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Yersinia pestis, the causative agent of plague and a biological threat agent, presents an urgent need for novel medical countermeasures due to documented cases of naturally acquired antibiotic resistance and potential person-to-person spread during a pneumonic infection. Immunotherapy has been proposed as a way to circumvent current and future antibiotic resistance. Here, we describe the development and characterization of two affinity matured human antibodies (αF1Ig AM2 and αF1Ig AM8) that promote survival of mice after exposure to aerosolized Y. pestis. We share details of the error prone PCR and yeast display technology-based affinity maturation process that we used. The resultant matured antibodies have nanomolar affinity for Y. pestis F1 antigen, are produced in high yield, and are resilient to 37°C stress for up to 6 months. Importantly, in vitro assays using a murine macrophage cell line demonstrated that αF1Ig AM2 and αF1Ig AM8 are opsonic. Even more importantly, in vivo studies using pneumonic plague mouse models showed that 100% of the mice receiving 500 μg of IgGs αF1Ig AM2 and αF1Ig AM8 survived lethal challenge with aerosolized Y. pestis CO92. Combined, these results provide evidence of the quality and robustness of αF1Ig AM2 and αF1Ig AM8 and support their development as potential medical countermeasures against plague.

According to WHO, plague, caused by Yersinia pestis, has resurged since 2000. Inner Mongolia, harboring a quarter of China's plague foci, has accounted for 80% of national plague cases in the past five years. Despite its pivotal role in Chinese plague epidemiology, the genetic diversity and transmission dynamics of Y. pestis in this region remain under-investigated. Our analysis of 585 Y. pestis strains from Inner Mongolia (1948–2021) revealed three primary lineages, with 2.MED3 being predominant. We further delineated seven sub-phylogroups in 2.MED3, with 2.MED3.1.2 and 2.MED3.1.4 showing recent dominance. These two subgroups reveal dual transmission patterns: localized short-distance spread and long-distance dispersals over 300 km. Xilingol League is highlighted as a key source and reservoir for Y. pestis, predominantly spreading from central-eastern to southwestern Inner Mongolia, including occasional reverse transmissions. These findings enhance understanding of Y. pestis diversity and transmission in Inner Mongolia, aiding in enhanced surveillance and control measures. Analysis of the genomes and spatiotemporal distribution of Yersinia pestis in Inner Mongolia, China, reveals its genetic diversity and dual transmission patterns, including localized short-distance spread and long-distance dispersals. [ABSTRACT FROM AUTHOR]

Pyroptosis is a gasdermin-mediated pro-inflammatory programmed cell death that, during microbial infections, aims to restrict the spreading of bacteria. Nevertheless, excessive pyroptosis activation leads to inflammation levels that are detrimental to the host. Pathogen-associated molecular patterns (PAMPs) present in bacteria and outer membrane vesicles (OMVs) can trigger pyroptosis pathways in different cell types with different outcomes. Moreover, some pathogens have evolved virulence factors that directly interfere with pyroptosis pathways, like Yersinia pestis YopM and Shigella flexneri IpaH7.8. Other virulence factors, such as those of Neisseria meningitidis, Neisseria gonorrhoeae, Salmonella enterica, and Helicobacter pylori affect pyroptosis pathways indirectly with important differences between pathogenic and commensal species of the same family. These pathogens deserve special attention because of the increasing antimicrobial resistance of S. flexneri and N. gonorrhoeae, the high prevalence of S. enterica and H. pylori, and the life-threatening diseases caused by N. meningitidis and Y. pestis. While inflammation due to macrophage pyroptosis has been extensively addressed, the effects of activation of pyroptosis pathways on modulation of cell cytoskeleton and cell–cell junctions in epithelia and endothelia and on the bacterial crossing of epithelial and endothelial barriers have only been partly investigated. Another important point is the diverse consequences of pyroptosis pathways on calcium influx, like activation of calcium-dependent enzymes and mitochondria dysregulation. This review will discuss the pyroptotic pathways activated by Gram-negative bacteria and their OMVs, analyzing the differences between pathogens and commensal bacteria. Particular attention will also be paid to the experimental models adopted and the main results obtained in the different models. Finally, strategies adopted by pathogens to modulate these pathways will be discussed with a perspective on the use of pyroptosis inhibitors as adjuvants in the treatment of infections. [ABSTRACT FROM AUTHOR]

Introduction: Marmots are recognized as host animals for plague caused by Yersinia pestis infection. It is unclear that why plague prevalent in marmot rather than other rodents like pikas in the same habitats. This study aims to analyze the differences of the soil characteristics around marmots and pikas burrows to explore the soils factors impacting on different epidemic intensities of Yersinia pestis in these two rodents. Methods: Soil samples were collected from within and around marmot and pika burrows, as well as from the nearby areas not inhabited by them and Chinese baseline soil properties as control groups, in the Qilian Mountains of Gansu Province, China. The physicochemical properties and the bacterial 16S rRNA were measured to analyze the characteristics of soils from different groups. Subsequently, the data were analyzed using R studio. Results: The analysis revealed that marmot habitats exhibited distinct soil characteristics, including lower organic matter and alkaline hydrolyzed nitrogen, but higher electrical conductivity and total soluble salts. And soil in marmot areas tended to have higher concentrations of nickel, chromium, and iron, also lower levels of zinc and selenium. Additionally, the alpha diversity of soil microorganisms in marmot habitats was significantly low. Simultaneously, redundancy analysis was conducted, which showed that the low alpha diversity of marmot-soil was influenced by its physicochemical properties. The alpha diversity of the soil was positively correlated with EC, TSS, Na, and Cr, etc., while it was negatively correlated with AHN, OM, Se, Zn, and Fe, etc. Conclusion: These characteristics in marmot habitats, including low levels of organic matter, alkaline hydrolyzed nitrogen, zinc, selenium, and bacterial alpha diversity, as well as high levels of electrical conductivity, total soluble salts, iron, and nickel, played a crucial role in the spread of plague. It was discovered that the unique characteristics ofmarmot-soils provided essential elements necessary for the survival of Yersinia pestis, including high levels of Fe and Ca, or facilitated the spread of plague. Thus, the transmission of the plague was facilitated. [ABSTRACT FROM AUTHOR]

Leukotriene B4 (LTB4) is an inflammatory lipid produced in response to pathogens that is critical for initiating the inflammatory cascade needed to control infection. However, during plague, Yersinia pestis inhibits the timely synthesis of LTB4 and subsequent inflammation. Using bacterial mutants, we previously determined that Y. pestis inhibits LTB4 synthesis via the action of the Yop effector proteins that are directly secreted into host cells through a type 3 secretion system (T3SS). Here, we show that the T3SS is the primary pathogen associated molecular pattern (PAMP) required for production of LTB4 in response to both Yersinia and Salmonella. However, we also unexpectantly discovered that T3SS-mediated LTB4 synthesis by neutrophils and macrophages require the activation of two distinctly different host signaling pathways. We identified that phagocytosis and the NLRP3/CASP1 inflammasome significantly impact LTB4 synthesis by macrophages but not neutrophils. Instead, the SKAP2/PLC signaling pathway is required for T3SS-mediated LTB4 production by neutrophils. Finally, while recognition of the T3SS is required for LTB4 production, we also discovered that a second unrelated PAMP-mediated signal activates the MAP kinase pathway needed for synthesis. Together, these data demonstrate significant differences in the host factors and signaling pathways required by macrophages and neutrophils to quickly produce LTB4 in response to bacteria. Moreover, while macrophages and neutrophils might rely on different signaling pathways for T3SS-dependent LTB4 synthesis, Y. pestis has evolved virulence mechanisms to counteract this response by either leukocyte to inhibit LTB4 synthesis and colonize the host. Author summary: The production of inflammatory lipid mediators by the host is essential for timely inflammation in response to invasion by bacterial pathogens. Therefore, defining how immune cells recognize pathogens and rapidly produce these lipids is essential for us to understand how our immune system effectively controls infection. In this study, we discovered that the host signaling pathways required for leukotriene B4 (LTB4) synthesis differ between neutrophils and macrophages, highlighting important differences in how immune cells respond to infection. Together, these data represent a significant improvement in our understanding of how neutrophils and macrophages rapidly react to bacteria and provide new insights into how Yersinia pestis manipulates leukocytes to evade immune recognition to cause disease. [ABSTRACT FROM AUTHOR]

Yersinia pestis, the causative agent of plague, is a highly virulent bacterium that poses a significant threat to human health. Preserving this bacterium in a viable state is crucial for research and diagnostic purposes. This paper presents and evaluates a simple lyophilization protocol for the long-term storage of Y. pestis strains from Fiocruz-CYP, aiming to explore its impact on viability and long-term stability, while replacing the currently used methodologies. The lyophilization tests were conducted using the non-virulent Y. pestis strain EV76, subjected to the lyophilization process under vacuum conditions. Viability assessment was performed to evaluate the effects of lyophilization and storage conditions on Y. pestis under multiple temperature conditions (− 80 °C, − 20 °C, 4–8 °C and room temperature). The lyophilization protocol employed in this study consistently demonstrated its efficacy in maintaining high viability rates for Y. pestis samples in a up to one year follow-up. The storage temperature that consistently exhibited the highest recovery rates was − 80 °C, followed by − 20 °C and 4–8 °C. Microscopic analysis of the post-lyophilized cultures revealed preserved morphological features, consistent with viable bacteria. The high viability rates observed in the preserved samples indicate the successful preservation of Y. pestis using this protocol. Overall, the presented lyophilization protocol provides a valuable tool for the long-term storage of Y. pestis, offering stability, viability, and functionality. By refining the currently used methods of lyophilization, this protocol can improve long-term preservation for Y. pestis strains collections, facilitating research efforts, diagnostic procedures, and the development of preventive and therapeutic strategies against plague. [ABSTRACT FROM AUTHOR]

This study focuses on Yersinia pestis, the bacterium responsible for plague, which posed a severe threat to public health in history. Despite the availability of antibiotics treatment, the emergence of antibiotic resistance in this pathogen has increased challenges of controlling the infections and plague outbreaks. The development of new drug targets and therapies is urgently needed. This research aims to identify novel protein targets from 28 Y. pestis strains by the integrative pan-genomic and subtractive genomics approach. Additionally, it seeks to screen out potential safe and effective alternative therapies against these targets via high-throughput virtual screening. Targets should lack homology to human, gut microbiota, and known human ‘anti-targets’, while should exhibit essentiality for pathogen’s survival and virulence, druggability, antibiotic resistance, and broad spectrum across multiple pathogenic bacteria. We identified two promising targets: the aminotransferase class I/class II domain-containing protein and 3-oxoacyl-[acyl-carrier-protein] synthase 2. These proteins were modeled using AlphaFold2, validated through several structural analyses, and were subjected to molecular docking and ADMET analysis. Molecular dynamics simulations determined the stability of the ligand-target complexes, providing potential therapeutic options against Y. pestis. [ABSTRACT FROM AUTHOR]

There is a compelling demand for approved plague vaccines due to the endemicity of Yersinia pestis and its potential for pandemic spread. Whilst substantial progress has been made, we recommend that the global funding and health security systems should work urgently to translate some of the efficacious vaccines reviewed herein to expedite clinical development and to prevent future disastrous plague outbreaks, particularly caused by antimicrobial resistant Y. pestis strains. Content includes material subject to Crown Copyright © 2024.This is an open access article under the Open Government License (http://www.nationalarchives.gov.uk/doc/open-government-licence/version/3/). [ABSTRACT FROM AUTHOR]

Plague is a deadly zoonosis that still poses a threat in many regions of the world. We combined epidemiologic, host, and vector surveillance data collected during 1961– 1980 from the Araripe Plateau focus in northeastern Brazil with ecologic, geoclimatic, and Yersinia pestis genomic information to elucidate how these factors interplay in plague activity. We identified well-delimited plague hotspots showing elevated plague risk in low-altitude areas near the foothills of the plateau’s concave sectors. Those locations exhibited distinct precipitation and vegetation coverage patterns compared with the surrounding areas. We noted a seasonal effect on plague activity, and human cases linearly correlated with precipitation and rodent and flea Y. pestis positivity rates. Genomic characterization of Y. pestis strains revealed a foundational strain capable of evolving into distinct genetic variants, each linked to temporally and spatially constrained plague outbreaks. These data could identify risk areas and improve surveillance in other plague foci within the Caatinga biome. [ABSTRACT FROM AUTHOR]

In the second half of the 20th century, through the efforts of scientists from many countries, a coherent theory of natural plague foci (sylvatic plague) was formulated, attempted to describe the history of the origin and evolution of the causative agent of plague infection, the microbeYersinia pestis. But the accumulated knowledge in this regard remained extremely limited. Envisioned by the modern phylogenetics, the methods of phylogenetic constructions in the pregenomic time were rather primitive, “manual”, characteristic of early empirico-intuitive Haeckel phylogenetics. Since the beginning of the 21st century, the introduction of genomic methodologies in the study of the plague pathogen allowed to detail the intraspecific diversity (subspecies, genovariants) of this particularly dangerous pathogen at the level of geographical and local populations (individual natural foci) around the world and to bring the diagnostics and description of intraspecific diversity to a high degree of perfection. Two important discoveries were made. First, the direct ancestor of the plague microbe was reliably established, it turned out to be the causative agent of intestinal infection — Far Eastern scarlet-like fever (Y. pseudotuberculosis0:1b). Secondly, the evolutionary youth of the plague pathogen was shown, the “molecular clock” showed the time of its divergence from the ancestral population no earlier than 30 thousand years ago. Thus, the root of the phylogenetic tree ofY. pestiswas fully characterized. Nevertheless, molecular genetic (MG) achievements do not yet allow to reveal the secrets of its phylogeny, i.e. the origin and sequence of world expansion. The most important reason is the high dependence of the MG of phylogeny reconstructions on the choice of evolutionary model for the analyzed characters: the model of neutral evolution is traditionally accepted, but its adequacy in relation toY. pestisphylogeny is questioned by many well-known ecological (in the broad sense) facts. At the same time, MG achievements contributed to the creation of an ecological (ECO) approach based on the provisions of the theory of natural plague foci in an updated version, according to which the plague pathogen is an evolutionarily young pathogen descended from a psychrophilic pseudotuberculous ancestor. The presumptive ECO scenario has no obvious natural-scientific and historical contradictions and can serve as a null hypothesis for improving the MG methodology of phylogenetic constructions for plague and other similar microbes. It is suggested that the creation of a real phylogeny of the plague microbe is possible only based on integration of MG and ECO approaches.

Plague is a deadly zoonosis that still poses a threat in many regions of the world. We combined epidemiologic, host, and vector surveillance data collected during 1961–1980 from the Araripe Plateau focus in northeastern Brazil with ecologic, geoclimatic, and Yersinia pestis genomic information to elucidate how these factors interplay in plague activity. We identified well-delimited plague hotspots showing elevated plague risk in low-altitude areas near the foothills of the plateau’s concave sectors. Those locations exhibited distinct precipitation and vegetation coverage patterns compared with the surrounding areas. We noted a seasonal effect on plague activity, and human cases linearly correlated with precipitation and rodent and flea Y. pestis positivity rates. Genomic characterization of Y. pestis strains revealed a foundational strain capable of evolving into distinct genetic variants, each linked to temporally and spatially constrained plague outbreaks. These data could identify risk areas and improve surveillance in other plague foci within the Caatinga biome.

The phylogenies of the plague microbe (Yersinia pestis), reconstructed on the basis of an advanced molecular genetic (MG) approach, are not congruent with the facts accumulated by classical scientific areas: ecology, biogeography, paleontology, epizootiology, and others. The MG approach cannot name the original host of the plague pathogen and reliably characterize the root of the phylogenetic tree. This deficiency is compensated by the ecological (in a broad sense) (ECO) approach, which operates with such ecological, phylogeographic, and biogeographic categories as geographical population, subspecies, range, ecological niche, and direct kinship. Y. pestis, the "blood dweller" of warm-blooded rodent hosts, is transmitted through flea bites and is unique in the family of predominantly intestinal bacteria Yersiniaceae (Enterobacteriaceae). According to the ECO approach, its uniqueness is associated with the origin in the populations of its primary host, the Mongolian marmot (Marmota sibirica), in unique circumstances, in which the marmot population was infected with pseudotuberculosis not by the traditional alimentary way in grasslands, but in a traumatic way during hibernation. The identification of the original host of the plague pathogen opens up broad prospects for studying its evolutionary history (speciation and intraspecific diversification) and improving the methodology of ecological, geographic, phylogeographic, and phylogenetic studies of this especially dangerous pathogen. [ABSTRACT FROM AUTHOR]

Plague is an important and serious zoonotic disease caused by the bacterium Yersinia pestis. This study is a narrative review. The required data for the research were gathered through a search in search engines such as Google Scholar and international scientific databases, including PubMed, Web of Science, ScienceDirect, Scopus, Elsevier, and Lilacs, as well as Persian databases like the Barakat Knowledge Network System (Barakatkns), the Scientific Information Database (SID), the Iranian Medical Library (Medlib), and Civilica. The search was conducted using both Persian and English keywords, including "Plague," "Yersinia pestis," "Plague outbreak," "Flea," and "Control," through individual and combined searches over a period from 1974 to June 2024. In the present article, the agents and types of plague, its transmission methods, diagnostic approaches, the status of plague in the world and Iran, fleas in Iran, and methods for prevention and control of the disease will be discussed. This disease has caused three major historical pandemics, including the Black Death in the 14th century, which claimed millions of lives in Europe, Asia, and Africa and profoundly affected the populations of the impacted countries. Recently, the highest number of human plague cases has been reported from Madagascar. The main vector of the disease in humans is known as Xenopsylla cheopis worldwide. In Iran, records of human plague date back to the past. Climate change and ecological shifts may influence the distribution of plague vectors and hosts, increasing the risk of its resurgence even in areas where the disease is currently dormant. In Iran, increasing interactions between human communities and wildlife, along with changes in agricultural practices influenced by climatic factors, could facilitate the spread of the plague. Reports of the pathogen in rodents and dogs in western Iran (Kurdistan and Hamadan) underscore the significance of the potential re-emergence of plague in the region. There are three clinical forms of this disease bubonic, septicemic, and pulmonary. The most common type of plague is bubonic plague, which occurs following the bite of a flea that has previously fed on an infected animal. It presents with symptoms including fever, headache, chills, weakness, fatigue, and swollen, inflamed lymph nodes. Delayed treatment increases the risk of mortality associated with the disease. Plague, with a history of horror and devastation, remains a global health challenge. Understanding the current situation of this disease and reviewing its prevention and control is very important to prevent its re-emergence. Considering the current plague situation in Iran, there is a need for continuous monitoring and more detailed studies on the ecology of vectors, as well as the development of effective strategies for disease control and prevention. International cooperation in scientific research and exchanging information about plague outbreaks and control strategies are among the other effective approaches in this field. Finally, strengthening the health infrastructure and developing educational programs for the community is one of the most important requirements to prevent the spread of plague and other infectious diseases in Iran.

Introduction. Based on data on the role of blood erythrocytes in the development and implementation of the vaccine and infectious processes in plague, it was of interest to evaluate changes in erythrocyte surface architecture from the position of searching for informative criteria for the preclinical evaluation of anti-plague vaccines. Aim — using atomic force microscopy to characterize the state of the blood erythrocyte membrane of guinea pigs in response to subcutaneous administration of the vaccine strain Yersinia pestis EV NIIEG and its isogenic derivatives. Materials and methods. For immunization of animals strain Y. pestis EV NIIEG (pYT+, pYV+, pYP+) and its isogenic derivatives Y. pestis KM216 (pYT–, pYV–, pYP+), Y. pestis KM217 (pYT–, pYV+, pYP–), Y. pestis KM218 (pYT–, pYV–, pYP–) were used. Analysis of the erythrocyte membrane was carried out using a Solver P47-PRO scanning probe microscope (NT-MDT, Russia). Results. The most pronounced changes in the surface architectonics of the membrane of guinea pig erythrocytes were established during the first three days of the formation of the immune response to the Y. pestis EV strain NIIEG and its isogenic variant Y. pestis KM217, in the genome of which the pYV plasmid is preserved, administered at a dose of 5 × 108 CFU. A significant increase (p 0.05) in the proportion of transformed cell forms (43.67 ± 3.63% and 37.83 ± 7.03% versus 4.08 ± 0.86% in the control group), root mean square roughness (319 ± 8 nm and 312 ± 7 nm versus 70 ± 6 nm in the control group), Young's modulus (125.73 ± 4.48 kPa and 113.8 ± 5.41 kPa versus 53.03 ± 1.47 kPa in the control group). By the 21st day, the value of these indicators decreased by an average of 2.7, 2.0 and 1.5 times, respectively, indicating restoration of the erythrocyte membrane. Conclusion. The dependence of the changes in the erythrocyte membrane and the rate of their restoration on the plasmid composition of Y. pestis strains has been established. The data obtained contribute to the understanding of the processes of interaction of Y. pestis with the erythrocyte membrane and can be used as additional characteristics in the development of new criteria for preclinical evaluation of plague candidate vaccines.

Russian Research Anti-Plague Institute “Microbe” of the Rospotrebnadzor develops and implements methods for obtaining and purifying biologically active substances of the causative agents of plague, cholera, tularemia and anthrax, which are used in the design of preventive and diagnostic preparations.The aim of the review is to summarize and systematize the accumulated data on the isolation, purification and assessment of antigens and toxins of plague, cholera, tularemia and anthrax pathogens. Attention is also paid to the application and prospects for using the obtained antigens for the design of medical immunobiological preparations (MIBPs). The Institute “Microbe” is a reference center for plague, hence modern immunodiagnostic drugs are necessary and in demand. Antigen-level preventive drugs against the infections listed above are also the subject of study by the institute’s staff. The main stages of isolation, purification and analysis of antigens include the selection or construction of a suitable strain; cultivation, extraction, concentration and purification of antigens and toxins using biotechnological techniques that allow for obtaining and preserving the biologically active substance of interest to the researcher. To study the antigenic activity of purified antigens, laboratory animals are involved, the immune response is recorded and analyzed, and antiserum is obtained. This is followed by the stage of investigating the physicochemical and immunobiological properties of the isolated antigen preparations and drawing up an antigen profile. The characterized antigens are used for the design of preventive and diagnostic drugs.

Post-translational addition of O-linked N-acetylglucosamine (O-GlcNAc) to proteins is commonly associated with a variety of stress responses and cellular processes in eukaryotes, but its potential roles in bacteria are unclear. Here, we show that protein HmwC acts as an O-GlcNAc transferase (OGT) responsible for O-GlcNAcylation of multiple proteins in Yersinia pestis, a flea-borne pathogen responsible for plague. We identify 64 O-GlcNAcylated proteins (comprising 65 sites) with differential abundance under conditions mimicking the mammalian host (Mh) and flea vector (Fv) environments. Deletion of hmwC, encoding a putative OGT, structurally distinct from any existing member of the GT41 family, results in reduced O-GlcNAcylation, reduced growth, and alterations in virulence properties and survival under stress. Purified HmwC can modify target proteins in vitro using UDP-GlcNAc as sugar donor. One of the target proteins, OsdY, promotes Y. pestis survival under oxidative stress conditions. Thus, our results support that regulation of antioxidative responses through O-GlcNAcylation may be a conserved process shared by prokaryotes and eukaryotes. Post-translational addition of O-linked N-acetylglucosamine (O-GlcNAc) to proteins is associated with stress responses in eukaryotes. Here, the authors identify a protein that acts as an O-GlcNAc transferase for modification of multiple proteins and regulates antioxidative stress responses in the bacterial pathogen Yersinia pestis. [ABSTRACT FROM AUTHOR]

Plague is an important and serious zoonotic disease caused by the bacterium Yersinia pestis. This study is a narrative review. The required data for the research were gathered through a search in search engines such as Google Scholar and international scientific databases, including PubMed, Web of Science, ScienceDirect, Scopus, Elsevier, and Lilacs, as well as Persian databases like the Barakat Knowledge Network System (Barakatkns), the Scientific Information Database (SID), the Iranian Medical Library (Medlib), and Civilica. The search was conducted using both Persian and English keywords, including "Plague," "Yersinia pestis," "Plague outbreak," "Flea," and "Control," through individual and combined searches over a period from 1974 to June 2024. In the present article, the agents and types of plague, its transmission methods, diagnostic approaches, the status of plague in the world and Iran, fleas in Iran, and methods for prevention and control of the disease will be discussed. This disease has caused three major historical pandemics, including the Black Death in the 14th century, which claimed millions of lives in Europe, Asia, and Africa and profoundly affected the populations of the impacted countries. Recently, the highest number of human plague cases has been reported from Madagascar. The main vector of the disease in humans is known as Xenopsylla cheopis worldwide. In Iran, records of human plague date back to the past. Climate change and ecological shifts may influence the distribution of plague vectors and hosts, increasing the risk of its resurgence even in areas where the disease is currently dormant. In Iran, increasing interactions between human communities and wildlife, along with changes in agricultural practices influenced by climatic factors, could facilitate the spread of the plague. Reports of the pathogen in rodents and dogs in western Iran (Kurdistan and Hamadan) underscore the significance of the potential re-emergence of plague in the region. There are three clinical forms of this disease bubonic, septicemic, and pulmonary. The most common type of plague is bubonic plague, which occurs following the bite of a flea that has previously fed on an infected animal. It presents with symptoms including fever, headache, chills, weakness, fatigue, and swollen, inflamed lymph nodes. Delayed treatment increases the risk of mortality associated with the disease. Plague, with a history of horror and devastation, remains a global health challenge. Understanding the current situation of this disease and reviewing its prevention and control is very important to prevent its re-emergence. Considering the current plague situation in Iran, there is a need for continuous monitoring and more detailed studies on the ecology of vectors, as well as the development of effective strategies for disease control and prevention. International cooperation in scientific research and exchanging information about plague outbreaks and control strategies are among the other effective approaches in this field. Finally, strengthening the health infrastructure and developing educational programs for the community is one of the most important requirements to prevent the spread of plague and other infectious diseases in Iran. [ABSTRACT FROM AUTHOR]

Surveillance for animal plague was conducted in the Marmota himalayana plague focus of the Qinghai-Tibet Plateau from 2020 to 2023. A 22.89% positive rate of serum F1 antibody was detected in live-caught marmots, alongside a 43.40% incidence of Yersinia pestis isolation from marmot carcasses. Marmot carcasses infected with plague exhibited a significantly higher spleen-somatic index (P < 0.05). Twenty-one Y. pestis-specific phages were isolated, among which one Y. pestis lytic phage (AKS2022HT87GU_phi) was isolated from the bone marrow of a marmot carcass (no. AKS2022HT87) and was found to be symbiotic with Y. pestis. Microscopy revealed the coexistence of lysed and non-lysed colonies of Y. pestis AKS2022HT87. Genome-wide analysis showed that certain strains of the Y. pestis AKS2022HT87 carried phage DNA fragments consistent with phage AKS2022HT87GU_phi. The rare symbiotic relationship between a lytic phage and Y. pestis observed in vitro was highlighted in this study, laying the basis for further exploring the relationship between Y. pestis and its bacteriophages. [ABSTRACT FROM AUTHOR]

The role of surface antigens of Yersinia pestis in reception of the phage L-413C was evaluated experimentally. Based on the methods of phage inactivation after its co-incubation with soluble or bead-bounded antigens, the importance of the plague microbe LPS in the phage reception was confirmed, and the inability to bind the capsular antigen F1, Ail protein, and two autotransporters YapF and YapM was shown. The native and recombinant PsaA, being solved, significantly inhibited the lytic activity of the phage in contrast to the bead-bound antigens. The knockout EV cells (ΔpsaA) are able to bind the phage particles as well as the wild strain. The use of three methods to evaluate the role of the PsaA antigen in phage L-413C reception gave contradictory results. On the one hand, the reactive domains of PsaA are able to interact with phage particles in solution. At the same time, these domains appear to determine nonspecific binding of the PsaA protein to the underlying bacterial cell structures or polystyrene microsphere, preventing phage adhesion. [ABSTRACT FROM AUTHOR]

Background: The control and prevention of rodent-borne diseases are mainly based on our knowledge of ecology and the infectious status of their reservoir hosts. This study aimed to evaluate the prevalence of Francisella tularensis, Yersinia pestis, and arenavirus infections in small mammals and to assess the potential of disease occurrence in East Azerbaijan, northwest of Iran, in 2017 and 2018. Methods: Spleen and lung samples were obtained from all trapped small mammals. The real-time quantitative PCR (qPCR) method was used to detect nucleic acid sequences of F. tularensis, Y. pestis, and arenaviruses. Serum samples were tested for antibodies indicating the host response to F. tularensis and Y. pestis infections using the standard tube agglutination test and enzyme-linked immunosorbent assay (ELISA), respectively. Results: A total of 205 rodents, four Eulipotyphla, and one carnivore were captured. The most common rodent species captured (123 of 205 rodents, 60%) belonged to the genus Meriones (mainly Persian jird, Meriones persicus). In total, 317 fleas were removed from trapped animals. Flea species belonged to Xenopsylla buxtoni, Xenopsylla nuttalli, Stenoponia tripectinata, Paraceras melis, Ctenophthalmus rettigi smiti, Rhadinopsylla bivirgis, Paradoxopsyllus grenieri, and Nosopsyllus iranus. Using the qPCR tests, five spleen samples from M. persicus were positive for F. tularensis. The qPCR tests were negative for the detection of Y. pestis and arenaviruses. Finally, all serum samples tested were negative for antibodies against Y. pestis and F. tularensis. Conclusions:F. tularensis was the only zoonotic agent detected in rodents captured in East Azerbaijan. However, the diversity of trapped rodents and fleas provides the potential for the spread of various rodent-borne viral and bacterial diseases in the studied areas. [ABSTRACT FROM AUTHOR]

In this pioneering in silico study in Peru, we aimed to analyze Escherichia coli (E. coli) genomes for antimicrobial resistance genes (ARGs) diversity and virulence and for its mobilome. For this purpose, 469 assemblies from human, domestic, and wild animal hosts were investigated. Of these genomes, three were E. coli strains (pv05, pv06, and sf25) isolated from chickens in our previous study, characterized for antimicrobial susceptibility profile, and sequenced in this study. Three other genomes were included in our repertoire for having rare cgMLSTs. The phenotypic analysis for antimicrobial resistance revealed that pv05, pv06, and sf25 strains presented multidrug resistance to antibiotics belonging to at least three classes. Our in silico analysis indicated that many Peruvian genomes included resistance genes, mainly to the aminoglycoside class, ESBL-producing E. coli, sulfonamides, and tetracyclines. In addition, through Multi-locus Sequence Typing, we found more than 180 different STs, with ST10 being the most prevalent among the genomes. Pan-genome mapping revealed that, with new lineages, the repertoire of accessory genes in E. coli increased, especially genes related to resistance and persistence, which may be carried by plasmids. The results also demonstrated several genes related to adhesion, virulence, and pathogenesis, especially genes belonging to the high pathogenicity island (HPI) from Yersinia pestis, with a prevalence of 42.2% among the genomes. The complexity of the genetic profiles of resistance and virulence in our study highlights the adaptability of the pathogen to different environments and hosts. Therefore, our in silico analysis through genome sequencing enables tracking the epidemiology of E. coli from Peru and the future development of strategies to mitigate its survival. [ABSTRACT FROM AUTHOR]

Proliferative enteropathy is an enteric disease caused by the bacterium Lawsonia intracellularis, which affects several species of domestic and wild animals. The mechanisms underlying the mechanisms employed by L. intracellularis to cause host cell proliferation are poorly understood, mostly because this bacterium is extremely difficult to isolate and propagate in vitro. Comparative genomics methods for searching for genes orthologous to genes known to be associated with pathogenesis allow identification of genes potentially involved in pathogenesis by the pathogen of interest. The goal of this study was to carry out in silico research on L. intracellularis genes orthologous to genes required for intracellular invasion and survival present in other pathogenic bacteria, particularly Brucella abortus, B. melitensis, B. suis, Listeria monocytogenes, Mycobacterium tuberculosis, Mycobacterium avium subspecies paratuberculosis, Salmonella enterica, Yersinia pestis, Y. enterocolitica, and Y. pseudotuberculosis. A total of 127 genes associated with invasion and intracellular survival from five known intracellular bacteria were mapped against the predicted proteomes of all L. intracellularis strains publicly available on GenBank, using the OrthoFinder program. A total of 45 L. intracellularis genes were orthologous to genes associated with pathogenesis of other intracellular bacteria. Genes putatively associated with signal the transduction of chemotaxis and cell motility were identified. Genes related to DNA binding and repair were also identified, with some of them supporting a possible association of bacteria with macrophages or inducing pro-inflammatory responses. The homology-based identification of these genes suggests their potential involvement in the virulence and pathogenicity of L. intracellularis, opening avenues for future research and insights into the molecular mechanisms of Lawsonia-elicited proliferative enteropathy. [ABSTRACT FROM AUTHOR]

COVID-19 brought back to the attention of the scientific community that males are more susceptible to infectious diseases. What is clear for other infections—that sex and gender differences influence both risk of infection and mortality—is not yet fully elucidated for plague, particularly bubonic plague, although this knowledge can help find specific defences against a disease for which a vaccine is not yet available. To address this question, we analysed data on plague from hospitals in different parts of the world since the early eighteenth century, which provide demographic information on individual patients, diagnosis and course of the disease in the pre-antibiotic era. Assuming that the two sexes were equally represented, we observe a worldwide prevalence of male cases hospitalized at any age, a result which seems better explained by gender-biased (thus cultural) behaviours than biological sex-related factors. Conversely, case fatality rates differ among countries and geographic macro-areas, while globally, lethality appears slightly prevalent in young females and older adults (regardless of sex). Logistic regression models confirm that the main risk factor for bubonic plague death was the geographical location of the cases and being older than 50 years, whereas sex only showcased a slight trend. [ABSTRACT FROM AUTHOR]

Field-forward analytical technologies, such as portable mass spectrometry (MS), enable essential capabilities for real-time monitoring and point-of-care diagnostic applications. Significant and recent investments improving the features of miniaturized mass spectrometers enable various new applications outside of small molecule detection. Most notably, the addition of tandem mass spectrometry scans (MS/MS) allows the instrument to isolate and fragment ions and increase the analytical specificity by measuring unique chemical signatures for ions of interest. Notwithstanding these technological advancements, low-cost, portable systems still struggle to confidently identify clinically significant organisms of interest, such as bacteria, viruses, and proteinaceous toxins, due to the limitations in resolving power. To overcome these limitations, we developed a novel multidimensional mass fingerprinting technique that uses tandem mass spectrometry to increase the chemical specificity for low-resolution mass spectral profiles. We demonstrated the method's capabilities for differentiating four different bacteria, including attentuated strains of Yersinia pestis. This approach allowed for the accurate (>92%) identification of each organism at the strain level using de-resolved matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) data to mimic the performance characteristics of miniaturized mass spectrometers. This work demonstrates that low-resolution mass spectrometers, equipped with tandem MS acquisition modes, can accurately identify clinically relevant bacteria. These findings support the future application of these technologies for field-forward and point-of-care applications where high-performance mass spectrometers would be cost-prohibitive or otherwise impractical. [ABSTRACT FROM AUTHOR]

Background: Antibodies have been proven effective as diagnostic agents for detecting zoonotic diseases. The variable domain of camel heavy chain antibody (VHH), as an antibody derivative, may be used as an alternative for traditional antibodies in existing immunodiagnostic reagents for detecting rapidly spreading infectious diseases. Objectives: To expedite the isolation of specific antibodies for diagnostic purposes, we constructed a semi‐synthetic camel single domain antibody library based on the phage display technique platform (PDT) and verified the validity of this study. Methods: The semi‐synthetic single domain antibody sequences consist of two parts: one is the FR1‐FR3 region amplified by RT‐PCR from healthy camel peripheral blood lymphocytes (PBLs), and the other part is the CDR3‐FR4 region synthesised as an oligonucleotide containing CDR3 randomised region. The two parts were fused by overlapping PCR, resulting in the rearranged variable domain of heavy‐chain antibodies (VHHs). Y. pestis low‐calcium response V protein (LcrV) is an optional biomarker to detect the Y. pestis infection. The semi‐synthetic library herein was screened using recombinant (LcrV) as a target antigen. Results: After four cycles of panning the library, four VHH binders targeting 1–270 aa residues of LcrV were isolated. The four VHH genes with unique sequences were recloned into an expression vector and expressed as VHH‐hFc chimeric antibodies. The purified antibodies were identified and used to develop a lateral flow immunoassay (LFA) test strip using latex microspheres (LM) for the rapid and visual detection of Y. pestis infection. Conclusions: These data demonstrate the great potential of the semi‐synthetic library for use in isolation of antigen‐specific nanobodies and the isolated specific VHHs can be used in antigen‐capture immunoassays. [ABSTRACT FROM AUTHOR]

Background: Plague, a zoonotic disease caused by Yersinia pestis, was responsible for 3 historical human pandemics that killed millions of people. It remains endemic in rodent populations in Africa, Asia, North America, and South America but human plague is rare in most of these locations. However, human plague is still highly prevalent in Madagascar, which typically records a significant part of all annual global cases. This has afforded an opportunity to study contemporary human plague in detail using various typing methods for Y. pestis. Aim: This review aims to summarize the methods that have been used to type Y. pestis in Madagascar along with the major discoveries that have been made using these approaches. Methods: Pubmed and Google Scholar were used to search for the keywords: "typing Yersinia pestis Madagascar," "evolution Yersinia pestis Madagascar," and "diversity Yersinia pestis Madagascar." Eleven publications were relevant to our topic and further information was retrieved from references cited in those publications. Results: The history of Y. pestis typing in Madagascar can be divided in 2 periods: the pre-genomics and genomics eras. During the pre-genomics era, ribotyping, direct observation of plasmid content and plasmid restriction fragment length polymorphisms (RFLP) were employed but only revealed a limited amount of diversity among Malagasy Y. pestis strains. Extensive diversity only started to be revealed in the genomics era with the use of clustered regularly interspaced palindromic repeats (CRISPR), multiple-locus variable number tandem repeats (VNTR) analysis (MLVA), and single-nucleotide polymorphisms (SNPs) discovered from whole genome sequences. These higher-resolution genotyping methods have made it possible to highlight the distribution and persistence of genotypes in the different plague foci of Madagascar (Mahajanga and the Central and Northern Highlands) by genotyping strains from the same locations across years, to detect transfers between foci, to date the emergence of genotypes, and even to document the transmission of antimicrobial resistant (AMR) strains during a pneumonic plague outbreak. Despite these discoveries, there still remain topics that deserve to be explored, such as the contribution of horizontal gene transfer to the evolution of Malagasy Y. pestis strains and the evolutionary history of Y. pestis in Madagascar. Conclusions: Genotyping of Y. pestis has yielded important insights on plague in Madagascar, particularly since the advent of whole-genome sequencing (WGS). These include a better understanding of plague persistence in the environment, antimicrobial AMR and multi-drug resistance in Y. pestis, and the person-to-person spread of pneumonic plague. Considering that human plague is still a significant public health threat in Madagascar, these insights can be useful for controlling and preventing human plague in Madagascar and elsewhere, and also are relevant for understanding the historical pandemics and the possible use of Y. pestis as a biological weapon. Author summary: Plague remains a major public health concern in Madagascar and has been since its introduction to the country in 1898. The scale and rapid spread of the 2017 pneumonic plague epidemic in urban settings in Madagascar reminds us of the relevance of improving our understanding of this disease as well as its monitoring. In this review, we revisit the studies conducted in Madagascar that utilized genotyping approaches to understand the evolution, geographic distribution, and emergence of Y. pestis, the etiologic agent of plague. These studies greatly contributed to the comprehension of the origin, the transmission and the dynamics of the circulation of plague in Madagascar, providing useful information for preparing for future plague outbreaks in Madagascar and elsewhere. [ABSTRACT FROM AUTHOR]

Objective To provide a theoretical basis for optimizing and improving the culture medium of Yersinia pestis, the effects of different species and different concentrations of animal blood on the growth of Yersinia pestis were investigated. Methods Ten kinds of animal blood including rat blood, rabbit blood and guinea pig blood at four concentrations of 0. 1 %, 0. 5 %, 1 % and 2 % were used as blood stimulators to culture Yersinia pestis in the medium. The culture results were observed and the corresponding statistical analysis was carried out. Results The effect of 10 kinds of animal blood on the number of Yersinia pestis colonies at 1 % concentration was different (P = 0. 015 < 0. 05), and pig blood was better than chicken blood (P = 0. 026 < 0. 05). The effect of four concentrations of pig blood on the number of Yersinia pestis colonies was statistically significant (P = 0. 037 < 0. 05), and it was better than the blank at 1 % concentration (P = 0. 002 < 0. 05). The effects of ten kinds of animal blood at four concentrations on the length of Yersinia pestis were statistically significant (P < 0. 001). The difference in the effect of four concentrations of blood on the length of Yersinia pestis in ten animals was also statistically significant (P < 0. 001). Conclusion When observing the number of colonies, the optimal choice is 1% pig blood. In addition, low concentration and easy - to - obtain blood stimulants can be selected as far as possible. To cultivate a better length of Yersinia pestis, the optimal choice is 0. 1% rabbit blood, which can achieve the desired experimental results and save costs. In addition, using the Herxheimer s sensitive medium with only blood stimulator to culture Yersinia pestis, the characteristics of the strain will not undergo L 一 type transformation, and the overall characteristics of the strain are stable, which is of great significance for the stable passage and preservation of the strain. [ABSTRACT FROM AUTHOR]

Background: Yersinia pestis is the etiological agent of plague, which can manifest as bubonic, septicemic, and/or pneumonic disease. Plague is a severe and rapidly progressing illness that can only be successfully treated with antibiotics initiated early after infection. There are no FDA-approved vaccines for plague, and some vaccine candidates may be less effective against pneumonic plague than bubonic plague. Y. pestis is not known to impact males and females differently in mechanisms of pathogenesis or severity of infection. However, one previous study reported sex-biased vaccine effectiveness after intranasal Y. pestis challenge. As part of developing a safe and effective vaccine, it is essential that potential sex differences are characterized. Methods: In this study we evaluated novel vaccines in male and female BALB/c mice using a heterologous prime-boost approach and monitored survival, bacterial load in organs, and immunological correlates. Our vaccine strategy consisted of two subcutaneous immunizations, followed by challenge with aerosolized virulent nonencapsulated Y. pestis. Mice were immunized with a combination of live Y. pestis pgm- pPst-Δcaf1, live Y. pestis pgm- pPst-Δcaf1/ΔyopD, or recombinant F1-V (rF1-V) combined with adjuvants. Results: The most effective vaccine regimen was initial priming with rF1-V, followed by boost with either of the live attenuated strains. However, this and other strategies were more protective in female mice. Males had higher bacterial burden and differing patterns of cytokine expression and serum antibody titers. Male mice did not demonstrate synergy between vaccination and antibiotic treatment as repeatedly observed in female mice. Conclusions: This study provides new knowledge about heterologous vaccine strategies, sex differences in plague-vaccine efficacy, and the immunological factors that differ between male and female mice. [ABSTRACT FROM AUTHOR]

Background: Yersinia pestis is a bacterium that causes the disease plague. It has caused the deaths of many people throughout history. The bacterium possesses several virulence factors (pPla, pFra, and PYV). PFra plasmid encodes fraction 1 (F1) capsular antigen. F1 protein protects the bacterium against host immune cells through phagocytosis process. This protein is specific for Y. pestis. Many diagnostic techniques are based on molecular and serological detection and quantification of F1 protein in different food and clinical samples. Aptamers are small nucleic acid sequences that can act as specific ligands for many targets.This study, aimed to isolate the high-affinity ssDNA aptamers against F1 protein. Methods and results: In this study, SELEX was used as the main strategy in screening aptamers. Moreover, enzyme-linked aptamer sorbent assay (ELASA) and surface plasmon resonance (SPR) were used to determine the affinity and specificity of obtained aptamers to F1 protein. The analysis showed that among the obtained aptamers, the three aptamers of Yer 21, Yer 24, and Yer 25 were selected with a KD value of 1.344E − 7, 2.004E − 8, and 1.68E – 8 M, respectively. The limit of detection (LoD) was found to be 0.05, 0.076, and 0.033 μg/ml for Yer 21, Yer 24, and Yer 25, respectively. Conclusion: This study demonstrated that the synthesized aptamers could serve as effective tools for detecting and analyzing the F1 protein, indicating their potential value in future diagnostic applications. [ABSTRACT FROM AUTHOR]

The aim of this work was to determine the role of periplasmic chaperone SurA in the pathogenesis of bubonic and pneumonic plague. Yersinia pestis surA knockout mutants were generated by RedGam mutagenesis with the use of a suicide vector. The generated strains were characterized by growth rate and susceptibility to antibiotics, bile acid salts, and sodium dodecyl sulfate, as well as to the bactericidal action of normal human serum. The intracellular localization of the SurA protein was determined by SDS-PAGE and immunoblot methods. The virulence of the ΔsurA mutant was studied in subcutaneously or intranasally infected mice or rats. The SurA-negative Y. pestis strain was characterized by an increase in the permeability of the outer membrane and susceptibility to antibiotics, as well as a decrease in virulence in a model of bubonic and pneumonic plague in two laboratory animal species. The SurA protein can be considered as a molecular target for plague therapy. Reducing the virulence of a strain and its resistance to antibiotics by disrupting the biogenesis of outer-membrane proteins may become a new strategy for solving the problem of combating multidrug resistance, and the widespread distribution of the protein among Gram-negative pathogens makes its use promising for the development of virulence inhibitors with a broad spectrum of action. [ABSTRACT FROM AUTHOR]

ABSTRACT Yersinia pestis, one of the deadliest bacterial pathogens ever known, is responsible for three plague pandemics and several epidemics, with over 200 million deaths during recorded history. Due to high genomic plasticity, Y. pestis is amenable to genetic mutations as well as genetic engineering that can lead to the emergence or intentional development of pan-drug-resistant strains. Indeed, antibiotic-resistant strains (e.g., strains carrying multidrug-resistant or MDR plasmids) have been isolated in various countries and endemic areas. Thus, there is an urgent need to develop novel, safe, and effective treatment approaches for managing Y. pestis infections. This includes infections by antigenically distinct strains for which vaccines (none FDA approved yet) may not be effective and those that cannot be managed by currently available antibiotics. Lytic bacteriophages provide one such alternative approach. In this study, we examined post-exposure efficacy of a bacteriophage cocktail, YPP-401, to combat pneumonic plague caused by Y. pestis CO92. YPP-401 is a four-phage preparation effective against a panel of at least 68 genetically diverse Y. pestis strains. Using a pneumonic plague aerosol challenge model in gender-balanced Brown Norway rats, YPP-401 demonstrated ~88% protection when delivered 18 h post-exposure for each of two administration routes (i.e., intraperitoneal and intranasal) in a dose-dependent manner. Our studies provide proof-of-concept that YPP-401 could be an innovative, safe, and effective approach for managing Y. pestis infections, including those caused by naturally occurring or intentionally developed multidrug-resistant strains.IMPORTANCECurrently, there are no FDA-approved plague vaccines. Since antibiotic-resistant strains of Y. pestis have emerged or are being intentionally developed to be used as a biothreat agent, new treatment modalities are direly needed. Phage therapy provides a viable option against potentially antibiotic-resistant strains. Additionally, phages are nontoxic and have been approved by the FDA for use in the food industry. Our study provides the first evidence of the protective effect of a cocktail of four phages against pneumonic plague, the most severe form of disease. When treatment was initiated 18 h post infection by either the intranasal or intraperitoneal route in Brown Norway rats, up to 87.5% protection was observed. The phage cocktail had a minimal impact on a representative human microbiome panel, unlike antibiotics. This study provides strong proof-of-concept data for the further development of phage-based therapy to treat plague.

ABSTRACT Outer membrane vesicles (OMVs) from Gram-negative bacteria can be used as a vaccine platform to deliver heterologous antigens. Here, the major protective antigens of Yersinia pestis, F1 and LcrV, were fused either with the leader sequence or the transmembrane domain of the outer membrane protein A (OmpA), resulting in chimeric proteins OmpA-ls-F1V and OmpA46-159-F1V, respectively. We show that OmpA-ls-F1V and OmpA46-159-F1V can be successfully delivered into the lumen and membrane of the OMVs of Escherichia coli, respectively. Mutation of ompA but not tolR in E. coli enhanced the delivery efficiency of OmpA-ls-F1V into OMVs. The OmpA-ls-F1V protein comprises up to 20% of the total protein in OMVs derived from the ompA mutant (OMVdA-ALS-F1V), a proportion significantly higher than the 1% observed for OmpA46-159-F1V in OMVs produced by an ompA mutant that expresses OmpA46-159-F1V, referred to as OMVdA-LATM5-F1V. Intramuscular (i.m.) immunization of mice with OMVdA-ALS-F1V induced significantly higher levels of serum anti-LcrV and anti-F1 IgG, and provided higher efficacy in protection against subcutaneous (s.c.) Y. pestis infection compared to OMVdA-LATM5-F1V and the purified recombinant F1V (rF1V) protein adsorbed to aluminum hydroxide. The three-dose i.m. immunization with OMVdA-ALS-F1V, administered at 14-day intervals, provides complete protection to mice against s.c. infection with 130 LD50 of Y. pestis 201 and conferred 80% against intranasal (i.n.) challenge with 11.4 LD50 of Y. pestis 201. Taken together, our findings indicate that the engineered OMVs containing F1V fused with the leader sequence of OmpA provide significantly higher protection than rF1V against both s.c. and i.n. infection of Y. pestis and more balanced Th1/Th2 responses.IMPORTANCEThe two major protective antigens of Y. pestis, LcrV and F1, have demonstrated the ability to elicit systemic and local mucosal immune responses as subunit vaccines. However, these vaccines have failed to provide adequate protection against pneumonic plague in African green monkeys. Here, Y. pestis F1 and LcrV antigens were successfully incorporated into the lumen and the surface of the outer membrane vesicles (OMVs) of E. coli by fusion either with the leader sequence or the transmembrane domain of OmpA. We compared the humoral immune response elicited by these OMV formulations and their protective efficacy in mice against Y. pestis. Our results demonstrate that the plague OMV vaccine candidates can induce robust protective immunity against both s.c. and i.n. Y. pestis infections, surpassing the effectiveness of rF1V. In addition, immunization with OMVs generated a relatively balanced Th1/Th2 immune response compared to rF1V immunization. These findings underscore the potential of OMVs-based plague vaccines for further development.

BackgroundThe virulence of Yersinia pestis strains in the Rattus flavipectus plague focus is relatively low. The purpose of this study was to investigate the undetected, sporadic plague cases in plague foci and provide the basis for plague prevention and control.MethodsA 3-year-old plague-confirmed case was investigated in the R. flavipectus plague focus of Yunnan Province in 2020 due to the intensive screening for fever symptoms during the coronavirus disease 2019 (COVID-19) pandemic. Epidemiological investigation, laboratory testing, and clinical treatment were conducted for the case. The expanded survey was carried out around the case within a 7-km radius, including the resident population, domesticated dogs, and rats. PCR and indirect hemagglutination tests were performed on the collected samples.ResultsThe isolation rates of Y. pestis were 100.0% (7 out of 7) in dead rats and 4.00% (3 out of 75) in live rats in the survey area of the foci. A total of 5.00% (6 out of 120) of the domesticated dogs were F1 antibody positive. Nine local people were determined for plague infection recently (0.92%, 9 out of 978). The locations of human cases coincided with the Y. pestis epidemic area among the animals.ConclusionThis study discovered the existence of plague cases that had not been detected by routine surveillance in the R. flavipectus plague focus, and the actual epidemic of human infection may be underestimated.

Abstract Plague, as an ancient zoonotic disease caused by Yersinia pestis, has brought great disasters. The natural plague focus of Marmota himalayana in the Qinghai-Tibet Plateau is the largest, which has been constantly active and the leading source of human plague in China for decades. Understanding the population genetics of M. himalayana and relating that information to the biogeographic distribution of Yersinia pestis and plague outbreaks are greatly beneficial for the knowledge of plague spillover and arecrucial for pandemic prevention. In the present research, we assessed the population genetics of M. himalayana. We carried out a comparative study of plague outbreaks and the population genetics of M. himalayana on the Qinghai-Tibet Plateau. We found that M. himalayana populations are divided into two main clusters located in the south and north of the Qinghai-Tibet Plateau. Fourteen DFR genomovars of Y. pestis were found and exhibited a significant region-specific distribution. Additionally, the increased genetic diversity of plague hosts is positively associated with human plague outbreaks. This insight gained can improve our understanding of biodiversity for pathogen spillover and provide municipally directed targets for One Health surveillance development, which will be an informative next step toward increased monitoring of M. himalayana dynamics.

Yersinia pestis, the causative agent of plague, is a highly lethal vector-borne pathogen responsible for killing large portions of Europe's population during the Black Death of the Middle Ages. In the wild, Y. pestis cycles between fleas and rodents; occasionally spilling over into humans bitten by infectious fleas. For this reason, fleas and the rats harboring them have been considered the main epidemiological drivers of previous plague pandemics. Human ectoparasites, such as the body louse (Pediculus humanus humanus), have largely been discounted due to their reputation as inefficient vectors of plague bacilli. Using a membrane-feeder adapted strain of body lice, we show that the digestive tract of some body lice become chronically infected with Y. pestis at bacteremia as low as 1 × 105 CFU/ml, and these lice routinely defecate Y. pestis. At higher bacteremia (≥1 × 107 CFU/ml), a subset of the lice develop an infection within the Pawlowsky glands (PGs), a pair of putative accessory salivary glands in the louse head. Lice that developed PG infection transmitted Y. pestis more consistently than those with bacteria only in the digestive tract. These glands are thought to secrete lubricant onto the mouthparts, and we hypothesize that when infected, their secretions contaminate the mouthparts prior to feeding, resulting in bite-based transmission of Y. pestis. The body louse's high level of susceptibility to infection by gram-negative bacteria and their potential to transmit plague bacilli by multiple mechanisms supports the hypothesis that they may have played a role in previous human plague pandemics and local outbreaks. Yersinia pestis, the causative agent of plague, uses fleas and rodents as vectors for transmission. This study reports a new vector for Y. pestis, showing that it can infect the Pawlowsky glands of human body lice and be transmitted by louse bite. [ABSTRACT FROM AUTHOR]

Background: Plague, caused by the bacterium Yersinia pestis, is a zoonotic disease that poses considerable threats to human health. Nucleic acid tests are crucial for plague surveillance and the rapid detection of Y. pestis. However, inhibitors in complex samples such as soil and animal tissues often hamper nucleic acid detection, leading to a reduced rate of identifying low concentrations of Y. pestis. To address this challenge, we developed a sensitive and specific droplet digital polymerase chain reaction (ddPCR) assay for detecting Y. pestis DNA from soil and animal tissue samples. Methods: Three genes (ypo2088, caf1, and pla) from Y. pestis were used to develop a multi-target ddPCR assay. The limits of detection (LoD), reproducibility, and specificity were assessed for bacterial genomic DNA samples. The ability of the assay to detect low concentrations of Y. pestis DNA from simulated soil and mouse liver tissue samples was respectively evaluated and compared with that of quantitative real-time PCR (qPCR). Results: The results showed that the ddPCR LoDs ranged from 6.2 to 15.4 copies/reaction for the target genes, with good reproducibility and high specificity for Y. pestis. By testing 130 soil and mouse liver tissue samples spiked with Y. pestis, the ddPCR assay exhibited a better sensitivity than that of the qPCR assay used in the study, with LoDs of 102 colony forming units (CFU)/100 mg soil and 103 CFU/20 mg liver. Moreover, the assay presented good quantitative linearity (R2 = 0.99) for Y. pestis at 103–106 CFU/sample for soil and liver samples. Conclusion: The ddPCR assay presented good performance for detecting Y. pestis DNA from soil and mouse tissue samples, showing great potential for improving the detection rate of low concentrations of Y. pestis in plague surveillance and facilitating the early diagnosis of plague cases. Author summary: Yersinia pestis is the causative agent of three historic plague pandemics. Presently, the plague remains endemic or sporadic in many countries. The wide distribution of natural plague foci in Asia, Africa, and the Americas reminds us of the threat of a plague outbreak. The timely detection and surveillance of Y. pestis prevalence in animals and the environment in natural epidemic foci is essential for preventing and controlling plague outbreaks. In this study, we developed a sensitive and specific ddPCR assay for detecting low concentrations of Y. pestis DNA from soil and mouse tissue samples. The ddPCR assay presented a considerable improvement in sensitivity compared to the qPCR assay used in the study, thus providing a promising tool for detecting Y. pestis DNA from complex samples and facilitating the accurate diagnosis of plague cases. [ABSTRACT FROM AUTHOR]

Plague is an endemic infectious disease caused by Yersinia pestis. In this study, we isolated fourteen phages with similar sequence arrangements to phage 186; these phages exhibited different lytic abilities in Enterobacteriaceae strains. To illustrate the phylogenetic relationships and evolutionary relationships between previously designated 186-type phages, we analysed the complete sequences and important genes of the phages, including whole-genome average nucleotide identity (ANI) and collinearity comparison, evolutionary analysis of four conserved structural genes (V, T, R, and Q genes), and analysis of the regulatory genes (cI, apl, and cII) and integrase gene (int). Phylogenetic analysis revealed that thirteen of the newly isolated phages belong to the genus Eganvirus and one belongs to the genus Felsduovirus in the family Peduoviridae, and these Eganvirus phages can be roughly clustered into three subgroups. The topological relationships exhibited by the whole-genome and structural genes seemed similar and stable, while the regulatory genes presented different topological relationships with the structural genes, and these results indicated that there was some homologous recombination in the regulatory genes. These newly isolated 186-type phages were mostly isolated from dogs, suggesting that the resistance of Canidae to Y. pestis infection may be related to the wide distribution of phages with lytic capability. [ABSTRACT FROM AUTHOR]

This study evaluated a depot-formulated cytokine-based adjuvant to improve the efficacy of the recombinant F1V (rF1V) plague vaccine and examined the protective response following aerosol challenge in a murine model. The results of this study showed that co-formulation of the Alhydrogel-adsorbed rF1V plague fusion vaccine with the depot-formulated cytokines recombinant human interleukin 2 (rhuIL-2) and/or recombinant murine granulocyte macrophage colony-stimulating factor (rmGM-CSF) significantly enhances immunogenicity and significant protection at lower antigen doses against a lethal aerosol challenge. These results provide additional support for the coapplication of the depot-formulated IL-2 and/or GM-CSF cytokines to enhance vaccine efficacy. [ABSTRACT FROM AUTHOR]

Plague is a flea-borne fatal disease caused by the bacterium Yersinia pestis, which persists in rural Madagascar. Although fleas parasitizing rats are considered the primary vectors of Y. pestis, the human flea, Pulex irritans, is abundant in human habitations in Madagascar, and has been found naturally infected by the plague bacterium during outbreaks. While P. irritans may therefore play a role in plague transmission if present in plague endemic areas, the factors associated with infestation and human exposure within such regions are little explored. To determine the socio-ecological risk factors associated with P. irritans infestation in rural households in plague-endemic areas of Madagascar, we used a mixed-methods approach, integrating results from P. irritans sampling, a household survey instrument, and an observational checklist. Using previously published vectorial capacity data, the minimal P. irritans index required for interhuman bubonic plague transmission was modeled to determine whether household infestations were enough to pose a plague transmission risk. Socio-ecological risk factors associated with a high P. irritans index were then identified for enrolled households using generalized linear models. Household flea abundance was also modeled using the same set of predictors. A high P. irritans index occurred in approximately one third of households and was primarily associated with having a traditional dirt floor covered with a plant fiber mat. Interventions targeting home improvement and livestock housing management may alleviate flea abundance and plague risk in rural villages experiencing high P. irritans infestation. As plague-control resources are limited in developing countries such as Madagascar, identifying the household parameters and human behaviors favoring flea abundance, such as those identified in this study, are key to developing preventive measures that can be implemented at the community level. Author summary: Plague is a bacterial disease transmitted by flea bites, and the rat fleas are the main vectors of Yersinia pestis, the plague bacterium. Households in plague endemic-areas of Madagascar are frequently infested by Pulex irritans, the human flea, which does become naturally infected with the plague bacterium during epidemic. The intensity of flea infestation varies among households, but the reasons for such disparities are poorly understood. This study identifies factors associated with P. irritans infestation in rural households in plague-endemic areas of Madagascar. Infestation risk was more pronounced for poor households living in homes made with organic construction materials and flea density did not show a seasonal pattern. One third of the household experienced high flea infestation, putting inhabitants at risk of sustained interhuman plague transmission, should the fleas or a household member become infected. While P. irritans may be a secondary vector, this additional route of plague transmission deserves more attention from epidemiologists. The factors identified in this analysis suggest that improvement of housing and better management of livestock would alleviate flea burden and potential plague risk in rural plague-endemic villages experiencing high flea infestation. [ABSTRACT FROM AUTHOR]

Plague, as an ancient zoonotic disease caused by Yersinia pestis, has brought great disasters. The natural plague focus of Marmota himalayana in the Qinghai-Tibet Plateau is the largest, which has been constantly active and the leading source of human plague in China for decades. Understanding the population genetics of M. himalayana and relating that information to the biogeographic distribution of Yersinia pestis and plague outbreaks are greatly beneficial for the knowledge of plague spillover and arecrucial for pandemic prevention. In the present research, we assessed the population genetics of M. himalayana. We carried out a comparative study of plague outbreaks and the population genetics of M. himalayana on the Qinghai-Tibet Plateau. We found that M. himalayana populations are divided into two main clusters located in the south and north of the Qinghai-Tibet Plateau. Fourteen DFR genomovars of Y. pestis were found and exhibited a significant region-specific distribution. Additionally, the increased genetic diversity of plague hosts is positively associated with human plague outbreaks. This insight gained can improve our understanding of biodiversity for pathogen spillover and provide municipally directed targets for One Health surveillance development, which will be an informative next step toward increased monitoring of M. himalayana dynamics. [ABSTRACT FROM AUTHOR]

The Serratia marcescens is a Gram-negative bacterium from the Enterobacteriaceae family. Recently, S. marcescens have evolved to become a versatile and opportunistic pathogen. Furthermore, this bacterium is also a multi-drug resistant pathogen exhibiting Extended-Spectrum Beta-Lactamases (ESBL) activity. This bacterium is highly associated with infections in healthcare settings and even leads to death. Hence, an advanced approach based on non-protein coding RNA (npcRNA) of S. marcescens was considered in this study to understand its regulatory roles in virulence, pathogenesis, and the differential expression of these transcripts in various growth phases of the bacterium. BLASTn search of known npcRNAs from Salmonella typhi, Escherichia coli, and Yersinia pestis against S. marcescens was performed to discover putative conserved homologous transcripts. The novelty of these putative homologous npcRNAs was verified by screening through the Rfam web tool. The target mRNA for the homologs was predicted via the TargetRNA2 webtool to understand the possible regulatory roles of these transcripts. The npcRNA homologs, which were predicted to regulate virulence target mRNA were assessed for their expression profile at different growth stages via reverse transcription PCR and the band intensity was quantitatively analysed using the Image J tool. The known npcRNA ssrS, from S. typhi showed expression in S. marcescens during three growth stages (lag, log, and stationary). Expression was observed to be high during the lag phase followed by a similarly low-level expression during the log and no expression during stationary phase. This ssrS homolog was predicted to regulate mRNA that encodes for protein FliR, which is associated with virulence. This is a preliminary study that lay the foundation for further elucidation of more virulence-associated npcRNAs that are yet to be discovered from S. marcescens, which can be useful for diagnostics and therapeutic applications. [ABSTRACT FROM AUTHOR]