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The pathogenic actinomycete Rhodococcus equi harbors different types of virulence plasmids associated with specific nonhuman hosts. We determined the complete DNA sequence of a vapB + plasmid, typically associated with pig isolates, and compared it with that of the horse-specific vapA + plasmid type. pVAPB1593, a circular 79,251-bp element, had the same housekeeping backbone as the vapA + plasmid but differed over an ≈22-kb region. This variable region encompassed the vap pathogenicity island (PAI), was clearly subject to selective pressures different from those affecting the backbone, and showed major genetic rearrangements involving the vap genes. The pVAPB1593 PAI harbored five different vap genes ( vapB and vapJ to - M , with vapK present in two copies), which encoded products differing by 24 to 84% in amino acid sequence from the six full-length vapA + plasmid-encoded Vap proteins, consistent with a role for the specific vap gene complement in R. equi host tropism. Sequence analyses, including interpolated variable-order motifs for detection of alien DNA and reconstruction of Vap family phylogenetic relationships, suggested that the vap PAI was acquired by an ancestor plasmid via lateral gene transfer, subsequently evolving by vap gene duplication and sequence diversification to give different (host-adapted) plasmids. The R. equi virulence plasmids belong to a new family of actinobacterial circular replicons characterized by an ancient conjugative backbone and a horizontally acquired niche-adaptive plasticity region.

Infection with Rhodococcus (Corynebacterium) equi is a well-recognised condition in foals that represents a consistent and serious risk worldwide. The condition manifests itself primarily as one of pulmonary abscessation and bronchitis, hence the terminology of 'rattles' derived from its most obvious clinical sign, frequently terminal when first identified. This review addresses the clinical manifestation, bacteriology and pathogenesis of the condition together with recent developments providing knowledge of the organism in terms of virulence, epidemiology, transmission and immune responses. Enhanced understanding of R. equi virulence mechanisms and biology derived from the recently available genome sequence may facilitate the rational development of a vaccine and the improvement of farm management practices used to control R. equi on stud farms in the future. Reliance on vaccines alone, in the absence of management strategies to control the on-farm challenge is likely to be disappointing.

Objectives: In this study a large random collection (n = 378) of Irish thermophilic Campylobacter isolates were investigated for the presence of integrons, genetic elements associated with the dissemination of anti- microbial resistance. Methods: Purified genomic DNA from each isolate was analysed by PCR for the presence of class 1 inte- grons. Four gene cassette-associated amplicons were completely characterized. Results: Sixty-two of the isolates possessed a complete class 1 integron with a recombined gene cassette located within a 1.0 kb amplicon containing an aadA2 gene. This cassette was present in both Campylo- bacter jejuni and Campylobacter coli isolates and following sequence analysis was shown to be similar to sequences recently reported in Salmonella enterica Hadar and on an 85 kb plasmid conferring quinolone resistance in Escherichia coli. Conclusions: Aminoglycoside aadA2-encoding class 1 integrons were identified among unrelated Campy- lobacter spp. Amino acid sequence comparisons revealed identical structures in both Salmonella and E. coli. The presence of class 1 integrons in Campylobacter spp. may be significant should these organisms enter the food chain and especially when antimicrobial treatment for severe infections is being considered.

We report the genome of the facultative intracellular parasite Rhodococcus equi, the only animal pathogen within the biotechnologically important actinobacterial genus Rhodococcus. The 5.0-Mb R. equi 103S genome is significantly smaller than those of environmental rhodococci. This is due to genome expansion in nonpathogenic species, via a linear gain of paralogous genes and an accelerated genetic flux, rather than reductive evolution in R. equi. The 103S genome lacks the extensive catabolic and secondary metabolic complement of environmental rhodococci, and it displays unique adaptations for host colonization and competition in the short-chain fatty acid–rich intestine and manure of herbivores—two main R. equi reservoirs. Except for a few horizontally acquired (HGT) pathogenicity loci, including a cytoadhesive pilus determinant (rpl) and the virulence plasmid vap pathogenicity island (PAI) required for intramacrophage survival, most of the potential virulence-associated genes identified in R. equi are conserved in environmental rhodococci or have homologs in nonpathogenic Actinobacteria. This suggests a mechanism of virulence evolution based on the cooption of existing core actinobacterial traits, triggered by key host niche–adaptive HGT events. We tested this hypothesis by investigating R. equi virulence plasmid-chromosome crosstalk, by global transcription profiling and expression network analysis. Two chromosomal genes conserved in environmental rhodococci, encoding putative chorismate mutase and anthranilate synthase enzymes involved in aromatic amino acid biosynthesis, were strongly coregulated with vap PAI virulence genes and required for optimal proliferation in macrophages. The regulatory integration of chromosomal metabolic genes under the control of the HGT–acquired plasmid PAI is thus an important element in the cooptive virulence of R. equi., Author Summary Rhodococcus is a prototypic genus within the Actinobacteria, one of the largest microbial groups on Earth. Many of the ubiquitous rhodococcal species are biotechnologically useful due to their metabolic versatility and biodegradative properties. We have deciphered the genome of a facultatively parasitic Rhodococcus, the animal and human pathogen R. equi. Comparative genomic analyses of related species provide a unique opportunity to increase our understanding of niche-adaptive genome evolution and specialization. The environmental rhodococci have much larger genomes, richer in metabolic and degradative pathways, due to gene duplication and acquisition, not genome contraction in R. equi. This probably reflects that the host-associated R. equi habitat is more stable and favorable than the chemically diverse but nutrient-poor environmental niches of nonpathogenic rhodococci, necessitating metabolically more complex, expanded genomes. Our work also highlights that the recruitment or cooption of core microbial traits, following the horizontal acquistion of a few critical genes that provide access to the host niche, is an important mechanism in actinobacterial virulence evolution. Gene cooption is a key evolutionary mechanism allowing rapid adaptive change and novel trait acquisition. Recognizing the contribution of cooption to virulence provides a rational framework for understanding and interpreting the emergence and evolution of microbial pathogenicity.

Reasons for performing study: Rhodococcoccus equi is a significant cause of bronchopneumonia in foals worldwide. Infection of the lungs is believed to result from inhalation of virulent R. equi in dust from contaminated environments. A measure of infectious risk in an environment is the level of airborne contamination. Objectives: To assess and compare the level of airborne virulent R. equi in paddocks and stables. Methods: Air samples were collected sequentially over the 2003 foaling season from the paddocks and stables on 3 Irish horse breeding farms affected by R. equi pneumonia. Colony blotting and DNA hybridisation techniques allowed quantitation of virulent R. equi. Results: The odds of detecting airborne virulent R. equi in stables were 17.3 times greater than in paddocks. The median airborne concentration of virulent R. equi was significantly higher (P

Telomere length has been linked to disease stage and degree of (pan-)cytopenia in patients with bone marrow failure syndromes. The aim of the current study was to analyze the impact of replicative stress on telomere length in residual glycosylphosphatidylinositol-positive (GPI+) versus GPI– hematopoiesis in patients with paroxysmal nocturnal hemoglobinuria (PNH). Peripheral blood granulocytes from 16 patients and 22 healthy individuals were analyzed. For this purpose, we developed proaerolysin flow-FISH, a novel methodology that combines proaerolysin staining (for GPI expression) with flow-FISH (for telomere length measurement). We found significantly shortened telomeres in GPI– granulocytes (mean ± SE: 6.26 ± 0.27 telomere fluorescence units [TFU]), both compared with their GPI+ counterparts (6.88 ± 0.38 TFU; P = .03) as well as with age-matched healthy individuals (7.73 ± 0.23 TFU; P < .001). Our findings are in support of a selective growth advantage model of PNH assuming that damage to the GPI+ hematopoietic stem-cell (HSC) compartment leads to compensatory hyperproliferation of residual GPI–HSCs.

Objective—To compare isolates of Rhodococcus equi on the basis of geographic source and virulence status by use of pulsed-field gel electrophoresis (PFGE). Sample Population—290 isolates of R equi(218 virulent isolates from foals and 72 avirulent isolates from feces, soil, and respiratory tract samples) obtained between 1985 and 2000 from horses and horse farms from 4 countries. Procedure—DNA from isolates was digested with the restriction enzyme AseI and tested by use of PFGE. Products were analyzed for similarities in banding patterns by use of dendrograms. A similarity matrix was constructed for isolates, and the matrix was tested for nonrandom distributions of similarity values with respect to groupings of interest. Results—There was little grouping of isolates on the basis of country, virulence status, or region within Texas. Isolates of R equi were generally < 80% similar, as determined by use of PFGE. Isolates from the same farm generally were rarely of the same strain. Conclusions and Clinical Relevance—Considerable chromosomal variability exists among isolates of R equi obtained from the same farm, sites within Texas, or among countries from various continents. Only rarely will it be possible to link infections to a given site or region on the basis of analysis of isolates by use of PFGE of chromosomal DNA. (Am J Vet Res 2003;64:153–161)

Toxic blooms of cyanobacteria, including Anabaena species, which are neurotoxic, and Microcystis , which is hepatotoxic, are becoming increasingly common in potable water sources. Normal flocculation and chlorination processes do not remove the toxicity. The laboratory and pilot-plant experiments described in this article showed that the toxicity can be removed by both powdered and granular activated carbon, with and without chlorination, alum flocculation, and polyelectrolyte addition.

M. aeruginosa is a bloom-forming cyanobacterium which is common in fresh-water lakes. It contains a potent hepatotoxin which when purifed has been shown to be a heptapeptide of molecular weight 1019. The toxin was iodinated with 1251 using the lactoperoxidase method, the labelled toxin administered iI).travenously to adult female rats and the half-life and organ distribution measured. The blood half-life after redistribution into extracellular pools was 42 min. The liver and kidneys showed accumulation of 21�7 � 1�1 and 5�6 � 0�2% of the dose respectively after 30 min. Little accumulation was observed in other organs and tissues. Small-intestinal contents and urine contained 9�4 � 6� 1 and 2�9 � 1'2% of the dose respectively after 120 min. It was concluded that the liver is the main target organ for both accumulation and excretion of the toxin.