- Biology and Biochemistry

Research
Nick Waterfield
Nick Waterfield

Research Profiles

Nick Waterfield

Invertebrates and the evolution of bacterial pathogenicity

Biography

e-mail:bssnw@bath.ac.uk

Current Research

It is only recently that the important relationship between pathogens of vertebrates and invertebrates has been appreciated. Conservation of key features of the innate immune responses of invertebrates such as insects, and mammals suggests that universal offensive strategies can be employed by bacterial pathogens of these two groups. This observation is vital to the understanding of the evolution of bacterial virulence as the innate response is arguably the most important aspect of immunity in all animals, including mammals. Given the age of the invertebrate lineages, the enormous number of different species and individuals, and the diversity of the lifestyles they employ it seems likely that they have provided both a reservoir and a vector for the flow of horizontally transferred virulence factors between pathogens of very different host organisms.

Many important mammalian pathogens associate with or parasitise invertebrate hosts, such as the plague bacillus Yersinia pestis in the flea, and the Lyme disease spirochete Borrelia, in ticks. Furthermore, it has been suggested that only minor molecular adaptations of bacterial pathogens may give rise to major changes in host range. Examples again include Y. pestis, which appears to have recently evolved from an environmental Y. psuedotuberculosis-like ancestor and the Anthrax agent, Bacillus anthracis, which is very closely related to the important insect associated pathogens B. cereus and B. thuringiensis. The demonstration of high frequency conjugative transfer of antibiotic resistance genes from Escherichia coli to Y. pestis within the gut of the flea host provides one such example of the role insects may play in the evolution of mammalian pathogenicity.

Nevertheless, I believe a more important aspect of this relationship lies in the fact that invertebrate pathogens provide a massive reservoir of bacterial pathogens that are pre-adapted to a basic innate-immune response. In addition, it is clear from comparisons of the Y. pestis with Y. psuedotuberculosis genomes that the need to be transmitted by the invertebrate flea vector has sculpted the genome of Y. pestis into a form that is highly virulent to man.

Goals

To characterise the genes and processes important in insect and human pathogenic variants of Photorhabdus. Comparative genomics provides an excellent opportunity to begin dissecting the genes important in these diseases.

Photorhabdus

"Glowing Rods"
Insect Pathogen

Photorhabdus are the only known terrestrial bioluminescent bacteria. Most members of the Photorhabdus are however insect pathogens that live in a strict symbiotic relationship within the guts entomopathogenic Heterorhabditid nematodes. Infective juvenile nematodes search in the soil for insect prey until they encounter a suitable host. They then scratch their way into the insect's hemocoel (an "open" blood system ) and "vomit" up Photorhabdus cells directly into the blood. The Photorhabdus then set up a lethal septicemia, secreting toxins and virulence factors that rapidly kill the insect host. The bacteria replicate rapidly and bio-convert the insect tissues into more bacteria that serve as a food source for the reproducing nematodes. It is around the time of insect death that the bioluminescence of the insect corpse can be seen.


Manduca sexta glowing with Photorhabditis

Infective nematodes "burst" free

 

Bioluminescence is an oxygen and energy costly process and as yet we have no good explanation as to why they do this. Theories include some unknown biochemical role, a warning to scavenging nocturnal mammals or even that it serves as a lure to tempt fresh insect victims into range. When the insect resources have been exhausted, the bacteria provide the nematode with an unknown "food signal" which switches them into a developmental state known as an infective juvenile. At this point they re-package the bacteria before bursting from the insect corpse in search of fresh victims. Insects provide ideal host systems to study the interaction between bacterial pathogen and animal host.

 

Photorhabdus luminescens strain W14 on a insect midgut under the collagen sheath.

Immunogold conjugates may be used to reveal the expression of specific virulence factors in sectioned insects. Below left shows the location of the TcaC toxin on Photorhabdus luminescens strain W14 cells during infection . Below right shows Photorhabdus expressing the Jellyfish green fluorescent protein attaching to the insect midgut.

 

Human Pathogen

While Photorhabdus have never been isolated as free living in the environment, an increasing number of clinical isolates, designated Photorhabdus asymbiotica, are being identified from human patients in the United States, Australia and recently Nepal. Clinical collaborators suggest that many other cases are misdiagnosed due to the failure of clinical microbiology laboratories to recognise this unexpected organism.

 

Cases occur in warm wet months, usually after rain storms, and on extremities such as the feet. P. asymbiotica is associated with severe soft tissue and systemic infections, and has been considered an "emerging human pathogen". We have recently discovered that P. asymbiotica strains (at least in Australia) are vectored by a heterorhabditis-like nematode which can predate insects in the laboratory in a life cycle similar to that of the well characterized EPN strain complexes. We don't yet know if these P. asymbiotica EPN complexes deliberately seek out mammalian skin in the environment, although genetic and biological analysis of P. asymbiotica ATCC43949 does suggest it is well adapted for survival, at least in humans.

 

Images of a Photorhabdus asymbiotica infection kindly provided by Dr John Gerrard (Goldcoast Hospital, Queensland)

We are currently sequencing the genome of the human pathogen Photorhabdus asymbiotica ATCC43949 in collaboration with the Sanger center UK.

A comparison between the obligate insect pathogen P. luminescens and the closely related human pathogen P. asymbiotica provides a safe, inexpensive and experimentally tractable model organism for studying the close relationship between insect and human disease processes. Furthermore, an understanding of how Photorhabdus evades the germ-line immunity of insects is beginning to highlight common themes in ancient humoral responses such as complement (in vertebrates) and melanisation (in insects).

http://www.sanger.ac.uk/Projects/P_asymbiotica/

Selected publications

Submitted

Published