Molecular characterization of Paenibacillus antarcticus IPAC21, a bioemulsifier producer isolated from Antarctic soil.

Paenibacillus antarcticus IPAC21, an endospore-forming and bioemulsifier-producing strain, was isolated from King George Island, Antarctica. As psychrotolerant/psychrophilic bacteria can be considered promising sources for novel products such as bioactive compounds and other industrially relevant substances/compounds, the IPAC21 genome was sequenced using Illumina Hi-seq, and a search for genes related to the production of bioemulsifiers and other metabolic pathways was performed. The IPAC21 strain has a genome of 5,505,124 bp and a G + C content of 40.5%. Genes related to the biosynthesis of exopolysaccharides, such as the gene that encodes the extracellular enzyme levansucrase responsible for the synthesis of levan, the 2,3-butanediol pathway, PTS sugar transporters, cold-shock proteins, and chaperones were found in its genome. IPAC21 cell-free supernatants obtained after cell growth in trypticase soy broth at different temperatures were evaluated for bioemulsifier production by the emulsification index (EI) using hexadecane, kerosene and diesel. EI values higher than 50% were obtained using the three oil derivatives when IPAC21 was grown at 28°C. The bioemulsifier produced by P. antarcticus IPAC21 was stable at different NaCl concentrations, low temperatures and pH values, suggesting its potential use in lower and moderate temperature processes in the petroleum industry.

Microbially induced corrosion impacts on the oil industry.

The numerous structural impacts on oil installations caused by corrosion make this issue a concern in the oil industry. Although chemical corrosion is relevant in this sector, it is indisputable that the microbial corrosion or bio-corrosion plays a preponderant role, with considerable economic losses. Microbial corrosion invariably depends on the formation of a biofilm on the attacked surface. Biofilm structures provide the conditions that favor the development of microbial groups related to corrosion. Despite the several microbial species are described as corrosive, certain groups, such as sulfate- and nitrate-reducing bacteria, acetogenic bacteria, and methanogenic archaea are the most commonly related. In spite of environmental factors influence the prevalence of certain species, it is increasingly accepted that the relationships between different species are determinant in corrosion. Such relationships can be evidenced by several surveys of microbial communities involved in bio-corrosion. Here, the main microbes related to corrosion in metallic structures used in oil installations are presented, as well as their metabolisms involved in the deterioration of metallic surfaces.

The adaptations of the microbial communities of the savanna soil over a period of wildfire, after the first rains, and during the rainy season.

Annually, the Cerrado ecosystem alternates between dry periods and long rainy seasons. During the dry season, severe forest fires occur, consuming a considerable part of the native vegetation, which impacts directly on the microbiome of the soil. Evaluate the adaptations of the soil microbiome to drought, rain and wildfire. Sequencing of the 16S rRNA gene was carried out for three significant conditions: drought and forest fires ("Fire"), after the first recorded rains ("First_Rain"), and during the rainy season ("Rainy"). It has been shown that under the "Fire" condition, there was a predominance of Phylum Actinobacteria, followed by Proteobacteria and Firmicutes. With the advent of the rainy season, "First_Rain," there was a change in the predominant taxonomic groups, with a higher prevalence of members of Proteobacteria and Firmicutes. During the rainy season, Proteobacteria and Firmicutes continued as the most prevalent groups. However, it was noted that in this period, there was an increase in bacterial diversity when compared with other periods analyzed. These results show how environmental factors influence adaptations in microbial communities. This allows for a better understanding of how to link the structure of the microbial community to the performance of ecosystems, and assist in preventing the consequences of increased frequency of wildfires, and long periods of drought.

The influence of the marine Bacillus cereus over carbon steel, stainless corrosion, and copper coupons.

The present study evaluated the influence of the marine bacteria Bacillus cereus Mc-1 on the corrosion of 1020 carbon steel, 316L stainless steel, and copper alloy. The Mc-1 strain was grown in a modified ammoniacal citrate culture medium (CFA.ico-), CFA.ico- with sodium nitrate supplementation (NO3-), and CFA.ico- with sodium chloride supplementation (NaCl). The mass loss and corrosion rate were evaluated after the periods of 7, 15, and 30 days. The results showed that in CFA.ico- and CFA.ico- medium added NO3- the corrosion rates of carbon steel and copper alloy were high when compared to the control. Whereas the medium was supplemented with NaCl, despite the rates being above the averages of the control system, they were considerably below the previous results. In general, the corrosion rates induced by Mc-1 on 316L coupons were below the results compared to carbon steel and copper alloy. When analyzing the corrosion rate measurements, regardless of the culture medium, the corrosion levels decreased consistently after 15 days, being below the levels evaluated after 7 days of the experiment. Our analyses suggest that B. cereus Mc-1 has different influences on corrosion in different metals and environmental conditions, such as the presence of NO3- and NaCl. These results can help to better understand the influence of this bacteria genus on the corrosion of metals in marine environments.

Influence of Salt Water Flow on Structures and Diversity of Biofilms Grown on 316L Stainless Steel.

Salt water, in addition to being a naturally corrosive environment, also includes factors such as temperature, pressure, and the presence of the microbial community in the environment that influence degradation processes on metal surfaces. The presence or absence of water flow over the metal surfaces is also an important aspect that influences the corrosion of metals. The objective of this study was to evaluate the presence or absence of salt water flow in the formation of biofilms grown in 316L stainless steel coupons. For this, the 316L stainless steel coupons were exposed in two different microcosms, the first being a system with continuous salt water flow, and the second without salt water flow system. The results of the sequencing of the 16S rDNA genes showed a clear difference in structures and diversity between the evaluated biofilms. There was greater abundance and diversity in the "In Flux" system when compared to the "No Flux" biofilm. The analysis of bacterial diversity showed a predominance of the Gammaproteobacteria class in both systems. However, at lower taxonomic levels, there were considerable differences in representativeness. Representatives of Vibrionales, Alteromonadales, Oceanospirillales, and Flavobacteriales were predominant in "No Flux", whereas in "In Flux" there was a greater representation of Alteromonadales, Rhodobacterales, and Saprospirales. These findings help to understand how the flow of water influences the dynamics of the formation of microbial biofilms on metal surfaces, which will contribute to the choice of strategies used to mitigate microbial biofouling.

The profile of the soil microbiota in the Cerrado is influenced by land use.

Extensive areas of the Cerrado biome have been deforested by the rapid advance of agricultural frontiers, especially by agricultural monocultures, and cultivated pastures. The objective of this study was to characterize the soil microbial community of an environment without anthropogenic interference and to compare it with soybean soil and pasture areas. For that, metagenomic sequencing techniques of the 16S rRNA gene were employed. Consistent changes in the profiles of diversity and abundance were described between communities in relation to the type of soil. The soil microbiome of the native environment was influenced by the pH level and content of Al3+, whereas the soil microbiomes cultivated with soybean and pasture were associated with the levels of nutrients N and P and the ions Ca2+ and Mg2+, respectively. The analysis of bacterial communities in the soil of the native environment showed a high abundance of members of the Proteobacteria phylum, with emphasis on the Bradyrhizobium and Burkholderia genera. In addition, significant levels of species of the Bacillus genus, and Dyella ginsengisoli, and Edaphobacter aggregans of the Acidobacteria phylum were detected. In the soil community with soybean cultivation, there was a predominance of Proteobacteria, mainly of the Sphingobium and Sphingomonas genera. In the pasture, the soil microbiota was dominated by the Firmicutes, which was almost entirely represented by the Bacillus genus. These results suggest an adaptation of the bacterial community to the soybean and pasture cultivations and will support understanding how environmental and anthropogenic factors shape the soil microbial community. KEY POINTS: • The Cerrado soil microbiota is sensitive to impacts on the biome. • Microbial communities have been altered at all taxonomic levels.

Influence of Acidification and Warming of Seawater on Biofouling by Bacteria Grown over API 5L Steel.

The acidification and warming of seawater have several impacts on marine organisms, including over microorganisms. The influence of acidification and warming of seawater on biofilms grown on API 5L steel surfaces was evaluated by sequencing the 16S ribosomal gene. For this, three microcosms were designed, the first simulating the natural marine environment (MCC), the second with a decrease in pH from 8.1 to 7.9, and an increase in temperature by 2 °C (MMS), and the third with pH in around 7.7 and an increase in temperature of 4 °C (MES). The results showed that MCC was dominated by the Gammaproteobacteria class, mainly members of the Alteromonadales Order. The second most abundant group was Alphaproteobacteria, with a predominance of Rhodobacterales and Oceanospirillales. In the MMS system there was a balance between representatives of the Gammaproteobacteria and Alphaproteobacteria classes. In MES there was an inversion in the representations of the most prevalent classes previously described in MCC. In this condition, there was a predominance of members of the Alphaproteobacteria Class, in contrast to the decrease in the abundance of Gammaproteobacteria members. These results suggest that possible future climate changes may influence the dynamics of the biofouling process in surface metals. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12088-021-00925-7.

The oil spill and the use of chemical surfactant reduce microbial corrosion on API 5L steel buried in saline soil.

In order to evaluate the biocorrosion of API 5L metal buried in saline soils, three different conditions in microcosms were evaluated. The control microcosm contained only saline soil, the second had the addition of petroleum, and the third contained the addition of both petroleum and surfactant. The corrosion rate of the metals was measured by loss of mass after 30 days, and the microbial communities were delineated using 16S rRNA gene sequencing techniques. The species were dominated by halophiles in all samples analyzed. Among the bacteria, the predominant group was Proteobacteria, with emphasis on the Alphaproteobacteria and Gammaproteobacteria. Betaproteobacteria and Deltaproteobacteria members were also identified in a smaller number in all conditions. Firmicutes were especially abundant in the control system, although it was persistently present in other conditions evaluated. Bacteroidetes and Actinobacteria were also present in a considerable number of OTUs in the three microcosms. Halobacteria were predominant among archaea and were present in all conditions. The analysis pointed to a conclusion that in the control microcosm, the corrosion rate was higher, while the microcosm containing only oil had the lowest corrosion rate. These results suggest that, under these conditions, the entry of other carbon sources favors the presence of petroleum degraders, rather than samples involved in the corrosion of metals.

Marine bacterial community analysis on 316L stainless steel coupons by Illumina MiSeq sequencing.

In order to evaluate the corrosive action of microorganisms on 316L metal exposed directly to a marine environment, a system was designed to immerse coupons in seawater. After periods of 30, 60 and 90 days, the coupons were recovered, the corrosion rates evaluated and the biofilm samples on their surface were analyzed by 16S rRNA gene sequencing. The results of the corrosion rate showed an acceleration over the entire experimental period. Alpha diversity measurements showed higher rates after 60 days of the experiment, while abundance measurements showed higher rates after 90 days of exposure to the marine environment. The beta-diversity results showed a clear separation between the three conditions and proximity in the indices between replicates of the same experimental condition. The results of 16S rRNA gene sequencing showed that after 30 days of exposure to seawater, there was massive representativeness of the pioneer bacteria, Gamma and Alphaproteobacteria, with emphasis on the genera Alcanivorax, Oceanospirillum and Shewanella. At the 60-day analysis, the Gammaproteobacteria class remained dominant, followed by Alphaproteobacteria and Flavobacteria, and the main representatives were Flexibacter and Pseudoalteromonas. In the last analysis, after 90 days, a change in the described bacterial community profile was observed. The Gammaproteobacteria class was still the largest in diversity and OTUs. The most predominant genera in number of OTUs were Alteromonas, Bacteriovorax and, Nautella. Our results describe a change in the microbial community over coupons directly exposed to the marine environment, suggesting a redirection to the formation of a mature biofilm. The conditions created by the biofilm structure suggest said condition favor biocorrosion on the analyzed coupons.

Changes in microbial community in the presence of oil and chemical dispersant and their effects on the corrosion of API 5L steel coupons in a marine-simulated microcosm.

The influence of crude oil and chemical dispersant was evaluated over planktonic bacteria and biofilms grown on API 5L steel surfaces in microcosm systems. Three conditions were simulated, an untreated marine environment and a marine environment with the presence of crude oil and a containing crude oil and chemical dispersant. The results of coupon corrosion rates indicated that in the oil microcosm, there was a high corrosion rate when compared with the other two systems. Analysis of bacterial communities by 16S rRNA gene sequencing described a clear difference between the different treatments. In plankton communities, the Bacilli and Gammaproteobacteria classes were the most present in numbers of operational taxonomic unit (OTUs). The Vibrionales, Oceanospirillales, and Alteromonadales orders were predominant in the treatment with crude oil, whereas in the microcosm containing oil and chemical dispersant, mainly members of Bacillales order were detected. In the communities analyzed from biofilms attached to the coupons, the most preponderant class was Alphaproteobacteria, followed by Gammaproteobacteria. In the control microcosm, there was a prevalence of the orders Rhodobacterales, Aeromonadales, and Alteromonadales, whereas in the dispersed oil and oil systems, the members of the order Rhodobacterales were present in a larger number of OTUs. These results demonstrate how the presence of a chemical dispersant and oil influence the corrosion rate and bacterial community structures present in the water column and biofilms grown on API 5L steel surfaces in a marine environment. KEY POINTS: • Evaluation of the effects of oil and chemical surfactants on the corrosion of API 5L. • Changes in microbial communities do not present corrosive biofilm on API 5L coupons.

Adaptative transcriptional response of Dietzia cinnamea P4 strain to sunlight simulator.

Responses to sunlight exposure of the oil-degrading Dietzia cinnamea P4 strain were evaluated by transcriptional levels of SOS genes, photoreactivation and genes involved in tolerance to high levels of reactive oxygen species. The P4 strain was exposed for 1 and 2 h and the magnitude of level changes in the mRNA was evaluated by qPCR. The results described the activation of the SOS system, with the decline of the repressor lexA gene levels and the concomitant increase of recA and uvrAD genes levels. The genes that participate in the photoreactivation process were also responsive to sunlight. The phrB gene encoding deoxyribodipyrimidine photo-lyase had its expression increased after 1-h exposure, while the phytAB genes showed a progressive increase over the studied period. The protective genes against reactive oxygen species, catalases, superoxides, peroxidases, and thioredoxins, had their expression rates detected under the conditions validated in this study. These results show a fast and coordinated response of genes from different DNA repair and tolerance mechanisms employed by strain P4, suggesting a complex concerted protective action against environmental stressors.

The era of 'omics' technologies in the study of microbiologically influenced corrosion.

Efforts to elucidate the relationships between microorganisms and metal corrosion were mainly directed to understanding the formation of biofilm structures grown on corroded surfaces. The emergence of high throughput DNA sequencing techniques has helped in the description of microbial species involved directly and indirectly in the corrosion processes of alloys. Coupled with sequencing from environmental samples, other methodologies such as metatranscriptome, metaproteomics and metabolomics have allowed a new horizon to be opened on the understanding of the role of corrosive microbial biofilm. Several groups of bacteria and archaea were identified, showing the dominance of Proteobacteria in several samples analyzed and members of groups that previously received less attention, such as Firmicutes and Bacteroidetes. Our research also shows that metagenomic studies describe the presence of various Archaea domain thermophilic and methanogenic groups associated with metal corrosion. Thus, opening the prospect of describing new microbial groups as possible participants in this current global concern.

Distinct Profiles in Microbial Diversity on Carbon Steel and Different Welds in Simulated Marine Microcosm.

The main studies on the corrosion of metals induced by microorganisms are directed only to the surface of the metal, without considering the presence of welds between these surfaces. For this reason, we evaluated the difference of microbial community grown in carbon steel coupons, and two different types of welds, E7018 and Tungsten electrodes, exposed under simulated microcosm. After 30 days, they were recovered, the biofilms scraped and the microbial communities analyzed by 16S rRNA gene sequencing. The results showed that there was a differentiated distribution among the three samples collected. Proteobacteria phylum composed most of the species described in all samples. At the class level, Gammaproteobacteria was the most detected, followed by Alphaproteobacteria and Flavobacteriia. The most prevalent order was Alteromonadales, which was present in Weld2, followed by Rhodobacteriales, which was more prevalent in Fe1020 and Weld1. The orders Cytophagales, Sphingomonadales, and Burkholderiales were described in higher number in Fe1020, whereas Oceanospirillales, Thiotrichales, Flavobacteriales, Rhodospirillales, and Kordiimonadales were higher in samples Weld1 and Weld2. The analyses between the three evaluated conditions show the presences of bacterial groups preferred by different types of metal, suggesting that approaches in the control of biocorrosion should take into account the chemical composition of the metal.

The role of biofilms in the corrosion of steel in marine environments.

Metal corrosion is a major global concern in many economic sectors. The degradation of metal surfaces is responsible for losses in values that account for about 3% of gross domestic product (GDP) only in the US. Parts of all corrosion processes described in different environments are present mainly in marine environments. The marine environment is characterized as favoring the corrosion processes of several metallic alloys, damaging structures used in the construction of ships, ports, oil pipelines, and others. Despite chemical corrosion being the most frequently described in these environments, studies show the participation of microorganisms in direct corrosion processes or in the acceleration/influence of the corrosive action, through the formation of complex biofilms. These structures create favorable conditions for microorganisms to degrade metal surfaces, causing damage known as pitting and crevices. Currently, diverse technicians are employed in biocorrosion research, e.g. electronic microscopy, and DNA sequencing. These techniques have clarified the dynamic process of the formation of biofilm structures, allowing understanding of the succession of different species during the evolution of the structure. Improving the understanding of how this interaction between biofilm and metallic surface occurs will enable better evaluation of strategies to avoid or decelerate the degradation of metallic structures in marine environments.

Oxidative damage induced by H2O2 reveals SOS adaptive transcriptional response of Dietzia cinnamea strain P4.

The oxidative stress response of the highly resistant actinomycete Dietzia cinnamea P4 after treatment with hydrogen peroxide (H2O2) was assessed in order to depict the possible mechanisms underlying its intrinsic high resistance to DNA damaging agents. We used transcriptional profiling to monitor the magnitude and kinetics of changes in the mRNA levels after exposure to different concentrations of H2O2 at 10 min and 1 h following the addition of the stressor. Catalase and superoxide dismutase genes were induced in different ways, according to the condition applied. Moreover, alkyl hydroperoxide reductase ahpCF, thiol peroxidase, thioredoxin and glutathione genes were upregulated in the presence of H2O2. Expression of peroxidase genes was not detected during the experiment. Overall results point to an actinomycete strain endowed with a set of enzymatic defenses against oxidative stress and with the main genes belonging to a functional SOS system (lexA, recA, uvrD), including suppression of lexA repressor, concomitantly to recA and uvrD gene upregulation upon H2O2 challenge.

The influence of surface microbial diversity and succession on microbiologically influenced corrosion of steel in a simulated marine environment.

To explore how a succession of bacteria grown on steel coupons in a marine environment can influence their corrosion process, we designed a microcosm in laboratory to evaluate corrosion kinetics and microbial diversity over 30 days. The results described a clear influence of corrosion by a succession of different bacterial groups. During the initial period, 2-7 days, a sharp increase in the rate of corrosion was detected accompanied by the presence of Alteromonadaceae, Vibrionaceae, Oceanospirillaceae, Rhodobacteraceae, Rhodospirillaceae and Flavobacteriaceae bacteria families. After 15 days, representatives of families Piscirickettsiaceae and Pseudomonadaceae were also described, accompanied by a continuous corrosion process over the coupons. After 30 days, there was a sudden change in the profile of the bacteria present on the steel coupons, with a prevalence of Halomonadaceae family species, and establishment and continuity of the corrosion process by the biofilm grown on the coupons. The results describe differences in microbial diversity over the time, highlighting certain bacterial lithotrophic species that persisted for most of the experiment, through a complex association between bacteria and metal surfaces, which can be a new starting point for development and maintenance of a favorable microenvironment to accelerate corrosion processes.

Transcriptional profiling of genes involved in n-hexadecane compounds assimilation in the hydrocarbon degrading Dietzia cinnamea P4 strain.

The petroleum-derived degrading Dietzia cinnamea strain P4 recently had its genome sequenced and annotated. This allowed employing the data on genes that are involved in the degradation of n-alkanes. To examine the physiological behavior of strain P4 in the presence of n-alkanes, the strain was grown under varying conditions of pH and temperature. D. cinnamea P4 was able to grow at pH 7.0-9.0 and at temperatures ranging from 35 ºC to 45 ºC. Experiments of gene expression by real-time quantitative RT-PCR throughout the complete growth cycle clearly indicated the induction of the regulatory gene alkU (TetR family) during early growth. During the logarithmic phase, a large increase in transcriptional levels of a lipid transporter gene was noted. Also, the expression of a gene that encodes the protein fused rubredoxin-alkane monooxygenase was enhanced. Both genes are probably under the influence of the AlkU regulator.

The putative α/ß-hydrolases of Dietzia cinnamea P4 strain as potential enzymes for biocatalytic applications.

The draft genome of the soil actinomycete Dietzia cinnamea P4 reveals a versatile group of α/ß-hydrolase fold enzymes. Phylogenetic and comparative sequence analyses were used to classify the α/ß-hydrolases of strain P4 into six different groups: (i) lipases, (ii) esterases, (iii) epoxide hydrolases, (iv) haloacid dehalogenases, (v) C-C breaking enzymes and (vi) serine peptidases. The high number of lipases/esterases (41) and epoxide hydrolase enzymes (14) present in the relatively small (3.6 Mb) P4 genome is unusual; it is likely to be linked to the survival of strain P4 in its natural environment. Strain P4 is thus equipped with a large number of genes which would appear to confer survivability in harsh hot tropical soil. As such, this highly resilient soil bacterial strain provides an interesting genome for enzyme mining for applications in the field of biotransformations of polymeric compounds.

Transcriptional profiling of genes involved in n-hexadecane compounds assimilation in the hydrocarbon degrading Dietzia cinnamea P4 strain

The petroleum-derived degrading Dietzia cinnamea strain P4 recently had its genome sequenced and annotated. This allowed employing the data on genes that are involved in the degradation of n-alkanes. To examine the physiological behavior of strain P4 in the presence of n-alkanes, the strain was grown under varying conditions of pH and temperature. D. cinnamea P4 was able to grow at pH 7.0-9.0 and at temperatures ranging from 35 ºC to 45 ºC. Experiments of gene expression by real-time quantitative RT-PCR throughout the complete growth cycle clearly indicated the induction of the regulatory gene alkU (TetR family) during early growth. During the logarithmic phase, a large increase in transcriptional levels of a lipid transporter gene was noted. Also, the expression of a gene that encodes the protein fused rubredoxin-alkane monooxygenase was enhanced. Both genes are probably under the influence of the AlkU regulator.

Insight from the draft genome of Dietzia cinnamea P4 reveals mechanisms of survival in complex tropical soil habitats and biotechnology potential.

The draft genome of Dietzia cinnamea strain P4 was determined using pyrosequencing. In total, 428 supercontigs were obtained and analyzed. We here describe and interpret the main features of the draft genome. The genome contained a total of 3,555,295 bp, arranged in a single replicon with an average G+C percentage of 70.9%. It revealed the presence of complete pathways for basically all central metabolic routes. Also present were complete sets of genes for the glyoxalate and reductive carboxylate cycles. Autotrophic growth was suggested to occur by the presence of genes for aerobic CO oxidation, formate/formaldehyde oxidation, the reverse tricarboxylic acid cycle and the 3-hydropropionate cycle for CO(2) fixation. Secondary metabolism was evidenced by the presence of genes for the biosynthesis of terpene compounds, frenolicin, nanaomycin and avilamycin A antibiotics. Furthermore, a probable role in azinomycin B synthesis, an important product with antitumor activity, was indicated. The complete alk operon for the degradation of n-alkanes was found to be present, as were clusters of genes for biphenyl ring dihydroxylation. This study brings new insights in the genetics and physiology of D. cinnamea P4, which is useful in biotechnology and bioremediation.