Life Sciences Commons^™

A Draft Of The Genome Of The Gulf Coast Tick, Amblyomma Maculatum, Jose M.C. Ribeiro, Natalia J. Bayona-Vásquez, Khemraj Budachetri, Deepak Kumar, Julia Catherine Frederick, Faizan Tahir, Brant C. Faircloth, Travis C. Glenn, Shahid Karim

Faculty Publications

The Gulf Coast tick, Amblyomma maculatum, inhabits the Southeastern states of the USA bordering the Gulf of Mexico, Mexico, and other Central and South American countries. More recently, its U.S. range has extended West to Arizona and Northeast to New York state and Connecticut. It is a vector of Rickettsia parkeri and Hepatozoon americanum. This tick species has become a model to study tick/Rickettsia interactions. To increase our knowledge of the basic biology of A. maculatum we report here a draft genome of this tick and an extensive functional classification of its proteome. The DNA from a single …

Investigating The Involvement Of The Tick Vector In The Induction Of Alpha-Galactose Hypersensitivity (Alpha-Gal Syndrome, Red Meat Allergy) In The United States., Gary Crispell

Dissertations

Alpha-gal syndrome (AGS or sometimes called red meat allergy) is a result of the development of specific IgE antibodies to the oligosaccharide galactose-α-1,3-galactose (α-gal) after a person has had exposure to tick bites. This dissertation investigates four common tick species found in North America: the lone-star tick (Amblyomma americanum), the Gulf-Coast tick (Amblyomma maculatum), the American dog tick (Dermacentor variabilis), and the black-legged tick (Ixodes scapularis) for the presence of α-gal by utilizing a combination of immunoproteomic approaches and carbohydrate analysis techniques.

Anti-α-gal IgM antibodies (M86) were used in immunoblotting to detect …

Robust Network Stability Of Mosquitoes And Human Pathogens Of Medical Importance, Donald A. Yee, Catherine Dean Bermond, Limarie J. Reyes-Torres, Nicole S. Fijman, Nicole A. Scavo, Joseph Nelsen, Susan H. Yee

Faculty Publications

Background: The exact number of mosquito species relevant to human health is unknown, posing challenges in understanding the scope and breadth of vector–pathogen relationships, and how resilient mosquito vector–pathogen networks are to targeted eradication of vectors.

Methods: We performed an extensive literature survey to determine the associations between mosquito species and their associated pathogens of human medical importance. For each vector–pathogen association, we then determined the strength of the associations (i.e., natural infection, lab infection, lab dissemination, lab transmission, known vector). A network analysis was used to identify relationships among all pathogens and vectors. Finally, we examined how …

Characterizing Endogenous Dicer Products To Unravel Novel Rnai Biogenesis Pathways, Jacob Oche Peter

Dissertations

ABSTRACT

RNA interference (RNAi) is a pervasive gene regulatory mechanism in eukaryotes based on the action of multiple classes of small RNA (sRNA). Exploiting RNAi pathways in non-model systems have great potential for creating potent RNAi technologies. Here, we accessed RNAi-mediated control of gene expression in the two-spotted spider mite, Tetranychus urticae (T. urticae) using engineered dsRNA designed to modulate the host RNAi pathway and increase RNAi efficacy. Analysis of Dicer (Dcr) generated fragments revealed how exogenous RNAs access the host RNAi pathway in this animal, opening avenues for designing RNAi technology for their control. Further, some organisms …

Sensitivity Of A Lamp Assay For Detection Of The Dinoflagellate Amyloodinium Ocellatum In Simulated Field Conditions And Freeze Tolerance Of The Parasite, Robert Gonzales

Master's Theses

The obligate parasitic dinoflagellate Amyloodinium ocellatum causes amyloodiniosis in warm water marine fishes. The prolific parasite, which has a direct, three-stage life cycle, is highly infectious and can cause heavy losses in aquaculture. Prevention, biosecurity, and early detection are vital for control. In this work, microscopy and a loop-mediated isothermal amplification (LAMP) assay were compared for early diagnosis of A. ocellatum in cultured stocks, and the freeze tolerance of tomonts was assessed to determine if frozen wild fish used as fish food can serve as a potential vector for the parasite.

The lowest dinospore concentration that could be detected by …

An Exploratory Study On The Microbiome Of Northern And Southern Populations Of Ixodes Scapularis Ticks Predicts Changes And Unique Bacterial Interactions, Deepak Kumar, Latoyia P. Downs, Abdulsalam Adegoke, Erika Machtinger, Kelly Oggenfuss, Richard S. Ostfeld, Monica Embers, Shahid Karim

Faculty Publications

The black-legged tick (Ixodes scapularis) is the primary vector of Borrelia burgdorferi, the causative agent of Lyme disease in North America. However, the prevalence of Lyme borreliosis is clustered around the Northern States of the United States of America. This study utilized a metagenomic sequencing approach to compare the microbial communities residing within Ix. scapularis populations from northern and southern geographic locations in the USA. Using a SparCC network construction model, we performed potential interactions between members of the microbial communities from Borrelia burgdorferi–infected tissues of unfed and blood-fed ticks. A significant difference in bacterial composition …

Dicer Represses Antiviral Innate Immunity Pathways In Mouse Embryonic Stem Cells, Chandan Gurung

Dissertations

Recent studies have demonstrated that embryonic stem cells (ESCs) are deficient in expressing type I interferons (IFN), the cytokines that play key roles in antiviral responses. However, the underlying molecular mechanisms and biological implications of this finding are poorly understood. In this study, I used a synthetic RNA-based assay that can simultaneously assess multiple forms of antiviral responses in ESCs. Dicer is an enzyme essential for RNA interference (RNAi), which is used as a major antiviral mechanism in invertebrates but not clear in vertebrates. RNAi activity is detected in wild-type ESCs but is abolished in Dicer knockout ESCs (D−/−ESCs) as …

Tick Salivary Gland Extract Induces Alpha-Gal Syndrome In Alpha-Gal Deficient Mice, Shailesh K. Choudhary, Shahid Karim, Onyinye I. Iweala, Shivangi Choudhary, Gary Crispell, Surendra Raj Sharma, Claire T. Addison, Mike Kulis, Brian H. Herrin, Susan E. Little, Scott P. Commins

Faculty Publications

Introduction: Alpha-gal syndrome (AGS) is characterized by delayed hypersensitivity to non-primate mammalian meat in people having specific immunoglobulin E (sIgE) to the oligosaccharide galactose-alpha-1,3-galactose. AGS has been linked to tick bites from Amblyomma americanum (Aa) in the U.S. A small animal model of meat allergy is needed to study the mechanism of alpha-gal sensitization, the effector phase leading to delayed allergic responses and potential therapeutics to treat AGS.

Methods: Eight- to ten-weeks old mice with a targeted inactivation of alpha-1,3-galactosyltransferase (AGKO) were injected intradermally with 50 μg of Aa tick salivary gland extract (TSGE) on days …

Tick Saliva And The Alpha-Gal Syndrome: Finding A Needle In A Haystack, Surendra Raj Sharma, Shahid Karim

Faculty Publications

Ticks and tick-borne diseases are significant public health concerns. Bioactive molecules in tick saliva facilitate prolonged blood-feeding and transmission of tick-borne pathogens to the vertebrate host. Alpha-gal syndrome (AGS), a newly reported food allergy, is believed to be induced by saliva proteins decorated with a sugar molecule, the oligosaccharide galactose-⍺-1,3-galactose (α-gal). This syndrome is characterized by an IgE antibody-directed hypersensitivity against α-gal. The α-gal antigen was discovered in the salivary glands and saliva of various tick species including, the Lone Star tick (Amblyomma americanum). The underlying immune mechanisms linking tick bites with α-gal-specific IgE production are poorly understood and are …

Recent Advances In Understanding Tick And Rickettsiae Interactions, Shahid Karim, Deepak Kumar, Khemraj Budachetri

Faculty Publications

Ticks are haematophagous arthropods with unique molecular mechanisms for digesting host blood meal while acting as vectors for various pathogens of public health significance. The tick's pharmacologically active saliva plays a fundamental role in modulating the host's immune system for several days to weeks, depending on the tick species. The vector tick has also developed sophisticated molecular mechanisms to serve as a competent vector for pathogens, including the spotted fever group (SFG) rickettsiae. Evidence is still inadequate concerning tick–rickettsiae–host interactions and saliva-assisted transmission of the pathogen to the mammalian host. Rickettsia parkeri, of the SFG rickettsia, can cause a …

Effects Of Infection Of The Protist Parasite, Dermomycoides Sp., In Dusky Gopher Frog Tadpoles, Jaime Smith

Master's Theses

Infections of the protist parasite, Dermomycoides sp. are thought to have caused several years of low recruitment in the dusky gopher frog (Rana sevosa) populations. I evaluated the effects of density of the infective zoospores, host developmental stage, and tadpoles' ability to acquire resistance to Dermomycoides sp. on dusky gopher frog tadpoles. Tadpoles were exposed to zoospore densities of 0, 250, 500, and 750 zoospores/µL at Gosner stage 25, and we found no significant differences among treatments in tadpole mortality. In evaluating susceptibility by development stage, I exposed R. sevosa to 50 zoospores/µL as eggs, embryos, hatchlings, and …

Interleukin-17a Facilitates Chikungunya Virus Infection By Inhibiting Ifn-Α2 Expression, Biswas Neupane, Dhiraj Acharya, Farzana Nazneen, Gabriel Gonzalez-Fernandez, Alex Sutton Flynt, Fengwei Bai

Faculty Publications

Interferons (IFNs) are the key components of innate immunity and are crucial for host defense against viral infections. Here, we report a novel role of interleukin-17A (IL-17A) in inhibiting IFN-α2 expression thus promoting chikungunya virus (CHIKV) infection. CHIKV infected IL-17A deficient (Il17a^−/−) mice expressed a higher level of IFN-α2 and developed diminished viremia and milder footpad swelling in comparison to wild-type (WT) control mice, which was also recapitulated in IL-17A receptor-deficient (Il17ra^−/−) mice. Interestingly, IL-17A selectively blocked IFN-α2 production during CHIKV, but not West Nile virus (WNV) or Zika virus (ZIKV), infections. Recombinant IL-17A …

Laboratory Colonization By Dirofilaria Immitis Alters The Microbiome Of Female Aedes Aegypti Mosquitoes, Abdulsalam Adegoke, Erik Neff, Amie Geary, Montana Ciara Husser, Kevin Wilson, Shawn Michael Norris, Guha Dharmarajan, Shahid Karim

Faculty Publications

Background

The ability of blood-feeding arthropods to successfully acquire and transmit pathogens of medical and veterinary importance has been shown to be interfered with, or enhanced by, the arthropod’s native microbiome. Mosquitoes transmit viruses, protozoan and filarial nematodes, the majority of which contribute to the 17% of infectious disease cases worldwide. Dirofilaria immitis, a mosquito-transmitted filarial nematodes of dogs and cats, is vectored by several mosquito species including Aedes aegypti.

Methods

In this study, we investigated the impact of D. immitis colonization on the microbiome of laboratory reared female Ae. aegypti. Metagenomic analysis of the V3–V4 variable …

Generating Zika Vaccine Candidates Using Nhumirim Virus As A Backbone, Mariam A. Atobiloye

Honors Theses

Zika virus (ZIKV) is a mosquito-transmitted flavivirus that is causing significant world-wide health concern. There are currently no treatments or vaccines available for this virus, thus, there is an urgent need to develop a safe and effective vaccine to combat ZIKV infection. Nhumirim virus (NHUV) is also a mosquito-transmitted flavivirus, but it is unable to infect humans and other vertebrate animals, making it an ideal candidate to develop chimeric viral vaccines against other disease-causing flaviviruses, such as ZIKV. In this study, we generated chimeric viruses by replacing envelope (E) gene in the genome of NHUV with ZIKV E gene, which …

Characterizing Erad And Antioxidant Response In Ixodes Scapularis Ise6 Cells During Borrelia Infection, Latoyia Downs

Master's Theses

Ticks are hematophagous ectoparasites that transmit a multitude of diseases to humans. Borrelia burgdorferi (BB) and Borrelia miyamotoi (BM) are both tick-borne pathogens that cause disease in humans and are transmitted by the black-legged tick (Ixodes scapularis). A byproduct of blood digestion generates reactive oxygen species that are toxic and cause oxidative stress which promotes cellular damage and dysfunction. The endoplasmic reticulum (ER) is especially affected by oxidative stress, resulting in a buildup of improperly folded proteins in the ER lumen called ER stress. To prevent cellular damage, the tick utilizes an antioxidant system to neutralize ROS and mechanisms …

Development Of An Attenuated Zika Virus By Editing The 5’ Untranslated Region, E. Ashley Thompson

Master's Theses

Zika Virus (ZIKV) is a mosquito-transmitted flavivirus that usually causes no symptoms to mild febrile in humans, and it has been regarded as an insignificant pathogen to public health. However, recent outbreaks of ZIKV infection have revealed that ZIKV can cause severe neurological effects in adults, such as Guillain-Barre Syndrome (GBS), and in infants whose mothers acquired the virus during pregnancy, causing Congenital Zika Syndrome (CZS). Currently, no approved vaccine is available and there is a critical need to develop an effective and safe vaccine. While most vaccine developmental strategies target the viral prM-E protein of ZIKV, we aimed to …

Tumor Necrosis Factor-Alpha Signaling May Contribute To Chronic West Nile Virus Post-Infectious Proinflammatory State, A. Arturo Leis, Marie F. Grill, Brent P. Goodman, Syed B. Sadiq, David J. Sinclair, Parminder J.S. Vig, Fengwei Bai

Faculty Publications

Background: West Nile virus (WNV) causes a spectrum of human disease ranging from a febrile illness (WNV fever) to severe neuroinvasive disease (meningitis, encephalitis, acute flaccid paralysis). Since WNV gained entry into North America in 1999, clinicians caring for WNV survivors have observed persistent neurological symptoms occurring long-after the production of neutralizing antibodies and clearance of the virus. Accordingly, alternative pathogeneses other than direct viral invasion have been hypothesized to explain these post-infectious symptoms. The dominant hypothesis is that antiviral inflammatory responses triggered initially to clear WNV may persist to promote a post-infectious proinflammatory state.

Methods: In 4 serologically-confirmed …

Cysteine Dioxygenase Enzyme Activity And Gene Expression In The Dimorphic Pathogenic Fungus Histoplasma Capsulatum Is In Both The Mold And Yeast Morphotypes And Exhibits Substantial Strain Variation, Melissa Adams, Glen Shearer

Faculty Publications

In the dimorphism (mold/yeast) Histoplasma capsulatum (Hc) literature are reports that yeast (the so-called pathogenic form) uniquely expresses a cysteine dioxygenase (CDO, approx. 10,500 dal) activity which the mold morphotype (the so-called saprophytic soil form) does not express (C.F., Kumar et al., Biochem 22, 762, 1983). This yeast-specific CDO activity is postulated to play a critical role in the mold-to-yeast shift. A number of years ago, our lab isolated the gene encoding the Hc cysteine dioxygenase (CDO1, Genbank accession AY804144) and noted significant expression in the mold morphotype of several Histoplasma strains and also determined that the predicted protein would …

Phylogenetic Afficity Of Genolopa (Digenea: Monorchiidae) With Descriptions Of Two New Species, Apryle J. Panyi, Stephen S. Curran, Robin M. Overstreet

Faculty Publications

The validity of Genolopa Linton, 1910 has been controversial because the observation of presently recognized critical diagnostic morphological features (spines in the genital atrium and a bipartite, anteriorly spined terminal organ) were omitted from the original diagnosis, and these features were not universally appreciated as important diagnostic features until 2008. Modern taxonomists have been further challenged by inappropriate fixation techniques that have resulted in various interpretations of morphological features. Consequently, named species in the genus have fluctuated among other monorchiid genera depending on various interpretations by taxonomists, and a modern consensus on classifying these species is lacking. This study …

The Histoplasma Capsulatum Ddr48 Gene Is Required For Survival Within Macrophages And Resistance To Oxidative Stress And Antifungal Drugs, Logan Blancett

Dissertations

Histoplasma capsulatum(Hc)is a systemic, dimorphic fungal pathogen that affects upwards of 500,000 individuals in the United States annually. Hc grows as a multicellular mold at environmental temperatures; whereas, upon inhalation into a human or other mammalian host, it transforms into a unicellular, pathogenic yeast. The research presented in this dissertation is focused on characterizing the DNA damage-responsive gene HcDDR48. HcDDR48was originally isolated via a subtractive DNA library enriched for transcripts enriched in the mold-phase of Hcgrowth. Upon further analysis we found that HcDDR48is not just expressed in the mold morphotype, but both growth programs …

The Impact Of Msaabcr On Sara-Associated Phenotypes Is Different In Divergent Clinical Isolates Of Staphylococcus Aureus, Joseph S. Rom, Aura M. Ramirez, Karen E. Beenken, Gyan S. Sahukhal, Mohamed O. Elasri, Mark S. Smeltzer

Faculty Publications

The staphylococcal accessory regulator (sarA) plays an important role in Staphylococcus aureus infections including osteomyelitis, and the msaABCR operon has been implicated as an important factor in modulating expression of sarA. Thus, we investigated the contribution of msaABCR to sarA-associated phenotypes in the S. aureus clinical isolates LAC and UAMS-1. Mutation of msaABCR resulted in reduced production of SarA and a reduced capacity to form a biofilm in both strains. Biofilm formation was enhanced in a LAC msa mutant by restoring the production of SarA, but this was not true in a UAMS-1 msa mutant. Similarly, …

Attenuated Responses To Inflammatory Cytokines In Mouse Embryonic Stem Cells: Biological Implications And The Molecular Basis, Bohan Chen

Master's Theses

Embryonic stem cells (ESCs) have attracted intense interest due to their great potential for regenerative medicine. However, their immune property is an overlooked but a significant issue that needs to be thoroughly investigated not only to resolve the concern for therapeutic applications but also for further understanding the early stage of organismal development. Recent studies demonstrated that ESCs are deficient in innate immune responses to viral/bacterial infections and inflammatory cytokines. Inflammatory conditions generally inhibit cell proliferation, which could be detrimental to ESCs, since cell proliferation is their dedicated task during early embryogenesis. Thus, I hypothesize that the attenuated innate immunity …

Differential Expression Of Genes Related To Innate Immune Responses In Ex Vivo Spinal Cord And Cerebellar Slice Cultures Infected With West Nile Virus, Parminder J.S. Vig, Deyin Lu, Amber M. Paul, Ram Kuwar, Maria Lopez, Dobrivoje S. Stokic, A. Arturo Leis, Michael R. Garrett, Fengwei Bai

Faculty Publications

West Nile virus (WNV) infection results in a spectrum of neurological symptoms, ranging from a benign fever to severe WNV neuroinvasive disease with high mortality. Many who recover from WNV neuroinvasive infection present with long-term deficits, including weakness, fatigue, and cognitive problems. While neurons are a main target of WNV, other cell types, especially astrocytes, play an important role in promoting WNV-mediated central nervous system (CNS) damage. Conversely, it has been shown that cultured primary astrocytes secrete high levels of interferons (IFNs) immediately after WNV exposure to protect neighboring astrocytes, as well as neurons. However, how intrinsic responses to WNV …

Taxonomy And Systematics Of Plagioporus (Trematoda), With Descriptions Of 10 New Species From Freshwater Fishes Of The Nearctic, Thomas John Fayton

Dissertations

The Opecoelidae Ozaki, 1925 is one of the largest families of digenetic trematodes of fishes. While the family is mostly marine/estuarine, invasion of freshwater hosts has occurred at least two times. The only representative freshwater plagioporine sequenced to date is Plagiocirrus loboides Curran, Overstreet, & Tkach, 2007, which previous phylogenetic analyses resolved as being related to deep water marine opecoelids. The taxonomy of the freshwater plagioporines, particularly Plagioporus, has long been confused; homoplasy is rife within the family and has complicated the delineation of species and genera, and the freshwater species from marine forms. Here, I hypothesize that the …

A Novel Role For Osteopontin In Facilitating West Nile Virus Neuroinvasion, Amber M. Paul

Dissertations

West Nile virus (WNV) is a positive-sensed, single-stranded RNA flavivirus that can cause human neuroinvasive diseases, including encephalitis, meningitis, and flaccid paralysis. The mechanisms by which WNV enters the central nervous system and the host-factors that are involved in WNV-neuroinvasiveness are not completely understood. Osteopontin (OPN), a multifunctional glycoprotein, has been implicated as a bio-marker for a number of neuroinflammatory diseases. In particular, secreted (s)OPN has been implicated to participate in recruitment of polymorphonuclear neutrophils (PMN) to sites of its expression, while PMNs have been suggested to act as WNV reservoirs. Therefore, sOPN recruitment of PMNs may contribute to neuroinvasive …

Study Of Rickettsia Parkeri Colonization And Proliferation In The Tick Host Amblyomma Maculatum (Acari: Ixodidae), Khemraj Budachetri

Dissertations

Amblyomma maculatum (Gulf coast tick) ticks are prevalent across the Atlantic to Gulf Coast region of United States. These ticks are recognized vectors of Rickettsia parkeri, a spotted fever group of Rickettsia (SFGR) known to cause American boutonneuse fever associated with fever and eschar rashes localized to the site of bites. We hypothesized that Rickettsia parkeri colonization and proliferation in the tick vector involve pathogen-symbiont dynamics and tick-pathogen interactions, which influence rickettsial transmission to the victims after tick bites. The rickettsial infection is maintained across the tick life cycle for many generations due to transovarial and transstadial transmission of …

Elucidating The Role Of Interleukin-17a In West Nile Virus Infection, Dhiraj Acharya

Dissertations

West Nile virus (WNV) is a neurotropic flavivirus of significant public health importance for which no therapeutics and vaccine are currently available. Interleukin-17A (IL-17A) is an inflammatory cytokine that regulates diverse immune functions, while its role is unclear in host’s immune response to WNV. Furthermore, CD8⁺ T cells are crucial components of immunity and play a vital role in recovery from WNV infection. Here, we report a previously unrecognized function of IL-17A in regulating CD8⁺ T cell cytotoxicity. We show that WNV induces the expression of IL-17A in both mouse splenocytes and human peripheral blood mononuclear cells cultured …

A Microbiomic Approach To The Characterization Of The Impacts And Influences Of Viral, Bacterial, And Harmful Algal Bloom Toxins On The Bottlenose Dolphin, Corey David Russo

Dissertations

As apex predators that display high site fidelity Tursiops truncatus (bottlenose dolphin) are indicators of marine ecosystem health. Bottlenose dolphins, additionally, display pathogenesis and immune response similar to that of humans. Humans and coastal bottlenose dolphins, in particular, are constantly exposed to the same industrial, agricultural and domestic toxins and pathogens, contaminants and pollutants. Thus, studies on the bottlenose dolphin are also valuable in bridging the gap between ocean and human health. Bottlenose dolphins are susceptible to viral bacterial and toxin infection. Infection in the bottlenose dolphin manifests itself in the form of mass stranding events, unusual mortality events, chronic …

Real-Time Pcr Detection And Phylogenetic Relationships Of Neorickettsia In Digeneans From Egypt, Phillipines, Thailand, Vietnam And The United States, Stephen E. Greiman, Jefferson A. Vaughan, Rasha Elmahy, Poom Adisakwattana, Nguyen Van Ha, Thomas J. Fayton, Amal I. Khalil, Vasyl V. Tkach

Student Publications

Neorickettsia (Rickettsiales, Anaplasmataceae) is a genus of obligate intracellular bacterial endosymbionts of digeneans (Platyhelminthes, Digenea). Some Neorickettsia are able to invade cells of the digenean's vertebrate host and are known to cause diseases of domestic animals, wildlife, and humans. In this study we report the results of screening digenean samples for Neorickettsia collected from bats in Egypt and Mindoro Island, Philippines, snails and fishes from Thailand, and fishes from Vietnam and the USA. Neorickettsia were detected using a real-time PCR protocol targeting a 152bp fragment of the heat shock protein coding gene, GroEL, and verified with nested PCR and sequencing …

Endohelminth Parasites Of Some Midwater And Benthopelagic Stomiiform Fishes From The Northern Gulf Of Mexico, Michael J. Andres, Mark S. Peterson, Robin M. Overstreet

Gulf and Caribbean Research

: Mesopelagic fishes represent significant ecological links between mesozooplankton and the larger pelagic squids, fishes, and marine mammals. As such, these fishes also play a significant role as intermediate or paratenic hosts for parasites that require a crustacean intermediate host and mature in marine mammals or pelagic fishes. We examined a total of 208 individuals representing 5 species of Sternoptychidae and 88 individuals representing 2 species of Phosichthyidae from 20 locations in the northern Gulf of Mexico (nGOM). Six of the 7 species we examined are mesopelagic and one species was benthopelagic. We found the larval stages of Anisakis brevispiculata …