Changes in microflora and biochemical composition during the Baceman stage of traditional indonesian Kecap (soy sauce) production

Indonesian soy sauce (kecap) is made from black soybeans in a traditional way which involves two microbiological stages: a solid-state fermentation and a brine fermentation. This study is concerned with the brine fermentation, called baceman. Samples from different kecap producers were analyzed for (bio)chemical content and micro-organisms. It was found that the final composition of the baceman differed from manufacturer to manufacturer, and even within companies large differences were found in microflora and the amounts of fermentation products, formol nitrogen and salt concentration. The main fermentation products were lactate, acetate, glycerol and ethanol. Pediococcus halophilus, staphylococci, a coryneform bacterium and yeasts belonging to Candida, Debaromyces and Sterigmatomyces were isolated from the brines. Compared to Japanese soy sauce production, fermentation by yeasts does not play an important role in Indonesian kecap production. This is due to the fact that kecap is made from whole soybeans only, which are poor in sugars. After fermentation by P. halophilus no substrates are left for growth and ethanol production by yeasts. The presence of film forming yeasts can even lead to spoilage of the product.     

Influence of prebrining treatments on microbial and biochemical changes during the baceman stage in Indonesian kecap (soy sauce) production

Prior to the baceman stage or brine fermentation in Indonesian kecap (soy sauce) production, the soybeans undergo a number of treatments, such as soaking, cooking and a fungal solid state fermentation (bungkil stage). The influence of these prebrining steps on the baceman stage was investigated. Acidification during soaking inhibited bacterial growth during the solid state fermentation, causing lower initial numbers of bacteria in and a slower acidification of the baceman. Cooking time influenced the softness of the beans and thus fungal growth during the bungkil stage. Increasing the cooking time from 1 to 3 h resulted in a better solubility of macromolecules and gave higher final concentrations of fermentation products and formol nitrogen in baceman. When cooking time was increased from 3 to 5 h only slight differences could be observed. Length of the bungkil stage had some minor influences; the amount of fermentation products and residual monosaccharides decreased by 16% when the length was increased from 2 to 7 d. The highest amounts of formol nitrogen were found with 5 d of bungkil fermentation. The addition of rice flour showed that the absence of yeast fermentation in baceman is related to the lack of sugars after the previous lactic acid fermentation. In addition to yeast fermentation, the growth of fungi during the bungkil stage was also positively influenced by the addition of rice flour. Sun drying and crumbling bungkil caused lower enzyme activities and resulted in lower amounts of formol nitrogen and glycerol. Growth of Pediococcus halophilus during the baceman stage was faster when bungkil was sun dried, although the final concentrations of fermentation products were lower due to a lower content of amino acids which resulted in less buffering. Increasing the amount of bungkil added to salty water caused a similar increase in formol nitrogen and fermentation products.     

Modeling growth and off-flavours production of spoiled beer bacteria, Pectinatus frisingensis

The genus Pectinatus includes strictly anaerobic Gram-negative non-spore-forming mesophilic bacteria often referred to as beer-spoilage bacteria. Pectinatus frisingensis was chosen as the reference strain. The organisms were grown in batch cultures under stringent anaerobic conditions in a synthetic medium and with pH regulation. Various glucose and lactate concentrations were used, and a low inoculum reproduced spoilage conditions in bottled beer. Propionate and acetate are the major compounds responsible for the off-flavour of beer. Gompertz curves were fitted to acid-biomass production and glucose consumption; thereby the lag-phase, production rate and final concentrations were derived. Volatile fatty acids production began 19 h after biomass growth. There was no lineareffect of substrate on final concentration of propionate, acetate and biomass. An additive model is proposed for the prediction of bacterial growth and acid production on both glucose and lactate.     

Prevention of Yeast Spoilage in Feed and Food by the Yeast Mycocin HMK

The yeast Williopsis mrakii produces a mycocin or yeast killer toxin designated HMK; this toxin exhibits high thermal stability, high pH stability, and a broad spectrum of activity against other yeasts. We describe construction of a synthetic gene for mycocin HMK and heterologous expression of this toxin in Aspergillus niger. Mycocin HMK was fused to a glucoamylase protein carrier, which resulted in secretion of biologically active mycocin into the culture media. A partial purification protocol was developed, and a comparison with native W. mrakii mycocin showed that the heterologously expressed mycocin had similar physiological properties and an almost identical spectrum of biological activity against a number of yeasts isolated from silage and yoghurt. Two food and feed production systems prone to yeast spoilage were used as models to assess the ability of mycocin HMK to act as a biocontrol agent. The onset of aerobic spoilage in mature maize silage was delayed by application of A. niger mycocin HMK on opening because the toxin inhibited growth of the indigenous spoilage yeasts. This helped maintain both higher lactic acid levels and a lower pH. In yoghurt spiked with dairy spoilage yeasts, A. niger mycocin HMK was active at all of the storage temperatures tested at which yeast growth occurred, and there was no resurgence of resistant yeasts. The higher the yeast growth rate, the more effective the killing action of the mycocin. Thus, mycocin HMK has potential applications in controlling both silage spoilage and yoghurt spoilage caused by yeasts.     

Oxygen Requirements of the Food Spoilage Yeast Zygosaccharomyces bailii in Synthetic and Complex Media

Most yeast species can ferment sugars to ethanol, but only a few can grow in the complete absence of oxygen. Oxygen availability might, therefore, be a key parameter in spoilage of food caused by fermentative yeasts. In this study, the oxygen requirement and regulation of alcoholic fermentation were studied in batch cultures of the spoilage yeast Zygosaccharomyces bailii at a constant pH, pH 3.0. In aerobic, glucose-grown cultures, Z. bailii exhibited aerobic alcoholic fermentation similar to that of Saccharomyces cerevisiae and other Crabtree-positive yeasts. In anaerobic fermentor cultures grown on a synthetic medium supplemented with glucose, Tween 80, and ergosterol, S. cerevisiae exhibited rapid exponential growth. Growth of Z. bailii under these conditions was extremely slow and linear. These linear growth kinetics indicate that cell proliferation of Z. bailii in the anaerobic fermentors was limited by a constant, low rate of oxygen leakage into the system. Similar results were obtained with the facultatively fermentative yeast Candida utilis. When the same experimental setup was used for anaerobic cultivation, in complex YPD medium, Z. bailii exhibited exponential growth and vigorous fermentation, indicating that a nutritional requirement for anaerobic growth was met by complex-medium components. Our results demonstrate that restriction of oxygen entry into foods and beverages, which are rich in nutrients, is not a promising strategy for preventing growth and gas formation by Z. bailii. In contrast to the growth of Z. bailii, anaerobic growth of S. cerevisiae on complex YPD medium was much slower than growth in synthetic medium, which probably reflected the superior tolerance of the former yeast to organic acids at low pH.     

Effects of culture conditions on Pectinatus cerevisiiphilus and Pectinatus frisingensis metabolism: a physiological and statistical approach

The genus Pectinatus has been often reported in beer spoilage with off-flavours. The bacteria are strictly anaerobic, Gram-negative rods. Propionate and acetate are the main fermentation products from glucose in the two species belonging to the genus, P. cerevisiiphilus and P. frisingensis. Amino acids routinely present at a high level in beer were not growth substrates for both species, and a significant accumulation of succinate was observed with lactate as growth substrate. Both Pectinatus ssp. showed almost identical fermentation balances on glucose. Growth kinetics of both glucose-grown species were unchanged under a N₂, H₂ or 20% CO₂-containing atmosphere. Combinations of culture medium pH values from pH 3·9 to pH 7·2, of glucose levels between 5 and 55 mmol l^-1, and of lactate concentrations varied from 4 to 40 mmol l^-1 demonstrated that biomass and volatile fatty acids production were proportional to glucose concentration for both Pectinatus species. A significant increase of volatile fatty acid production was measured for both species at the lowest pH values with a lactate or a glucose concentration increase. The maximum biomass production was observed at pH 6·2 for P. cerevisiiphilus , and between pH 4·5 and pH 4·9 for P. frisingensis. Glucose and lactate or pH value were dependent with regard to propionate and acetate production in P. frisingensis. On the other hand, the variations of these three parameters were independent with regard to biomass production for both strains, and to volatile fatty acids production for P. cerevisiiphilus. Addition of ethanol to glucose-grown cultures completely inhibited growth at 1·3 mol l^-1 ethanol for P. cerevisiiphilus , and at 1·8 mol l^-1 for P. frisingensis.     

Nisin Production by a Mixed-Culture System Consisting of Lactococcus lactis and Kluyveromyces marxianus

To control the pH during antimicrobial peptide (nisin) production by a lactic acid bacterium, Lactococcus lactis subsp. lactis (ATCC11454), a novel method involving neither addition of alkali nor a separation system such as a ceramic membrane filter and electrodialyzer was developed. A mixed culture of L. lactis and Kluyveromyces marxianus, which was isolated from kefir grains, was utilized in the developed system. The interaction between lactate production by L. lactis and its assimilation by K. marxianus was used to control the pH. To utilize the interaction of these microorganisms to maintain high-level production of nisin, the kinetics of growth of, and production of lactate, acetate, and nisin by, L. lactis were investigated. The kinetics of growth of and lactic acid consumption by K. marxianus were also investigated. Because the pH of the medium could be controlled by the lactate consumption of K. marxianus and the specific lactate consumption rate of K. marxianus could be controlled by changing the dissolved oxygen (DO) concentration, a cascade pH controller coupled with DO control was developed. As a result, the pH was kept constant because the lactate level was kept low and nisin accumulated in the medium to a high level compared with that attained using other pH control strategies, such as with processes lacking pH control and those in which pH is controlled by addition of alkali.     

Killer activity of Saccharomyces cerevisiae strains: partial characterization and strategies to improve the biocontrol efficacy in winemaking

Killer yeasts are considered potential biocontrol agents to avoid or reduce wine spoilage by undesirable species. In this study two Saccharomyces cerevisiae strains (Cf8 and M12) producing killer toxin were partially characterized and new strategies to improve their activity in winemaking were evaluated. Killer toxins were characterized by biochemical tests and growth inhibition of sensitive yeasts. Also genes encoding killer toxin were detected in the chromosomes of both strains by PCR. Both toxins showed optimal activity and production at conditions used during the wine-making process (pH 3.5 and temperatures of 15–25 °C). In addition, production of both toxins was higher when a nitrogen source was added. To improve killer activity different strategies of inoculation were studied, with the sequential inoculation of killer strains the best combination to control the growth of undesired yeasts. Sequential inoculation of Cf8–M12 showed a 45 % increase of killer activity on sensitive S. cerevisiae and spoilage yeasts. In the presence of ethanol (5–12 %) and SO₂ (50 mg/L) the killer activity of both toxins was increased, especially for toxin Cf8. Characteristics of both killer strains support their future application as starter cultures and biocontrol agents to produce wines of controlled quality.     

Isolation and description ofHaloanaerobium praevalens gen. nov. and sp. nov., an obligately anaerobic halophile common to Great Salt Lake sediments

A new halophilic species is described that was isolated from the hypersaline (>20%) surface sediments of Great Salt Lake, Utah, via transfer from MPN end-dilution tubes that contained a complex organic medium. The organism was an obligate anaerobe that proliferated optimally at approximately 13% salt, but did not grow significantly at <2% or ≥30% salt. It stained Gram-negative, was nonmotile, nonsporing, and contained an outer-wall membranous layer. The complex lipids of the organism were fatty acid esters that did not change dramatically during growth at 5% or 25% NaCl. The DNA base composition was 27.0±1 mol% guanosine plus cytosine. The temperature range for growth was >5°C and <60°C, the pH range was between 6.0 and 9.0. The doubling time for growth in complex medium with 25% NaCl was 7 h. The organism utilized carbohydrates, peptides, and amino acids. Butyrate, acetate, propionate. H₂, and CO₂ were the major fermentation end products formed. Glucose, mannose, fructose,n-acetyl glucosamine, and pectin were used as energy sources for growth. Methylmercaptan was produced from methionine degradation. The nameHaloanaerobium praevalens gen. nov. sp. nov. is proposed for the type strain GSL which has been deposited as DSM 2228. The taxonomic relationships ofH. praevalens to other obligate halophiles and anaerobes, as well as its biological role in the Great Salt Lake microbial ecosystem, are discussed.     

一株新型同型产乙酸菌Clostridium sp.的自养和异养生长特性

【背景】同型产乙酸菌是一类利用乙酰辅酶A途径固定CO_2合成自身细胞物质并生成乙酸、乙醇等代谢产物的厌氧菌群,其分布广泛、种类繁多且代谢多样。深入研究同型产乙酸菌菌株的代谢能力及特性,对探索该种群的生理生化特性及其环境作用至关重要。【目的】研究一株同型产乙酸菌Clostridium sp. BXX的最适培养条件及其自养与异养生长特性。【方法】设置BXX菌株培养温度10-55°C、初始pH 6.0-9.0、NaCl浓度0-2.0%、不同氮源,测定菌体细胞含量和产物生成浓度,确定菌株最适培养条件。研究BXX菌株分别以H_2/CO_2、合成气、CO、葡萄糖、1,2-丙二醇、甲酸钠、乙二醇甲醚、甘油、丙酮酸和乳酸为底物时的底物消耗、产物生成、菌体细胞含量和pH等,探究其自养和异养生长特性。【结果】BXX菌株的最适培养温度为30°C,初始pH为7.0,NaCl浓度为1.0%,氮源为酵母粉。BXX菌株能以H2/CO2、合成气、葡萄糖、1,2-丙二醇、甲酸钠、乙二醇甲醚和甘油为底物生长,不能以CO、丙酮酸或乳酸为底物生长。【结论】BXX菌株既能自养生长产乙酸,又能异养生长产乙醇。BXX菌株是乙酸发酵的优良菌种资源,有较好的工业应用潜力。     

Identification and characterization of non-saccharomyces spoilage yeasts isolated from Brazilian wines

The industry of fine wines and also locally consumed table wines is emerging in Brazil with an increasing volume and economic impact. Enologists in this region currently lack information about the prevalence and characteristics of spoilage yeasts, which may contaminate and potentially undervalue Brazilian wines. Herein, we analyzed 50 local red wines including 27 fine wines (V. vinifera) and 23 table wines (V. labrusca). Presumptive spoilage yeasts were isolated on differential medium, and classified by RFLP-PCR and sequencing of the ITS1-5.8S-ITS2 and D1/D2 26S rDNA loci. The prevalence of spoilage yeasts in fine wines (11 %) was comparable to that reported in European and US wines, and significantly lower than that observed for local table wines (70 %). The majority of isolates belonged to Brettanomyces bruxelliensis, followed by Pichia guillermondii, and more rarely Candida wickerhamii and Trigonopsis cantarelli. The Brettanomyces isolates varied greatly in off-flavor production, displayed ethanol tolerance (>10 % by volume), tolerated sulfite (≥0.68 mg/l mSO₂), and 39 % of them grew on ethanol as sole carbon source. We discuss the causes and consequences of spoilage yeasts in relation to the Brazilian wine industry.     

Fate ofescherichia coli andE. coli O157∶H7 in apple juice treated with propolis extract

Fruit juices are targets of spoilage moulds, yeasts and acid tolerant bacteria. They might be contaminated with bacteria from raw materials, environment, packaging and during the handling of the product. These contaminations have frequently resulted in the spoilage of fruit juice and consequently commercial losses. The objective of this study was to determine the influence of propolis in apple juice againstEscherichia coli andE. coli O157:H7 strains of the spoilage and pathogenic bacteria. For this purpose, apple juice was obtained from fresh apples and then was pasteurised. The pH value, titrable acidity (as % malic acid) and Brix degree of this apple juice were 3.72±0.10, 0.67±0.05% and 12.1±0.01, respectively. Propolis extract at 1,2 and 5% concentrations were tested to determine ofE. coli andE. coli O157:H7 inhibition using paper disc diffusion method. The control treatment had no propolis extract. The apple juices were contaminated with these bacteria, and the activity of propolis was observed at first, 18^th, 24^th, 48^th and 72^nd hours at 4 and 25°C. The number of cells in the tubes was counted using serial dilution method. Results indicated that propolis extract at 2 and 5% concentrations had significant antimicrobial activity againstE. coli andE. coli O157:H7, therefore we can conclude that propolis extract is worthy of further study as a natural preservative for the foods prone to microbial spoilage.     

Effects of initial lactic acid concentration, HRTs, and OLRs on bio-hydrogen production from lactate-type fermentation

A batch test and continuous operation were performed to identify the effect of lactate on hydrogen production at pH 4.5. When the initial lactic acid concentration was increased from 0 to 8 g/L in the batch test, the hydrogen yield also increased from 1.41 to 1.72 mol-H₂/mol-glucose. The system exhibited a long lag time and an insignificant hydrogen yield with 16 g-lactic acid/L. A continuous stirred tank reactor (CSTR) was operated at different organic loading rates (OLRs: 10, 15, 20 and 40 g/L/day) and hydraulic retention times (HRTs: 6, 12 and 24 h). At an OLR of 20 g-glucose/L/day and 12 h of HRT, the hydrogen yield was 1.2 mol-H₂/mol-glucose. The yield decreased with a 24 h HRT. Even though lactate was one of the major constituents of volatile fatty acids (VFAs), hydrogen production was feasible throughout the operation. Clostridium sp. was the dominant hydrogen-producing bacteria in the system.     

Biosynthesis of Lactobionic Acid in Whey-Containing Medium by Microencapsulated and Free Bacteria of Pseudomonas taetrolens

The aim of this research was to develop a method of its production from whey using bacteria of the species Pseudomonas taetrolens. Analyses of the lactobionic acid production method from whey showed that the following factors have a significant effect on its efficiency: the frequency of whey batch feeding, pH and the type of bacteria application, i.e. microencapsulated vs. free. Lactose and lactobionic acid were assayed using high performance liquid chromatography (HPLC) and liquid chromatography-electrospray ionization-tandem mass spectrometry (LC–ESI–MS). The highest concentration of lactobionic acid of 22.03 mg/cm³ was obtained when whey was batch fed at 72-h intervals, pH was maintained at 6.25 and bacteria were enclosed in alginate microcapsules.Supplementary InformationThe online version contains supplementary material available at 10.1007/s12088-021-00944-4.     

Production of carotenoids and lipids by <Emphasis Type="Italic">Rhodococcus opacus</Emphasis> PD630 in batch and fed-batch culture

Production of carotenoids by Rhodococcus opacus PD630 is reported. A modified mineral salt medium formulated with glycerol as an inexpensive carbon source was used for the fermentation. Ammonium acetate was the nitrogen source. A dry cell mass concentration of nearly 5.4 g/L could be produced in shake flasks with a carotenoid concentration of 0.54 mg/L. In batch culture in a 5 L bioreactor, without pH control, the maximum dry biomass concentration was ~30 % lower than in shake flasks and the carotenoids concentration was 0.09 mg/L. Both the biomass concentration and the carotenoids concentration could be raised using a fed-batch operation with a feed mixture of ammonium acetate and acetic acid. With this strategy, the final biomass concentration was 8.2 g/L and the carotenoids concentration was 0.20 mg/L in a 10-day fermentation. A control of pH proved to be unnecessary for maximizing the production of carotenoids in this fermentation.     

Effect of vitamin concentration on the synthesis of lactate, ethanol, pyruvate, and ethyl acetate in cells of the yeast Dipodascus magnusii

In the yeast Dipodascus magnusii, which is auxotrophic for thiamine and biotin, during cultivation on glucose with excessive thiamine concentration, pyruvate metabolism was shown to result in the synthesis of fermentation products, namely, ethanol and, to a lesser extent, lactate. Substantial synthesis of ethyl acetate was also observed under these conditions. Introduction of nicotinic acid (NA) into the medium resulted in time separation of ethanol and lactate production. It was shown that cultivation of the yeast under biotin deficiency resulted in nearly complete suppression of aerobic production of ethanol and cessation of ethyl acetate synthesis, whereas lactate synthesis was activated as early as in the first hours of cultivation. Upon introduction of NA under these conditions, lactate concentration sharply increased. These results show that the combination of thiamine and biotin with other vitamins can stimulate utilization of the pyruvate pool in yeasts towards formation of considerable amounts of lactate, which is typical only of cells of higher eukaryotes and bacteria.     

Physiological Characteristics Underlying the Distribution Patterns of Luminous Bacteria in the Mediterranean Sea and the Gulf of Elat

Physiological characteristics of luminous bacteria isolated from the Mediterranean and Gulf of Elat were compared to determine their relationship to the specific seasonal and geographic distribution patterns of these bacteria. The effects of temperature on growth rate and yield, relative sensitivity to photooxidation, resistance to high salt concentration (8%), and ability to grow in nutrient-poor conditions appear to control these patterns. The winter appearance of Photobacterium fischeri and the succession of winter and summer types of Beneckea harveyi in the eastern Mediterranean are explained by different temperature requirements for growth. Sensitivity to photooxidation explains the disappearance of P. leiognathi, present in the main body of the Gulf of Elat throughout the year, from the shallow coastal strip. B. harveyi is present in this coastal strip which is higher in nutrients and in productivity than the open waters. Competition experiments between B. harveyi and P. leiognathi in batch and continuous culture indicate that the oligotrophic P. leiognathi is outcompeted by B. harveyi in rich and even in relatively poor media. The distribution pattern found in the Bardawil hypersaline lagoon is explained by selection of salinity-resistant mutants of B. harveyi from the Mediterranean Sea.     

Parameters Affecting Solvent Production by Clostridium pasteurianum

The effect of pH, growth rate, phosphate and iron limitation, carbon monoxide, and carbon source on product formation by Clostridium pasteurianum was determined. Under phosphate limitation, glucose was fermented almost exclusively to acetate and butyrate independently of the pH and growth rate. Iron limitation caused lactate production (38 mol/100 mol) from glucose in batch and continuous culture. At 15% (vol/vol) carbon monoxide in the atmosphere, glucose was fermented to ethanol (24 mol/100 mol), lactate (32 mol/100 mol), and butanol (36 mol/100 mol) in addition to the usual products, acetate (38 mol/100 mol) and butyrate (17 mol/100 mol). During glycerol fermentation, a completely different product pattern was found. In continuous culture under phosphate limitation, acetate and butyrate were produced only in trace amounts, whereas ethanol (30 mol/100 mol), butanol (18 mol/100 mol), and 1,3-propanediol (18 mol/100 mol) were the major products. Under iron limitation, the ratio of these products could be changed in favor of 1,3-propanediol (34 mol/100 mol). In addition, lactate was produced in significant amounts (25 mol/100 mol). The tolerance of C. pasteurianum to glycerol was remarkably high; growth was not inhibited by glycerol concentrations up to 17% (wt/vol). Increasing glycerol concentrations favored the production of 1,3-propanediol.     

Characterization of sulphate-reducing bacterial populations within marine and estuarine sediments with different rates of sulphate reduction

Viable counts of sulphate-reducing bacteria, able to use a range of different growth substrates were determined in sediments from two Sea Lochs (Etive and Eil) and an estuarine site (Tay), in Scotland. The composition of the sulphate-reducing bacterial population, in terms of substrate utilization, broadly corresponded to the in situ substrates for sulphate reduction and concentration of substrates at each site. Addition of acetate, lactate, propionate, butyrate, hydrogen and glutamate/serine (20 mM) to replicate slurries from each site resulted in stimulation of the corresponding population of sulphate-reducing bacteria and the in situ rates of sulphate reduction. The metabolism of the added substrates and changes in bacterial phospholipid fatty acids (PLFA) were quantified. With the exception of acetate and hydrogen, added substrates were incompletely oxidised, producing a mixture of further substrates, which predominantly were sequentially oxidised, and resulted in the stimulation of a mixed population of sulphate-reducing bacteria. There were significant changes in the PLFA of slurries with added substrate compared to controls. Acetate was completely removed at all sites and the small increase in even chain PLFA together with the absence of stimulation of any other biomarker, indicated that acetate was oxidised by sulphate-reducing bacteria distinctly different from those using other substrates. A biomarker for Desulfobacter, 10 Methyl 16:0, was not stimulated in any of the acetate slurries or in slurries where acetate was produced. Biomarkers for the propionate utilizing Desulfobulbus sp (17:1w6, 15:1w6) were always stimulated in propionate slurries and also in lactate slurries, where partial lactate fermentation produced propionate and acetate. In lactate and glutamate / serine slurries from the Tay estuary and lactate and hydrogen slurries from Loch Etive the biomarker for Desulfovibrio sp (i17:1w7) as well as those for Desulfobulbus were stimulated. This provides direct evidence for the significance of Desulfovibrio sp. within sediment slurries and demonstrates the competitive interaction between members of this genus and Desulfobulbus sp. for lactate, hydrogen and amino acid metabolism. At the estuarine site, sulphate reduction was limited at higher sulphate concentrations (about 3.5 mM) than the Sea Loch sites (<2 mM) and this had a significant effect on propionate and butyrate metabolism, as well as on methane production. These results demonstrate that although the sulphate-reducing bacterial population at each site could metabolise identical substrates, the types of sulphate-reducing bacteria involved and their sulphate thresholds were characteristically different.     

Staphylococci in Competition: III. Influence of pH and Salt on Staphylococcal Growth in Mixed Populations

Previous results showed definite repressive effects on the growth of staphylococci in mixed cultures due to the competitive growth of psychrophilic saprophytes. This study was continued, and the influence of other environmental factors, pH and salt, on the competition between staphylococci and saprophytes was investigated. Initial pH values varied from 5 to 9. At the extremes of the pH range, staphylococci failed to grow, while the saprophytes grew under all of the conditions tested. At pH 5, the growth curves for the saprophytes were markedly altered from those obtained at neutral pH. The lag phases were greatly lengthened at and below 20 C, but normal numbers of saprophytes were reached in the stationary phase. At pH 6 and 8, staphylococcal growth showed the same inhibition observed at pH 7, at and below 20 C; normal multiplication was observed above this temperature, but with accelerated death phases. Thus, pH did not primarily effect staphylococcal growth through its influence on saprophyte growth and competition, but rather directly affected the growth of Staphylococcus cultures. Salt concentrations from 3.5 to 9.5% were investigated for influence on staphylococcal growth in mixed populations. Above 3.5% salt, staphylococcal inhibition at and above 20 C was not as marked as in the controls, although normal numbers were never reached. The saprophytes were increasingly inhibited, and their lag phases materially lengthened as salt concentration was increased. Salt acted directly on the Staphylococcus population and also, by repressing saprophyte growth, decreased competition, which allowed the staphylococci to grow.