Enumeration and identification of dominant types of sulfate-reducing bacteria in pulp from a paper-recycling plant: a multiphasic approach

Multiple independent approaches were applied for monitoring the abundance and identity of sulfate-reducing bacteria (SRB) in pulp of a paper-recycling plant suffering from excessive sulfide emission. The methods applied included most-probable-number (MPN) enumeration of cultivable SRB, rate measurements, FISH and PCR-based retrieval of the functional marker genes dsrA and B (encoding the two major subunits of dissimilatory bisulfite reductase) and 16S rRNA genes. The SRB community was composed of phylogenetically highly different lineages all of low abundance relative to the total microbial community in the pulp, which hampered the applicability of FISH. It was also demonstrated that dsrA- or B -targeted PCR primers commonly used for denaturing gradient gel electrophoresis and real-time PCR analyses were biased. However, using a novel approach combining MPN-PCR and terminal restriction fragment length polymorphism analysis of dsrAB amplicons generated from serially diluted DNA extracts allowed the enumeration and identification of the quantitatively most important members of the SRB community. For fast quantification of SRB in the pulp, the dsrAB -MPN-PCR assay and sulfate reduction rate measurements were found to be most suitable.     

TRAMPR: AN R package for analysis and matching of terminal‐restriction fragment length polymorphism (TRFLP) profiles

trampr (TRFLP analysis and matching package for r ) is a package for matching multiple terminal restriction fragment length polymorphism (TRFLP) profiles between unknown samples and a database of known TRFLP profiles in order to infer the presence of species in environmental samples. It permits simultaneous analysis of multiple samples and facilitates direct workflow from electrophoresis output through to community analyses. trampr also resolves the issues of multiple TRFLP profiles within a species and (conversely) shared TRFLP profiles across species.     

Bacteriocidal effects of brevetoxin on natural microbial communities

The sensitivity of bacteria to the marine neurotoxins, brevetoxins, produced by the dinoflagellate Karenia brevis and raphidophytes Chattonella spp. remains an open question. We investigated the bacteriocidal effects of brevetoxin (PbTx-2) on the abundance and community composition of natural microbial communities by adding it to microbes from three coastal marine locations that have varying degrees of historical brevetoxin exposure: (1) Great Bay, New Jersey, (2) Rehoboth Bay, Delaware and (3) Sarasota Bay, Florida. The populations with limited or no documented exposure were more susceptible to the effects of PbTx-2 than the Gulf of Mexico populations which are frequently exposed to brevetoxins. The community with no prior documented exposure to brevetoxins showed significant (p = 0.03) changes in bacterial abundance occurring with additions greater than 2.5 μg PbTx-2 L⁻¹. Brevetoxin concentrations during K. brevis blooms range from ∼2.5 to nearly 100 μg L⁻¹ with typical concentrations of ∼10–30 μg L⁻¹. In contrast to the unexposed populations, there was no significant decrease in bacterial cell number for the microbial community that was frequently exposed to brevetoxins, which implies variable sensitivity in natural communities. The diversity in the bacterial communities that were sensitive to PbTx-2 declined upon exposure. This suggests that the PbTx-2 was selecting for or against specific species. Mortality was much higher in the 200 μg PbTx-2 L⁻¹ treatment after 48 h and >37% of the species disappeared in the bacterial communities with no documented exposure. These results suggest that toxic red tides may play a role in structuring bacterial communities.     

Microbial community structures in anoxic freshwater lake sediment along a metal contamination gradient

Contamination, such as by heavy metals, has frequently been implicated in altering microbial community structure. However, this association has not been extensively studied for anaerobic communities, or in freshwater lake sediments. We investigated microbial community structure in the metal-contaminated anoxic sediments of a eutrophic lake that were impacted over the course of 80 years by nearby zinc-smelting activities. Microbial community structure was inferred for bacterial, archaeal and eukaryotic populations by evaluating terminal restriction fragment length polymorphism (TRFLP) patterns in near-surface sediments collected in triplicate from five areas of the lake that had differing levels of metal contamination. The majority of the fragments in the bacterial and eukaryotic profiles showed no evidence of variation in association with metal contamination levels, and diversity revealed by these profiles remained consistent even as metal concentrations varied from 3000 to 27 000 mg kg⁻¹ total Zn, 0.125 to 11.2 μ pore water Zn and 0.023 to 5.40 μ pore water As. Although most archaeal fragments also showed no evidence of variation, the prevalence of a fragment associated with mesophilic Crenarchaeota showed significant positive correlation with total Zn concentrations. This Crenarchaeota fragment dominated the archaeal TRFLP profiles, representing between 35% and 79% of the total measured peak areas. Lake DePue 16S rRNA gene sequences corresponding to this TRFLP fragment clustered with anaerobic and soil mesophilic Crenarchaeota sequences. Although Crenarchaeota have been associated with metal-contaminated groundwater and soils, this is a first report (to our knowledge) documenting potential increased prevalence of Crenarchaeota associated with elevated levels of metal contamination.     

Influence of arbuscular mycorrhizal mycelial exudates on soil bacterial growth and community structure

Plant root systems colonized by arbuscular mycorrhizal (AM) fungi have previously been shown to influence soil bacterial populations; however, the direct influence of the AM extraradical mycelium itself on bacterial growth and community composition is not well understood. In this study, we investigated the effects of exudates produced by AM extraradical mycelia on the growth and development of an extracted soil bacterial community in vitro. The chemical composition of the mycelial exudates was analysed using proton nuclear magnetic resonance spectrometry. Following the addition of exudates to a bacterial community extracted from soil, bacterial growth and vitality were determined using a bacterial vitality stain and fluorescence microscopy. Changes in community composition were also analysed at various times over the course of 3 days by terminal restriction fragment length polymorphism analysis, in combination with cloning and sequencing of 16S rRNA genes. Mycelial exudates increased bacterial growth and vitality and changed bacterial community composition. Several Gammaproteobacteria, including a taxon within the Enterobacteriaceae, increased in frequency of occurrence in response to AM mycelial exudates. This study is the first attempt to identify carbohydrates from the extraradical mycelium of an AM fungus, and demonstrates the direct effects of mycelial exudates on a soil bacterial community.     

Microbial macroecology: highly structured prokaryotic soil assemblages in a tropical deciduous forest

Aim To assess the hypothesis that free‐living prokaryotes show a pattern of ‘no biogeography’ by examining the scaling of soil prokaryotic diversity and by comparing it with other groups’ biogeographical patterns. Location Two sites in the tropical deciduous forest of Chamela, Jalisco, on the western coast of Mexico. Methods We examined the diversity and distribution of soil prokaryotes in two 8 × 8 m quadrats divided in such manner that we could sample at four spatial scales. Restriction fragment length polymorphisms of 16S rRNA genes were used to define operational taxonomic units (OTUs) that we used in lieu of species to assess diversity. Results We found highly structured species assemblages that allowed us to reject multiple predictions of the hypothesis that soil bacteria show ‘no biogeography’. The frequency distribution of range size (measured as the occupancy of quadrats) of OTUs followed a hollow curve similar to that of vertebrates on continents. Assemblages showed high levels of beta diversity and a non‐random nested pattern of diversity. OTU diversity scaled with area followed a power function with slopes z = 0.42 and 0.47. Main conclusions We demonstrate a non‐ubiquitous dispersal for soil prokaryotes, which suggests a complex biogeography similar to that found for terrestrial vertebrates.     

Metabolite induction via microorganism co-culture: A potential way to enhance chemical diversity for drug discovery

Microorganisms have a long track record as important sources of novel bioactive natural products, particularly in the field of drug discovery. While microbes have been shown to biosynthesize a wide array of molecules, recent advances in genome sequencing have revealed that such organisms have the potential to yield even more structurally diverse secondary metabolites. Thus, many microbial gene clusters may be silent under standard laboratory growth conditions. In the last ten years, several methods have been developed to aid in the activation of these cryptic biosynthetic pathways. In addition to the techniques that demand prior knowledge of the genome sequences of the studied microorganisms, several genome sequence-independent tools have been developed. One of these approaches is microorganism co-culture, involving the cultivation of two or more microorganisms in the same confined environment. Microorganism co-culture is inspired by the natural microbe communities that are omnipresent in nature. Within these communities, microbes interact through signaling or defense molecules. Such compounds, produced dynamically, are of potential interest as new leads for drug discovery. Microorganism co-culture can be achieved in either solid or liquid media and has recently been used increasingly extensively to study natural interactions and discover new bioactive metabolites. Because of the complexity of microbial extracts, advanced analytical methods (e.g., mass spectrometry methods and metabolomics) are key for the successful detection and identification of co-culture-induced metabolites.     

Crenarchaeal heterotrophy in salt marsh sediments

Mesophilic Crenarchaeota (also known as Thaumarchaeota) are ubiquitous and abundant in marine habitats. However, very little is known about their metabolic function in situ. In this study, salt marsh sediments from New Jersey were screened via stable isotope probing (SIP) for heterotrophy by amending with a single ¹³C-labeled compound (acetate, glycine or urea) or a complex ¹³C-biopolymer (lipids, proteins or growth medium (ISOGRO)). SIP incubations were done at two substrate concentrations (30–150 μM; 2–10 mg ml⁻¹), and ¹³C-labeled DNA was analyzed by terminal restriction fragment length polymorphism (TRFLP) analysis of 16S rRNA genes. To test for autotrophy, an amendment with ¹³C-bicarbonate was also performed. Our SIP analyses indicate salt marsh crenarchaea are heterotrophic, double within 2–3 days and often compete with heterotrophic bacteria for the same organic substrates. A clone library of ¹³C-amplicons was screened to find matches to the ¹³C-TRFLP peaks, with seven members of the Miscellaneous Crenarchaeal Group and seven members from the Marine Group 1.a Crenarchaeota being discerned. Some of these crenarchaea displayed a preference for particular carbon sources, whereas others incorporated nearly every ¹³C-substrate provided. The data suggest salt marshes may be an excellent model system for studying crenarchaeal metabolic capabilities and can provide information on the competition between crenarchaea and other microbial groups to improve our understanding of microbial ecology.     

Toxicants inhibiting anaerobic digestion: A review

Anaerobic digestion is increasingly being used to treat wastes from many sources because of its manifold advantages over aerobic treatment, e.g. low sludge production and low energy requirements. However, anaerobic digestion is sensitive to toxicants, and a wide range of compounds can inhibit the process and cause upset or failure. Substantial research has been carried out over the years to identify specific inhibitors/toxicants, and their mechanism of toxicity in anaerobic digestion. In this review we present a detailed and critical summary of research on the inhibition of anaerobic processes by specific organic toxicants (e.g., chlorophenols, halogenated aliphatics and long chain fatty acids), inorganic toxicants (e.g., ammonia, sulfide and heavy metals) and in particular, nanomaterials, focusing on the mechanism of their inhibition/toxicity. A better understanding of the fundamental mechanisms behind inhibition/toxicity will enhance the wider application of anaerobic digestion.