The mutualistic symbiont Vibrio fischeri builds a symbiotic biofilm during colonization of squid hosts. Regulation of the exopolysaccharide component, termed Syp, has been examined in strain ES114, where production is controlled by a phosphorelay that includes the inner membrane hybrid histidine kinase RscS. Most strains that lack RscS or encode divergent RscS proteins cannot colonize a squid host unless RscS from a squid symbiont is heterologously expressed. In this study, we examine V. fischeri isolates worldwide to understand the landscape of biofilm regulation during beneficial colonization. We provide a detailed study of three distinct evolutionary groups of V. fischeri and find that while the RscS-Syp biofilm pathway is required in one of the groups, two other groups of squid symbionts require Syp independent of RscS. Mediterranean squid symbionts, including V. fischeri SR5, colonize without an RscS homolog encoded by their genome. Additionally, group A V. fischeri strains, which form a tightly related clade of Hawaii isolates, have a frameshift in rscS and do not require the gene for squid colonization or competitive fitness. These same strains have a frameshift in sypE, and we provide evidence that this group A sypE allele leads to an upregulation in biofilm activity. Thus, this work describes the central importance of Syp biofilm in colonization of diverse isolates and demonstrates that significant evolutionary transitions correspond to regulatory changes in the syp pathway. IMPORTANCE Biofilms are surface-associated, matrix-encased bacterial aggregates that exhibit enhanced protection to antimicrobial agents. Previous work has established the importance of biofilm formation by a strain of luminous Vibrio fischeri bacteria as the bacteria colonize their host, the Hawaiian bobtail squid. In this study, expansion of this work to many natural isolates revealed that biofilm genes are universally required, yet there has been a shuffling of the regulators of those genes. This work provides evidence that even when bacterial behaviors are conserved, dynamic regulation of those behaviors can underlie evolution of the host colonization phenotype. Furthermore, this work emphasizes the importance of investigating natural diversity as we seek to understand molecular mechanisms in bacteria.A fundamental question in studying host-associated bacterial communities is understanding how specific microbial taxa assemble reproducibly in their host. Key insights into these processes were first obtained by studying plant-associated microbes, and the discovery and characterization of Nod factors in rhizobia were valuable to understanding how partner choice between microbe and host could be mediated at the molecular level (1, 2). There are complex communities in humans and other vertebrate animals, yet metagenomic and imaging analyses of these communities have Citation Rotman ER,

Bacteria have evolved diverse strategies to compete for limited space and resources. Because these mechanisms can be costly to use, their expression and function are often restricted to specific environments where the benefits outweigh the costs.

Aspergillus fumigatus causes life-threatening infections, and treatment options remain limited. Thus, there is an urgent need to find new therapeutic targets to treat this deadly disease. Previously, we have shown that SREBP transcription factors and their regulatory components are critical for the pathobiology of A. fumigatus. Here we identify a role for RbdB, a rhomboid protease, as an essential component of SREBP activity. Our results indicate that mutants lacking rbdB have growth defects under hypoxic conditions, are hypersusceptible to voriconazole, lack extracellular siderophore production, and fail to cause disease in a murine model of invasive pulmonary aspergillosis. This study increases our understanding of the molecular mechanisms involved in SREBP activation in pathogenic fungi and provides a novel therapeutic target for future development.

Invasive aspergillosis (IA), most commonly caused by the filamentous fungus Aspergillus fumigatus, occurs in immune compromised individuals. The ability of A. fumigatus to proliferate in a multitude of environments is hypothesized to contribute to its pathogenicity and virulence. Transcription factors (TF) have long been recognized as critical proteins for fungal pathogenicity, as many are known to play important roles in the transcriptional regulation of pathways implicated in virulence. Such pathways include regulation of conidiation and development, adhesion, nutrient acquisition, adaptation to environmental stress, and interactions with the host immune system among others. In both murine and insect models of IA, TF loss of function in A. fumigatus results in cases of hyper- and hypovirulence as determined through host survival, fungal burden, and immune response analyses. Consequently, the study of specific TFs in A. fumigatus has revealed important insights into mechanisms of pathogenicity and virulence. Although in vitro studies have identified virulence-related functions of specific TFs, the full picture of their in vivo functions remain largely enigmatic and an exciting area of current research. Moreover, the vast majority of TFs remain to be characterized and studied in this important human pathogen. Here in this mini-review we provide an overview of selected TFs in A. fumigatus and their contribution to our understanding of this important human pathogen's pathogenicity and virulence.

Regulation of fungal cell wall biosynthesis is critical to maintain cell wall integrity in dynamic fungal infection microenvironments. Genes involved in this response that impact fungal fitness and host immune responses remain to be fully defined. In this study, we observed that a yeast ssd1 homolog, ssdA, in the filamentous fungus Aspergillus fumigatus is involved in trehalose and cell wall homeostasis. An ssdA null mutant strain exhibited an increase in trehalose levels and a reduction in fungal colony growth rate. In contrast, overexpression of ssdA perturbed trehalose biosynthesis and reduced germination of conidia. The ssdA null mutant strain was more resistant to cell wall-perturbing agents, while overexpression of ssdA increased sensitivity. Overexpression of ssdA significantly increased chitin levels, and both loss and overexpression of ssdA altered subcellular localization of the class V chitin synthase CsmA. Strikingly, overexpression of ssdA abolished adherence to abiotic surfaces and severely attenuated the virulence of A. fumigatus in a murine model of invasive pulmonary aspergillosis. Despite the severe in vitro fitness defects observed upon loss of ssdA, neither surface adherence nor murine survival was impacted. In conclusion, A. fumigatus SsdA plays a critical role in cell wall homeostasis impacting A. fumigatus-host interactions. IMPORTANCE The incidence of life-threatening infections caused by the filamentous fungus Aspergillus fumigatus is increasing along with an increase in the number of fungal strains resistant to contemporary antifungal therapies. The fungal cell wall and the associated carbohydrates required for its synthesis and maintenance are attractive drug targets given that many genes encoding proteins involved in cell wall biosynthesis and integrity are absent in humans. Importantly, genes and associated cell wall biosynthesis and homeostasis regulatory pathways remain to be fully defined in A. fumigatus. In this report, we identify SsdA as an important component of trehalose and fungal cell wall biosynthesis in A. fumigatus that consequently impacts the host immune response and fungal virulence in animal models of infection.