Phylogenetic evidence for inter-typic recombination in the emergence of human enterovirus 71 subgenotypes

Background  

Human enterovirus 71 (EV-71) is a common causative agent of hand, foot and mouth disease (HFMD). In recent years, the virus has caused several outbreaks with high numbers of deaths and severe neurological complications. Several new EV-71 subgenotypes were identified from these outbreaks. The mechanisms that contributed to the emergence of these subgenotypes are unknown.     

Sequence analysis of the first B5 subgenogroup strain of enterovirus 71 isolated in Korea

Journal of Microbiology - Enterovirus A71 (EV71), the main etiological agent of handfoot- mouth disease (HFMD), circulates in many areas of the world and has caused large epidemics since 1997,...     

The contribution of recombination to heterozygosity differs among plant evolutionary lineages and life-forms

Background  

Despite its role as a generator of haplotypic variation, little is known about how the rates of recombination evolve across taxa. Recombination is a very labile force, susceptible to evolutionary and life trait related processes, which have also been correlated with general levels of genetic diversity. For example, in plants, it has been shown that long-lived outcrossing taxa, such as trees, have higher heterozygosity (H _e) at SSRs and allozymes than selfing or annual species. However, some of these tree taxa have surprisingly low levels of nucleotide diversity at the DNA sequence level, which points to recombination as a potential generator of genetic diversity in these organisms. In this study, we examine how genome-wide and within-gene rates of recombination evolve across plant taxa, determine whether such rates are influenced by the life-form adopted by species, and evaluate if higher genome-wide rates of recombination translate into higher H _e values, especially in trees.     

人肠道病毒71型与宿主相互作用研究进展

自1969年首次从美国加利福尼亚患者分离出人肠道病毒71型(Human enterovirus 71,HEV71)后,该病毒已在世界范围内造成多次暴发流行,近年来又在中国大陆、中国台湾及其他亚太地区流行。该肠道病毒主要引起婴幼儿手足口病,但在某些患儿可表现为严重的神经系统感染和呼吸、循环功能     

肠道病毒71型的功能基因组学研究进展

肠道病毒71型（enterovirus type 71,EV71）感染通常引起婴幼儿手足口病（hand,foot and mouth disease,HFMD）,但少数可引起无菌性脑膜炎（asepic meningitis）、脑炎（encephalitis）和类脊髓灰质炎的麻痹性疾病（poliomyelitis-like paralysis）等严重的神经系统疾病。功能基因组学研究对于探讨EV71的感染及复制过程、药物及疫苗的研制具有重大意义。该文就EV71的基因组结构及其功能的研究进展作简要的概述。     

The essential role of clathrin-mediated endocytosis in the infectious entry of human enterovirus 71

Little is currently known about the infectious entry process of human enterovirus 71 (HEV71) into host cells, which may represent potential anti-viral targeting sites. In this study a targeted small-interfering RNA (siRNA) screening platform assay was established and validated to identify and profile key cellular genes involved in processes of endocytosis, cytoskeletal dynamics, and endosomal trafficking essential for HEV71 infection. Screen evaluation was conducted via the expression of well characterized dominant-negative mutants, bioimaging studies (double-labeled immunofluorescence assays, transmission electron microscopy analysis), secondary siRNA-based dosage dependence studies, and drug inhibition assays. The infectious entry of HEV71 into rhabdomyosarcoma cells was shown to be significantly inhibited by siRNAs targeting genes associated with clathrin-mediated endocytosis (CME) that include AP2A1, ARRB1, CLTC, CLTCL1, SYNJ1, ARPC5, PAK1, ROCK1, and WASF1. The functional role of CME was verified by the observation of strong co-localization between HEV71 particles and clathrin as well as dose-dependent inhibition of HEV71 infection upon siRNA knockdown of CME-associated genes. HEV71 entry by CME was further confirmed via inhibition by dominant-negative EPS15 mutants and treatment of CME drug inhibitors, with more than 80% inhibition observed at 20 μm chlorpromazine. Furthermore, HEV71 infection was shown to be sensitive to the disruption of human genes in regulating early to late endosomal trafficking as well as endosomal acidic pH. The identification of clathrin-mediated endocytosis as the entry pathway for HEV71 infection of susceptible host cells contributes to a better understanding of HEV71 pathogenesis and enables future development of anti-viral strategies against HEV71 infection.     

2008～2010年河南省人肠道病毒71型基因特征和重组分析

对河南省2008～2010年河南省人肠道病毒71型进行基因特征及重组特点研究。对河南省2008～2010年分离的5株肠道病毒EV71型构建VP1序列系统进化树并分析其全基因组序列的重组特点。VP1序列系统进化分析显示2008～2010年河南株均属于C4基因型的C4a亚群,Bootscan分析和5’NCR、P1、P2、P3区的进化树证实C4基因型在2A～2B处存在EV71的B基因型和C基因型的型内重组及在3B～3C处存在EV71的B基因型和CA16/G-10间的型间重组。2008～2010年河南EV71分离株为C4基因型的C4a亚群,与2004年以来的中国大陆优势株流行趋势完全一致,EV71C4基因型存在基因型内和型间双重组现象。     

Comparison of Francisella tularensis genomes reveals evolutionary events associated with the emergence of human pathogenic strains

Background

Francisella tularensis subspecies tularensis and holarctica are pathogenic to humans, whereas the two other subspecies, novicida and mediasiatica, rarely cause disease. To uncover the factors that allow subspecies tularensis and holarctica to be pathogenic to humans, we compared their genome sequences with the genome sequence of Francisella tularensis subspecies novicida U112, which is nonpathogenic to humans.

Results

Comparison of the genomes of human pathogenic Francisella strains with the genome of U112 identifies genes specific to the human pathogenic strains and reveals pseudogenes that previously were unidentified. In addition, this analysis provides a coarse chronology of the evolutionary events that took place during the emergence of the human pathogenic strains. Genomic rearrangements at the level of insertion sequences (IS elements), point mutations, and small indels took place in the human pathogenic strains during and after differentiation from the nonpathogenic strain, resulting in gene inactivation.

Conclusion

The chronology of events suggests a substantial role for genetic drift in the formation of pseudogenes in Francisella genomes. Mutations that occurred early in the evolution, however, might have been fixed in the population either because of evolutionary bottlenecks or because they were pathoadaptive (beneficial in the context of infection). Because the structure of Francisella genomes is similar to that of the genomes of other emerging or highly pathogenic bacteria, this evolutionary scenario may be shared by pathogens from other species.     

Epidemiology and seroepidemiology of human enterovirus 71 among Thai populations

Background

Human enterovirus 71 (EV71) is an important pathogen caused large outbreaks in Asian-Pacific region with severe neurological complications and may lead to death in young children. Understanding of the etiological spectrum and epidemic changes of enterovirus and population’s immunity against EV71 are crucial for the implementation of future therapeutic and prophylactic intervention.

Results

A total of 1,182 patients who presented with the symptoms of hand foot and mouth disease (67.3%) or herpangina (HA) (16.7%) and admitted to the hospitals during 2008-2013 were tested for enterovirus using pan-enterovirus PCR targeting 5′-untranslated region and specific PCR for viral capsid protein 1 gene. Overall, 59.7% were pan-enterovirus positive comprising 9.1% EV71 and 31.2% coxsackievirus species A (CV-A) including 70.5% CV-A6, 27.6% CV-A16, 1.1% CV-A10, and 0.8% CV-A5. HFMD and HA occurred endemically during 2008-2011. The number of cases increased dramatically in June 2012 with the percentage of the recently emerged CV-A6 significantly rose to 28.4%. Co-circulation between different EV71 genotypes was observed during the outbreak. Total of 161 sera obtained from healthy individuals were tested for neutralizing antibodies (NAb) against EV71 subgenotype B5 (EV71-B5) using microneutralization assay. The seropositive rate of EV71-B5 was 65.8%. The age-adjusted seroprevalence for individuals was found to be lowest in children aged >6 months to 2 years (42.5%). The seropositive rate remained relatively low in preschool children aged > 2 years to 6 years (48.3%) and thereafter increased sharply to more than 80% in individuals aged > 6 years.

Conclusions

This study describes longitudinal data reflecting changing patterns of enterovirus prevalence over 6 years and demonstrates high seroprevalences of EV71-B5 NAb among Thai individuals. The rate of EV71 seropositive increased with age but without gender-specific significant difference. We identified that relative lower EV71 seropositive rate in early 2012 may demonstrate widely presented of EV71-B5 in the population before account for a large outbreak scale epidemic occurred in 2012 with due to a relatively high susceptibility of the younger population.     

Crystal structure of human enterovirus 71 3C protease

Human enterovirus 71 (EV71) is the major pathogen that causes hand, foot and mouth disease that particularly affects young children. Growing hand, foot and mouth disease outbreaks were observed worldwide in recent years and caused devastating losses both economically and politically. However, vaccines or effective drugs are unavailable to date. The genome of EV71 consists of a positive sense, single-stranded RNA of ∼ 7400 bp, encoding a large precursor polyprotein that requires proteolytic processing to generate mature viral proteins. The proteolytic processing mainly depends on EV71 3C protease (3C^pro) that possesses both proteolysis and RNA binding activities, which enable the protease to perform multiple tasks in viral replication and pathogen-host interactions. The central roles played by EV71 3C^pro make it an appealing target for antiviral drug development. We determined the first crystal structure of EV71 3C^pro and analyzed its enzymatic activity. The crystal structure shows that EV71 3C^pro has a typical chymotrypsin-like fold that is common in picornaviral 3C^pro. Strikingly, we found an important surface loop, also denoted as β-ribbon, which adopts a novel open conformation in EV71 3C^pro. We identified two important residues located at the base of the β-ribbon, Gly123 and His133, which form hinges that govern the intrinsic flexibility of the ribbon. Structure-guided mutagenesis studies revealed that the hinge residues are important to EV71 3C^pro proteolytic activities. In summary, our work provides the first structural insight into EV71 3C^pro, including a mobile β-ribbon, which is relevant to the proteolytic mechanism. Our data also provides a framework for structure-guided inhibitor design against EV71 3C^pro.     

An evolutionary view of human recombination

Recombination has essential functions in mammalian meiosis, which impose several constraints on the recombination process. However, recent studies have shown that, in spite of these roles, recombination rates vary tremendously among humans, and show marked differences between humans and closely related species. These findings provide important insights into the determinants of recombination rates and raise new questions about the selective pressures that affect recombination over different genomic scales, with implications for human genetics and evolutionary biology.     

Emergence and transmission pathways of rapidly evolving evolutionary branch C4a strains of human enterovirus 71 in the Central Plain of China

Background

Large-scale outbreaks of hand, foot, and mouth disease (HFMD) occurred repeatedly in the Central Plain of China (Shandong, Anhui, and Henan provinces) from 2007 until now. These epidemics have increased in size and severity each year and are a major public health concern in mainland China.

Principal Findings

Phylogenetic analysis was performed and a Bayesian Markov chain Monte Carlo tree was constructed based on the complete VP1 sequences of HEV71 isolates. These analyses showed that the HFMD epidemic in the Central Plain of China was caused by at least 5 chains of HEV71 transmission and that the virus continued to circulate and evolve over the winter seasons between outbreaks. Between 1998 and 2010, there were 2 stages of HEV71 circulation in mainland China, with a shift from evolutionary branch C4b to C4a in 2003–2004. The evolution rate of C4a HEV71 was 4.99×10^-3 substitutions per site per year, faster than the mean of all HEV71 genotypes. The most recent common ancestor estimates for the Chinese clusters dated to October 1994 and November 1993 for the C4a and C4b evolutionary branches, respectively. Compared with all C4a HEV71 strains, a nucleotide substitution in all C4b HEV71 genome (A to C reversion at nt2503 in the VP1 coding region, which caused amino acid substitution of VP1–10: Gln to His) had reverted.

Conclusions

The data suggest that C4a HEV71 strains introduced into the Central Plain of China are responsible for the recent outbreaks. The relationships among HEV71 isolates determined from the combined sequence and epidemiological data reveal the underlying seasonal dynamics of HEV71 circulation. At least 5 HEV71 lineages circulated in the Central Plain of China from 2007 to 2009, and the Shandong and Anhui lineages were found to have passed through a genetic bottleneck during the low-transmission winter season.     

Antiviral effect of geraniin on human enterovirus 71 in vitro and in vivo

Human enterovirus 71 infection causes hand, foot and mouth disease in children under 6 years of age and has caused mortalities in large-scale outbreaks in the Asia-Pacific region. No effective vaccine or antiviral drugs currently exist against enterovirus 71 in the clinic. In this study, we investigated the antiviral effect of geraniin on enterovirus 71 both in vitro and in vivo. The results showed that geraniin effectively inhibited virus replication in rhabdomyosarcoma cells with an IC(50) of 10 μg/ml. Moreover, geraniin treatment of mice that were challenged with a lethal dose of enterovirus 71 resulted in a reduction of mortality, relieved clinical symptoms, and inhibited virus replication in muscle tissues. The results suggest that geraniin may be used as a potential drug for anti-enterovirus 71.     

On the minimum number of recombination events in the evolutionary history of DNA sequences

In representing the evolutionary history of a set of binary DNA sequences by a connected graph, a set theoretical approach is introduced for studying recombination events. We show that set theoretical constraints have direct implications on the number of recombination events. We define a new lower bound on the number of recombination events and demonstrate the usefulness of our new approach through several explicit examples.     

Immunity to avirulent enterovirus 71 and coxsackie A16 virus protects against enterovirus 71 infection in mice

In this study, we sought to determine whether intratypic and intertypic cross-reactivity protected against enterovirus 71 (EV71) infection in a murine infection model. We demonstrate that active immunization of 1-day-old mice with avirulent EV71 strain or coxsackie A16 virus (CA16) by the oral route developed anti-EV71 antibodies with neutralizing activity (1:16 and 1:2, respectively). Splenocytes from both EV71- and CA16-immunized mice proliferated upon EV71 or CA16, but not coxsackie B3 virus (CB3), antigen stimulation. Immunized mice became more resistant to virulent EV71 strain challenge than nonimmunized mice. There was an increase in the percentage of activated splenic T cells and B cells in the immunized mice 2 days after EV71 challenge. The CA16 immune serum reacted with EV71 antigens in an enzyme-linked immunosorbent assay and neutralized EV71 but not CB3 or poliovirus at a titer of 1:4. Passive immunization with the CA16 immune serum reduced the clinical score, diminished the organ viral load, and increased the survival rate of mice upon EV71 challenge. CB3 neither shared in vitro cross-reactivity with EV71 nor provided in vivo protection after both active and passive immunization. These results illustrated that live vaccine is feasible for EV71 and that intertypic cross-reactivity of enteroviruses may provide a way to determine the prevalence of EV71.     

The founding mitochondrial DNA lineages of Tristan da Cunha Islanders

Genealogical histories show that the inhabitants of Tristan da Cunha are derived from a known number of founders. Using the transmission of mitochondrial DNA (mtDNA) from mother to offspring pairs, we traced the mtDNA types found in 161 extant individuals to five female founders. Although the historical data claimed that two pairs of sisters were among the founding females, mtDNA data showed support for only one pair of sisters. We also studied the fidelity of mtDNA transmission in conjunction with the genealogical data. We did not detect any mutations from 698 base pairs of sequence data from 75 individuals, which together accounted for 108 independent transmissions of mtDNA from mother to offspring. Based on this observation, we estimate that the mtDNA mutation rate is no more than one new mutation every 36 transmissions. These results indicate a high fidelity of maternal mtDNA transmission and support the utility of mtDNA in evolutionary and forensic studies. Am J Phys Anthropol 104:157–166, 1997. © 1997 Wiley-Liss, Inc.     

The evolutionary emergence of pandemic influenza

Pandemic influenza remains a serious public health threat and the processes involved in the evolutionary emergence of pandemic influenza strains remain incompletely understood. Here, we develop a stochastic model for the evolutionary emergence of pandemic influenza, and use it to address three main questions. (i) What is the minimum annual number of avian influenza virus infections required in humans to explain the historical rate of pandemic emergence? (ii) Are such avian influenza infections in humans more likely to give rise to pandemic strains if they are driven by repeated cross-species introductions, or by low-level transmission of avian influenza viruses between humans? (iii) What are the most effective interventions for reducing the probability that an influenza strain with pandemic potential will evolve? Our results suggest that if evolutionary emergence of past pandemics has occurred primarily through viral reassortment in humans, then thousands of avian influenza virus infections in humans must have occurred each year for the past 250 years. Analyses also show that if there is epidemiologically significant variation among avian influenza virus genotypes, then avian virus outbreaks stemming from repeated cross-species transmission events result in a greater likelihood of a pandemic strain evolving than those caused by low-level transmission between humans. Finally, public health interventions aimed at reducing the duration of avian virus infections in humans give the greatest reduction in the probability that a pandemic strain will evolve.     

Multiple introductions and recombination events underlie the emergence of a hyper-transmissible Cryptosporidium hominis subtype in the USA

1. Download : Download high-res image (249KB)
2. Download : Download full-size image