Microbiology Commons^™

Pharky, Phedro, Phractured, And Stagephright: Complete Genome Sequences Of Four Ek2 Cluster Microbacterium Phages, Susan M. R. Gurney, Medha Gupta, Deeksha Reddy, Gloria Zacharias, Aleda S. Jomy, Carly J. Smith, Matthew T. Mcdonald, Brett M. Condon

Student Papers, Posters & Projects

We report the complete genome sequence of four Microbacterium phages, Pharky, Phedro, Phractured, and StagePhright, isolated using the bacterial host Microbacterium foliorum from soil samples collected in Philadelphia. All four phages are genetically very similar and contain a gene of unknown function, which is uncommon in the EK2 cluster.

Results Of Tor001: An Open-Label Single Patient Study Using Targeted Bacteriophage Therapy For The Treatment Of A Chronic Urinary Tract Infection, Jonathan D Cook, Payton B Hooey, Keiko C Salazar, Justin R Clark, Umesh Jain, Carlos Fernando, Hanjeong Harvey, Alexander Cervantes, Stephanie Omagari, Udi Blankstein, Andrew M Kropinski, Alan R Davidson, Karen L Maxwell, Lori L Burrows, Austen L Terwilliger, Anthony W Maresso, Greg J German

Faculty, Staff and Students Publications

Chronic urinary tract infections are persistent bacterial infections with the potential to drive antibiotic resistance. Like other persistent bacterial infections, intracellular bacterial reservoirs and biofilm formation hinder the clearance of pathogens despite long courses of antibiotic therapy. New strategies for treatment of these persistent infections are needed. Here we describe the results of an open-label individual patient study using bacteriophage therapy to treat a chronic urinary tract infection in an immunocompetent 72-year-old woman without underlying urolithiasis or indwelling devices. We co-administered HP3.1, HP3, and ES19 bacteriophages with demonstrated in vitro activity via bladder instillation, orally, and as a topical formulation. …

Seasonal Detection Of Coliphages In Freshwater Bodies At Coastal Carolina University, Ella Caldwell

Honors Theses

Bacteriophages, commonly referred to as phages, are non-living viruses that only infect bacteria. As the most abundant biological entities on Earth, understanding their role in regulating microbial populations is essential. This ongoing research aims to identify naturally occurring bacteriophages that could effectively control bacteria blooms. The primary objective of this study is to detect coliphages on Coastal Carolina University's campus and determine their viral families. Coliphages are a subgroup of bacteriophages that specifically infect coliform bacteria. Water samples were collected across multiple sites on campus, and polymerase chain reaction (PCR) was employed to amplify the viral DNA from the potential …

An In-Depth Investigation Into Yersinia Pestis Phages: Isolating A Yersinia Pestis Phage From Soil And Constructing A Comprehensive Phylogenetic Analysis Of Known Yersinia Pestis Phages, Jacqueline Chavez

All HCAS Student Capstones, Theses, and Dissertations

Yersinia pestis, the causative agent of the plague, has become increasingly resistant to many antibiotics; therefore, alternatives such as phage therapy are necessary. Phage therapy uses combinations of bacteriophages or phages, which are viruses that only infect bacteria. However, phages—especially Y. pestis phages —are severely understudied. Thus, this research has sought to specifically isolate a phage from soil that infects Y. pestis. There were 146 samples that were collected from across the US, focusing on those with reported cases of plague. These samples were enriched with Y. pestis to maximize the amount of potential phage that target Y. pestis. …

Construction And Characterization Of Dna Libraries From Cultured Phages And Environmental Viromes, Carmen Gu Liu, Brianna E Thompson, James D Chang, Lorna Min, Anthony W Maresso

Faculty, Staff and Students Publications

Despite many efforts to understand and leverage the functional potential of environmental viromes, most bacteriophage genes are largely uncharacterized. To explore novel biology from uncultivated microbes like phages, metagenomics has emerged as a powerful tool to directly mine new genes without the need to culture the diverse microbiota and the viruses within. When a pure computational approach cannot infer gene function, it may be necessary to create a DNA library from environmental genomic DNA, followed by the screening of that library for a particular function. However, these screens are often initiated without a metagenomic analysis of the completed DNA library …

A Case Of Persistent Intra-Abdominal Stenotrophomonas Maltophilia Infection Despite Bacteriophage Therapy, Grace D Cullen, Keiko Christine Salazar, Austen Lee Terwilliger, Saima Aslam, Justin R Clark, Anthony W Maresso, Paul L Bollyky, Jenny R Aronson

Faculty, Staff and Students Publications

Multidrug resistant infections are a challenge in the health care setting and a cause of patient morbidity and mortality. Bacteriophages (phages) are viruses that target and kill bacteria and have been used in patients to treat bacterial infections. We present a case of disseminated Stenotrophomonas maltophilia infection, with pulmonary, intra-abdominal and bloodstream involvement. The patient was treated with a combination of antibiotics and personalized phage therapy, administered daily for 12 days both intravenously as well as via intra-abdominal drains. Phage therapy was well-tolerated, the patient cleared S. maltophilia from their bloodstream and their intra-abdominal abscesses were stable or decreased in …

Appelmans Protocol – A Directed In Vitro Evolution Enables Induction And Recombination Of Prophages With Expanded Host Range, Thao Nguyen Vu, Justin Ryan Clark, Eris Jang, Roshan D'Souza, Le Phuong Nguyen, Naina Adren Pinto, Seongjun Yoo, Ricardo Abadie, Anthony William Maresso, Dongeun Yong

Faculty, Staff and Students Publications

Infections caused by carbapenem-resistant Acinetobacter baumannii (CRAB) present significant healthcare challenges due to limited treatment options. Bacteriophage (phage) therapy offers potential as an alternative treatment. However, the high host specificity of phages poses challenges for their therapeutic application. To broaden the phage spectrum, laboratory-based phage training using the Appelmans protocol was employed in this study. As a result, the protocol successfully expanded the host range of a phage cocktail targeting CRAB. Further analysis revealed that the expanded host range phages isolated from the output cocktail were identified as recombinant derivatives originating from prophages induced from encountered bacterial strains. These findings …

Using Single Cell Genomics To Explore The Impact Of Marine Viruses On Microbial Respiration., Paxton Tomko

MCB Articles

Viral metabolic reprograming of marine prokaryotes, through the use of virally encoded auxiliary metabolic genes (AMGs), plays a critical role in marine ecosystem function by influencing biochemical cycles and genetic diversity in these environments. Despite the fundamental role viruses play in global environmental ecosystems, they remain an understudied aspect of microbial ecology and evolution, in part due to the methods available for studying virus host interactions in natural systems. Thus far, metagenomic analyses have been used to study the interactions of virus host pairs, but these types of analyses have their limitations in accurately linking viruses to hosts, or culture-based …

Harnessing Antagonistic Pleiotropy: Tolc-Dependent Coliphages Can Drive Reduced Antibiotic Resistance In E. Coli, Cooper Mckenna

CMC Senior Theses

The rise of multidrug-resistant (MDR) bacterial pathogens has been a growing concern in healthcare around the world, and bacteriophage (phage) therapy is a promising method to combat MDR infections. One defense mechanism of E. coli against antibiotics is the TolC-AcrAB efflux pump. At the same time, some phages use TolC as a surface receptor to infect the cell. This puts TolC at a crossroads in which it is beneficial to the bacteria to protect against antibiotics but harmful by allowing phage infection, creating the potential for evolutionary trade-offs. Previous research has identified only three TolC-dependent phages and shown phage-resistant E. …

A Novel Subcluster Of Closely Related Bacillus Phages With Distinct Tail Fiber/Lysin Gene Combinations, Rachel E. Loney, Veronique A. Delesalle, Brianne E. Chaudhry, Megan Czerpak, Alexandra A. Guffey, Leo Goubet-Mccall, Michael Mccarty, Madison S. Strine, Natalie T. Tanke, Albert C. Vill, Gregory P. Krukonis

Biology Faculty Publications

Bacteriophages (phages) are the most numerous entities on Earth, but we have only scratched the surface of describing phage diversity. We isolated seven Bacillus subtilis phages from desert soil in the southwest United States and then sequenced and characterized their genomes. Comparative analyses revealed high nucleotide and amino acid similarity between these seven phages, which constitute a novel subcluster. Interestingly, the tail fiber and lysin genes of these phages seem to come from different origins and carry out slightly different functions. These genes were likely acquired by this subcluster of phages via horizontal gene transfer. In conjunction with host range …

Analyzing Pseudomonas Aeruginosa With Bacteriophage Tags Using Photoacoustic Flow Cytometry, Jennifer C. Schinke

Electronic Theses and Dissertations

The number of daily bacterial infections is climbing and the CDC explains that this is due to the antibiotic-resistant threat in the United States. Finding a faster way of bacterial identification is necessary as it currently takes 1-4 days for a medical lab to culture and identify bacteria. Photoacoustic flow cytometry (PAFC) can be used as an alternative method resulting in swift identification within an hour (Edgar, 2019). Pseudomonas aeruginosa, cell line PA01, will be coated in up to a few hundred red dyed phages making it detectible by the photoacoustic flow cytometry system. Bacteriophages (phages) are viruses that …

The Persistence And Stabilization Of Auxiliary Genes In The Human Skin Virome, Ema H. Graham, Wesley A. Tom, Alison C. Neujahr, Michael Adamowicz, Jennifer Clarke, Joshua R. Herr, Samodha C. Fernando

Nebraska Center for Virology: Faculty Publications

Background The human skin contains a diverse microbiome that provides protective functions against environmental pathogens. Studies have demonstrated that bacteriophages modulate bacterial community composition and facilitate the transfer of host-specific genes, potentially influencing host cellular functions. However, little is known about the human skin virome and its role in human health. Especially, how viral-host relationships influence skin microbiome structure and function is poorly understood.

Results Population dynamics and genetic diversity of bacteriophage communities in viral metagenomic data collected from three anatomical skin locations from 60 subjects at five different time points revealed that cutaneous bacteriophage populations are mainly composed of …

Isolation And Characterization Of Chi-Like Salmonella Bacteriophages Infecting Two Salmonella Enterica Serovars, Typhimurium And Enteritidis, Addisu D. Teklemariam, Mona G. Alharbi, Rashad R. Al-Hindi, Ibrahim Alotibi, Abdullah A. Aljaddawi, Sheren A. Azhari, Ahmed Esmael

Nebraska Center for Virology: Faculty Publications

Salmonella enterica Serovar Typhimurium and Salmonella enterica Serovar Enteritidis are well-known pathogens that cause foodborne diseases in humans. The emergence of antibiotic-resistant Salmonella serovars has caused serious public health problems worldwide. In this study, two lysogenic phages, STP11 and SEP13, were isolated from a wastewater treatment plant in Jeddah, KSA. Transmission electron microscopic images revealed that both phages are new members of the genus “Chivirus” within the family Siphoviridae. Both STP11 and SEP13 had a lysis time of 90 min with burst sizes of 176 and 170 PFU/cell, respectively. The two phages were thermostable (0 ^◦C …

Forty Years Without Family: Three Novel Bacteriophages With High Similarity To Spp1 Reveal Decades Of Evolutionary Stasis Since The Isolation Of Their Famous Relative, Veronique A. Delesalle, Brianne E. Tomko, Albert C. Vill, Katherine B. Lichty, Gregory P. Krukonis

Biology Faculty Publications

SPP1, an extensively studied bacteriophage of the Gram-positive Bacillus subtilis, is a model system for the study of phage–host interactions. Despite progress in the isolation and characterization of Bacillus phages, no previously fully sequenced phages have shared more than passing genetic similarity to SPP1. Here, we describe three virulent phages very similar to SPP1; SPP1 has greater than 80% nucleotide sequence identity and shares more that 85% of its protein coding genes with these phages. This is remarkable, given more than 40 years between the isolation of SPP1 and these phages. All three phages have somewhat larger genomes and more …

Complete Genome Sequences Of Two Temperate Bacillus Subtilis Phages Isolated At Tumamoc Hill Desert Laboratory, Gregory P. Krukonis, Amanda K. Kemp, Katie F. Storrie, Vivian R. Chavira, Hayden W. Lantrip, Victoria D. Perez, Desiree A. Reyes, Julian A. Truax, Rachel Loney, Veronique A. Delesalle

Biology Faculty Publications

Bacteriophages are important in structuring bacterial communities, including desert soils dominated by Bacillus species. Here, we describe two genetically similar temperate phages isolated on a Bacillus subtilis strain from soil in Tucson, Arizona. Their double-stranded DNA (dsDNA) genomes contain 98 and 102 genes, with a set of 4 genes being found in only one phage.

Qualitative Proteomic And Genomic Analysis Of Klebsiella Pneumoniae Bacteriophage Kaomega, Emilee Carr

Undergraduate Honors Theses

Klebsiella pneumoniae is a Gram-negative Enterobacteriaceae that is a common nosocomial pathogen causing pneumonia, infections in the bloodstream, wound infections, and meningitis. It has developed natural resistance to multiple antibiotics, most notably carbapenems which are often seen as the last line of defense against multi-drug resistant pathogens. Bacteriophages are being investigated as a promising alternative treatment to antibiotics in fighting these resistant pathogens. KaOmega, a Klebsiella pneumoniae bacteriophage, was isolated, sequenced, and annotated to characterize and understand its potential for use in a phage therapy. Characterization included Phyre2 analysis to predict putative protein functions based on structural homology, burst size …

Genomic And Functional Characterization Of Vancomycin-Resistant Enterococci-Specific Bacteriophages In The Galleria Mellonella Wax Moth Larvae Model, Lynn El Haddad, Georgios Angelidakis, Justin R Clark, Jesus F Mendoza, Austen L Terwilliger, Christopher P Chaftari, Mark Duna, Serena T Yusuf, Cynthia P Harb, Mark Stibich, Anthony Maresso, Roy F Chemaly

Faculty, Staff and Students Publications

Phages are naturally occurring viruses that selectively kill bacterial species without disturbing the individual’s normal flora, averting the collateral damage of antimicrobial usage. The safety and the effectiveness of phages have been mainly confirmed in the food industry as well as in animal models. In this study, we report on the successful isolation of phages specific to Vancomycin-resistant Enterococci, including Enterococcus faecium (VREfm) and Enterococcus faecalis from sewage samples, and demonstrate their efficacy and safety for VREfm infection in the greater wax moth Galleria mellonella model. No virulence-associated genes, antibiotic resistance genes or integrases were detected in …

An Investigation Of The Novel Use Of Bacteriophages To Diagnose And Treat Johne's Disease In Cattle, Max Kevane-Campbell

ORBioM (Open Research BioSciences Meeting)

Johne’s disease is a scourge to dairy farmers all over the world. It is an infectious disease that causes chronic inflammation and lesions along the inside of the small intestine of, primarily, ruminant animals (i.e., cattle and sheep). It is an incurable disease and urgently requires new and radical intervention strategies. Apart from careful on-site farm management practices, little can be offered to farmers to reduce the risk of infection, and nothing short of livestock culling is effective once an animal becomes infected. Currently, there are no vaccines licensed in Ireland or antibiotic treatment strategies available for Johne’s disease. This …

Diversity Of Bacteriophage In Burkholderia Species, Abigail Price

Honors Projects

Bacteriophages are viruses that infect and replicate within bacteria and offer the potential of a therapeutic alternative to chronic infections that do not respond to antibiotic-based therapies. B. vietnamiensis is one of a number of Burkholderia species involved with chronic drug resistant infections in the lungs of individuals with compromised respiratory systems, as found in those with chronic obstructive pulmonary disease and, most especially, are of particular significance in patients with cystic fibrosis. The diversity of the Burkholderia species is explored by using online databases and looking at bacteriophage or phage-encoding viruses found in B. vietnamiensis. The open reading frames …

Aeromonas Phage Research And The Public Health Impact Of Antibiotics In Aquaculture Workers, Madelyn Merchant

Honors Projects

One of the most common fish diseases in aquaculture is Aeromonas infection. The most common way to treat this infection is through antibiotics. The bacteria in the fish can become antibiotic-resistant and perpetuate the disease. The diseases in fish create a huge financial loss and the industry loses $6 billion per year due to diseases in fish. An alternative to antibiotics is bacteriophage which causes less environmental degradation and is better for human gut flora. In aquaculture there have been examples of aquaculture workers becoming sick from the water in aquaculture ponds as well as from people eating the fish. …

Examining Bacteriophage Reliance On The Tola Protein Of Escherichia Coli, Ellen Davis

Honors Projects

With the ever-growing crisis of antibiotic resistant bacteria, bacteriophage provide an important potential alternative therapy to treat infections when antibiotics are ineffective. For this reason, it is important to understand the mechanisms bacteriophage use to enter their bacterial host cell. The cellular envelope of the gram-negative bacterium E. coli contains a protein system known as Tol-Pal, which utilizes energy from the Proton Motive Force to carry out several cellular functions, including some associated with cellular division and outer-membrane stability. Certain bacteriophage take advantage of the energy produced by this system to cross the bacterial cell membrane. One important protein of …

The Isolation And Characterization Of Bacteriophage Hasitha, Gillian Brown

Mahurin Honors College Capstone Experience/Thesis Projects

Microbacteriophage Hasitha is a virus that infects Microbacterium foliorum, a bacterium associated with grasses that was first discovered in Germany. Hasitha was isolated from an enriched compost sample and is of particular interest due to its unusual growth pattern. Most bacteriophages require actively growing host cells to produce new phage progeny. However, Hasitha can infect and kill stationary (non-replicating) bacterial cells. We discovered this unusual characteristic through a fortuitous observation of infected lawns that were allowed to incubate in the lab workspace for approximately one month. During this time, a noticeable “halo” grew around the initial site of infection …

Isolating Bacteriophage For Potential Treatment Of Chronic Multi-Drug Resistant Escherichia Coli Infections, Leila Oswalt

Honors Projects

The misuse and overuse of antibiotics has led to the intense rise in antibiotic resistance. As society transitions into the post antibiotic era, there will be a great need for new therapeutic strategies to address multiple drug resistant bacterial infections. One such method, called bacteriophage therapy, allows for specific targeting of certain pathogenic bacteria through the use of viruses that attack bacteria; termed “bacteriophage” or simply “phage”. Urinary tract infections are among the most common pathological human infections that rely heavily on the use of antibiotics, the major cause of which is the bacterium Escherichia coli. During the Spring 2021-Spring …

Complete Genome Sequence Of The Pantoea Phage Ah07, Gregory P. Krukonis, Sam J. Roth, Veronique A. Delesalle

Biology Faculty Publications

Bacteriophages of the phyllosphere have not been extensively described, despite their role in bacterial communities on this plant organ. Here, we describe a temperate Pantoea phage, AH07, that was isolated from the leaves of horse chestnut trees. The 37,859-bp linear double-stranded DNA genome contains 58 putative genes, including an integration cassette.

Complete Genome Sequences Of Four Phages Of The Horse Chestnut Phyllosphere, Gregory P. Krukonis, Sam J. Roth, Veronique A. Delesalle

Biology Faculty Publications

Bacteriophages play important roles in determining bacterial communities, including plant microbiota. Here, we describe four lytic phages, three Siphoviridae and one Podoviridae, isolated from four different bacterial species found on the leaves of horse chestnut trees. Their double-stranded DNA (dsDNA) genomes range from 39,095 to 46,062 bp and contain 51 to 70 genes.

Genome Sequences Of Erwinia Phyllophages Ah04 And Ah06, Gregory P. Krukonis, Sam J. Roth, Veronique A. Delesalle

Biology Faculty Publications

Although crucial in shaping bacterial communities, few bacteriophages of the phyllosphere have been described. We provide genome data for two Myoviridae phages, AH04 and AH06, isolated on Erwinia billingiae strains. AH04 shares limited genetic similarity with previously described phages, while AH06 shares over 75% similarity with various Erwinia phages.

F Plasmids In Escherichia Coli Decrease Permissivity To Coliphage, Cesar Eber Montelongo Hernandez

Dissertations

The urinary tract contains a community of bacteria called the urinary microbiota (urobiota) thatmay be relevant to health; the genomic component of the urobiota is the urinary microbiome (urobiome). Urinary bacteria have been associated with both asymptomatic states and disease conditions, such as urinary tract infection (UTI), overactive bladder (OAB), and urge urinary incontinence (UUI). Some bacteria, such as E. coli, are considered urinary pathogens (uropathogens) but also can be commensals. Bacteriophage (phage) are ubiquitous in nature and likely shape bacterial populations in every niche; thus, phage may be one factor that modulates the urobiota. Phages have a specific host …

Innovative Approaches In The Discovery Of Aquatic Mycobacteriophages, Janis H. Doss

Biomedical Sciences Theses & Dissertations

Bacteriophages (phages), viruses that infect bacteria, have many applications in medicine, agriculture, molecular biology, and other fields. As antibiotic resistance becomes an increasing problem, interest in phages has grown. The traditional techniques of phage discovery are successful for some phages, but others require modified procedures to achieve detectable host infection.

Mycobacterium is a diverse bacterial genus characterized by a unique cell wall containing mycolic acids, which aids in survival and pathogenesis. The aims of the present research were to isolate mycobacteriophages, use bioinformatics techniques to analyze mycobacterial prophages, and combine genetic analysis with multi-well plate host range studies to identify …

Lysogeny And Use Of Mycobacteriophage Pita2, Eleanor Behling, Neocles B. Leontis, Jill Zeilstra-Ryalls

Honors Projects

The CDC has classified antibiotic resistance as the biggest health challenge of our era; every year 2 million lives are impacted and even lost due to resistant bacteria. Bacteriophages provide an alternative route to fighting infections that does not further the development of antibiotic resistance among bacterial species. A bacteriophage replicates inside a bacterial cell and then causes that cell to lyse, an event that kills the bacterial host. However, some phage can integrate their genomes into the host chromosome without causing lysis. The HHMI SEA-PHAGES program has generated a collection of bacteriophage that infect Actinobacteria species. Over 13,000 phages …

Isolation, Genetic Characterization And Clinical Application Of Bacteriophages Of Pathogenic Bacterial Species, Trever Leon Thurgood

Theses and Dissertations

Bacteriophages (phages) are the smallest biological entity on the planet. They provide vast amounts of valuable knowledge to biologists. Phage genomes are relatively simple compared to the organisms they infect (prokaryotes) and yet continually point to the complexity surrounding molecular- and microbiological mechanisms of life. By studying phages we can learn of the systems of gene expression, protein interaction and DNA organization. Phages are useful not only from an academic perspective, but may also have useful clinical applications. In the face of the rise of antibiotic-resistant bacterial “super pathogens”, scientists and researchers turn to phages as alternative treatments to these …