Isolation and characterization of a novel acidophilic zero-valent sulfur- and ferric iron-respiring Firmicute

A novel, obligately anaerobic, acidophilic bacterium (strain I2511), isolated from sediment in an abandoned copper mine, was shown to couple the oxidation of organic electron donors to the reduction of both zero-valent sulfur and ferric iron in acidic media. The isolate was an obligate heterotroph that used a variety of organic compounds as electron donors and required yeast extract for growth. Alternative electron acceptors (sulfate, tetrathionate, thiosulfate and nitrate) were not used by the novel isolate. The strain grew as motile, endospore-forming rods, and was mesophilic and moderately acidophilic, with a growth rate of 0.01 h⁻¹ at optimum pH (3.7) and temperature (35 °C). Analysis of its 16S rRNA gene sequence placed strain I2511 within the phylum Firmicutes, distantly related to validated species. Phylogenetic analysis and physiological traits indicate that the novel strain represents a species of a candidate novel genus. Strain I2511 was included in a microbial consortium in a low pH “hybrid” sulfidogenic bioreactor designed to remove chalcophilic metals from metal-contaminated liquors and was present in >50% relative abundance when bioreactor was operated at pH ∼ 2.0. Results indicate that the novel isolate could be applied in biotechnologies to treat acidic and neutral pH, metal-rich effluents.     

Quantitative proteomics using SWATH-MS identifies mechanisms of chloride tolerance in the halophilic acidophile Acidihalobacter prosperus DSM 14174

In this study, the differential protein expression of the acidophilic halophile, Acidihalobacter prosperus DSM 14174 (strain V6) was studied with the aim of understanding its mechanisms of tolerance to high chloride ion stress in the presence of low pH, using Sequential Window Acquisition of all Theoretical Mass Spectra (SWATH-MS). In acidophiles, chloride stress results in both osmotic stress as well as acidification of the cytoplasm due to the ability of chloride to permeate the cell membrane and disrupt the reversed transmembrane potential which normally extrudes protons. The proteomic response of A. prosperus DSM 14174 to elevated chloride concentrations included the production of osmotic stress regulators that potentially induced the production of compatibles solutes, of which the most significant increase was in the synthesis of ectoine. Other responses directly related to the increased chloride and acid stress, included the increased synthesis of glutathione, changes in carbon flux, the increased production of amino acids, the decreased production of ribosomal proteins, the efflux of metals and protons, and the increase in proteins involved in DNA repair and membrane biosynthesis. Energy generation through iron oxidation and sulphur oxidation were decreased, and energy was probably obtained from the metabolism of glycogen stores. Overall, these studies have helped to create a model of tolerance to elevated chloride under acidic conditions by A. prosperus DSM 14174 that differs from the previous model developed for the type strain, A. prosperus DSM 5130^T.     

Variables Affecting Results of Sodium Chloride Tolerance Test for Identification of Rapidly Growing Mycobacteria

The sodium chloride tolerance test is often used in the identification of rapidly growing mycobacteria, particularly for distinguishing between Mycobacterium abscessus and Mycobacterium chelonae. This test, however, is frequently unreliable for the identification of some species. In this study we examined the following variables: medium manufacturer, inoculum concentration, and atmosphere and temperature of incubation. Results show that reliability is improved if the test and control slants are inoculated with an organism suspension spectrophotometrically equal to a 1 McFarland standard. Slants should be incubated at 35°C in ambient air and checked weekly for 4 weeks. Growth on control slants should be critically evaluated to determine the adequacy of the inoculum; colonies should number greater than 50. Salt-containing media should be examined carefully to detect pinpoint or tiny colonies, and colonies should number greater than 50 for a positive reaction. Concurrent use of a citrate slant may be helpful for distinguishing between M. abscessus and M. chelonae. Molecular methodologies are probably the most reliable means for the identification of rapidly growing mycobacteria and should be used, if possible, when unequivocal species identification is of particular importance.     

Isolation and characterisation of mineral-oxidising “Acidibacillus” spp. from mine sites and geothermal environments in different global locations

Eight strains of acidophilic bacteria, isolated from mine-impacted and geothermal sites from different parts of the world, were shown to form a distinct clade (proposed genus “Acidibacillus”) within the phylum Firmicutes, well separated from the acidophilic genera Sulfobacillus and Alicyclobacillus. Two of the strains (both isolated from sites in Yellowstone National Park, USA) were moderate thermophiles that oxidised both ferrous iron and elemental sulphur, while the other six were mesophiles that also oxidised ferrous iron, but not sulphur. All eight isolates reduced ferric iron to varying degrees. The two groups shared <95% similarity of their 16S rRNA genes and were therefore considered to be distinct species: “Acidibacillus sulfuroxidans” (moderately thermophilic isolates) and “Acidibacillus ferrooxidans” (mesophilic isolates). Both species were obligate heterotrophs; none of the eight strains grew in the absence of organic carbon. “Acidibacillus” spp. were generally highly tolerant of elevated concentrations of cationic transition metals, though “A. sulfuroxidans” strains were more sensitive to some (e.g. nickel and zinc) than those of “A. ferrooxidans”. Initial annotation of the genomes of two strains of “A. ferrooxidans” revealed the presence of genes (cbbL) involved in the RuBisCO pathway for CO₂ assimilation and iron oxidation (rus), though with relatively low sequence identities.     

Detection,identification and typing of Acidithiobacillus species and strains: a review

The genus Acidithiobacillus comprises several species of Gram-negative acidophilic bacteria that thrive in natural and man-made low pH environments in a variety of geo-climatic contexts. Beyond their fundamental interest as model extreme acidophiles, these bacteria are involved in the processing of minerals and the desulfurization of coal and natural gas, and are also sources of environmental pollution due to their generation of acid mine drainage and corrosion of cement and concrete structures. Acidithiobacillus spp. are therefore considered a biotechnologically relevant group of bacteria, and their identification and screening in natural and industrial environments is of great concern.Several molecular typing methodologies have been instrumental in improving knowledge of the inherent diversity of acidithiobacilli by providing information on the genetic subtypes sampled in public and private culture collections; more recently, they have provided specific insight into the diversity of acidithiobacilli present in industrial and natural environments. The aim of this review is to provide an overview of techniques used in molecular detection, identification and typing of Acidithiobacillus spp. These methods will be discussed in the context of their contribution to the general and specific understanding of the role of the acidithiobacilli in microbial ecology and industrial biotechnology. Emerging opportunities for industrial and environmental surveillance of acidithiobacilli using next-generation molecular typing methodologies are also reviewed.     

Effect of inoculum history,growth substrates and yeast extract addition on inhibition of Sulfobacillus thermosulfidooxidans by NaCl

This study reports on the effect of inoculum history, growth substrates, and yeast extract on sodium chloride tolerance of Sulfobacillus thermosulfidooxidans DSM 9293^T. The concentrations of NaCl for complete inhibition of Fe²⁺ oxidation by cells initially grown with ferrous iron sulfate, or tetrathionate, or pyrite as energy sources were 525 mM, 725 mM, and 800 mM, respectively. Noticeably, regardless of NaCl concentrations, oxygen consumption rates of S. thermosulfidooxidans with 20 mM tetrathionate were higher than with 50 mM FeSO₄. NaCl concentrations of higher than 400 mM strongly inhibited the iron respiration of S. thermosulfidooxidans. In contrast, the presence of NaCl was shown to stimulate tetrathionate oxidation. This trend was especially pronounced in NaCl-adapted cells where respiration rates at 200 mM NaCl were threefold of those in the absence of NaCl. In NaCl-adapted cultures greater respiration rates for tetrathionate were observed than in non-NaCl-adapted cultures, especially at concentrations ≥ 200 mM NaCl. At concentrations of ≤ 200 mM NaCl, cell growth and iron oxidation were enhanced with the addition of increasing concentrations of yeast extract. Thus, cell numbers in cultures with 0.05% yeast extract were ∼5 times higher than without yeast extract addition. At NaCl concentration as high as 400 mM, however, iron oxidation rates improved compared to control assays without yeast extract, but there was no clear dependence on yeast extract concentrations. The initial growth of bacteria with and without yeast extract in the presence of different NaCl concentrations was shown to impact leaching of copper from chalcopyrite. Copper dissolution was enhanced in the presence of 200 mM NaCl and absence of yeast extract, while the addition of 0.02% yeast extract was shown to promote copper solubilization in the presence of 500 mM NaCl.     

Expression of the atpD gene in probiotic Lactobacillus plantarum strains under in vitro acidic conditions using RT-qPCR

F₁F₀-ATPase has been identified as an operon directly involved in the tolerance of probiotic bacteria towards a hostile acidic environment encountered in the stomach. Expression of atpD (a key part of the F₁F₀-ATPase operon) gene of the two putative probiotic Lactobacillus plantarum isolates (Lp9 and Lp91) under different in vitro pH conditions which closely mimic the physiological environment prevalent in the human gut was investigated by quantitative real-time PCR (RT-qPCR). A battery of housekeeping genes, i.e. gapB, dnaG, gyrA, ldhD, rpoD and 16S rRNA, were evaluated using geNorm 3.4 Excel-based application for normalizing atpD gene expression in Lp9 and Lp91. The most stably expressed genes were found to be gapB, gyrA and ldhD. Although both putative probiotic L. plantarum isolates investigated in this study were able to survive acid stress under in vitro conditions, amongst the two, Lp91 exhibited relatively greater acid tolerance, as revealed by 4.7-fold upregulation of the atpD gene as well as higher log counts at pH 2.5 after 90 min These results clearly demonstrate that expression of the ‘atp’ operon was chiefly instrumental in in vitro survival and tolerance of test cultures at acidic conditions encountered in the stomach.     

Microorganisms in subterranean acidic waters within Europe's deepest metal mine

The Pyhäsalmi mine, central Finland, has operated as a deep metal mine since 1967. It currently reaches a depth of almost 1500 m, making it the deepest mining operation in Europe. Around 900,000 m³ of metal-rich, extremely acidic water are pumped out of the mine each year. The near constant air temperature of ∼24 °C together with exposure of sulfidic rock surfaces to air and water, have created an environment that is highly suitable for colonization by acidophilic mineral-oxidizing microorganisms. Using a combined cultivation-dependent and molecular approach, indigenous bacteria in waters at two depths within the mine, and of an acid streamer sample were identified and isolated. Iron-oxidizing chemolithotrophs (Acidithiobacillus and Leptospirillum spp., and “Ferrovum myxofaciens” were the most abundant bacteria in mine water samples, whereas the acid streamer community contained a greater proportion of heterotrophic acidophiles (Ferrimicrobium acidiphilum and a gammaproteobacterium related to Metallibacterium scheffleri). The most abundant isolates obtained from both water and streamer samples were all strains of Acidithiobacillus Group IV, a proposed separate species of iron-oxidizing acidithiobacilli that has not yet been classified as such. Archaea were also detected in water and streamer samples using molecular methods, but most were not identified and no isolates were obtained.     

Flagellin from Marinobacter algicola and Vibrio vulnificus activates the innate immune response of gilthead seabream

Adjuvants have emerged as the best tools to enhance the efficacy of vaccination. However, the traditional adjuvants used in aquaculture may cause adverse alterations in fish making necessary the development of new adjuvants able to stimulate the immune system and offer strong protection against infectious pathogens with minimal undesirable effects. In this respect, flagellin seems an attractive candidate due to its ability to strongly stimulate the immune response of fish. In the present study, we have evaluated the ability of recombinant flagellin from Marinobacter algicola (MA) and Vibrio vulnificus (Vvul), a non-pathogenic and a pathogenic bacteria, respectively, to stimulate the innate immune system of gilthead seabream (Sparus aurata L.) and compare the effect with that of the classical flagellin from Salmonella enterica serovar Typhimurium (Salmonella Typhimurium, STF). Intraperitoneal injection of MA and Vvul resulted in a strong inflammatory response characterized by increased reactive oxygen species production and the infiltration of acidophilic granulocytes at the injection site. Interestingly, however, only flagellin from MA consistently induced the expression of the gene encoding pro-inflammatory interleukin-1β. These effects were further confirmed in vitro, where a dose-dependent activation of macrophages and acidophilic granulocytes by MA and Vvul flagellins was observed. In contrast, STF flagellin was found to be less potent in both in vivo and in vitro experiments. Our results suggest the potential use of MA and Vvul flagellins as immunostimulants and adjuvants for fish vaccination.     

Sulfoacidibacillus ferrooxidans,gen. nov., sp. nov., Sulfoacidibacillus thermotolerans,gen. nov., sp. nov., and Ferroacidibacillus organovorans,gen. nov., sp. nov.: Extremely acidophilic chemolitho-heterotrophic Firmicutes

Ten strains of extremely acidophilic bacteria, isolated from different environments form a distinct monophyletic clade within the phylum Firmicutes. Comparison of complete genomes of the proposed type strains confirm that they comprise two genera (proposed names Sulfoacidibacillus and Ferroacidibacillus), and at least three species (Sulfoacidibacillus ferrooxidans, Sulfoacidibacillus thermotolerans and Ferroacidibacillus organovorans). The bacterial strains share some physiological traits, including catalysing the dissimilatory oxidation and reduction of iron, and in being obligately heterotrophic. Both species of Sulfoacidibacillus are also able to oxidise elemental sulfur and tetrathionate. Both S. ferrooxidans and Ferroacidibacillus spp. are mesophilic, while S. thermotolerans isolates are moderate thermophiles. The isolates display different degrees of acid-tolerance: Ferroacidibacillus spp. are the most acid-sensitive while the type strain of S. ferrooxidans grows at pH 0.9. MK7 was detected as the sole menaquinone present in all three nominated type strains, and their peptidoglycans all contain meso-2,6 diaminopimelic acid type A1γ. The chromosomal DNA of the strains examined contain between 44 and 52 mol% G + C. The nominated type strains of the new species are S. ferrooxidans S⁰AB^T (= DSM 105355^T = JCM 33225^T); S. thermotolerans Y002^T (= ATCC TSD-104^T = JCM 31946^T); F. organovorans SLC66^T (= ATCC TSD-103^T = JCM 31945^T).     

Microbiology of hip and knee periprosthetic joint infections: a database study

ObjectivesKnowledge of the microbiological aetiology of periprosthetic joint infection (PJI) is essential to its management. Contemporary literature from the United States on this topic is lacking. This study aimed to identify the most common microorganisms associated with types of arthroplasty, the timing of infection, and clues to polymicrobial infection.MethodsWe performed an analytical cross-sectional study of patients 18 years of age or older with hip or knee PJI diagnosed at our institution between 2010 and 2019. PJI was defined using the criteria adapted from those of the Musculoskeletal Infection Society. Cases included PJI associated with primary or revision arthroplasty and arthroplasty performed at our institution or elsewhere.ResultsA total of 2067 episodes of PJI in 1651 patients were included. Monomicrobial infections represented 70% of episodes (n = 1448), with 25% being polymicrobial (n = 508) and the rest (5%, n = 111) culture-negative. The most common group causing PJI was coagulase-negative Staphylococcus species (other than S. ludgunensis) (37%, n = 761). The distribution of most common organisms was similar regardless of arthroplasty type. The S. aureus complex, Gram-negative bacteria, and anaerobic bacteria (other than Cutibacterium species) were more likely to be isolated than other organisms in the first year following index arthroplasty (OR 1.7, 95%CI 1.4–2.2; OR 1.5, 95%CI 1.1–2.0; and OR 1.5, 95%CI 1.0–2.2, respectively). The proportion of culture-negative PJIs was higher in primary than revision arthroplasty (6.5% versus 3%, p 0.0005). The presence of a sinus tract increased the probability of the isolation of more than one microorganism by almost three-fold (OR 2.6, 95%CI 2.0–3.3).ConclusionsJoint age, presence of a sinus tract, and revision arthroplasties influenced PJI microbiology.     

Biofilm formation,communication and interactions of leaching bacteria during colonization of pyrite and sulfur surfaces

Bioleaching of metal sulfides is an interfacial process where biofilm formation is considered to be important in the initial steps of this process. Among the factors regulating biofilm formation, molecular cell-to-cell communication such as quorum sensing is involved. A functional LuxIR-type I quorum sensing system is present in Acidithiobacillus ferrooxidans. However, cell-to-cell communication among different species of acidophilic mineral-oxidizing bacteria has not been studied in detail. These aspects were the scope of this study with emphasis on the effects exerted by the external addition of mixtures of synthetic N-acyl-homoserine-lactones on pure and binary cultures. Results revealed that some mixtures had inhibitory effects on pyrite leaching. Some of them correlated with changes in biofilm formation patterns on pyrite coupons. We also provide evidence that A. thiooxidans and Acidiferrobacter spp. produce N-acyl-homoserine-lactones. In addition, the observation that A. thiooxidans cells attached more readily to pyrite pre-colonized by living iron-oxidizing acidophiles than to heat-inactivated or biofilm-free pyrite grains suggests that other interactions also occur. Our experiments show that pre-cultivation conditions influence A. ferrooxidans attachment to pre-colonized pyrite surfaces. The understanding of cell-to-cell communication may consequently be used to develop attempts to influence biomining/bioremediation processes.     

Arsenic‐tolerant microbial consortia from sediments of Copahue geothermal system with potential applications in bioremediation

Although arsenic (As) is recognized as a toxic element for living species, some microorganisms have the ability to tolerate and transform it; recent studies have proposed to take advantage of such capacity to develop sustainable bioremediation strategies. In this study, we evaluated the adaptation to increasing concentrations of As(III) and As(V) of three metabolically different microbial cultures (heterotrophic, autotrophic–acidophilic, and anaerobic) obtained from a sample with low‐soluble As content from the Copahue geothermal system. At the end of the adaptation process, the heterotrophic culture was able to grow at 20 mM and 450 mM of As(III) and As(V), respectively; the autotrophic–acidophilic culture showed tolerance to 15 mM of As(III) and 150 mM of As(V), whereas the anaerobic culture only developed in As(V) at concentrations up to 50 mM. The most tolerant consortia were characterized by their growth performance, complexity, and the presence of genes related to As metabolism and resistance. Regarding the consortia complexity, the predominant genera identified were: Paenibacillus in both heterotrophic consortia, Acidithiobacillus in the autotrophic–acidophilic consortium tolerant to As(III), Acidiphilium in the autotrophic–acidophilic consortium tolerant to As(V), and Thiomonas and Clostridium in the anaerobic consortium. This study is the first report of As tolerance microorganisms obtained from Copahue and reasserts the versatility and flexibility of the community of this natural extreme environment; also, it opens the door to the study of possible uses of these consortia in the design of biotechnological processes where the As concentration may fluctuate.     

The effect of acid treatment on survival and protein expression of a laboratory K-12 strain Escherichia coli

Pre-exposure of log phase enteric bacteria to nonlethal acidic pH induces phenotypic changes that protect the organisms against subsequent lethal acidity. Studies have revealed that when Salmonella typhimurium is grown in minimal medium at pH 5.5 and 4.3 the organism develops a biphasic acid tolerance. This two-stage response has not been reported at present in Escherichia coli; rather it is thought that when this organism is grown in rich medium there is a single stress response throughout the pH range of 4 to 6. We believe that the evidence for such a report is lacking; therefore, in this study the acid response of log phase E. coli was examined in rich medium (LB). The pH 3.0 acid survival assays of a laboratory strain of E. coli K-12 MG1655, after cultures had been exposed to LB acidified to pH 5.5 or pH 4.3 indicate that like S. typhimurium, E. coli shows both an acid tolerance and an acid-shock response to pH 5.5 and 4.3 exposure, respectively. It was consistently found, however, that longer pre-exposure (60 min rather than 15 min) at either pH afforded better protection against the lethal pH 3.0 challenge. Analysis of polypeptide induction at pH 5.5 and 4.3 by two-dimensional gel electrophoresis clearly shows different profiles. Together the results show that in E. coli, pre-treatment between pH 4 and 6 does not result in a flat protective response.     

Microbial diversity at the moderate acidic stage in three different sulfidic mine tailings dumps generating acid mine drainage

In freshly deposited sulfidic mine tailings the pH is alkaline or circumneutral. Due to pyrite or pyrrhotite oxidation the pH is dropping over time to pH values <3 at which acidophilic iron- and sulfur-oxidizing prokaryotes prevail and accelerate the oxidation processes, well described for several mine waste sites. The microbial communities at the moderate acidic stage in mine tailings are only scarcely studied. Here we investigated the microbial diversity via 16S rRNA gene sequence analysis in eight samples (pH range 3.2–6.5) from three different sulfidic mine tailings dumps in Botswana, Germany and Sweden. In total 701 partial 16S rRNA gene sequences revealed a divergent microbial community between the three sites and at different tailings depths. Proteobacteria and Firmicutes were overall the most abundant phyla in the clone libraries. Acidobacteria, Actinobacteria, Bacteroidetes, and Nitrospira occurred less frequently. The found microbial communities were completely different to microbial communities in tailings at <pH 3 described in the literature.     

A new genome of Acidithiobacillus thiooxidans provides insights into adaptation to a bioleaching environment

Acidithiobacillus thiooxidans is a sulfur oxidizing acidophilic bacterium found in many sulfur-rich environments. It is particularly interesting due to its role in bioleaching of sulphide minerals. In this work, we report the genome sequence of At. thiooxidans Licanantay, the first strain from a copper mine to be sequenced and currently used in bioleaching industrial processes. Through comparative genomic analysis with two other At. thiooxidans non-metal mining strains (ATCC 19377 and A01) we determined that these strains share a large core genome of 2109 coding sequences and a high average nucleotide identity over 98%. Nevertheless, the presence of 841 strain-specific genes (absent in other At. thiooxidans strains) suggests a particular adaptation of Licanantay to its specific biomining environment. Among this group, we highlight genes encoding for proteins involved in heavy metal tolerance, mineral cell attachment and cysteine biosynthesis. Several of these genes were located near genetic motility genes (e.g. transposases and integrases) in genomic regions of over 10 kbp absent in the other strains, suggesting the presence of genomic islands in the Licanantay genome probably produced by horizontal gene transfer in mining environments.     

Influence of various uropathogens on crystallization of urine mineral components caused by Proteus mirabilis

Infectious urolithiasis is a consequence of long-standing urinary tract infections with urease-positive bacteria, especially Proteus spp. However, because of the often mixed nature of urinary tract infections, in the case of urinary stones formation, several species of bacteria may be involved in the process. The purpose of the study was to determine the impact of the bacterial species: Escherichia coli, Klebsiella pneumoniae, Providencia stuartii, Pseudomonas aeruginosa and Staphylococcus aureus on the crystallization caused by Proteus mirabilis. The studies were conducted in synthetic urine with the addition of P. mirabilis and a representative of another species. During the experiments the viability of bacteria, pH, presence and morphology of crystals, and the intensity of crystallization were assessed. Crystallization of calcium and magnesium phosphates occurred in all investigated configurations. However, there were differences observed in the course and intensity of crystallization between the mixed culture and the P. mirabilis culture. Although most intense crystallization took place in the pure culture of P. mirabilis it was also demonstrated that the presence of other uropathogens increased the survival of P. mirabilis. This synergistic effect could be responsible for the persistence and recurrence of urolithiasis in the urinary tract.     

The multicopper oxidase of Mycobacterium tuberculosis (MmcO) exhibits ferroxidase activity and scavenges reactive oxygen species in activated THP-1 cells

The MmcO protein of Mycobacterium tuberculosis is a membrane-associated multicopper oxidase. Its natural substrate(s) and its role in pathogenesis are not well characterized. A recent report proposes that MmcO contributes to copper resistance in M. tuberculosis during infection. We have expressed and reconstituted the active enzyme from inclusion bodies in E. coli. MmcO exhibits maximal activity against the experimental substrate 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) or ABTS, at pH 4. The enzyme also exhibits ferroxidase activity at pH 4. Most notable was the finding that MmcO is able to scavenge the reactive oxygen species (ROS) generated by the xanthine/xanthine oxidase enzyme system. This ROS scavenging activity of MmcO was also evident against ROS generated by THP-1 cells. We propose that MmcO protects M. tuberculosis during infection against ROS attack in addition to providing copper resistance to the pathogen.     

Effects of in vitro conditions on the survival of Alaria alata mesocercariae

The aim of this study was to determine the effect of different concentrations of table salt (NaCl) and ethanol (v/v) solutions on the viability of Alaria alata mesocercariae. Furthermore, the survival of A. alata mesocercariae during simulated human gastric digestion was evaluated. For this purpose, A. alata mesocercariae migration technique (AMT) was used for the isolation of the parasite from high-positive A. alata mesocercariae meat from wild boar, raccoon, raccoon dog, and badger meat. In total, we have studied the behavior of 582 larvae under different conditions (NaCl, ethanol, and artificial gastric juice) in three independent in vitro experiments. The larvae survived at a NaCl concentration of up to 2.0 % until day 21 with a median survival time of 11 days. At 3.0 % NaCl concentration, the larvae lost their vitality after less than 24 h. In addition, it was found that ethanol concentrations from 8.0 to 70.0 % were effective at reducing survival of A. alata mesocercariae within a short period of time (<1 min). Finally, our studies have revealed that it required 120 min to reliably inactivate all A. alata mesocercariae within HCl-pepsin digestion solution with a pH of 1.5–2.0 at 37 °C. Consequently, the results showed that 3.0 % is the minimum concentration of NaCl in meat products recommended for human consumption because at lower NaCl concentration the parasite survived for a substantial period of time. Finally, the common concentrations of ethanol used for the disinfection of surfaces in household and/or laboratory, are sufficient for the inactivation of A. alata mesocercariae.     

pH-linked control of energy charge in Acetobacter methanolicus sp. MB 70

The influence of the extracellular pH on the adenylate system of the acidophilic Acetobacter methanolicus sp. MB 70 was investigated. The ATP level was maintained over a pH range from 3.0 to 6.0. An increased endogenous respiration was observed after a raise of the pH from 4.0 upwards. Above pH 6.0 the level of ATP decreased and the level of ADP and even more that of AMP increased. This resulted in a drop of the energy charge from 0.78 to values below 0.2. At pH 4.0 the level of ATP and AMP decreased only after prolonged incubation without an external energy source but the energy charge remained constant at about 0.7. In the range of external pH values where the energy charge is maintained the intracellular pH value is expected to be stable. The investigation of the reasons for the displacement of this range to low extracellular pH values may help to more profoundly understand the phenomenon of acidophily.