Bacteriovorax Research Articles

Microbial defluorination of TFA, PFOA, and HFPO-DA by a native microbial consortium under anoxic conditions

In this study, the biodegradability of trifluoroacetate (TFA), perfluorooctanoic acid (PFOA), and perfluoro-2-methyl-3-oxahexanoic acid (HFPO-DA) by a native microbial community was evaluated over a 10-month incubation period. The observed microbial defluorination ratios and removal efficiency were 3.46 ( ± 2.73) % and 8.03 ( ± 3.03) %, 8.44 ( ± 1.88) % and 13.52 ( ± 4.96) %, 3.02 ( ± 0.62) % and 5.45 ( ± 2.99) % for TFA, PFOA and HFPO-DA, respectively. The biodegradation intermediate products, TFA and pentafluoropropionic acid (PFA), of PFOA and HFPO-DA were detected in their biodegradation treatment groups. Furthermore, the concentrations of the PFOA metabolites, perfluorohexanoic acid (PFHxA) and perfluoroheptanoic acid (PFHpA), in the aqueous solutions after incubation were quantified to be 0.21 and 4.14 µg/L. TFA, PFOA and HFPO-DA significantly reduced the microbial diversity and changed the structure of the community. The co-occurrence network analysis showed that low abundance species, such as Flexilinea flocculi, Bacteriovorax stolpii, and g_Sphingomonas, are positively correlated with the generation of fluoride ion, implying their potential collaborative functions contributing to the observed biodefluorination. The findings in this study can provide insights for the biodegradation of perfluoroalkyl carboxylic acids and their emerging alternatives by indigenous microorganisms in the environment.

Characterization of Two Novel Predatory Bacteria, Bacteriovorax stolpii HI3 and Myxococcus sp. MH1, Isolated from a Freshwater Pond: Prey Range, and Predatory Dynamics and Efficiency.

Predatory bacteria, which prey on other bacteria, have significant functions in microbial ecosystems and have attracted increasing attention for their biotechnological use. However, knowledge of the characteristics of wild-type environmental predatory bacteria remains limited. This study isolated two predatory bacteria, Bacteriovorax stolpii HI3 and Myxococcus sp. MH1, from a freshwater pond and characterized their predation capabilities. Determination of the prey range using 53 potential prey strains, including 52 environmental strains, revealed that B. stolpii HI3 and Myxococcus sp. MH1 could prey on a wide spectrum of Gram-negative bacteria and a broader range of bacteria, irrespective of phylogeny, in accordance with the common characteristics of Bdellovibrio and like organisms and myxobacteria, respectively. Liquid culture assays also found that although predation by B. stolpii HI3 rapidly and largely occurred, the prey bacteria regrew, possibly through plastic phenotypic resistance to predation. In contrast, predation by Myxococcus sp. MH1 occurred at relatively low efficiency but was longer lasting. The two strains exhibited slightly distinct temperature preferences but commonly preferred slightly alkaline pH. The novel findings of this study provide evidence for the coexistence of predatory bacteria with diverse predation capabilities in the natural aquatic environment.

Investigating the Responses of Human Epithelial Cells to Predatory Bacteria.

One beguiling alternative to antibiotics for treating multi-drug resistant infections are Bdellovibrio-and-like-organisms (BALOs), predatory bacteria known to attack human pathogens. Consequently, in this study, the responses from four cell lines (three human and one mouse) were characterized during an exposure to different predatory bacteria, Bdellovibrio bacteriovorus HD100, Bacteriovorus BY1 and Bacteriovorax stolpii EB1. TNF-α levels were induced in Raw 264.7 mouse macrophage cultures with each predator, but paled in comparison to those obtained with E. coli. This was true even though the latter strain was added at an 11.1-fold lower concentration (p < 0.01). Likewise, E. coli led to a significant (54%) loss in the Raw 264.7 murine macrophage viability while the predatory strains had no impact. Tests with various epithelial cells, including NuLi-1 airway, Caco2, HT29 and T84 colorectal cells, gave similar results, with E. coli inducing IL-8 production. The viabilities of the NuLi-1 and Caco-2 cells were slightly reduced (8%) when exposed to the predators, while T84 viability remained steady. In no cases did the predatory bacteria induce actin rearrangement. These results clearly demonstrate the gentle natures of predatory bacteria and their impacts on human cells.

POTENTIAL OF PREDATORY BACTERIA AS BIOCONTROL AGENTS FOR FOODBORNE AND PLANT PATHOGENS

Foodborne pathogens such as Escherichia coli O157:H7, Salmonella spp., Listeria monocytogenes are responsible for frequent occurrences of illness and mortality in humans. Several economically important plant diseases are caused by pathogenic bacteria. Economically important plant diseases and post-harvest losses and decay are also incited by plant pathogens such as Erwinia, Pectobacterium, Botrytis, and Pseudomonas spp. This paper discusses the potential of predatory bacteria for biocontrol of foodborne and plant pathogens with emphasis on the deltaproteobacteria group. Deltaproteobacteria (Bdellovibrio and Bacteriovorax sp. as well as Daptobacter, and Myxobacteria) are Gram-negative predatory microorganisms which prey mainly on Gram-negative bacteria and occur in diverse soil, marine and fresh water ecologies. The predation of Gram-negative bacteria by Bdellovibrio and Bdellovibrio-like organisms (BALOs) may enhance their potential application for biocontrol of foodborne and plant pathogens. The difficulty in controlling foodborne and plant pathogenic bacteria is due to the paucity of available bactericidal chemicals and regulations limiting the use of antibiotics in pathogen control. In this review, the types and mechanisms of predation by predatory bacteria (PB) are discussed with respect to potential utilization as biocontrol agents. Their diversity and relationships is highlighted by genomic research. Potential for biocontrol of foodborne and plant pathogens by PB is discussed in terms of attributes and limitations. Although research in animal systems (Salmonella/chicken) indicates significant biocontrol potential when PB are applied as probiotics, their application to foodborne and plant pathogens are limited. With increased research in metagenomics on PB effects on animal cells /tissues, a better understanding of regulation of cellular metabolism and their association with growth of PB and prey degradation will ultimately enhance the utility of PB as biocontrol agents of foodborne and plant pathogens.

Bacterial Predators Possess Unique Membrane Lipid Structures

Bdellovibrio-and-like organisms (BALO) are a phylogenetically diverse group of predatory prokaryotes that consists of the two families Bdellovibrionaceae and Bacteriovoracaceae. We investigated the phospholipid composition of the three important BALO strains Bacteriovorax stolpii (DSM 12778), Bdellovibrio bacteriovorus HD100 (DSM 50701) and Peredibacter starrii (DSM 17039). We confirmed the presence of sphingophosphonolipids in B. stolpii, while we characterized sphingophosphonolipids with a 2-amino-3-phosphonopropanate head group for the first time. In B. bacteriovorus HD100 phosphatidylthreonines were found and, thus, B. bacteriovorus is the second prokaryote investigated so far possessing this rare lipid class. In the third analyzed organism, P. starrii, we observed phosphatidylethanolamine structures with an additional N-glutamyl residue, which form the first reported class of amino acid-containing phosphatidylethanolamines.

Chemical Structure of Bacteriovorax stolpii Lipid A

Bdellovibrionales is a phylogenetically diverse group of predatory prokaryotes, which consists of the two families Bdellovibrionaceae and Bacteriovoracaceae. We describe LPS and lipid A of the type strain Bacteriovorax stolpii DSM 12778, representing the first characterized endotoxin of a Bacteriovoracaceae member. It has a smooth form LPS, which was identified by SDS-polyacrylamide gel electrophoresis. The lipid A structure was determined by combined gas chromatography-mass spectrometry, electrospray ionization mass spectrometry and NMR spectroscopy. Its backbone consists of two beta-(1-->6)-linked 2,3-diamino-2,3-dideoxy-D-glucopyranoses (GlcpN3N) carrying a pyrophosphoethanolamine at O-4' of the non-reducing sugar and a phosphate group linked to O-1 of the reducing GlcpN3N. Positions 2, 3, 2' and 3' of the two GlcpN3N are acylated with primary 3-hydroxy fatty acids and one of those carries a secondary fatty acid.

Phylogenetic relationships amongst the saltwater members of the genus Bacteriovorax using rpoB sequences and reclassification of Bacteriovorax stolpii as Bacteriolyticum stolpii gen. nov., comb. nov.

Members of the saltwater genus Bacteriovorax, formerly known as the marine Bdellovibrio, are obligate predatory bacteria that prey selectively on other Gram-negative bacteria. Previous phylogenetic analysis based on the 16S rRNA genes of saltwater Bacteriovorax isolates from environmental samples revealed 11 distinct phylogenetic clusters based on > or =96.5 % gene sequence similarity. In other micro-organisms, the gene coding for the beta-subunit of RNA polymerase (rpoB) has been shown to be more discriminating than 16S rRNA genes. In this study, rpoB sequences from Bacteriovorax isolates were analysed to determine whether the results would be consistent with those based on 16S rRNA gene sequences. A 1242 bp region of the rpoB gene from 74 saltwater Bacteriovorax strains and two freshwater isolates, Bacteriovorax stolpii Uki2T and Peredibacter starrii A3.12T, was amplified by PCR and analysed. The sequences were aligned and phylogenetic trees were constructed using a neighbour-joining algorithm. The resulting tree showed that the rpoB sequences produced smaller subdivisions of isolates, but were nevertheless consistent with the clusters determined using 16S rRNA gene sequences. Thus, the highly conserved 16S rRNA gene sequences provided good phylogenetic information and the rpoB gene sequences permitted greater differentiation in order to further subdivide phylogenetically distinct groups within the genus Bacteriovorax. Also, on the basis of the extensive diversity and large distance between the saltwater members of the genus Bacteriovorax and the freshwater/soil Bacteriovorax, a reclassification of Bacteriovorax stolpii as Bacteriolyticum stolpii gen. nov., comb. nov. is proposed. A new family, Peredibacteraceae fam. nov., is also described.

Bacteriovorax stolpii proliferation and predation without sphingophosphonolipids

Bacteriovorax stolpii strain UKi2, a facultative predator-parasite of larger Gram-negative bacteria, synthesizes distinct sphingophosphonolipids. These lipids are characterized by a direct P–C bond, the novel head group 1-hydroxy-2-aminoethylphosphonate, iso-branched long chain bases and fatty acids, and fatty acids dominated by those with α-hydroxy groups. Myriocin, an inhibitor of serine:fatty acyl CoA transferase, reversibly blocked sphingophosphonolipid synthesis in B. stolpii UKi2. However, the inhibitor did not block cell proliferation indicating that these lipids are not vital for B. stolpii UKi2 viability and growth. When mixed with Escherichia coli prey cells, control predator-parasite bacteria were effective in forming large E. coli bdelloplasts and cleared the suspension of the prey cells. Although myriocin-treated cells could attack prey cells and form bdelloplasts, their locomotory behavior was altered and fewer and smaller bdelloplasts were produced. These observations open up the possibility for a role of sphingophosphonolipids in B. stolpii UKi2 complex behavior.

Detection and Identification of Bacteriovorax stolpii UKi2 Sphingophosphonolipid Molecular Species

Bacteriovorax stolpii is a predator of larger gram-negative bacteria and lives as a parasite in the intraperiplasmic space of the host cell. This bacterium is unusual among prokaryotes in that sphingolipids comprise a large proportion of its lipids. We here report the presence of 18 molecular species of B. stolpii UKi2 sphingophosphonolipids (SPNLs). (31)P NMR spectroscopy and analysis of P(i) released by a differential hydrolysis protocol confirmed the phosphonyl nature of these lipids. The SPNLs were dominated by those with 1-hydroxy-2-aminoethane phosphonate (hydroxy-aminoethylphosphonate) polar head groups; aminoethylphosphonate was also detected in minor SPNL components. The long-chain bases (LCBs) were dominated by C(17) iso-branched phytosphingosine; C(17) iso-branched dihydrosphingosine was also present in some SPNLs. The N-linked fatty acids were predominantly iso-branched and most contained an alpha-hydroxy group (C(15) alpha-hydroxy fatty acid was the major fatty acid). Minor molecular species containing nonhydroxy fatty acids were also detected. The definitive iso-structures of the predominant fatty acids and LCBs present in the B. stolpii SPNLs were established using (13)C and (3)H nuclear magnetic resonance spectroscopy; less than 20% were unbranched. Detection and analyses of intact compounds by MS-MS were performed by a hybrid quadrupole time-of-flight (Q-TOF-II) MS equipped with an electrospray ionization source. Analyses of peracetylated derivatives verified the structural assignments of these lipids.

Characterization of outer membrane protein fractions of Bdellovibrionales

Bdellovibrio-and-like organisms (BALOs) are predatory bacteria that prey upon Gram-negative bacteria and are taxonomically subsumed in the order Bdellovibrionales. Despite their unique lifestyle, these bacteria show remarkable genotypic diversities. The outer membrane of the predators is likely to play an important role during the recognition and invasion stage, as well as in the intraperiplasmic growth phase. In this study, the outer membrane protein fractions of type strains of Bdellovibrio, Bacteriovorax and Peredibacter were investigated, revealing the presence of outer membrane proteins (Omps) similar to the major Omps of Bdellovibrio bacteriovorus. The primary structures of these Omps of Bdellovibrio sp. W, Bacteriovorax stolpii and Peredibacter starrii were elucidated by a combined mass spectrometric–reverse genetic approach. The similarity between the analyzed Omps of the investigated BALOs ranges from 32% to 89% showing conserved amino acid regions in their primary structure.

Taxonomic Studies of Predatory Bdellovibrios Based on 16S rRNA Analysis, Ribotyping and the hit Locus and Characterization of Isolates from the Gut of Animals

The aim of our study was to obtain data for the molecular characterization of bdellovibrio bacteria, which were recently split into the genus Bdellovibrio and the newly designated genus Bacteriovorax. We determined the 16S rDNA sequences of five reference strains and performed a phylogenetic analysis including published 16S rRNA sequences of bdellovibrios. A comparison of the secondary structure showed significant differences in two regions of the 16S rRNAs of the species Bdellovibrio bacteriovorus, Bacteriovorax starrii, and Bacteriovorax stolpii. In addition, ribotyping techniques gave specific hybridization patterns and revealed that two rRNA operons are present in the investigated strains. A hybridization probe derived from the genetic locus hit, associated with the host independent (HI) phenotype of B. bacteriovorus, was found to be specific for this species. Sequence comparison of the hit locus revealed few base pair changes between host independent (HI) and host dependent (HD) strains. Ribotyping and hybridization experiments using the hit probe were applied to characterize bdellovibrio strains isolated from the gut of animals and humans and one isolate from sewage.

A proposal for the reclassification of Bdellovibrio stolpii and Bdellovibrio starrii into a new genus, Bacteriovorax gen. nov. as Bacteriovorax stolpii comb. nov. and Bacteriovorax starrii comb. nov., respectively.

Bdellovibrios are unique bacteria with the ability to prey upon a wide variety of susceptible Gram-negative bacteria. Micro-organisms exhibiting this trait have been included in the genus Bdellovibrio despite their isolation from diverse habitats and relatively unstudied taxonomic relatedness. In this study, 16S rDNA sequences were compared from known terrestrial Bdellovibrio species, Bdellovibrio bacteriovorus 100T, Bdellovibrio stolpii Uki2T and Bdellovibrio starrii A3.12T in order to study their phylogenetic relationship. The two sequences from B. stolpii Uki2T and B. starrii A3.12T were 90.0% similar to each other but exhibited only 81.7% and 81.2% similarity, respectively to B. bacteriovorus 100T. Phylogenetic analysis indicated that B. bacteriovorus 100T clustered in a separate clade from B. starrii A3.12T and B. stolpii Uki2T, demonstrating only a distant relationship between B. bacteriovorus 100T and the other two recognized type species. DNA-DNA hybridization experiments also demonstrated <4% hybridization between these three species. On the basis of the results obtained from the phylogenetic analysis and DNA-DNA hybridization studies, it is proposed that B. stolpii Uki2T and B. starrii A3.12T should be transferred to a new genus, Bacteriovorax gen. nov. as Bacteriovorax stolpii comb. nov. and Bacteriovorax starrii comb. nov., respectively. It is also proposed that the type species for the new genus Bacteriovorax should be Bacteriovorax stolpii comb. nov.