Fluorescence Characteristics of Coalbed Methane Produced Water and Its Influence on Freshwater Bacteria in the South Qinshui Basin, China

Production of coalbed methane (CBM) resources commonly requires using hydraulic fracturing and chemical production well additives. Concern exists for the existence of chemical compounds in CBM produced water, due to the risk of environmental receptor contamination. In this study, parallel factor method analysis (PARAFAC), fluorescence index, and the fluorescence area integral methods were used to analyse the properties of CBM produced water sampled from Shizhuang Block (one of the most active CBM-producing regions in the Qinshui Basin). A culture experiment was designed to determine the effect of discharged CBM produced water on microorganisms in freshwater. Water quality analysis shows the hydrochemistry of most water samples as Na-HCO3 type produced water of CBM appears as a generally weak alkaline (pH 8.69 ± 0.185) with high salinity, high alkalinity, and a high chemical oxygen demand (COD) value. Three individual components were identified by using parallel factor method analysis as humic-like components (C1), fulvic-like components (C2), and amino acid-like substances (C3). The fluorescence characteristic index comprehensively explains that the fluorescent substances in CBM produced water has the characteristics of a low degree of humification and a high recent self-generating source. The region integration results of characteristic peaks show that tyrosine-like and tryptophan-like materials account for more than 67% of fluorescent substances in CBM produced water. The addition of produced water from coalbed methane promotes the growth of freshwater bacteria, and this process is accompanied by the decrease of the proportion of fulvic acid, humic acid, and the increase of the proportion of soluble microbial metabolites. This paper proposes a convenient method for organic matter identification of CBM produced water and provides some theoretical support and reference for the improvement of CBM water treatment and utilization.

Severe bilateral pleuropneumonia caused by Legionella sainthelensi: a case report

Background Legionella spp. are ubiquitous freshwater bacteria responsible for rare but potentially severe cases of Legionnaires’ disease (LD). Legionella sainthelensi is a non-pneumophila Legionella species that was first isolated in 1980 from water near Mt. St-Helens (USA). Although rare cases of LD caused by L. sainthelensi have been reported, very little data is available on this pathogen. Case presentation We describe the first documented case of severe bilateral pleuropneumonia caused by L. sainthelensi. The patient was a 35-year-old woman with Sharp’s syndrome treated with long-term hydroxychloroquine and corticosteroids who was hospitalized for an infectious illness in a university hospital in Reunion Island (France). The patient’s clinical presentation was complicated at first (bilateral pneumonia, multiloculated pleural effusion, then bronchopleural fistula) but her clinical condition eventually improved with the reintroduction of macrolides (spiramycin) in intensive care unit. Etiological diagnosis was confirmed by PCR syndromic assay and culture on bronchoalveolar lavage. Conclusions To date, only 14 documented cases of L. sainthelensi infection have been described worldwide. This pathogen is difficult to identify because it is not or poorly detected by urinary antigen and molecular methods (like PCR syndromic assays that primarily target L. pneumophila and that have only recently been deployed in microbiology laboratories). Pneumonia caused by L. sainthelensi is likely underdiagnosed as a result. Clinicians should consider the possibility of non-pneumophila Legionella infection in patients with a compatible clinical presentation when microbiological diagnostic tools targeted L. pneumophila tested negative.

CARP EDEMA VIRUS DISEASE IN SERBIA - DISEASE OUT OF CONTROL

A poxvirus named carp edema virus (CEV), is the causative agent of carp edema virus disease (CEVD), which is an emerging disease of global concern that may cause high rates of morbidity and mortality in ornamental koi and common carp and. Common carp (Cyprinus carpio) is the most important fish species for warm‐water aquaculture in Serbia. CEVD was first time detected in Serbia in 2017. During 2017-2020 period, increasing number of CEVD outbreaks in carp farms is reported. Carp were collected from farms in different regions of Serbia from 2017 to 2020. The fish were sampled for disease diagnosis because they exhibited lethargy and anorexia, which eventually led to mortality. Mortality started with clinical signs of hypoxia while fish swam slowly without escaping reactions. The gills were pale and covered with a thick mucus layer. In advanced cases, the lesions in the gills turned into a necrotizing form. A moderate to high amount of opportunistic freshwater bacteria were isolated from gills of diseased fish. By performing real-time PCR, CEV was detected in 38 samples of diseased carps from 21 carp farms. These outbreaks further confirm the spread of CEVD and the need for practitioners to be vigilant for outbreaks of this disease. To prevent further spreading of the disease, it is very important to introduce CEV testing before fish movement. To avoid further transmission of the virus to common carp populations in Serbia, testing of CEV should become part of fish disease surveillance programs. Fish health service should be aware of the presence of CEV in Serbia which may result in high losses in carp aquaculture. Action should be taken also to prevent transmission of CEV to carp populations in open waters.

Laboratory In-Situ Production of Autochthonous and Allochthonous Fluorescent Organic Matter by Freshwater Bacteria

This work investigates the origin and range of fluorescent organic matter (FOM) produced in-situ by environmentally sourced freshwater bacteria. Aquatic FOM is an essential component in global carbon cycling and is generally classified as either autochthonous, produced in-situ via microbial processes, or allochthonous, transported into aquatic systems from external sources. We have demonstrated that, within laboratory model systems, environmentally sourced mixed microbial communities and bacterial isolates can produce and/or export FOM associated with both autochthonous and allochthonous material. This study focuses on fluorescence peak B, T, M, C and C+, exploring (1) the cellular nature of FOM produced, (2) FOM exported as extracellular material into the water column and (3) the impact of physical cell lysis on FOM signature. For the laboratory model systems studied, Peak T fluorescence is retained within bacterial cells (>68%), while Peak C fluorescence is mainly observed as extracellular material (>80%). Peak M is identified as both cellular and extracellular FOM, produced by all isolated freshwater microorganisms investigated. The origin of Peak C+ is postulated to originate from functional metabolites associated with specific microorganisms, seen specifically within the Pseudomonas sp. monoculture here. This work challenges the binary classification of FOM as either allochthonous or autochthonous, suggesting that FOM processing and production occurs along a dynamic continuum. Within this study, fluorescence intensity data for the environmental bacteria isolate monocultures are presented as enumeration corrected data, for the first time providing quantitative fluorescence data per bacterial colony forming unit (cfu). From this, we are able to assess the relative contribution of different bacteria to the autochthonous FOM pool and if this material is cellular or extracellular.

Protein Model and Function Analysis in Quorum-Sensing Pathway of Vibrio qinghaiensis sp.-Q67

Bioluminescent bacteria are mainly found in marine habitats. Vibrio qinghaiensis sp.-Q67 (Q67), a nonpathogenic freshwater bacterium, has been a focus due to its wide use in the monitoring of environmental pollution and the assessment of toxicity. However, the lack of available crystal structures limits the elucidation of the structures of the functional proteins of the quorum-sensing (QS) system that regulates bacterial luminescence in Q67. In this study, 19 functional proteins were built through monomer and oligomer modeling based on their coding proteins in the QS system of Q67 using MODELLER. Except for the failure to construct LuxM due to the lack of a suitable template, 18 functional proteins were successfully constructed. Furthermore, the relationships between the function and predicted structures of 19 functional proteins were explored one by one according to the three functional classifications: autoinducer synthases and receptors, signal transmission proteins (phosphotransferases, an RNA chaperone, and a transcriptional regulator), and enzymes involved in bacterial bioluminescence reactions. This is the first analysis of the whole process of bioluminescence regulation from the perspective of nonpathogenic freshwater bacteria at the molecular level. It provides a theoretical basis for the explanation of applications of Q67 in which luminescent inhibition is used as the endpoint.

Living in a Puddle of Mud: Isolation and Characterization of Two Novel Caulobacteraceae Strains Brevundimonas pondensis sp. nov. and Brevundimonas goettingensis sp. nov.

Brevundimonas is a genus of freshwater bacteria belonging to the family Caulobacteraceae. The present study describes two novel species of the genus Brevundimonas (LVF1T and LVF2T). Both were genomically, morphologically, and physiologically characterized. Average nucleotide identity analysis revealed both are unique among known Brevundimonas strains. In silico and additional ProphageSeq analyses resulted in two prophages in the LVF1T genome and a remnant prophage in the LVF2T genome. Bacterial LVF1T cells form an elliptical morphotype, in average 1 µm in length and 0.46 µm in width, with a single flagellum. LVF2T revealed motile cells approximately 1.6 µm in length and 0.6 µm in width with a single flagellum, and sessile cell types 1.3 µm in length and 0.6 µm in width. Both are Gram-negative, aerobic, have optimal growth at 30 °C (up to 0.5 to 1% NaCl). Both are resistant towards erythromycin, meropenem, streptomycin, tetracycline and vancomycin. Anaerobic growth was observed after 14 days for LVF1T only. For LVF1T the name Brevundimonas pondensis sp. nov. and for LVF2T the name Brevundimonas goettingensis sp. nov. are proposed. Type strains are LVF1T (=DSM 112304T = CCUG 74982T = LMG 32096T) and LVF2T (=DSM 112305T = CCUG 74983T = LMG 32097T).

Continental-scale gene flow prevents allopatric divergence of pelagic freshwater bacteria

Allopatric divergence is one of the principal mechanisms for speciation of macro-organisms. Microbes by comparison are assumed to disperse more freely and to be less limited by dispersal barriers. However, thermophilic prokaryotes restricted to geothermal springs have shown clear signals of geographic isolation, but robust studies on this topic for microbes with less strict habitat requirements are scarce. Furthermore, it has only recently been recognized that homologous recombination among conspecific individuals provides species coherence in a wide range of prokaryotes. Recombination barriers thus may define prokaryotic species boundaries, yet, the extent to which geographic distance between populations gives rise to such barriers is an open question. Here, we investigated gene flow and population structure in a widespread species of pelagic freshwater bacteria, Polynucleobacter paneuropaeus. Through comparative genomics of 113 conspecific strains isolated from freshwater lakes and ponds located across a North-South range of more than 3000 km, we were able to reconstruct past gene flow events. The species turned out to be highly recombinogenic as indicated by significant signs of gene transfer and extensive genome mosaicism. While genomic differences increased with spatial distance on a regional scale (< 170 km), such correlations were mostly absent on larger scales up to 3400 km. We conclude that allopatric divergence in European P. paneuropaeus is minor, and that effective gene flow across the sampled geographic range in combination with a high recombination efficacy maintains species coherence.

Osmotic Adaptation and Compatible Solute Biosynthesis of Phototrophic Bacteria as Revealed from Genome Analyses

Osmotic adaptation and accumulation of compatible solutes is a key process for life at high osmotic pressure and elevated salt concentrations. Most important solutes that can protect cell structures and metabolic processes at high salt concentrations are glycine betaine and ectoine. The genome analysis of more than 130 phototrophic bacteria shows that biosynthesis of glycine betaine is common among marine and halophilic phototrophic Proteobacteria and their chemotrophic relatives, as well as in representatives of Pirellulaceae and Actinobacteria, but are also found in halophilic Cyanobacteria and Chloroherpeton thalassium. This ability correlates well with the successful toleration of extreme salt concentrations. Freshwater bacteria in general lack the possibilities to synthesize and often also to take up these compounds. The biosynthesis of ectoine is found in the phylogenetic lines of phototrophic Alpha- and Gammaproteobacteria, most prominent in the Halorhodospira species and a number of Rhodobacteraceae. It is also common among Streptomycetes and Bacilli. The phylogeny of glycine-sarcosine methyltransferase (GMT) and diaminobutyrate-pyruvate aminotransferase (EctB) sequences correlate well with otherwise established phylogenetic groups. Most significantly, GMT sequences of cyanobacteria form two major phylogenetic branches and the branch of Halorhodospira species is distinct from all other Ectothiorhodospiraceae. A variety of transport systems for osmolytes are present in the studied bacteria.