Applied and Environmental Microbiology RG Impact & Description | ResearchGate | Impact Rankings (
and 2017 )
Applied and Environmental Microbiology publishes a substantial share of the most significant current research in the areas of biotechnology, microbial ecology, food microbiology, and industrial microbiology. Highlighting research findings applicable to the development of new processes or products, AEM is a broad-based journal that is indispensable to those whose basic work has implications of near-term practical benefit.RG Journal Impact: 3.12 **This value is calculated using ResearchGate data and is based on average citation counts from work published in this journal. The data used in the calculation may not be exhaustive.2017 RG Journal impactAvailable summer 20182015 / 2016 RG Journal impact3.122014 RG Journal impact4.602013 RG Journal impact2.972012 RG Journal impact2.792011 RG Journal impact4.272010 RG Journal impact4.062009 RG Journal impact5.342008 RG Journal impact4.952007 RG Journal impact4.672006 RG Journal impact4.272005 RG Journal impact4.402004 RG Journal impact4.522003 RG Journal impact4.532002 RG Journal impact4.152001 RG Journal impact3.842000 RG Journal impact3.21RG Journal impact over timeRG Journal impactRG Journal impact over timeGraph showing a linear path with a yearly representation of impact points of the journalPublisher detailsThis journal may support self-archiving.Publications in this journalNov 2017Applied and Environmental Microbiology[...]Nov 2017Applied and Environmental Microbiology[...]Staphylococcus aureus is part of the nasal microbiome of many humans and has become a significant public health burden due to infections with antibiotic resistant strains, including methicillin resistant S. aureus (MRSA). Several lineages of S. aureus including MRSA are found in livestock species and can be acquired by humans through contact with animals. These livestock associated MRSA (LA-MRSA) isolates raise public health concerns because of the potential for livestock to act as reservoirs for MRSA outside of the hospital setting. In the United States, swine harbor a mixed population of LA-MRSA isolates with the sequence type (ST) 398, ST9, and ST5 lineages being detected. LA-MRSA ST5 isolates are particularly concerning to the public health community because, unlike ST398 and ST9 lineages, the ST5 lineage is a significant cause of human disease in both the hospital and community setting globally. The ability of swine associated LA-MRSA ST5 isolates to adhere to human keratinocytes in vitro was investigated and adherence genes harbored by these isolates were evaluated and compared to clinical MRSA ST5 isolates from humans with no swine contact. The two subsets of isolates adhered equivalently to human keratinocytes in vitro and contained an indistinguishable complement of adherence genes that possessed a high degree of sequence identity. Collectively our data indicate that, unlike LA-MRSA ST398 isolates, LA-MRSA ST5 isolates did not exhibit a reduced genotypic or phenotypic capacity to adhere to human keratinocytes.
IMPORTANCE
Our data indicate swine associated livestock-associated methicillin resistant Staphylococcus aureus (LA-MRSA) ST5 isolates are equally capable of adhering to human skin and have the same genetic potential to adhere as clinical MRSA ST5 isolates from humans. This suggests humans in contact with livestock have the potential to become colonized with LA-MRSA ST5 however, genes that contribute to persistence of S. aureus on human skin were absent in LA-MRSA ST5 isolates. The data presented here is important evidence in evaluating potential risks LA-MRSA ST5 isolates pose to humans with livestock contact.Expand abstractNov 2017Applied and Environmental Microbiology[...]Virulence factors of mammary pathogenic Escherichia coli (MPEC) have not been identified, and it is not known how bacterial gene content influences severity of mastitis. Here, we report the first genome-wide identification of genes that contribute to fitness of MPEC in conditions relevant to the natural history of the disease. A highly virulent clinical isolate (M12) was identified that killed Galleria mellonella at low infectious doses and which replicated to high numbers in mouse mammary glands and spread to spleens. Genome sequencing was combined with transposon insertion site sequencing to identify MPEC genes that contribute to growth in unpasteurized whole milk, as well as during G. mellonella and mouse mastitis infections. These analyses show that strain M12 possesses a unique genomic island encoding a group III polysaccharide capsule that greatly enhances virulence in G. mellonella. Several genes appear critical for MPEC survival in both G. mellonella and in mice, including nutrient scavenging systems and resistance to cellular stress. Insertions in the ferric dicitrate receptor gene fecA caused significant fitness defects in all conditions (milk, G. mellonella and in mice). This gene was highly expressed during growth in milk. Targeted deletion of fecA from strain M12 caused attenuation in G. mellonella larvae, and reduced growth in unpasteurized cow's milk and lactating mouse mammary glands. Our results confirm that iron scavenging by the ferric dicitrate receptor, which is strongly associated with MPEC strains, is required for MPEC growth and may influence disease severity in mastitis infections.
Importance
Mastitis caused by E. coli inflicts substantial burdens on the health and productivity of dairy animals. Strains causing mastitis may express genes that distinguish them from other E. coli and promote infection of mammary glands, but these have not been identified. Using a highly virulent strain, we employed genome-wide mutagenesis and sequencing to discover genes that contribute to mastitis. This extensive data set is the first screen for mastitis-associated E. coli fitness factors and provides the following contributions to the field: 1) Global comparison of genes required for different aspects of mastitis infection, 2) Discovery of a unique capsule that contributes to virulence, 3) Conclusive evidence for the crucial role of iron scavenging systems in mastitis, particularly the ferric dicitrate transport system. Similar approaches applied to other mastitis-associated strains will uncover conserved targets for prevention or treatment, and provide a better understanding of their relationship to other E. coli pathogens.Expand abstractNov 2017Applied and Environmental Microbiology[...]Nitrate-reducing iron(II)-oxidizing bacteria have been known for approximately 20 years. There has been much debate to which extent the reduction of nitrate and the oxidation of ferrous iron are coupled via enzymatic pathways or via abiotic processes induced by nitrite formed by heterotrophic denitrification. The aim of the present study was to assess the coupling of nitrate reduction and iron(II) oxidation by monitoring changes in substrate concentrations as well as in the activity of nitrate-reducing bacteria in natural littoral freshwater sediment as a response to a stimulation with nitrate and iron(II). In substrate-amended microcosms, we found that the biotic oxidation of ferrous iron depended on the simultaneous microbial reduction of nitrate. Additionally, the abiotic oxidation of ferrous iron by nitrite in sterilized sediment was not fast enough to explain the iron oxidation rates observed in microbially active sediment. Furthermore, the expression levels of genes coding for enzymes crucial for nitrate reduction were in some setups stimulated by the presence of ferrous iron. These results indicate that there is a direct influence of ferrous iron on bacterial denitrification and support the hypothesis that microbial nitrate reduction is stimulated by biotic iron(II) oxidation.
Importance
The coupling of nitrate reduction and Fe(II) oxidation affects the environment at a local scale, e.g. by changing nutrient or heavy metal mobility in soils due to the formation of Fe(III) minerals, as well as at a global scale, e.g. by the formation of the primary greenhouse gas nitrous oxide. Although the coupling of nitrate reduction and Fe(II) oxidation was reported twenty years ago and has been studied intensively since then, the underlying mechanisms still remain unknown. One of the main knowledge gaps is the extent of enzymatic Fe(II) oxidation coupled to nitrate reduction which has frequently been questioned in the literature. In the present study we provide evidence for microbially mediated nitrate-reducing Fe(II) oxidation in freshwater sediments. This evidence is based on the rates of nitrate reduction and Fe(II) oxidation determined in microcosm incubations and the effect of iron on the expression of genes required for denitrification.Expand abstractNov 2017Applied and Environmental Microbiology[...]The haloalkane dehalogenase enzyme DmmA was identified by marine metagenomic screening. Determination of its crystal structure revealed an unusually large active site compared to previously characterized haloalkane dehalogenases. Here we present a biochemical characterization of this interesting enzyme with emphasis on its structure-function relationships. DmmA exhibited an exceptionally broad substrate specificity and degraded several halogenated environmental pollutants that are resistant to other members of this enzyme family. In addition to this unique substrate specificity, the enzyme was highly tolerant to organic co-solvents such as dimethyl sulfoxide, methanol, and acetone. Its broad substrate specificity, high over-expression yield (200 mg of protein per litre o 50% of total protein), good tolerance to organic co-solvents and a broad range of pH make DmmA an attractive biocatalyst for various biotechnological applications.
IMPORTANCE We present a thorough biochemical characterization of the haloalkane dehalogenase DmmA from a marine metagenome. The enzyme with unusually large active site shows remarkably broad substrate specificity, high overexpression, significant tolerance to organic co-solvents and activity at a broad range of pH conditions. DmmA is an attractive catalyst for sustainable biotechnology applications, e.g., biocatalysis, biosensing and biodegradation of halogenated pollutants. We also report its ability to convert multiple-halogenated compounds to corresponding polyalcohols.Expand abstractNov 2017Applied and Environmental Microbiology[...]The aim of the current work was to identify key features of the fungal proteome involved in active decay of beech wood blocks, by the white rot fungus Bjerkandera adusta at 20°C and 24°C. A combination of protein and domain analyses ensured a high level of annotation, which revealed that while variation in the proteins identified was high between replicates, there was a considerable degree of functional conservation between the two temperatures. Further analysis revealed differences in the pathways and processes employed by the fungus at the different temperatures, particularly in relation to nutrient acquisition and xenobiotic mitigation. Key features showing temperature-dependent variation in mechanisms for both lignocellulose decomposition and sugar utilisation were found, alongside differences in the enzymes involved in mitigation against damage caused by toxic phenolic compounds and oxidative stress.
Importance
This work was conducted using the wood decay fungus B. adusta , grown on solid wood blocks to closely mimic the natural environment, and gives greater insight into the proteome of an important environmental fungus during active decay. We show that a change in incubation temperature from 20°C to 24°C altered protein profile. Proteomic studies in the field of white-rotting basidiomycetes have thus far been hampered by poor annotation of protein databases, with a large proportion of proteins simply ‘unknown function'. This study was enhanced by extensive protein domain analysis, enabling a higher level of functional assignment and greater understanding of the proteome composition. This work revealed a strong inter-dependence between the primary process of nutrient acquisition and specialised metabolic processes for detoxification of plant extractives and the phenolic breakdown products of lignocellulose.Expand abstractNov 2017Applied and Environmental Microbiology[...]Lysobacter enzymogenes is a Gram-negative, environmental ubiquitous bacterium that produces a secondary metabolite, called HSAF (Heat-stable antifungal factor), as an antifungal factor against plant and animal fungal pathogens. 4-hydroxybenzoic acid (4-HBA) is a newly identified diffusible factor that regulates HSAF synthesis via LysRLe, a LysR-type transcription factor (TF). Here, to identify additional TFs within the 4-HBA regulatory pathway that control HSAF production, we re-analyzed the LenB2-based transcriptomic data, in which LenB2 is the enzyme responsible for 4-HBA production. This survey led to identification of three TFs (Le4806, Le4969, and Le3904). Of them, LarR (Le4806), a member of the MarR-family proteins, was identified as a new TF that participated in the 4-HBA-dependent regulation of HSAF production. Our data show that: (i) LarR is a downstream component of the 4-HBA regulatory pathway controlling the HSAF level, while LysRLe is the receptor of 4-HBA; (ii) 4-HBA and LysRLe play an opposite regulatory effect on larR transcription, where larR transcript was negatively modulated by 4-HBA, while LysRLe, in contrast, exerted a positive transcriptional regulation by directly binding to the larR promoter without being affected by 4-HBA (iii) LarR, similar to LysRLe, could bind to promoter of the HSAF biosynthetic gene operon, leading to a positive regulation of HSAF (iv) LarR and LysRLe could not interact, and instead control HSAF biosynthesis independently. These results outline a previously uncharacterized mechanism by which the antibiotic HSAF biosynthesis in L. enzymogenes is modulated by the interplay of 4-HBA, a diffusible molecule, and two different TFs.
IMPORTANCE
Bacteria use diverse chemical signaling molecules to regulate a wide range of physiological and cellular processes. 4-HBA is an “old” chemical molecule that was produced by diverse bacterial species, but its regulatory function and working mechanism remain largely unknown. We previously found that 4-HBA in L. enzymogenes could serve as a diffusible factor regulating HSAF synthesis via LysRLe. Here, we further identified LarR, a MarR-family protein as a second TF that participates in the 4-HBA-dependent regulation of HSAF biosynthesis. Our results dissected how LarR acts as a protein linker to connect 4-HBA and HSAF synthesis, where LarR also has a cross-talk with LysRLe. Thus, our findings not only provide fundamental insight regarding how a diffusible molecule (4-HBA) adopts two different types of TFs for coordinating HSAF biosynthesis, but also present applied microbiology knowledge for increasing the antibiotic HSAF yield by modification of the 4-HBA regulatory pathway in L. enzymogenes.Expand abstractNov 2017Applied and Environmental Microbiology[...]Sucrose and glycogen syntheses in cyanobacteria share the common precursor glucose-1-phosphate. It is generally assumed that lowering glycogen synthesis could drive more carbon towards sucrose synthesis that can be induced by salt stress among cyanobacteria. By using a theophyllin-dependent riboswitch system, the expression of glgC , a key gene in glycogen synthesis, was down-regulated in a quantitative manner in a sucrose-secreting strain of Synechococcus elongatus PCC 7942. We observed that the stepwise suppression of glycogen synthesis limited rather than stimulated sucrose production in the salt stressed cells, suggesting that glycogen could serve as a carbon pool for the synthesis of sucrose. Accordingly, we generated glycogen-overproducing strains, but the increased glycogen pool alone did not stimulate sucrose production, indicating that alternative steps limit the carbon flux towards the synthesis of sucrose. Consistent with previous studies that showed that sucrose-phosphate synthase (SPS) catalyzes the rate-limiting step in sucrose synthesis, the combination of glycogen overproduction and sps overexpression resulted in increased sucrose production. Our results indicate that the glycogen and sucrose pools are closely linked in Synechococcus elongatus PCC 7942, and we propose that enhancing glycogen pool could be a promising strategy for the improvement of sucrose production by cyanobacteria in the presence of a strong sucrose synthesis sink.
IMPORTANCE
Many cyanobacteria naturally synthesize and accumulate sucrose when stressed by NaCl, which provides novel possibilities for obtaining sugar feedstock by engineering of cyanobacteria. It has been assumed that glycogen synthesis competes with sucrose synthesis for the carbon flux. However, our results showed that the suppression of glycogen synthesis decreased rather than stimulated sucrose production in the sucrose-secreting strain of Synechococcus elongatus PCC 7942. This result suggests that glycogen could serve as a supportive rather than a competitive carbon pool for the synthesis of sucrose, providing new insights about the relation between glycogen synthesis and sucrose synthesis in cyanobacteria. This finding is also useful to guide the metabolic engineering work to optimize the production of sucrose and possibly other products by cyanobacteria.Expand abstractNov 2017Applied and Environmental MicrobiologyRenée M. Petri[...]Chickens of good and poor feed efficiency (FE) have been shown to differ in their intestinal microbiota composition. This study investigated differences in the fecal bacterial community of good and poor feed efficient chickens at 16 and 29 days post-hatch (dph) and evaluated whether a fecal microbiota transplant (FMT) from feed efficient donors early in life can affect the fecal microbiota in chickens at 16 and 29 dph, chicken's FE and nutrient retention at four weeks of age. One-hundred-ten chickens were inoculated with a FMT or a control transplant (CT) on 1, 6 and 9 dph and ranked according to residual feed intake (RFI; metric for FE) on 30 dph. Fifty-six chickens across both inoculation groups were selected as the extremes in RFI (29 low, 27 high). RFI-related fecal bacterial profiles were discernible at 16 and 29 dph. Particularly, Lactobacillus salivarius -, Lactobacillus crispatus - and Anaerobacterium -operational taxonomic units were associated with low RFI (good FE). Multiple administration of the FMT only slightly changed the fecal bacterial composition, which was supported by weighted UniFrac analysis, showing similar bacterial communities in feces of both inoculation groups at 16 and 29 dph. Moreover, the FMT did not change RFI and nutrient retention of good and poor feed efficient recipients, whereas it tended to increase feed intake and body weight gain in female chickens. This may suggest that host- and environment-related factors may more strongly affect chicken's fecal microbiota and FE than the FMT.
IMPORTANCE
Modulating the chicken's early microbial colonization using a FMT from highly feed efficient donor chickens may be a promising tool to establish a more desirable bacterial profile in recipient chickens, thereby improving host FE. Although FE-associated fecal bacterial profiles at 16 and 29 dph could be established, the microbiota composition of a FMT, when administrated early in life, may not be a strong factor modulating the fecal microbiota at two to four weeks of life and reducing the variation in chicken's FE. Nevertheless, the present FMT may have potential benefits on growth performance in female chickens.Expand abstractNov 2017Applied and Environmental Microbiology[...]Ectoine and hydroxyectoine are widely synthesized by members of the Bacteria and a few Archaea as potent osmostress protectants. We have studied the salient features of the osmostress-responsive promoter directing the transcription of the ectoine/hydroxyectoine biosynthetic gene cluster from the plant-roots-associated bacterium Pseudomonas stutzeri by transferring it into Escherichia coli , an enterobacterium that does not produce ectoines naturally. Using ect-lacZ reporter fusions, we found that the heterologous ect promoter reacted with exquisite sensitivity in its transcriptional profile to graded increases in sustained high salinity, responded to a true osmotic signal, and required the build-up of an osmotically effective gradient across the cytoplasmic membrane for its induction. The involvement of the -10, -35, and spacer regions of the Sigma-70 type ect promoter in setting promoter strength and response to osmotic stress was assessed through site-directed mutagenesis. Moderate changes in the ect promoter sequence that increase its resemblance to house-keeping Sigma-70-type promoters of E. coli afforded substantially enhanced expression, both in absence and in the presence of osmotic stress. Building on this set of ect promoter mutants, we engineered an E. coli chassis strain for the heterologous production of ectoines. This synthetic cell factory lacks the genes for the osmostress-responsive synthesis of trehalose and the compatible solute importers ProP and ProU and it continuously excretes ectoines into the growth medium. By combining appropriate host strains and different plasmid variants, excretion of ectoine, hydroxyectoine, or a mixture of both compounds was achieved under mild osmotic stress conditions.
IMPORTANCE
Ectoines are compatible solutes, organics osmolytes that are used by microorganisms to fend off the negative consequences of high environmental osmolarity on cellular physiology. An understanding of the salient features of osmostress-responsive promoters directing the expression of the ectoine/hydroxyectoine biosynthetic gene clusters is lacking. We exploited the ect promoter from an ectoine/hydroxyectoine-producing soil bacterium for such a study by transferring it into a surrogate bacterial host. Despite the fact that E. coli does not synthesize ectoines naturally, the ect promoter retained its exquisitely sensitive osmotic control, indicating that osmoregulation of ect transcription is an inherent feature of the promoter and its flanking sequences. These sequences were narrowed to a 116 bp DNA fragment. Ectoines have interesting commercial applications. Building on data from a site-directed mutagenesis study of the ect promoter, we designed a synthetic cell factory that secretes ectoine, hydroxyectoine, or a mixture of both compounds, into the growth medium.Expand abstractNov 2017Applied and Environmental MicrobiologyHigh pressure processing is a non-thermal method of food preservation that uses pressure to inactivate microorganisms. To ensure the effective validation of process parameters, it is important that the design of challenge protocols consider the potential for resistance in a particular species. Herein, the response of 99 diverse Salmonella enterica strains to high pressure is reported. Members of this population belong to one of 24 serovars and were isolated from various Canadian sources over a period of 26 years. When exposed to 600 MPa for 3 min, the average reduction in cell numbers for this population was 5.6 log 10 CFU/mL with a range of 0.9 log 10 CFU/mL to 6 log 10 CFU/mL. Eleven strains from 5 serovars with variable levels of pressure resistance were selected for further studies. The membrane characteristics (propidium iodide uptake during and after pressure treatment, sensitivity to membrane active agents, and membrane fatty acid composition) and the response of this panel to stressors (heat, nutrient deprivation, desiccation, acid) suggested potential roles for the cell membrane and RpoS regulon in mediating pressure resistance in S. enterica . The data indicate a heterogeneous and multifactorial response to high pressure that cannot be predicted for individual S. enterica strains.
IMPORTANCE The response of foodborne pathogens to increasingly popular minimal food decontamination methods are not understood and therefore difficult to predict. This report shows the response of Salmonella enterica to high pressure processing is diverse. The magnitude of inactivation does not depend on how closely related the strains are and where they were isolated from. Moreover, strains that are resistant to high pressure do not behave similarly to other stresses suggesting that more than one mechanism might be responsible for resistance to high pressure and the mechanisms used may vary from one strain to another.Expand abstractNov 2017Applied and Environmental Microbiology[...]Gut microbiota associations through habitat transitions are fundamentally important, yet poorly understood. One such habitat transition is the migration from fresh to salt water for anadromous fish such as salmon. The aim of the current work was therefore to determine the fresh- to saltwater impact on the gut microbiota in farmed Atlantic salmon, with dietary interventions resembling that of fresh- and salt water diets with respect to fatty acid composition. Using deep 16S rRNA gene sequencing, and quantitative PCR, we found that the fresh- to salt water transition both had a major association with the microbiota composition and quantity, while diet did not show significantly associations with the microbiota. In salt water there was a 100-fold increase in bacterial quantity, with a relative increase of Firmicutes and a relative decrease of both Actinobacteria and Proteobacteria. Irrespective of an overall shift in microbiota composition from fresh to salt water we identified three core clostridia and one Lactobacillus -affiliated phylotype with wide geographic distribution that were highly prevalent and co-occurring. Taken together, our results support the importance of the dominating bacteria in the salmon gut, with the fresh water microbiota being immature. Due to the low number of potentially host associated bacterial species in the salmon gut, we believe farmed salmon can represent an important model for future understanding of host-bacterial interactions in aquatic environments.
IMPORTANCE
Little is known about factors affecting the inter-individual distribution of gut bacteria in aquatic environments. We have shown that there is a core of four highly prevalent and co-occurring bacteria irrespective of feed and fresh- to saltwater transition. The potential host interactions of the core bacteria, however, need to be elucidated further.Expand abstractNov 2017Applied and Environmental Microbiology[...]Mycotoxin contamination of cereal grains causes well-recognized toxicities in animals and humans, but the fate of plant-bound masked mycotoxins in the gut is less well understood. Masked mycotoxins have been found to be stable under conditions prevailing in the small intestine, but are rapidly hydrolyzed by fecal microbiota. This study aims to assess the hydrolysis of the masked mycotoxin deoxynivalenol-3-glucoside (DON3Glc) by microbiota of different regions of the porcine intestinal tract.
Intestinal digesta samples were collected from the jejunum, ileum, caecum, colon and feces of 5 pigs and immediately frozen under anaerobic conditions. Sample slurries were prepared in M2 culture medium, spiked with DON3Glc or free DON (2 nmoles/mL) and incubated anaerobically for up to 72 hours. Mycotoxin concentrations were determined using LC-MS/MS and microbiota composition was determined using qPCR methodology.
Jejunal microbiota hydrolyzed DON3Glc very slowly, while samples from the ileum, caecum, colon and feces rapidly and efficiently hydrolyzed DON3Glc. No further metabolism of DON was observed in any sample. Microbial load and microbiota composition was significantly different in the ileum, but similar in caecum, colon and feces.
Importance
Results from this study clearly demonstrate that the masked mycotoxin DON3Glc is hydrolyzed efficiently in the distal small intestine and large intestine of pigs. Once DON is released, toxicity and absorption in the distal intestinal tract are likely to occur in vivo . This study further supports the need to include masked metabolites into mycotoxin risk assessments and regulatory actions for feed and food.Expand abstractNov 2017Applied and Environmental Microbiology[...]The dynamics of individual microbial populations and their gene functions in agricultural soils, especially after major activities such as nitrogen (N) fertilization, remain elusive but are important for better understanding nutrient cycling. Here, we analyzed 20 short-read metagenomes collected at four time points across one year from two depths (0-5 and 20-30 cm) in two Midwestern agricultural sites representing contrasting soil textures (sandy versus silty-loam), with similar cropping histories. Although microbial community taxonomic and functional compositions differed between the two locations and depths, they were more stable within a depth/site throughout the year than communities in natural water-based ecosystems. For example, among the 69 population genomes assembled from the metagenomes, 75% showed less than 2-fold change in abundance between any two sampling points. Interestingly, six deep-branching Thaumarchaeota and three complete ammonia oxidizer (comammox) Nitrospira populations increased up to 5-fold in abundance upon the addition of N fertilizer. These results indicated that indigenous archaeal ammonia oxidizers may respond faster (more copiotrophic) to N fertilization than previously thought. None of 29 recovered putative denitrifier genomes encoded the complete denitrification pathway, suggesting that denitrification is carried out by a collection of different populations. Altogether, our study identified novel microbial populations and genes responding to seasonal and human-induced perturbations in agricultural soils that should facilitate future monitoring efforts and N-related studies.
Importance
Even though the impact of agricultural management on the microbial community structure has already been recognized, understanding of the dynamics of individual microbial populations and what functions each population encodes are limited. Yet, this information is important for better understanding nutrient cycling, with potentially important implications for preserving nitrogen in soils and sustainability. Here we show that reconstructed metagenome-assembled genomes (MAGs) are relatively stable in their abundance and functional gene content year-round, and seasonal nitrogen fertilization has selected for novel Thaumarchaeota and comammox Nitrospira nitrifiers that are potentially less oligotrophic compared to their marine counterparts previously studied.Expand abstractNov 2017Applied and Environmental MicrobiologyMembers of the genus Caldicellulosiruptor have the ability to deconstruct and grow on lignocellulosic biomass without conventional pretreatment and a genetically tractable species, C. bescii , was recently engineered to produce ethanol directly from switchgrass. C. bescii contains more than 50 glycosyl-hydrolases and a suite of extracellular enzymes for biomass deconstruction, most prominently CelA, a multi-domain cellulase that uses a novel mechanism to deconstruct plant biomass. Accumulation of cellobiose, a product of CelA during growth on biomass, inhibits cellulase activity. Here we show that heterologous expression of a cellobiose phosphorylase from Thermotoga maritima improves the phosphorolytic pathway in C. bescii and results in synergistic activity with endogenous enzymes, including CelA, to increase cellulolytic activity and growth on crystalline cellulose.
Importance
CelA is the only known cellulase to function well on highly crystalline cellulose and uses a mechanism distinct from other cellulases, including fungal cellulases. Also unlike fungal cellulases, it functions at high temperature out preforms commercial cellulase cocktails. Factors that inhibit CelA during biomass deconstruction are significantly different for those that impact the performance of fungal cellulases and commercial mixtures. This work contributes to the understanding of cellulase inhibition, enzyme function and will suggest a rational approach to engineering optimal activity.Expand abstractOct 2017Applied and Environmental Microbiology[...]Polyvinyl alcohol (PVA) is used widely in industry, and associated environmental pollution is a serious problem. Herein, we report a novel, efficient PVA degrader, Stenotrophomonas rhizophila QL-P4, isolated from fallen leaves from virgin forest in the Qinling Mountains. The complete genome was obtained using single-molecule real-time (SMRT) technology and corrected using Illumina sequencing. Bioinformatics analysis revealed eight PVA/OVA (vinyl alcohol oligomer)-degrading genes. Of these, seven genes were predicted to be involved in the classical intracellular PVA/OVA degradation pathway, and one (BAY15_3292) was identified as a novel PVA oxidase. Five PVA/OVA-degrading enzymes were purified and characterised. Among which, BAY15_1712, a PVA dehydrogenase (PVADH), displayed high catalytic efficiency towards PVA and OVA substrate. All reported PVADHs only have PVA-degrading ability. Most importantly, we discovered a novel PVA oxidase (BAY15_3292) that exhibited highest PVA-degrading efficiency than the reported PVADHs. Further investigation indicated that BAY15_3292 plays a crucial role in PVA degradation in S. rhizophila QL-P4. Knocking out BAY15_3292 resulted in a significant decline in PVA-degrading activity in S. rhizophila QL-P4. Interestingly, we found that BAY15_3292 possesses exocrine activity, which distinguishes it from classical PVADHs. Transparent circle experiments further proved that BAY15_3292 greatly affects extracellular PVA degradation in S. rhizophila QL-P4. The exocrine characteristics of BAY15_3292 facilitate its potential application to PVA bioremediation. In addition, we report three new efficient secondary alcohol dehydrogenases (SADHs) with OVA-degrading ability in S. rhizophila QL-P4, compared with only one OVA-degrading SADH as reported previously.
Importance
With the widespread application of PVA in industry, PVA-related environmental pollution is an increasingly serious issue. Because PVA is difficult to degrade, it accumulates in aquatic environments and causes chronic toxicity to aquatic organisms. Biodegradation of PVA, as an economical and environment-friendly method, has attracted much interest. To date, effective and applicable PVA-degrading bacteria/enzymes have not been reported. Herein, we report a new efficient PVA degrader (S. rhizophila QL-P4) that has five PVA/OVA-degrading enzymes with high catalytic efficiency, among which BAY15_1712 is the only reported PVADH with both PVA- and OVA-degrading abilities. Importantly, we discovered a novel PVA oxidase (BAY15_3292) that is not only more efficient than other reported PVA-degrading PVADHs, but also has exocrine activity. Overall, our findings provide new insight into PVA-degrading pathways in microorganisms, and suggest S. rhizophila QL-P4 and its enzymes have potential for application to PVA bioremediation to reduce or eliminate PVA-related environmental pollution.Expand abstractOct 2017Applied and Environmental Microbiology[...]Although biocatalytic transformation has shown great promise in chemical synthesis, there remain significant challenges in controlling high selectivity without the formation of undesirable byproducts. For instance, few attempts have been successful in constructing biocatalyst for de novo synthesis of pure flavin mononucleotide (FMN) due to riboflavin (RF) accumulating inside the cytoplasm and being secreted with FMN. To address this problem, we show here a novel biosynthesis strategy, compartmentalizing the final FMN biosynthesis step in the periplasm of an engineered Escherichia coli strain. This construct is able to over-produce FMN with high specificity (92.4% of total excreted flavins). Such biosynthesis approach allows isolation of the final biosynthesis step from the cytoplasm so as to eliminate undesirable by-products, providing a new route to develop biocatalysts for high-purity chemicals.
IMPORTANCE: The periplasm of Gram-negative bacterial hosts is engineered to compartmentalize the final biosynthesis step from the cytoplasm. This strategy is promising for the overproduction of high-value product with high specificity. We demonstrate the successful implementation of this strategy in microbial production of highly pure flavin mononucleotide (FMN).Expand abstractOct 2017Applied and Environmental Microbiology[...]Heat-stable toxin (STa)-producing enterotoxigenic Escherichia coli (ETEC) strains are a top cause of moderate-to-severe diarrhea in children from developing countries and a common cause of travelers' diarrhea. Recent progress in using STa toxoids and toxoid fusions to induce neutralizing anti-STa antibodies accelerates ETEC vaccine development. However, concern remains whether the derived anti-STa antibodies cross-react with STa-like guanylin and uroguanylin, two GC-C ligands regulating the fluid and electrolyte transportation in human intestinal and renal epithelial cells. To further divert STa from guanylin and uroguanylin structurally and antigenically and to eliminate anti-STa antibody cross-reactivity with guanylin and uroguanylin, we mutated STa toxin at the 9th (leucine), the 12th (asparagine) and the 14th (alanine) residues for double and triple mutants STaL9A/N12S, STaL9A/A14H, STaN12S/A14T and STaL9A/N12S/A14H. We then fused each STa mutant (three copies) to a monomeric LT mutant (mnLTR192G/L211A) for toxoid fusions 3xSTaL9A/N12S-mnLTR192G/L211A, 3xSTaL9A/A14H-mnLTR192G/L211A, 3xSTaN12S/A14T-mnLTR192G/L211A and 3xSTaL9A/N12S/A14H-mnLTR192G/L211A, examined each fusion for anti-STa immunogenicity, and assessed derived antibodies for in vitro neutralization activity against STa toxicity and for cross-reactivity with guanylin and uroguanylin. Mice subcutaneously immunized with each fusion protein developed anti-STa antibodies, and the antibodies derived from 3xSTaN12S-mnLTR192G/L211A, 3xSTaL9A/N12S-mnLTR192G/L211A or 3xSTaN12S/A14T-mnLTR192G/L211A prevented STa from stimulation of intracellular cGMP in T-84 cells. Competitive ELISAs showed guanylin and uroguanylin hardly blocked the binding of anti-STa antibodies to the coated STa-ovalbumin conjugate. These results indicated that the antibodies derived from 3xSTaN12S-mnLTR192G/L211A, 3xSTaL9A/N12S-mnLTR192G/L211A or 3xSTaN12S/A14T-mnLTR192G/L211A neutralized STa toxin and had little cross-reactivity with guanylin and uroguanylin, suggesting these toxoid fusions are suitable antigens for ETEC vaccines.
Importance
Enterotoxigenic Escherichia coli (ETEC) strains are a leading cause of children' diarrhea and travelers' diarrhea. Currently, there is no licensed vaccine against ETEC diarrhea. One key challenge is to identify safe antigens to induce antibodies neutralizing the key STa toxin without cross-reacting with guanylin and uroguanylin, two important ligands controlling the homeostasis in human intestinal and renal epithelial cells. In this study, we generated nontoxic fusion antigens that induced antibodies neutralizing STa enterotoxicity in vitro and not cross-reacting with guanylin or uroguanylin. These fusions become the preferred antigens for developing ETEC vaccines to potentially prevent the deaths of hundreds of thousands of young children and hundreds of millions of diarrheal cases each year.Expand abstractOct 2017Applied and Environmental MicrobiologyTannerella forsythia and Fusobacterium nucleatum are dental plaque bacteria implicated in the development of periodontitis. These two species have been shown to form synergistic biofilms and found to be closely associated in dental plaque biofilms. A number of genetic loci for TonB-dependent membrane receptors (TDR) for glycan acquisition, with many existing in association with genes coding for enzymes involved in the breakdown of complex glycans, have been identified in T. forsythia. In this study, we focused on a locus, BFO_0186 -- BFO_0188, that codes for a predicted TDR-SusD transporter along with a putative β-glucan hydrolyzing enzyme (BFO_0186). This operon is located immediately downstream of a two-gene operon that codes for a putative stress-responsive extracytoplasmic function (ECF) Sigma factor and an anti-Sigma factor. Here, we show that BFO_0186 expresses a β-glucanase that cleaves glucans with β-1,6 and -1, 3 linkages. Furthermore, BFO_0186-BFO_0188 locus is upregulated, with an induction of β-glucanase activity, in co-biofims of T. forsythia and F. nucleatum. The β-glucanase activity in mixed biofilms in turn leads to an enhanced hydrolysis of β-glucans and release of glucose monomers and oligomers as nutrients for F. nucleatum. In summary, our study highlights the role of T. forsythia β-glucanase expressed by the asaccharolytic oral bacterium T. forsythia in the development of T. forsythia - F. nucleatum mixed species biofilms, and suggest that dietary β-glucans might contribute in plaque development and periodontal disease pathogenesis.
IMPORTANCE
The development of dental plaque biofilm is a complex process in which metabolic, chemical and physical interactions between bacteria take a central role. Previous studies have shown that the dental pathogen T. forsythia and F. nucleatum form synergistic biofilms and are closely associated in the human dental plaque. In this study, we show that β-glucanase from the periodontal pathogen T. forsythia plays a role in the formation of T. forsythia-F. nucleatum co-biofilms by hydrolyzing β-glucans to glucose as a nutrient. We also unveiled that the expression of T. forsythia β-glucanase is induced in response to F. nucleatum sensing. This study highlights the involvement of β-glucanase activity in the development of T. forsythia - F. nucleatum biofilms, and suggest that intake of dietary β-glucans might be a contributing risk factor in plaque development and periodontal disease pathogenesis.Expand abstractOct 2017Applied and Environmental MicrobiologyEnterococcus faecalis is a commensal of the human gastrointestinal tract that can persist in the external environment and is a leading cause of hospital acquired infections. Given its diverse habitats, the organism has developed numerous strategies to survive a multitude of environmental conditions. Previous studies have demonstrated that E. faecalis will incorporate fatty acids from bile and serum into its membrane, resulting in an induced tolerance to membrane damaging agents. To discern whether all fatty acids induce membrane stress protection, we examined how E. faecalis responded to individually supplied fatty acids. E. faecalis readily incorporated fatty acids 14-18 carbons in length into its membrane, but poorly incorporated fatty acids shorter or longer than this length. Supplementation with saturated fatty acids tended to increase generation time and lead to altered cellular morphology in most cases. Further, exogenously supplied saturated fatty acids did not induce tolerance to the membrane damaging antibiotic daptomycin. Supplementation with unsaturated fatty acids produced variable growth effects, with some impacting generation time and morphology. Exogenously supplied unsaturated fatty acids that are normally produced by E. faecalis and those that are found in bile or serum, could restore growth in the presence of a fatty acid biosynthetic inhibitor. However, only the eukaryotic derived fatty acids, oleic acid and linoleic acid, provided protection from daptomycin. Thus, exogenous fatty acids do not lead to a common physiological effect on E. faecalis: the organism responds uniquely to each, and only host-derived fatty acids induce membrane protection.
Importance
Enterococcus faecalis is a commonly acquired hospital infectious agent with resistance to many antibiotics, including those that target its cellular membrane. We previously demonstrated that E. faecalis will incorporate fatty acids found in human fluids, like serum, into its cellular membrane, thereby altering its membrane composition. In turn, the organism is better able to survive membrane damaging agents, including the antibiotic daptomycin. We examined fatty acids commonly found in serum and those normally produced by E. faecalis to determine which fatty acids can induce protection from membrane damage. Supplementation with individual fatty acids produced a myriad of different effects on cellular growth, morphology, and stress response. However, only host-derived unsaturated fatty acids provided stress protection. Future studies are aimed to understand how these specific fatty acids induce protection from membrane damage.Expand abstractOct 2017Applied and Environmental MicrobiologySoil and plant inoculation with heterotrophic Zinc Solubilizing Bacteria (ZSB) is considered as a promising approach for increasing zinc (Zn) phytoavailability and enhance crop growth and nutritional quality. It is nevertheless necessary to understand the underlying bacterial solubilization processes in order to predict their repeatability in inoculation strategies. Acidification via gluconic acid production remains the most reported process. In this study, wheat rhizosphere soil serial dilutions were plated onto several solid microbiological media supplemented with scarcely soluble Zn oxide (ZnO) and 115 putative Zn solubilizing isolates were directly detected based on the formation of solubilization halo around the colonies. Eight strains were selected based on their Zn solubilization efficiency and siderophore production capacity. These included one Curtobacterium, two Plantibacter, three Pseudomonas, one Stenotrophomonas and one strain of Streptomyces. In ZnO liquid solubilization assays, the presence of glucose clearly stimulated organic acid production, leading to media acidification and ZnO solubilization. While solubilization by Streptomyces and Curtobacterium was attributed, respectively, to the accumulated production of seven and eight different organic acids, the other strains solubilized Zn via gluconic, malonic and oxalic acid, exclusively. In contrast, in absence of glucose, ZnO dissolution resulted from protons extrusion (e.g. via ammonia consumption by Plantibacter strains) and complexation processes (i.e. complexation with glutamic acid in cultures of Curtobacterium). Therefore, while gluconic acid production was described as a major Zn solubilization mechanism in literature, this study goes beyond and shows that solubilization mechanisms are various among ZSB and strongly affected by growth conditions.
IMPORTANCE
Barriers towards a better understanding of the mechanisms underlying Zn solubilization by bacteria include the lack of methodological tools for isolation, discrimination and identification of such organisms. Our study proposes a direct bacterial isolation procedure, which prevent the need of screening numerous bacterial candidates (i.e. which ability to solubilize Zn is unknown) for recovering ZSB. Moreover, we confirm the potential of Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry [MALDI-TOF MS] as a quick and accurate tool for the identification and discrimination of environmental bacterial isolates. This work also describes various Zn solubilization processes used by wheat rhizosphere bacteria, including proton extrusion and production of different organic acids among bacterial strains. These processes were also clearly affected by growth conditions (i.e. solid versus liquid cultures, presence and absence of glucose). Although highlighted mechanisms may have significant effects at the soil-plant interface, these should only be transposed to real ecological situations cautiously.Expand abstractOct 2017Applied and Environmental Microbiology[...]Pseudomonas sp. strains C5pp and C7 degrade carbaryl as the sole carbon source. Carbaryl hydrolase (CH) catalyzes the hydrolysis of carbaryl to 1-naphthol and methylamine. Bioinformatic analysis of mcbA encoding CH in C5pp predicted it to have a transmembrane domain (Tmd) and a signal peptide (Sp). In these isolates, the activity of CH was found to be 4 to 6-fold higher in the periplasm as compared to the cytoplasm. The recombinant CH (rCH) showed 4-fold higher activity in the periplasm of Escherichia coli. The deletion of Tmd showed activity in the cytoplasmic fraction, while deletion of Tmd+Sp resulted in expression of the inactive protein. Confocal microscopic analysis of E. coli expressing (Tmd+Sp)-GFP fusion protein revealed the localization of GFP into the periplasm. Altogether, these results indicate that Tmd probably helps in anchoring of polypeptide to the inner membrane, while Sp assists folding and release of CH in the periplasm. N-terminal sequence of the mature periplasmic CH confirms the absence of Tmd+Sp region and the signal peptidase cleavage site as Ala-Leu-Ala. CH purified from strains C5pp, C7 and rCHΔ(Tmd)a were found to be monomeric with molecular mass of ~68-76 kDa, catalyze hydrolysis of ester bond with apparent Km and Vmax in the range of 98-111 μM and 69-73 μmol.min??.mg??, respectively. The presence of low-affinity CH in the periplasm and 1-naphthol metabolizing enzymes in the cytoplasm of Pseudomonas spp. suggests the compartmentalization of metabolic pathway as a strategy for efficient degradation of carbaryl at higher concentration without cellular toxicity of 1-naphthol.
IMPORTANCE
Proteins in the periplasmic space of bacteria play an important role in various cellular processes like solute transport, nutrient binding, antibiotic resistance, substrate hydrolysis and detoxification of xenobiotics. Carbaryl is one of the most widely used carbamate pesticide. Carbaryl hydrolase (CH), the first enzyme of the degradation pathway which converts carbaryl to 1-naphthol, was found to be localized in the periplasm of Pseudomonas spp. Predicted transmembrane domain and signal peptide sequences of Pseudomonas were found to be functional in Escherichia coli and translocate CH and GFP into the periplasm. The localization of low affinity CH into the periplasm indicates controlled formation of toxic and recalcitrant 1-naphthol, thus minimizing its accumulation and interaction with various cellular components thereby reducing the cellular toxicity. This study highlights the significance of compartmentalization of metabolic pathway enzymes for efficient removal of toxic compounds.Expand abstractOct 2017Applied and Environmental Microbiology[...]Commensal Streptococcus sanguinis and Streptococcus gordonii are pioneer oral biofilm colonizers. Characteristic for both is the SpxB-dependent production of H2O2, which is crucial for inhibiting competing biofilm members, especially the cariogenic species Streptococcus mutans. H2O2 production is strongly impacted by environmental conditions, yet few mechanisms are known. Dental plaque pH is one of the key parameters dictating dental plaque ecology, and ultimately oral health status. Therefore, the objective of the current study was to characterize the effect of environmental pH upon H2O2 production by S. sanguinis and S. gordonii. S. sanguinis H2O2 production was not found to be affected by moderate changes in environmental pH, whereas S. gordonii H2O2 production declined markedly in response to lower pH. Further investigation into the pyruvate node, the central metabolic switch modulating H2O2 or lactic acid production, revealed increased lactic acid levels from S. gordonii at pH6. The bias for lactic acid production at pH6 resulted in a concomitant improvement in the survival of S. gordonii at low pH and seems to comprise part of its acid tolerance response. Additionally, the differential response to pH similarly affects other oral streptococcal species suggesting that the observed results are part of a larger phenomenon linking environmental pH, central metabolism, and the capacity to produce antagonistic amounts of H2O2.
IMPORTANCE
The oral biofilm is subject to frequent and dramatic changes in pH. S. sanguinis and S. gordonii can compete with caries/periodontitis-associated pathogens by generating H2O2. Therefore, it is crucial to understand how S. sanguinis and S. gordonii adapt to low pH and maintain their competitiveness under acid stress. The present study provides evidence that certain oral bacteria respond to environmental pH changes by tuning the metabolic output in favor of lactic acid production to increase their acid survival, while others maintain their H2O2 production at a constant level. The differential control of H2O2 production provides important insights into the role of environmental conditions for growth competition among the oral flora.Expand abstractOct 2017Applied and Environmental MicrobiologyVegetable seeds contaminated with bacterial pathogens have been linked to fresh produce-associated outbreaks of gastrointestinal infections. This study was undertaken to observe the physiological behavior of Salmonella enterica and enterohemorrhagic Escherichia coli (EHEC) cells, artificially internalized into vegetable seeds during the germination process. Surface-decontaminated seeds of alfalfa, fenugreek, lettuce, and tomato were vacuum-infiltrated with four individual strains of Salmonella or EHEC. Contaminated seeds were germinated at 25°C for 9 days and different sprout/seedling tissues were microbiologically analyzed every other day. Internalization of Salmonella and EHEC cells into vegetable seeds was confirmed by the absence of pathogens in seed-rinsing water and presence of pathogens in seed homogenates after post-internalization seed surface decontamination. Results show that 317 (62%) and 343 (67%) out of the 512 collected sprout/seedling tissue samples were positive for Salmonella and EHEC, respectively. Average Salmonella populations were significantly higher (p&0.05) than the EHEC populations. Significantly higher Salmonella populations were recovered from the cotyledon and seed coat tissues, followed by the root tissues, but the mean EHEC populations from all sampled tissue sections were statistically similar, except pre-germinated seeds. Three Salmonella, and two EHEC, strains had significantly higher cell populations on sprout/seedling tissues than other strains used in the study. Salmonella and EHEC populations from fenugreek and alfalfa tissues were significantly higher than those from tomato and lettuce tissues. The study observed the fate of internalized human pathogens on germinating vegetable seeds and sprout/seedling tissues and emphasized the importance of using pathogen-free seeds for sprout production.
IMPORTANCE
Internalization of microorganisms into vegetable seeds could occur naturally and represents a possible pathway of vegetable seed contamination by human pathogens. The present study investigated the ability of two important bacterial pathogens, Salmonella and EHEC, when artificially internalized into vegetable seeds, to grow and disseminate along vegetable sprouts/seedlings during germination. Data of the study revealed that the pathogen cells artificially internalized into vegetable seeds caused the contamination of different tissues of sprouts/seedlings, and pathogen growth on germinating seeds is bacterial species- and vegetable seed type-dependent. These results further stress the necessity of using pathogen-free vegetable seeds for edible sprout production.Expand abstractOct 2017Applied and Environmental Microbiology[...]Homology searches indicate that Saccharomyces cerevisiae strain BY4741 contains seven redundant genes that encode putative aryl-alcohol dehydrogenases (AAD). Yeast AADs are located in subtelomeric regions of different chromosomes, and their functional role(s) remain enigmatic. Here, we show that two of these genes, AAD4 and AAD14, encode functional enzymes that reduce aliphatic and aryl-aldehydes concomitant with oxidation of cofactor NADPH, and that Aad4p and Aad14p exhibit different substrate preference patterns. Other yeast AADs are undergoing pseudogenization. The 5’ sequence of AAD15 has been deleted from the genome. Repair of an AAD3 missense mutation at the catalytically essential Tyr?? residue did not result in a functional enzyme. However, ancestral state reconstruction by fusing Aad6 with Aad16, and by N-terminal repair of Aad10 restores NADPH-dependent, aryl-alcohol dehydrogenase activities. Phylogenetic analysis indicates that AAD genes are narrowly distributed in wood-saprophyte fungi and in yeast that occupy lignocellulosic niches. Because yeast AADs exhibit activity on veratraldehyde, cinnamaldehyde and vanillin they could serve to detoxify aryl-aldehydes released during lignin degradation. However, none of these compounds induce yeast AAD gene expression, and Aad activities do not relieve aryl-aldehyde growth inhibition. Our data suggest an ancestral role for AAD genes in lignin degradation that is degenerating as a result of yeast's domestication and use in brewing, baking and other industrial applications.
IMPORTANCE
Functional characterization of hypothetical genes remains one of the chief tasks of the post-genomic era. Although the first Saccharomyces cerevisiae genome sequence was published over 20 years ago, 22% of its estimated 6603 ORFs remain unverified. One outstanding example of this category of genes is the enigmatic seven-member AAD family. Here we demonstrate that proteins encoded by two members of this family exhibit aliphatic and aryl-aldehydes reductase activity, and further that such activity can be recovered from pseudogenized AADs via ancestral state reconstruction. The phylogeny of yeast AADs suggests that these proteins may have played an important ancestral role in detoxifying aromatic aldehydes in ligninolytic fungi. However, in yeast adapted to niches rich in sugars AADs become subject to mutational erosion. Our findings shed new light on the selective pressures and molecular mechanisms by which genes undergo pseudogenization.Expand abstractOct 2017Applied and Environmental MicrobiologyIn soy sauce manufacturing, Candida versatilis plays a role in the production of volatile flavor compounds, such as volatile phenols, but limited accessible information on its genome has prevented further investigation regarding aroma production and breeding. Although the draft genome sequence data of two strains of C. versatilis have recently been reported, these strains are not similar to each other. Here, we reassess the draft genome sequence data for strain t-1, which was originally reported to be C. versatilis, and conclude that strain t-1 is most probably not C. versatilis but a gamete of hybrid Zygosaccharomyces rouxii. Phylogenetic analysis of the D1/D2 region of the 26S rDNA sequence indicated that strain t-1 is more similar to the genus Zygosaccharomyces than C. versatilis. Moreover, we found that the genome of strain t-1 is composed of haploid genome content and divided into two regions that show approximately 100% identity with the T- or P-subgenome derived from the natural hybrid Zygosaccharomyces rouxii, such as NBRC110957 and NBRC1876. We also found a chromosome crossing-over signature in the scaffolds of strain t-1. These results suggest that strain t-1 is a gamete of the hybrid Z. rouxii, generated by either meiosis or chromosome loss following reciprocal translocation between the T- and P-subgenomes. Although it is unclear why strain t-1 was misidentified as C. versatilis, the genome of strain t-1 has broad implications for considering the evolutionary fate of an allodiploid.
Importance
In yeast, crossing between different species sometimes leads to interspecies hybrids. The hybrid generally cannot produce viable spores because dissimilarity of parental genomes prevents normal chromosome segregation during meiotic division, leading to a dead end. Thus, only a handful of natural cases of homoploid hybrid speciation, which requires mating between 1n gametes of hybrids, have been described. However, a recent study provided strong evidence that homoploid hybrid speciation initiates in natural populations of the budding yeast, suggesting the potential presence of viable 1n gametes of hybrids. The significance of our study is finding that the strain t-1, which had been misidentified as Candida versatilis, is a viable 1n gamete derived from hybrid Zygosaccharomyces rouxii.Expand abstractOct 2017Applied and Environmental Microbiology[...]The critically endangered elkhorn coral, Acropora palmata, is affected by white pox disease (WPX) throughout the Florida Reef Tract and wider Caribbean. The bacterium Serratia marcescens was previously identified as one etiologic agent of WPX, but is no longer consistently detected in contemporary outbreaks. It is now believed that multiple etiologic agents cause WPX; however, to date, no other potential pathogens have been thoroughly investigated. This study examined the association of Vibrio bacteria with WPX occurrence from August
at Looe Key Reef in the Florida Keys, USA. The concentration of cultivable Vibrio was consistently greater in WPX samples compared to healthy samples. The relative abundance of Vibrio bacteria to total bacteria was four times higher in samples from WPX lesions compared to adjacent apparently healthy regions of diseased corals based on quantitative PCR (qPCR). Multilocus sequence analysis (MLSA) was used to assess the diversity of 69 Vibrio isolates collected from diseased and apparently healthy A. palmata colonies and the surrounding seawater. Vibrio species with known pathogenicity to corals were detected in both apparently healthy and diseased samples. While the causative agent(s) of contemporary WPX outbreaks remain elusive, our results suggest that Vibrio may be part of a non-specific heterotrophic bacterial bloom rather than acting as primary pathogens. This study highlights the need for highly resolved temporal sampling in situ to further elucidate the role of Vibrio during WPX onset and progression.
Importance
Coral diseases are increasing worldwide and are now considered a major contributor to coral reef decline. In particular, the Caribbean has been noted as a coral disease hotspot owing to the dramatic loss of framework-building acroporid corals due to tissue loss diseases. The pathogenesis of contemporary white pox disease (WPX) outbreaks in Acropora palmata remains poorly understood. This study investigates the association of Vibrio bacteria with WPX.Expand abstractOct 2017Applied and Environmental Microbiology[...]Norovirus (NoV) is the leading cause of gastroenteritis outbreaks linked to oyster consumption. In this study, we investigated the potential of F-specific RNA bacteriophages (FRNAPH) as indicators of viral contamination in oysters, by focusing especially on the subgroup FRNAPH-II. These viral indicators have been neglected because of their sometimes different behavior from that of NoV in shellfish, especially during the depuration processes usually performed before marketing. However, a significant bias needs to be taken into account which is that, in the absence of routine culture methods, NoV is targeted by genome detection while FRNAPH presence is usually investigated by isolation of infectious particles. In this study, by targeting both viruses using genome detection, a significant correlation was observed between FRNAPH-II and NoV in shellfish collected from various European harvesting areas impacted by fecal pollution. Moreover, during their depuration, while high persistence of NoV was confirmed, similar or even greater persistence was observed for FRNAPH-II genome over 30 days. Such striking genome persistence calls into question the relevance of molecular methods in assessing viral hazard. Targeting the same virus (i.e. FRNAPH-II) by culture and genome detection in specimens coming from harvesting areas as well as during depuration, we concluded that the presence of genomes in shellfish does not provide any information on the presence of the corresponding infectious particles. In view of these results, infectious FRNAPH detection should be reconsidered as a valuable indicator in oysters and its potential for assessing viral hazard needs to be investigated.
IMPORTANCE
This work brings new data about the behavior of viruses in shellfish, as well as concerning the relevance of molecular methods to detect them and evaluate the viral hazard. Firstly, a high correlation has been observed between F-specific RNA bacteriophages of subgroup II (FRNAPH-II) and norovirus (NoV) in shellfish impacted by fecal contamination, when both are detected using molecular approaches. Secondly, when using RT-PCR and culture to detect FRNAPH-II in shellfish, it appears that genome of viruses presents higher persistence compared to infectivity, and thus their detection fails to give information about the concomitant presence of infectious viruses. At last, this study shows that persistence of FRNAPH is at least as long as that of NoV. These data are major arguments to reconsider the potential of FRNAPH as indicators of shellfish viral quality.Expand abstractOct 2017Applied and Environmental MicrobiologyTrichoderma reesei can produce up to 100 g/l of extracellular proteins. The major an