Causative Agent Of Plague Research Articles

Low cost, low tech SNP genotyping tools for resource-limited areas: Plague in Madagascar as a model.

BackgroundGenetic analysis of pathogenic organisms is a useful tool for linking human cases together and/or to potential environmental sources. The resulting data can also provide information on evolutionary patterns within a targeted species and phenotypic traits. However, the instruments often used to generate genotyping data, such as single nucleotide polymorphisms (SNPs), can be expensive and sometimes require advanced technologies to implement. This places many genotyping tools out of reach for laboratories that do not specialize in genetic studies and/or lack the requisite financial and technological resources. To address this issue, we developed a low cost and low tech genotyping system, termed agarose-MAMA, which combines traditional PCR and agarose gel electrophoresis to target phylogenetically informative SNPs.Methodology/Principal findingsTo demonstrate the utility of this approach for generating genotype data in a resource-constrained area (Madagascar), we designed an agarose-MAMA system targeting previously characterized SNPs within Yersinia pestis, the causative agent of plague. We then used this system to genetically type pathogenic strains of Y. pestis in a Malagasy laboratory not specialized in genetic studies, the Institut Pasteur de Madagascar (IPM). We conducted rigorous assay performance validations to assess potential variation introduced by differing research facilities, reagents, and personnel and found no difference in SNP genotyping results. These agarose-MAMA PCR assays are currently employed as an investigative tool at IPM, providing Malagasy researchers a means to improve the value of their plague epidemiological investigations by linking outbreaks to potential sources through genetic characterization of isolates and to improve understanding of disease ecology that may contribute to a long-term control effort.ConclusionsThe success of our study demonstrates that the SNP-based genotyping capacity of laboratories in developing countries can be expanded with manageable financial cost for resource constraint laboratories. This is a practical formula that reduces resource-driven limitations to genetic research and promises to advance global collective knowledge of infectious diseases emanating from resource limited regions of the world.

The crystal structure of the Yersinia pestis iron chaperone YiuA reveals a basic triad binding motif for the chelated metal.

Biological chelating molecules called siderophores are used to sequester iron and maintain its ferric state. Bacterial substrate-binding proteins (SBPs) bind iron-siderophore complexes and deliver these complexes to ATP-binding cassette (ABC) transporters for import into the cytoplasm, where the iron can be transferred from the siderophore to catalytic enzymes. In Yersinia pestis, the causative agent of plague, the Yersinia iron-uptake (Yiu) ABC transporter has been shown to improve iron acquisition under iron-chelated conditions. The Yiu transporter has been proposed to be an iron-siderophore transporter; however, the precise siderophore substrate is unknown. Therefore, the precise role of the Yiu transporter in Y. pestis survival remains uncharacterized. To better understand the function of the Yiu transporter, the crystal structure of YiuA (YPO1310/y2875), an SBP which functions to present the iron-siderophore substrate to the transporter for import into the cytoplasm, was determined. The 2.20 and 1.77 Å resolution X-ray crystal structures reveal a basic triad binding motif at the YiuA canonical substrate-binding site, indicative of a metal-chelate binding site. Structural alignment and computational docking studies support the function of YiuA in binding chelated metal. Additionally, YiuA contains two mobile helices, helix 5 and helix 10, that undergo 2-3 Å shifts across crystal forms and demonstrate structural breathing of the c-clamp architecture. The flexibility in both c-clamp lobes suggest that YiuA substrate transfer resembles the Venus flytrap mechanism that has been proposed for other SBPs.

Protein abundances can distinguish between naturally-occurring and laboratory strains of Yersinia pestis, the causative agent of plague.

The rapid pace of bacterial evolution enables organisms to adapt to the laboratory environment with repeated passage and thus diverge from naturally-occurring environmental (“wild”) strains. Distinguishing wild and laboratory strains is clearly important for biodefense and bioforensics; however, DNA sequence data alone has thus far not provided a clear signature, perhaps due to lack of understanding of how diverse genome changes lead to convergent phenotypes, difficulty in detecting certain types of mutations, or perhaps because some adaptive modifications are epigenetic. Monitoring protein abundance, a molecular measure of phenotype, can overcome some of these difficulties. We have assembled a collection of Yersinia pestis proteomics datasets from our own published and unpublished work, and from a proteomics data archive, and demonstrated that protein abundance data can clearly distinguish laboratory-adapted from wild. We developed a lasso logistic regression classifier that uses binary (presence/absence) or quantitative protein abundance measures to predict whether a sample is laboratory-adapted or wild that proved to be ~98% accurate, as judged by replicated 10-fold cross-validation. Protein features selected by the classifier accord well with our previous study of laboratory adaptation in Y. pestis. The input data was derived from a variety of unrelated experiments and contained significant confounding variables. We show that the classifier is robust with respect to these variables. The methodology is able to discover signatures for laboratory facility and culture medium that are largely independent of the signature of laboratory adaptation. Going beyond our previous laboratory evolution study, this work suggests that proteomic differences between laboratory-adapted and wild Y. pestis are general, potentially pointing to a process that could apply to other species as well. Additionally, we show that proteomics datasets (even archived data collected for different purposes) contain the information necessary to distinguish wild and laboratory samples. This work has clear applications in biomarker detection as well as biodefense.

Characterization of Yersinia pestis Interactions with Human Neutrophils In vitro.

Yersinia pestis is a gram-negative, zoonotic, bacterial pathogen, and the causative agent of plague. The bubonic form of plague occurs subsequent to deposition of bacteria in the skin by the bite of an infected flea. Neutrophils are recruited to the site of infection within the first few hours and interactions between neutrophils and Y. pestis have been demonstrated in vivo. In contrast to macrophages, neutrophils have been considered non-permissive to Y. pestis intracellular survival. Several studies have shown killing of the vast majority of Y. pestis ingested by human neutrophils. However, survival of 10–15% of Y. pestis after phagocytosis by neutrophils is consistently observed. Furthermore, these surviving bacteria eventually replicate within and escape from the neutrophils. We set out to further characterize the interactions between Y. pestis and human neutrophils by (1) determining the effects of known Y. pestis virulence factors on bacterial survival after uptake by neutrophils, (2) examining the mechanisms employed by the neutrophil to kill the majority of intracellular Y. pestis, (3) determining the activation phenotype of Y. pestis-infected neutrophils, and (4) characterizing the Y. pestis-containing phagosome in neutrophils. We infected human neutrophils in vitro with Y. pestis and assayed bacterial survival and uptake. Deletion of the caf1 gene responsible for F1 capsule production resulted in significantly increased uptake of Y. pestis. Surprisingly, while the two-component regulator PhoPQ system is important for survival of Y. pestis within neutrophils, pre-induction of this system prior to infection did not increase bacterial survival. We used an IPTG-inducible mCherry construct to distinguish viable from non-viable intracellular bacteria and determined the association of the Y. pestis-containing phagosome with neutrophil NADPH-oxidase and markers of primary, secondary and tertiary granules. Additionally, we show that inhibition of reactive oxygen species (ROS) production or Src family kinases increased survival of intracellular bacteria indicating that both ROS and granule-phagosome fusion contribute to neutrophil killing of Y. pestis. The data presented here further our understanding of the Y. pestis neutrophil interactions and suggest the existence of still unknown virulence factors involved in Y. pestis survival within neutrophils.

Crystal structure of Yersinia pestis virulence factor YfeA reveals two polyspecific metal-binding sites.

Gram-negative bacteria use siderophores, outer membrane receptors, inner membrane transporters and substrate-binding proteins (SBPs) to transport transition metals through the periplasm. The SBPs share a similar protein fold that has undergone significant structural evolution to communicate with a variety of differentially regulated transporters in the cell. In Yersinia pestis, the causative agent of plague, YfeA (YPO2439, y1897), an SBP, is important for full virulence during mammalian infection. To better understand the role of YfeA in infection, crystal structures were determined under several environmental conditions with respect to transition-metal levels. Energy-dispersive X-ray spectroscopy and anomalous X-ray scattering data show that YfeA is polyspecific and can alter its substrate specificity. In minimal-media experiments, YfeA crystals grown after iron supplementation showed a threefold increase in iron fluorescence emission over the iron fluorescence emission from YfeA crystals grown from nutrient-rich conditions, and YfeA crystals grown after manganese supplementation during overexpression showed a fivefold increase in manganese fluorescence emission over the manganese fluorescence emission from YfeA crystals grown from nutrient-rich conditions. In all experiments, the YfeA crystals produced the strongest fluorescence emission from zinc and could not be manipulated otherwise. Additionally, this report documents the discovery of a novel surface metal-binding site that prefers to chelate zinc but can also bind manganese. Flexibility across YfeA crystal forms in three loops and a helix near the buried metal-binding site suggest that a structural rearrangement is required for metal loading and unloading.

Novel CTL epitopes identified through a Y. pestis proteome-wide analysis in the search for vaccine candidates against plague.

The causative agent of Plague, Yersinia pestis, is a highly virulent pathogen and a potential bioweapon. Depending on the route of infection, two prevalent occurrences of the disease are known, bubonic and pneumonic. The latter has a high fatality rate. In the absence of a licensed vaccine, intense efforts to develop a safe and efficacious vaccine have been conducted, and humoral-driven subunit vaccines containing the F1 and LcrV antigens are currently under clinical trials. It is well known that a cellular immune response might have an essential additive value to immunity and protection against Y. pestis infection. Nevertheless, very few documented epitopes eliciting a protective T-cell response have been reported. Here, we present a combined high throughput computational and experimental effort towards identification of CD8 T-cell epitopes. All 4067 proteins of Y. pestis were analyzed with state-of-the-art recently developed prediction algorithms aimed at mapping potential MHC class I binders. A compilation of the results obtained from several prediction methods revealed a total of 238,000 peptide candidates, which necessitated downstream filtering criteria. Our previously established and proven approach for enrichment of true positive CTL epitopes, which relies on mapping clusters rich in tandem or overlapping predicted MHC binders ("hotspots"), was applied, as well as considerations of predicted binding affinity. A total of 1532 peptides were tested for their ability to elicit a specific T-cell response by following the production of IFNγ from splenocytes isolated from vaccinated mice. Altogether, the screen resulted in 178 positive responders (11.8%), all novel Y. pestis CTL epitopes. These epitopes span 113 Y. pestis proteins. Substantial enrichment of membrane-associated proteins was detected for epitopes selected from hotspots of predicted MHC binders. These results considerably expand the repertoire of known CTL epitopes in Y. pestis and pave the way to attest their protective potential, and hence their contribution to a future potent subunit vaccine.

Swabbing Prairie Dog Burrows for Fleas That Transmit Yersinia pestis: Influences on Efficiency.

Scientists and health-care professionals sometimes use a swabbing technique to collect fleas from rodent burrows, and later test the fleas for Yersinia pestis, the causative agent of plague. Detection of Y. pestis is enhanced when large pools of fleas are available. The following study investigated factors that might affect the rate at which fleas are collected from burrows in colonies of black-tailed prairie dogs (Cynomys ludovicianus). Data were collected from 13 colonies in New Mexico during 0600-1000 hours, June-August 2010-2011. Fleas were scarce on swabs inserted into burrows that were not actively used by prairie dogs; fleas are presumably suppressed in burrows that are void of hosts and might have begun to collapse due to a lack of maintenance. Fleas were scarce on swabs inserted into burrows with little sunlight entering the tunnel; many species of fleas use changes in light intensity to locate objects, but if light is limited, it might be difficult to locate a swab. Fleas were scarce on swabs inserted to shallow depths underground, especially during hot mornings, and during the hottest portions of mornings; when conditions are hot above ground, ectothermic fleas might migrate into the deep components of burrows, or become less willing to jump onto hosts, making it difficult to collect the fleas with swabs. If the swabbing technique is used to survey for Y. pestis on colonies of black-tailed prairie dogs, investigators might use the results of this study to modify their methods and increase the number of fleas collected.

SUBSPECIES DIFFERENTIATION OF YERSINIA PESTIS STRAINS BY PCR WITH HYBRIDIZATION-FLUORESCENT DETECTION

Develop a method of differentiation of Y.pestis strains of different subspecies based on PCR with hybridization-fluorescent detection in real-time. DNA target search for differentiation of subspecies of plague causative agent was carried out by Mauve 2.3.1, Mega 5.0 and BLAST algorithm based on comparison of full-genome sequenc- es of Y.pestis strains. Primers and TAqMan probes were calculated for the DNA targets found, conditions of PCR with hybridization-fluorescent detection - optimized. DNA targets carrying marker mutations for the caucasus, altai, gissar, ulegei subspecies, strains from Talass alpine plague reservoir were detected. The effectiveness of the DNA targets found and the developed approach of subspecies differentiation is confirmed on 101 Ypestis strains of different subspecies, isolated from natural foci of Russia, near and far abroad. The developed approach based on PCR with real-time detection allows for a rapid and effective differentiation of Ypestis strains of various subspecies.

Defining the Ail Ligand-Binding Surface: Hydrophobic Residues in Two Extracellular Loops Mediate Cell and Extracellular Matrix Binding To Facilitate Yop Delivery.

Yersinia pestis, the causative agent of plague, binds host cells to deliver cytotoxic Yop proteins into the cytoplasm that prevent phagocytosis and generation of proinflammatory cytokines. Ail is an eight-stranded β-barrel outer membrane protein with four extracellular loops that mediates cell binding and resistance to human serum. Following the deletion of each of the four extracellular loops that potentially interact with host cells, the Ail-Δloop 2 and Ail-Δloop 3 mutant proteins had no cell-binding activity while Ail-Δloop 4 maintained cell binding (the Ail-Δloop 1 protein was unstable). Using the codon mutagenesis scheme SWIM (selection without isolation of mutants), we identified individual residues in loops 1, 2, and 3 that contribute to host cell binding. While several residues contributed to the binding of host cells and purified fibronectin and laminin, as well as Yop delivery, three mutations, F80A (loop 2), S128A (loop 3), and F130A (loop 3), produced particularly severe defects in cell binding. Combining these mutations led to an even greater reduction in cell binding and severely impaired Yop delivery with only a slight defect in serum resistance. These findings demonstrate that Y. pestis Ail uses multiple extracellular loops to interact with substrates important for adhesion via polyvalent hydrophobic interactions.

A Noise Trimming and Positional Significance of Transposon Insertion System to Identify Essential Genes in Yersinia pestis

Massively parallel sequencing technology coupled with saturation mutagenesis has provided new and global insights into gene functions and roles. At a simplistic level, the frequency of mutations within genes can indicate the degree of essentiality. However, this approach neglects to take account of the positional significance of mutations - the function of a gene is less likely to be disrupted by a mutation close to the distal ends. Therefore, a systematic bioinformatics approach to improve the reliability of essential gene identification is desirable. We report here a parametric model which introduces a novel mutation feature together with a noise trimming approach to predict the biological significance of Tn5 mutations. We show improved performance of essential gene prediction in the bacterium Yersinia pestis, the causative agent of plague. This method would have broad applicability to other organisms and to the identification of genes which are essential for competitiveness or survival under a broad range of stresses.

Simulation of the use of Yersinia pestis as a Biological Weapon in Nigeria

A simulation study using the Spatiotemporal Epidemiological Modeler was carried out on the potential usage of Yersinia pestis, the causative agent of plague (‘black death’), as a biological weapon of terror. The study revealed widespread infections, incidences and deaths due to the infection all over Nigeria with bioweapon attacks originating from 2 Nigerian cities. Instituting an effective intervention program against the infection could save as many as 3.6 million lives within 10 days of the onset of the intervention program. Intervention programs could include social distancing policies and the use of antibiotics in addition to controlling the rodents and fleas vector population. Because of the relative ease of development of bioweapons and the desperation by terrorists to use any weapon at their disposal to achieve terror, there is an urgent need for an effective preparedness plan that can stop or limit the use of this category A bio-agent for biowarfare.

New Insights into Autoinducer-2 Signaling as a Virulence Regulator in a Mouse Model of Pneumonic Plague.

The Enterobacteriaceae family members, including the infamous Yersinia pestis, the causative agent of plague, have a highly conserved interbacterial signaling system that is mediated by the autoinducer-2 (AI-2) quorum-sensing molecule. The AI-2 system is implicated in regulating various bacterial virulence genes in diverse environmental niches. Deletion of the gene encoding the synthetic enzyme for the AI-2 substrate, luxS, leads to either no significant change or, paradoxically, an increase in in vivo bacterial virulence. We showed that deletion of the rbsA and lsrA genes, components of ABC transport systems that interact with AI-2, synergistically disrupted AI-2 signaling patterns and resulted in a more-than-50-fold decrease in Y.pestis strain CO92 virulence in a stringent pneumonic plague mouse model. Deletion of luxS or lsrK (encoding AI-2 kinase) from the ΔrbsA ΔlsrA background strain or complementation of the ΔrbsA ΔlsrA mutant with the corresponding gene(s) reverted the virulence phenotype to that of the wild-type Y.pestis CO92. Furthermore, the administration of synthetic AI-2 in mice infected with the ΔrbsA ΔlsrA ΔluxS mutant strain attenuated this triple mutant to a virulence phenotype similar to that of the ΔrbsA ΔlsrA strain in a pneumonic plague model. Conversely, the administration of AI-2 to mice infected with the ΔrbsA ΔlsrA ΔluxS ΔlsrK mutant did not rescue animals from lethality, indicating the importance of the AI-2-LsrK axis in regulating bacterial virulence. By performing high-throughput RNA sequencing, the potential role of some AI-2-signaling-regulated genes that modulated bacterial virulence was determined. We anticipate that the characterization of AI-2 signaling in Y.pestis will lead to reexamination of AI-2 systems in other pathogens and that AI-2 signaling may represent a broad-spectrum therapeutic target to combat antibiotic-resistant bacteria, which represent a global crisis of the 21st century. IMPORTANCEYersinia pestis is the bacterial agent that causes the highly fatal disease plague. The organism represents a significant concern because of its potential use as a bioterror agent, beyond the several thousand naturally occurring human infection cases occurring globally each year. While there has been development of effective antibiotics, the narrow therapeutic window and challenges posed by the existence of antibiotic-resistant strains represent serious concerns. We sought to identify novel virulence factors that could potentially be incorporated into an attenuated vaccine platform or be targeted by novel therapeutics. We show here that a highly conserved quorum-sensing system, autoinducer-2, significantly affected the virulence of Y.pestis in a mouse model of pneumonic plague. We also identified steps in autoinducer-2 signaling which had confounded previous studies and demonstrated the potential for intervention in the virulence mechanism(s) of autoinducer-2. Our findings may have an impact on bacterial pathogenesis research in many other organisms and could result in identifying potential broad-spectrum therapeutic targets to combat antibiotic-resistant bacteria, which represent a global crisis of the 21st century.

Complete Genome Sequence of Pigmentation-Negative Yersinia pestis Strain Cadman.

Here, we report the genome sequence of Yersinia pestis strain Cadman, an attenuated strain lacking the pgm locus. Y. pestis is the causative agent of plague and generally must be worked with under biosafety level 3 (BSL-3) conditions. However, strains lacking the pgm locus are considered safe to work with under BSL-2 conditions.

Effect of natural polymorphism on structure and function of the Yersinia pestis outer membrane porin F (OmpF protein): a computational study

The Yersinia pestis outer membrane porin F (OmpF) is a transmembrane protein located in the outer membrane of this Gram-negative bacterium which is the causative agent of plague, where it plays a significant role in controlling the selective permeability of the membrane. The amino acid sequences of OmpF proteins from 48 Y. pestis strains representing all currently available phylogenetic groups of this Gram-negative bacterium were recently deduced. Comparison of these amino acid sequences revealed that the OmpF can be present in four isoforms, the pestis-pestis type, and the pestis-microtus types I, II, and III. OmpF of the most recent pestis-pestis type has an alanine residue at the position 148, where all the pestis-microtus types have threonine there (T148A polymorphism). The variability of different pestis-microtus types is caused by an additional polymorphism at the 193rd position, where the OmpFs of the pestis-microtus type II and type III have isoleucine-glycine (IG+193) or isoleucine-glycine-isoleucine-glycine (IGIG+193) insertions, respectively (IG+193 and IGIG+193 polymorphism). To investigate potential effects of these sequence polymorphisms on the structural properties of the OmpF protein, we conducted multi-level computational analysis of its isoforms. Analysis of the I-TASSER-generated 3D-models revealed that the Yersinia OmpF is very similar to other non-specific enterobacterial porins. The T148A polymorphism affected a loop located in the external vestibule of the OmpF channel, whereas IG+193 and IGIG+193 polymorphisms affected one of its β-strands. Our analysis also suggested that polymorphism has moderate effect on the predicted local intrinsic disorder predisposition of OmpF, but might have some functional implementations.

Proteolysis of plasminogen activator inhibitor‐1 by Yersinia pestis remodulates the host environment to promote virulence

Essentials Effect of plasminogen activator inhibitor (PAI)‐1 on plague and its Y. pestis cleavage is unknown.An intranasal mouse model of infection was used to determine the role of PAI‐1 in pneumonic plague.PAI‐1 is cleaved and inactivated by the Pla protease of Y. pestis in the lung airspace.PAI‐1 impacts both bacterial outgrowth and the immune response to respiratory Y. pestis infection. Click to hear Dr Bock discuss pathogen activators of plasminogen.SummaryBackgroundThe hemostatic regulator plasminogen activator inhibitor‐1 (PAI‐1) inactivates endogenous plasminogen activators and aids in the immune response to bacterial infection. Yersinia pestis, the causative agent of plague, produces the Pla protease, a virulence factor that is required during plague. However, the specific hemostatic proteins cleaved by Pla in vivo that contribute to pathogenesis have not yet been fully elucidated.ObjectivesTo determine whether PAI‐1 is cleaved by the Pla protease during pneumonic plague, and to define the impact of PAI‐1 on Y. pestis respiratory infection in the presence or absence of Pla.MethodsAn intranasal mouse model of pneumonic plague was used to assess the levels of total and active PAI‐1 in the lung airspace, and the impact of PAI‐1 deficiency on bacterial pathogenesis, the host immune response and plasmin generation following infection with wild‐type or ∆pla Y. pestis.ResultsWe found that Y. pestis cleaves and inactivates PAI‐1 in the lungs in a Pla‐dependent manner. The loss of PAI‐1 enhances Y. pestis outgrowth in the absence of Pla, and is associated with increased conversion of plasminogen to plasmin. Furthermore, we found that PAI‐1 regulates immune cell recruitment, cytokine production and tissue permeability during pneumonic plague.ConclusionsOur data demonstrate that PAI‐1 is an in vivo target of the Pla protease in the lungs, and that PAI‐1 is a key regulator of the pulmonary innate immune response. We conclude that the inactivation of PAI‐1 by Y. pestis alters the host environment to promote virulence during pneumonic plague.

ROLE OF NEUTROPHIL EXTRACELLULAR TRAPS IN ESPECIALLY DANGEROUS BACTERIAL INFECTIONS

Novel data on neutrophil extracellular traps (NET), carrying out capture and killing of pathogenic microorganisms with higher effectiveness than during phagocytosis, are presented. A contemporary view on how neutrophils choose intracellular (phagocytosis) or extracellular (NETosis) mechanism of bactericidity during interaction with pathogenic microorganisms is given. Experimental data on the presence in causative agents of plague, cholera and melioidosis of mechanisms of protection from bactericidal effect of NET, as well as NET’S role in regulation of immune response and sepsis development are analyzed.

The effect of growth temperature on the nanoscale biochemical surface properties of Yersinia pestis.

Yersinia pestis, the causative agent of plague, has been responsible for several recurrent, lethal pandemics in history. Currently, it is an important pathogen to study owing to its virulence, adaptation to different environments during transmission, and potential use in bioterrorism. Here, we report on the changes to Y. pestis surfaces in different external microenvironments, specifically culture temperatures (6, 25, and 37°C). Using nanoscale imaging coupled with functional mapping, we illustrate that changes in the surfaces of the bacterium from a morphological and biochemical standpoint can be analyzed simultaneously using atomic force microscopy. The results from functional mapping, obtained at a single cell level, show that the density of lipopolysaccharide (measured via terminal N-acetylglucosamine) on Y. pestis grown at 37°C is only slightly higher than cells grown at 25°C, but nearly three times higher than cells maintained at 6°C for an extended period of time, thereby demonstrating that adaptations to different environments can be effectively captured using this technique. This nanoscale evaluation provides a new microscopic approach to study nanoscale properties of bacterial pathogens and investigate adaptations to different external environments.

Yersinia adhesins: An arsenal for infection.

The Yersiniae are a group of Gram-negative coccobacilli inhabiting a wide range of habitats. The genus harbors three recognized human pathogens: Y. enterocolitica and Y. pseudotuberculosis, which both cause gastrointestinal disease, and Y. pestis, the causative agent of plague. These three organisms have served as models for a number of aspects of infection biology, including adhesion, immune evasion, evolution of pathogenic traits, and retracing the course of ancient pandemics. The virulence of the pathogenic Yersiniae is heavily dependent on a number of adhesin molecules. Some of these, such as the Yersinia adhesin A and invasin of the enteropathogenic species, and the pH 6 antigen of Y. pestis, have been extensively studied. However, genomic sequencing has uncovered a host of other adhesins present in these organisms, the functions of which are only starting to be investigated. Here, we review the current state of knowledge on the adhesin molecules present in the Yersiniae, and their functions and putative roles in the infection process.

Host-Parasite Associations in Small Mammal Communities in Semiarid Savanna Ecosystems of East Africa.

Despite the established importance of rodents as reservoirs of vector-borne zoonoses in East Africa, there is relatively limited information regarding the infestation parameters and host associations of ectoparasites that vector many such pathogens among small mammals in this region. Between 2009 and 2013, small mammals were live-trapped in the semiarid savanna of Kenya. A subset of these individual hosts, including 20 distinct host taxa, was examined for ectoparasites, which were identified to species. Species of fleas, ticks, mites, and sucking lice were recorded. Based on these data, we calculated host-specific infestation parameters, documented host preferences among ectoparasites, conducted a rarefaction analysis and extrapolation to determine if ectoparasites were adequately sampled, and assessed nestedness for fleas to understand how pathogens might spread in this system. We found that the flea community structure was significantly nested. Understanding the ectoparasite network structure may have significant human relevance, as at least seven of the ectoparasite species collected are known vectors of pathogens of medical importance in the region, including Yersinia pestis, Rickettsia spp., and Theileria parva, the causative agents of plague, spotted fevers and other rickettsial illnesses in humans, and theileriosis, respectively.

MiRNA profiling in blood and tissue samples in non‐human primates in response to Yersinia pestis infection using next generation sequencing approach

MicroRNAs (miRNAs) are a class of small non‐coding RNA molecules (20–30 nucleotides long) that regulate gene expression negatively by transcriptional inhibition or post‐translational degradation. It is increasingly clear that miRNAs are an important part of the host response to microbes. In the current study, African green monkeys (Cercopithecus aethiops) were challenged intranasally with aerosolized Yersinia pestis strain CO92. Blood and tissue samples were collected post‐exposure from 6 hr through 42 hr time points. We used a next generation sequencing approach to profile miRNA expression patterns in different tissues and blood samples. Y. pestis is a gram‐negative facultative anaerobe and the causative agent of plague, which throughout history has had devastating effects on human populations. Our data enabled us to examine miRNA expression profiles during infection and identify tissue‐specific expression patterns during progression of the disease. We observed 180 upregulated miRNAs and 272 downregulated miRNAs across all time points in blood samples. At the 24 hr time point there were 52 differentially expressed (DE) miRNAs in blood samples that targeted 5381 genes. Functional analysis of DE miRNAs and their targets showed activation of TGF‐β and STAT3 pathways. TGF‐ β activated kinase 1 (TAK1, also called MAP3K7) is a known key regulator that inhibits innate immune signaling. In the kidney tissues there were a total of 32 DE miRNAs across all time points. At the 24 hr time point there were 11 unique mature DE miRNAs in exposed animal tissues as compared with non‐exposed animal tissues. In the lung tissues there were 67 DE miRNAs across all time points. At the 24 hr time point there were seven mature DE miRNAs when compared with non‐exposed animal tissues. These seven miRNAs were also DE in the blood samples. In the mediastinal lymph node tissues we had 52 DE miRNAs, eight of which were in common with blood samples. In the submandibular lymph nodes there were 25 DE miRNAs, three of which were in common with blood. We found that blood, lung, submandibular lymph and mediastinal lymph tissues shares two common miRNAs: mml‐miR‐590‐5p, which has 301 gene targets, and mml‐miR‐451, which has 17 gene targets. Both of these miRNAs are known to contribute to cytokine and grown factor activity, and miR‐590‐5p is a known circulating biomarker miRNA in cardiovascular and inflammatory diseases. Follow‐up comprehensive investigations are currently underway to further elucidate disease progression.Support or Funding InformationThis work was supported by the Defense Threat Reduction Agency, contractTMTI0029_09_WR_T