Genome mining for the search and discovery of bioactive compounds: the Streptomyces paradigm.

The need for new antimicrobials is indisputable. The flight from natural products in drug discovery was unfortunate; however, the revolution that is genome mining, enabled by the explosion in sequencing technology, is a cause for hope. Nevertheless, renewed search and discovery is still a challenge. We explore novel metabolite diversity and the challenges in Streptomyces. Estimating the extent of novel bioactive metabolites remaining to be discovered is an important driver for future investment. Frequent re-discovery of known natural products was a major factor in big pharma exiting search and discovery, and remains a reality. We explore whether this is due to exhaustive isolation and frequent lateral gene transfer. Analysing all biosynthetic gene clusters across all genomes is challenging. Therefore, representative examples of the patterns of secondary metabolite diversity suggest that re-discovery is linked to frequent expression in frequently isolated (and frequently misidentified) strains. Lateral gene transfer of complete biosynthetic clusters is less frequent than might be perceived but frequent gene exchange implies a massive combinatorial biosynthesis experiment. Genome sequencing emphasises rare expression of many secondary metabolite gene clusters and diversification at the finest levels of phylogenetic discrimination. In addition, we are only just beginning to unravel the impact of ecology. The hidden diversity suggests that cluster cloning and heterologous expression in microbial cell factories will explore this diversity more effectively.

Identification and characterisation of Staphylococcus aureus on low cost screen printed carbon electrodes using impedance spectroscopy.

Staphylococcus aureus infections are a cause of significant morbidity and mortality, in addition to representing a considerable economic burden. The aim of this study was to explore a low cost screen printed electrode as a sensor for the detection of S. aureus using impedance spectroscopy. S. aureus was incubated in chambers containing the electrodes and the results analysed using a novel normalisation approach. These results show that it is possible to detect the presence of S. aureus in LB media after 30 min incubation of a 1% growth culture, in addition to being able to see immediate cell concentration dependant changes in 0.9% NaCl. These observations imply that a number of electrochemical mechanisms cause a change in the impedance as a result of the presence of S. aureus, including adsorption to the electrode surface and the metabolism of the bacteria during growth. The study suggests that this detection approach would be useful in a number of clinical scenarios where S. aureus leads to difficult to treat infections.

Vitamin D insufficiency in the first 6 months of infancy and challenge-proven IgE-mediated food allergy at 1 year of age: a case-cohort study.

BACKGROUND: Ecological evidence suggests vitamin D insufficiency (VDI) due to lower ambient ultraviolet radiation (UVR) exposure may be a risk factor for IgE-mediated food allergy. However, there are no studies relating directly measured VDI during early infancy to subsequent challenge-proven food allergy. OBJECTIVE: To prospectively investigate the association between VDI during infancy and challenge-proven food allergy at 1 year. METHODS: In a birth cohort (n = 1074), we used a case-cohort design to compare 25-hydroxyvitamin D3 (25(OH)D3 ) levels among infants with food allergy vs a random subcohort (n = 274). The primary exposures were VDI (25(OH)D3 <50 nM) at birth and 6 months of age. Ambient UVR and time in the sun were combined to estimate UVR exposure dose. IgE-mediated food allergy status at 1 year was determined by formal challenge. Binomial regression was used to examine associations between VDI, UVR exposure dose and food allergy and investigate potential confounding. RESULTS: Within the random subcohort, VDI was present in 45% (105/233) of newborns and 24% (55/227) of infants at 6 months. Food allergy prevalence at 1 year was 7.7% (61/786), and 6.5% (53/808) were egg-allergic. There was no evidence of an association between VDI at either birth (aRR 1.25, 95% CI 0.70-2.22) or 6 months (aRR 0.93, 95% CI 0.41-2.14) and food allergy at 1 year. CONCLUSIONS: There was no evidence that VDI during the first 6 months of infancy is a risk factor for food allergy at 1 year of age. These findings primarily relate to egg allergy, and larger studies are required.

Bioprocess intensification of antibiotic production by Streptomyces coelicolor A3(2) in micro-porous culture.

A novel functionalized micro-porous matrix was developed with well-controlled physicochemical proprieties such as pore size and surface chemistry. The matrix was used as a solid support in the growth of "Streptomyces coelicolor" A3(2) to enhance the production of antibiotics. The results shown support a higher production of prodigiosin and actinorhodin with overall production increase of 2-5 and 6-17, respectively, compared to conventional submerged liquid culture, offering a potential improvement in volumetric productivity. Scanning Electron Microscopy was used to evaluate pore size as well as bacterial adhesion, penetration, proliferation and migration within the micro-porous matrix.

Granulocyte colony-stimulating factor receptor signalling via Janus kinase 2/signal transducer and activator of transcription 3 in ovarian cancer.

BACKGROUND: Ovarian cancer remains a major cause of cancer mortality in women, with only limited understanding of disease aetiology at the molecular level. Granulocyte colony-stimulating factor (G-CSF) is a key regulator of both normal and emergency haematopoiesis, and is used clinically to aid haematopoietic recovery following ablative therapies for a variety of solid tumours including ovarian cancer. METHODS: The expression of G-CSF and its receptor, G-CSFR, was examined in primary ovarian cancer samples and a panel of ovarian cancer cell lines, and the effects of G-CSF treatment on proliferation, migration and survival were determined. RESULTS: G-CSFR was predominantly expressed in high-grade serous ovarian epithelial tumour samples and a subset of ovarian cancer cell lines. Stimulation of G-CSFR-expressing ovarian epithelial cancer cells with G-CSF led to increased migration and survival, including against chemotherapy-induced apoptosis. The effects of G-CSF were mediated by signalling via the downstream JAK2/STAT3 pathway. CONCLUSION: This study suggests that G-CSF has the potential to impact on ovarian cancer pathogenesis, and that G-CSFR expression status should be considered in determining appropriate therapy.

Zebrafish as a genetic model in pre-clinical drug testing and screening.

The traditional drug discovery pipeline for the identification and development of compounds that selectively target specific molecules to ameliorate disease remains a major focus for medical research. However, the zebrafish is increasingly providing alternative strategies for various components of this pipeline. Zebrafish and their embryos are small, easily accessible and relatively low cost, making them applicable to high-throughput, small molecule screening. Zebrafish can also be manipulated by a range of forward and reverse genetics techniques to facilitate gene discovery and functional studies. Moreover, their physiological and developmental complexity provides accurate models of human disease to underpin mechanism of action and in vivo validation studies. Finally, several of these biological characteristics make zebrafish eminently suitable for toxicity testing, including eco-toxicology. Here we review the application of zebrafish to preclinical drug development and toxicity testing, including recent advances in mutant generation, drug screening and toxicology that serve to further enhance the capabilities of this valuable model organism in drug discovery.

Phylogenetic study of the species within the family Streptomycetaceae.

Species of the genus Streptomyces, which constitute the vast majority of taxa within the family Streptomycetaceae, are a predominant component of the microbial population in soils throughout the world and have been the subject of extensive isolation and screening efforts over the years because they are a major source of commercially and medically important secondary metabolites. Taxonomic characterization of Streptomyces strains has been a challenge due to the large number of described species, greater than any other microbial genus, resulting from academic and industrial activities. The methods used for characterization have evolved through several phases over the years from those based largely on morphological observations, to subsequent classifications based on numerical taxonomic analyses of standardized sets of phenotypic characters and, most recently, to the use of molecular phylogenetic analyses of gene sequences. The present phylogenetic study examines almost all described species (615 taxa) within the family Streptomycetaceae based on 16S rRNA gene sequences and illustrates the species diversity within this family, which is observed to contain 130 statistically supported clades, as well as many unsupported and single member clusters. Many of the observed clades are consistent with earlier morphological and numerical taxonomic studies, but it is apparent that insufficient variation is present in the 16S rRNA gene sequence within the species of this family to permit bootstrap-supported resolution of relationships between many of the individual clusters.

Cross talk of signals between EGFR and IL-6R through JAK2/STAT3 mediate epithelial-mesenchymal transition in ovarian carcinomas.

Epidermal growth factor receptor (EGFR) is overexpressed in ovarian carcinomas, with direct or indirect activation of EGFR able to trigger tumour growth. We demonstrate significant activation of both signal transducer and activator of transcription (STAT)3 and its upstream activator Janus kinase (JAK)2, in high-grade ovarian carcinomas compared with normal ovaries and benign tumours. The association between STAT3 activation and migratory phenotype of ovarian cancer cells was investigated by EGF-induced epithelial-mesenchymal transition (EMT) in OVCA 433 and SKOV3 ovarian cancer cell lines. Ligand activation of EGFR induced a fibroblast-like morphology and migratory phenotype, consistent with the upregulation of mesenchyme-associated N-cadherin, vimentin and nuclear translocation of beta-catenin. This occurred concomitantly with activation of the downstream JAK2/STAT3 pathway. Both cell lines expressed interleukin-6 receptor (IL-6R), and treatment with EGF within 1 h resulted in a several-fold enhancement of mRNA expression of IL-6. Consistent with that, EGF treatment of both OVCA 433 and SKOV3 cell lines resulted in enhanced IL-6 production in the serum-free medium. Exogenous addition of IL-6 to OVCA 433 cells stimulated STAT3 activation and enhanced migration. Blocking antibodies against IL-6R inhibited IL-6 production and EGF- and IL-6-induced migration. Specific inhibition of STAT3 activation by JAK2-specific inhibitor AG490 blocked STAT3 phosphorylation, cell motility, induction of N-cadherin and vimentin expression and IL6 production. These data suggest that the activated status of STAT3 in high-grade ovarian carcinomas may occur directly through activation of EGFR or IL-6R or indirectly through induction of IL-6R signalling. Such activation of STAT3 suggests a rationale for a combination of anti-STAT3 and EGFR/IL-6R therapy to suppress the peritoneal spread of ovarian cancer.

Multiple pathways contribute to the hyperproliferative responses from truncated granulocyte colony-stimulating factor receptors.

Mutations in the granulocyte colony-stimulating factor receptor (G-CSF-R) gene leading to a truncated protein have been identified in a cohort of neutropenia patients highly predisposed to acute myeloid leukemia. Such mutations act in a dominant manner resulting in hyperproliferation but impaired differentiation in response to G-CSF. This is due, at least in part, to defective internalization and loss of binding sites for several negative regulators, leading to sustained receptor activation. However, those signaling pathways responsible for mediating the hyperproliferative function have remained unclear. In this study, analysis of an additional G-CSF-R mutant confirmed the importance of residues downstream of Box 2 as important contributors to the sustained proliferation. However, maximal proliferation correlated with the ability to robustly activate signal transducer and activator of transcription (STAT) 5 in a sustained manner, whereas co-expression of dominant-negative STAT5, but not dominant-negative STAT3, was able to inhibit G-CSF-stimulated proliferation from a truncated receptor. Furthermore, a Janus kinase (JAK) inhibitor also strongly reduced the proliferative response, whereas inhibitors of mitogen-activated protein kinase/extracellular signal-regulated kinase kinase (MEK) or phosphatidylinositol (PI) 3-kinase reduced proliferation to a lesser degree. These data suggest that sustained JAK2/STAT5 activation is a major contributor to the hyperproliferative function of truncated G-CSF receptors, with pathways involving MEK and PI 3-kinase playing a reduced role.

Application of oligonucleotide probes for the detection of Thiothrix spp. in activated sludge plants treating paper and board mill wastes.

Filamentous bacteria belonging to the genus Thiothrix were detected in activated sludge samples using the fluorescent in situ hybridisation (FISH) technique. A 16S rRNA-targeted oligonucleotide probe was developed for the detection of members of the T. fructosivorans group, and the performance of probe TNI for the detection of Thiothrix nivea group was enhanced by using an unlabeled competitor. A set of 5 probes covering all phylogenetic groups of Thiothrix were used to examine samples taken from selected activated sludge plants treating paper and board mill wastes. Members of the T. eikelboomii group formed the predominant filamentous bacterial population in plants experiencing poor sludge settleability, whereas members of the T. nivea group were commonly found but not dominantly in the remaining plants. Members of the T. fructosivorans group were not detected at any significant level in any of the samples. The distribution of the main Thiothrix types remained unchanged throughout the investigation period. It was evident that mixed populations of Thiothrix spp. were present in all activated sludge samples investigated, the observed differences were in the relative abundance of the various groups. These findings were supported by the results obtained using conventional microscopy.

Morphologic and functional characterization of granulocytes and macrophages in embryonic and adult zebrafish.

The zebrafish is a useful model organism for developmental and genetic studies. The morphology and function of zebrafish myeloid cells were characterized. Adult zebrafish contain 2 distinct granulocytes, a heterophil and a rarer eosinophil, both of which circulate and are generated in the kidney, the adult hematopoietic organ. Heterophils show strong histochemical myeloperoxidasic activity, although weaker peroxidase activity was observed under some conditions in eosinophils and erythrocytes. Embryonic zebrafish have circulating immature heterophils by 48 hours after fertilization (hpf). A zebrafish myeloperoxidase homologue (myeloid-specific peroxidase; mpx) was isolated. Phylogenetic analysis suggested it represented a gene ancestral to the mammalian myeloperoxidase gene family. It was expressed in adult granulocytes and in embryos from 18 hpf, first diffusely in the axial intermediate cell mass and then discretely in a dispersed cell population. Comparison of hemoglobinized cell distribution, mpx gene expression, and myeloperoxidase histochemistry in wild-type and mutant embryos confirmed that the latter reliably identified a population of myeloid cells. Studies in embryos after tail transection demonstrated that mpx- and peroxidase-expressing cells were mobile and localized to a site of inflammation, indicating functional capability of these embryonic granulocytes. Embryonic macrophages removed carbon particles from the circulation by phagocytosis. Collectively, these observations have demonstrated the early onset of zebrafish granulopoiesis, have proved that granulocytes circulate by 48 hpf, and have demonstrated the functional activity of embryonic granulocytes and macrophages. These observations will facilitate the application of this genetically tractable organism to the study of myelopoiesis.

Copper/zinc superoxide dismutase is phosphorylated and modulated specifically by granulocyte-colony stimulating factor in myeloid cells.

Using two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (2-D SDS-PAGE) of 32P-labeled cytosolic and membrane extracts, we identified a 21.5 kDa phosphoprotein with an isoelectric point of 6.0 in NFS-60 cells that was phosphorylated maximally at 15 min by treatment with granulocyte-colony stimulating factor (G-CSF) but not with interlevkin-3 (IL-3) or colony-stimulating factor-1 (macrophage-colony stimulating factor (CSF-1 (M-CSF)). The phosphorylation of this protein, designated 21.5/6.0, was unaffected by a series of antiproliferative agents [32]. These findings suggested that the 21.5/6.0 phosphoprotein may be involved in specific G-CSF-mediated biological responses such as activation and/or differentiation. We sought to characterize this 21.5/6.0 by a novel combination of 2-D SDS-PAGE and hydroxyapatite (HTP)-chromatography. Amino acid sequence determination of 21.5/6.0 revealed it to share a high level of homology with copper/zinc superoxide dismutase (Cu/Zn-SOD), indicating that a Cu/Zn-SOD is phosphorylated following treatment with G-CSF. This is the first report of the phosphorylation and possible involvement of Cu/Zn-SOD protein in granulocyte activation/differentiation events. In addition, Cu/Zn-SOD levels and activity were diminished by G-CSF but not IL-3 treatment. This new protocol combining 2-D SDS-PAGE and HTP-chromatography allows the characterization of low abundance phosphoproteins involved in the cellular responses to G-CSF and presumably to other cytokines/growth factors.

Corynebacterium mooreparkense sp. nov. and Corynebacterium casei sp. nov., isolated from the surface of a smear-ripened cheese.

Ten isolates each of two different bacterial species isolated from the surface of a smear-ripened cheese were found to exhibit many characteristics of the genus Corynebacterium. The isolates were Gram-positive, catalase-positive, non-spore-forming rods that did not undergo a rod/coccus transformation when grown on complex media. Chemotaxonomic investigation revealed that the strains belonged unambiguously to the genus Corynebacterium. Their cell walls contained arabinose, galactose and short-chain mycolic acids (C22 to C36) and their peptidoglycan contained meso-diaminopimelic acid. The G+C content of the DNA was 51-60 mol%. MK-9 (H2) was the principal menaquinone. The 16S rDNA sequences of four isolates of each bacterium were determined and aligned with those of other members of the coryneform group. Phylogenetic analysis showed that the strains represented two new sublines within the genus Corynebacterium; Corynebacterium variabile and Corynebacterium ammoniagenes were their nearest known phylogenetic neighbours. Corynebacterium variabile and Corynebacterium ammoniagenes showed the highest levels of sequence homology with the isolates; however, DNA-DNA hydridization studies indicated that the Corynebacterium strains isolated from the cheese smear did not belong to either Corynebacterium variabile or Corynebacterium ammoniagenes (26 and 46% chromosomal similarity, respectively). On the basis of the phylogenetic and phenotypic distinctiveness of the unknown isolates, it is proposed that the bacteria be classified as two new Corynebacterium species, for which the names Corynebacterium mooreparkense sp. nov. and Corynebacterium casei sp. nov. are proposed. Type strains have been deposited in culture collections as Corynebacterium mooreparkense LMG S-19265T (= NCIMB 30131T) and Corynebacterium casei LMG S-19264T (= NCIMB 30130T).

C/EBPalpha and G-CSF receptor signals cooperate to induce the myeloperoxidase and neutrophil elastase genes.

To assess cooperation between G-CSF signals and C/EBPalpha, we characterized Ba/F3 pro-B cell lines expressing C/EBPalphaWT-ER and the G-CSF receptor (GCSFR). In these lines, GCSFR signals can be evaluated independent of their effect on C/EBPalpha levels. G-CSF alone did not induce the MPO, NE, LF, or PU.1 RNAs, and C/EBPalphaWT-ER alone stimulated low-level MPO and high-level PU.1 expression. Simultaneous activation of the GCSFR and C/EBPalphaWT-ER markedly increased MPO and NE induction at 24 h, and LF mRNA was detected at 48 h. G-CSF did not increase endogenous GCSFR, endogenous C/EBPalpha or exogenous C/EBPalphaWT-ER levels, and C/EBPalphaWT-ER did not induce endogenous or exogenous GCSFR. Several GCSFR mutants were also co-expressed with C/EBPalphaYWT-ER. Mutation of all four cytoplasmic tyrosines prevented NE induction but enhanced MPO induction. Mutation of Y704 was required for increased MPO induction. Consistent with this finding, removing IL-3 without G-CSF addition enabled MPO, but not NE, induction by C/EBPalphaWT-ER. GCSFR signals or related signals from other receptors may cooperate with C/EBPalpha to direct differentiation of normal myeloid stem cells.

Proteomic analysis of macrophage differentiation. p46/52(Shc) Tyrosine phosphorylation is required for CSF-1-mediated macrophage differentiation.

Macrophage colony stimulating factor (M-CSF or CSF-1) acts to regulate the development and function of cells of the macrophage lineage. Murine myeloid FDC-P1 cells transfected with the CSF-1 receptor (FD/WT) adopt a macrophage-like morphology when cultured in CSF-1. This process is abrogated in FDC-P1 cells transfected with the CSF-1 receptor with a tyrosine to phenyalanine substitution at position 807 (FD/807), suggesting that a molecular interaction critical to differentiation signaling is lost (Bourette, R. P., Myles, G. M., Carlberg, K., Chen, A. R., and Rohrschneider, L. R. (1995) Cell Growth Differ. 6, 631--645). A detailed examination of lysates of CSF-1-treated FD/807 cells by two-dimensional SDS-polyacrylamide gel electrophoresis (PAGE) revealed a number of proteins whose degree of tyrosine phosphorylation was modulated by the Y807F mutation. Included in this category were three phosphorylated proteins that co-migrated with p46/52(Shc). Immunoprecipitation, Western blotting, and in vitro binding studies suggest that they are indeed p46/52(Shc). A key regulator of differentiation in a number of cell systems, ERK was observed to exhibit an activity that correlated with the relative degree of differentiation induced by CSF-1 in the two cell types. Transfection of cells with a non-tyrosine-phosphorylatable form of p46/52(Shc) prevented the normally observed CSF-1-mediated macrophage differentiation as determined by adoption of macrophage-like morphology and expression of the monocyte/macrophage lineage cell surface marker, Mac-1. These results are the first to suggest that p46/52(Shc) may play a role in CSF-1-induced macrophage differentiation. Additionally, a number of proteins were identified by two-dimensional SDS-PAGE whose degree of tyrosine phosphorylation is also modulated by the Y807F substitution. This group of molecules may contain novel signaling molecules important in macrophage differentiation.

Seasonal incidence and antibiotic susceptibility patterns of Pasteurellaceae isolated from American bison (Bison bison).

Ninety pharyngeal tonsils were collected from 2-year-old American bison (Bison bison) bulls and sampled for members of the Pasteurellaceae family. Particular attention was paid to seasonal incidence and antimicrobial resistance in serotypes and biovariants. Multiple strains of Pasteurella haemolytica (39%), P. trehalosi (68%), P. multocida (34%) and Haemophilus somnus (13%) were cultured from 86 out of the 90 (96%) tonsil samples. Pasteurella trehalosi was the most common and evenly distributed of the organisms recovered. Pasteurella haemolytica was found in fewer numbers than P. trehalosi, but showed an increase in number of isolates recovered with each sampling period. Pasteurella multocida, both A and D capsular types, was recovered from all sampling periods. No serotype pattern was observed in any of the animal groups sampled. One hundred twenty-seven of 147 (86%) of the isolates were resistant to at least 1 antibiotic, 95/147 (65%) to at least 2 different antibiotics, and 16/147 (11%) to at least 3 antibiotics. The most common resistance pattern observed was to neomycin and spectinomycin (73/147) (49%).

Somatostatin modulates G-CSF-induced but not interleukin-3-induced proliferative responses in myeloid 32D cells via activation of somatostatin receptor subtype 2.

INTRODUCTION: Somatostatin, originally identified as a peptide involved in neurotransmission, functions as an inhibitor of multiple cellular responses, including hormonal secretion and proliferation. Somatostatin acts through activation of G-protein-coupled receptors of which five subtypes have been identified. We have recently established that human CD34/c-kit expressing hematopoietic progenitors and acute myeloid leukemia (AML) cells exclusively express SSTR2. A major mechanism implicated in the antiproliferative action of somatostatin involves activation of the SH2 domain-containing protein tyrosine phosphatase SHP-1. While 0.1-1 x 10(-9) M of somatostatin, or its synthetic stable analog octreotide, can inhibit G-CSF-induced proliferation of AML cells, little or no effects are seen on GM-CSF- or IL-3-induced responses. MATERIALS AND METHODS: To study the mechanisms underlying the antiproliferative responses of myeloblasts to somatostatin, clones of the IL-3-dependent murine cell line 32D that stably express SSTR2 and G-CSF receptors were generated. RESULTS: Similar to AML cells, octreotide inhibited G-CSF-induced but not IL-3-induced proliferative responses of 32D[G-CSF-R/SSTR2] cells. Somatostatin induced SHP-1 activity and inhibited G-CSF-induced, but not IL-3-induced, activation of the signal transducer and activator of transcription proteins STAT3 and STAT5. CONCLUSION: Based on these data and previous results, we propose a model in which recruitment and activation of the tyrosine phosphatase SHP-1 by SSTR2 is involved in the selective negative action of somatostatin on G-CSF-R signaling.

Microbacterium gubbeenense sp. nov., from the surface of a smear-ripened cheese.

Phenotypic and phylogenetic studies were performed on 11 strains of a Microbacterium-like organism isolated from the surface of a smear-ripened cheese. The isolates were Gram-positive, catalase-positive, facultatively anaerobic, oxidase-negative, non-spore-forming, non-motile, small, slender rods and grew in 12% (w/v) NaCl. Chemotaxonomic investigation revealed that all the isolates belonged unambiguously to the genus Microbacterium. They contained type B1 peptidoglycans with L-lysine as the diamino acid and glycolyl acyl types; rhamnose and galactose were the cell wall sugars. The G+C content ranged from 69 to 72 mol%. The major menaquinones were MK-11 and MK-12 and the major fatty acids were anteiso C15:0 and C17:0 and iso C16:0. Phylogenetic analysis of the 16S rRNA sequences of four isolates showed that they represented a new subline in the genus Microbacterium, with Microbacterium barkeri as their nearest phylogenetic neighbour. M. barkeri showed the highest sequence similarity to the isolates; however, DNA-DNA hybridization showed that the isolates had only 38% chromosomal similarity to M. barkeri. Based on the phylogenetic and phenotypic distinctiveness of the isolates, it is proposed that they be classified as a new Microbacterium species, for which the name Microbacterium gubbeenense sp. nov. is suggested. The type strain has been deposited as LMG S-19263T (= NCIMB 30129T). The GenBank accession number for the 16S rDNA sequence of the type strain is AF263563.