Studies on Infectious Mononucleosis: A Progress Report

The authors recently reported the occurrence of an unstable cytopathic effect and a “first passage” interference phenomenon in tissue culture as well as the detection of antigens by immunofluorescence following inoculation of material from patients with infectious mononucleosis.These tissue cultures have since been discovered to be contaminated by a bacterial L-form. The part played by the contaminant in the results obtained has been determined: it can explain only one of the patterns shown by immunofluorescence.The same labile interference phenomenon has been observed by two independent investigators.     

Spontaneous splenic rupture in infectious mononucleosis: a review.

Spontaneous rupture of the spleen is a rare complication of infectious mononucleosis (IM) occurring in 0.1-0.5 percent of patients with proven IM [1]. Although splenectomy has been advocated as the definitive therapy in the past, numerous recent reports have documented favorable outcomes with non-operative management. A review of the literature suggests that non-operative management can be successful if appropriate criteria, such as hemodynamic stability and transfusion requirements are applied in patient selection. We report the case of a 36 year old man with infectious mononucleosis who had a spontaneous splenic rupture and who was successfully managed by splenectomy. Based on review of the literature, an approach to management of a spontaneously ruptured spleen secondary to IM is suggested.     

Studies on Persistent Infections of Tissue Culture VI. Reversible Changes in Newcastle Disease Virus Populations as a Result of Passage in L Cells or Chick Embryos

Populations of the Victoria strain of Newcastle disease virus (NDV), reisolated from persistently infected L-cell cultures and passed twice in the embryonated hen's egg (NDV_L-E-2), were found to differ strikingly from the original, chick embryo-adapted virus (NDV_o). After exposure of L cells to NDV_o at high multiplicities of infection, all cells became abortively infected; they produced only small aggregates of viral antigen and few, if any, infectious virus particles, but they yielded large amounts of interferon. No cytopathic effects (CPE) were noted, and the cultures survived readily as viral carriers. In contrast, NDV_L-E-2 yielded under similar conditions large quantities of viral antigen and infectious virus particles, but no detectable interferon, and the cultures were rapidly destroyed. This change in “virulence” was at least partially reversible by further serial passages of NDV_L-E-2 in chick embryos, as was evident from a consecutive decrease in CPE with a concomitant increasingly rapid recovery of the L-cell cultures, gradually diminishing yields of infectious viral progeny, and the returning of a capacity to induce interferon synthesis. Thus, NDV_L-E-16 resembled NDV_o in many aspects, except for a less striking reduction in its ability to replicate in L cells. Although a selection of viral variants under the given sets of conditions has not been entirely excluded, the establishment of “avirulence” appears to be largely explained by a gradual accumulation of noninfectious, interferon-inducing components in the course of serial passages in the embryonated hen's egg, and the acquisition of “virulence” by a loss of these components. The evidence is as follows. (i) By a step-wise decrease in the dose of virus and restriction of the analyses to the first infectious cycle, a multiplicity of infection was ultimately reached for all “avirulent” populations at which infected cells produced normal yields of infectious viral progeny; i.e., the interferon-inducing components were diluted to noneffective levels. The lowest multiplicity which resulted in a measurable reduction in infectious virus replication was also the last one to induce detectable interferon synthesis. (ii) All viral clones derived from “avirulent” populations behaved like NDV_L-E-2 rather than like the parent viral suspensions, except that some of them elicited small amounts of interferon in L cells. The interferon-inducing components were reduced or lost in the cloning procedures. The nature of the interferon-inducing components has not been established. These components, which were neutralized by rabbit sera against “virulent” NDV_L-E-2 populations, may represent largely inactive or incomplete virus particles; however, the infectious virus-hemagglutinin ratios of “avirulent” populations were mostly of an order similar to those of “virulent” populations. The interferon-inducing components aborted the infectious process in cells simultaneously invaded by infectious virus particles. The implications of these findings are discussed.     

The Epidemiologist's Dilemma: A Position Paper on the Role of Epidemiology in Canadian Research

Canadian epidemiology is currently in transition from being primarily interested in infectious disease to becoming active in investigating causes of non-infectious disease. Generally, epidemiologists limit their work to “field”, “basic”, or “theoretical” epidemiology. In all three fields there appears to be a shortage of qualified personnel which is likely to become even more acute because of new roles that epidemiologists will probably play in the future. A minimum of 40 full-time epidemiologists is currently required in Canada.Departments other than departments of preventive medicine at medical schools and teaching hospitals are currently spending 79.5% of all funds allocated for non-microbiological epidemiological research in Canada. Since epidemiology is by its very nature population orientated, rather than clinically orientated, clinicians require consultative advice from epidemiologists at many stages of their research. Epidemiological facilities in departments of preventive medicine should therefore be strengthened in order to provide research training and didactic courses needed as the numbers of physicians, nurses and paramedical persons are increased.     

Selecting Essential Information for Biosurveillance—A Multi-Criteria Decision Analysis

The National Strategy for Biosurveillancedefines biosurveillance as “the process of gathering, integrating, interpreting, and communicating essential information related to all-hazards threats or disease activity affecting human, animal, or plant health to achieve early detection and warning, contribute to overall situational awareness of the health aspects of an incident, and to enable better decision-making at all levels.” However, the strategy does not specify how “essential information” is to be identified and integrated into the current biosurveillance enterprise, or what the metrics qualify information as being “essential”. Thequestion of data stream identification and selection requires a structured methodology that can systematically evaluate the tradeoffs between the many criteria that need to be taken in account. Multi-Attribute Utility Theory, a type of multi-criteria decision analysis, can provide a well-defined, structured approach that can offer solutions to this problem. While the use of Multi-Attribute Utility Theoryas a practical method to apply formal scientific decision theoretical approaches to complex, multi-criteria problems has been demonstrated in a variety of fields, this method has never been applied to decision support in biosurveillance.We have developed a formalized decision support analytic framework that can facilitate identification of “essential information” for use in biosurveillance systems or processes and we offer this framework to the global BSV community as a tool for optimizing the BSV enterprise. To demonstrate utility, we applied the framework to the problem of evaluating data streams for use in an integrated global infectious disease surveillance system.     

Phase I trial of a CD8+ T-cell peptide epitope-based vaccine for infectious mononucleosis

A single blind, randomized, placebo-controlled, single-center phase I clinical trial of a CD8⁺ T-cell peptide epitope vaccine against infectious mononucleosis was conducted with 14 HLA B*0801-positive, Epstein-Barr virus (EBV)-seronegative adults. The vaccine comprised the HLA B*0801-restricted peptide epitope FLRGRAYGL and tetanus toxoid formulated in a water-in-oil adjuvant, Montanide ISA 720. FLRGRAYGL-specific responses were detected in 8/9 peptide-vaccine recipients and 0/4 placebo vaccine recipients by gamma interferon enzyme-linked immunospot assay and/or limiting-dilution analysis. The same T-cell receptor Vβ CDR3 sequence that is found in FLRGRAYGL-specific T cells from most EBV-seropositive individuals could also be detected in the peripheral blood of vaccine recipients. The vaccine was well tolerated, with the main side effect being mild to moderate injection site reactions. After a 2- to 12-year follow-up, 1/2 placebo vaccinees who acquired EBV developed infectious mononucleosis, whereas 4/4 vaccinees who acquired EBV after completing peptide vaccination seroconverted asymptomatically. Single-epitope vaccination did not predispose individuals to disease, nor did it significantly influence development of a normal repertoire of EBV-specific CD8⁺ T-cell responses following seroconversion.     

Experimental transmission of infectious mononucleosis to monkeys,and the hemagglutination of erythrocytes treated with Newcastle Disease virus

Summary The intramuscular and subcutaneous inoculation of Cynomolgus monkeys with bacteriologically sterile blood, suspensions of swollen lymph glands, and with throat-washings treated with penicillin and streptomycin, from patients suffering from infectious mononucleosis, and the inoculation with cerebrospinal fluid from a patient presenting symptoms of meningoencephalitis in the course of infectious mononucleosis, resulted in an experimental disease, which resembled abortive cases of infectious mononucleosis in man. The principal features of the experimental disease were a rise of thePaul-Bunnell titre, an increase of the percentage of mononuclear cells, a slight leucocytosis followed by a slight to moderate leucopenia, and sometimes a febrile reaction.These results are suggestive for the viral etiology of infectious mononucleosis, and they indicate that the agent is present in the pharyngeal secretions. This may give an explanation for the method of spread of the disease.Human O and chicken erythrocytes were brought in contact with the virus of Newcastle Disease, and cells treated in that way were agglutinated by 27 of 28 normal human sera to a titre of 1:64 or less. From 39 mononucleosis sera 70 per cent caused hemagglutination of N.C.D. treated human red cells and 46 per cent caused hemagglutination of N.C.D. treated chicken red cells to a titre of 1:128 or higher (maximum 1:1024). No correlation could be demonstrated between the titre of thePaul-Bunnell reaction and that of the N.C.D. treated red cells.     

PATHOLOGY AND THE PATHOLOGIST IN THE CANCER PROGRAM

Cancer as one of the most important human diseases does not present as formidable a problem as infectious disease did a century ago. The diversified cancer program, combining voluntary and governmental agencies in support of research, education and coordinated teamwork in the clinical care of the patient, presents a varied although unified approach to the problem that has never before been available for the study of any single human disease. Pathology, with its applied methods from the basic sciences, has a singular role in the scientific aspect of the cancer program. Representing a new specialty in medicine and embodying an inquiring approach to the study of human disease, pathology has a leading role to play. The pathologist, assuming the new role of “pathologist-physician” brings to the clinical care of the cancer patient the most precise methods of cancer diagnosis. The “pathologist-physician” should be a pivotal member of the “clinical team” in the immediate diagnosis, care and treatment of the cancer patient.     

Infectious mononucleosis and Epstein-Barr virus

Epstein-Barr virus (EBV) is a gamma-herpesvirus that infects over 90% of the human population worldwide. It is usually transmitted between individuals in saliva, and establishes replicative infection within the oropharynx as well as life-long latent infection of B cells. Primary EBV infection generally occurs during early childhood and is asymptomatic. If delayed until adolescence or later, it can be associated with the clinical syndrome of infectious mononucleosis (also known as glandular fever or 'mono'), an illness characterised by fevers, pharyngitis, lymphadenopathy and malaise. EBV infection is also associated with the development of EBV-associated lymphoid or epithelial cell malignancies in a small proportion of individuals. This review focuses on primary EBV infection in individuals suffering from infectious mononucleosis. It discusses the mechanism by which EBV establishes infection within its human host and the primary immune response that it elicits. It describes the spectrum of clinical disease that can accompany primary infection and summarises studies that are leading to the development of a vaccine designed to prevent infectious mononucleosis.     

The Cost of Simplifying Air Travel When Modeling Disease Spread

Background

Air travel plays a key role in the spread of many pathogens. Modeling the long distance spread of infectious disease in these cases requires an air travel model. Highly detailed air transportation models can be over determined and computationally problematic. We compared the predictions of a simplified air transport model with those of a model of all routes and assessed the impact of differences on models of infectious disease.

Methodology/Principal Findings

Using U.S. ticket data from 2007, we compared a simplified “pipe” model, in which individuals flow in and out of the air transport system based on the number of arrivals and departures from a given airport, to a fully saturated model where all routes are modeled individually. We also compared the pipe model to a “gravity” model where the probability of travel is scaled by physical distance; the gravity model did not differ significantly from the pipe model. The pipe model roughly approximated actual air travel, but tended to overestimate the number of trips between small airports and underestimate travel between major east and west coast airports. For most routes, the maximum number of false (or missed) introductions of disease is small (<1 per day) but for a few routes this rate is greatly underestimated by the pipe model.

Conclusions/Significance

If our interest is in large scale regional and national effects of disease, the simplified pipe model may be adequate. If we are interested in specific effects of interventions on particular air routes or the time for the disease to reach a particular location, a more complex point-to-point model will be more accurate. For many problems a hybrid model that independently models some frequently traveled routes may be the best choice. Regardless of the model used, the effect of simplifications and sensitivity to errors in parameter estimation should be analyzed.     

Severe Neutropenia in Infectious Mononucleosis

Mild neutropenia is a well-known concomitant of infectious mononucleosis caused by the Epstein-Barr virus (EBV) occurring in the first weeks of illness. However, severe neutropenia (less than 200 polymorphonuclear leukocytes per μl) is not generally regarded as a complication of infectious mononucleosis. Three patients were seen with severe neutropenia and EBV infection, and an additional eight cases were found in the literature. In two of the latter cases the neutropenia was fatal.In the 11 cases the severe neutropenia began 14 to 40 days after illness and usually lasted for three to seven days. At the time of severe neutropenia, studies of marrow specimens showed increased proportions of promyelocytes and myelocytes. Our data suggest that EBV infection is the proximate cause of the severe neutropenia in some patients with infectious mononucleosis and that in such cases close observation and early treatment of suspected superinfections is necessary.     

“Complications” of the Landry-Guillain-Barré-Strohl syndrome

Prognosis for life in the Landry-Guillain-Barré-Strohl syndrome is dependent upon the development of respiratory and non-respiratory “complications” and their successful management. Review of the literature, a case history, and a study of 14 patients with this syndrome at the University Hospital, Edmonton, indicate that “complications” can be anticipated in virtually all areas of acute care management, including respiratory, gastrointestinal, urinary tract, central and autonomic nervous systems, metabolic, cardiovascular, and infectious disease. The proper management of patients with the Landry-Guillain-Barré-Strohl syndrome demands an awareness of the totality of care required and the presence of a hospital system that provides for vital system monitoring and support, and for ready interdisciplinary consultation.     

The Application of the Tissue Microarray (TMA) Technology to Analyze Cerebral Organoids

“Multi-Omics” technologies have contributed greatly to the understanding of various diseases by enabling researchers to accurately and rapidly investigate the molecular circuitry that connects cellular systems. The tissue-engineered, three-dimensional (3D), in vitro disease model “organoid” integrates the “omics” results in a model system, elucidating the complex links between genotype and phenotype. These 3D structures have been used to model cancer, infectious disease, toxicity, and neurological disorders. Here, we describe the advantage of using the tissue microarray (TMA) technology to analyze human-induced pluripotent stem cell–derived cerebral organoids. Compared with the conventional processing of individual samples, sectioning and staining of TMA slides are faster and can be automated, decreasing labor and reagent costs. The TMA technology faithfully captures cell morphology variations and detects specific biomarkers. The use of this technology can scale up organoid research results in at least two ways: (1) in the number of specimens that can be analyzed simultaneously and (2) in the number of consecutive sections that can be produced for analysis with different probes and antibodies.     

Infectious Mononucleosis

A short review of past and recent works pertinent to the etiology and pathogenesis of infectious mononucleosis is presented. Epidemiological studies have led to the elaboration of hypotheses concerning the etiology, the length of the incubation period and the mode of transmission of the disease. An unusual type of infectious mononucleosis of rickettsial origin has been reported by Japanese workers. Studies of accidental and experimental transmission suggest that more than one agent may give rise to the same disease. Isolation attempts in tissue cultures have been unrewarding except for the uncovering of possible agents by interference and immunofluorescence.The atypical lymphocyte is the site of increased RNA and DNA synthesis. It does not seem to be involved in antibody synthesis. The heterophile agglutinins and other mononucleosis-associated antibodies apparently account for only part of the excess 19S antibody material found in mononucleosis sera. The origin and function of these antibodies and of the atypical lymphocyte are the subject of speculation.The final elucidation of the pathogenesis of the disease and the confirmation of the reviewed hypotheses are all dependent on the eventual discovery of the elusive etiological agent(s) of infectious mononucleosis.     

Quantifying Trends in Disease Impact to Produce a Consistent and Reproducible Definition of an Emerging Infectious Disease

The proper allocation of public health resources for research and control requires quantification of both a disease''s current burden and the trend in its impact. Infectious diseases that have been labeled as “emerging infectious diseases” (EIDs) have received heightened scientific and public attention and resources. However, the label ‘emerging’ is rarely backed by quantitative analysis and is often used subjectively. This can lead to over-allocation of resources to diseases that are incorrectly labelled “emerging,” and insufficient allocation of resources to diseases for which evidence of an increasing or high sustained impact is strong. We suggest a simple quantitative approach, segmented regression, to characterize the trends and emergence of diseases. Segmented regression identifies one or more trends in a time series and determines the most statistically parsimonious split(s) (or joinpoints) in the time series. These joinpoints in the time series indicate time points when a change in trend occurred and may identify periods in which drivers of disease impact change. We illustrate the method by analyzing temporal patterns in incidence data for twelve diseases. This approach provides a way to classify a disease as currently emerging, re-emerging, receding, or stable based on temporal trends, as well as to pinpoint the time when the change in these trends happened. We argue that quantitative approaches to defining emergence based on the trend in impact of a disease can, with appropriate context, be used to prioritize resources for research and control. Implementing this more rigorous definition of an EID will require buy-in and enforcement from scientists, policy makers, peer reviewers and journal editors, but has the potential to improve resource allocation for global health.     

Blood Pressure: A Consideration of Terminology

Concepts of hypertension have changed and changes in terminology to reflect this state of affairs are suggested. Statistically, the best mortality experience is associated with blood pressure commonly regarded as subnormal, and increments of blood pressure above this level are associated with progressive increases in mortality. The terms “normal”, “benign” and “essential” in relation to blood pressure should be abandoned. “Optimal”, “acceptable” and “hypertensive” ranges of blood pressure are suggested. Hypertension is regarded as a symptom of disease, rather than as a disease in itself, and “hypertension”, when used as a diagnostic label, should be qualified always by the primary disease, if known, or by the modifying phrase, “of unknown cause”, if not known.     

Coxsackievirus B Exits the Host Cell in Shed Microvesicles Displaying Autophagosomal Markers

Coxsackievirus B3 (CVB3), a member of the picornavirus family and enterovirus genus, causes viral myocarditis, aseptic meningitis, and pancreatitis in humans. We genetically engineered a unique molecular marker, “fluorescent timer” protein, within our infectious CVB3 clone and isolated a high-titer recombinant viral stock (Timer-CVB3) following transfection in HeLa cells. “Fluorescent timer” protein undergoes slow conversion of fluorescence from green to red over time, and Timer-CVB3 can be utilized to track virus infection and dissemination in real time. Upon infection with Timer-CVB3, HeLa cells, neural progenitor and stem cells (NPSCs), and C2C12 myoblast cells slowly changed fluorescence from green to red over 72 hours as determined by fluorescence microscopy or flow cytometric analysis. The conversion of “fluorescent timer” protein in HeLa cells infected with Timer-CVB3 could be interrupted by fixation, suggesting that the fluorophore was stabilized by formaldehyde cross-linking reactions. Induction of a type I interferon response or ribavirin treatment reduced the progression of cell-to-cell virus spread in HeLa cells or NPSCs infected with Timer-CVB3. Time lapse photography of partially differentiated NPSCs infected with Timer-CVB3 revealed substantial intracellular membrane remodeling and the assembly of discrete virus replication organelles which changed fluorescence color in an asynchronous fashion within the cell. “Fluorescent timer” protein colocalized closely with viral 3A protein within virus replication organelles. Intriguingly, infection of partially differentiated NPSCs or C2C12 myoblast cells induced the release of abundant extracellular microvesicles (EMVs) containing matured “fluorescent timer” protein and infectious virus representing a novel route of virus dissemination. CVB3 virions were readily observed within purified EMVs by transmission electron microscopy, and infectious virus was identified within low-density isopycnic iodixanol gradient fractions consistent with membrane association. The preferential detection of the lipidated form of LC3 protein (LC3 II) in released EMVs harboring infectious virus suggests that the autophagy pathway plays a crucial role in microvesicle shedding and virus release, similar to a process previously described as autophagosome-mediated exit without lysis (AWOL) observed during poliovirus replication. Through the use of this novel recombinant virus which provides more dynamic information from static fluorescent images, we hope to gain a better understanding of CVB3 tropism, intracellular membrane reorganization, and virus-associated microvesicle dissemination within the host.     

H-rosette formation in T-cell-proliferative diseases.

In health up to 6% of human lymphocytes will form rosettes with homologous group O rhesus-negative erythrocytes (H rosettes). Increased numbers of H rosettes were found in T-cell-proliferative diseases-- namely, infectious mononucleosis, Sézary''s syndrome, and T-cell leukaemia. H-rosette formation is thus a marker for a subpopulation of T lymphocytes, and this easily performed test of T-cell activation may have clinical value in characterising changes in blood lymphocyte populations in disease.     

Microbiome-immune interactions in tuberculosis

Tuberculosis (TB) remains an infectious disease of global significance and a leading cause of death in low- and middle-income countries. Significant effort has been directed towards understanding Mycobacterium tuberculosis genomics, virulence, and pathophysiology within the framework of Koch postulates. More recently, the advent of “-omics” approaches has broadened our appreciation of how “commensal” microbes have coevolved with their host and have a central role in shaping health and susceptibility to disease. It is now clear that there is a diverse repertoire of interactions between the microbiota and host immune responses that can either sustain or disrupt homeostasis. In the context of the global efforts to combatting TB, such findings and knowledge have raised important questions: Does microbiome composition indicate or determine susceptibility or resistance to M. tuberculosis infection? Is the development of active disease or latent infection upon M. tuberculosis exposure influenced by the microbiome? Does microbiome composition influence TB therapy outcome and risk of reinfection with M. tuberculosis? Can the microbiome be actively managed to reduce risk of M. tuberculosis infection or recurrence of TB? Here, we explore these questions with a particular focus on microbiome-immune interactions that may affect TB susceptibility, manifestation and progression, the long-term implications of anti-TB therapy, as well as the potential of the host microbiome as target for clinical manipulation.