The Nishiguchi Lab Members

Most microorganisms experience variable environments which at least occasionally involve periods of harsh and extreme conditions that impose stress and hinder optimal performance of microbial physiology, metabolism, survival, and fitness. Microorganisms have evolved sophisticated, elaborate, and complex stress responses to regulate and mitigate the negative effects of stress in attempt to maintain homeostasis. The different stress responses (e.g., heat shock and starvation) involve crosstalk to each other and are cross-connected in manifold ways to determinants of symbiosis, defined here to include mutualisms, parasitisms, host-pathogen relationships, and commensalisms. How stress impacts microbial diversity, ecology, and evolution is not entirely clear and is an interesting question.

The sepiolid squid-Vibrio mutualism is a marvelous model system to explore this problem. The aim of this dissertation was to investigate how environmental variation, abiotic factors, exposure to novel hosts, and stressors influence the evolutionary ecology of the symbiont Vibrio fischeri and what the potential consequences are to its free-living and host-associated phases, along with the accompanying overall effect on the sepiolid squid-Vibrio symbiosis itself. Growth curve studies showed temperature, salinity, and nutrient availability can synergistically interact and affect symbiont reproduction, abundance, competition, and allelopathy prior to their encounter with host squid. Furthermore, these results suggested the distribution patterns of the hosts could be a contributor in shaping how V. fischeri biomass production responds to abiotic factors and stress during the planktonic lifestyle.

Subsequent co-infection studies with dissimilar strains in squid showed how proportions of different symbiont types can modify the outcome of host colonization, including the countervailing of competitive dominance. Microbial experimental evolution indicated that V. fischeri is capable of rapid adaptation to environmentally stressful and novel host environments. Both of these perspectives are related, since host immunity imparts its own particular set of advosarial stresses that are themselves well coordinated and integrated into a single vigorous defense. Temporal population genetic studies of V. fischeri isolated from animals provided a macroscale evolutionary viewpoint into the cospeciation of the symbionts with their host squid. Genomic resequencing of experimentally and naturally evolved V. fischeri in E. tasmanica identified genes potentially responsible for the adaptation to this host.
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I am interested in cooperation both within and between species. Cooperative behavior has long posed a special problem for evolutionary theory, though it forms the basis of many of life's major transitions: cohesive genomes, organelles within cells, and multicellularity itself. I am currently working with the squid-Vibrio symbiosis, which has been studied from a medical perspective as a model system of host-bacterial mutualism for over 20 years. This particular mutualism is often mentioned in social evolution literature, but has never been explored in detail from this perspective. My current projects fall into two main categories:
1. Between species: the use of molecular evolution tools to address influence of the mutualistic host on the bacterial partner.
2. Within species: cheating and competitive interactions among Vibrio fischeri strains.
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In nature, most bacteria grow in communities enclosed by a matrix attached to a surface, this microniche is known as a biofilm. This ability to form communities appears to be critical for the environmental survival and host colonization. I am currently exploring what environmental factors affect biofilm formation by Vibrio fischeri isolates and the molecular mechanisms utilized by this bacterium to form and maintain biofilms in order to predict bacterial community dynamics and persistence under extreme habitats which is a dominant factor in controlling squid and Vibrio distribution.
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My current research focus on fish, in the genus Gila, that are endemic to the Colorado River Basin. The Gila robusta complex consists of 7 species. Five of which are federally listed as endangered, one is currently a candidate for listing and the other is undergoing a status review. Over the years there has been taxonomic uncertainty about several of these species. The goals of my project are to: 1) develop definitive set of genetic markers that can be used to identify each species, any hybrids, and allow comparative population assessments 2) Resolve and explore the phylogeny of the G. robusta complex 3) Use landscape genetics to assess how the Colorado River has played a role in the current and historical population structure of the G. robusta complex.
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My name is Laila Rajabi and I am a Master’s student in the Marine Symbiosis Lab of Dr. Michele Nishiguchi. My current project focuses on mechanisms that promote infectious relationships between two organisms. The genus Vibrio consists of disease-causing (pathogenic) as well as mutualistic (non-pathogenic) strains of bacteria. Factors such as biochemical parameters of the environment, specific genes involved in initiating and maintaining symbiosis and how the symbiont and host interact upon infection are just a few key issues that are crucial for the association to be maintained.

My project deals with the mutualistic relationship between sepiolid squids (Cephalopoda: Sepiolidae) and their Vibrio bacteria. Studies have shown that in vibrios, chiP is responsible for expressing an outer membrane chitin oligosaccharide specific porin known as chitoporin (Li et al., 2004). Our lab has previously shown that in Vibrio strain ES114, the chiP gene is only expressed inside the host (Jones et al., 2006). I am using this gene to create viable knock out mutants with disrupted chitoporin function. I predict that chitoporin disruption will decrease survival in the host light organ. These results should provide evidence as to whether or not chitin is being utilized as a biomolecule inside the host light organ for the maintenance of this specific symbiosis.
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Brief Description of Bioinformatics project
for Anne Jacobs
Euprymna scolopes, the Hawaiian bobtail squid, is a small nocturnal (to just 35 mm ml) sepiolid squid indigenous to the Hawaiian Islands and ranges in very shallow water just 2-4 cm deep. E. tasmanica, the southern dumpling squid, is found along the continental shelf surrounding southern Australia.

I am following a couple of genes in three strains of Vibrio fischeri (V. fischeri ES114, V. fischeri 2B1 and V. fischeri ETJB5C). V. fischeri ES114 and possibly V. fischeri MJ11 (possible) shall serve as reference genomes for this project. V. fischeri ES114 was isolated from Euprymna scolopes while V. fischeri ETJB5C was isolated from E. tasmanica. V. fischeri 2B1 is the 400th generation of V. fischeri ES114 (ancestral stain is V. fischeri JRM200).

I am investigating whether or not the environment influenced any changes in these gene sequences. I am also investigating why there would be no changes in these gene sequences if any remain the same in all three strains.
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Grady Easterling works closely with graduate student Will Soto on the abiotic effects on V. fischeri. Grady's project is the experimental evolution of V. fischeri at extreme temperatures.
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Vibrio fischeri can survive harsh environments by forming biofilms. The related changes in gene expression when the bacterium faces extreme conditions (like high salinity, temperature fluctuation and different carbon sources) have not been described. My work focuses on developing a real-time reverse transcription-PCR method to determine whether selective genes are upregulated or downregulated under different environmental stresses.
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Microbiology is becoming increasingly important for solving environmental issues around the world. New technologies based on microbial physiology, such as biofuels and toxicity assays, are being developed to address a wide variety of the issues we face today. My work focuses on the physiology of Vibrio fischeri to develop applications for biofilm formation and toxicity screening of water supplies.
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My work focuses on construction of insertion mutations by using a novel suicide vector specific for Vibrio fischeri (pEVS122) with the purpose of testing for genotypic changes on biofilm formation when different novel genes are disrupted. In addition, I am working on complementing these mutations by developing an operon mobilization technique (gap repair) which uses the Saccharomyces cereviceae recombinatinal machinery to construct Bacterial Artificial Chromosomes and mobilize them by triparental mating.
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Vibrio fischeri is a symbiotic bioluminescent bacterium found in both sepiolid squids and monocentrid fishes V. fischeri colonizes the light organs of sepiolid squids, resulting in a mutualistic relationship in which the squid provides the bacteria with a hospitable microenvironment and the bacteria in return provides the ability for counterillumination as an anti-predatory behavior. The presence of multiple Vibrio populations within and between host squid and is genetically diverse, yet its genetic diversity and spatially distributed populations are not fully implicit. One cause for this difference may be due to bacterial antagonistic interactions such as antibiosis or bacterial allelopathy. Antagonism is a type of interaction in which one bacterial strain produces toxic substances to inhibit similar strains as a competitive mechanism.

Previous research using E. coli as a modeled system shows that this type of antagonistic interaction may be a factor to V. fischeri’s ecological spatial distribution and genetic diversity. Thus, the goal of our study is to understand if such strategies exist within V. fischeri’s genetic breadth, and to identify if biotic factors play a role in the diversification of V. fischeri. Our study also expands to better understand the chemical interactions that Vibrio bacterium possess such as autoinducers which plays a major factor in bioluminescence. By measuring growth and luminescence of specific V. fischeri strains in spent media of competing strains, we hope to identify whether such antagonistic interactions occur prior or during host colonization as well as understand the chemical factors in this bacterium.
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