Hemorrhagic fever viruses are a major public health threat in Sub-Saharan Africa. These kinds of viruses cause symptoms ranging from non-specific fevers and body aches to severe, life-threatening bleeding, shock, and multi-organ failure. Previously discovered hemorrhagic fever viruses can cause recurrent or seasonal outbreaks, but new ones continue to emerge. In order to combat these viruses, we need to better understand the aspects of pathogenesis that lead to mortality or survival. I will present analysis of the host immune response to two hemorrhagic fever viruses, Lassa virus and Bundibugyo virus, and how the host response can be used to predict mortality in these diseases. Lassa virus (LASV) was identified over 50 years ago, but it remains understudied and has hence been denoted a “Neglected Tropical Disease”. Clinical studies and experiments were run by our collaborators in Nigeria and Germany. In all, longitudinal blood samples were collected for over two hundred Nigerian Lassa Fever patients and concentrations of over 60 proteins analyzed. I processed the datasets, performed statistical testing, and created logistic regression models for each protein. This modeling allowed me to determine which proteins could be used as a predictive biomarker of mortality and the level of that protein that could best stratify patients who died and survived. I then compared protein levels for the best biomarkers and other markers in the same biological pathways with those of healthy and other febrile illness (non-Lassa Fever) controls. I examined the best biomarkers over time for their utility as biomarkers at later timepoints in hospitalization. Finally, I produced an application using RShiny that incorporated these and other exploratory analyses of the data, which allows users to visualize all the data we had in addition to the plots that were published. The filovirus Bundibugyo ebolavirus (BDBV), a relative of the more well-known Ebola virus (EBOV), first caused an outbreak in people fifteen years ago. Animal models are still being developed and characterized for this virus. Our collaborators in Texas experimentally infected cynomolgus macaques with BDBV and gave them post-exposure treatment with a VSV-based vaccine. These collaborators performed RNA-Seq on longitudinal samples from the infected macaques and sent me these data for analysis. I wrote pipelines to perform RNA-Seq and differential expression analyses on over 600 samples, of which I will focus on a subset here. I found differentially expressed genes for different subsets of the data, and I examined these gene lists using gene set enrichment analysis. I then generated logistic regression models to find differentially expressed genes that could predict mortality or survival. Many of these genes could accurately predict outcome at either late or early timepoints. I then used the top genes found by logistic regression to generate random forest models that could predict mortality over the entire course of disease. / 2025-03-20T00:00:00Z

Rabbit Passage of the Virus The hog cholera virus was carried through six alternate passages from pig to rabbit and then top five serial passages in rabbits. The virus could not be demonstrated in rabbit-spleen suspension prepared for the sixth serial passage in rabbits. The lack of pathogenic or antigenic properties for the pig made it impossible to detect the virus, assuming that it was present in the rabbit-spleen suspension, at the sixth serial passage. The virus did not become pathogenic for the rabbit during the alternate and serial passages, although it survived for at least 72 hours in the rabbit. No evidence or attenuation of the virus for the pig was obtained in either the alternate or serial passages. Ultraviolet Irradiation or the Virus The hog cholera virus in the form of a pig-spleen suspension was irradiated by the Westinghouse lamps CH4 and Wl793. The virus was irradiated at a distance of 15 centimeters from the lamp. The suspension was placed in an open petri dish, to a depth or l to 2 millimeters, and was constantly agitated during the irradiation. Irradiation by the CH4 lamp for 30 and 60 minutes and irradiation by the WL793 lamp for 15, 30, and 60 minutes failed to attenuate the virus for the pig. Typical hog cholera was produced in pigs injected with the irradiated virus. Nitrogen Mustard Treatment of the Virus The hog cholera virus in the form of blood virus was mixed with the nitrogen mustard, methyl-bis (β-chloroethyl)-amine, in the proportion or five milliliters of blood virus to ten milligrams of nitrogen mustard. In one lot of blood virus so treated, the virus was completely destroyed, whereas in a second lot the virus was not affected. Treatment of five milliliters of the virus, in the form of pig-spleen suspension, with ten milligrams of the nitrogen mustard completely attenuated the virus for the pig, so that, when the virus was injected into pigs, no ill effects appeared. Pigs injected with five milliliters of the mustard-treated pig-spleen virus developed a resistance to the virulent virus. The injected pigs successfully withstood the intramuscular injection or two milliliters of virulent virus, or pen exposure to cholera-infected pigs. The mustard-treated pig-spleen virus produced a satisfactory immunity in pigs injected with the treated virus 24 hours after its preparation. The mustard-treated pig-spleen virus did not spread from injected pigs to non-injected pigs kept in close contact with them. / Ph. D.

M.S.

M.S.

Valley fever, or coccidioidomycosis, is a disease caused by the Coccidioides immitis and Coccidioides posadasii fungal species that dwell in the soil but can become airborne and infect a human or mammalian host through their respiratory tract. Disease rates in the western U.S. have significantly increased over the past two decades, creating an emerging public health burden. Studies have been conducted that attempt to elucidate the association between environmental conditions and the growth and dispersal of the pathogen, yet the specific ecology of and environmental precursors to the disease remain uncertain. This research project investigates the relationship between environmental variables and valley fever by modeling the spatial and temporal dynamics of the disease using varying techniques. Chapter 1 discusses relevant literature before discussing the challenges associated with studying valley fever. Chapter 2 analyzes the temporal relationships between valley fever and climatic variables, focusing on Kern County, California, an understudied region in the U.S. where valley fever is highly endemic. Chapter 3 focuses on a regional spatial analysis using ecological niche modeling to better understand the environmental factors that influence the overall spatial distribution of valley fever in the U.S. Finally, combining both spatial and temporal components, Chapter 4 uses a hierarchical Bayesian spatio-temporal model to investigate the patterns and drivers of this disease, focusing on state of California, which saw an approximate 200% increase in cases from 2014 to 2018. Cumulatively, this work offers new insights on relationships between climate, landcover, and valley fever disease risk. Significant findings include climate variables explaining up to 76% of valley fever variability in Kern County, California, the significance of both climatic and landcover variables in characterizing the geographic distribution of the disease, and identification of patterns increasing risk in geographic regions of California not currently considered highly endemic. These findings advance scholarly understandings of valley fever's environmental disease drivers. The results of this research can be applied by public health officials in the allocation of surveillance and public education resources, focusing upon regions that are most likely to encounter the illness. / Doctor of Philosophy / Valley fever is a fungal disease that causes illness in over ten thousand people in the western U.S. every year. Disease rates have been increasing for the past two decades for unknown reasons, although previous research suggests that climatic variations are likely contributing factors. This research evaluated environmental factors with hypothesized relationships to valley fever disease rates. First, this dissertation explored time-series relationships between climatic factors and valley fever incidence in an understudied county in California. Research findings identified that climatic factors including precipitation from previous seasons and temperature were significantly associated with valley fever incidence in this county. Second, this dissertation assessed where valley fever is found in the western U.S. The likely spatial distribution for the disease was mapped and environmental variables influential to this distribution were identified; they included both climate and landcover variables. Finally, a model was developed to analyze patterns of disease risk in California that considered both space and time, and environmental risk factors potentially contributing to the observed patterns were assessed. Counties with increased risk were identified and significant environmental relationships with valley fever risk were confirmed. The results of this research can be applied by public health officials in allocating surveillance and public education resources, focusing upon regions that are most likely to encounter the illness.

Introdução: A febre sem sinais localizatórios é definida como presença de febre de até 7 dias de duração, sem identificação da causa após anamnese e exame físico detalhados. A maioria destas crianças apresenta doença infecciosa aguda autolimitada ou está em fase prodrômica de uma doença infecciosa benigna. Poucas têm infecção bacteriana grave: bacteremia oculta, pneumonia oculta, infecção urinária, meningite bacteriana, artrite séptica, osteomielite ou celulite. Embora a febre seja uma das queixas mais comuns nos serviços de emergência, a abordagem da criança febril permanece controversa. Objetivos: avaliar a aplicabilidade de um protocolo padronizado para o atendimento e seguimento das crianças até 36 meses de idade com febre sem sinais localizatórios e analisar os fatores de risco para infecção bacteriana grave nestas crianças. Métodos: estudo prospectivo em crianças até 36 meses de idade que procuraram o pronto socorro do Hospital Universitário da Universidade de São Paulo, com quadro de febre sem sinais localizatórios, durante o período de um ano (junho/2006 a maio/2007). As crianças foram atendidas conforme protocolo que estratifica o risco de infecção bacteriana grave de acordo com a presença ou não de toxemia, idade e valor da temperatura. Conforme a avaliação de risco, indicava-se triagem laboratorial (hemograma, hemocultura, sedimento urinário, urocultura e, se necessário, radiografia torácica, liquor e coprocultura). Os fatores de risco para infecção bacteriana grave estudados foram: sexo, idade, presença de toxemia, temperatura, número total de leucócitos, número total de neutrófilos e número total de neutrófilos jovens. Resultados: Foram seguidas 215 crianças, sendo 111 (51,6%) do sexo feminino. A média de idade foi de 11,85 meses (DP ± 8,91). Vinte crianças, na avaliação inicial, apresentavam algum grau de toxemia, e 195 estavam em bom estado geral. Nas crianças de 3 a 36 meses não toxêmicas, 95 tinham temperatura axilar >39ºC. Em 107 crianças (49,8%), houve melhora espontânea do quadro febril; em 88 (40,9%), foi identificada doença benigna autolimitada; e em 20 (9,3%), infecção bacteriana grave. Dentre as infecções bacterianas graves, identificou-se 16 infecções urinárias, três pneumonias e uma bacteremia oculta. Das 215 crianças, 129 (60%) não receberam qualquer antibioticoterapia, e 86 receberam antibiótico em algum momento. O antibiótico empírico foi mantido por, em média, 72 horas. Na análise univariada, dos fatores utilizados para estratificação de risco para infecção bacteriana grave, apresentaram níveis descritivos inferiores a 0,05 a temperatura >39oC, o número total de leucócitos >15.000/mm3 e >20.000/mm3 e o número total de neutrófilos >10.000/mm3. Na análise multivariada apenas o número total de neutrófilos >10.000/mm3 mostrou-se estatisticamente significante. Conclusão: o protocolo aplicado mostrou-se adequado para o atendimento e seguimento destas crianças utilizando para busca de infecção bacteriana grave exames simples e passíveis de serem realizados na maioria dos serviços. A maioria das crianças apresentou resolução espontânea do quadro de febre. Todas as infecções bacterianas graves foram identificadas, sendo a infecção urinária a mais comum. Dentre os fatores de risco o número total de neutrófilos >10.000/mm3 associou-se de maneira estatisticamente significante com infecção bacteriana grave, tanto na análise univariada como na multivariada. / Introduction: Fever without localizing signs is defined as the presence of fever of up to 7 days duration, without identifying the cause after history and physical examination. Most of these children presented self-limited acute infectious disease or is in the prodromal phase of a benign infectious disease. Few have serious bacterial infection: occult bacteremia, occult pneumonia, urinary tract infection, bacterial meningitis, septic arthritis, osteomyelitis or cellulitis. Although fever is a common complaint in emergency departments, the approach to febrile children remains controversial. Objectives: To evaluate the applicability of a standardized guideline for the management of children up to 36 months of age with fever without localizing signs and examine the risk factors for serious bacterial infection in these children. Methods: Prospective study involving children up to 36 months of age with fever without localizing signs treated at the emergency department of Hospital Universitário, Universidade de São Paulo, Brazil, from June 2006 to May 2007. The children were treated according to the guideline that classifies the risk of serious bacterial infection according to the presence or absence of toxemia, age and temperature. The laboratory screening was based on risk assessment (blood test, blood culture, urine sediment, urine culture and, if necessary, chest radiograph, cerebrospinal fluid and stool culture). Risk factors for severe bacterial infection were studied: sex, age, presence of toxemia, temperature, total number of leukocytes, total number of neutrophils and total number of young neutrophils. Results: We studied 215 children, 111 (51.6%) females. The mean age was 11.85 months (SD ± 8.91). Toxemia was found in 20 children, and 195 were well-appearing. Among the children from 3 to 36 months without toxemia, 95 had axillary temperature >39ºC. In 107 (49.8%) children, there was spontaneous resolution of fever; in 88 (40.9%), benign self-limited disease was identified; and in 20 (9.3%), there was serious bacterial infectious. Among the serious bacterial infections, we identified 16 urinary infections, three cases of pneumonia and one occult bacteremia. Of the 215 children, 129 (60%) received no therapy, and 86 received antibiotics at some point. Empirical antibiotic treatment was maintained for an average of 72 hours. The temperature >39°C, the total number of leukocytes >15.000/mm3 and >20.000/mm3 and the total number of neutrophils >10,000/mm3 were statistically significant (p<0,05) in univariate analysis of the factors used for risk stratification for serious bacterial infection. In multivariate analysis only the total number of neutrophils >10.000/mm3 was statistically significant. Conclusion: The guideline was shown to be appropriate to follow up these children using simple laboratory tests that can be carried out at most health facilities. Most of the children had spontaneous resolution of fever. All serious bacterial infections were identified, and the urinary tract infection was the most common. Among the risk factors studied the total number of neutrophils >10.000/mm3 was statistically significant with serious bacterial infection in both, univariate and multivariate analysis.

Introdução: A febre sem sinais localizatórios é definida como presença de febre de até 7 dias de duração, sem identificação da causa após anamnese e exame físico detalhados. A maioria destas crianças apresenta doença infecciosa aguda autolimitada ou está em fase prodrômica de uma doença infecciosa benigna. Poucas têm infecção bacteriana grave: bacteremia oculta, pneumonia oculta, infecção urinária, meningite bacteriana, artrite séptica, osteomielite ou celulite. Embora a febre seja uma das queixas mais comuns nos serviços de emergência, a abordagem da criança febril permanece controversa. Objetivos: avaliar a aplicabilidade de um protocolo padronizado para o atendimento e seguimento das crianças até 36 meses de idade com febre sem sinais localizatórios e analisar os fatores de risco para infecção bacteriana grave nestas crianças. Métodos: estudo prospectivo em crianças até 36 meses de idade que procuraram o pronto socorro do Hospital Universitário da Universidade de São Paulo, com quadro de febre sem sinais localizatórios, durante o período de um ano (junho/2006 a maio/2007). As crianças foram atendidas conforme protocolo que estratifica o risco de infecção bacteriana grave de acordo com a presença ou não de toxemia, idade e valor da temperatura. Conforme a avaliação de risco, indicava-se triagem laboratorial (hemograma, hemocultura, sedimento urinário, urocultura e, se necessário, radiografia torácica, liquor e coprocultura). Os fatores de risco para infecção bacteriana grave estudados foram: sexo, idade, presença de toxemia, temperatura, número total de leucócitos, número total de neutrófilos e número total de neutrófilos jovens. Resultados: Foram seguidas 215 crianças, sendo 111 (51,6%) do sexo feminino. A média de idade foi de 11,85 meses (DP ± 8,91). Vinte crianças, na avaliação inicial, apresentavam algum grau de toxemia, e 195 estavam em bom estado geral. Nas crianças de 3 a 36 meses não toxêmicas, 95 tinham temperatura axilar >39ºC. Em 107 crianças (49,8%), houve melhora espontânea do quadro febril; em 88 (40,9%), foi identificada doença benigna autolimitada; e em 20 (9,3%), infecção bacteriana grave. Dentre as infecções bacterianas graves, identificou-se 16 infecções urinárias, três pneumonias e uma bacteremia oculta. Das 215 crianças, 129 (60%) não receberam qualquer antibioticoterapia, e 86 receberam antibiótico em algum momento. O antibiótico empírico foi mantido por, em média, 72 horas. Na análise univariada, dos fatores utilizados para estratificação de risco para infecção bacteriana grave, apresentaram níveis descritivos inferiores a 0,05 a temperatura >39oC, o número total de leucócitos >15.000/mm3 e >20.000/mm3 e o número total de neutrófilos >10.000/mm3. Na análise multivariada apenas o número total de neutrófilos >10.000/mm3 mostrou-se estatisticamente significante. Conclusão: o protocolo aplicado mostrou-se adequado para o atendimento e seguimento destas crianças utilizando para busca de infecção bacteriana grave exames simples e passíveis de serem realizados na maioria dos serviços. A maioria das crianças apresentou resolução espontânea do quadro de febre. Todas as infecções bacterianas graves foram identificadas, sendo a infecção urinária a mais comum. Dentre os fatores de risco o número total de neutrófilos >10.000/mm3 associou-se de maneira estatisticamente significante com infecção bacteriana grave, tanto na análise univariada como na multivariada. / Introduction: Fever without localizing signs is defined as the presence of fever of up to 7 days duration, without identifying the cause after history and physical examination. Most of these children presented self-limited acute infectious disease or is in the prodromal phase of a benign infectious disease. Few have serious bacterial infection: occult bacteremia, occult pneumonia, urinary tract infection, bacterial meningitis, septic arthritis, osteomyelitis or cellulitis. Although fever is a common complaint in emergency departments, the approach to febrile children remains controversial. Objectives: To evaluate the applicability of a standardized guideline for the management of children up to 36 months of age with fever without localizing signs and examine the risk factors for serious bacterial infection in these children. Methods: Prospective study involving children up to 36 months of age with fever without localizing signs treated at the emergency department of Hospital Universitário, Universidade de São Paulo, Brazil, from June 2006 to May 2007. The children were treated according to the guideline that classifies the risk of serious bacterial infection according to the presence or absence of toxemia, age and temperature. The laboratory screening was based on risk assessment (blood test, blood culture, urine sediment, urine culture and, if necessary, chest radiograph, cerebrospinal fluid and stool culture). Risk factors for severe bacterial infection were studied: sex, age, presence of toxemia, temperature, total number of leukocytes, total number of neutrophils and total number of young neutrophils. Results: We studied 215 children, 111 (51.6%) females. The mean age was 11.85 months (SD ± 8.91). Toxemia was found in 20 children, and 195 were well-appearing. Among the children from 3 to 36 months without toxemia, 95 had axillary temperature >39ºC. In 107 (49.8%) children, there was spontaneous resolution of fever; in 88 (40.9%), benign self-limited disease was identified; and in 20 (9.3%), there was serious bacterial infectious. Among the serious bacterial infections, we identified 16 urinary infections, three cases of pneumonia and one occult bacteremia. Of the 215 children, 129 (60%) received no therapy, and 86 received antibiotics at some point. Empirical antibiotic treatment was maintained for an average of 72 hours. The temperature >39°C, the total number of leukocytes >15.000/mm3 and >20.000/mm3 and the total number of neutrophils >10,000/mm3 were statistically significant (p<0,05) in univariate analysis of the factors used for risk stratification for serious bacterial infection. In multivariate analysis only the total number of neutrophils >10.000/mm3 was statistically significant. Conclusion: The guideline was shown to be appropriate to follow up these children using simple laboratory tests that can be carried out at most health facilities. Most of the children had spontaneous resolution of fever. All serious bacterial infections were identified, and the urinary tract infection was the most common. Among the risk factors studied the total number of neutrophils >10.000/mm3 was statistically significant with serious bacterial infection in both, univariate and multivariate analysis.

<p>Abstract</p><p>Crimean-Congo Hemorrhagic Fever virus (CCHF) and Lassa virus are giving sources illness to humans. In addition to zoonotic transmission, CCHF and Lassa virus can spread from person to person. After a short incubation period, CCHF and Lassa virus infections are characterized by a sudden onset of high fever, chills, headache and cough just like flu. Even some people are vomiting and have diarrhoea. After a few days of illness hemorrhagic manifestations occur. Treatment options for CCHF and Lassa viruses are limited, and there is no vaccine available for use in humans. The purpose of the present study was to produce recombinant nucleocapsid protein of Lassavirus and CCHF virus including an aminoterminal His-tag by a Semliki Forest Virus Replicon (pSFV 4.2). The recombinant proteins are planned to be used in future development of diagnostic methods.</p>

Abstract Crimean-Congo Hemorrhagic Fever virus (CCHF) and Lassa virus are giving sources illness to humans. In addition to zoonotic transmission, CCHF and Lassa virus can spread from person to person. After a short incubation period, CCHF and Lassa virus infections are characterized by a sudden onset of high fever, chills, headache and cough just like flu. Even some people are vomiting and have diarrhoea. After a few days of illness hemorrhagic manifestations occur. Treatment options for CCHF and Lassa viruses are limited, and there is no vaccine available for use in humans. The purpose of the present study was to produce recombinant nucleocapsid protein of Lassavirus and CCHF virus including an aminoterminal His-tag by a Semliki Forest Virus Replicon (pSFV 4.2). The recombinant proteins are planned to be used in future development of diagnostic methods.

Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3460. Adviser: Carl S. Hacker. Includes bibliographical references.