Advances In Molecular Epidemiology Research Articles

Towards a rapid sequencing-based molecular surveillance and mosaicism investigation of Toxoplasma gondii.

Advances in molecular epidemiology of Toxoplasma gondii are hampered by technical and cost-associated hurdles underlying the acquisition of genomic data from parasites. In order to implement an enhanced genotyping approach for molecular surveillance of T. gondii, we applied a multi-locus amplicon-based sequencing strategy to samples associated with human infection. This approach, targeting genome-dispersed polymorphic loci potentially involved in adaptation and virulence, genetically discriminated almost all 68 studied strains and revealed a scenario of marked genomic mosaicism. Two-thirds (n = 43) of all strains were classified as recombinant, although recombination seemed to be linked to the classical archetypal lineage. While 92% of the Sag2 archetype I strains revealed genetic mosaicism, only 45% of Sag2 archetype II strains were identified as recombinant. Contrarily to the virulence-associated archetype I, most type II strains (regardless of their recombination background) were non-virulent in mouse. Besides Sag2, some of the newly studied loci (namely the type I/I-like alleles of Sag1, B17, PK1, and Sag3 and type III/III-like alleles of TgM-A) constitute promising candidates to rapidly infer T. gondii mouse virulence. Our successful attempt to capture microsatellite length variation launches good perspectives for the straightforward transition from the laborious intensive historical method to more informative next-generation sequencing (NGS)/bioinformatics-based methodologies. Overall, while T. gondii whole-genome sequencing will be hardly feasible in most laboratories, this study shows that a discrete loci panel has the potential to improve the molecular epidemiology of T. gondii towards a better monitoring of circulating genotypes with clinical importance.

Population Structure and Dynamics of Helminthic Infection: Schistosomiasis.

While disease and outbreaks are mainly clonal for bacteria and other asexually reproducing organisms, sexual reproduction in schistosomes and other helminths usually results in unique individuals. For sexually reproducing organisms, the traits conserved in clones will instead be conserved in the group of organisms that tends to breed together, the population. While the same tools are applied to characterize DNA, how results are interpreted can be quite different at times (see another article in this collection, http://www.asmscience.org/content/journal/microbiolspec/10.1128/microbiolspec.AME-0002-2018). It is difficult to know what the real effect any control program has on the parasite population without assessing the health of this population, how they respond to the control measure, and how they recover, if they do. This review, part of the Microbiology Spectrum Curated Collection: Advances in Molecular Epidemiology of Infectious Diseases, concentrates on one approach using pooled samples to study schistosome populations and shows how this and other approaches have contributed to our understanding of this parasite family's biology and epidemiology. *This article is part of a curated collection.

Differentiating Epidemic from Endemic or Sporadic Infectious Disease Occurrence.

One important scope of work of epidemiology is the investigation of infectious diseases that cluster in time and place. Clusters of infectious disease may represent outbreaks or epidemics in which the cases share in common a point source exposure or an infectious agent in a chain of transmission pathways. Investigations of outbreaks of an illness can facilitate identification of a source, risk, or cause of the illness. However, most infectious disease episodes occur not as part of any apparent outbreaks but as sporadic infections. Multiple sporadic infections that occur steadily in time and place are referred to as endemic disease. How does one investigate sources and risk factors for sporadic or endemic infections? As part of the Microbiology Spectrum Curated Collection: Advances in Molecular Epidemiology of Infectious Diseases, this review discusses limitations of traditional approaches and advantages of molecular epidemiology approaches to investigate sporadic and endemic infections. Using specific examples, the discussions show that most sporadic infections are actually part of unrecognized outbreaks and that what appears to be endemic disease occurrence is actually comprised of multiple small outbreaks. These molecular epidemiologic investigations have unmasked modes of transmission of infectious agents not known to cause outbreaks. They have also raised questions about the traditional ways to measure incidence and assess sources of drug-resistant infections in community settings. The discoveries made by the application of molecular microbiology methods in epidemiologic investigations have led to creation of new public health intervention strategies that have not been previously considered. *This article is part of a curated collection.

Malaria Molecular Epidemiology: An Evolutionary Genetics Perspective.

Malaria is a vector-borne disease that involves multiple parasite species in a variety of ecological settings. However, the parasite species causing the disease, the prevalence of subclinical infections, the emergence of drug resistance, the scale-up of interventions, and the ecological factors affecting malaria transmission, among others, are aspects that vary across areas where malaria is endemic. Such complexities have propelled the study of parasite genetic diversity patterns in the context of epidemiologic investigations. Importantly, molecular studies indicate that the time and spatial distribution of malaria cases reflect epidemiologic processes that cannot be fully understood without characterizing the evolutionary forces shaping parasite population genetic patterns. Although broad in scope, this review in the Microbiology Spectrum Curated Collection: Advances in Molecular Epidemiology highlights the need for understanding population genetic concepts when interpreting parasite molecular data. First, we discuss malaria complexity in terms of the parasite species involved. Second, we describe how molecular data are changing our understanding of malaria incidence and infectiousness. Third, we compare different approaches to generate parasite genetic information in the context of epidemiologically relevant questions related to malaria control. Finally, we describe a few Plasmodium genomic studies as evidence of how these approaches will provide new insights into the malaria disease dynamics. *This article is part of a curated collection.

Phylogenetic Concepts and Tools Applied to Epidemiologic Investigations of Infectious Diseases.

In this review, which is a part of the Microbiology Spectrum Curated Collection: Advances in Molecular Epidemiology of Infectious Diseases, I present an overview of the principles used to classify organisms in the field of phylogenetics, highlight the methods used to infer the interrelationships of organisms, and summarize how these concepts are applied to molecular epidemiologic analyses. I present steps in analyses that come downstream of the assembly of a set of genomes or genes and the production of a multiple-sequence alignment or other matrices of putative orthologs for comparison. I focus on the history of the problem of phylogenetic reconstruction and debates within the field about the most appropriate methods. I illustrate methods that bridge the gap between molecular epidemiology and traditional epidemiology, including phylogenetic character evolution and geographic visualization. Finally, I provide practical advice on how to conduct an example analysis in the appendix. *This article is part of a curated collection.

Advances in Molecular Epidemiology of Infectious Diseases: Definitions, Approaches, and Scope of the Field.

Molecular epidemiology is a discipline that uses molecular microbiology tools to study the distribution and determinants of diseases in human populations and veterinary animals. Our understanding of epidemiology of infectious diseases has evolved with technological advancements made in molecular biology that refine our perception of the identity and dynamics of microorganisms. This review is an introduction to the Microbiology Spectrum Curated Collection: Advances in Molecular Epidemiology of Infectious Diseases that will discuss how these advancements have contributed to investigations of infectious disease outbreaks/epidemics, surveillance, transmission dynamics, risk factor identification, pathogenesis, and etiologic attribution of bacterial, viral, protozoan, and helminthic pathogens to a disease. Here we define "molecular epidemiology" and distinguish it from other disciplines that use many of the same molecular biology tools-taxonomy, phylogenetics, and molecular evolution of microorganisms. The Curated Collection will be spread throughout multiple issues of Microbiology Spectrum and will be divided into four general sections: (i) laboratory methods used to strain type microbial pathogens, (ii) methods used to analyze genotyping data, (iii) examples of molecular epidemiologic investigations of bacterial, viral, and parasitic diseases, and (iv) applications of molecular epidemiology to address new research questions in communicable and noncommunicable diseases. The major theme of this Curated Collection is to address the following question frequently asked by clinicians, clinical microbiologists, and public health professionals: what is the advantage or unique contribution of molecular epidemiology in solving infectious disease problems in the clinical and public health arenas? *This article is part of a curated collection.

Laboratory Methods in Molecular Epidemiology: Viral Infections.

Viruses, which are the most abundant biological entities on the planet, have been regarded as the "dark matter" of biology in the sense that despite their ubiquity and frequent presence in large numbers, their detection and analysis are not always straightforward. The majority of them are very small (falling under the limit of 0.5 μm), and collectively, they are extraordinarily diverse. In fact, the majority of the genetic diversity on the planet is found in the so-called virosphere, or the world of viruses. Furthermore, the most frequent viral agents of disease in humans display an RNA genome, and frequently evolve very fast, due to the fact that most of their polymerases are devoid of proofreading activity. Therefore, their detection, genetic characterization, and epidemiological surveillance are rather challenging. This review (part of the Curated Collection on Advances in Molecular Epidemiology of Infectious Diseases) describes many of the methods that, throughout the last few decades, have been used for viral detection and analysis. Despite the challenge of having to deal with high genetic diversity, the majority of these methods still depend on the amplification of viral genomic sequences, using sequence-specific or sequence-independent approaches, exploring thermal profiles or a single nucleic acid amplification temperature. Furthermore, viral populations, and especially those with RNA genomes, are not usually genetically uniform but encompass swarms of genetically related, though distinct, viral genomes known as viral quasispecies. Therefore, sequence analysis of viral amplicons needs to take this fact into consideration, as it constitutes a potential analytic problem. Possible technical approaches to deal with it are also described here. *This article is part of a curated collection.

Laboratory Methods in Molecular Epidemiology: Bacterial Infections.

In infectious disease epidemiology, the laboratory plays a critical role in diagnosis, outbreak investigations, surveillance, and characterizing biologic properties of microbes associated with their transmissibility, resistance to anti-infectives, and pathogenesis. The laboratory can inform and refine epidemiologic study design and data analyses. In public health, the laboratory functions to assess effect of an intervention. In addition to research laboratories, the new-generation molecular microbiology technology has been adapted into clinical and public health laboratories to simplify, accelerate, and make precise detection and identification of infectious disease pathogens. This technology is also being applied to subtype microbes to conduct investigations that advance our knowledge of epidemiology of old and emerging infectious diseases. Because of the recent explosive progress in molecular microbiology technology and the vast amount of data generated from the applications of this technology, this Microbiology Spectrum Curated Collection: Advances in Molecular Epidemiology of Infectious Diseases describes these methods separately for bacteria, viruses, and parasites. This review discusses past and current advancements made in laboratory methods used to conduct epidemiologic studies of bacterial infections. It describes methods used to subtype bacterial organisms based on molecular microbiology techniques, following a discussion on what is meant by bacterial "species" and "clones." Discussions on past and new genotyping tests applied to epidemiologic investigations focus on tests that compare electrophoretic band patterns, hybridization matrices, and nucleic acid sequences. Applications of these genotyping tests to address epidemiologic issues are detailed elsewhere in other reviews of this series. *This article is part of a curated collection.

Population Genetics and Molecular Epidemiology of Eukaryotes.

Molecular epidemiology uses the distribution and organization of a pathogen's DNA to understand the distribution and determinants of disease. Since the biology of DNA for eukaryotic pathogens differs substantially from that of bacteria, the analytic approach to their molecular epidemiology can also differ. While many of the genotyping techniques presented earlier in this series, "Advances in Molecular Epidemiology of Infectious Diseases," can be applied to eukaryotes, the output must be interpreted in the light of how DNA is distributed from one generation to the next. In some cases, parasite populations can be evaluated in ways reminiscent of bacteria. They differ, however, when analyzed as sexually reproducing organisms, where all individuals are unique but the genetic composition of the population does not change unless a limited set of events occurs. It is these events (migration, mutation, nonrandom mating, selection, and genetic drift) that are of interest. At a given time, not all of them are likely to be equally important, so the list can easily be narrowed down to understand the driving forces behind the population as it is now and even what it will look like in the future. The main population characteristics measured to assess these events are differentiation and diversity, interpreted in the light of what is known about the population from observation. The population genetics of eukaryotes is important for planning and evaluation of control measures, surveillance, outbreak investigation, and monitoring of the development and spread of drug resistance. *This article is part of a curated collection.

First report of Cryptosporidium canis in foxes (Vulpes vulpes) and raccoon dogs (Nyctereutes procyonoides) and identification of several novel subtype families for Cryptosporidium mink genotype in minks (Mustela vison) in China

Despite the rapid and extensive advances in molecular epidemiology of Cryptosporidium in humans and a variety of animals, the prevalence and genetic traits of the parasite in wildlife bred in captivity and the role of the neglected hosts in zoonotic transmission of human cryptosporidiosis are rarely understood. This study investigated the prevalence, species/genotype, and subtype of Cryptosporidium in farmed fur animals in China and assessed the possibility of zoonotic transmission. Three of 191 (1.6%) foxes (Vulpes vulpes), 17 of 162 (10.5%) raccoon dogs (Nyctereutes procyonoides), and 48 of 162 (29.6%) minks (Mustela vison) were positive for Cryptosporidium by nested PCRs targeting the small subunit rRNA gene. Sequence analysis indicated the presence of only Cryptosporidium canis in foxes and raccoon dogs. There is no significant difference in prevalence between young and adult foxes (or raccoon dogs). Three Cryptosporidium species or genotype including C. canis, Cryptosporidium meleagridis, and mink genotype were determined in minks aged five to six months. Subtyping based on nucleotide and amino acid sequence polymorphisms of the 60kDa glycoprotein facilitated identification of three novel subtype families named as Xb to Xd for Cryptosporidium mink genotype. The presence of zoonotic C. canis, C. meleagridis, and Cryptosporidium mink genotype in captive-bred fur animals is of public health concerns. The findings expanded the host ranges of C. canis and C. meleagridis and confirmed genetic diversity at the subtype level in Cryptosporidium mink genotype. This is the first study reporting Cryptosporidium infections in foxes and raccoon dogs in China.

Advances in molecular epidemiology of Treponema pallidum and its neurotropic subtypes

Since the development of molecular typing system for Treponema pallidum based on acidic repeat protein(arp)gene and Treponema pallidum repeat(tpr)gene in 1998, there have been reports on the epidemiological characteristics of Treponema pallidum subtypes from various countries in the world. The tp0548 and rpsA genes also have been used for enhancing the molecular typing of Treponema pallidum. Predominant subtypes of Treponema pallidum have been reported in various regions, and there is a high variety in the subtypes of Treponema pallidum, with 14d and 14f as the most frequent subtypes. Further more, some specific subtypes of Treponema pallidum are associated with clinical manifestations of syphilis. Studies have found that some neurotropic subtypes of Treponema pallidum, for example, 14a, 14d/f and 19d/c types are related to neurosyphilis. Determining specific Treponema pallidum subtypes associated with neurosyphilis is of great significance for the clinical diagnosis, treatment and prediction of prognosis of syphilis. Key words: Syphilis; Treponema pallidum; Neurosyphilis; Epidemiology; Molecular typing

Geotemporal Analysis of Neisseria meningitidis Clones in the United States: 2000–2005

BackgroundThe detection of meningococcal outbreaks relies on serogrouping and epidemiologic definitions. Advances in molecular epidemiology have improved the ability to distinguish unique Neisseria meningitidis strains, enabling the classification of isolates into clones. Around 98% of meningococcal cases in the United States are believed to be sporadic.MethodsMeningococcal isolates from 9 Active Bacterial Core surveillance sites throughout the United States from 2000 through 2005 were classified according to serogroup, multilocus sequence typing, and outer membrane protein (porA, porB, and fetA) genotyping. Clones were defined as isolates that were indistinguishable according to this characterization. Case data were aggregated to the census tract level and all non-singleton clones were assessed for non-random spatial and temporal clustering using retrospective space-time analyses with a discrete Poisson probability model.ResultsAmong 1,062 geocoded cases with available isolates, 438 unique clones were identified, 78 of which had ≥2 isolates. 702 cases were attributable to non-singleton clones, accounting for 66.0% of all geocoded cases. 32 statistically significant clusters comprised of 107 cases (10.1% of all geocoded cases) were identified. Clusters had the following attributes: included 2 to 11 cases; 1 day to 33 months duration; radius of 0 to 61.7 km; and attack rate of 0.7 to 57.8 cases per 100,000 population. Serogroups represented among the clusters were: B (n = 12 clusters, 45 cases), C (n = 11 clusters, 27 cases), and Y (n = 9 clusters, 35 cases); 20 clusters (62.5%) were caused by serogroups represented in meningococcal vaccines that are commercially available in the United States.ConclusionsAround 10% of meningococcal disease cases in the U.S. could be assigned to a geotemporal cluster. Molecular characterization of isolates, combined with geotemporal analysis, is a useful tool for understanding the spread of virulent meningococcal clones and patterns of transmission in populations.

Trends in plant virus epidemiology: Opportunities from new or improved technologies

This review focuses on new or improved technologies currently being applied, or likely to be applied in the future, to worldwide research on plant virus epidemiology. Recent technological advances and innovations provide many opportunities to improve understanding of the way diverse types of plant virus epidemics develop and how to manage them. The review starts at the macro level by considering how recent innovations in remote sensing and precision agriculture can provide valuable information about (i) virus epidemics occurring at continental, regional or district scales (via satellites) and within individual crops (mostly via lightweight unmanned aerial vehicles), and (ii) exactly where to target control measures. It then considers recent improvements in information systems and innovations in modelling that improve (i) understanding of virus epidemics and ability to predict them, and (ii) delivery to end-users of critical advice on control measures, such as Internet-based Decision Support Systems. The review goes on to discuss how advances in analysis of spatiotemporal virus spread patterns within crops can help to enhance understanding of how virus epidemics develop and validate potentially useful virus control measures. At the micro level, the review then considers the many insights that advances in molecular epidemiology can provide about genetic variation within plant virus populations involved in epidemics, and how this variation drives what occurs at the macro level. Next, it describes how recent innovations in virus detection technologies are providing many opportunities to collect and analyse new types, and ever increasing amounts, of data about virus epidemics, and the genetic variability of the virus populations involved. Finally, the implications for plant virus epidemiology of technologies likely to be important in the future are considered. To address looming world food insecurity and threats to plant biodiversity resulting from climate change and rapid population growth, it is important that new and improved technologies that help understand and control epidemics of damaging plant viruses are adopted as smoothly and speedily as possible.

Comparison of Extended Virulence Genotypes for Bacteria Isolated From Pediatric Patients With Urosepsis, Acute Pyelonephritis, and Acute Lobar Nephronia

Despite recent advances in molecular epidemiology and pathogenecity analyses of extraintestinal Escherichia coli infections, detailed analyses identifying virulence factors of E. coli isolates from pediatric urosepsis patients have not been reported. This study was conducted to explore and differentiate bacterial virulence factors associated with urosepsis and 2 other severe parenchymal infections, acute pyelonephritis (APN) and acute lobar nephronia (ALN), in pediatric patients. Patients included in this study were those who fulfilled the diagnostic criteria of urosepsis, APN, and ALN, without underlying disease or structural anomalies, excluding those with vesicoureteral reflux. Patients with cystitis were included as controls. E. coli isolates from urine (cystitis, APN, and ALN) or blood (urosepsis) specimens were analyzed using polymerase chain reaction (PCR) for 25 virulence genes. A total of 147 children (24 cystitis, 45 APN, 48 ALN, and 30 urosepsis) were enrolled in the study. Distinct syndrome-specific differences in the distribution for certain virulence genes, but conservation across syndromes for others, were found. In addition, urosepsis isolates presented higher aggregate virulence factor scores (P < 0.0001) compared with cystitis, APN, and ALN isolates. By contrast, cystitis isolates showed significantly lower aggregate virulence factor scores than all 3 invasive urinary bacterial infections; APN (P < 0.01), ALN (P < 0.01), and urosepsis (P < 0.0001). Our findings suggested that urosepsis isolates carry more virulence factors and are likely more urovirulent compared with cystitis, APN, and ALN isolates.

Molecular epidemiology, cancer-related symptoms, and cytokines pathway

The Human Genome Project and HapMap have led to a better appreciation of the importance of common genetic variation in determining cancer risk, created potential for predicting response to therapy, and made possible the development of targeted prevention and therapeutic interventions. Advances in molecular epidemiology can be used to explore the role of genetic variation in modulating the risk for severe and persistent symptoms, such as pain, depression, and fatigue, in patients with cancer. The same genes that are implicated in cancer risk might also be involved in the modulation of therapeutic outcomes. For example, polymorphisms in several cytokine genes are potential markers for genetic susceptibility both for cancer risk and for cancer-related symptoms. These genetic polymorphisms are stable markers and easily and reliably assayed to explore the extent to which genetic variation might prove useful in identifying patients with cancer at high-risk of symptom development. Likewise, they could identify subgroups who might benefit most from symptom intervention, and contribute to developing personalized and more effective therapies for persistent symptoms.

Treatment of parasitic diarrhea in HIV-infected patients

Parasitic infections responsible for diarrhea have a worldwide distribution, overlapping with AIDS in most countries. Indeed, highly active antiretroviral therapy has markedly reduced the incidence of most parasitic opportunistic infections, but parasite-related diarrhea remains frequent and probably underestimated in developing countries. In this review, we focus on the advances in molecular epidemiology, diagnosis and current treatment of the most prevalent parasitic infections in HIV-infected patients. Most of these parasites are protozoa, whose diagnosis at the laboratory requires some adapted technique and expertise. We highlight the importance of diagnosis and the skill of the laboratory of parasitology, since most parasitic infections responsible for diarrhea in AIDS patients can be treated.

Assessing Biomarker use in Risk Assessment—A Survey of Practitioners

Advances in molecular epidemiology and mechanistic toxicology have provided increased opportunities for incorporating biomarkers in the human health risk assessment process. For years, the published literature has lauded the concept of incorporating biomarkers into risk assessments as a means to reduce uncertainty in estimating health risk. For all the potential benefits, one would think that markers of effective dose, markers of early biological effects, and markers of human susceptibility are frequently selected as the basis for quantitative human health risk assessments. For this article, we sought to determine the degree to which this evolution in risk assessment has come to pass. The extent to which biomarkers are being used in current human health risk assessment was determined through an informal survey of leading risk assessment practitioners. Case studies highlighting the evolution of risk assessment methods to include biomarkers are also described. The goal of this review was to enhance the implementation of biomarker technology in risk assessment by (1) highlighting successes in biomarker implementation, (2) identifying key barriers to overcome, and (3) describing evolutions in risk assessment methods.

Recent advances in molecular epidemiology and detection of Taylorella equigenitalis associated with contagious equine metritis (CEM)

In the present review article, recent molecular advances relating to studies with Taylorella equigenitalis, as well as the recently described second species of the genus Taylorella, namely Taylorella asinigenitalis, have been described. Molecular genotyping of T. equigenitalis strains by pulsed-field gel electrophoresis (PFGE) after digestion with the suitable restriction enzyme(s) enabled the effective discrimination of strains, thus allowing the examination of the scientific mechanism(s) for its occurrence and transmission of contagious equine metritis (CEM). Alternatively, polymerase chain reaction (PCR) amplification and nucleotide sequencing of the 16S ribosomal DNA sequence and/or the other species specific sequence(s) as targets were confirmed to be effective for identification of T. equigenitalis. These new analytical methods at the genomic DNA level also enabled the discrimination of the newly discovered donkey-related T. asinigenitalis from T. equigenitalis, and moreover, the performance of phylogenetic analysis of genus Taylorella organisms with other closely related genera. Furthermore, detailed analysis of the genes responsible for CEM within the T. equigenitalis genome would be useful to help elucidate the pathogenic virulence and transmission mechanisms associated with the important equine pathogen associated with CEM.

Chemoprevention of head and neck cancer.

Squamous cell carcinoma of the head and neck is the most common epithelial neoplasm of the upper aerodigestive tract and represents a major health concern in the United States and worldwide. Invasive squamous cell carcinoma is the end result of a multiyear, multistep process of accumulation of genetic and phenotypic damage. Chemoprevention is defined as the use of pharmacologic or natural agents that inhibit the development of invasive cancer whether by blocking the DNA damage that initiates carcinogenesis or by arresting or reversing the progression of premalignant cells in which such damage has already occurred. Chemoprevention is widely recognized as an important area of research in head and neck cancer. This article reviews the field of chemoprevention and recent advances in molecular epidemiology and genetics. Current clinical trials are described.

Molecular epidemiology of Neisseria meningitidis.

Neisseria meningitidis poses a major disease burden on human beings. Meningococcal typing has a longstanding tradition for epidemiological surveillance of the disease. Genetic and antigenetic variability resulting from horizontal genetic exchange has been exploited for this purpose. Neisseria meningitidis served as the bacterial prototype organism for the development of multi locus sequence typing, which has replaced multi locus enzyme electrophoresis as the gold standard for meningococcal typing. Due to the rapid emergence of new porin variants serotyping by monoclonal antibodies is currently being replaced by DNA sequencing of variable regions of the porA gene. Sequence data were used to characterize the population structure of meningococci in carriage and disease. The advances in molecular epidemiology of meningococcal disease, and the rapid exchange of DNA sequence data via curated internet websites have resulted in an interactive international network, which is capable of identifying newly emerging clones within weeks.