Harald Huber’s research while affiliated with University of Regensburg and other places

Background: The colonization of skin with pathogenic, partially antibiotic resistant bacteria is frequently a severe problem in dermatological therapies. For instance, skin colonisation with Staphylococcus aureus is even a disease-promoting factor in atopic dermatitis. The photodynamic inactivation (PDI) of bacteria could be a new antibacterial procedure. Upon irradiation with visible light, a special photosensitizer exclusively generates singlet oxygen. This reactive oxygen species kills bacteria via oxidation independent of species or strain and their antibiotic resistance profile causing no bacterial resistance on its part. Objective: To investigate the antibacterial potential of a photosensitizer, formulated in a new hydrogel, on human skin ex vivo. Methods: The photochemical stability of the photosensitizer and its ability to generate singlet oxygen in the hydrogel was studied. Antimicrobial efficacy of this hydrogel was tested step by step, firstly on inanimate surfaces and then on human skin ex vivo against S. aureus and Pseudomonas aeruginosa using standard colony counting. NBTC staining and TUNEL assays were performed on skin biopsies to investigate potential necrosis and apoptosis effects in skin cells possibly caused by PDI. Results: None of the hydrogel components affected the photochemical stability and the life time of singlet oxygen. On inanimate surfaces as well as on the human skin, the number of viable bacteria was reduced with up to 4.8 log10 being more effective than most other antibacterial topical agents. Histology and assays showed that PDI against bacteria on the skin surface caused no harmful effects in the underlying skin cells. Conclusion: PDI hydrogel proved to be effective for decolonization of human skin including the potential to act against superficial skin infections. Being a water-based formulation, the hydrogel should be also suitable on the mucosa. The results of the present ex vivo study form a good basis for conducting clinical studies in vivo.

A novel interdomain consortium composed of a methanogenic Archaeon and a sulfate-reducing bacterium was isolated from a microbial biofilm in an oil well in Cahuita National Park, Costa Rica. Both organisms can be grown in pure culture or as stable co-culture. The methanogenic cells were non-motile rods producing CH 4 exclusively from H 2 /CO 2. Cells of the sulfate-reducing partner were motile rods forming cell aggregates. They utilized hydrogen, lactate, formate, and pyru-vate as electron donors. Electron acceptors were sulfate, thiosulfate, and sulfite. 16S rRNA sequencing revealed 99% gene sequence similarity of strain CaP3V-M-L2A T to Methanobacterium subterraneum and 98.5% of strain CaP3V-S-L1A T to Desulfomicrobium baculatum. Both strains grew from 20 to 42 °C, pH 5.0-7.5, and 0-4% NaCl. Based on our data, type strains CaP3V-M-L2A T (= DSM 113354 T = JCM 39174 T) and CaP3V-S-L1A T (= DSM 113299 T = JCM 39179 T) represent novel species which we name Methanobacterium cahuitense sp. nov. and Desulfomicrobium aggregans sp. nov.

Photodynamic inactivation of microorganisms (PDI) finds use in a variety of applications. Several studies report on substances enhancing or inhibiting PDI. In this study, we analyzed the inhibitory potential of ubiquitous salts like CaCl2 and MgCl2 on PDI against Staphylococcus aureus and Pseudomonas aeruginosa cells using five cationic photosensitizers methylene blue, TMPyP, SAPYR, FLASH-02a and FLASH-06a. TMPyP changed its molecular structure when exposed to MgCl2, most likely due to complexation. CaCl2 substantially affected singlet oxygen generation by MB at small concentrations. Elevated concentrations of CaCl2 and MgCl2 impaired PDI up to a total loss of bacterial reduction, whereas CaCl2 is more detrimental for PDI than MgCl2. Binding assays cannot not explain the differences of PDI efficacy. It is assumed that divalent ions tightly bind to bacterial cells hindering close binding of the photosensitizers to the membranes. Consequently, photosensitizer binding might be shifted to outer compartments like teichoic acids in Gram-positives or outer sugar moieties of the LPS in Gram-negatives, attenuating the oxidative damage of susceptible cellular structures. In conclusion, CaCl2 and MgCl2 have an inhibitory potential at different phases in PDI. These effects should be considered when using PDI in an environment that contains such salts like in tap water or different fields of food industry.

Many studies show that photodynamic inactivation (PDI) is a powerful tool for the fight against pathogenic, multi‐resistant bacteria and the closing of hygiene gaps. However, PDI studies have been frequently performed under standardized in vitro conditions comprising artificial laboratory settings. Under real life conditions, however, PDI encounters substances like ions, proteins, amino acids, and fatty acids, potentially hampering the efficacy PDI to an unpredictable extent. Thus, we investigated PDI with the phenalene‐1‐one based photosensitizer SAPYR against Escherichia coli and Staphylococcus aureus in the presence of calcium or magnesium ions, which are ubiquitous in potential fields of PDI applications like in tap water or on tissue surfaces. The addition of citrate should elucidate the potential as a chelator. The results indicate that PDI is clearly affected by such ubiquitous ions depending on its concentration and the type of bacteria. The application of citrate enhanced PDI especially for Gram‐negative bacteria at certain ionic concentrations (e.g. CaCl2 or MgCl2: 7.5 to 75 mmol l‐1). Citrate also improved PDI efficacy in tap water (especially for Gram‐negative bacteria) and synthetic sweat solution (especially for Gram‐positive bacteria). In conclusion, the use of chelating agents like citrate may facilitate the application of PDI under real life conditions.

A novel methanogenic strain, CaP3V-MF-L2AT, was isolated from an exploratory oil well from Cahuita National Park, Costa Rica. The cells were irregular cocci, 0.8–1.8 μm in diameter, stained Gram-negative and were motile. The strain utilized H2/CO2, formate and the primary and secondary alcohols 1-propanol and 2-propanol for methanogenesis, but not acetate, methanol, ethanol, 1-butanol or 2-butanol. Acetate was required as carbon source. The novel isolate grew at 25–40 °C, pH 6.0–7.5 and 0–2.5% (w/v) NaCl. 16S rRNA gene sequence analysis revealed that the strain is affiliated to the genus Methanofollis. It shows 98.8% sequence similarity to its closest relative Methanofollis ethanolicus. The G + C content is 60.1 mol%. Based on the data presented here type strain CaP3V-MF-L2AT (= DSM 113321T = JCM 39176T) represents a novel species, Methanofollis propanolicus sp. nov.

Securing new sources of renewable energy and achieving national self-sufficiency in natural gas have become increasingly important in recent times. The study described in this paper focuses on three geologically diverse underground gas reservoirs (UGS) that are the natural habitat of methane-producing archaea, as well as other microorganisms with which methanogens have various ecological relationships. The objective of this research was to describe the microbial metabolism of methane in these specific anoxic environments during the year. DNA sequencing analyses revealed the presence of different methanogenic communities and their metabolic potential in all sites studied. Hydrogenotrophic Methanobacterium sp. prevailed in Lobodice UGS, members of the hydrogenotrophic order Methanomicrobiales predominated in Dolní Dunajovice UGS and thermophilic hydrogenotrophic members of the Methanothermobacter sp. were prevalent in Tvrdonice UGS. Gas composition and isotope analyses were performed simultaneously. The results suggest that the biotechnological potential of UGS for biomethane production cannot be neglected.

The antibiotic crisis increasingly threatens the health systems world-wide. Especially as there is an innovation gap in the development of novel antibiotics, treatment options for bacterial infections become fewer. The photodynamic inactivation (PDI) of bacteria appears to be a potent, new technology that may support the treatment of colonized or infected skin. In photodynamic inactivation, a dye - called photosensitizer - absorbs light and generates reactive singlet oxygen. This singlet oxygen is then capable of killing bacteria independent of species or strain and their antibiotic resistance profile. In order to provide a practical application for the skin surface, the photosensitizer was included in an aqueous hydrogel (photodynamically active hydrogel). The efficacy of this gel was initially tested on an inanimate surface and then on the human skin ex vivo. NBTC staining and TUNEL assays were carried out on skin biopsies to investigate potential harmful effects of the surface PDI to the underlying skin cells. The photosensitizer in the gel sufficiently produced singlet oxygen while showing only little photobleaching. On inanimate surfaces as well as on the human skin, the number of viable bacteria was reduced by over or nearly up to 4 log10 steps, equal to 99.99% reduction or even more. Furthermore, histological staining showed no harmful effects of the gel towards the tissue. The application of this hydrogel represents a valuable method in decolonizing human skin including the potential to act against superficial skin infections. The presented results are promising and should lead to further investigation in a clinical study to check the effectivity of the photodynamically active hydrogel on patients.

Most of our knowledge on microbial physiology and biochemistry is based on studies performed under laboratory conditions. For growing hydrogen-oxidizing anaerobic, autotrophic prokaryotes, an H 2 :CO 2 (80:20, v/v) gas mixture is typically used. However, hydrogen concentrations in natural environments are usually low, but may vary in a wide range. Here we show that the thermophilic anaerobic bacterium Ammonifex degensii balances its autotrophic carbon fixation between two pathways depending on the H 2 partial pressure. At 80% H 2 , favoring ferredoxin reduction, it uses the ferredoxin-dependent Wood-Ljungdahl pathway. In contrast, during growth at 10% H 2 , it switches to the more ATP-demanding, ferredoxin-independent Calvin-Benson cycle. The study reveals that the H 2 redox potential is an important factor influencing the usage of different autotrophic pathways. This type of metabolic adjustment may be widespread in the microbial world.

The hydrogen gas-to-liquid mass transfer is the limiting factor in biological methanation. In trickle-bed reactors, mass transfer can be increased by high flow velocities in the liquid phase, by adding a packing material with high liquid hold-up or by using methanogenic archaea with a high methane productivity. This study developed a polyphasic approach to address all methods at once. Various methanogenic strains and packings were investigated from a microbial and hydrodynamic perspective. Analyzing the ability to produce high-quality methane and to form biofilms, pure cultures of Methanothermobacter performed better than those of the genus Methanothermococcus. Liquid and static hold-up of a packing material and its capability to facilitate attachment was not attributable to a single property. Consequently, it is recommended to carefully match organism and packing for optimized performance of trickle-bed reactors. The ideal combination for the ORBIT-system was identified as Methanothermobacter thermoautotrophicus IM5 and DuraTop®.

Employing deep reservoirs as UGS (underground gas storage) has a long history across continents. In 2018, 689 underground gas reservoirs with a total volume of 417 bcm were in operation worldwide. It is known that many microbial processes take place in the deep underground, even under the conditions of underground gas reservoirs. In this review, we focus mainly on methanogenesis and discuss related topics such as optimal environmental conditions, description of different types of UGS and microbial communities inhabiting these environments. We elucidate the potential of UGS as natural bioreactors for non-fossil methane production in the context of Power to Methane technology and the extension/expansion of the low-carbon economy. The role of carbon-neutral methane in the energy mix is likely to play a significant role in the coming decades. The safe production, transportation and storage of methane are well managed as well the existing infrastructure has been in place for a long time without problems. We also have experience in the long-term operation of underground gas storage systems. Methane technology thus appears to be a very promising approach and, together with the functioning UGS infrastructure, could be an important step towards the potential use of biomethanation in underground gas storage facilities as a way to reduce greenhouse gas emissions in the future.