Age influences resistance of Caenorhabditis elegans to killing by pathogenic bacteria

Caenorhabditis elegans has previously been proposed as an alternative host for models of infectious disease caused by human pathogens. When exposed to some human pathogenic bacteria, the life span of nematodes is significantly reduced. We have shown that mutations in the age-1, and/or age-2 genes of C. elegans, that normally enhance life expectancy, can also increase resistance to killing by the bacterial pathogens Pseudomonas aeruginosa, Salmonella enterica var. Typhimurium, Burkholderia cepacia or Yersinia pseudotuberculosis. We also found that the rate at which wild-type C. elegans was killed by the bacterial pathogens tested increased as nematodes aged. In the case of P. aeruginosa infection, the difference in life span of wild type and age-1 mutants of C. elegans was not due to differences in the level of bacterial colonisation of the gut.     

The worm has turned--microbial virulence modeled in Caenorhabditis elegans

The nematode Caenorhabditis elegans is emerging as a facile and economical model host for the study of evolutionarily conserved mechanisms of microbial pathogenesis and innate immunity. A rapidly growing number of human and animal microbial pathogens have been shown to injure and kill nematodes. In many cases, microbial genes known to be important for full virulence in mammalian models have been shown to be similarly required for maximum pathogenicity in nematodes. C. elegans has been used in mutation-based screening systems to identify novel virulence-related microbial genes and immune-related host genes, many of which have been validated in mammalian models of disease. C. elegans-based pathogenesis systems hold the potential to simultaneously explore the molecular genetic determinants of both pathogen virulence and host defense.     

Caenorhabditis elegans is a model host for Salmonella typhimurium

The idea of using simple, genetically tractable host organisms to study the virulence mechanisms of pathogens dates back at least to the work of Darmon and Depraitère [1]. They proposed using the predatory amoeba Dictyostelium discoideum as a model host, an approach that has proved to be valid in the case of the intracellular pathogen Legionella pneumophila [2]. Research from the Ausubel laboratory has clearly established the nematode Caenorhabditis elegans as an attractive model host for the study of Pseudomonas aeruginosa pathogenesis [3]. P. aeruginosa is a bacterium that is capable of infecting plants, insects and mammals. Other pathogens with a similarly broad host range have also been shown to infect C. elegans [3,4]. Nevertheless, the need to determine the universality of C. elegans as a model host, especially with regards pathogens that have a naturally restricted host specificity, has rightly been expressed [5]. We report here that the enterobacterium Salmonella typhimurium, generally considered to be a highly adapted pathogen with a narrow range of target hosts [6], is capable of infecting and killing C. elegans. Furthermore, mutant strains that exhibit a reduced virulence in mammals were also attenuated for their virulence in C. elegans, showing that the nematode may constitute a useful model system for the study of this important human pathogen.     

The nematode Caenorhabditis elegans as a model for the study of host-pathogen interactions

For certain pathogens capable of infecting a broad range of organisms, there exist universal virulence factors, necessary for full pathogenicity regardless of the host. This has been most clearly demonstrated by Ausubel and colleagues for the human opportunistic pathogen Pseudomonas aeruginosa. As a consequence, one can use non-mammalian model systems, including the nematode worm Caenorhabditis elegans, to assay for such virulence factors. A significant number of pathogens of C. elegans, that provoke a range of diseases, are now known, including the opportunistic human pathogen Serratia marcescens. After explaining the practical advantages associated with the use of C. elegans, and briefly reviewing previous studies, the results of a screen for S. marcescens virulence factors will be presented.     

DAF-16-dependent suppression of immunity during reproduction in Caenorhabditis elegans

To further understand how the nematode Caenorhabditis elegans defends itself against pathogen attack, we analyzed enhanced pathogen resistance (epr) mutants obtained from a forward genetic screen. We also examined several well-characterized sterile mutants that exhibit an Epr phenotype. We found that sterility and pathogen resistance are highly correlated and that resistance in both epr and sterile mutants is dependent on DAF-16 activity. Our data indicate that a DAF-16-dependent signaling pathway distinct from previously described pathways is involved in the activation of genes that confer resistance to bacterial pathogens. The timing of DAF-16-dependent gene activation in sterile mutants coincides with the onset of embryonic development in wild-type animals, suggesting that signals from developing embryos normally downregulate the immune response.     

Resistance to antimicrobial peptides contributes to persistence of Salmonella typhimurium in the C. elegans intestine

The human pathogen Salmonella typhimurium can colonize, proliferate and persist in the intestine causing enteritis in mammals and mortality in the nematode Caenorhabditis elegans. Using C. elegans as a model, we determined that the Salmonella pathogenicity islands-1 and -2 (SPI-1 and SPI-2), PhoP and the virulence plasmid are required for the establishment of a persistent infection. We observed that the PhoP regulon, SPI-1, SPI-2 and spvR are induced in C. elegans and isogenic strains lacking these virulence factors exhibited significant defects in the ability to persist in the worm intestine. Salmonella infection also leads to induction of two C. elegans antimicrobial genes, abf-2 and spp-1, which act to limit bacterial proliferation. The SPI-2, phoP and Delta pSLT mutants are more sensitive to the cationic peptide polymyxin B, suggesting that resistance to worm's antimicrobial peptides might be necessary for Salmonella to persist in the C. elegans intestine. Importantly, we showed that the persistence defects of the SPI-2, phoP and Delta pSLT mutants could be rescued in vivo when expression of C. elegans spp-1 was reduced by RNAi. Together, our data suggest that resistance to host antimicrobials in the intestinal lumen is a key mechanism for Salmonella persistence.     

Virulence factors of the human opportunistic pathogen Serratia marcescens identified by in vivo screening

The human opportunistic pathogen Serratia marcescens is a bacterium with a broad host range, and represents a growing problem for public health. Serratia marcescens kills Caenorhabditis elegans after colonizing the nematode's intestine. We used C.elegans to screen a bank of transposon-induced S.marcescens mutants and isolated 23 clones with an attenuated virulence. Nine of the selected bacterial clones also showed a reduced virulence in an insect model of infection. Of these, three exhibited a reduced cytotoxicity in vitro, and among them one was also markedly attenuated in its virulence in a murine lung infection model. For 21 of the 23 mutants, the transposon insertion site was identified. This revealed that among the genes necessary for full in vivo virulence are those that function in lipopolysaccharide (LPS) biosynthesis, iron uptake and hemolysin production. Using this system we also identified novel conserved virulence factors required for Pseudomonas aeruginosa pathogenicity. This study extends the utility of C.elegans as an in vivo model for the study of bacterial virulence and advances the molecular understanding of S.marcescens pathogenicity.     

A liquid-based method for the assessment of bacterial pathogenicity using the nematode Caenorhabditis elegans

Caenorhabditis elegans has previously been used as an alternative to mammalian models of infection with bacterial pathogens. We have developed a liquid-based assay to measure the effect of bacteria on the feeding ability of C. elegans. Using this assay we have shown that Pseudomonas aeruginosa strain PA14, Burkholderia pseudomallei and Yersinia pestis were able to inhibit feeding of C. elegans strain N2. An increase in sensitivity of the assay was achieved by using C. elegans mutant phm-2, in place of the wild-type strain. Using this assay,P. aeruginosa PA01 inhibited the feeding of C. elegans mutant phm-2. Such liquid-based feeding assays are ideally suited to the high-throughput screening of mutants of bacterial pathogens.     

The importance of being regular: Caenorhabditis elegans and Pristionchus pacificus defecation mutants are hypersusceptible to bacterial pathogens

Bacterial pathogens have shaped the evolution and survival of organisms throughout history, but little is known about the evolution of virulence mechanisms and the counteracting defence strategies of host species. The nematode model organisms, Caenorhabditis elegans and Pristionchus pacificus, feed on a wealth of bacteria in their natural soil environment, some of which can cause mortality. Previously, we have shown that these nematodes differ in their susceptibility to a range of human and insect pathogenic bacteria, with P. pacificus showing extreme resistance compared with C. elegans. Here, we isolated 400 strains of Bacillus from soil samples and fed their spores to both nematodes. Spores of six Bacillus strains were found to kill C. elegans but not P. pacificus. While the majority of Bacillus strains are benign to nematodes, observed pathogenicity is restricted to either the spore or the vegetative stage. We used the rapid C. elegans killer strain (Bacillus sp. 142) to conduct a screen for hypersusceptible P. pacificus mutants. Two P. pacificus mutants with severe muscle defects and an extended defecation cycle that die rapidly on Bacillus spores were isolated. These genes were identified to be homologous to C. elegans, unc-22 and unc-13. To test whether a similar relationship between defecation and bacterial pathogenesis exists in C. elegans, we used five known defecation mutants. Quantification of the defecation cycle in mutants also revealed a severe effect on survival in C. elegans. Thus, intestinal peristalsis is critical to nematode health and contributes significantly to survival when fed Gram-positive bacteria.     

Characterization of Burkholderia pseudomallei infection and identification of novel virulence factors using a Caenorhabditis elegans host system

The environmental saphrophyte Burkholderia pseudomallei is the causative agent of melioidosis, a systemic, potentially life-threatening condition endemic to many parts of south-east Asia and northern Australia. We have used the soil nematode Caenorhabditis elegans as a model host to characterize the mechanisms by which this bacterium mounts a successful infection. We find that C. elegans is susceptible to a broad range of Burkholderia species, and that the virulence mechanisms used by this pathogen to kill nematodes may be similar to those used to infect mammals. We also find that the specific dynamics of the C. elegans-B. pseudomallei host-pathogen interaction can be highly influenced by environmental factors, and that nematode killing results at least in part from the presence of a diffusible toxin. Finally, by screening for bacterial mutants attenuated in their ability to kill C. elegans, we genetically identify several new potential virulence factors in B. pseudomallei. The use of C. elegans as a model host should greatly facilitate future investigations into how B. pseudomallei can interact with host organisms.     

A reverse genetic analysis of components of the Toll signaling pathway in Caenorhabditis elegans

BACKGROUND: Both animals and plants respond rapidly to pathogens by inducing the expression of defense-related genes. Whether such an inducible system of innate immunity is present in the model nematode Caenorhabditis elegans is currently an open question. Among conserved signaling pathways important for innate immunity, the Toll pathway is the best characterized. In Drosophila, this pathway also has an essential developmental role. C. elegans possesses structural homologs of components of this pathway, and this observation raises the possibility that a Toll pathway might also function in nematodes to trigger defense mechanisms or to control development. RESULTS: We have generated and characterized deletion mutants for four genes supposed to function in a nematode Toll signaling pathway. These genes are tol-1, trf-1, pik-1, and ikb-1 and are homologous to the Drosophila melanogaster Toll, dTraf, pelle, and cactus genes, respectively. Of these four genes, only tol-1 is required for nematode development. None of them are important for the resistance of C. elegans to a number of pathogens. On the other hand, C. elegans is capable of distinguishing different bacterial species and has a tendency to avoid certain pathogens, including Serratia marcescens. The tol-1 mutants are defective in their avoidance of pathogenic S. marcescens, although other chemosensory behaviors are wild type. CONCLUSIONS: In C. elegans, tol-1 is important for development and pathogen recognition, as is Toll in Drosophila, but remarkably for the latter r?le, it functions in the context of a behavioral mechanism that keeps worms away from potential danger.     

The genetics of pathogen avoidance in Caenorhabditis elegans

Much attention is rightly focused on how microbes cause disease, but they can also affect other aspects of host physiology, including behaviour. Indeed, pathogen avoidance behaviours are seen across animal taxa and are probably of major importance in nature. Here, we review what is known about the molecular genetics underlying pathogen avoidance in the nematode Caenorhabditis elegans. In its natural environment, the soil, this animal feeds on microbes and is continuously exposed to a diverse mix of microorganisms. Nematodes that develop efficient behavioural responses that enhance their attraction to sources of food and avoidance of pathogens will have an evolutionary advantage. C. elegans can specifically detect natural products of bacteria, including surfactants (such as serrawettin) and acylated homoserine lactone autoinducers, and it can learn to avoid pathogenic species. To date, several distinct mechanisms have been shown to be involved in pathogen avoidance. They are based on G protein-like, insulin-like and neuronal serotonin signalling. We discuss recent findings on the mechanisms of pathogen recognition in C. elegans, the relationship between alternative behavioural defences and also between these and other life-history traits. We propose that the selective pressure associated with avoidance behaviours influence both pathogen and host evolution.     

The pursuit of cryptococcal pathogenesis: heterologous hosts and the study of cryptococcal host-pathogen interactions

Analysis of the molecular mechanisms by which a pathogen interacts with the human host is most commonly performed using a mammalian model of infection. However, several virulence-related genes previously shown to be involved in mammalian infection with Cryptococcus neoformans have also been shown to play a role in the interaction of these pathogens with invertebrates, such as Acanthamoeba castellanii, Caenorhabditis elegans, Dictyostelium discoideum, Drosophila melanogaster and Galleria mellonella. The study of host-pathogen interactions using these model hosts has allowed rapid screening of mutant libraries and can be used for the study of evolutionarily preserved aspects of microbial virulence and host response.     

Neural regulation of immunity: Role of NPR-1 in pathogen avoidance and regulation of innate immunity

The nervous and immune systems consist of complex networks that have been known to be closely interrelated. However, given the complexity of the nervous and immune systems of mammals, including humans, the precise mechanisms by which the two systems influence each other remain understudied. To cut through this complexity, we used the nematode Caenorhabditis elegans as a simple system to study the relationship between the immune and nervous systems using sophisticated genetic manipulations. We found that C. elegans mutants in G-protein coupled receptors (GPCRs) expressed in the nervous system exhibit aberrant responses to pathogen infection. The use of different pathogens, different modes of infection, and genome-wide microarrays highlighted the importance of the GPCR NPR-1 in avoidance to certain pathogens and in the regulation of innate immunity. The regulation of innate immunity was found to take place at least in part through a mitogen-activated protein kinase signaling pathway similar to the mammalian p38 MAPK pathway. Here, the results that support the different roles of the NPR-1 neural circuit in the regulation of C. elegans responses to pathogen infection are discussed.     

Yersinia pestis kills Caenorhabditis elegans by a biofilm-independent process that involves novel virulence factors

It is known that Yersinia pestis kills Caenorhabditis elegans by a biofilm-dependent mechanism that is similar to the mechanism used by the pathogen to block food intake in the flea vector. Using Y. pestis KIM 5, which lacks the genes that are required for biofilm formation, we show that Y. pestis can kill C. elegans by a biofilm-independent mechanism that correlates with the accumulation of the pathogen in the intestine. We used this novel Y. pestis-C. elegans pathogenesis system to show that previously known and unknown virulence-related genes are required for full virulence in C. elegans. Six Y. pestis mutants with insertions in genes that are not related to virulence before were isolated using C. elegans. One of the six mutants carried an insertion in a novel virulence gene and showed significantly reduced virulence in a mouse model of Y. pestis pathogenesis. Our results indicate that the Y. pestis-C. elegans pathogenesis system that is described here can be used to identify and study previously uncharacterized Y. pestis gene products required for virulence in mammalian systems.     

A two-gene balance regulates Salmonella typhimurium tolerance in the nematode Caenorhabditis elegans

Lysozymes are antimicrobial enzymes that perform a critical role in resisting infection in a wide-range of eukaryotes. However, using the nematode Caenorhabditis elegans as a model host we now demonstrate that deletion of the protist type lysozyme LYS-7 renders animals susceptible to killing by the fatal fungal human pathogen Cryptococcus neoformans, but, remarkably, enhances tolerance to the enteric bacteria Salmonella Typhimurium. This trade-off in immunological susceptibility in C. elegans is further mediated by the reciprocal activity of lys-7 and the tyrosine kinase abl-1. Together this implies a greater complexity in C. elegans innate immune function than previously thought.