Item Open AccessTherapeutic efficacy of antimalarial drugs targeting DosRS signaling in Mycobacterium abscessus(Colorado State University. Libraries, 2022-02-23) Belardinelli, Juan Manuel, author; Verma, Deepshikha, author; Li, Wei, author; Avanzi, Charlotte, author; Wiersma, Crystal J., author; Williams, John T., author; Johnson, Benjamin K., author; Zimmerman, Matthew, author; Whittel, Nicholas, author; Angala, Bhanupriya, author; Wang, Han, author; Jones, Victoria, author; Dartois, Veronique, author; de Moura, Vinicius C. N., author; Gonzalez-Juarrero, Mercedes, author; Pearce, Camron, author; Schenkel, Alan R., author; Malcolm, Kenneth C., author; Nick, Jerry A., author; Charman, Susan A., author; Wells, Timothy N. C., author; Podell, Brendan K., author; Vennerstrom, Jonathan L., author; Ordway, Diane J., author; Abramovitch, Robert B., author; Jackson, Mary, authorA search for alternative Mycobacterium abscessus treatments led to our interest in the two-component regulator DosRS, which, in Mycobacterium tuberculosis, is required for the bacterium to establish a state of nonreplicating, drug-tolerant persistence in response to a variety of host stresses. We show here that the genetic disruption of dosRS impairs the adaptation of M. abscessus to hypoxia, resulting in decreased bacterial survival after oxygen depletion, reduced tolerance to a number of antibiotics in vitro and in vivo, and the inhibition of biofilm formation. We determined that three antimalarial drugs or drug candidates, artemisinin, OZ277, and OZ439, can target DosS-mediated hypoxic signaling in M. abscessus and recapitulate the phenotypic effects of genetically disrupting dosS. OZ439 displayed bactericidal activity comparable to standard-of-care antibiotics in chronically infected mice, in addition to potentiating the activity of antibiotics used in combination. The identification of antimalarial drugs as potent inhibitors and adjunct inhibitors of M. abscessus in vivo offers repurposing opportunities that could have an immediate impact in the clinic. Item Open AccessMolecular epidemiology of leprosy: an update(Colorado State University. Libraries, 2020-12) Avanzi, Charlotte, author; Singh, Pushpendra, author; Truman, Richard W, author; Suffys, Philip N, authorMolecular epidemiology investigations are notoriously challenging in the leprosy field mainly because the inherent characteristics of the disease as well as its yet uncultivated causative agents, Mycobacterium leprae and M. lepromatosis. Despite significant developments in understanding the biology of leprosy bacilli through genomic approaches, the exact mechanisms of transmission is still unclear and the factors underlying pathological variation of the disease in different patients remain as major gaps in our knowledge about leprosy. Despite these difficulties, the last two decades have seen the development of genotyping procedures based on PCR-sequencing of target loci as well as by the genome-wide analysis of an increasing number of geographically diverse isolates of leprosy bacilli. This has provided a foundation for molecular epidemiology studies that are bringing a better understanding of strain evolution associated with ancient human migrations, and phylogeographical insights about the spread of disease globally. This review discusses the advantages and drawbacks of the main tools available for molecular epidemiological investigations of leprosy and summarizes various methods ranging from PCR-based genotyping to genome-typing techniques. We also describe their main applications in analyzing the short-range and long-range transmission of the disease. Finally, we summarise the current gaps and challenges that remain in the field of molecular epidemiology of leprosy. Item Open AccessQuantifying proximity, confinement, and interventions in disease outbreaks: a decision support framework for air-transported pathogens(Colorado State University. Libraries, 2021-02-19) Bond, Tami C, author; Bosco-Lauth, Angela, author; Farmer, Delphine K., author; Francisco, Paul W., author; Pierce, Jeﬀrey R., author; Fedak, Kristen M., author; Ham, Jay M., author; Jathar, Shantanu H., author; VandeWoude, Sue, author; Environmental Science & Technology, publisherThe inability to communicate how infectious diseases are transmitted in human environments has triggered avoidance of interactions during the COVID-19 pandemic. We define a metric, Effective ReBreathed Volume (ERBV), that encapsulates how infectious pathogens, including SARS-CoV-2, transport in air. ERBV separates environmental transport from other factors in the chain of infection, allowing quantitative comparisons among situations. Particle size affects transport, removal onto surfaces, and elimination by mitigation measures, so ERBV is presented for a range of exhaled particle diameters: 1, 10, and 100 μm. Pathogen transport depends on both proximity and confinement. If interpersonal distancing of 2 m is maintained, then confinement, not proximity, dominates rebreathing after 10–15 min in enclosed spaces for all but 100 μm particles. We analyze strategies to reduce this confinement effect. Ventilation and filtration reduce person-to-person transport of 1 μm particles (ERBV1) by 13–85% in residential and office situations. Deposition to surfaces competes with intentional removal for 10 and 100 μm particles, so the same interventions reduce ERBV10 by only 3–50%, and ERBV100 is unaffected. Prior knowledge of size-dependent ERBV would help identify transmission modes and effective interventions. This framework supports mitigation decisions in emerging situations, even before other infectious parameters are known. Item Open AccessEukaryotic microbiology(Colorado State University. Libraries, 2020-09-21) Huseby, Medora, authorThis chapter details cell-cell communication highlighting studies performed in the model organism Dictyostelium discoideum. Topics covered include signaling molecules, cell surface receptors, and subversion of host cell signaling by pathogens. Item Open AccessMedical and molecular virology(Colorado State University. Libraries, 2020-06-30) Suchman, Erica, authorThis book was created for upper division microbiology students studying virology. It will describe the molecular biology and major diseases of virus families that cause significant disease in animals and humans. This book is by no means meant to be exhaustive. In fact, because virology can be so overwhelming, the author has tried to keep the book as simple as possible, while still giving the reader a solid understanding of the molecular mechanisms of viral replication and pathogenesis. This book is different than other virology textbooks in that it is laid out grouping viruses by how they replicate. This should hopefully allow you, the reader, to think about why these viruses replicate similarly, and why replication strategies may be different than those used by other viruses. Item Open AccessThe mmpL3 interactome reveals a complex crosstalk between cell envelope biosynthesis and cell elongation and division in mycobacteria(Colorado State University. Libraries, 2019-07-24) Belardinelli, Juan Manuel, author; Stevens, Casey M., author; Li, Wei, author; Tan, Yong Zi, author; Jones, Victoria, author; Mancia, Filippo, author; Zgurskaya, Helen I., author; Jackson, Mary, author; Springer Nature Publishing, publisherIntegral membrane transporters of the Mycobacterial Membrane Protein Large (MmpL) family and their interactome play important roles in the synthesis and export of mycobacterial outer membrane lipids. Despite the current interest in the mycolic acid transporter, MmpL3, from the perspective of drug discovery, the nature and biological significance of its interactome remain largely unknown. We here report on a genome-wide screening by two-hybrid system for MmpL3 binding partners. While a surprisingly low number of proteins involved in mycolic acid biosynthesis was found to interact with MmpL3, numerous enzymes and transporters participating in the biogenesis of peptidoglycan, arabinogalactan and lipoglycans, and the cell division regulatory protein, CrgA, were identified among the hits. Surface plasmon resonance and co-immunoprecipitation independently confirmed physical interactions for three proteins in vitro and/or in vivo. Results are in line with the focal localization of MmpL3 at the poles and septum of actively-growing bacilli where the synthesis of all major constituents of the cell wall core are known to occur, and are further suggestive of a role for MmpL3 in the coordination of new cell wall deposition during cell septation and elongation. This novel aspect of the physiology of MmpL3 may contribute to the extreme vulnerability and high therapeutic potential of this transporter. Item Open AccessEvidence of zoonotic leprosy in Pará, Brazilian Amazon, and risks associated with human contact or consumption of armadillos(Colorado State University. Libraries, 2018-06-28) da Silva, Moises B., author; Portela, Juliana M., author; Li, Wei, author; Jackson, Mary, author; Gonzalez-Juarrero, Mercedes, author; Hidalgo, Andrea Sánchez, author; Belisle, John T., author; Bouth, Raquel C., author; Gobbo, Angélica R., author; Barreto, Josafá G., author; Minervino, Antonio H. H., author; Cole, Stewart T., author; Avanzi, Charlotte, author; Busso, Philippe, author; Frade, Marco A. C., author; Geluk, Annemieke, author; Salgado, Claudio G., author; Spencer, John S., author; Public Library of Science, publisherMycobacterium leprae (M. leprae) is a human pathogen and the causative agent for leprosy, a chronic disease characterized by lesions of the skin and peripheral nerve damage. Zoonotic transmission of M. leprae to humans by nine-banded armadillos (Dasypus novemcinctus) has been shown to occur in the southern United States, mainly in Texas, Louisiana, and Florida. Nine-banded armadillos are also common in South America, and residents living in some areas in Brazil hunt and kill armadillos as a dietary source of protein. This study examines the extent of M. leprae infection in wild armadillos and whether these New World mammals may be a natural reservoir for leprosy transmission in Brazil, similar to the situation in the southern states of the U.S. The presence of the M. leprae-specific repetitive sequence RLEP was detected by PCR amplification in purified DNA extracted from armadillo spleen and liver tissue samples. A positive RLEP signal was confirmed in 62% of the armadillos (10/16), indicating high rates of infection with M. leprae. Immunohistochemistry of sections of infected armadillo spleens revealed mycobacterial DNA and cell wall constituents in situ detected by SYBR Gold and auramine/rhodamine staining techniques, respectively. The M. leprae-specific antigen, phenolic glycolipid I (PGL-I) was detected in spleen sections using a rabbit polyclonal antibody specific for PGL-I. Anti-PGL-I titers were assessed by ELISA in sera from 146 inhabitants of Belterra, a hyperendemic city located in western Pará state in Brazil. A positive anti-PGL-I titer is a known biomarker for M. leprae infection in both humans and armadillos. Individuals who consumed armadillo meat most frequently (more than once per month) showed a significantly higher anti-PGL-I titer than those who did not eat or ate less frequently than once per month. Armadillos infected with M. leprae represent a potential environmental reservoir. Consequently, people who hunt, kill, or process or eat armadillo meat are at a higher risk for infection with M. leprae from these animals. Item Open AccessPathogenesis of oral FIV infection(Colorado State University. Libraries, 2017) Miller, Craig, author; Boegler, Karen, author; Carver, Scott, author; MacMillan, Martha, author; Bielefeldt-Ohmann, Helle, author; VandeWoude, Susan, authorFeline immunodeficiency virus (FIV) is the feline analogue of human immunodeficiency virus (HIV) and features many hallmarks of HIV infection and pathogenesis, including the development of concurrent oral lesions. While HIV is typically transmitted via parenteral transmucosal contact, recent studies prove that oral transmission can occur, and that saliva from infected individuals contains significant amounts of HIV RNA and DNA. While it is accepted that FIV is primarily transmitted by biting, few studies have evaluated FIV oral infection kinetics and transmission mechanisms over the last 20 years. Modern quantitative analyses applied to natural FIV oral infection could significantly further our understanding of lentiviral oral disease and transmission. We therefore characterized FIV salivary viral kinetics and antibody secretions to more fully document oral viral pathogenesis. Our results demonstrate that: (i) saliva of FIV-infected cats contains infectious virus particles, FIV viral RNA at levels equivalent to circulation, and lower but significant amounts of FIV proviral DNA; (ii) the ratio of FIV RNA to DNA is significantly higher in saliva than in circulation; (iii) FIV viral load in oral lymphoid tissues (tonsil, lymph nodes) is significantly higher than mucosal tissues (buccal mucosa, salivary gland, tongue); (iv) salivary IgG antibodies increase significantly over time in FIV-infected cats, while salivary IgA levels remain static; and, (v) saliva from naïve Specific Pathogen Free cats inhibits FIV growth in vitro. Collectively, these results suggest that oral lymphoid tissues serve as a site for enhanced FIV replication, resulting in accumulation of FIV particles and FIV-infected cells in saliva. Failure to induce a virus-specific oral mucosal antibody response, and/or viral capability to overcome inhibitory components in saliva may perpetuate chronic oral cavity infection. Based upon these findings, we propose a model of oral FIV pathogenesis and suggest alternative diagnostic modalities and translational approaches to study oral HIV infection. Item Open AccessThe method of attachment influences accelerometer-based activity data in dogs(Colorado State University. Libraries, 2017-02-10) Martin, Kyle W., author; Olsen, Anastasia M., author; Duncan, Colleen G., author; Duerr, Felix M., author; BioMed Central, publisherBackground: Accelerometer-based activity monitoring is a promising new tool in veterinary medicine used to objectively assess activity levels in dogs. To date, it is unknown how device orientation, attachment method, and attachment of a leash to the collar holding an accelerometer affect canine activity data. It was our goal to evaluate whether attachment methods of accelerometers affect activity counts. Eight healthy, client-owned dogs were fitted with two identical neck collars to which two identical activity monitors were attached using six different methods of attachment. These methods of attachment evaluated the use of a protective case, positioning of the activity monitor and the tightness of attachment of the accelerometer. Lastly, the effect of leash attachment to the collar was evaluated. For trials where the effect of leash attachment to the collar was not being studied, the leash was attached to a harness. Activity data obtained from separate monitors within a given experiment were compared using Pearson correlation coefficients and across all experiments using the Kruskal-Wallis Test. Results: There was excellent correlation and low variability between activity monitors on separate collars when the leash was attached to a harness, regardless of their relative positions. There was good correlation when activity monitors were placed on the same collar regardless of orientation. There were poor correlations between activity monitors in three experiments: when the leash was fastened to the collar that held an activity monitor, when one activity monitor was housed in the protective casing, and when one activity monitor was loosely zip-tied to the collar rather than threaded on using the provided metal loop. Follow-up, pair-wise comparisons identified the correlation associated with these three methods of attachment to be statistically different from the level of correlation when monitors were placed on separate collars. Conclusions: While accelerometer-based activity monitors are useful tools to objectively assess physical activity in dogs, care must be taken when choosing a method to attach the device. The attachment of the activity monitor to the collar should utilize a second, dedicated collar that is not used for leash attachment and the attachment method should remain consistent throughout a study period.