found to harbour plasmids.     

Utilization of Unfermented Cassava Flour for the Production of an Indigenous African Fermented Food, Agbelima

Summary Unfermented cassava flour, also called high quality cassava flour (HQCF) is used as a partial substitute for wheat flour in the baking industry. This work was carried out to produce an indigenous fermented cassava dough, agbelima, from high quality cassava flour in order to diversify uses for the flour. An isolate of each of the dominant species of lactic acid bacteria isolated from agbelima, was used to ferment reconstituted HQCF dough into agbelima, whilst pH changes, population on MRS, and the organoleptic quality were assessed. The antimicrobial properties of the fermenting samples against three enteric pathogens were also investigated. Lactobacillus plantarum, Lactobacillus brevis, Lactobacillus fermentum and Leuconostoc mesenteroides were isolated at levels of 10¹⁰, 10⁹, 10⁹ and 10⁸ c.f.u. g⁻¹ respectively in agbelima. All isolates and a control sample, produced agbelima with pH of between 4.11 and 3.82 in 48 h, and ANOVA showed no significant difference between the pH of the samples at P ≤ 0.05. Spontaneous fermentation of the reconstituted dough was possible, because the flour was found to contain Lactobacillus plantarum at a level of 10⁶ c.f.u. g⁻¹ as well as the other species isolated from agbelima. Counts of Gram-positive catalase-negative rods and cocci on MRS in all the fermented samples were at levels of 10⁸ to 10⁹ c.f.u. g⁻¹, with ANOVA showing no significant difference between the populations at P ≤ 0.05. Three enteric pathogens, Salmonella typhimurium 9, Echerichia coli D2188 and Vibrio cholerae C-230, inoculated into the samples at 10⁶ to 10⁷ c.f.u. g⁻¹ at the start of fermentation, could not be detected in 10 g of any of the samples at the end of 48 h fermentation. A taste panel preferred banku - a stiff porridge made from a combination of fermented maize dough and fermented cassava dough – prepared with fermented HQCF dough to banku prepared with a market sample of agbelima or reconstituted agbelima flour. Processing the highly perishable cassava roots into high quality cassava flour, therefore, offers a means for preserving cassava, which can subsequently be used for both industrial and traditional purposes.     

Effect of starter culture inoculation on feed hygiene and microbial population development in fermented pig feed composed of a cereal grain mix with wet wheat distillers' grain

Aims:  Investigating the influence of an added starter culture on the properties of fermented liquid pig feed.
Methods and Results:  Diets of cereal grain blended with wet wheat distillers' grain that were either not inoculated (WWDG), inoculated with a silage starter culture at start (WWDGsc1) or at start and at each backslopping (replacement of 80% the content with fresh mixture, simulating feed outtake, WWDGsc5) were fermented for 5 days, followed by 5 days of daily backslopping. Numbers of undesirable micro-organisms (enterobacteria, moulds) were reduced in all fermentations; particularly enterobacteria in the starter culture inoculated diets. Lactobacillus plantarum present in the starter culture became dominant in diets WWDGsc1 and WWDGsc5. However, Lactobacillus panis that was dominating WWDG was also abundant in WWDGsc1 and WWDGsc5. Yeast populations were not influenced by the starter culture, with Pichia fermentans dominating all fermentations. All diets had similar chemical characteristics with the exception of a significant increase of all tested organic acids in WWDGsc5.
Conclusions:  The addition of a starter culture influences the bacterial population in fermented liquid feed, but there is also a strong impact of the flora already present in the feed ingredients. The yeast population is not influenced by adding a lactic acid bacteria (LAB) starter culture. A consortium of LAB and yeast strains adapted to the fermentation should be used as starter culture.
Significance and Impact of the Study:  The results suggest that it is possible to influence the current unpredictable and spontaneous process of feed fermentation when appropriate starter cultures are used. For this purpose, LAB and yeasts with desirable characteristics should be isolated.     

益生乳酸菌Lactobacillus casei Zhang和Lactobacillus plantarum P8对全价饲料pH及微生物类群变化的研究

研究了益生乳酸菌干酪乳杆菌Zhang(Lactobacillus casei Zhang)和植物乳杆菌P8(Lactobacillus planta-rum P8)对全价饲料pH及微生物类群变化的影响。分别将L.casei Zhang、L.plantarum P8单一菌种及复合菌种(11)以6.30 lg cfu/g的接种总量发酵全价饲料,测定25℃10 d发酵期间全价饲料pH和微生物类群的变化,应用选择培养基测定发酵饲料中的乳酸菌及杂菌(酵母菌、霉菌、大肠菌群、芽胞杆菌和梭状芽胞杆菌)的动态变化,应用RT-PCR技术测定试验组中的L.casei Zhang和L.plantarum P8的动态变化。结果显示,试验组pH下降显著,发酵10 d时,L.casei Zhang、L.plantarum P8单一菌种和复合菌种发酵饲料的pH分别为4.23、4.24和4.22,显著低于对照组(P0.05);L.casei Zhang、L.plantarum P8单一菌种和复合菌种发酵饲料中的L.casei Zhang、L.plantarum P8活菌数分别为8.91、8.89、6.58和8.69 lg cfu/g。发酵期间,试验组中酵母菌、霉菌、大肠菌群、芽胞杆菌及梭状芽胞杆菌活菌数显著低于对照组(P0.05),其中L.plantarum P8单一菌种发酵和复合菌种发酵对杂菌抑制效果显著优于L.casei Zhang单一菌种发酵(P0.05)。结果表明,全价饲料经L.casei Zhang、L.plantarum P8发酵可以显著降低其pH,抑制其中杂菌的生长,同时L.casei Zhang、L.plantarum P8在饲料中具有良好的稳定性。     

The effect of Lactobacillus buchneri and L. plantarum, applied at ensiling, on the ensiling fermentation and aerobic stability of wheat and sorghum silages

The effect of applying Lactobacillus buchneri (LB), alone or in combinations with L. plantarum (LP) and yeasts at ensiling, on the ensiling fermentation and aerobic stability of wheat and sorghum silages was studied under laboratory conditions. Treatments comprised LB, LP, yeasts, LB + yeasts, LP + yeasts, LB + LP and B-589 (a lactic acid bacterial strain isolated from wheat silage in Israel) alone. The treatments were also applied to sterilized aqueous extracts of wheat which were incubated at 30°C for 10 days. The pH of all treatments was below 4.0 already on day 4 of the experiment. Silages treated with LB had higher acetic acid concentrations than those treated with LP: 32–34 vs 16–18, and 28–34 vs 4–7 g kg⁻¹ in the experiments with wheat and sorghum, respectively. Similar results were obtained in wheat extracts. In the aqueous phase, marked differences in pH decrease were noticed among the treatments: 4.4 in LB, 6.0 in the yeast, and 3.7 in LP and B-589 (from day 3 and onwards). In both crops LB resulted in aerobically stable silages when applied alone or with LP and yeasts, whereas LP resulted in unstable silages upon aerobic exposure; the stability of the LB-treated silages is attributed to the higher acetic acid concentrations. The isolated strain (B-589) did not exhibit any advantage with regard to aerobic stability. Received 26 April 1999/ Accepted in revised form 05 July 1999     

Bacterial community dynamics during the ensilage of wilted grass

Aims:  Grass silage is the product formed by a natural lactic acid bacterial fermentation when grass is stored under anaerobic conditions, and represents an important ruminant feedstuff on farms during winter. Of the two commonly employed methods of ensiling forage, baled silage composition frequently differs from that of comparable precision-chop silage reflecting a different ensiling environment. The aim of this study was to investigate the dynamics of the silage fermentation in wilted grass and between ensiling systems.
Methods and Results:  Fermentation dynamics were examined using traditional methods of silage analyses, including microbial enumeration and analysis of fermentation products, and culture-independent terminal restriction fragment length polymorphism (T-RFLP). A successful fermentation was achieved in both systems, with the fermentation (increase in lactic acid bacteria and lactic acid concentration, decrease in pH) proceeding rapidly once the herbage was ensiled.
Conclusions:  Under controlled conditions, little difference in silage quality and microbial composition were observed between ensiling systems and this was further reflected in the T-RFLP community analysis.
Significance and Impact of the Study:  T-RFLP proved a potentially useful tool to study the ensilage process and could provide valid support to traditional methods, or a viable alternative to these methods, for investigating the dynamics of the bacterial community over the course of the fermentation.     

Exopolysaccharides produced by mixed culture of yeast Rhodotorula rubra GED10 and yogurt bacteria (Streptococcus thermophilus 13a + Lactobacillus bulgaricus 2-11)

AIMs: The studies of the production of exopolysaccharides by lactose-negative yeast and a yogurt starter co-cultivated in a natural substrate containing lactose may be considered of interest because they reveal the possibilities for high-efficiency synthesis of biopolymers by mixed cultivation. METHODS AND RESULTS: The mixed culture Rhodotorula rubra GED10 + (Streptococcus thermophilus 13a + Lactobacillus bulgaricus 2-11) was cultivated in cheese whey ultrafiltrate (WU) (44.0 g lactose l(-1)) at initial pH 6.0, 28 degrees C, under intensive aeration (air-flow rate 1.0 l l(-1) min(-1), agitation 220 rev min(-1)) in a MBR AG fermentor. The mixed culture manifested the highest activity for synthesis of exopolysaccharides (19.3 g l(-1)) and cell mass (21.0 g l(-1)) at the 84th hour. The yogurt starter synthesized neutral exopolysaccharides, while the mixed culture yeast + yogurt starter produced acidic exopolysaccharides containing uronic acid (6%). The neutral sugar composition was identified as mannose, glucose, galactose, xylose and arabinose. Mannose dominated in the polymer composition (83%) that was produced only by the yeast (97%). CONCLUSIONS: Lactose in the WU can be effectively utilized by a co-culture of lactose-negative yeast-yogurt starter for synthesis of exopolysaccharides. SIGNIFICANCE AND IMPACT OF THE STUDY: The present findings propose an alternative use of WU as a cost-effective carbohydrate substrate, and suggest that the lactose-negative yeast Rhodotorula rubra can have industrial application as producers of exopolysaccharides.     

Experiences with the use of a starter culture in the fermentation of maize for ‘kenkey’ production in Ghana

Controlled fermentation of maize was carried out using six strains of Lactobacillus fermentum and one strain of yeast, Saccharomyces cerevisiae, isolated from traditionally fermented maize dough as starter cultures for inoculum enrichement. The fermentations were monitored by pH, acidity, microbiological analysis and taste panel evaluation of two products, kenkey and koko, prepared from the fermented doughs. The strains of L. fermentum used as starter culture dominated the microflora during fermentation and in most inoculated doughs the required pH was attained by 24 h instead of 48 h of dough fermentation. Higher contents of lactic acid bacteria and yeasts were observed in inoculated doughs at the initial stages of fermentation but the spontaneously fermented doughs attained similar lactic acid bacteria and yeasts counts by 24 h of dough fermentation. The organoleptic quality of kenkey and koko prepared from doughs fermented with starter culture for 48 h was not significantly different from the traditional products. Kenkey prepared from doughs fermented for 24 h with starter culture were found to be unacceptable by the taste panel although similarly produced koko was acceptable.The authors are with the Food Research Institute, Council for Scientific and Industrial Research, P.O Box M 20. Accra, Ghana.     

Survival, growth and persistence under farm conditions of a Lactobacillus plantarum strain inoculated into liquid pig feed

AIMS: To investigate the survival and persistence of Lactobacillus plantarum REB1 in the fermentation process of liquid pig feed on a working farm in standard production conditions. METHODS AND RESULTS: Two feed types, a control diet [nonfermented liquid feed (NFLF)] and a fermented diet [fermented liquid feed (FLF)], were compared. A rifampicin-resistant mutant L. plantarum REB1-Rif was used to initiate the fermentation of the feed. Inoculation with the experimental strain was repeated one or two times per week throughout the three month growing period. Four microbial groups were followed using standard microbiological techniques, as well as pH. Lactic acid bacteria (LAB) counts were high already at the beginning in FLF, while it took nine days to reach the corresponding LAB levels in NFLF. Yeasts were stable in FLF, whereas in NFLF there were occasional high counts during the first week. The numbers of Enterobacteriaceae were low, although in NFLF they were variable. The average pH in NFLF was 4.5 and 4 in FLF. CONCLUSIONS: The inoculation of L. plantarum REB1-Rif provided a LAB population of log 9 colony forming units (CFU) ml(-1) from the first feeding day, stable numbers of yeast and pH, and a drastic reduction of Enterobacteriaceae. SIGNIFICANCE AND IMPACT OF THE STUDY: The inoculation by L. plantarum REB1-Rif offered a FLF microbiologically stable from the first week in actual production conditions.     

Clostridia spore formation during aerobic deterioration of maize and sorghum silages as influenced by Lactobacillus buchneri and Lactobacillus plantarum inoculants

Aims:  The effect of the inoculation of maize and sorghum silages with Lactobacillus plantarum (LP) and Lactobacillus buchneri (LB) on the clostridia spore formation during aerobic deterioration has been studied.
Methods and results:  The crops were ensiled in 30 l jars, without a lactic acid bacteria inoculant (C), and with an LP or LB inocula (theoretical rate of 1 × 10⁶). After 90 days of conservation, the silages were analysed for the chemical and microbiological characteristics and subjected to an aerobic stability test, during which pH, temperature, nitrate, yeast, mould and clostridia spores were measured. Compared to the C and LP silages, yeasts were reduced in the LB silages, resulting in an increased aerobic stability. Clostridia spores, determined by most probable number (MPN) procedure, increased to 6 log₁₀ MPN g⁻¹ in the C and LP maize silages, whereas they reached 3 log₁₀ MPN g⁻¹ in C and LP sorghum silages.
Conclusions:  Clostridia spore count only slightly increased in the LB maize silages after 342 h (2·59 log₁₀ MPN g⁻¹), whereas it did not show any increase in the LB sorghum silages for the whole period of air exposure.
Significance and impact of the study:  The data indicated that clostridia spore outgrowth can take place during silo feedout in aerobic-deteriorated silages and that LB inoculation reduces the risk of clostridia outgrowth after silage opening by increasing the aerobic stability.