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PubMed2026

Identification and full genome sequencing of previously unknown sandfly-borne phleboviruses using a newly established capture-based next-generation sequencing approach.

Sandfly-borne phleboviruses cause febrile illness and neuroinvasive disease in humans. While infections are reported in the Mediterranean region, the discovery of previously unknown phleboviruses in sandflies from Kenya suggests a wider geographic distribution. Detection and characterization of novel phleboviruses are often hindered by low-quality and low-viral-load samples. We developed a capture-based target enrichment next-generation sequencing approach that showed a 99%-100% fold enrichment of viral genomes from primary material and provides a robust tool for generating complete genomes of both known and previously unknown viruses. From a collection of 15,652 sandflies in Kenya, we recovered seven complete coding sequences of Embossos, Bogoria, and Kiborgoch viruses, and of two previously unknown phleboviruses, which were named Sosoik and Shable viruses. Sosoik virus shared 83% amino acid identity in its RdRp gene with that of Bogoria virus, while Shable virus shared ca. 88% amino acid identity with viruses of the Salehabad serocomplex. Additionally, a reassortant of Shable virus was detected that possessed an M segment from an undescribed Ponticelli-like virus. DNA barcoding of blood-fed sandflies revealed several potentially novel Sergentomyia species and evidence of host-feeding on humans, livestock, and reptiles, suggesting possibilities for zoonotic transmission. Overall, our findings increase the known genetic diversity of Old World sandfly-borne phlebovirus species from 18 to 25 (by 38.9%), including the detection of viruses from all pathogenic sandfly-borne phlebovirus serocomplexes in East Africa, opening new horizons in disease ecology research.IMPORTANCEKnowledge of the genetic diversity of circulating pathogens is crucial for providing appropriate diagnostics and disease management. This study established a novel capture-based target enrichment next-generation sequencing approach that enabled the near-complete viral genome recovery from primary samples, while native NGS yielded negative or poor-quality results. In addition to the five recently discovered sandfly-borne phleboviruses in Kenya, two previously unknown phleboviruses were detected in sandflies from the same region. The viruses were detected in several sandfly species, which showed diverse host-feeding behaviors, including mixed feeding on humans and chickens. The study significantly advances the understanding of sandfly-borne phleboviruses by uncovering their broader geographic distribution and genetic diversity, particularly in East Africa, highlighting the importance of expanding surveillance efforts beyond traditionally studied regions.

Phlebovirus↗

PubMed2003

Examination of diverse toxin-coregulated pilus-positive Vibrio cholerae strains fails to demonstrate evidence for Vibrio pathogenicity island phage.

The major virulence factors of toxigenic Vibrio cholerae are cholera toxin, which is encoded by a lysogenic filamentous bacteriophage (CTXPhi), and toxin-coregulated pilus (TCP), an essential colonization factor that is also the receptor for CTXPhi. The genes involved in the biosynthesis of TCP reside in a pathogenicity island, which has been reported to correspond to the genome of another filamentous phage (designated VPIPhi) and to encode functions necessary for the production of infectious VPIPhi particles. We examined 46 V. cholerae strains having diverse origins and carrying different genetic variants of the TCP island for the production of the VPIPhi and CTXPhi in different culture conditions, including induction of prophages with mitomycin C and UV irradiation. Although 9 of 10 V. cholerae O139 strains and 12 of 15 toxigenic El Tor strains tested produced extracellular CTXPhi, none of the 46 TCP-positive strains produced detectable VPIPhi in repeated assays, which detected as few as 10 particles of a control CTX phage per ml. These results contradict the previous report regarding VPIPhi-mediated horizontal transfer of the TCP genes and suggest that the TCP island is unable to support the production of phage particles. Further studies are necessary to understand the mechanism of horizontal transfer of the TCP island.

Bacteriophages↗

PubMed2004

Genetic characterization of Erwinia amylovora strains by amplified fragment length polymorphism.

AIMS: Erwinia amylovora is one of the most important pathogens of pear and apple and is subject to strict quarantine regulations worldwide, although its patterns of dispersal are largely unknown. Previous attempts to fingerprint E. amylovora strains by molecular techniques have detected very little polymorphism because of the high genetic homogeneity of this bacterium. Our aim was to establish and test a typing method to quantify genetic diversity among strains of this plant pathogen. METHODS AND RESULTS: Twenty-two strains from different hosts and geographical locations were examined by PCR fingerprinting with four primers and by amplified fragment length polymorphism (AFLP) with four selected combinations of primers with a single base extension. PCR fingerprinting revealed little polymorphism producing the same amplification patterns for 17 strains, while the combined AFLP patterns yielded 78 polymorphic bands (34% of total bands) and allowed the differentiation of all but two strains. Clustering of strains in the resulting dendrogram was not correlated with host, year or country of isolation, and questions previous genealogies based on PFGE patterns. CONCLUSIONS: The AFLP technique allowed the detection of an unprecedented number of genetic markers in E. amylovora and proved to be the most useful tool so far for discriminating among strains of this pathogen. The results obtained in this study strongly suggest the occurrence of multiple introductions of the pathogen in Spain and other European countries. SIGNIFICANCE AND IMPACT OF THE STUDY: A major limitation in understanding the ecology of fire blight is the lack of typing techniques with a high power of discrimination. This study demonstrates the high resolution and the usefulness of the AFLP technique to differentiate among E. amylovora strains.

Biodiversity↗

PubMed2006

Assessing the effects of human mixing patterns on human immunodeficiency virus-1 interhost phylogenetics through social network simulation.

Geneticists seeking to understand HIV-1 evolution among human hosts generally assume that hosts represent a panmictic population. Social science research demonstrates that the network patterns over which HIV-1 spreads are highly nonrandom, but the effect of these patterns on the genetic diversity of HIV-1 and other sexually transmitted pathogens has yet to be thoroughly examined. In addition, interhost phylogenetic models rarely account explicitly for genetic diversity arising from intrahost dynamics. This study outlines a graph-theoretic framework (exponential random graph modeling, ERGM) for the estimation, inference, and simulation of dynamic partnership networks. This approach is used to simulate HIV-1 transmission and evolution under eight mixing patterns resembling those observed in empirical human populations, while simultaneously incorporating intrahost viral diversity. Models of parametric growth fit panmictic populations well, yielding estimates of total viral effective population on the order of the product of infected host size and intrahost effective viral population size. Populations exhibiting patterns of nonrandom mixing differ more widely in estimates of effective population size they yield, however, and reconstructions of population dynamics can exhibit severe errors if panmixis is assumed. I discuss implications for HIV-1 phylogenetics and the potential for ERGM to provide a general framework for addressing these issues.

Evolution, Molecular↗

PubMed1996

Animal mycoplasmoses: a general introduction.

The nature of the smallest prokaryotes with autonomous replication, i.e. the mycoplasmas, which constitute the class Mollicutes, is presented in brief. These micro-organisms are extracellular parasites of the mucous membranes in animals. Mycoplasmas may cause infections, known as mycoplasmoses. The author describes current knowledge of the pathogenicity of mycoplasmas, especially in regard to the diverse effects on the immune system and the variability of the superficial antigens. Relatively few species of pathogenic mycoplasmas cause mycoplasmoses (both sporadic and endemic diseases) which have significant socio-economic consequences. The control and prevention of endemic mycoplasmoses can only be achieved with effective diagnostic tools and an efficient structure of epidemiological surveillance. The author recommends the encouragement of biological and genetic research on the mycoplasmas and the co-ordination of the development of diagnostic tests, including evaluation and validation in various epidemiological field situations.

Animals↗

PubMed2006

Plant pathogens drive density-dependent seedling mortality in a tropical tree.

One explanation for the extraordinary diversity of tropical forest trees is that density-dependent mortality from herbivores or pathogens puts locally rare species at an advantage. Density-dependent mortality of seeds and small seedlings is particularly intense in tropical forests, but its causes remain uncertain. Here, we show experimentally that pathogens from the Oomycota are associated with intense mortality in seedlings of a neotropical tree, Sebastiana longicuspis. Seedlings in untreated plots experienced eight times higher mortality compared with seedlings in plots treated with fungicide. Mortality was strongly density dependent: in fungicide-treated plots survival was unaffected by density, but survival in unsprayed plots was over three times higher at low density. Density-dependent mortality observed in a simultaneous, non-manipulative study was highly transient, suggesting that short-term observational studies may underestimate the intensity and form of pathogen-induced mortality. If such effects are widespread, plant pathogens may play a key role in maintaining and structuring tropical diversity.

Biodiversity↗

PubMed2001

Biogeographic range expansion into South America by Coccidioides immitis mirrors New World patterns of human migration.

Long-distance population dispersal leaves its characteristic signature in genomes, namely, reduced diversity and increased linkage between genetic markers. This signature enables historical patterns of range expansion to be traced. Herein, we use microsatellite loci from the human pathogen Coccidioides immitis to show that genetic diversity in this fungus is geographically partitioned throughout North America. In contrast, analyses of South American C. immitis show that this population is genetically depauperate and was founded from a single North American population centered in Texas. Variances of allele distributions show that South American C. immitis have undergone rapid population growth, consistent with an epidemic increase in postcolonization population size. Herein, we estimate the introduction into South America to have occurred within the last 9,000-140,000 years. This range increase parallels that of Homo sapiens. Because of known associations between Amerindians and this fungus, we suggest that the colonization of South America by C. immitis represents a relatively recent and rapid codispersal of a host and its pathogen.

Base Sequence↗

PubMed1998

Evolution of a simian immunodeficiency virus pathogen.

Analysis of disease induction by simian immunodeficiency viruses (SIV) in macaques was initially hampered by a lack of molecularly defined pathogenic strains. The first molecularly cloned SIV strains inoculated into macaques, SIVmacBK28 and SIVmacBK44 (hereafter designated BK28 and BK44, respectively), were cases in point, since they failed to induce disease within 1 year postinoculation in any inoculated animal. Here we report the natural history of infection with BK28 and BK44 in inoculated rhesus macaques and efforts to increase the pathogenicity of BK28 through genetic manipulation and in vivo passage. BK44 infection resulted in no disease in four animals infected for more than 7 years, whereas BK28 induced disease in less than half of animals monitored for up to 7 years. Elongation of the BK28 transmembrane protein (TM) coding sequence truncated by prior passage in human cells marginally increased pathogenicity, with two of four animals dying in the third year and one dying in the seventh year of infection. Modification of the BK28 long terminal repeat to include four consensus nuclear factor SP1 and two consensus NF-kappaB binding sites enhanced early virus replication without augmenting pathogenicity. In contrast, in vivo passage of BK28 from the first animal to die from immunodeficiency disease (1.5 years after infection) resulted in a consistently pathogenic strain and a 50% survival time of about 1.3 years, thus corresponding to one of the most pathogenic SIV strains identified to date. To determine whether the diverse viral quasispecies that evolved during in vivo passage was required for pathogenicity or whether a more virulent virus variant had evolved, we generated a molecular clone composed of the 3' half of the viral genome derived from the in vivo-passaged virus (H824) fused with the 5' half of the BK28 genome. Kinetics of disease induction with this cloned virus (BK28/H824) were similar to those with the in vivo-passaged virus, with four of five animals surviving less than 1.7 years. Thus, evolution of variants with enhanced pathogenicity can account for the increased pathogenicity of this SIV strain. The genetic changes responsible for this virulent transformation included at most 59 point mutations and 3 length-change mutations. The critical mutations were likely to have been multiple and dispersed, including elongation of the TM and Nef coding sequences; changes in RNA splice donor and acceptor sites, TATA box sites, and Sp1 sites; multiple changes in the V2 region of SU, including a consensus neutralization epitope; and five new N-linked glycosylation sites in SU.

Amino Acid Sequence↗

PubMed2006

Subepithelial myofibroblasts are novel nonprofessional APCs in the human colonic mucosa.

The human gastrointestinal mucosa is exposed to a diverse normal microflora and dietary Ags and is a common site of entry for pathogens. The mucosal immune system must respond to these diverse signals with either the initiation of immunity or tolerance. APCs are important accessory cells that modulate T cell responses which initiate and maintain adaptive immunity. The ability of APCs to communicate with CD4+ T cells is largely dependent on the expression of class II MHC molecules by the APCs. Using immunohistochemistry, confocal microscopy, and flow cytometry, we demonstrate that alpha-smooth muscle actin(+), CD90+ subepithelial myofibroblasts (stromal cells) constitutively express class II MHC molecules in normal colonic mucosa and that they are distinct from professional APCs such as macrophages and dendritic cells. Primary isolates of human colonic myofibroblasts (CMFs) cultured in vitro were able to stimulate allogeneic CD4+ T cell proliferation. This process was dependent on class II MHC and CD80/86 costimulatory molecule expression by the myofibroblasts. We also demonstrate that CMFs, engineered to express a specific DR4 allele, can process and present human serum albumin to a human serum albumin-specific and DR4 allele-restricted T cell hybridoma. These studies characterize a novel cell phenotype which, due to its strategic location and class II MHC expression, may be involved in capture of Ags that cross the epithelial barrier and present them to lamina propria CD4+ T cells. Thus, human CMFs may be important in regulating local immunity in the colon.

Actins↗

PubMed2006

[The new-world hantaviruses. Ecology and epidemiology of an emerging virus in Latin America].

The hantaviruses are a group of emerging rodent-borne pathogens (family Bunyaviridae; Genus Hantavirus) that are etiologic agents for hemorrhagic fever with renal syndrome (HFRS) in Europe and Asia and hantavirus cardiopulmonary syndrome (HCPS) in the Americas. HFRS is associated with rodents of the family Muridae, subfamilies Murinae and Arvicolinae; HPS is associated with rodents of the subfamily Sigmodontinae. Since the identification of HCPS in USA in 1993, a large number of cases of HPS and an increasing number of hantaviruses and rodent reservoir hosts have been identified in Central and South America. Epidemiologic studies have demonstrated important differences in frequency of infection with hantaviruses in both human and rodent host populations. Antibody prevalences in rodent and human populations may vary from less than 1% to more than 40%. Currently, more than 1500 cases of HCPS have been reported and more than 15 genetically distinct variants of hantaviruses, all associated with sigmodontine rodents, have been identified throughout the Americas. Several characteristics distinguish Latin American HCPS cases from the classical HCPS described for the first time in the USA. These include a variation in severity of disease from moderate and self-limiting to severe, the demonstration of person-to-person transmission, and a somewhat higher incidence of extrapulmonary clinical manifestations in the South American form of HCPS. Nevertheless, our understanding of hantaviruses in the Americas is still far from complete. The factors involved in the dynamics of these viruses in nature, their establishment and transmission within host populations and from hosts to humans, and the variable pathology of these viruses in humans are complex. It is likely that more hantaviruses will be described in the future, and much more data will be required in order to describe the diversity and evolution of this group of pathogens. Latin America, as the center of diversity for Sigmodontine rodents and their hantaviruses is presented with the unique opportunity as well as the challenge of being center stage for continued studies of the dynamics of hantaviruses in natural host populations and the links of host and virus to human populations.

Animals↗

PubMed2001

Use of ribotyping and random amplified polymorphic DNA to differentiate isolates of Burkholderia andropogonis.

AIMS: The aim of this study was to determine the genetic diversity among isolates of Burkholderia andropogonis from various host plant species and geographic locations. METHODS AND RESULTS: Both random amplified polymorphic DNA (RAPD) and ribotyping analyses were used to assess the diversity of B. andropogonis isolates and compare these results with pathogenicity assays carried out on a number of common hosts of the organism. CONCLUSIONS: Both RAPD and ribotyping analyses revealed a high level of genetic diversity between isolates of B. andropogonis. Both methods demonstrated a similar clustering of isolates. However, there was no strict correlation between the genetic diversity revealed and the original host, geographic location or pathogenicity of the isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on the genetic diversity of isolates of B. andropogonis. The great degree of diversity revealed in this study contrasts with the lack of phenotypic diversity within this species.

Bacterial Typing Techniques↗

PubMed2005

Genetic diversity of anaplasma species major surface proteins and implications for anaplasmosis serodiagnosis and vaccine development.

The genus Anaplasma (Rickettsiales: Anaplasmataceae) includes several pathogens of veterinary and human medical importance. An understanding of the diversity of Anaplasma major surface proteins (MSPs), including those MSPs that modulate infection, development of persistent infections, and transmission of pathogens by ticks, is derived in part, by characterization and phylogenetic analyses of geographic strains. Information concerning the genetic diversity of Anaplasma spp. MSPs will likely influence the development of serodiagnostic assays and vaccine strategies for the control of anaplasmosis.

Anaplasma marginale↗

PubMed2006

Phylogenetic origins of African and Neotropical Beauveria bassiana s.l. pathogens of the coffee berry borer, Hypothenemus hampei.

A phylogenetic epidemiological study of Beauveria bassiana s.l. was conducted for African and Neotropical pathogens of the coffee berry borer (CBB), Hypothenemus hampei, based on inferences from two nuclear intergenic regions, EFutr and Bloc. CBB pathogens were distributed among four terminal clades, however, the majority of African and Neotropical isolates cluster in a well-supported monophyletic group, informally designated AFNEO_1. Although the relationship between African and Neotropical AFNEO_1 is unresolved, the majority of alleles detected were exclusive to either the African or the Neotropical populations. These fixed genetic differences suggest that their disjunction predates the world trade in coffee. Neotropical AFNEO_1 have a broad host range and CBB pathogens are intermixed phylogenetically with isolates from diverse indigenous insects. Several Neotropical AFNEO_1 isolates were isolated from coffee plants as epiphytes or endophytes, thus plants themselves may potentially serve as reservoirs of pathogens against their insect pests. Topological incongruence between the EFutr and Bloc phylogenies of Neotropical AFNEO_1 may signify that individuals within this population are recombining.

Africa↗

PubMed2000

Citrobacter rodentium, the causative agent of transmissible murine colonic hyperplasia, exhibits clonality: synonymy of C. rodentium and mouse-pathogenic Escherichia coli.

Citrobacter rodentium (formerly Citrobacter freundii biotype 4280 and Citrobacter genomospecies 9) was described on the basis of biochemical characterization and DNA-DNA hybridization data and is the only Citrobacter species known to possess virulence factors homologous to those of the human pathogens enteropathogenic Escherichia coli and enterohemorrhagic E. coli. These virulence factors are encoded on the locus of enterocyte effacement (LEE), a pathogenicity island required for the characteristic attaching and effacing (AE) pathology seen in infection with these three pathogens. C. rodentium, which apparently infects only mice, provides a useful animal model for studying the molecular basis of AE pathology. No work has been done to assess differences in pathogenicity between C. rodentium isolates from diverse sources. Here, we report the examination of 15 C. rodentium isolates using a battery of genetic and biochemical approaches. No differences were observed between the isolates by repetitive-element sequence-based PCR analysis, biochemical analysis, and possession of LEE-specific virulence factors. These data suggest that members of the species are clonal. We further characterized an atypical E. coli strain from Japan called mouse-pathogenic E. coli (MPEC) that, in our hands, caused the same disease as C. rodentium. Applying the same battery of tests, we found that MPEC possesses LEE-encoded virulence factors and is indistinguishable from the previously characterized C. rodentium isolate DBS100. These results demonstrate that MPEC is a misclassified C. rodentium isolate and that members of this species are clonal and represent the only known attaching and effacing bacterial pathogen of mice.

Animals↗

PubMed1993

[Absence of distinctive characteristics between Staphylococcus epidermidis strains responsible of non-hospital infectious endocarditis and commensal strains].

In order to recognize particular characteristics of pathogenic strains, epidemiologic markers of 27 Staphylococcus epidermidis strains (9 pathological and 18 commensal) were studied. Nine strains were responsible of infective endocarditis (8 on native valves, and 1 on prosthetic valve). No case occurred after admission to hospital or surgery. Eighteen commensal strains were isolated from control subjects who had had no contact with the hospital environment and who had not received a recent antibiotic treatment. The microbiological characteristics were so diverse that no differentiation between the pathogenic strains and commensal strains could be done and no particular pathogenic clone was recognized.

Anti-Bacterial Agents↗

PubMed1997

Genetic relationships among Pasteurella trehalosi isolates based on multilocus enzyme electrophoresis.

Genetic diversity among 60 British Pasteurella trehalosi isolates representing the four recognized capsular serotypes, T3, T4, T10 and T15, and recovered predominantly from sheep suffering from systemic pasteurellosis, was estimated by analysing electrophoretically demonstrable allelic variation at structural genes encoding 19 enzymes. Thirteen of the locl were polymorphic and 20 distinctive multilocus genotypes (electrophoretic types, ETs) were identified. The population structure of P. trehalosi is clonal and its genetic diversity is limited compared with most other pathogenic bacteria. ETs represent clones, and isolates of the same ET were generally associated with the same combination of serotype, LPS type and outer-membrane protein (OMP) type. The genetic diversity of isolates within each of the capsular serotypes varied. Serotype T10 was represented by 18 isolates in two related ETs and exhibited little diversity. By contrast, serotype T15 was represented by 18 isolates in nine ETs and was almost as diverse as the species as a whole Serotype T4 was represented by 18 isolates in five ETs and was less diverse than serotype T15. Although serotype T3 was more diverse than serotype T15 it was represented by only three isolates. With the exception of the T10 isolates and those recovered from healthy sheep, 35 disease isolates belonged to 16 ETs, each of which was represented by only one to four isolates. The fact that a high proportion of disease is caused by a relatively large number of clones suggests that P. trehalosi is essentially an opportunistic pathogen. In addition to having the same capsular structure, isolates belonging to the two T10 clones were characterized by possession of similar, if not identical, O-antigens (LPS types 2 and 4). The occurrence of 18 serotype T10 isolates in only two ETs suggests that the T10 capsule and type 2/4 O-antigen confer enhanced virulence on members of these two clones. Multilocus enzyme electrophoresis (MLEE) had greater resolving power than did capsule/LPS/OMP analysis, being able to distinguish 20 rather than 14 sub-divisions within P. trehalosi. The technique demonstrated genetic identity or non-identity among strains of the same or different serotypes from different geographic localities within the UK and was a useful epidemiological tool.

Alleles↗

PubMed1996

Parietal occipital edema in hypertensive encephalopathy: a pathogenic mechanism.

Eight patients with hypertensive encephalopathy from diverse etiologies developed cerebral edema in the vertebrobasilar distribution which resolved after blood pressure was lowered. Parietal occipital edema is a recognized feature of hypertensive encephalopathy. The explanation for this regional pathological variation in hypertensive encephalopathy remains undefined. Some evidence suggests that sympathetic innervation of the anterior cerebral vasculature may be protective, and conversely, the relative lack of sympathetic innervation in the vertebrobasilar vasculature may predispose the parietal occipital region to the development of cerebral edema in hypertensive encephalopathy.

Adolescent↗

PubMed2005

Human astrovirus diagnosis and typing: current and future prospects.

Human astroviruses (HAstV) are important human pathogens causing gastroenteritis worldwide. The increased recognition of astroviruses as the cause of sporadic outbreaks of disease is due to the recent availability of improved diagnostic methods. During the last decade, most epidemiological surveys have chosen astrovirus-specific RT-PCR as screening methods. In addition to serotyping by molecular techniques, new typing methods are being developed that may also identify other viral properties related to virulence. The information provided by different typing assays is required for a better understanding of both the antigenic diversity and the molecular mechanisms of pathogenicity.

Astroviridae Infections↗