Pseudomonas aeruginosa is a ubiquitous environmental bacterium and a clinically significant opportunistic human pathogen. Central to the ability of P. aeruginosa to colonise both environmental and host niches is the acquisition of zinc. Here we show that P. aeruginosa PAO1 acquires zinc via an ATP-binding cassette (ABC) permease in which ZnuA is the high affinity, zinc-specific binding protein. Zinc uptake in Gram-negative organisms predominantly occurs via an ABC permease, and consistent with this expectation a P. aeruginosa ΔznuA mutant strain showed an ~60% reduction in cellular zinc accumulation, while other metal ions were essentially unaffected. Despite the major reduction in zinc accumulation, minimal phenotypic differences were observed between the wild-type and ΔznuA mutant strains. However, the effect of zinc limitation on the transcriptome of P. aeruginosa PAO1 revealed significant changes in gene expression that enable adaptation to low-zinc conditions. Genes significantly up-regulated included non-zinc-requiring paralogs of zinc-dependent proteins and a number of novel import pathways associated with zinc acquisition. Collectively, this study provides new insight into the acquisition of zinc by P. aeruginosa PAO1, revealing a hitherto unrecognized complexity in zinc homeostasis that enables the bacterium to survive under zinc limitation.

SummaryThe psaBCA locus of Streptococcus pneumoniae encodes a putative ABC Mn 2+ + + + -permease complex. Downstream of the operon is psaD , which may be cotranscribed and encodes a thiol peroxidase. Previously, there has been discordance concerning the phenotypic impact of mutations in the psa locus, resolution of which has been complicated by differences in mutant construction and the possibility of polar effects. Here, we constructed unmarked, in frame However, we found that deleting psaB resulted in decreased expression of PsaA relative to that in D39, whereas deleting both psaB and psaC resulted in at least wild-type levels of PsaA.

S. pneumoniae is a component of the commensal nasopharyngeal microflora of humans, but from this reservoir, it can progress to localized or invasive disease with a frequency that translates into massive global morbidity and mortality. However, the factors that govern the switch from commensal to pathogen, as well as those that determine disease tropism, are poorly understood. Here we show that capacity to utilize raffinose can determine the nature of the disease caused by a given pneumococcal strain. Moreover, our findings provide an interesting example of convergent evolution, whereby pneumococci belonging to two unrelated serotypes/lineages exhibit SNPs in separate genes affecting raffinose uptake and utilization that correlate with distinct pathogenic profiles in vivo. This further underscores the critical role of differential carbohydrate metabolism in the pathogenesis of localized versus invasive pneumococcal disease.

Communication between bacterial cells is crucial for the coordination of diverse cellular processes that facilitate environmental adaptation and, in the case of pathogenic species, virulence. This is achieved by the secretion and detection of small signaling molecules called autoinducers, a process termed quorum sensing. To date, the only signaling molecule recognized by both Gram-positive and Gram-negative bacteria is autoinducer 2 (AI-2), synthesized by the metabolic enzyme LuxS (S-ribosylhomocysteine lyase) as a by-product of the activated methyl cycle. Homologues of LuxS are ubiquitous in bacteria, suggesting a key role in interspecies, as well as intraspecies, communication. Gram-negative bacteria sense and respond to AI-2 via the Lsr ABC transporter system or by the LuxP/LuxQ phosphorelay system. However, homologues of these systems are absent from Gram-positive bacteria and the AI-2 receptor is unknown. Here we show that in the major human pathogen Streptococcus pneumoniae, sensing of exogenous AI-2 is dependent on FruA, a fructose-specific phosphoenolpyruvate-phosphotransferase system that is highly conserved in Gram-positive pathogens. Importantly, AI-2 signaling via FruA enables the bacterium to utilize galactose as a carbon source and upregulates the Leloir pathway, thereby leading to increased production of capsular polysaccharide and a hypervirulent phenotype.

The capsular serotype has long been associated with the virulence of Streptococcus pneumoniae. Here we present an in-depth study of phenotypic and genetic differences between serotype 3 and serogroup 11 S. pneumoniae clinical isolates from both the general and indigenous populations of Australia. Both serotypes/ groups included clonally unrelated strains with differences in well-known polymorphic virulence genes, such as nanA and pspA, as demonstrated by multilocus sequence typing and Western blot analysis. Nonetheless, the serotype 3 strains were consistently and significantly more virulent in mice than the serogroup 11 strains. Despite extensive genomic analysis, noncapsular genes common to one serotype/group but not the other were not identified. Nevertheless, following the conversion of a serotype 11A isolate to serotype 3 and subsequent analysis in an intranasal infection model, it was evident that both capsular and noncapsular factors determine the virulence phenotype in mice. However, it appears that these noncapsular factors vary from strain to strain.Streptococcus pneumoniae (the pneumococcus) is a formidable pathogen, being responsible for a broad spectrum of diseases, including pneumonia, meningitis, bacteremia, and otitis media (OM), and it accounts for more deaths worldwide than any other single pathogen (48). In Australia, the overall rate of invasive pneumococcal disease (IPD) in the indigenous population is over 4 times the rate seen in nonindigenous Australians (54). Furthermore, chronic suppurative OM, which is normally rare in developed countries, is present in remote Aboriginal communities at 10 times the rate defined as a major public health concern by the World Health Organization (9). Not only do children living in these communities suffer the highest rates of tympanic membrane perforation in the world, but 80% are estimated to suffer from some degree of OMrelated hearing impairment (45).Significant research on the pneumococcus has already been undertaken, but there are still many unanswered questions due to the complex nature of its pathogenicity. The polysaccharide capsule that surrounds the pneumococcus and determines the serotype, of which there are more than 90, is the most wellestablished virulence factor. Certain serotypes have been shown to have a higher association with disease or carriage than others (7,19,20,60). In addition, a number of conserved virulence proteins, including neuraminidase A (NanA), pneumolysin (Ply), autolysin (LytA), and pneumococcal surface proteins A and C (PspA and PspC), have been widely characterized and shown to be critical in pathogenesis (29, 50). However, these are only a subset of potential virulence factors. Like other pathogens, such as Haemophilus influenzae, another common cause of invasive disease and OM, S. pneumoniae has a pangenome (23, 24). Core genes, which are conserved across pneumococcal strains, represent only 70 to 80% of the genome, and a gene pool of over 5,000 orthologous clusters is estimated to be available to this naturally tran...