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Journal of Laboratory Physicians Jul 2013Paenibacilli, the Gram positive, aerobic spore bearing bacilli are found normally in the environment. Though these organisms were not known to cause human disease, until...
Paenibacilli, the Gram positive, aerobic spore bearing bacilli are found normally in the environment. Though these organisms were not known to cause human disease, until recently; few species of this genus have been reported to cause infections in humans. We report here, a case of urinary tract infection in a 60-year-old chronic kidney disease patient due to this rare bacterium. The patient presented with complains of fever, dysuria, and flank pain. Routine and microscopic examination of urine revealed no abnormality except plenty of pus cells and albumin (1+). Bacterial culture showed significant bacteruria and the isolated bacteria was identified to be Paenibacillus alvei based on standard biochemical reactions.
PubMed: 24701110
DOI: 10.4103/0974-2727.119872 -
World Journal of Microbiology &... Apr 2014A wild-type, Gram-positive, rod-shaped, endospore-forming and motile bacteria has been isolated from palm oil mill sludge in Malaysia. Molecular identification using 16S...
A wild-type, Gram-positive, rod-shaped, endospore-forming and motile bacteria has been isolated from palm oil mill sludge in Malaysia. Molecular identification using 16S rRNA gene sequence analysis indicated that the bacteria belonged to genus Paenibacillus. With 97 % similarity to P. alvei (AUG6), the isolate was designated as P. alvei AN5. An antimicrobial compound was extracted from P. alvei AN5-pelleted cells using 95 % methanol and was then lyophilized. Precipitates were re-suspended in phosphate buffered saline (PBS), producing an antimicrobial crude extract (ACE). The ACE showed antimicrobial activity against Salmonella enteritidis ATCC 13076, Escherichia coli ATCC 29522, Bacillus cereus ATCC 14579 and Lactobacillus plantarum ATCC 8014. By using SP-Sepharose cation exchange chromatography, Sephadex G-25 gel filtration and Tricine SDS-PAGE, the ACE was purified, which produced a ~2-kDa active band. SDS-PAGE and infrared (IR) spectroscopy indicated the proteinaceous nature of the antimicrobial compound in the ACE, and liquid chromatography electrospray ionization mass spectroscopy and de novo sequencing using an automatic, Q-TOF premier system detected a peptide with the amino acid sequence F-C-K-S-L-P-L-P-L-S-V-K (1,330.7789 Da). This novel peptide was designated as AN5-2. The antimicrobial peptide exhibited stability from pH 3 to 12 and maintained its activity after being heated to 90 °C. It also remained active after incubation with denaturants (urea, SDS and EDTA).
Topics: Antimicrobial Cationic Peptides; Chromatography; DNA, Bacterial; DNA, Ribosomal; Electrophoresis, Polyacrylamide Gel; Environmental Microbiology; Gram-Negative Bacteria; Gram-Positive Bacteria; Hydrogen-Ion Concentration; Malaysia; Microbial Sensitivity Tests; Molecular Sequence Data; Molecular Weight; Paenibacillus; Protein Stability; RNA, Ribosomal, 16S; Sequence Analysis, DNA; Spectrum Analysis; Temperature
PubMed: 24272828
DOI: 10.1007/s11274-013-1558-z -
PloS One 2013Swarming and biofilm formation have been studied for a variety of bacteria. While this is well investigated for Gram-negative bacteria, less is known about Gram-positive...
BACKGROUND
Swarming and biofilm formation have been studied for a variety of bacteria. While this is well investigated for Gram-negative bacteria, less is known about Gram-positive bacteria, including Paenibacillus alvei, a secondary invader of diseased honeybee colonies infected with Melissococcus pluton, the causative agent of European foulbrood (EFB).
METHODOLOGY
Paenibacillus alvei CCM 2051(T) is a Gram-positive bacterium which was recently shown to employ S-layer homology (SLH) domains as cell wall targeting modules to display proteins on its cell surface. This study deals with the newly identified 1335-amino acid protein SlhA from P. alvei which carries at the C‑terminus three consecutive SLH-motifs containing the predicted binding sequences SRGE, VRQD, and LRGD instead of the common TRAE motif. Based on the proof of cell surface location of SlhA by fluorescence microscopy using a SlhA-GFP chimera, the binding mechanism was investigated in an in vitro assay. To unravel a putative function of the SlhA protein, a knockout mutant was constructed. Experimental data indicated that one SLH domain is sufficient for anchoring of SlhA to the cell surface, and the SLH domains of SlhA recognize both the peptidoglycan and the secondary cell wall polymer in vitro. This is in agreement with previous data from the S-layer protein SpaA, pinpointing a wider utilization of that mechanism for cell surface display of proteins in P. alvei. Compared to the wild-type bacterium ΔslhA revealed changed colony morphology, loss of swarming motility and impaired biofilm formation. The phenotype was similar to that of the flagella knockout Δhag, possibly due to reduced EPS production influencing the functionality of the flagella of ΔslhA.
CONCLUSION
This study demonstrates the involvement of the SLH domain-containing protein SlhA in swarming and biofilm formation of P. alvei CCM 2051(T).
Topics: Amino Acid Motifs; Bacterial Proteins; Biofilms; Flagella; Paenibacillus; Protein Structure, Tertiary
PubMed: 24058714
DOI: 10.1371/journal.pone.0076566 -
Genome Announcements Aug 2013Here, we report draft genomes of Paenibacillus alvei strains A6-6i and TS-15, which were isolated, respectively, from plant material and soil in the Virginia Eastern...
Here, we report draft genomes of Paenibacillus alvei strains A6-6i and TS-15, which were isolated, respectively, from plant material and soil in the Virginia Eastern Shore (VES) tomato growing area. An array of genes related to antimicrobial biosynthetic pathways have been identified with whole-genome analyses of these strains.
PubMed: 23990585
DOI: 10.1128/genomeA.00673-13 -
Colloids and Surfaces. B, Biointerfaces Sep 2013The demulsifying performance of Paenibacillus alvei ARN63 (P. alvei), as a biodemulsifier-producing bacterium, for breaking water-in-heavy crude oil emulsion has been...
The demulsifying performance of Paenibacillus alvei ARN63 (P. alvei), as a biodemulsifier-producing bacterium, for breaking water-in-heavy crude oil emulsion has been investigated. The produced lipopeptide biodemulsifier showed the potential to be used in the petroleum industry as an environmentally friendly and non-toxic material. To optimize the biodemulsifier production, the impacts of parameters such as temperature, pH, carbon source and carbon concentration at a constant agitation speed of 180 rpm and with ammonium sulfate as the sole nitrogen source (1.0 g/l) were studied in detail. Several normal paraffin compounds, vegetable oils and motor oil revealed the ability to be used as the carbon source for synthesis of biodemulsifier. The best biodemulsifier production was obtained employing motor oil as the carbon source with a concentration of 42.5 g/l at 37°C and pH 7.0 after 72 h of incubation. Under these conditions, the surface tension of the medium reduced from 58 mN/m to 24.7 mN/m and the biodemulsifier yield reached a value of 2.1 g/l. The demulsification ratio approached 77% and the produced biodemulsifier by P. alvei strain effectively broke water-in-heavy crude oil emulsion. According to biodemulsifier production and growth time course profiles, the biosynthesis was growth associated. Besides, the produced biodemulsifier had good stability during exposure to salinities up to 20%, temperatures up to 80°C and a wide pH range of 2-12.
Topics: Emulsions; Hydrogen-Ion Concentration; Lipopeptides; Molecular Conformation; Paenibacillus; Petroleum; Temperature
PubMed: 23660310
DOI: 10.1016/j.colsurfb.2013.03.029 -
Journal of Industrial Microbiology &... Jun 2013An antimicrobial substance produced by the Paenibacillus alvei strain AN5 was detected in fermentation broth. Subsequently, cell-free culture supernatant (CFCS) was...
An antimicrobial substance produced by the Paenibacillus alvei strain AN5 was detected in fermentation broth. Subsequently, cell-free culture supernatant (CFCS) was obtained by medium centrifugation and filtration, and its antimicrobial activity was tested. This showed a broad inhibitory spectrum against both Gram-positive and -negative bacterial strains. The CFCS was then purified and subjected to SDS-PAGE and infrared spectroscopy, which indicated the proteinaceous nature of the antimicrobial compound. Some de novo sequencing using an automatic Q-TOF premier system determined the amino acid sequence of the purified antimicrobial peptide as Y-S-K-S-L-P-L-S-V-L-N-P (1,316 Da). The novel peptide was designated as peptide AN5-1. Its mode of action was bactericidal, inducing cell lysis in E. coli ATCC 29522 and S. aureus, and non-cell lysis in both S. marcescens and B. cereus ATCC 14579. Peptide AN5-1 displayed stability at a wide range of pH values (2-12) and remained active after exposure to high temperatures (100 °C). It also maintained its antimicrobial activity after incubation with chemicals such as SDS, urea and EDTA.
Topics: Amino Acid Sequence; Anti-Bacterial Agents; Bacteria; Bacterial Proteins; Culture Media, Conditioned; Fermentation; Microbial Sensitivity Tests; Paenibacillus; Peptides; Spectroscopy, Fourier Transform Infrared
PubMed: 23508455
DOI: 10.1007/s10295-013-1259-5 -
Probiotics and Antimicrobial Proteins Mar 2013The important viscosity of the respiratory tract mucus of Cystic fibrosis (CF) patients impairs the mucociliary transport system and allows the growth of numerous...
The important viscosity of the respiratory tract mucus of Cystic fibrosis (CF) patients impairs the mucociliary transport system and allows the growth of numerous micro-organisms. Among them, Pseudomonas aeruginosa and Staphylococcus aureus are known to be responsible for pulmonary infections. We imagined that CF microflora could also harbour micro-organisms naturally equipped to compete with these pathogens. A method was developed to recover these antibiotic-producing strains within 20 CF sputum. Using this approach, we have isolated an unusual Gram-positive bacterium identified as Paenibacillus alvei by Api galleries and 16S rRNA gene sequence analysis. This strain has inhibited the growth of P. aeruginosa, S. aureus and Klebsiella pneumoniae, in co-cultures. A liquid mineral medium named MODT50 was designed and optimised for the production and the recovery of the antimicrobial compounds. The supernatant has inhibited the growth of all Gram-positive strains tested, even Methicillin-resistant S. aureus. One antimicrobial compound with a peptide structure (mainly active against S. aureus, Micrococcus luteus, and Pseudomonas stutzeri) has been purified and characterised by liquid chromatography-mass spectrometry. The new active molecule (m/z 786.6) named depsipeptide L possesses a 15-guanidino-3-hydroxypentadecanoic acid side chain (m/z 298) linked on a cyclic part of four amino acids residues (Ser, two Leu/Ile, Arg). This work reports for the first time the production of such a molecule by a P. alvei strain in a mineral medium. The CF lung microflora might represent a valuable source for the discovery of new antimicrobial-producing strains.
PubMed: 26782601
DOI: 10.1007/s12602-012-9121-z -
International Journal of Biochemistry... 2012Prospection of cellulose-degrading bacteria in natural environments allows the identification of novel cellulases and hemicellulases that could be useful in...
Prospection of cellulose-degrading bacteria in natural environments allows the identification of novel cellulases and hemicellulases that could be useful in second-generation bioethanol production. In this work, cellulolytic bacteria were isolated from decaying native forest soils by enrichment on cellulose as sole carbon source. There was a predominance of Gram positive isolates that belonged to the phyla Proteobacteria and Firmicutes. Many primary isolates with cellulolytic activity were not pure cultures. From these consortia, isolation of pure constituents was attempted in order to test the hypothesis whether microbial consortia are needed for full degradation of complex substrates. Two isolates, CB1-2-A-5 and VG-4-A-2, were obtained as the pure constituents of CB1-2 and VG-4 consortia, respectively. Based on 16S RNA sequence, they could be classified as Variovorax paradoxus and Paenibacillus alvei. Noteworthy, only VG-4 consortium showed measurable xylan degrading capacity and signs of filter paper degradation. However, no xylan or filter paper degrading capacities were observed for the pure cultures isolated from it, suggesting that other members of this consortium were necessary for these hydrolyzing activities. Our results indicated that Paenibacillus sp. and Variovorax sp. as well as VG-4 consortium, might be a useful source of hydrolytic enzymes. Moreover, although Variovorax sp. had been previously identified in metagenomic studies of cellulolytic communities, this is the first report on the isolation and characterization of this microorganism as a cellulolytic genus.
PubMed: 23301200
DOI: No ID Found -
Journal of Bacteriology Feb 2013Paenibacillus alvei CCM 2051(T) cells are decorated with a two-dimensional (2D) crystalline array comprised of the glycosylated S-layer protein SpaA. At its N terminus,...
Paenibacillus alvei CCM 2051(T) cells are decorated with a two-dimensional (2D) crystalline array comprised of the glycosylated S-layer protein SpaA. At its N terminus, SpaA possesses three consecutive surface layer (S-layer) homology (SLH) domains containing the amino acid motif TRAE, known to play a key role in cell wall binding, as well as the TVEE and TRAQ variations thereof. SpaA is predicted to be anchored to the cell wall by interaction of the SLH domains with a peptidoglycan (PG)-associated, nonclassical, pyruvylated secondary cell wall polymer (SCWP). In this study, we have analyzed the role of the three predicted binding motifs within the SLH domains by mutating them into TAAA motifs, either individually, pairwise, or all of them. Effects were visualized in vivo by homologous expression of chimeras made of the mutated S-layer proteins and enhanced green fluorescent protein and in an in vitro binding assay using His-tagged SpaA variants and native PG-containing cell wall sacculi that either contained SCWP or were deprived of it. Experimental data indicated that (i) the TRAE, TVEE, and TRAQ motifs are critical for the binding function of SLH domains, (ii) two functional motifs are sufficient for cell wall binding, regardless of the domain location, (iii) SLH domains have a dual-recognition function for the SCWP and the PG, and (iv) cell wall anchoring is not necessary for SpaA glycosylation. Additionally, we showed that the SLH domains of SpaA are sufficient for in vivo cell surface display of foreign proteins at the cell surface of P. alvei.
Topics: Amino Acid Motifs; Amino Acid Sequence; Bacterial Proteins; Cell Membrane; Cell Wall; Gene Expression Regulation, Bacterial; Glycoproteins; Glycosylation; Membrane Glycoproteins; Paenibacillus; Plasmids; Protein Structure, Tertiary
PubMed: 23204458
DOI: 10.1128/JB.01487-12 -
Advances in Microbiology Dec 2012Surface (S)-layer proteins are model systems for studying protein glycosylation in bacteria and simultaneously hold promises for the design of novel,...
Surface (S)-layer proteins are model systems for studying protein glycosylation in bacteria and simultaneously hold promises for the design of novel, glyco-functionalized modules for nanobiotechnology due to their 2D self-assembly capability. Understanding the mechanism governing S-layer glycan biosynthesis in the Gram-positive bacterium CCM 2051 is necessary for the tailored glyco-functionalization of its S-layer. Here, the putative oligosaccharyl:S-layer protein transferase WsfB from the S-layer glycosylation gene locus is characterized. The enzyme is proposed to catalyze the final step of the glycosylation pathway, transferring the elongated S-layer glycan onto distinct tyrosine -glycosylation sites. Genetic knock-out of WsfB is shown to abolish glycosylation of the S-layer protein SpaA but not that of other glycoproteins present in CCM 2051, confining its role to the S-layer glycosylation pathway. A transmembrane topology model of the 781-amino acid WsfB protein is inferred from activity measurements of green fluorescent protein and phosphatase A fused to defined truncations of WsfB. This model shows an overall number of 13 membrane spanning helices with the Wzy_C domain characteristic of -oligosaccharyl:protein transferases (-OTases) located in a central extra-cytoplasmic loop, which both compares well to the topology of OTases from Gram-negative bacteria. Mutations in the Wzy_C motif resulted in loss of WsfB function evidenced in reconstitution experiments in ΔWsfB cells. Attempts to use WsfB for transferring heterologous oligosaccharides to its native S-layer target protein in CWG702 and SL3749, which should provide lipid-linked oligosaccharide substrates mimicking to some extent those of the natural host, were not successful, possibly due to the stringent function of WsfB. Concluding, WsfB has all features of a bacterial -OTase, making it the most probable candidate for the oligosaccharyl:S-layer protein transferase of , and a promising candidate for the first -OTase reported in Gram-positives.
PubMed: 25893145
DOI: 10.4236/aim.2012.24069