Last data update: Oct 28, 2024. (Total: 48004 publications since 2009)
Records 1-26 (of 26 Records) |
Query Trace: Metcalfe MG[original query] |
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A tale of two viruses: Coinfections of monkeypox and varicella zoster virus in the Democratic Republic of Congo
Hughes CM , Liu L , Davidson WB , Radford KW , Wilkins K , Monroe B , Metcalfe MG , Likafi T , Lushima RS , Kabamba J , Nguete B , Malekani J , Pukuta E , Karhemere S , Muyembe Tamfum JJ , Wemakoy EO , Reynolds MG , Schmid DS , McCollum AM . Am J Trop Med Hyg 2020 104 (2) 604-611 Recent enhanced monkeypox (MPX) surveillance in the Democratic Republic of Congo, where MPX is endemic, has uncovered multiple cases of MPX and varicella zoster virus (VZV) coinfections. The purpose of this study was to verify if coinfections occur and to characterize the clinical nature of these cases. Clinical, epidemiological, and laboratory results were used to investigate MPX/VZV coinfections. A coinfection was defined as a patient with at least one Orthopoxvirus/MPX-positive sample and at least one VZV-positive sample within the same disease event. Between September 2009 and April 2014, 134 of the 1,107 (12.1%) suspected MPX cases were confirmed as MPX/VZV coinfections. Coinfections were more likely to report symptoms than VZV-alone cases and less likely than MPX-alone cases. Significantly higher lesion counts were observed for coinfection cases than for VZV-alone but less than MPX-alone cases. Discernible differences in symptom and rash severity were detected for coinfection cases compared with those with MPX or VZV alone. Findings indicate infection with both MPX and VZV could modulate infection severity. Collection of multiple lesion samples allows for the opportunity to detect coinfections. As this program continues, it will be important to continue these procedures to assess variations in the proportion of coinfected cases over time. |
A fatal case of disseminated microsporidiosis due to Anncaliia algerae in a renal and pancreas allograft recipient
Anderson NW , Muehlenbachs A , Arif S , Bruminhent J , Deziel PJ , Razonable RR , Wilhelm MP , Metcalfe MG , Qvarnstrom Y , Pritt BS . Open Forum Infect Dis 2019 6 (7) ofz285 Microsporidiosis is an emerging opportunistic infection in immunocompromised patients. We report a case of fatal disseminated Anncaliia algerae infection in a profoundly immunosuppressed pancreas and kidney transplant recipient. |
Influenza virus exploits tunneling nanotubes for cell-to-cell spread.
Kumar A , Kim JH , Ranjan P , Metcalfe MG , Cao W , Mishina M , Gangappa S , Guo Z , Boyden ES , Zaki S , York I , Garcia-Sastre A , Shaw M , Sambhara S . Sci Rep 2017 7 40360 Tunneling nanotubes (TNTs) represent a novel route of intercellular communication. While previous work has shown that TNTs facilitate the exchange of viral or prion proteins from infected to naive cells, it is not clear whether the viral genome is also transferred via this mechanism and further, whether transfer via this route can result in productive replication of the infectious agents in the recipient cell. Here we present evidence that lung epithelial cells are connected by TNTs, and in spite of the presence of neutralizing antibodies and an antiviral agent, Oseltamivir, influenza virus can exploit these networks to transfer viral proteins and genome from the infected to naive cell, resulting in productive viral replication in the naive cells. These observations indicate that influenza viruses can spread using these intercellular networks that connect epithelial cells, evading immune and antiviral defenses and provide an explanation for the incidence of influenza infections even in influenza-immune individuals and vaccine failures. |
Oblitimonas alkaliphila gen. nov., sp. nov., a novel genus in the Pseudomonadaceae family recovered from a historical collection of previously unidentified clinical strains
Drobish AM , Emery BD , Whitney AM , Lauer AC , Metcalfe MG , McQuiston JR . Int J Syst Evol Microbiol 2016 66 (8) 3063-3070 Eight Gram-negative bacteria (B4199T, C6819, C6918, D2441, D3318, E1086, E1148 and E5571) were identified during a retrospective study of unidentified strains from a historical collection held in the Special Bacteriology Reference Laboratory at the Centers for Disease Control and Prevention. The strains were isolated from eight patients: five female, two male and one not specified. No ages were indicated for the patients. The sources were urine (3), leg tissue (2), foot wound, lung tissue and deep liver. The strains originated from seven different states across the United States of America (Colorado, Connecticut (2), Indiana, North Carolina, Oregon and Pennsylvania). The strains grew at 10 to 42 degrees C, were non-motile, alkalitolerant, slightly halophilic, microaerophilic, catalase and oxidase positive. The DNA G + C content was 47.3 to 47.8 mol%. The major cellular fatty acids were tetradecanoic acid (C14:0), hexadecanoic acid (C16:0) and 11-octadecenoic acid (C18:1omega7c). Polar lipids detected were phosphatidylglycerol (PG), phosphatidylethanolamine (PE), diphosphatidylglycerol (DPG) and an unknown phospholipid; the only respiratory quinone detected was the ubiquinone Q-9 (100 %). 16S rRNA gene sequence analysis produced results with 95.6 % similarity to Pseudomonas caeni (DSM 24390T) and 95.2 % similarity to Thiopseudomonas denitrificans (X2T). The results of the biochemical, chemotaxonomic and phylogenetic analyses between the study strains and some related type strains indicate that these strains represent a novel species of a new genus within the family Pseudomonadaceae, for which the name Oblitimonas alkaliphila gen. nov., sp. nov. is proposed. The type strain is B4199T (=DSM 100830T = CCUG 67636T). |
Lawsonella clevelandensis gen. nov., sp. nov., a new member of the suborder Corynebacterineae isolated from human abscesses
Bell ME , Bernard KA , Harrington SM , Patel NB , Tucker TA , Metcalfe MG , McQuiston JR . Int J Syst Evol Microbiol 2016 66 (8) 2929-2935 Gram-positive, partially acid-fast, non-spore forming, anaerobic, catalase-positive, pleomorphic bacteria were isolated from human abscesses. Strains X1036T, X1698, and NML 120705, were recovered from a spinal abscess, a peritoneal abscess, and a breast abscess respectively. A phylogenetic analysis of the 16S rRNA gene sequences showed that the strains shared 100% similarity and the nearest phylogenetic neighbor is Dietzia timorensis DSM 45568T (95%). Chemotaxonomic characteristics of the strains were consistent with those described for members of the suborder Corynebacterineae. Mycolic acids were detected using HPLC and one-dimensional TLC; whole cell hydrolysates yielded meso-DAP with arabinose and galactose as the predominant sugars; the muramic acid acyl type was acetylated; the major menaquinone was MK-9 (96.3%); polar lipids detected were phosphatidylglycerol, phosphatidylinositol, and an unknown glycophospholipid. Cellular fatty acids were hexadecanoic acid (C16:0), octadecanoic acid (C18:1omega9c), and decanoic acid (C10:0). Tuberculostearic acid was not detected. Based on the results of this polyphasic study, we conclude that these strains represent a novel genus and species within the suborder Corynebacterineae for which we propose the name Lawsonella clevelandensis gen. nov., sp. nov., with the type strain X1036T (=DSM 45743T = CCUG 66657T). |
Aggregation of adenovirus 2 in source water and impacts on disinfection by chlorine
Kahler AM , Cromeans TL , Metcalfe MG , Humphrey CD , Hill VR . Food Environ Virol 2016 8 (2) 148-55 It is generally accepted that viral particles in source water are likely to be found as aggregates attached to other particles. For this reason, it is important to investigate the disinfection efficacy of chlorine on aggregated viruses. A method to produce adenovirus particle aggregation was developed for this study. Negative stain electron microscopy was used to measure aggregation before and after addition of virus particles to surface water at different pH and specific conductance levels. The impact of aggregation on the efficacy of chlorine disinfection was also examined. Disinfection experiments with human adenovirus 2 (HAdV2) in source water were conducted using 0.2 mg/L free chlorine at 5 degrees C. Aggregation of HAdV2 in source water (≥3 aggregated particles) remained higher at higher specific conductance and pH levels. However, aggregation was highly variable, with the percentage of particles present in aggregates ranging from 43 to 71 %. Upon addition into source water, the aggregation percentage dropped dramatically. On average, chlorination CT values (chlorine concentration in mg/L x time in min) for 3-log10 inactivation of aggregated HAdV2 were up to three times higher than those for dispersed HAdV2, indicating that aggregation reduced the disinfection rate. This information can be used by water utilities and regulators to guide decision making regarding disinfection of viruses in water. |
Clinicopathologic, immunohistochemical, and ultrastructural findings of a fatal case of Middle East respiratory syndrome coronavirus infection in United Arab Emirates, April 2014
Ng DL , Al Hosani F , Keating MK , Gerber SI , Jones TL , Metcalfe MG , Tong S , Tao Y , Alami NN , Haynes LM , Mutei MA , Abdel-Wareth L , Uyeki TM , Swerdlow DL , Barakat M , Zaki SR . Am J Pathol 2016 186 (3) 652-8 Middle East respiratory syndrome coronavirus (MERS-CoV) infection causes an acute respiratory illness and is associated with a high case fatality rate; however, the pathogenesis of severe and fatal MERS-CoV infection is unknown. We describe the histopathologic, immunohistochemical, and ultrastructural findings from the first autopsy performed on a fatal case of MERS-CoV in the world, which was related to a hospital outbreak in the United Arab Emirates in April 2014. The main histopathologic finding in the lungs was diffuse alveolar damage. Evidence of chronic disease, including severe peripheral vascular disease, patchy cardiac fibrosis, and hepatic steatosis, was noted in the other organs. Double staining immunoassays that used anti-MERS-CoV antibodies paired with immunohistochemistry for cytokeratin and surfactant identified pneumocytes and epithelial syncytial cells as important targets of MERS-CoV antigen; double immunostaining with dipeptidyl peptidase 4 showed colocalization in scattered pneumocytes and syncytial cells. No evidence of extrapulmonary MERS-CoV antigens were detected, including the kidney. These results provide critical insights into the pathogenesis of MERS-CoV in humans. |
Haematospirillum jordaniae gen. nov., sp. nov., isolated from human blood samples.
Humrighouse BW , Emery BD , Kelly AJ , Metcalfe MG , Mbizo J , McQuiston JR . Antonie Van Leeuwenhoek 2016 109 (4) 493-500 A Gram-negative, aerobic, motile, spiral-shaped bacterium, strain H5569T, was isolated from a human blood sample. Phenotypic and molecular characteristics of the isolate were investigated. Optimal growth was found to occur at 35 degrees C under aerobic conditions on Heart Infusion Agar supplemented with 5 % rabbit blood. The major fatty acids present in the cells were identified as C16:0, C16:1omega7c and C18:1omega7c. The predominant respiratory quinone was found to be ubiquinone-Q10. The G+C content of genomic DNA for strain H5569T was found to be 49.9 %. Based on 16S rRNA gene sequence analysis results, 13 additional isolates were also analysed in this study. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the organism, represented by strain H5569T, forms a distinct lineage within the family Rhodospirillaceae, closely related to two Novispirillum itersonii subspecies (93.9-94.1 %) and two Caenispirillum sp. (91.2-91.6 %). Based on these results, the isolate H5569T is concluded to represent a new genus and species for which the name Haematospirillum jordaniae gen. nov., sp. nov. is proposed. The type strain is H5569T (=DSMT 28903 = CCUG 66838T). |
Assessment of Inhibitors of Pathogenic Crimean-Congo Hemorrhagic Fever Virus Strains Using Virus-Like Particles.
Zivcec M , Metcalfe MG , Albarino CG , Guerrero LW , Pegan SD , Spiropoulou CF , Bergeron E . PLoS Negl Trop Dis 2015 9 (12) e0004259 Crimean-Congo hemorrhagic fever (CCHF) is an often lethal, acute inflammatory illness that affects a large geographic area. The disease is caused by infection with CCHF virus (CCHFV), a nairovirus from the Bunyaviridae family. Basic research on CCHFV has been severely hampered by biosafety requirements and lack of available strains and molecular tools. We report the development of a CCHF transcription- and entry-competent virus-like particle (tecVLP) system that can be used to study cell entry and viral transcription/replication over a broad dynamic range (~4 orders of magnitude). The tecVLPs are morphologically similar to authentic CCHFV. Incubation of immortalized and primary human cells with tecVLPs results in a strong reporter signal that is sensitive to treatment with neutralizing monoclonal antibodies and by small molecule inhibitors of CCHFV. We used glycoproteins and minigenomes from divergent CCHFV strains to generate tecVLPs, and in doing so, we identified a monoclonal antibody that can prevent cell entry of tecVLPs containing glycoproteins from 3 pathogenic CCHFV strains. In addition, our data suggest that different glycoprotein moieties confer different cellular entry efficiencies, and that glycoproteins from the commonly used strain IbAr10200 have up to 100-fold lower ability to enter primary human cells compared to glycoproteins from pathogenic CCHFV strains. |
Malignant Transformation of Hymenolepis nana in a Human Host.
Muehlenbachs A , Bhatnagar J , Agudelo CA , Hidron A , Eberhard ML , Mathison BA , Frace MA , Ito A , Metcalfe MG , Rollin DC , Visvesvara GS , Pham CD , Jones TL , Greer PW , Velez Hoyos A , Olson PD , Diazgranados LR , Zaki SR . N Engl J Med 2015 373 (19) 1845-52 Neoplasms occur naturally in invertebrates but are not known to develop in tapeworms. We observed nests of monomorphic, undifferentiated cells in samples from lymph-node and lung biopsies in a man infected with the human immunodeficiency virus (HIV). The morphologic features and invasive behavior of the cells were characteristic of cancer, but their small size suggested a nonhuman origin. A polymerase-chain-reaction (PCR) assay targeting eukaryotes identified Hymenolepis nana DNA. Although the cells were unrecognizable as tapeworm tissue, immunohistochemical staining and probe hybridization labeled the cells in situ. Comparative deep sequencing identified H. nana structural genomic variants that are compatible with mutations described in cancer. Invasion of human tissue by abnormal, proliferating, genetically altered tapeworm cells is a novel disease mechanism that links infection and cancer. |
Identification of Influenza A/PR/8/34 Donor Viruses Imparting High Hemagglutinin Yields to Candidate Vaccine Viruses in Eggs.
Johnson A , Chen LM , Winne E , Santana W , Metcalfe MG , Mateu-Petit G , Ridenour C , Hossain MJ , Villanueva J , Zaki SR , Williams TL , Cox NJ , Barr JR , Donis RO . PLoS One 2015 10 (6) e0128982 One of the important lessons learned from the 2009 H1N1 pandemic is that a high yield influenza vaccine virus is essential for efficient and timely production of pandemic vaccines in eggs. The current seasonal and pre-pandemic vaccine viruses are generated either by classical reassortment or reverse genetics. Both approaches utilize a high growth virus, generally A/Puerto Rico/8/1934 (PR8), as the donor of all or most of the internal genes, and the wild type virus recommended for inclusion in the vaccine to contribute the hemagglutinin (HA) and neuraminidase (NA) genes encoding the surface glycoproteins. As a result of extensive adaptation through sequential egg passaging, PR8 viruses with different gene sequences and high growth properties have been selected at different laboratories in past decades. The effect of these related but distinct internal PR8 genes on the growth of vaccine viruses in eggs has not been examined previously. Here, we use reverse genetics to analyze systematically the growth and HA antigen yield of reassortant viruses with 3 different PR8 backbones. A panel of 9 different HA/NA gene pairs in combination with each of the 3 different lineages of PR8 internal genes (27 reassortant viruses) was generated to evaluate their performance. Virus and HA yield assays showed that the PR8 internal genes influence HA yields in most subtypes. Although no single PR8 internal gene set outperformed the others in all candidate vaccine viruses, a combination of specific PR8 backbone with individual HA/NA pairs demonstrated improved HA yield and consequently the speed of vaccine production. These findings may be important both for production of seasonal vaccines and for a rapid global vaccine response during a pandemic. |
Identification of Giardia duodenalis and Enterocytozoon bieneusi in an epizoological investigation of a laboratory colony of prairie dogs, Cynomys ludovicianus.
Roellig DM , Salzer JS , Carroll DS , Ritter JM , Drew C , Gallardo-Romero N , Keckler MS , Langham G , Hutson CL , Karem KL , Gillespie TR , Visvesvara GS , Metcalfe MG , Damon IK , Xiao L . Vet Parasitol 2015 210 91-7 Since 2005, black-tailed prairie dogs (Cynomys ludovicianus) have been collected for use as research animals from field sites in Kansas, Colorado, and Texas. In January of 2012, Giardia trophozoites were identified by histology, thin-section electron microscopy, and immunofluorescent staining in the lumen of the small intestine and colon of a prairie dog euthanized because of extreme weight loss. With giardiasis suspected as the cause of weight loss, a survey of Giardia duodenalis in the laboratory colony of prairie dogs was initiated. Direct immunofluorescent testing of feces revealed active shedding of Giardia cysts in 40% (n=60) of animals held in the vivarium. All tested fecal samples (n=29) from animals in another holding facility where the index case originated were PCR positive for G. duodenalis with assemblages A and B identified from sequencing triosephosphate isomerase (tpi), glutamate dehydrogenase (gdh), and beta-giardin (bg) genes. Both assemblages are considered zoonotic, thus the parasites in prairie dogs are potential human pathogens and indicate prairie dogs as a possible wildlife reservoir or the victims of pathogen spill-over. Molecular testing for other protozoan gastrointestinal parasites revealed no Cryptosporidium infections but identified a host-adapted Enterocytozoon bieneusi genotype group. |
Human infection with a zoonotic orthopoxvirus in the country of Georgia.
Vora NM , Li Y , Geleishvili M , Emerson GL , Khmaladze E , Maghlakelidze G , Navdarashvili A , Zakhashvili K , Kokhreidze M , Endeladze M , Mokverashvili G , Satheshkumar PS , Gallardo-Romero N , Goldsmith CS , Metcalfe MG , Damon I , Maes EF , Reynolds MG , Morgan J , Carroll DS . N Engl J Med 2015 372 (13) 1223-30 During 2013, cutaneous lesions developed in two men in the country of Georgia after they were exposed to ill cows. The men had never received vaccination against smallpox. Tests of lesion material with the use of a quantitative real-time polymerase-chain-reaction assay for non-variola virus orthopoxviruses were positive, and DNA sequence analysis implicated a novel orthopoxvirus species. During the ensuing epidemiologic investigation, no additional human cases were identified. However, serologic evidence of exposure to an orthopoxvirus was detected in cows in the patients' herd and in captured rodents and shrews. A third case of human infection that occurred in 2010 was diagnosed retrospectively during testing of archived specimens that were originally submitted for tests to detect anthrax. Orthopoxvirus infection should be considered in persons in whom cutaneous lesions develop after contact with animals. |
Novel poxvirus infection in two patients from the United States
Osadebe LU , Manthiram K , McCollum AM , Li Y , Emerson GL , Gallardo-Romero NF , Doty JB , Wilkins K , Zhao H , Drew CP , Metcalfe MG , Goldsmith CS , Muehlenbachs A , Googe P , Dunn J , Duenckel T , Henderson H , Carroll DS , Zaki SR , Denison M , Reynolds MG , Damon IK . Clin Infect Dis 2014 60 (2) 195-202 BACKGROUND: Some human poxvirus infections can be acquired through zoonotic transmission. We report a previously unknown poxvirus infection in two patients, one of whom was immunocompromised, and both patients had known equine contact. METHODS: The patients were interviewed and clinical information was abstracted from the patients' medical files. Biopsies of the skin lesions were collected from both patients for histopathology, immunohistochemistry, and transmission electron microscopy analysis. Oral and skin swabs were collected from animals with frequent contact with the patients and environmental sampling including rodent trapping was performed on the farm where the immunosuppressed patient was employed. 'Pan-pox and high GC' PCR assays were performed on patient, animal, and environmental isolates. Amplicon sequences of the viral DNA were used for agent identification and phylogenetic analysis. RESULTS: Specimens from both human cases revealed a novel poxvirus. The agent shares 88% similarity to viruses in the Parapoxvirus genus and 78% to those in the Molluscipoxvirus genus but is sufficiently divergent to resist classification as either. All animal and environmental specimens were negative for poxvirus and both patients had complete resolution of lesions. CONCLUSION: This report serves as a reminder that poxviruses should be considered in cutaneous human infections especially in individuals with known barnyard exposures. The clinical course of the patients was similar to that of parapoxvirus infections and the source of this virus is currently unknown but is presumed to be zoonotic. This report also demonstrates the importance of a comprehensive approach to diagnosis of human infections caused by previously unknown pathogens. |
Novel paramyxovirus associated with severe acute febrile disease, South Sudan and Uganda, 2012
Albarino CG , Foltzer M , Towner JS , Rowe LA , Campbell S , Jaramillo CM , Bird BH , Reeder DM , Vodzak ME , Rota P , Metcalfe MG , Spiropoulou CF , Knust B , Vincent JP , Frace MA , Nichol ST , Rollin PE , Stroher U . Emerg Infect Dis 2014 20 (2) 211-6 In 2012, a female wildlife biologist experienced fever, malaise, headache, generalized myalgia and arthralgia, neck stiffness, and a sore throat shortly after returning to the United States from a 6-week field expedition to South Sudan and Uganda. She was hospitalized, after which a maculopapular rash developed and became confluent. When the patient was discharged from the hospital on day 14, arthralgia and myalgia had improved, oropharynx ulcerations had healed, the rash had resolved without desquamation, and blood counts and hepatic enzyme levels were returning to reference levels. After several known suspect pathogens were ruled out as the cause of her illness, deep sequencing and metagenomics analysis revealed a novel paramyxovirus related to rubula-like viruses isolated from fruit bats. |
Alteration of the phenotypic and pathogenic patterns of Burkholderia pseudomallei that persist in a soil environment
Chen YS , Shieh WJ , Goldsmith CS , Metcalfe MG , Greer P , Zaki SR , Chang HH , Chan H , Chen YL . Am J Trop Med Hyg 2014 90 (3) 469-79 Melioidosis is caused by the soil-borne pathogen Burkholderia pseudomallei. To investigate whether the distinct phenotypic and virulent characteristics result from environmental adaptations in the soil or from the host body, two pairs of isogenic strains were generated by passages in soil or mice. After cultivation in soil, the levels of 3-hydroxytetradecanoic acid, biofilm formation, flagellar expression, and ultrastructure were altered in the bacteria. Uniformly fatal melioidosis developed as a result of infection with mouse-derived strains; however, the survival rates of mice infected with soil-derived strains prolonged. After primary infection or reinfection with soil-derived strains, the mice developed a low degree of bacterial hepatitis and bacterial colonization in the liver and bone marrow compared with mice that were infected with isogenic or heterogenic mouse-derived strains. We suggest that specific phenotypic and pathogenic patterns can be induced through infection with B. pseudomallei that has been cultured in different (soil versus mouse) environments. |
Novel poxvirus in big brown bats, northwestern United States
Emerson GL , Nordhausen R , Garner MM , Huckabee JR , Johnson S , Wohrle RD , Davidson WB , Wilkins K , Li Y , Doty JB , Gallardo-Romero NF , Metcalfe MG , Karem KL , Damon IK , Carroll DS . Emerg Infect Dis 2013 19 (6) 1002-4 A wildlife hospital and rehabilitation center in northwestern United States received several big brown bats with necrosuppurative osteomyelitis in multiple joints. Wing and joint tissues were positive by PCR for poxvirus. Thin-section electron microscopy showed poxvirus particles within A-type inclusions. Phylogenetic comparison supports establishment of a new genus of Poxviridae. |
Exserohilum infections associated with contaminated steroid injections: a clinicopathologic review of 40 cases
Ritter JM , Muehlenbachs A , Blau DM , Paddock CD , Shieh WJ , Drew CP , Batten BC , Bartlett JH , Metcalfe MG , Pham CD , Lockhart SR , Patel M , Liu L , Jones TL , Greer PW , Montague JL , White E , Rollin DC , Seales C , Stewart D , Deming MV , Brandt ME , Zaki SR . Am J Pathol 2013 183 (3) 881-92 September 2012 marked the beginning of the largest reported outbreak of infections associated with epidural and intra-articular injections. Contamination of methylprednisolone acetate with the black mold, Exserohilum rostratum, was the primary cause of the outbreak, with >13,000 persons exposed to the potentially contaminated drug, 741 confirmed drug-related infections, and 55 deaths. Fatal meningitis and localized epidural, paraspinal, and peripheral joint infections occurred. Tissues from 40 laboratory-confirmed cases representing these various clinical entities were evaluated by histopathological analysis, special stains, and IHC to characterize the pathological features and investigate the pathogenesis of infection, and to evaluate methods for detection of Exserohilum in formalin-fixed, paraffin-embedded (FFPE) tissues. Fatal cases had necrosuppurative to granulomatous meningitis and vasculitis, with thrombi and abundant angioinvasive fungi, with extensive involvement of the basilar arterial circulation of the brain. IHC was a highly sensitive method for detection of fungus in FFPE tissues, demonstrating both hyphal forms and granular fungal antigens, and PCR identified Exserohilum in FFPE and fresh tissues. Our findings suggest a pathogenesis for meningitis involving fungal penetration into the cerebrospinal fluid at the injection site, with transport through cerebrospinal fluid to the basal cisterns and subsequent invasion of the basilar arteries. Further studies are needed to characterize Exserohilum and investigate the potential effects of underlying host factors and steroid administration on the pathogenesis of infection. |
The pox in the North American backyard: Volepox virus pathogenesis in California mice (Peromyscus californicus)
Gallardo-Romero NF , Drew CP , Weiss SL , Metcalfe MG , Nakazawa YJ , Smith SK , Emerson GL , Hutson CL , Salzer JS , Bartlett JH , Olson VA , Clemmons CJ , Davidson WB , Zaki SR , Karem KL , Damon IK , Carroll DS . PLoS One 2012 7 (8) e43881 Volepox virus (VPXV) was first isolated in 1985 from a hind foot scab of an otherwise healthy California vole (Microtus californicus). Subsequent surveys in San Mateo County, CA, revealed serological evidence suggesting that VPXV is endemic to this area, and a second viral isolate from a Pinyon mouse (Peromyscus truei) was collected in 1988. Since then, few studies have been conducted regarding the ecology, pathology, and pathogenicity of VPXV, and its prevalence and role as a potential zoonotic agent remain unknown. To increase our understanding of VPXV disease progression, we challenged 24 California mice (Peromyscus californicus) intranasally with 1.6x10(3) PFU of purified VPXV. By day five post infection (pi) we observed decreased activity level, conjunctivitis, ruffled hair, skin lesions, facial edema, and crusty noses. A mortality rate of 54% was noted by day eight pi. In addition, internal organ necrosis and hemorrhages were observed during necropsy of deceased or euthanized animals. Viral loads in tissues (brain, gonad, kidney, liver, lung, spleen, submandibular lymph node, and adrenal gland), bodily secretions (saliva, and tears), and excretions (urine, and/or feces) were evaluated and compared using real time-PCR and tissue culture. Viral loads measured as high as 2x10(9) PFU/mL in some organs. Our results suggest that VPXV can cause extreme morbidity and mortality within rodent populations sympatric with the known VPXV reservoirs. |
A new phlebovirus associated with severe febrile illness in Missouri.
McMullan LK , Folk SM , Kelly AJ , MacNeil A , Goldsmith CS , Metcalfe MG , Batten BC , Albarino CG , Zaki SR , Rollin PE , Nicholson WL , Nichol ST . N Engl J Med 2012 367 (9) 834-41 Two men from northwestern Missouri independently presented to a medical facility with fever, fatigue, diarrhea, thrombocytopenia, and leukopenia, and both had been bitten by ticks 5 to 7 days before the onset of illness. Ehrlichia chaffeensis was suspected as the causal agent but was not found on serologic analysis, polymerase-chain-reaction (PCR) assay, or cell culture. Electron microscopy revealed viruses consistent with members of the Bunyaviridae family. Next-generation sequencing and phylogenetic analysis identified the viruses as novel members of the phlebovirus genus. Although Koch's postulates have not been completely fulfilled, we believe that this phlebovirus, which is novel in the Americas, is the cause of this clinical syndrome. |
Single-dose immunization with virus replicon particles confers rapid robust protection against Rift Valley fever virus challenge
Dodd KA , Bird BH , Metcalfe MG , Nichol ST , Albarino CG . J Virol 2012 86 (8) 4204-12 Rift Valley fever virus (RVFV) causes outbreaks of severe disease in people and livestock throughout Africa and the Arabian Peninsula. The potential for RVFV introduction outside the area of endemicity highlights the need for fast-acting, safe, and efficacious vaccines. Here, we demonstrate a robust system for the reverse genetics generation of a RVF virus replicon particle (VRP(RVF)) vaccine candidate. Using a mouse model, we show that VRP(RVF) immunization provides the optimal balance of safety and single-dose robust efficacy. VRP(RVF) can actively synthesize viral RNA and proteins but lacks structural glycoprotein genes, preventing spread within immunized individuals and reducing the risk of vaccine-induced pathogenicity. VRP(RVF) proved to be completely safe following intracranial inoculation of suckling mice, a stringent test of vaccine safety. Single-dose subcutaneous immunization with VRP(RVF), although it is highly attenuated, completely protected mice against a virulent RVFV challenge dose which was 100,000-fold greater than the 50% lethal dose (LD(50)). Robust protection from lethal challenge was observed by 24 h postvaccination, with 100% protection induced in as little as 96 h. We show that a single subcutaneous VRP(RVF) immunization initiated a systemic antiviral state followed by an enhanced adaptive response. These data contrast sharply with the much-reduced survivability and immune responses observed among animals immunized with nonreplicating viral particles, indicating that replication, even if confined to the initially infected cells, contributes substantially to protective efficacy at early and late time points postimmunization. These data demonstrate that replicon vaccines successfully bridge the gap between safety and efficacy and provide insights into the kinetics of antiviral protection from RVFV infection. |
Involvement of the autophagy pathway in trafficking of Mycobacterium tuberculosis bacilli through cultured human type II epithelial cells
Fine KL , Metcalfe MG , White E , Virji M , Karls RK , Quinn FD . Cell Microbiol 2012 14 (9) 1402-14 Interactions between Mycobacterium tuberculosis bacilli and alveolar macrophages have been extensively characterized, while similar analyses in epithelial cells have not been performed. In this study, we microscopically examined endosomal trafficking of M. tuberculosis strain Erdman in A549 cells, a human type II pneumocyte cell line. Immuno-electron microscopic (IEM) analyses indicate that M. tuberculosis bacilli are internalized to a compartment labeled first with Rab5 and then with Rab7 small GTPase proteins. This suggests that, unlike macrophages, M. tuberculosis bacilli traffic to late endosomes in epithelial cells. However, fusion of lysosomes with the bacteria-containing compartment appears to be inhibited, as illustrated by IEM studies employing LAMP-2 and Cathepsin-L antibodies. Examination by transmission electron microscopy and IEM revealed M. tuberculosis-containing compartments surrounded by double-membranes and labeled with antibodies against the autophagy marker Lc3, providing evidence for involvement and intersection of the autophagy and endosomal pathways. Interestingly, inhibition of the autophagy pathway using 3-methyladenine improved host cell viability and decreased numbers of viable intracellular bacteria recovered after 72 hours post infection. Collectively, these data suggest that trafficking patterns for M. tuberculosis bacilli in alveolar epithelial cells differ from macrophages, and that autophagy is involved this process. (c) 2012 Blackwell Publishing Ltd. |
Ultrastructural characterization of pandemic (H1N1) 2009 virus
Goldsmith CS , Metcalfe MG , Rollin DC , Shieh WJ , Paddock CD , Xu XY , Zaki SR . Emerg Infect Dis 2011 17 (11) 2056-2059 We evaluated pandemic influenza A (H1N1) 2009 virus isolates and respiratory tissues collected at autopsy by electron microscopy. Many morphologic characteristics were similar to those previously described for influenza virus. One of the distinctive features was dense tubular structures in the nuclei of infected cells. |
Tubulinosema sp. microsporidian myositis in immunosuppressed patient
Choudhary MM , Metcalfe MG , Arrambide K , Bern C , Visvesvara GS , Pieniazek NJ , Bandea RD , Deleon-Carnes M , Adem P , Zaki SR , Saeed MU . Emerg Infect Dis 2011 17 (9) 1727-30 The Phylum Microsporidia comprises >1,200 species, only 15 of which are known to infect humans, including the genera Trachipleistophora, Pleistophora, and Brachiola. We report an infection by Tubulinosema sp. in an immunosuppressed patient. |
Reduced virus replication, proinflammatory cytokine production, and delayed macrophage cell death in human PBMCs infected with the newly discovered Bundibugyo ebolavirus relative to Zaire ebolavirus
Gupta M , Goldsmith CS , Metcalfe MG , Spipopoulou CF , Rollin PE . Virology 2010 402 (1) 203-8 Bundibugyo ebolavirus is a newly identified Ebolavirus species. The virus was responsible for a recent hemorrhagic fever outbreak in Uganda with an approximate 30% case fatality rate. In this study, we compared the pathogenesis of Bundibugyo with highly lethal Zaire Ebolavirus by using in vitro human PBMCs. We found that PBMCs infected with Bundibugyo ebolaviruses resulted in 1 to 2 log lower virus yields compared to Zaire ebolavirus and produced 2- to 10-fold lower levels of TNF-alpha, MCP-1, IL-1beta, MIP1-alpha and IL-10 than PBMCs infected with Zaire ebolavirus. In addition, flow cytometric studies have shown lower levels and delay of the macrophage cell death in Bundibugyo ebolavirus compared to Zaire ebolavirus infection. The findings of slower Bundibugyo ebolavirus replication, lower production of proinflammatory cytokines and delay in macrophage cell death provide insight into the basis of the lower case fatality observed with Bundibugyo ebolavirus. |
Isolation of Rickettsia parkeri and identification of a novel spotted fever group Rickettsia sp. from gulf coast ticks (Amblyomma maculatum) in the United States
Paddock CD , Fournier PE , Sumner JW , Goddard J , Elshenawy Y , Metcalfe MG , Loftis AD , Varela-Stokes A . Appl Environ Microbiol 2010 76 (9) 2689-96 Until recently, Amblyomma maculatum, (the Gulf Coast tick), has garnered little attention relative to other species of human-biting ticks in the United States. A. maculatum is now recognized as the principal vector of Rickettsia parkeri, a pathogenic spotted fever group rickettsia (SFGR) that causes an eschar-associated illness in humans that resembles Rocky Mountain spotted fever. A novel SFGR, distinct from other recognized Rickettsia spp., has also been detected recently in A. maculatum specimens collected from several regions of the southeastern United States. In this study, 198 questing adult Gulf Coast ticks were collected from 4 locations in Florida and Mississippi; 28% were infected with R. parkeri and 3% with a novel SFGR. Seventeen isolates of R. parkeri were cultivated in Vero E6 cells from individual specimens of A. maculatum; however, all attempts to isolate the novel SFGR were unsuccessful. Partial genetic characterization of the novel SFGR revealed identity with several recently described, incompletely characterized, and non-cultivated SFGR including Candidatus 'Rickettsia andeanae' and Rickettsia sp. 'Argentina', detected in several species of Neotropical ticks from Argentina and Peru. These findings suggest that each of these 'novel' rickettsiae represent the same species. Our study expands considerably the number of low-passage, A. maculatum-derived isolates of R. parkeri, and characterizes a second and sympatrically distributed Rickettsia sp. found in Gulf Coast ticks. |
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