Microbiology Commons^™

Extracellular Proteases Are Key Mediators Of Staphylococcus Aureus Virulence Via The Global Modulation Of Virulence-Determinant Stability, Stacey L. Kolar, J. Antonio Ibarra, Frances E. Rivera, Joe M. Mootz, Jessica E. Davenport, Stanley M. Stevens Jr., Alexander R. Horswill, Lindsey N. Shaw

Molecular Biosciences Faculty Publications

Staphylococcus aureus is a highly virulent and successful pathogen that causes a diverse array of diseases. Recently, an increase of severe infections in healthy subjects has been observed, caused by community-associated methicillin-resistant S. aureus (CA-MRSA). The reason for enhanced CA-MRSA virulence is unclear; however, work suggests that it results from hypersecretion of agr-regulated toxins, including secreted proteases. In this study, we explore the contribution of exo-proteases to CA-MRSA pathogenesis using a mutant lacking all 10 enzymes. We show that they are required for growth in peptide-rich environments, serum, in the presence of antimicrobial peptides (AMPs), and in human blood. …

A Transient Α-Helical Molecular Recognition Element In The Disordered N-Terminus Of The Sgs1 Helicase Is Critical For Chromosome Stability And Binding Of Top3/Rmi1, Jessica A. Kennedy, Gary W. Daughdrill, Kristina H. Schmidt

Molecular Biosciences Faculty Publications

The RecQ-like DNA helicase family is essential for the maintenance of genome stability in all organisms. Sgs1, a member of this family in Saccharomyces cerevisiae, regulates early and late steps of double-strand break repair by homologous recombination. Using nuclear magnetic resonance spectroscopy, we show that the N-terminal 125 residues of Sgs1 are disordered and contain a transient α-helix that extends from residue 25 to 38. Based on the residue-specific knowledge of transient secondary structure, we designed proline mutations to disrupt this α-helix and observed hypersensitivity to DNA damaging agents and increased frequency of genome rearrangements. In vitro binding assays …

Identification Of Proteins At Active, Stalled, And Collapsed Replication Forks Using Isolation Of Proteins On Nascent Dna (Ipond) Coupled With Mass Spectrometry, Bianca M. Sirbu, W. Hayes Mcdonald, Huzefa Dungrawala, Akosua Badu-Nkansah, Gina M. Kavanaugh, Yaoyi Chen, David L. Tabb, David Cortez

Molecular Biosciences Faculty Publications

Both DNA and chromatin need to be duplicated during each cell division cycle. Replication happens in the context of defects in the DNA template and other forms of replication stress that present challenges to both genetic and epigenetic inheritance. The replication machinery is highly regulated by replication stress responses to accomplish this goal. To identify important replication and stress response proteins, we combined isolation of proteins on nascent DNA (iPOND) with quantitative mass spectrometry. We identified 290 proteins enriched on newly replicated DNA at active, stalled, and collapsed replication forks. Approximately 16% of these proteins are known replication or DNA …

Free-Living Amoebae In Sediments From The Lascaux Cave In France, Angela M. Garcia-Sanchez, Concepcion Ariza, Jose M. Ubeda, Pedro M. Martin-Sanchez, Valme Jurado, Fabiola Bastian, Claude Alabouvette, Cesareo Saiz Jimenez

International Journal of Speleology

The Lascaux Cave in France is an old karstic channel where the running waters are collected in a pool and pumped to the exterior. It is well-known that water bodies in the vicinity of humans are suspected to be reservoirs of amoebae and associated bacteria. In fact, the free-living amoebae Acanthamoeba astronyxis, Acanthamoeba castellanii, Acanthamoeba sp. and Hartmannella vermiformis were identified in the sediments of the cave using phylogenetic analyses and morphological traits. Lascaux Cave sediments and rock walls are wet due to a relative humidity near saturation and water condensation, and this environment and the presence of abundant bacterial …

Cure From The Cave: Volcanic Cave Actinomycetes And Their Potential In Drug Discovery, Naowarat (Ann) Cheeptham Dr., Tara Sadoway, Devon Rule, Kent Watson, Paul Moote, Laiel C. Soliman, Nicholas Azad, Kingsley Donkor, Derrick Horne

International Journal of Speleology

Volcanic caves have been little studied for their potential as sources of novel microbial species and bioactive compounds with new scaffolds. We present the first study of volcanic cave microbiology from Canada and suggest that this habitat has great potential for the isolation of novel bioactive substances. Sample locations were plot ted on a contour map that was compiled in ArcView 3.2. Over 400 bacterial isolates were obtained from the Helmcken Falls cave in Wells Gray Provincial Park, British Columbia. From our preliminary screen, of 400 isolates tested, 1% showed activity against extended spectrum ß-lactamase E. coli, 1.75% against Escherichia …

Diversity And Biosynthetic Potential Of Culturable Aerobic Heterotrophic Bacteria Isolated From Magura Cave, Bulgaria, Iva Tomova, Irina Lazarkevich, Anna Tomova, Margarita Kambourova, Evgenia Vasileva-Tonkova

International Journal of Speleology

Biocapacity of bacteria inhabiting karstic caves to produce valuable biologically active compounds is still slightly investigated. A total of 46 culturable heterotrophic bacteria were isolated under aerobic conditions from the Gallery with pre-historical drawings in MaguraCave, Bulgaria. Phylogenetic analysis revealed that most of bacterial isolates affiliated with Proteobacteria (63%), followed by Actinobacteria (10.9%), Bacteroidetes (10.9%), and Firmicutes (6.5%). A strong domination of Gram-negative bacteria (total 81%) belonging to nine genera: Serratia, Pseudomonas, Enterobacter, Sphingobacterium, Stenotrophomonas, Commamonas, Acinetobacter, Obesumbacterium, and Myroides, was observed. Gram-positive isolates were represented by the genera Bacillus, Arthrobacter, and Micrococcus. One isolate …

The Effects Of Uv Light On The Antimicrobial Activities Of Cave Actinomycetes, Devon Rule, Naowarat Cheeptham

International Journal of Speleology

The goal of this study was to determine whether actinomycetes isolated from a volcanic cave in western Canada could produce novel antimicrobial compounds against six multidrugresistant pathogens when exposed to UV light. One hundred and seventy-six actinomycete strains isolated from Helmcken Falls Cave, Wells Gray Provincial Park, BC, were screened against six pathogens using the “plug assay” in UV light and no light conditions. Of the 176 strains tested, 100 or 57% of the cave actinomycete strains had antimicrobial activities against the pathogens in 124 different instances: 35 instances when exposed to UV and no light, 30 when exposed to …

A World Review Of Fungi, Yeasts, And Slime Molds In Caves, Karen J. Vanderwolf, David Malloch, Donald F. Mcalpine, Graham J. Forbes

International Journal of Speleology

We provide a review of fungi, yeasts, and slime molds that have been found in natural solution caves and mines worldwide. Such habitats provide frequent roost sites for bats, and in eastern North America the environmental conditions that support white-nose syndrome, a lethal fungal disease currently devastating bat populations. A list of 1029 species of fungi, slime moulds, and yeasts in 518 genera have been documented from caves and mines worldwide in 225 articles. Ascomycota dominate the cave environment. Most research has been conducted in temperate climates, especially in Europe. A mean of 17.9±24.4SD fungal species are reported per study. …

La Serreta Endokarst (Se Spain): A Sustainable Value?, Antonia D. Asencio, Teodoro Espinosa

International Journal of Speleology

La Serreta endokarst (SE Spain), which UNESCO declared a World Heritage Site in 1998, was considered a sanctuary with cave art and one of the most important archaeological sites in the Mediterranean region for both the remains it hosts and the spectacular karstic landscape at the site.

To coincide with the 40th anniversary of its discovery, the La Serreta cave-chasm was adapted for public use with the intention of showing visitors the remains, which date back to prehistoric times. The solution included attempts to minimize contact with the valuables in the cave in order to alter the existing remains as …

Assessment Of A Modified Double Agar Layer Method To Detect Bacteriophage For Assessing The Potential Of Wastewater Reuse In Rural Bolivia, Sakira N. Hadley

USF Tampa Graduate Theses and Dissertations

Water scarcity is a global concern that impacts many developing countries, forcing people to depend on unclean water sources for domestic, agricultural, and industrial needs. Wastewater is an alternative water source that contains nutrients needed for crop growth. Wastewater reuse for agriculture can cause public health problems because of human exposure to pathogens. Pathogen monitoring is essential to evaluate the compliance of wastewater with established World Health Organization (WHO) and U.S. Environmental Protection Agency (EPA) wastewater reuse guidelines. Indicator organisms are commonly used to detect pathogens in water and wastewater because they are quick and easy to measure, non-pathogenic, and …

Direct And Indirect Effects Of Agrochemicals On Bacterial Pathogens And Fecal Indicator Bacteria, Zachery Staley

USF Tampa Graduate Theses and Dissertations

The presence of agrochemical residues in both urban and agricultural water bodies has become ubiquitous, often producing deleterious effects in the impacted watershed including reductions in biodiversity, alterations in species interactions, and toxicity to non-target organisms. While these effects have been studied on metazoan consumers, the consequences of agrochemical contamination on microorganisms, such as bacteria, protozoa, and viruses, are poorly understood. Agrochemicals could act directly on microorganisms, including pathogens, by either facilitating their survival or decreasing their abundance. Further, a multitude of indirect effects of agrochemicals on microorganisms are possible, whereby agrochemicals alter predation, competition, or parasitism on or available …

Prenatal Stress, Depression, And Herpes Viral Titers, Pao-Chu Hsu

USF Tampa Graduate Theses and Dissertations

Recent studies suggest that some cases of prenatal depression may be associated with reactivation of latent infections of the herpesvirus family. The possible relationships among stress, prenatal depression, and herpes viral reactivation in pregnancy are understudied and the molecular pathways such as the neuroimmune biogenic amine pathway are unidentified. Chronic stress shifts the T helper-1 cell (Th1) cytokine profile to a Th2 profile, which favors virus induced pathogenesis and survival. Pregnancy is also associated with a similar Th2 dominance. In non-pregnant individuals, exposure to psychological or physical stress may be associated with latent herpes viral reactivation and could result in …

Innate Immune Responses To Respiratory Syncytial Virus: Age-Associated Changes, Terianne Maiko Wong

USF Tampa Graduate Theses and Dissertations

Respiratory syncytial virus (RSV) infection causes ~64 million cases of respiratory disease and 200,000 deaths annually worldwide, yet there is no broadly effective prophylactic or treatment regimen. RSV can produce acute respiratory illness in patients of all ages but strikes the age extremes, infants and the elderly, with highest frequency presumably due to innate immune deficiencies. A higher morbidity and mortality has been reported for the elderly above 65 years of age, which has been attributed to immune senescence. Efforts to generate an effective vaccine have thus far been unsuccessful.

The innate immune system provides the first line of defense …

Persistent Infection With Human Herpesvirus-6 In Patients With An Inherited Form Of The Virus: A Newly Described Disease, Shara Pantry

USF Tampa Graduate Theses and Dissertations

Human Herpesvirus 6A (HHV-6A) and 6B (HHV-6B) are ubiquitous betaherpesviruses. Both viruses are associated with a variety of adult disorders including neurological disorder, such as multiple sclerosis and chronic fatigue syndrome. HHV-6 viruses are capable of establishing latency by integration into the telomeres of the host chromosome and are transmitted in a Mendelian manner in approximately one percent of the population. To date little is known about the immunological and neurological consequences of HHV-6 inheritance. This study focused on a unique population of individuals that inherited HHV-6 and present with chronic fatigue-like symptoms, including hypersomnia, generalized fatigue, headache, and short …

Complete Genome Sequences And Phylogeny Of West Nile Virus Isolates From Southeastern United States, 2003-2012, Crystal Wedin

USF Tampa Graduate Theses and Dissertations

The study of the evolution and phylogeny of West Nile virus (WNV) has been an important area of research since the introduction of WNV in 1999. However, genome sequencing of isolates from the Southeastern part of the United States has been somewhat limited. To determine how WNV has evolved at a more localized level, ten isolates from Florida and Georgia from 2003-2012 were completely sequenced using Illumina's next-generation technology. In addition, a phylogenetic comparison of both the complete genome and select partial genomes was completed to ensure consistency among the results. This study further demonstrated the dominance of the North …

The Role And Regulation Of Nsars: A Cell-Envelope Stress Sensing Two-Component System In Staphylococcus Aureus, Stacey Lynn Kolar

USF Tampa Graduate Theses and Dissertations

Abstract

S. aureus has 16 predicted two-component systems (TCS) that respond to a range of environmental stimuli, and allow for adaptation to stresses. Of these 16, three have no known function, and are not homologous to any other TCS found in closely related organisms. NsaRS is one such element, and belongs to the intramembrane-sensing histidine kinase (IM-HK) family, which is conserved within the Firmicutes. The regulators are defined by a small sensing domain within their histidine kinase, suggesting that they do not sense external signals, but stress in or at the membrane. Our characterization of NsaRS in this work reveals …

Sigma Factor N (Σn): A Novel Regulator Of Extreme Acid Resistance In Enterohemorrhagic E. Coli O157:H7, Pamela Ann Fay

USF Tampa Graduate Theses and Dissertations

Extreme acid resistance contributes to the successful transmission of enterohemorrhagic E. coli (EHEC) through acidic food matrices and the stomach, allowing it to gain access to the intestine and elicit disease in humans. Alternative sigma factor N (σ^N, encoded by rpoN) was previously identified as a novel regulator of extreme acid resistance in EHEC. This study investigated the role for σ^N and co-expressed products of the rpoN operon in the acid resistance phenotype of EHEC. The results revealed that σ^N primarily controls acid resistance through repression of the glutamate-dependent acid resistance (GDAR) system through control …

Investigation Into The Mechanism Of Salicylate-Associated Genotypic Antibiotic Resistance In Staphylococcus Aureus, Nada Salah Helal

USF Tampa Graduate Theses and Dissertations

Growth of Staphylococcus aureus with the NSAID salicylate increases phenotypic resistance (SAPAR), and the frequency at which heritable resistance occurs to various antibiotics (SAGAR). This study describes the effect of salicylate on heritable and phenotypic resistance to a set of antibiotics for laboratory and multi-drug resistant strains of S. aureus and investigates the link between resistance and SAGAR. Drug gradient plates were used to determine phenotypic resistance to antibiotics targeting DNA replication, transcription, translation and the cell wall in the presence or absence of salicylate. To measure heritable resistance, mutation frequencies were determined for each antibiotic in the presence and …

Assessing The Microbial Consequences Of Remediation: Surrogate Microbial Screening And Native Metabolic Signatures In Tc(Vii) Contaminated Sediments, Kathryn Lafaye Bailey

USF Tampa Graduate Theses and Dissertations

The chemical and physical processes controlling contaminant fate and transport in the vadose zone limit the options for application of many remedial technologies. Foam delivery technology (FDT) has been developed as a potential solution to overcome these limitations for remediating subsurface and deep vadose zone environments using reactive amendments. Although there are many advantages to utilizing FDT for treatment in the deep vadose zone, little information is available on how the addition of these surfactants and remedial amendments affect the indigenous microbial communities in the deep vadose zone as well as the impact of biological transformations of surfactant-based foams on …

Evidence For Viral Infection In The Copepods Labidocera Aestiva And Acartia Tonsa In Tampa Bay, Florida, Darren Stephenson Dunlap

USF Tampa Graduate Theses and Dissertations

Mesozooplankton are of critical importance to marine food webs by transferring energy from the microbial food web to higher trophic levels and depositing energy to the deeper ocean layers through fecal deposition. While decades of research have shown that viruses have significant impacts in the oceans, and infect a wide range of organisms from bacteria to whales, there is still little known about the impacts of viruses on the mesozooplankton community. As copepods are the most abundant mesozooplankton group, this study sought to characterize the viruses present in natural populations of the calanoid copepods Acartia tonsa and Labidocera aestiva in …

Cell Wall Antibiotics Provoke Accumulation Of Anchored Mcherry In The Cross Wall Of Staphylococcus Aureus, Wenqi Yu, Friedrich Götz

Molecular Biosciences Faculty Publications

A fluorescence microscopy method to directly follow the localization of defined proteins in Staphylococcus was hampered by the unstable fluorescence of fluorescent proteins. Here, we constructed plasmid (pCX) encoded red fluorescence (RF) mCherry (mCh) hybrids, namely mCh-cyto (no signal peptide and no sorting sequence), mCh-sec (with signal peptide), and mCh-cw (with signal peptide and cell wall sorting sequence). The S. aureus clones targeted mCh-fusion proteins into the cytosol, the supernatant and the cell envelope respectively; in all cases mCherry exhibited bright fluorescence. In staphylococci two types of signal peptides (SP) can be distinguished: the +YSIRK motif SPlip and the −YSIRK …

Discovery Of Marinopyrrole A (Maritoclax) As A Selective Mcl-1 Antagonist That Overcomes Abt-737 Resistance By Binding To And Targeting Mcl-1 For Proteasomal Degradation*, Kenichiro Doi, Rongshi Li, Shen-Shu Sung, Hongwei Wu, Yan Liu, Wanda Manieri, Gowdahalli Krishnegowda, Andy Awwad, Alden Dewey, Xin Liu, Shantu Amin, Chunwei Cheng, Yong Qin, Ernst Schonbrunn, Gary W. Daughdrill, Thomas P. Loughran Jr., Said M. Sebti, Hong-Gang Wang

Molecular Biosciences Faculty Publications

The anti-apoptotic Bcl-2 family of proteins, including Bcl-2, Bcl-XL and Mcl-1, are well-validated drug targets for cancer treatment. Several small molecules have been designed to interfere with Bcl-2 and its fellow pro-survival family members. While ABT-737 and its orally active analog ABT-263 are the most potent and specific inhibitors to date that bind Bcl-2 and Bcl-XL with high affinity but have a much lower affinity for Mcl-1, they are not very effective as single agents in certain cancer types because of elevated levels of Mcl-1. Accordingly, compounds that specifically target Mcl-1 may overcome this resistance. In this study, we identified …

An Ims/Atp Assay For The Detection Of Mycobacterium Tuberculosis In Urine, Dawn M. Hunter, Daniel V. Lim

Molecular Biosciences Faculty Publications

Background. Although sputum smears are the gold standard for diagnosis of tuberculosis, sensitivity in HIV/TB coinfection cases is low, indicating a need for alternative methods. Urine is being increasingly evaluated. Materials and Methods. A novel method for detecting Mycobacterium tuberculosis (MTB) in synthetic urine using a combined IMS/ATP assay was evaluated. Preliminary work established standard ATP conditions and the sensitivity and specificity of the MTB antibody. Eighty-four blinded samples in four replicate assays were evaluated for the presence of MTB using labeled immunomagnetic beads for capture. Beads were separated, washed, and resuspended in broth and added to a …

Identification Of New Cell Size Control Genes In S. Cerevisiae, Huzefa Dungrawala, Hui Hua, Jill Wright, Lesley Abraham, Thivakorn Kasemsri, Anthony Mcdowell, Jessica Stilwell, Brandt L. Schneider

Molecular Biosciences Faculty Publications

Cell size homeostasis is a conserved attribute in many eukaryotic species involving a tight regulation between the processes of growth and proliferation. In budding yeast S. cerevisiae, growth to a “critical cell size” must be achieved before a cell can progress past START and commit to cell division. Numerous studies have shown that progression past START is actively regulated by cell size control genes, many of which have implications in cell cycle control and cancer. Two initial screens identified genes that strongly modulate cell size in yeast. Since a second generation yeast gene knockout collection has been generated, we screened …

The Two-Component Signal Transduction System Arlrs Regulates Staphylococcus Epidermidis Biofilm Formation In An Ica-Dependent Manner, Yang Wu, Jiaxue Wang, Tao Xu, Jingran Liu, Wenqi Yu, Qiang Lou, Tao Zhu, Nianan He, Haijing Ben, Jian Hu, Friedrich Götz, Di Qu

Molecular Biosciences Faculty Publications

Due to its ability to form biofilms on medical devices, Staphylococcus epidermidis has emerged as a major pathogen of nosocomial infections. In this study, we investigated the role of the two-component signal transduction system ArlRS in regulating S. epidermidis biofilm formation. An ArlRS-deficient mutant, WW06, was constructed using S. epidermidis strain 1457 as a parental strain. Although the growth curve of WW06 was similar to that of SE1457, the mutant strain was unable to form biofilms in vitro. In a rabbit subcutaneous infection model, sterile disks made of polymeric materials were implanted subcutaneously followed with inoculation of WW06 or SE1457. …

Wipi-1 Positive Autophagosome-Like Vesicles Entrap Pathogenic Staphylococcus Aureus For Lysosomal Degradation, Mario Mauthe, Wenqi Yu, Oleg Krut, Martin Krönke, Friedrich Götz, Horst Robenek, Tassula Proikas-Cezanne

Molecular Biosciences Faculty Publications

Invading pathogens provoke the autophagic machinery and, in a process termed xenophagy, the host cell survives because autophagy is employed as a safeguard for pathogens that escaped phagosomes. However, some pathogens can manipulate the autophagic pathway and replicate within the niche of generated autophagosome-like vesicles. By automated fluorescence-based high content analyses, we demonstrate that Staphylococcus aureus strains (USA300, HG001, SA113) stimulate autophagy and become entrapped in intracellular PtdIns(3)P-enriched vesicles that are decorated with human WIPI-1, an essential PtdIns(3)P effector of canonical autophagy and membrane protein of both phagophores and autophagosomes. Further, agr-positive S. aureus (USA300, HG001) strains were more efficiently …

Chemoorganotrophic Bacteria Isolated From Biodeteriorated Surfaces In Cave And Catacombs, Filomena De Leo, Agnese Iero, Gabrielle Zammit, Clara E. Urzi

International Journal of Speleology

The main objective of this work was the comparative analysis of a large number of bacterial strains isolated from biodeteriorated surfaces in three different sites, namely the catacombs of St. Callistus in Rome, Italy, the catacombs dedicated to St. Agatha in Rabat, Malta and the Cave of Bats in Zuheros, Spain. Our results showed that even considering only culturable chemoorganotrophic bacteria the variability is very high, reflecting the great variety of microhabitats present. Hence any strategies to prevent, control or eliminate the biofilm-embedded microbiota from an archeological surface should take into account a number of considerations as stipulated in our …

Bacterial Community Survey Of Sediments At Naracoorte Caves, Australia, Eric M. Adetutu, Krystal Thorpe, Esmaeil Shahsavari, Steven Bourne, Xiangsheng Cao, Ramin Mazaheri Nezhad Fard, Greg Kirby, Andrew S. Ball

International Journal of Speleology

Bacterial diversity in sediments at UNESCO World Heritage listed Naracoorte Caves was surveyed as part of an investigation carried out in a larger study on assessing microbial diversity in caves. Cave selection was based on tourist accessibility; Stick Tomato and Alexandra Cave (> 15000 annual visits) and Strawhaven Cave was used as control (no tourist access). Microbial analysis showed that Bacillus was the most commonly detected microbial genus by culture dependent and independent survey of tourist accessible and inaccessible areas of show (tourist accessible) and control caves. Sediment bacterial groups were assigned to the Firmicutes, Actinobacteria and Proteobacteria. The survey …

Characterization Of The Lone Extracytoplasmic Function Sigma Factor, Ós, And Its Role In The Staphylococcus Aureus Virulence And Stress Responses, Halie Kay Miller

USF Tampa Graduate Theses and Dissertations

Previously our laboratory had identified a novel component of the Staphylococcus aureus regulatory network, an extracytoplasmic function ó factor, óS, involved in stress response and disease causation. Here we present additional characterization of óS, demonstrating a role for it in protection against DNA damage, cell wall disruption and interaction with components of the innate immune system. Promoter mapping reveals the existence of four unique sigS start sites, one of which appears to be subject to auto-regulation. Transcriptional profiling revealed that sigS expression remains low in a number of S. aureus wild-types, but is upregulated in the highly mutated strain RN4220. …

Investigation Of Reservoirs Of Fecal Indicator Bacteria And Water Quality On The Presence Of Allochthonous Pathogens And The Ecology And Virulence Of Vibrio Vulnificus, Christopher Staley

USF Tampa Graduate Theses and Dissertations

The quality of recreational and shellfishing waters has historically been monitored using commensal, allochthonous bacteria shed in feces (fecal indicator bacteria, FIB). The fate of FIB in the environment should mimic that of bacterial, protozoan, and viral human pathogens, which may also be allochthonous (e.g. Salmonella, Cryptosporidium, or enteric viruses) or autochthonous (e.g. Vibrio spp.) to aquatic environments. FIB are contributed to water from human and animal sources; however, pollution source cannot be determined by conventional FIB measurements. Because fecal source determination is important for pollution remediation and assessment of human health risks, microbial source tracking (MST) methods are …