Plague Strains Research Articles

Pathogenicity studies and molecular characterization of DPV infection in ducklings

In the spring of 2023, 10 to 21-day-old chicks in a broiler duck farm in Shandong Province, China, developed swelling of the head and neck, moist eyes with mucous discharge, difficulty in walking, shrinking of the neck, and loose and disorganized coat. Anatomical observation revealed hemorrhages in the esophageal mucosa, myocardium, and liver, and severe hemorrhages in the trachea with copious inflammatory secretions. Soon after, similar symptoms appeared in a large number of ducks in the flock, which eventually led to the elimination of all the 20,000-odd newly introduced ducklings on the farm, resulting in huge economic losses. We detected duck plague virus in the tissues of liver, spleen and lungs of diseased and dead ducks, and successfully isolated the pathogenic strain, named SD423, by inoculating duck embryos and inoculating duck embryo fibroblasts. We successfully conducted animal regression experiments with the isolated strain, and the experimental animals in the 1 d of age group showed symptoms of swollen eyes and tearing, shrinking of the neck, crouching, and hemorrhage in organs such as the liver and intestines successively from the 3rd d. We sequenced the whole genome of the isolated duck plague strain, and by comparing the homology with the published duck plague virus whole sequences in Genbank, the virus strain obtained in this study had the highest homology with the Chinese virulent strain SD (MN518864.1), with nucleotide (nt) homology of about 99.90% and amino acid (aa) homology of about 99.75%, which indicated that the isolate is a virulent strain. Previously, it was reported that the natural infection of duck plague virus mainly occurs above 30 d of age, but the duck plague virus found in this study can naturally infect ducklings up to 20 d of age, and the mortality rate is as high as 100%. In this study, the pathogenicity test and whole genome sequence analysis of this isolate provided data support and theoretical basis for further research on pathogenicity and virulence-related gene analysis of duck plague virus.

The Influence of the Synthetic Organoselenium Preparation 974zh on the Immunogenic Activity of Yersinia pestis EV Vaccine Strain NIIEG.

In the present study, a stimulating effect of a new synthetic organoselenium compound 2,6-dipyridinium-9-selenabicyclo[3.3.1]nonandibromide (974zh) on the immunogenic activity of the vaccine strain Yersinia pestis EV NIIEG was revealed. After infection with the virulent plague strain, the survival rate of laboratory mice immunized with the vaccine strain grown on Hottinger's agar in the presence of 974zh (300 μg/ml) increased in comparison with control animals immunized with the Y. pestis EV NIIEG culture grown on agar without the studied compound. Plasmid screening of cultures grown on medium with and without 974zh showed that plasmid DNA of Y. pestis EV culture grown in the presence of 974zh had broader bands in the control grown without 974zh. This phenomenon can indicate activation of replication of plasmid DNA of Y. pestis EV under the influence of the experimental compound.

Spatiotemporal Variations of Plague Risk in the Tibetan Plateau from 1954-2016.

Simple SummaryClimate variability has influence on plague outbreaks worldwide. Usually, plague cases increase with increasing precipitation. Currently there are many studies on the epidemics of plague in human beings, whereas there are few studies on the dynamic of plague in animal. Nevertheless, animal plague is key in the natural epidemiological cycle of plague. We identified spatiotemporal changes of the plague territories in the Tibetan Plateau only using animal plague records. Our risky plague maps are far superior to the county-based maps used currently and have valuable applications for directly informing conservation and management decisions locally and regionally.Plague persists in the plague natural foci today. Although previous studies have found climate drives plague dynamics, quantitative analysis on animal plague risk under climate change remains understudied. Here, we analyzed plague dynamics in the Tibetan Plateau (TP) which is a climate-sensitive area and one of the most severe animal plague areas in China to disentangle variations in marmot plague enzootic foci, diffusion patterns, and their possible links with climate and anthropogenic factors. Specifically, we developed a time-sharing ecological niche modelling framework to identify finer potential plague territories and their temporal epidemic trends. Models were conducted by assembling animal records and multi-source ecophysiological variables with actual ecological effects (both climatic predictors and landscape factors) and driven by matching plague strains to periods corresponding to meteorological datasets. The models identified abundant animal plague territories over the TP and suggested the spatial patterns varied spatiotemporal dimension across the years, undergoing repeated spreading and contractions. Plague risk increased in the 1980s and 2000s, with the risk area increasing by 17.7 and 55.5 thousand km2, respectively. The 1990s and 2010s were decades of decreased risk, with reductions of 71.9 and 39.5 thousand km2, respectively. Further factor analysis showed that intrinsic conditions (i.e., elevation, soil, and geochemical landscape) provided fundamental niches. In contrast, climatic conditions, especially precipitation, led to niche differentiation and resulted in varied spatial patterns. Additionally, while increased human interference may temporarily reduce plague risks, there is a strong possibility of recurrence. This study reshaped the plague distribution at multiple time scales in the TP and revealed multifactorial synergistic effects on the spreading and contraction of plague foci, confirming that TP plague is increasingly sensitive to climate change. These findings may facilitate groups to take measures to combat the plague threats and prevent potential future human plague from occurring.

Physiological performance of native and invasive crayfish species in a changing environment: insights from Dynamic Energy Budget models.

Crayfish are keystone species important for maintaining healthy freshwater ecosystems. Crayfish species native to Europe, such as Astacus astacus and Austropotamobius torrentium, are facing decline and are increasingly endangered by changing climate and invasions of non-native crayfish, such as Pacifastacus leniusculus and Procambarus virginalis. The success of these invasions largely depends on differences in ontogeny between the native species and the invaders and how changes in the environment will affect the ontogeny. Dynamic Energy Budget (DEB) models can be used to investigate such differences because the models capture dependence of metabolism, and therefore ontogeny, on environmental conditions. We develop DEB models for all four species and investigate key elements of ontogeny and metabolism affecting interspecific competition. We then use the DEB models to predict individual growth and reproduction in current and new conditions that are expected to arise from climate change. Although observations suggest that P. leniusculus poses the major threat to native species, our analysis identifies P. virginalis, in spite of its smaller size, as the superior competitor by a large margin—at least when considering metabolism and ontogeny. Our simulations show that climate change is set to increase the competitive edge of P. virginalis even further. Given the prospects of P. virginalis dominance, especially when considering that it is able to withstand and spread at least some crayfish plague strains that severely affect native species, additional research into P. virginalis is necessary.

Конструирование системы мультиплексных ПЦР с гибридизационно-флуоресцентным учетом результатов на твердой подложке для индикации и идентификации штаммов возбудителя чумы

Plague agent strains differ in intraspecific appurtenance, place of origin, as well as their virulence. Development of a rapid and reliable means of Y. pestis strain indication, their identification by intra-specific appurtenance and assessment of their virulence is a relevant issue. To solve the task, we have constructed a system of multiplex PCR with hybridization-fluorescent registration of results on solid substrate, for indication and identification of strains by their belonging to Y. pestis species, sub-species, biovars, phylogenetic lines, also by the presence of the main gene-factors of pathogenicity of chromosomal and plasmid localization. Its effectiveness is confirmed on 114 plague agent strains of various origins. The developed system of multiplex PCR with hybridization-fluorescent registration of results on solid substrate, can be applied for molecular-genetic identification of strains from natural plague foci with specification of their appurtenance to the main phylogenetic branches and assessment of virulence, for enhancement of sanitary-epidemiological control of RF territories and CIS member-states. plague pathogen, strains, indication and identification, PCR

Marmots and Yersinia pestis Strains in Two Plague Endemic Areas of Tien Shan Mountains.

The main purpose of this study was to clarify the role of gray marmots (Marmota baibacina) in the long-term maintenance of highly virulent strains of Yersinia pestis in two plague endemic foci of the Tien Shan Mountains in Kyrgyzstan. We present data from regular observations of populations of M. baibacina and small rodents cohabiting with marmots in the mountainous grasslands of the Sari-Dzhas (east of Issyk-Kul Lake) and the Upper-Naryn (south of Issyk-Kul Lake) natural foci. During 2012–2017, an abundance of marmots and their ectoparasites (fleas and ticks) was significantly higher in Upper-Naryn comparing to Sari-Dzhas, although there were no differences in a number and diversity of small rodents cohabiting with marmots. The plague bacterium was detected either in marmots or in their ectoparasites collected during 4 of 6 years of observation in Sari-Dzhas and during 2 of 4 years of observation in Upper-Naryn. Plague was found in three sectors situated closely to each other in Sari-Dzhas and in 1 of 8 repeatedly surveyed sectors in Upper-Naryn. During 6 years, we isolated 9 strains of Y. pestis from marmots, two from their fleas Oropsylla silantiewi, one from an unidentified tick, and one from the gray hamster (Cricetulus migratorius). All plague strains isolated from the rodents and their ectoparasites in this study were similar to Antiqua biovar specific for marmots. The results indicate that plague can circulate continuously in the Tien Shan Mountains in populations of gray marmots and their ectoparasites with a facultative involvement of other rodent species after significant changes in rodent communities that happened in Kyrgyzstan during the previous two decades. The simultaneous field survey of two natural foci of plague, Sari-Dzhas, and Upper-Naryn, would be important for further analysis of circulation of Y. pestis strains belonging to Antiqua biovar in the Tien Shan Mountains.

The rise of pneumonic plague in Madagascar: current plague outbreak breaks usual seasonal mould.

Madagascar has just emerged from the grip of an acute urban pneumonic plague outbreak, which began in August 2017, before the usual plague season of October-April and outside the traditional plague foci in the northern and central highlands. The World Health Organization reported a total of 2417 confirmed, probable and suspected cases, including 209 deaths between 1 August and 26 November 2017. The severity and scope of this outbreak, which has affected those in higher socioeconomic groups as well as those living in poverty, along with factors including the potential for use of multi-drug-resistant strains of plague in bioterrorism, highlights the ongoing threat posed by this ancient disease. Factors likely to have contributed to transmission include human behaviour, including burial practices and movement of people, poor urban planning leading to overcrowding and ready transmission by airborne droplets, climatic factors and genomic subtypes. The outbreak demonstrates the importance of identifying targeted pneumonic plague therapies and of developing vaccines that can be administered in planned programmes in developing countries such as Madagascar where plague is endemic. The dominance of pneumonic plague in this outbreak suggests that we need to focus more urgently on the danger of person-to-person transmission, as well as the problem of transmission of plague from zoonotic sources.

Bubonic Panic: When Plague Invaded America by G. Jarrow

Jarrow, Gail. Bubonic Panic: When Plague Invaded America. Calkins Creek, 2015.If public health seems like it would be one of those topics that would send you to sleep, then Bubonic Plague: When Plague Invaded America by Gail Jarrow will change your mind. This is the final book in her trilogy about Deadly Diseases for middle grades and higher.Jarrow is fairly succinct in presenting the history, transmission, and trajectory of various waves of plague around the world. She briefly charts its first appearance in 541 in Turkey, then vividly describes the second wave that started in 1346 and at its most virulent was named the Black Death, killing millions in Europe and parts of Asia and North Africa. The majority of the book focuses on the third wave, when it reached North America.The third pandemic began in the mid-1800s when China became ground zero for this next wave, which spread to Hong Kong by 1894. Hong Kong was a busy port town and trade and travel on steamships allowed for rapid dispersion of the disease. Researchers from a number of countries sought feverishly to identify the source of the epidemic and learn how it was spread. By the late 1890s two of them had proven it was rat fleas. Unfortunately almost nobody believed them, which became problematic when in 1900 San Francisco saw its first deaths in Chinatown.Jarrow provides a fascinating look at the political and social climate of this period in relation to the attitudes of Americans towards Chinese immigrants and the impact quarantining San Francisco’s Chinatown would have on businesses reliant on trade and tourism. It became a complicated and fraught tug-o-war between politicians, businessmen, doctors and public health officials, fighting about whether to recognize and publicize the deaths and quarantine when the evidence seemed inconclusive as to their cause. Even after proof was offered action was surprisingly slow to follow and the disease was able to spread, although the number of deaths was comparatively low, being in the low hundreds.This well researched book also includes information about contemporary cases in the United States, ongoing research and treatments for all three strains of plague. There are extensive source notes and bibliography, a glossary, timeline, index and an author’s note explaining her keen interest in public health and the importance it had in the past,and will have when the next global pandemic hits. Also included are numerous photographs (some a little gruesome), newspaper clippings, cartoons, posters and illustrations to engage readers’ interest.This will pair perfectly with a middle grade novel, Chasing Secrets by Gennifer Choldenko, 2015 that gives a fictional account of the outbreak in San Francisco.This is a strong finish to a fascinating series that combines history, social issues, scientific research, technological developments and culture in America, showing long term implications for today’s government policies towards health.Highly Recommended: 4 out of 4 starsReviewer: Tammy FlandersTammy is the Reference Coordinator in the Doucette Library of Teaching Resources at the University of Calgary. She also reviews juvenile resources with an eye to classroom use in her blog, Apples with Many Seeds.

Analysis of complete sequence of cryptic plasmid pTP33 from Yersinia pestis isolated in Tuva natural focus of plague

This paper studies a full nucleotide sequence of cryptic plasmid pTP33, which was isolated from the typical plague strain of the Tuvinian natural focus, Yersinia pestis I-2638. Sequencing was carried out using the 454 GS Junior platform (Roche). In analysis using the software package GS De Novo Assembler v. 2.7 (Roche) and the algorithm Newbler v. 2.7, 1855 nucleotide reads, which contained 1101246 nucleotides, were assembled to a contig of 33 978 bp. The GC content of the obtained nucleotide sequence was 50.25%. During annotation, we found 56 open reading frames. Homologs of the predicted reading frames were sought in the BLAST databases. We detected 22 reading frames coding hypothetical proteins, 23 frames coding phagerelated proteins, and 11 frames coding proteins with known functions, including toxin–antitoxin system YefM-YoeB, nucleic acids and polysaccharides metabolism proteins (exopolysaccharide production protein ExoZ, exodeoxyribonuclease VIII), and replication proteins (ParA). Some predicted pTP33 proteins were found to be homologs (from 45 to 75%) with sequences of phage-related proteins of certain microorganisms—endosymbionts of insects (Sodalis glossinidius) and endosymbionts of entomopathogenic nematodes (Photorhabdus luminescens, P. asymbiotica, Xenorhabdus bovienii).

Early Divergent Strains of Yersinia pestis in Eurasia 5,000 Years Ago

SummaryThe bacteria Yersinia pestis is the etiological agent of plague and has caused human pandemics with millions of deaths in historic times. How and when it originated remains contentious. Here, we report the oldest direct evidence of Yersinia pestis identified by ancient DNA in human teeth from Asia and Europe dating from 2,800 to 5,000 years ago. By sequencing the genomes, we find that these ancient plague strains are basal to all known Yersinia pestis. We find the origins of the Yersinia pestis lineage to be at least two times older than previous estimates. We also identify a temporal sequence of genetic changes that lead to increased virulence and the emergence of the bubonic plague. Our results show that plague infection was endemic in the human populations of Eurasia at least 3,000 years before any historical recordings of pandemics.

Inhibition of Yersinia pestis DNA adenine methyltransferase in vitro by a stibonic acid compound: identification of a potential novel class of antimicrobial agents.

Multiple antibiotic resistant strains of plague are emerging, driving a need for the development of novel antibiotics effective against Yersinia pestis. DNA adenine methylation regulates numerous fundamental processes in bacteria and alteration of DNA adenine methlytransferase (Dam) expression is attenuating for several pathogens, including Y. pestis. The lack of a functionally similar enzyme in humans makes Dam a suitable target for development of novel therapeutics for plague. Compounds were evaluated for their ability to inhibit Dam activity in a high-throughput screening assay. DNA was isolated from Yersinia grown in the presence of lead compounds and restricted to determine the effect of inhibitors on DNA methylation. Transcriptional analysis was undertaken to determine the effect of an active inhibitor on virulence-associated phenotypes. We have identified a series of aryl stibonic acids which inhibit Dam in vitro. The most active, 4-stibonobenzenesulfonic acid, exhibited a competitive mode of inhibition with respect to DNA and a K(i) of 6.46 nM. One compound was found to inhibit DNA methylation in cultured Y. pestis. The effects of this inhibition on the physiology of the cell were widespread, and included altered expression of known virulence traits, including iron acquisition and Type III secretion. We have identified a novel class of potent Dam inhibitors. Treatment of bacterial cell cultures with these inhibitors resulted in a decrease in DNA methylation. Expression of virulence factors was affected, suggesting these inhibitors may attenuate bacterial infectivity and function as antibiotics.

Yersinia pestis Lineages in Mongolia

BackgroundWhole genome sequencing allowed the development of a number of high resolution sequence based typing tools for Yersinia (Y.) pestis. The application of these methods on isolates from most known foci worldwide and in particular from China and the Former Soviet Union has dramatically improved our understanding of the population structure of this species. In the current view, Y. pestis including the non or moderate human pathogen Y. pestis subspecies microtus emerged from Yersinia pseudotuberculosis about 2,600 to 28,600 years ago in central Asia. The majority of central Asia natural foci have been investigated. However these investigations included only few strains from Mongolia.Methodology/Principal FindingsClustered Regularly Interspaced Short Prokaryotic Repeats (CRISPR) analysis and Multiple-locus variable number of tandem repeats (VNTR) analysis (MLVA) with 25 loci was performed on 100 Y. pestis strains, isolated from 37 sampling areas in Mongolia. The resulting data were compared with previously published data from more than 500 plague strains, 130 of which had also been previously genotyped by single nucleotide polymorphism (SNP) analysis. The comparison revealed six main clusters including the three microtus biovars Ulegeica, Altaica, and Xilingolensis. The largest cluster comprises 78 isolates, with unique and new genotypes seen so far in Mongolia only. Typing of selected isolates by key SNPs was used to robustly assign the corresponding clusters to previously defined SNP branches.Conclusions/SignificanceWe show that Mongolia hosts the most recent microtus clade (Ulegeica). Interestingly no representatives of the ancestral Y. pestis subspecies pestis nodes previously identified in North-western China were identified in this study. This observation suggests that the subsequent evolution steps within Y. pestis pestis did not occur in Mongolia. Rather, Mongolia was most likely re-colonized by more recent clades coming back from China contemporary of the black death pandemic, or more recently in the past 600 years.

Comparative Genomics of 2009 Seasonal Plague (Yersinia pestis) in New Mexico

Plague disease caused by the Gram-negative bacterium Yersinia pestis routinely affects animals and occasionally humans, in the western United States. The strains native to the North American continent are thought to be derived from a single introduction in the late 19th century. The degree to which these isolates have diverged genetically since their introduction is not clear, and new genomic markers to assay the diversity of North American plague are highly desired. To assay genetic diversity of plague isolates within confined geographic areas, draft genome sequences were generated by 454 pyrosequencing from nine environmental and clinical plague isolates. In silico assemblies of Variable Number Tandem Repeat (VNTR) loci were compared to laboratory-generated profiles for seven markers. High-confidence SNPs and small Insertion/Deletions (Indels) were compared to previously sequenced Y. pestis isolates. The resulting panel of mutations allowed clustering of the strains and tracing of the most likely evolutionary trajectory of the plague strains. The sequences also allowed the identification of new putative SNPs that differentiate the 2009 isolates from previously sequenced plague strains and from each other. In addition, new insertion points for the abundant insertion sequences (IS) of Y. pestis are present that allow additional discrimination of strains; several of these new insertions potentially inactivate genes implicated in virulence. These sequences enable whole-genome phylogenetic analysis and allow the unbiased comparison of closely related isolates of a genetically monomorphic pathogen.

Genetic diversity of Yersinia pestis in Brazil

Plague outbreaks are occasionally reported in Brazil. Unfortunately, due to great genetic similarity, molecular subtyping of Yersinia pestis strains is difficult. Analysis of multiple-locus variable number of tandem repeats (VNTR), also known as MLVA, has been found to be a valuable tool to discriminate among strains. To check for genetic differences, strains obtained from two different ecological complexes in Brazil collected during two different epidemiological events, an epizootic in Sítio Alagoinha in 1967 and an outbreak in Planalto da Borborema in 1986, were subtyped through MLVA using 12 VNTR loci. Three clusters (A, B and C) were observed. Of the 20 strains from the epizootic, 18 fit into cluster A. Cluster A was divided into two subgroups: A(1) (15 strains) and A(2) (3 strains). Of the 17 strains from the outbreak, 15 fit into cluster B. Cluster B was divided into three subgroups: B(1) (4 strains), B(2) (4 strains) and B(3) (7 strains). Cluster C is a singleton with one epizootic strain. The external standards, Y. pestis CO92 and Y. pseudotuberculosis IP32953, formed two clusters of singletons. The stability of 12 VNTR loci of three unrelated cultures included in this study was assessed. The 12 VNTR loci were stable through multiple serial subcultures in the laboratory. MLVA revealed that Y. pestis populations in Brazil are not monomorphic, and that there is intraspecific genetic diversity among Brazilian plague strains. We conclude that there is some correlation among genetic groups of this species, related to the temporal and geographic origin of isolates.

An additional step in the transmission of Yersinia pestis?

Plague, caused by the bacterium Yersinia pestis, is a mammalian vector-borne disease, transmitted by fleas that serve as the vector between rodent hosts. For many pathogens, including Y. pestis, there are strong evolutionary pressures that lead to a reduction in 'useless genes', with only those retained that reflect function in the specific environment inhabited by the pathogen. Genetic traits critical for survival and transmission between two environments, the rodent and the flea, are conserved in epizootic/epidemic plague strains. However, there are genes that remain conserved for which no function in the flea-rodent cycle has yet been observed, indicating an additional environment may exist in the transmission cycle of plague. Here, we present evidence for highly conserved genes that suggests a role in the persistence of Y. pestis after death of its host. Furthermore, maintenance of these genes points to Y. pestis traversing a post-mortem path between, and possibly within, epizootic periods and offering insight into mechanisms that may allow Y. pestis an alternative route of transmission in the natural environment.

Development of Nutrient Media out of Vegetable Material for Culturing Particularly Dangerous Infections Agents

5 experimental nutritious bases were prepared out of plant materials (soybeans, soymilk, sugar-beet molasses - sugar syrup), and their physical and chemical parameters and amino-acid composition were studied. Nutrient media developed on experimental basis met the nutritious demands of studied strains of plague, cholera, anthrax microbes and sporogenous saprophytes.

Comparative Analysis of <i>inv</i> Gene Structure in Strains of Plague and Pseudotuberculosis Agents

Gene inv encoding one of the important pathogenicity factors of pseudotuberculosis agent was analyzed as regards its structure and function in plague microbe of the main and non-main subspecies and pseudotuberculosis microbe. The structure of gene inv was shown to be affected by introduction of insertion sequence IS1541 in all studied Yersinia pestis strains of non-main and main subspecies. This evidences that the loss of functional activity of this gene took place at initial stage of plague agent evolution.

Biological and physicochemical properties of Yersinia pseudotuberculosis cultures carrying the Yersinia pestis fra operon

The biological and physicochemical properties of cultures of two isogenic recombinant variants of Y. pseudotuberculosis were studied. The cell genomes of the studied cultures differed only with respect to the presence or absence of the fra operon responsible for capsule formation in Y. pestis. The expression of the above-mentioned trait is amplified by an increase in the cultivation temperature of microbial biomass and stimulates a decrease in the viability and adaptation potential of bacteria in vitro. In a warm-blooded host organism, the capsule-forming recombinant variant is characterized by greater residual pathogenicity and immunogenicity to the experimental plague strains of laboratory animals as compared to the reference variant cells. These differences can be attributed to the more increased colonizing ability of the capsule-forming microbes caused by the resistance of the host cells to phagocytosis.

Yersinia pestis caf1Variants and the Limits of Plague Vaccine Protection

Yersinia pestis, the highly virulent agent of plague, is a biological weapon. Strategies that prevent plague have been sought for centuries, and immunization with live, attenuated (nonpigmented) strains or subunit vaccines with F1 (Caf1) antigen is considered effective. We show here that immunization with live, attenuated strains generates plague-protective immunity and humoral immune responses against F1 pilus antigen and LcrV. Y. pestis variants lacking caf1 (F1 pili) are not only fully virulent in animal models of bubonic and pneumonic plague but also break through immune responses generated with live, attenuated strains or F1 subunit vaccines. In contrast, immunization with purified LcrV, a protein at the tip of type III needles, generates protective immunity against the wild-type and the fully virulent caf1 mutant strain, in agreement with the notion that LcrV can elicit vaccine protection against both types of virulent plague strains.

Natural history of plague: perspectives from more than a century of research.

For more than a century, scientists have investigated the natural history of plague, a highly fatal disease caused by infection with the gram-negative bacterium Yersinia pestis. Among their most important discoveries were the zoonotic nature of the disease and that plague exists in natural cycles involving transmission between rodent hosts and flea vectors. Other significant findings include those on the evolution of Y. pestis; geographic variation among plague strains; the dynamics and maintenance of transmission cycles; mechanisms by which fleas transmit Y. pestis; resistance and susceptibility among plague hosts; the structure and typology of natural foci; and how landscape features influence the focality, maintenance, and spread of the disease. The knowledge gained from these studies is essential for the development of effective prevention and control strategies.