It is time for a new type of type to facilitate naming the microbial world.

Since January 1, 2001, the only acceptable nomenclatural type for species under the International Code of Nomenclature of Prokaryotes (ICNP) has been pure cultures. Here, we argue that this requirement is discordant with the more inclusive nature of nomenclatural types accepted under other codes of nomenclature and posit that the unique rigidity of the ICNP has failed to serve the broad research community and has stifled progress. This case is based on the axiom that many archaea and bacteria are interdependent in nature and therefore difficult, if not impossible, to grow, preserve, and distribute as pure cultures. As such, a large proportion of Earth's biodiversity cannot be named under the current system, which limits our ability to communicate about microbial diversity within and beyond the microbiology research community. Genome sequence data are now encouraged for valid publication of new taxa in microbial systematics journals, and metagenome-assembled genomes and single cell-amplified genomes are being generated rapidly from every biome on Earth. Thus, genome sequences are available for both cultivated and uncultivated microorganisms and can readily serve as a new category of nomenclatural type, allowing for a unified nomenclature for all archaea and bacteria, whether or not they are available as pure cultures. Ideally this would be under a single code of nomenclature but, as we review here, the newly established SeqCode will operate in parallel with the ICNP as a first step toward this goal.

Addressing the sublime scale of the microbial world: reconciling an appreciation of microbial diversity with the need to describe species.

There are fewer than 20,000 prokaryotic species with validly published names, meaning >99% of a reasonable estimate of microbial diversity remains formally unnamed. Here we explore the damaging consequences of the current practice in which each new species is described in a standardized publication, most typically a 'single strain species description'. This approach is both an impediment to scaling up progress in naming the microbial world and also a significant factor in the poor reputation of the discipline of microbial taxonomy. We conclude that significant changes in author habits are needed and make constructive suggestions as to how author practice should adapt.

Genomic analysis of a novel nontoxigenic Corynebacterium diphtheriae strain isolated from a cancer patient.

The genome of a novel nontoxigenic Corynebacterium diphtheriae, strain 5015, isolated from a patient with adenoid cystic carcinoma was sequenced and compared with 117 publically available genomes. This strain is phylogenetically distinct and lacks virulence genes encoding the toxin, BigA and Sdr-like adhesins. Strain 5015 possesses spaD-type and spaH-type pilus gene clusters with a loss of some gene functions, and 31 unique genes that need molecular characterization to understand their potential role in virulence characteristics.

Genomic analyses reveal two distinct lineages of Corynebacterium ulcerans strains.

Corynebacteriumulcerans is an important zoonotic pathogen which is causing diphtheria-like disease in humans globally. In this study, the genomes of three recently isolated C. ulcerans strains, 4940, 2590 and BR-AD 2649, respectively from an asymptomatic carrier, a patient with pharyngitis and a canine host, were sequenced to investigate their virulence potential. A comparative analysis was performed including the published genome sequences of 16 other C. ulcerans isolates. C. ulcerans strains belong to two lineages; 13 strains are grouped together in lineage 1, and six strains comprise lineage 2. Consistent with the zoonotic nature of C. ulcerans infections, isolates from both the human and canine hosts clustered in both the lineages. Most of the strains possessed spaDEF and spaBC gene clusters along with the virulence genes cpp, pld, cwlH, nanH, rpfI, tspA and vsp1. The gene encoding Shiga-like toxin was only present in one strain, and 11 strains carried the tox gene encoding the diphtheria-like toxin. However, none of strains 4940, 2590 and BR-AD 2649 carried any toxin genes. These strains varied in the number of prophages in their genomes, which suggests that they play an important role in introducing diversity in C. ulcerans. The pan-genomic analyses revealed a variation in the number of membrane-associated and secreted proteins that may contribute to the variation in pathogenicity among different strains.

Is investigation of patients with haemoptysis and normal chest radiograph justified?

BACKGROUND: Haemoptysis is a common clinical symptom. A small proportion of patients present with haemoptysis and normal chest radiograph. The investigation strategy for this group of patients is unclear. The aim of this study is to see whether further investigations for this group of patients are justified. METHODS: A retrospective analysis was conducted of consecutive patients presenting with haemoptysis and normal chest radiograph over a period of 56 months irrespective of their smoking status. These patients were investigated by CT of the thorax and fibreoptic bronchoscopy. RESULTS: 275 episodes of haemoptysis with normal chest radiograph were investigated further in 270 patients (60% males). The median age was 60 years. Twenty-six patients were diagnosed to have respiratory tract malignancies (larynx, 1; trachea, 1; lung, 22; carcinoid, 1; and leiomyoma, 1). Eight (31%) of the 26 patients with respiratory tract malignancy had radical treatment. Fibreoptic bronchoscopy was diagnostic of cancer in 14 (54%) of the 26 patients with malignancy. CT of the thorax was suggestive of cancer in 24 (96%) of the 25 patients with malignancy. CONCLUSION: It is concluded that further investigation of haemoptysis in smokers is justified regardless of the amount or frequency of haemoptysis based on a 9.6% rate of malignancy in this consecutive series. It is recommended that these patients are investigated with CT of the thorax followed by fibreoptic bronchoscopy.

Phenotypic variation in Streptomyces sp. DSM 40537, a lipoteichoic acid producing actinomycete.

AIMS: To reinvestigate the production of lipoteichoic acid (LTA) by the actinomycete strain Streptomyces sp. DSM 40537 (=ATCC 3351). METHODS AND RESULTS: LTA was extracted and purified from strain Streptomyces sp. DSM 40537. The identification of the LTA was confirmed by Western blotting with a monoclonal antibody. During these studies, two stable phenotypic variants of DSM 40537 were obtained, one of which released a distinctive orange pigment. 16S rRNA gene sequencing of each variant yielded identical sequences and allowed phylogenetic analysis to be performed. CONCLUSIONS: Streptomyces sp. DSM 40537 was shown to exhibit stable morphological variation. The strain was confirmed to be a LTA-producing actinomycete and to belong to the Streptomyces albidoflavus cluster within the genus Streptomyces. SIGNIFICANCE AND IMPACT OF THE STUDY: These data provide important support for the hypothesis that the distribution of LTA is linked to that of wall teichoic acids and emphasizes the need to reinvestigate LTA distribution in actinomycetes.

Tuberculosis: an under-diagnosed aetiological agent in uveitis with an effective treatment.

PURPOSE: To highlight the diversity of clinical presentations with tubercular uveitis in a nonendemic setting, and discuss the diagnostic approach and an effective treatment. METHOD: Descriptive case series. RESULTS: A total of 12 cases of varied presentations of tubercular uveitis diagnosed over a period of 1 year of which six cases are described in detail. Presentations included choroidal tuberculomas, multifocal choroiditis, recurrent granulomatous uveitis, panuveitis with cystoid macular oedema, and serpiginous choroiditis. All cases had a chronic or recurrent course and responded very well to antitubercular treatment. Diagnosis was mainly assisted by positive tuberculin testing. CONCLUSION: A high index of suspicion helps diagnose ocular tuberculosis in areas of low prevalence of the disease. It forms part of the differential diagnosis of any chronic or recurrent uveitis, especially in an at-risk patient. Antitubercular treatment seems highly effective.

Effects of moderate hypothermia on IL-1 beta-induced leukocyte rolling and adhesion in pial microcirculation of mice and on proinflammatory gene expression in human cerebral endothelial cells.

SUMMARY: Although the neuroprotective effects of hypothermia have been known for a long time, the molecular correlates of this neuroprotection are poorly understood. In this study, the authors investigated how hypothermia affects inflammatory responses in the brain elicited by systemic injection of IL-1 beta. Leukocyte rolling and adhesion were quantified in pial venules (20 to 50 microm) of C57/Bl6 mice 4 hours after intraperitoneal injection of IL-1 beta (5 microg/kg) using an open cranial window and intravital microscopy. Animals were subjected to moderate hypothermia (32 degrees C) or normothermia (37 degrees C) for 1 or 4 hours after IL-1 beta injection. Significant increases in leukocyte rolling and adhesion were observed in IL-1 beta-injected animals as compared with sham controls. Whereas 1-hour hypothermia did not affect IL-1 beta-induced leukocyte rolling and adhesion, 4-hour hypothermia caused a reduction in both rolling and adhesion. Molecular mechanisms of hypothermic effects were investigated in cultured human cerebral endothelial cells exposed to IL-1 beta (50 U/mL) for 4 hours at 37 degrees C or 32 degrees C followed by 18 hours at 37 degrees C. Human cerebral endothelial cells exposed to IL-1 beta at 32 degrees C showed attenuated NF-kappa B activation determined by the Luciferase yellow reporter gene assay and reduced expression of IL-8 and IL-1 beta measured by reverse transcriptase-polymerase chain reaction and enzyme-linked immunosorbent assay. Intracellular adhesion molecule-1 was induced to similar levels (threefold over control) at both temperatures. The expression of CD18 on neutrophils in vitro was not affected by either IL-1 beta or hypothermia. These findings suggest that mechanisms by which hypothermia reduces leukocyte rolling and adhesion include suppression of inflammatory gene transcription in brain endothelial cells.

Reclassification of [Pseudomonas] doudoroffii (Baumann et al. 1983) into the genus Oceanomonas gen. nov. as Oceanomonas doudoroffii comb. nov., and description of a phenol-degrading bacterium from estuarine water as Oceanomonas baumannii sp. nov.

A bacterium, isolate GB6T, capable of degrading phenol in the presence of elevated salinity was isolated from the estuary of the River Wear, UK. The bacterium was subjected to biochemical and molecular analysis to determine its taxonomic status. These studies indicated that the bacterium was a distinct species closely related to [Pseudomonas] doudoroffii. However, the phylogenetic analysis indicated that [Pseudomonas] doudoroffii was misclassified, as noted previously. Analysis of the characteristics of isolate GB6T and the type strain of [Pseudomonas] doudoroffii confirmed that these bacteria belonged to the same novel genus, which we have named Oceanomonas gen. nov. The type strain of Oceanomonas doudoroffii (Baumann et al. 1983) comb. nov. is ATCC 27123T (= DSM 7028T. The DNA-DNA homology between isolate GB6T and [Pseudomonas] doudoroffii is low and phenotypic differences between the two organisms are evident. Isolate GB6T (= ATCC 700832T = NCIMB 13685T) is therefore proposed as the type strain of a new species, Oceanomonas baumannii sp. nov.

Surface immunolocalisation of HPr in the equine pathogen Streptococcus equi.

We have investigated the surface localisation of the phosphotransferase system protein HPr in the equine pathogen Streptococcus equi subsp. equi using immunogold localisation and transmission electron microscopy. Like the LppC acid phosphatase lipoprotein, a reference surface antigen, the S. equi HPR could be clearly detected on the surfaces of intact cells. This study is consistent with previous reports that some streptococcal HPr is cell surface associated and suggests that the extracytoplasmic mobilisation and transfer of phosphate groups by streptococci warrant further investigation.

Characterization of acid phosphatase activities in the equine pathogen Streptococcus equi.

Acid phosphatases hydrolyse phosphomonoesters at acidic pH in a variety of physiological contexts. The recently defined class C family of acid phosphatases includes the 32 kDa LppC lipoprotein of Streptococcus equisimilis. To define further the distribution of acid phosphatases in the genus Streptococcus we have examined the equine pathogens Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus. Whole cell assays indicated that these organisms possess two acid phosphatases with activity optima at pH 5.0 and pH 6.0-6.5 and that only the former of these was, like LppC, resistant to EDTA. Western blotting with a polyclonal anti-LppC antiserum revealed the presence of a cross-reactive 32 kDa protein in both organisms. The cross-reactive protein in S. equi was shown to be a surface accessible lipoprotein as its processing was inhibited by the antibiotic globomycin and it was released from whole cells by treatment with trypsin. The presence of DNA sequences homologous to the S. equisimilis lppC gene were confirmed by PCR. These data strongly suggest that Streptococcus equi subsp. equi and Streptococcus equi subsp. zooepidemicus produce a lipoprotein acid phosphatase homologous to LppC of S. equisimilis.

Identification of methionine-processed HPr in the equine pathogen Streptococcus equi.

Using preparative electrophoresis, a low molecular weight protein has been partially purified from a cell extract of the equine pathogen Streptococcus equi susp. equi. N-terminal sequence analysis and Western blotting revealed the protein to be HPr, a central component of the phosphoenolpyruvate:sugar phosphotransferase system (PTS). Interestingly, the only form of the HPr protein detected in S. equi was one with the amino-terminal methionine removed, a modification that has previously been associated with surface localization of streptococcal HPr proteins.

Identification of lipoprotein homologues of pneumococcal PsaA in the equine pathogens Streptococcus equi and Streptococcus zooepidemicus.

Streptococcus equi and Streptococcus zooepidemicus are major etiological agents of upper and lower airway disease in horses. Despite the considerable animal suffering and economic burden associated with these diseases, the factors that contribute to the virulence of these equine pathogens have not been extensively investigated. Here we demonstrate the presence of a homologue of the Streptococcus pneumoniae PsaA protein in both of these equine pathogens. Inhibition of signal peptide processing by the antibiotic globomycin confirmed the lipoprotein nature of the mature proteins, and surface exposure was confirmed by their release from intact cells by mild trypsinolysis.

The modification of the membrane of Oceanomonas baumannii when subjected to both osmotic and organic solvent stress.

Oceanomonas baumanniioff a novel halotolerant bacterium which was isolated from the estuary of the river Wear (Sunderland, UK). When grown in tryptone soya broth it can tolerate high levels of phenol, which is not utilised as a carbon source in this medium. However, the level of tolerance was reduced from 35 mM to 3 mM phenol as salinity increased from 1% to 12% NaCl (w/v). Increasing salinity up to 12% NaCl also decreased the growth rate 8-fold and caused modifications to the cytoplasmic membrane particularly anionic phosphatidylglycerol levels, which doubled at the expense of zwitterionic phosphatidylethanolamine. In addition, changes in the phospholipid fatty acid composition were noted, cis-vaccenic acid decreased significantly at higher salinities. Intracellular solute levels also increased with increasing salinity and there was an accumulation of the compatible solutes ectoine, glycine betaine and glutamate. The addition of phenol to osmotically compromised cultures led to a further modification of the cytoplasmic membrane phospholipid composition, in particular, that the decrease in zwitterionic phosphatidylethanolamine and the increase of anionic phospholipid species was much less pronounced. A further decrease in unsaturation, particularly in the proportion of cis-vaccenic acid, and the mean chain length of the fatty acids suggested that this response was important in maintaining membrane integrity in the presence of phenol.

A role for platelets and endothelial selectins in tumor necrosis factor-alpha-induced leukocyte recruitment in the brain microvasculature.

The mechanisms mediating leukocyte recruitment into the cerebral nervous system during inflammation are still poorly understood. The objective of this study was to investigate the leukocyte recruitment in the brain microcirculation by intravital microscopy. Superfusion of the brain with artificial cerebrospinal fluid did not induce leukocyte rolling or adhesion. However, intraperitoneal tumor necrosis factor-alpha (TNF-alpha) caused marked leukocyte rolling and adhesion in the brain microcirculation. Histology revealed that the recruitment was primarily of neutrophils. Both E- and P-selectin were required for TNF-alpha-induced leukocyte recruitment, as rolling was reduced after treatment with either anti-E- or anti-P-selectin antibody and eliminated in E- or P-selectin-deficient mice. A significant increase in brain P- and E-selectin expression was seen after TNF-alpha treatment, but both were an order of magnitude less than in any other tissue. We observed significant platelet paving of TNF-alpha-stimulated endothelium and found that anti-platelet antibody reduced leukocyte rolling and adhesion, as did acetylsalicylic acid (aspirin). However, depletion of platelets did not reduce cerebral P-selectin expression. Moreover, chimeric mice lacking P-selectin on endothelium but not platelets had significantly decreased P-selectin expression and reduced leukocyte recruitment in the brain. This suggests a role for endothelial P-selectin in cerebral leukocyte recruitment. In conclusion, TNF-alpha-induced neutrophil recruitment into the brain requires both endothelial E-selectin and P-selectin as well as platelets, but platelet P-selectin was not a major contributor to this process.

Characterisation of a lipomannan lipoglycan from the mycolic acid containing actinomycete Dietzia maris.

Lipoglycans such as the mycobacterial lipoarabinomannans (LAM) are important cell envelope components of actinomycetes. To further our understanding of the diversity of these enigmatic macromolecules the lipoglycan composition of Dietzia maris has been investigated. Phenol-water extraction and hydrophobic interaction chromatography were used to purify a lipoglycan which was unusually small and predominantly lipomannan in nature. The presence of minor levels of arabinose along with components consistent with the presence of a phosphatidylinositol anchor suggest that this lipoglycan is a novel representative of the lipomannan/LAM structural archetype. This was further supported by the observed cross-reaction of the D. maris lipoglycan with an antiserum raised against LAM from Mycobacterium tuberculosis. These findings reveal a previously unsuspected diversity in the lipoglycan composition of the mycolic acid containing actinomycetes and are further discussed in relation to the apparent absence of phosphatidylinositolmannoside glycolipids in D. maris.

Identification of a lipoarabinomannan-like lipoglycan in Gordonia rubropertincta.

Lipoarabinomannans are well characterised lipoglycans present in the cell envelopes of mycobacteria and closely related bacteria. To define further the distribution of these lipoglycans we have investigated the mycolic acid-containing bacterium Gordonia rubropertincta and, by analysis of carbohydrate composition and antigenic cross-reactivity, demonstrated the presence of a lipoarabinomannan-like lipoglycan. A fraction that appeared to contain higher phosphatidylinositolmannosides was also isolated.

Cell envelope composition and organisation in the genus Rhodococcus.

A knowledge of the organisation of the rhodococcal cell envelope is of fundamental importance if the environmental and biotechnological significance of these bacteria are to be understood and successfully exploited. The genus Rhodococcus belongs to a distinctive suprageneric taxon, the mycolata, which includes among others the genera Corynebacterium, Mycobacterium and Nocardia. Members of this taxon exhibit an unusual complexity in their cell envelope composition and organisation compared to other Gram-positive bacteria. Models that describe the architecture of the mycobacterial cell envelope are extrapolated here to provide a model of the rhodococcal cell envelope. The rhodococcal cell envelope is dominated by the presence of an arabinogalactan cell wall polysaccharide and large 2-alkyl 3-hydroxy branched-chain fatty acids, the mycolic acids, which are covalently assembled into a peptidoglycan-arabinogalactan-mycolic acid matrix. This review further emphasises that the mycolic acids in this complex form the basis of an outer lipid permeability barrier. The localisation and roles of other cell envelope components, notably complex free lipids, lipoglycans, proteins and lipoproteins are also considered.

Macroamphiphilic cell envelope components of Rhodococcus equi and closely related bacteria.

Recent progress towards an understanding of the architecture of the mycobacterial cell envelope (P.J. Brennan and H. Nikaido, Annual Review of Biochemistry 64 (1995) 29-63) provides a model with features more generally applicable to cell envelope organisation in other mycolic acid-containing bacteria. Using this archetype, a model for the organisation of the rhodococcal cell envelope is presented here, with particular reference to cell envelope composition in Rhodococcus equi. The likelihood that mycolic acids bound to the cell wall arabinogalactan contribute to the formation of a distinct outer lipid layer is emphasised. Furthermore, the model incorporates recent work which has characterised rhodococcal macroamphiphiles (lipoglycans and lipoproteins), including the VapA virulence-associated lipoproteins of R. equi.