Vibrio fischeri

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Vibrio fischeri
Scientific classification
Kingdom: Bacteria Phylum: Proteobacteria Class: Gammaproteobacteria Order: Vibrionales Family: Vibrionaceae Genus: Aliivibrio Species: V. fischeri
Binomial name
Vibrio fischeri

Description and significance

Vibrio fischeri is a gram-negative bioluminescent marine bacterium that forms mutualistic symbiosis with various species of fish and squid. V. fischeri is a member of the Vibrionacea family of marine γ-proteobacteria which includes many species that have evolved both beneficial and harmful relationships with animals. The motile bacterium can be found living freely in the oceans or inside the microhabitats of its host's light organs. The proteins necessary for the production of bioluminescence are encoded in a set of genes called the lux operon. The bioluminescent bacteria produces light in a chemical reaction where luciferin, a substrate molecule, is oxidized by an enzyme, luciferase. As a result, energy in the form of blue-green light (480-490nm) is emitted. Bioluminescence and other metabolic functions in V. fischeri are regulated by a cell density dependent system called quorum sensing. Quorum sensing occurs through the production and accumulation of a signal molecule, N-acyl homoserine lactone (AHL); as a consequence individual bacteria do not luminesce. Since its discovery in V. fischeri, quorum sensing and it's respective signal molecules have been shown to regulate genes in many gram-negative and gram-positive bacteria. More importantly, it has been shown that quorum sensing regulates a variety of genes whose proteins are required for virulence factors, symbiosis, biofilm formation, plasmid transfer and morphogenesis.. The symbiotic relationship between a strain of V. fischeri and its host, the Hawaiian bobtail squid Euprymna scolopes, has been studied extensively and represents a model of bacteria -animal symbiosis. Studies on V. fischeri and E. scolopes have provided insights into the mechanisms of colonization by bacteria as well as mechanisms for bacteria/host specificity. Within the genus Vibrios, comparative studies between symbiotic species like V. fischeri and pathogenic species like Vibrio cholerae are being examined to detect patterns and similarities in bacteria/host relationships. The isolation and cloning of the lux genes from V. fischeri, and their use as a reporter, have provided scientists with another valuable visual tool to examine living organisms at the cellular level. Similarly, V. fischeri cells have been made commercially available and are used in the field of ecotoxicology to detect contaminants in the environment.

Bioluminescence

Bioluminescence, the ability of living organisms to produce light has evolved independently multiple times and occurs in a variety of marine species as well as some terrestrial forms. There are various mechanisms leading to light production and different forms of luciferin and luciferase are used but they all require the use of molecular oxygen . In the bacterial bioluminescent pathway, luciferase catalyses the dual oxidation of luciferin, a reduced riboflavin mononucleotide (FMNH₂) and an associated molecule, a long chain aldehyde. ATP is required to catalyze this reaction and the resulting products are light, water, oxyluciferin and carboxyl group. Bacterial luciferase is also required for aerobic respiration and luciferase has a high affinity for molecular oxygen. It is hypothesized that bacterial luciferase evolved from an enzyme whose original function was to provide protection from reactive oxygen species (ROS) in a dark pathway. Wild type bioluminescent bacteria survive UV radiation and hydrogen peroxide better than lux mutants.

Juvenile E. scolopes, symbiotic host of V. fischeri

Ecology

V. fischeri can be located in the microhabitats of its hosts’ light organs, but can also be found living freely as ‘marine snow’, in fecal pellets, as saprohytes , and amongst the microbial flora in the guts of marine animals. The bacterium is distributed in the pelagic zone of temperate and subtropical waters . However, higher cell populations of V. fischeri is achieved when it is associated with its host.

Symbiosis

The symbiosis benefits E.scolopes by using the bioluminescent bacteria in a camouflage strategy called ‘counter-illumination’. The light produced by the bacterium is projected ventrally by the squid, which mimics down dwelling moonlight when viewed from below. The squid effectively projects no shadow, and it can regulate the amount of bacterial illumination with the light organ . The squid provides housing and nutrients, especially carbon and nitrogen, in the form of proteins and peptides inside the crypts of the light organ . In providing a nutrient rich environment, the bacteria population increases to 1.0⁹ cells of which 90-95% are expelled every morning by the squid. The remaining bacteria in the light organ are replenished daily, and the squid becomes a vehicle to produce an abundance of V. fischeri . The symbiotic relationship between strains of V. fischeri and their particular hosts is highly specific . Newly hatched squids are born without the bacterium and must acquire them from the surrounding seawaters. In waters inhabited by E.scolopes, the abundance of V. fischeri constitutes only 0.1% of total microbes per mL of seawater, yet only the specific strain of V. fischeri can effectively colonize and remain in the light organ of the squid. The bacterium is housed inside the crypts of the light organ located in the squid’s mantle cavity. Once the bacterium is acquired and colonizes the crypts, morphogenesis occurs for both the bacterium and its host. The bacteria lose the flagella, and the cells decrease in size. As for the host, the ciliated fields used to obtain the bacterium undergo programmed cell death, and the light organ swells Colonization by the bacterium concurrently alters gene expression in the squid and the bacterium.

Inoculation pathway of V. fischeri into light organ of juvenile E. scolopes

Genome

To date, the complete genomes of two strains of V. fischeri, ES114 and MJII have been sequenced. MJ11 is isolated from the japanese pinecone fish, Monocentris japonica. Strain ES114 isolated from E.scolopes contains two circular chromosomes and a circular plasmid pES100. The genetic material is composed of DNA and the total genome is 4.284 Mbp in length. Chromosome 1, the larger of the two chromosomes contains 2586 genes, while Chromosome 2 and the plasmid contains 1175 and 57 genes respectively. The lux operon system, located on chromosome 2, contains luxI, luxR and luxCDABEG. LuxI encodes for AHL synthase, the enzyme that produces the autoinducer AHL. The Lux R gene produces the AHL dependent transcriptional activator protein that binds to the AHL and promotes the transcription of the lux operon. Luciferase, the enzyme responsible for catalyzing the light reaction, is encoded by luxA and lux B. LuxA produces the alpha chain subunit and luxB, the beta subunit of luciferase. LuxCDE are required for aldehyde synthesis (BioUK). LuxG encodes flavin mononucleotide reductase.

Pathology

V. fischeri is non-pathogenic to humans but is pathogenic to some marine invertebrates. Within the Vibrios family there are three human pathogens V. cholerae, V.parahaemolytus, V. vulnificus.

Application to Biotechnology

The lux genes isolated from various bioluminescent organisms, in combination with the use of recombinant DNA technology, have had wide applications in the field of science. Applications for bioluminescent genes have reinforced the many innovative applications for green fluorescent protein (GFP), which is isolated from jellyfish. Differences in specific applications have to do with the different chemistries of light production. One of the primary uses of bioluminescent genes is as a reporter gene. A reporter gene allows scientists to visually track proteins and molecular processes occurring inside of living organisms. Scientists use reporter genes to uncover various characteristics of proteins, determine the functions of genes, and to describe regulatory regions of the genome (methods Gheysens). In clinical research, bioluminescence and fluorescence imaging is used to observe physiological changes during disease progression in patients and to observe how disease alters molecular processes in patients. The V. fischeri bacteria and its isolated lux operon are being used to detect pollution in the environment. In using the whole-cell approach, non-specific contamination is detected by the decrease in cell luminescence brought upon by cell death or metabolic failure. Methods to detect for specific contaminants utilize the lux operon and a regulatory region that is specific to the target compound.

Current Research

Dr. E.G. Ruby has been studying V. fischeri in the symbiosis model to provide insight into the molecular and biochemical pathways of pathogenic bacterial colonization of animal tissue. In his recent paper, he and his colleagues found that a regulatory gene in V. fischeri, the rscS, was necessary for establishment for symbiosis by V. fischeri and its host E.scolopes. Comparative genomic studies of the rscS gene in different strains of V. fischeri isolated from squid and fish hosts revealed that the squid isolates all had a conserved rscS gene. From the fish isolates, five out of the ten fish also had a conserved rscS gene, but highly divergent from the squid isolates. In the experiment, the rscs gene was inserted into the MJ11 strains isolated from fish that cannot effectively colonize squid. With the transformation, the MJ11 strains were able to colonize the light organ as effectively as the natural symbionts.

References

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Herring, P.J. and Widder, E.A.2001. Bioluminescence in Plankton and Nekton. In: Steele, J.H., Thorpe, S.A. and Turekian, K.K. editors, Encyclopedia of Ocean Science, Vol. 1, 308-317. Academic Press, San Diego. Accessed from "International society of bioluminescence and chemiluminescence", <http://www.isbc.unibo.it/Files/BC_PlanktonNekton.htm> (updated 10/2009; accessed 03/20/09)

McFall-Ngai, M. J. 2000. Negotiations between animals and bacteria: the 'diplomacy' of the squid-vibrio symbiosis. Comp. Biochem. Physiol. A. Mol. Integr. Physiol., 126(4), 471-480.

NCBI. Entrez genome project. <http://www.ncbi.nlm.nih.gov/sites/entrez?Db=genomeprj&Cmd=Retrieve&list_uids=12986>(updated 2/09;accessed 03/29/09)

Ruby, E. G., M. Urbanowski, J. Campbell, A. Dunn, M. Faini, R. Gunsalus, P. Lostroh, C. Lupp, J. McCann, D. Millikan, A. Schaefer, E. Stabb, A. Stevens, K. Visick, C. Whistler, and E. P. Greenberg. 2005. Complete genome sequence of Vibrio fischeri: a symbiotic bacterium with pathogenic congeners. Proc. Natl. Acad. Sci. USA 102:3004-3009

Ruby, E. G., McFall-Ngai, M. J. Oxygen-utilizing reactions and symbiotic colonization of the squid light organ by Vibrio fischeri. Trends in microbiology. 415 Volume 7, issue , 10October 1999, Pages 414-420. Accessed from ScienceDirect

Thompson, F.L., Austin, B., Swings, J. The biology of Vibrios. 2006. ASM press. Virginia.

Visick, K., Ruby, E. G. Vibrio fischeri and its host: it takes two to tango. Current Opinion in Microbiology. Volume 9, Issue 6, December 2006, Pages 632-638. Accessed from ScienceDirect.

Willey, J.M., Sherwood, L.M., Woolverton, C. J. Prescott, Harley, and Klein's Microbiology. 7th Ed. 2008. McGraw-Hill. New York.