Bonnie E. Cone Distinguished Professor

Ecology, physiology, and metabolism of aquatic bacteria

Biology of the human pathogen,Vibrio vulnificus.

Academic Degrees and Honors

• B.S.   Microbiology, University of Arizona (1968)
• Ph.D. Microbiology, Georgetown University (1973)
• National Research Council (Canada) Postdoctoral Fellowship (1973-1974)
• Academy of Sciences Exchange Scientist, Romania (1982)
• First Citizens Bank Scholar Award for Excellence in Research (1988)
• Fellow, American Academy of Microbiology (1990-)
• Member, Phi Kappa Phi (Academic Excellence), Phi Beta Delta (Honor Society for International Scholars)
• Visiting Professor, University of Göteborg, Sweden (1990)
• Visiting Professor, Duke University Marine Laboratory (1991, 1992)
• Member, Editorial Board, Applied and Environmental Microbiology (1988-1993)
• Visiting Professor, North Carolina State University (1994,1995,1996)
• Bonnie E. Cone Distinguished Professor for Teaching (1998)
• Visiting Professor, Royal Veterinary and Agricultural University, Copenhagen, Denmark (1998)
• Burroughs Welcome Fund Visiting Professor in the Microbiological Sciences (1999)
• Member, Standard Methods Committee, Standard Council, American Water Works Association (2000-)
• Member, Working Party on Culture Media, International Committee on Food Microbiology and Hygiene, International Union of Microbiological Sciences (2000-)
• Harshini V. de Silva Graduate Student Mentoring Award (2002)
• Senior Faculty Fellow, Global Institute for Energy and Environmental Systems (2001-)
• Member, Editorial Board, FEMS Microbiology Ecology (2005-
• Visiting Professor National University of Ireland, Galway (2006)
• First “Jay and Beverly Grimes Distinguished Lecturer”, Gulf Coast Research Lab, Univ. Southern Mississippi, 2007
• Visiting Professor (Sabbatical) University of Aberdeen, Scotland (2008)
• Member, Editorial Board, Advances in Microbiology (2008-)
• Member, Editorial Board, Advanced Studies in Biology (2009-)
• Member, Working Group on Vibrio Taxonomy, Subcommittee on the Taxonomy of Aeromonadaceae, Vibrionaceae and related organisms, International Committee on Systematics of Prokaryotes (2009-)

Courses Taught

• BIOL 4250 Microbiology
• BIOL 4257 Microbial Physiology and Metabolism

Areas of Research

The major areas of study in my laboratory are Vibrio vulnificus and other pathogenic marine Vibrio spp., the “viable but nonculturable” state, and bacterial stress responses and their relationship to survival and virulence.

Vibrio vulnificus

First described in 1976, Vibrio vulnificus is a halophilic bacterial species that causes primary septicemia and wound infections (Fig. 1). Disease in humans results from contamination of a skin lesion or ingestion of contaminated seafood. A striking feature is that many victims die with a few hours of symptom development) and the high mortality rate (>50% and 25%, respectively). The infectious dose of V. vulnificus is believed to be low and fatal infections have been reported after the ingestion of a single oyster (the primary source in 93% of all cases). Fatal wound infections acquired by contamination of ant bite lesions have also been reported. Between 1989 and 2008, 520 ingestion cases with 263 deaths (51%) were reported. Most cases (>85%) occur in males over the age of 40 (Fig. 2), as estrogen appears to block the endotoxin produced by this species. This is an opportunistic pathogen, with infections largely restricted (>95% of cases) to those having underlying liver diseases (such as liver cirrhosis or hepatitis). Three biotypes of V. vulnificus exist, and among biotype 1, the primary type in human infections, several very distinct genotypes are known to occur.  One area of our study involves these genotypes.  The “C-genotype” correlates strongly with a clinical origin, and these strains appear to be highly virulent. In contrast, the “E-genotype” correlates with isolation from the environment, and appears to be of lower virulence.  At the DNA level, the two genotypes differ significantly from each other (some genes differ in >30% of their sequence), yet the two genotypes are highly similar within the genotype (Fig. 3). One enigma we are investigating is a result of our observation that, while the distribution of the two genotypes in estuarine waters appears to be approximately equal (Fig. 4), the oysters that inhabit those waters carry a high proportion (ca 85:15) of the E-genotype.

Some Recent Papers on this Topic:

• Vibrio vulnificus. 2006.  Oliver, J.D. pp. 349-366 In: Biology of Vibrios.  F.L.Thompson, B. Austin, and J. Swing (eds).  Amer. Soc. Microbiol. Press, Washington, D.C.
• Vibrio vulnificus.  2006. Oliver, J.D. In: Oceans and Health: Pathogens in the Marine Environment. (pp. 253-276). S. Belkin and R.R. Colwell (eds.). Springer Science, New York.
• Wound infections caused by Vibrio vulnificus and other marine bacteria. 2005. Oliver, J.D.  Epidemiol. Infect. 133:383-391.
• Vibrio vulnificus: Disease and pathogenesis.  2009. Jones, M.K. and J.D. Oliver. Infect. Immun. 77:1723-1733.
• Role of iron in human serum resistance of the clinical and environmental Vibrio vulnificus genotypes. 2007. Bogard, R. and J.D. Oliver. Appl. Environ. Microbiol. 73:7501-7505.
• Essential role for estrogen in protection against Vibrio vulnificus induced endotoxic shock.  2001. Merkel, S.M., S. Alexander, J.D. Oliver, and Y.M. Huet-Hudson.  Infect. Immun. 69:6119-6122.
• Ecology of Vibrio vulnificus in estuarine waters of eastern North Carolina. 2003. Pfeffer, C.S., M.F. Hite and J.D. Oliver.  Appl. Environ. Microbiol. 69:3526-3531.
• A rapid and simple PCR analysis indicates there are two subgroups of Vibrio vulnificus which correlate with clinical or environmental isolation. 2005. Rosche, T.M., Y. Yano, and J.D. Oliver. Microbiol. Immunol. 49:381-389.
• Population structure of two genotypes of Vibrio vulnificus in oysters (Crassostrea virginica) and sea water. 2008. Warner, E.B. and J.D. Oliver. Appl. Environ. Microbiol. 74:80-85.

Bacterial Stress Responses

Many of the so-called heat shock proteins, such as chaperone proteins (e.g. DnaK and GroEL) and proteases (e.g. Clp, Lon), are also induced by other environmental changes, such as high osmolarity, nutrient starvation, exposure to low temperature, the presence of heavy metals, oxidative agents or pollutants, and interaction with eukaryotic hosts. Therefore, the heat shock response can more accurately be considered a general stress response. Through a process termed cross protection, this response improves tolerance to otherwise lethal temperatures, high or low salt levels, heavy metals, UV exposure, and starvation, and plays a critical role in bacterial pathogenesis. Such stress responses are critical for bacterial adaptation to the constant changes in the environment experienced by estuarine bacteria, such as most Vibrio spp., and are therefore a major link between microbial ecology, physiology, and microbial pathogenesis. One example is the htrA system, the product of which is essential for bacterial growth only at elevated (e.g. human body) temperatures. This system is activated by the alternate sigma factor, σ^E, encoded by the rpoE gene. This gene has been shown to control mucoidy in cystic fibrosis isolates of Pseudomonas aeruginosa. In E. coli, transcriptional activation of the heat shock genes is induced by the alternate sigma factor, σ³² (product of the rpoH gene). Our lab is interested in the bacterial response to a variety of environmental stresses, especially starvation, low temperature, high salinity (osmotic stress), and oxidative (e.g. peroxide) stress. The bacterium under greatest investigation I this lab is V. vulnificus. We study the mechanisms by which this pathogen survives in the estuarine environment, the molecular regulatory mechanisms which occur during these survival responses. Of special interest in this regard is the use of membrane diffusion chambers to examine in situ gene expression by cells in their natural aquatic environment.

Some Recent Papers on this Topic:

• RpoS involvement in osmotically-induced cross protection in Vibrio vulnificus.  2005. Rosche, T.M., T.C. Bates, D.J. Smith, E.E. Parker, and J.D. Oliver.  FEMS. Microbiol. Ecol. 53:455-462.
• Survival and in situ gene expression of Vibrio vulnificus at varying salinities in estuarine environments. 2008. Jones, M.K., E. Warner, and J.D. Oliver. Appl. Environ. Microbiol. 74:182-187.

The Viable but Nonculturable State

Microbial ecologists have long recognized that large proportions of the microbial populations inhabiting natural habitats appear to be nonculturable. Indeed, plate counts of bacteria in soil, rivers and oceans typically indicate that far less than 1% of the total bacteria observed by direct microscopic examination can be grown on culture media.  Further, it has also long been known that certain portions of bacterial populations in natural environments seem to disappear during certain seasons, only to reappear at other times.  We now understand that at least part of the explanation for these observations is not due to seasonal die-off of the cells, but to their entry into a physiological state most commonly called the “viable but nonculturable” state.

A bacterial cell in the viable but nonculturable (VBNC) state may be defined as one which fails to grow on the routine bacteriological media on which it would normally grow and develop into a colony, but which is in fact alive and metabolically active. Bacteria enter into this dormant state in response to one or more environmental stresses which might otherwise be lethal to the cell. Thus, the VBNC state should be considered a means of cell survival. Eventually, when the inducing stress is removed, these cells are able to emerge from the VBNC state and again become culturable on routine media.

The typical VBNC response is seen in this figure, which shows the response of Vibrio vulnificus to exposure to low temperature (5^oC). Such a temperature is below that at which this aquatic bacterium can grow and, if it were not for the VBNC response, is a temperature which would eventually lead to death of the population.

As is evident from the figure, cells lose their ability to be cultured (open squares) in a rather linear manner, eventually reaching a point where plate counts suggest a total lack of any living cells. However, whereas death of a bacterial population generally leads to lysis of the cells and loss of cell structure, direct examination of cells entering the VBNC state indicates that the cells remain intact (closed symbols). Such cells could, of course, have died, but simply not undergone lysis. The primary evidence that such cells are alive, even if nonculturable, is from data obtained when a “direct viability” assay is applied to such cultures (Fig. 2), or continued production of mRNA is detected. Such assays allow direct determination of individual cell viability, without the need for culture. As seen in the figure (open circles), such assays generally indicate that a large portion of the apparently dead population is, in fact, alive.

Cells entering the VBNC state generally undergo a reduction in size, and during this time, significant changes in membrane structure, protein composition, ribosomal content, and possibly even DNA arrangement are experienced. However, decreases macromolecular synthesis do not mean that all synthesis has ceased.  Indeed, protein synthesis appears to be essential for entry into this state, and under these conditions V. vulnificus produces some 40 new proteins not seen during growth at “normal” temperatures. At the same time, dramatic decreases in membrane fatty acid composition, and in nutrient transport and respiration rates, have generally been reported to occur as cells enter this dormant state. Cell wall synthesis, or at least metabolism of the constituents of these structures, also apparently continues.

Some Recent Papers on this Topic:

• Recent findings on the viable but nonculturable state in pathogenic bacteria. 2009. Oliver, J.D. FEMS Microbiology Rev. DOI:10.1111/j.1574-6976.2009.00200.x
• Role of catalase and oxyR in the viable but nonculturable state of Vibrio vulnificus.  2004. Kong, I.-S., T.C. Bates, A. Hülsmann, H. Hassan, and J.D. Oliver.  FEMS Microbiol. Ecol. 50:133-142.
• Effect of weak acids on Listeria monocytogenes survival: evidence for a viable but-nonculturable state in response to low pH.  2009. Cunningham, E., C. O’Byrne, and J.D. Oliver. J. Food Control 20:1141-1144.
• In situ and in vitro gene expression by Vibrio vulnificus during entry into, persistence within, and resuscitation from the viable but nonculturable state.  2006. Smith, B.E. and J.D. Oliver. Appl. Environ. Microbiol. 72:1445-1451.
• Changes in membrane fatty acid composition during entry of Vibrio vulnificus in the viable but nonculturable state. 2004. Day, A.P. and J.D. Oliver.  J. Microbiol. 42:69-73.
• The viable but nonculturable state in bacteria. 2005. Oliver, J.D. J. Microbiol. 43:93-100.

Brett Froelich (PhD Candidate)

Traditionally found in Gulf, East, and Pacific coast estuaries, Vibrio vulnificus is responsible for 95% of seafood-related deaths in the U.S. Most of these infections result from the consumption of raw or undercooked oysters that contain V. vulnificus cells.  While the concentration of V. vulnificus in seawater is relatively low, filter-feeding shellfish, such as oysters, can concentrate them to >10⁶ CFU/g. Vibrio vulnificus can be separated into three biotypes, with biotype I being of greatest health concern.  This biotype can be further divided into two genotypes, a C-genotype corresponding highly with clinical isolation, and an E-type that corresponds with environmental isolation.  Our lab has previously shown that while the ratio of C and E-types in seawater is nearly even, isolates recovered from oysters are predominately (>80%) E-type, a disparity we have recently shown to be unexplained by uptake or depuration of planktonic V. vulnificus cells (see uptake/depuration curves at left).  Filter-feeding bivalves, such as the Eastern Oyster, Crassostrea virginica, have particle capture efficiencies that vary depending on particle size.  C. virginica retains 90% of particles 5µm in diameter, yet only 5% of free bacteria.  However bacteria are rarely planktonic, and prefer attachment to surfaces.  Marine aggregates, also known as marine snow, naturally form in aquatic environments when smaller particles flocculate into larger particles.  Smaller particles integrated into these aggregates have been shown to exhibit increased uptake and retention rates in oysters.  My work involves the in vitro incorporation of V. vulnificus cells (of both C- and E-genotypes) into laboratory created marine aggregates, and observing the uptake and depuration of these cells into oysters, via dissection and plating onto selective media.

Naturally, C-type strains that cause human infection must have originated from environmental sources, such as oysters or estuarine water (the major reservoirs of V. vulnificus).  I have found that mannitol fermentation is highly correlated with C-type strains of this bacterium. I am currently attempting to show that that C-type strains can be divided into two subgroups, where only one of which appears to be infectious, based on the arrangement of genes around, and including the mannitol fermentation operon (100% C-genotype positive vs 0% of the E-genotype).

Tiffany Williams (PhD Candidate)

My studies involve examining differences in in situ gene expression between the clinical and environmental genotypes of V. vulnificus. Specifically, I am trying to elucidate the distribution anomaly between the C- and E- genotypes in the aquatic environment. To do this, I am examining in situ gene expression of several attachment, stress, and putative virulence genes in a natural estuarine environment by C- and E-strains. These studies are carried out using membrane diffusion chambers, an apparatus that contains pores which hold the bacterial cells inside but still exposes them to the aquatic environment. These chambers are suspended in the water column over a period of time in which each strain is analyzed periodically for culturability, using a plate count method, and gene expression using RT-PCR. Most recently, I have been utilizing an indoor aquarium tank setup in which I compare gene expression of C- and E-genotypes in the presence and absence of oysters.

In addition to this, I have also been working on collaborative studies with Dr. Matt Parrow of this Department, investigating quorum sensing interactions between microalgae and associated marine bacteria. Specifically, we have investigated the potential for select harmful algal species to produce autoinducers (or AI inhibitors) involved in quorum sensing. To detect the presence of AI-2 inducers or inhibitors, we use a Vibrio harveyi bioluminescence assay. Future studies will employ Chromobacterium violaceum and Agrobacterium tumefaciens reporter assays to detect the presence of HSL/AHL (AI-1) quorum sensing molecules.

Kristi Doyle (MS Candidate)

My research focuses on the human pathogen, Vibrio vulnificus ,which the leading cause of all seafood related deaths in the United States. V. vulnificus is a natural inhabitant of the eastern oyster, Crassostrea virginica. There are two genotypes of V. vulnificus; one which is associated with human virulence and is correlated with clinical isolation (C-type) and one which correlates with environmental isolation (E-type). To date, there has been no studies reported elucidating the association between these two genotypes and oysters, and especially why the E-genotype predominates in oysters when the two genotypes are in equal numbers in surrounding waters. Thus, the aim of my first project is to determine if there is preferential uptake of the E-genotype by oysters. The second project that I am currently working on involves the long term starvation effects on V. vulnificus. This bacterium is a natural inhabitant of estuarine waters, which is a nutrition limited, oligotrophic environment. However, there have not been any data published regarding the survival of V. vulnificus in seawater over great lengths of time. Thus, this aspect of my research focuses on examining several phenotypic and biochemical properties of V. vulnificus cells which are subjected to long-term (up to 30 months) starvation conditions in artificial seawater. In addition, I am examining which genes are expressed during the starvation response. These results will be examined to ascertain if there are any phenotypic/biochemical/gene expression differences between the two genotypes.

Eric A. Binder (MS Candidate)

My research is focused on the autoinducer-2 (AI-2) quorum sensing molecule produced by the bacterial pathogen, Vibrio vulnificus.  AI-2 has been shown to be an inter-species communication molecule that is concentration dependent.  In V. vulnificus, AI-2 is involved in the regulation of several virulence factors, as well as motility and attachment. I’m investigating the levels of AI-2 under different environmentally-relevant parameters such as salinity and temperature, and following various gene mutations.  At present I am focusing on how AI-2 production is effected by the varying salinity concentrations that V. vulnificus encounters in its estuarine environment. I have observed that AI-2 production in V. vulnificus increases proportionally as salinity increases from 1/8x – 1x artificial seawater.

In addition, I am examining an AI-2 mutant strain, JDO1. Using this mutant and its compliment, JDO2, I am observing attachment and biofilm formation.

Casey Taylor (MS Candidate)

My work focuses mainly on the ability of V. vulnificus to break down and utilize the components of the mucin that is commonly found in animal’s gut.  Mucins are highly glycosylated, large glycoproteins that are components of tissue-lining mucus. The ability to use mucus is an important trait in the survival, growth, and attachment of pathogens to host organisms. I use cultural techniques to monitor the extent to which V. vulnificus can break down mucin proteins. An extension of this is a study on the breakdown and utilization of the nine-carbon sugars known as sialic acids. My work primarily deals with N-acetylneuraminic acid (Nan), which is s a component of mucin proteins. Sialic acids are typically found at the terminal ends of the carbohydrate side chains of these proteins. There is a cluster of genes, including nanA, nanE, and nanK, whose function is to break Nan down to a byproduct that can be directly inserted into the respiration cycle. I am particularly interested in NanA and I use PCR and gel electrophoresis to detect the presence of the nanA gene (responsible for the catabolism of N-acetylneuraminic acid) in V. vulnificus. I intend to compare the results of the C- and E- strains to determine if there is a correlation which could provide some explanation as to why E strains are found predominately in oysters, while the C strains are found predominantly in human infections.

V. vulnificus is motile by way of a single, polar flagellum. Because motility is known to be a virulence factor in this pathogen, it is of particular interest to determine whether there is a difference in the motility of C and E strains of this organism. Any differences in motility could account for the difference in numbers of these two genotypes found in oysters and their surrounding waters. I use a specialized medium to allow the bacteria to swarm out from the inoculation site, and measure the distance that they swim. Part of this research also includes observations on whether or not salt levels have any effect on the motility levels of this organism; salinity has been observed to have an effect on quorum-sensing, which is also involved in regulating motility.

Joanna Nowakowska (MS Candidate)

I am investigating attachment to chitin by Vibrio vulnificus.  Chitin is composed of β-1,4–linked units of N-acetylglucosamine, and this polysaccharide is present in large amounts in the ocean. It appears as both free particles as well as being a major component of the exoskeletons of copepods, crustaceans, and some algae.

Attachment by bacteria is a crucial step in the colonization of host tissues. It is known that bacteria are able to attach to of chitin and to survive longer in often stressful oceanic conditions. It has been found that bacterial adhesion to this complex polysaccharide surface is a very specific process involving CBPs (chitin binding proteins) which are located in the outer membrane and in some cases this adhesion is arbitrated by sugar-binding proteins termed lectins (Montgomery et al., 1993). It also has been found that adhesion of some Vibrio spp. to chitin decreases with increasing salinity (Kaneko et al., 1975). The salinity of the seawater is higher (around 35 parts per thousand) than in estuaries which often vary considerably.  Salinity levels in estuaries usually decline during cold months and increase during summer. These observations are essential in terms of distribution of pathogenic microorganisms, including several Vibrio spp., which live in these waters.  My studies will test if there are differences in attachment to chitin between the clinical and environmental genotypes of V. vulnificus, as well as possible variations in adhesion between opaque (encapsulated) and translucent (non-encapsulated) forms of this pathogen. This information could be useful in understanding the ecology and pathogenicity of this bacterium.

Alexandria Reinhart (Honors Undergraduate)

My research centers on the difference in biofilm formation between different strains of Vibrio vulnificus, and on gene expression in V. vulnificus cells when in a biofilm. I am also currently researching the differences in biofilm formation of C versus E genotypes of V. vulnificus, and its opaque (encapsulated) versus translucent (non-encapsulated) strains.  This information will be compared to expression of a variety of relevant genes present in different V. vulnificus strains when present in a biofilm.

Recent Publications (2000-2005)

• Effect of starvation and the viable but nonculturable state on green fluorescent protein (GFP) in GFP-tagged Pseudomonas fluorescens. 2000. Lowder, M.A., A. Unge, N. Maraha, J.K. Jansson, J. Swiggertt, and J.D. Oliver. Appl. Environ. Microbiol. 66:3160-3165.
• The viable but nonculturable state and cellular resuscitation. 2000. Oliver, J.D. Intern. Symp. Microb. Ecol. 723-730.
• The public health significance of viable but nonculturable bacteria. 2000. Oliver, J.D. In: "Nonculturable Microorganisms in the Environment", R.R. Colwell and D.J. Grimes (ed.). Amer. Soc. Microbiol. Press, Washington, D.C.
• Problems in detecting dormant (VBNC) cells and the role of DNA elements in this response. 2000. Oliver, J.D. pp. 1-15, In: Marker and Reporter Genes, J.K. Jansson, J.D. van Elsas, and M.J. Bailey (eds.). Landes Biosciences, Georgetown, TX.
• Culture media for the isolation and enumeration of pathogenic Vibrio species in foods and environmental samples. 2001. Oliver, J.D. In: Culture Media for Food Microbiology, 2nd Rev. Ed. J.E.L. Corry, G.D.W. Curtis, and R.M. Baird (eds.), Elsevier Science, Netherlands.
• Essential role for estrogen in protection against Vibrio vulnificus induced endotoxic shock. 2001. Merkel, S.M., S. Alexander, J.D. Oliver, and Y.M. Huet-Hudson. Infect. Immun. 69:6119-6122.
• The use of GFP as a reporter for metabolic activity in Pseudomonas putida. 2001. Lowder, M. and J.D. Oliver. Microbiol. Ecol. 41:310-313.
• Vibrio species. Oliver, J.D. and J. Kaper. 2001. pp. 263-300 In: Food Microbiology: Fundamentals and Frontiers, 2nd ed. M.P. Doyle, L.R. Beuchat, T.J. Montville (ed.). Amer. Soc. Microbiol.
• Effects of refrigeration and alcohol on the load of Aeromonas hydrophila in oysters. 2002. Birkenhauer, J.B. and J.D. Oliver. J. Food Protect. 65:560–562.
• Use of diacetyl to reduce the load of Vibrio vulnificus in the Eastern oyster, Crassostrea virginica. 2003. Birkenhauer, J.B. and J.D. Oliver. J. Food Protect. 66:38-43.
• A comparison of thiosulphate-citrate-bile salts-sucrose (TCBS) agar and thiosulphate-chloride-iodide (TCI) agar for the isolation of Vibrio species from estuarine environments. 2003. Pfeffer, C. and J.D. Oliver. Lett. Appl. Microbiol. 36:150-151.
• Analysis of Vibrio vulnificus from market oysters and septicemia cases for virulence markers. 2003. DePaola, A. et al. (11 authors). Appl. Environ. Microbiol. 69: 4006-4011.
• The ecology of Vibrio vulnificus in estuarine waters of eastern North Carolina. 2003. Pfeffer, C.S. and J.D. Oliver. Appl. Environ. Microbiol. 69:3526-3531.
• Culture media for the isolation and enumeration of pathogenic Vibrio species in foods and environmental samples. 2003. Oliver, J.D. pp. 249-269 In: Handbook of Culture media for Food Microbiology, 2nd ed. J.E.L. Corry, G.D.W. Curtis, and R.M. Baird (eds.). Vol. 37 of Progress in Industrial Microbiology. Elsevier. Amsterdam.
• RpoS-dependent stress response and exoenzyme production in Vibrio vulnificus. Huelsmann, A., T.M. Rosche, I.-S. Kong, H.M. Hassan, D.M Beam, and J.D. Oliver. Appl. Environ. Microbiol. 69:6114-6120.
• Survival of Helicobacter pylori in a natural freshwater environment. 2003. Adams, B.L., T.C. Bates, and J.D. Oliver. Appl. Environ. Microbiol. 69:7462-7466.
• Effects of temperature on detection of plasmid or chromosomally encoded gfp- and lux­-labeled Pseudomonas fluorescens in soil. 2004. Bunker, S.T., T.C. Bates, and J.D. Oliver. Environ. Biosaf. Res. 3:83-90.
• Biochemical and virulence characterization of viable but nonculturable cells of Vibrio parahaemolyticus. 2004. Wong, H.-C. and J.D. Oliver. J. Food Prot. 67:2430-2435.
• The viable but nonculturable state of Vibrio parahaemolyticus. 2004. Bates, T.C. and J.D. Oliver. J. Microbiol. 42:74-79.
• Role of catalase and oxyR in the viable but nonculturable state of Vibrio vulnificus. 2004. Kong, I.-S., T.C. Bates, A. Hülsmann, , H. Hassan, and J.D. Oliver. FEMS Microbiol. Ecol. 50:133-142.
• Pulsed-field electrophoresis analysis of Vibrio vulnificus strains isolated from Taiwan and United States. 2004. Wong, H.-c., S.Y. Chen, M.-Y. Chen, J.D. Oliver, L.-I. Hor, and W.-Ch. Tsai. Appl. Environ. Microbiol. 70:5153-5158.
• Changes in membrane fatty acid composition during entry of Vibrio vulnificus in the viable but nonculturable state. 2004. Day, A.P. and J.D. Oliver. J. Microbiol. 42:69-73.
• Induction of Escherichia coli and Salmonella typhimurium into the viable but nonculturable state following chlorination of wastewater. 2005. Oliver, J.D., M. Dagher, and K. Linden. J. Water and Health 3.3:249-257.
• Wound infections caused by Vibrio vulnificus and other marine bacteria. “Special Article”. Oliver, J.D. 2005. Epidemiol. Infect. 133:383-391.
• A rapid and simple PCR analysis indicates there are two subgroups of Vibrio vulnificus which correlate with clinical or environmental isolation. 2005. Rosche, T.M., Y. Yano, and J.D. Oliver. Microbiol. Immunol. 49:381-389.
• The viable but nonculturable state in bacteria. 2005. Oliver, J.D. J. Microbiol. 43:93-100.
• Cloning, sequencing and expression of a GroEL-like protein gene of Vibrio vulnificus. 2005. Wong, H.-c., K.H. Lu, and J.D. Oliver. Taiw. J. Agric. Chem. Food Sci. 43:1-7.
• RpoS involvement in osmotically-induced cross protection in Vibrio vulnificus. Rosche, T.M., T.C. Bates, D.J. Smith, E.E. Parker, and J.D. Oliver. FEMS. Microbiol. Ecol. 53:455-462.
• Intrapopulational variation in Vibrio vulnificus levels in Crassostrea virginica is associated with the host size but not with disease status or developmental stability. 2005. Sokolova, I.M., L. Leamy, M. Harrison, and J.D. Oliver. J. Shellfish Res. 24:503-508.
• Vibrio vulnificus. 2005. Oliver, J.D. In: Biology of Vibrios. F.L. Thompson, B. Austin, and J. Swing. (eds.). Amer. Soc. Microbiol. Press, Washington, D.C. (in press).
• In situ and in vitro gene expression by Vibrio vulnificus during entry into, persistence within, and resuscitation from the viable but nonculturable state. Smith, B.E. and J.D. Oliver. Appl. Environ. Microbiol. (submitted).
• In situ and in vitro gene expression by Vibrio vulnificus during the starvation-survival state. Smith, B.E. and J.D. Oliver. Appl. Environ. Microbiol. (submitted).
• An AFLP approach to identify genetic markers associated with resistance to Vibrio vulnificus and Perkinsus marinus in eastern oysters. Sokolova, I.M., J.D. Oliver, and L.J. Leamy. J. Shellfish Res. (submitted).
• Viable but nonculturable bacteria in food environments. 2005. Oliver, J.D. In: Food-borne pathogens: Microbiology and Molecular Biology. P.M. Fratamico, A.K. Bhunia, and J.L. Smith (eds.). Caister Academic Press, Norfolk, UK.
• Vibrio vulnificus. 2005. Oliver, J.D. In: Bacterial Pathogens in Sea Water. Belkin, S. (ed.). Plenum Publishing. (In press).
• Gene expression by Helicobacter pylori in a natural freshwater environment. Adams, B. L. and J. D. Oliver. Appl. Environ. Microbiol. (submitted).
• The role of cross protection in survival of Helicobacter pylori during stress. Adams, B.L. and J.D. Oliver. Helicobacter (submitted).
• In situ and in vitro gene expression by Vibrio vulnificus during entry into, persistence within, and resuscitation from the viable but nonculturable state.  2006. Smith, B.E. and J.D. Oliver. Appl. Environ. Microbiol. 72:1445-1451.
• In situ gene expression by Vibrio vulnificus.  2006. Smith, B.E. and J.D. Oliver.  Appl. Environ. Microbiol. 72:2244-2246.
• An AFLP approach to identify genetic markers associated with resistance to Vibrio vulnificus and Perkinsus marinus in eastern oysters.  2006. Sokolova, I.M., J.D. Oliver, and L.J. Leamy. J. Shellfish Res. 25: 95-100.
• Evidence for an intermediate colony morphology of Vibrio vulnificus.  2006. Rosche, T.M., B. Smith, and J.D. Oliver.  Appl. Environ. Microbiol.  72: 4356-4359.
• Capsular polysaccharide phase variation in Vibrio vulnificus.  2006. Hilton, T., T. Rosche, B. Froelich, B. Smith, and J.D. Oliver. Appl. Environ. Microbiol. 72:6986-6993.
• Refined medium for direct isolation of Vibrio vulnificus from oyster tissue and sea water. 2007. Warner, E. and J.D. Oliver. Appl. Environ. Microbiol. 73:3098-3100.
• Emergence of a virulent clade of Vibrio vulnificus and correlation with the presence of a 33-kilobase genomic island.  2007. Cohen, A.L.V., J.D. Oliver, A. DePaola, E. J. Feil, and E.F. Boyd. Appl. Environ. Microbiol. 73:5553-5565.
• Role of iron in human serum resistance of the clinical and environmental Vibrio vulnificus genotypes.  2007. Bogard, R. and J.D. Oliver. Appl. Environ. Microbiol. 73:7501-7505.
• Survival and in situ gene expression of Vibrio vulnificus at varying salinities in estuarine environments. 2008. Jones, M.K., E. Warner, and J.D. Oliver. Appl. Environ. Microbiol. 74:182-187.
• Population structure of two genotypes of Vibrio vulnificus in oysters (Crassostrea virginica) and sea water.  2008. Warner, E.B. and J.D. Oliver. Appl. Environ. Microbiol. 74:80-85.
• The ecology of Vibrio vulnificus, Vibrio cholerae, and Vibrio parahaemolyticus in North Carolina estuaries.  2008. Blackwell, K.D. and J.D. Oliver.  J. Microbiology. 46:146-153.
• 128.  Horizontal transfer of lux genes in Vibrionaceae.  2008. Urbanczyk, H., J.C. Ast, A.J. Kaeding, J.D. Oliver, and P.V. Dunlap. J. Bacteriol. 190:3494-3504.
• Multi-site analysis reveals widespread antibiotic resistance in the marine pathogen
  Vibrio vulnificus.  2008.  Baker-Austin, C., J.V. McArthur, A.H. Lindell, M.S. Wright, R.C. Tuckfield, J. Gooch, L. Warner, J.D. Oliver, and R. Stepanauskas.  Microb. Ecol. 57: 151-159.
• Multiplex PCR assay for detection and simultaneous differentiation of genotypes of Vibrio vulnificus biotype 1.  2008.  Warner, E.B. and J.D. Oliver. Foodborne Path. Dis. 5: 691-693.
• csrA inhibits biofilm formation in Vibrio vulnificus. Jones, M. and J.D. Oliver. 2008.  Appl. Environ. Microbiol. 74:7064-7066.
• Vibrio vulnificus: Disease and pathogenesis.  2009. Jones, M.K. and J.D. Oliver. Infect. Immun. 77:1723-1733.
• Evaluation of genotypic and phenotypic methods to distinguish clinical from environmental Vibrio vulnificus strains.  2009. Sanjuan, E., J.D. Oliver, and C. Amaro.  Appl. Environ. Microbiol. 75:1594-1598.
• Effect of weak acids on Listeria monocytogenes survival: evidence for a viable but-nonculturable state in response to low pH.  2009. Cunningham, E., C. O’Byrne, and J.D. Oliver. J. Food Control 20:1141-1144.
• Rapid in situ detection of virulent Vibrio vulnificus strains in raw oyster matrices using real-time PCR. 2009. Baker-Austin, C., A. Gore, J. D. Oliver, R. Rangdale, J.V. McArthur and D. N. Lees. Env. Microbiol. Rep. doi:10.1111/j.1758-2229.2009.00092.x
• Uptake and depuration of the C- and E-genotype of Vibrio vulnificus by the Eastern oyster (Crassostrea virginica). 2009. Froelich, B., A. Ringwood, I. Sokolova, and J.D. Oliver. Environ. Microbiol. Rep. (n press).
• Recent findings on the viable but nonculturable state in pathogenic bacteria. 2009. Oliver, J.D. FEMS Microbiology Rev. DOI:10.1111/j.1574-6976.2009.00200.x
• Role of RpoS in the susceptibility of low salinity-adapted Vibrio vulnificus to environmental stresses. 2010. Tan, H.-J., S.-H. Liu, J.D. Oliver, and H.-c. Wong. Intern. J. Food Microbiol. 137:137-142.
• Vibrio vulnificus genome suggests two distinct ecotypes. 2010. Rosche, T.M., E.A. Binder, and J.D. Oliver. Environ. Microbiol. Rep. (in press).
• Survival of spinach-associated Helicobacter pylori in the viable but nonculturable state. 2010. Buck, A. and J.D. Oliver. Food Control (in press).

Recent Graduate Students

• Bunker, Stephen. 2000. Effects of environmental stresses on culturability of genetically modified Pseudomonas fluorescens in soil.
• Bates, Tonya. 2001. The VBNC state in Vibrio parahaemolyticus.
• Birkenhauer, Jennifer. 2001. Use of GRAS compounds to reduce loads of Vibrio vulnificus in oysters.
• Day, Ashley. 2002. Membrane fatty acid changes in Vibrio vulnificus induced by low temperature and osmotic shock.
• Courtney Pfeffer. 2002. Involvement of Vibrio vulnificus in wound infections in the Neuse River area of North Carolina.
• Daren Beam. 2004. Quorum sensing and pathogenesis in Vibrio vulnificus.
• Ben Smith. 2005. In situ and in vitro gene expression by Vibrio vulnificus during the VBNC and starvation-survival states.
• Tamara Hilton. 2005. Capsular switching among the clinical- and environmental-genotypes of Vibrio vulnificus. 2005.
• Karen Dyer Blackwell. 2005. The viable but nonculturable states of Vibrio vulnificus, Vibrio cholerae, and Vibrio parahaemolyticus in the natural environment. 2005.
• Smith, Ben. 2005.  In situ and in vitro gene expression by Vibrio vulnificus during the VBNC and starvation-survival states.
• Dyer Blackwell, Karen. 2005. The viable but nonculturable states of Vibrio vulnificus, Vibrio cholerae, and Vibrio parahaemolyticus in the natural environment. 
• Hilton, Tamara.  2006. Capsular switching among the clinical- and environmental-genotypes of Vibrio vulnificus.
• Bogard, Ryan.  2007. Serum sensitivity of two genotypes of Vibrio vulnificus.
• Buck, Alan.  2008. Survival of, and gene expression by, Helicobacter pylori on plant surfaces.
• Kim, Erica. 2008.  Expression of Vibrio vulnificus catecholate and hydroxamate siderophores in natural and human environments.