Evolution of immune genes is associated with the Black Death.

Infectious diseases are among the strongest selective pressures driving human evolution1,2. This includes the single greatest mortality event in recorded history, the first outbreak of the second pandemic of plague, commonly called the Black Death, which was caused by the bacterium Yersinia pestis3. This pandemic devastated Afro-Eurasia, killing up to 30-50% of the population4. To identify loci that may have been under selection during the Black Death, we characterized genetic variation around immune-related genes from 206 ancient DNA extracts, stemming from two different European populations before, during and after the Black Death. Immune loci are strongly enriched for highly differentiated sites relative to a set of non-immune loci, suggesting positive selection. We identify 245 variants that are highly differentiated within the London dataset, four of which were replicated in an independent cohort from Denmark, and represent the strongest candidates for positive selection. The selected allele for one of these variants, rs2549794, is associated with the production of a full-length (versus truncated) ERAP2 transcript, variation in cytokine response to Y. pestis and increased ability to control intracellular Y. pestis in macrophages. Finally, we show that protective variants overlap with alleles that are today associated with increased susceptibility to autoimmune diseases, providing empirical evidence for the role played by past pandemics in shaping present-day susceptibility to disease.

The NLRP12 inflammasome recognizes Yersinia pestis.

Yersinia pestis, the causative agent of plague, is able to suppress production of inflammatory cytokines IL-18 and IL-1ß, which are generated through caspase-1-activating nucleotide-binding domain and leucine-rich repeat (NLR)-containing inflammasomes. Here, we sought to elucidate the role of NLRs and IL-18 during plague. Lack of IL-18 signaling led to increased susceptibility to Y. pestis, producing tetra-acylated lipid A, and an attenuated strain producing a Y. pseudotuberculosis-like hexa-acylated lipid A. We found that the NLRP12 inflammasome was an important regulator controlling IL-18 and IL-1ß production after Y. pestis infection, and NLRP12-deficient mice were more susceptible to bacterial challenge. NLRP12 also directed interferon-γ production via induction of IL-18, but had minimal effect on signaling to the transcription factor NF-κB. These studies reveal a role for NLRP12 in host resistance against pathogens. Minimizing NLRP12 inflammasome activation may have been a central factor in evolution of the high virulence of Y. pestis.

Human ectoparasites and the spread of plague in Europe during the Second Pandemic.

Plague, caused by the bacterium Yersinia pestis, can spread through human populations by multiple transmission pathways. Today, most human plague cases are bubonic, caused by spillover of infected fleas from rodent epizootics, or pneumonic, caused by inhalation of infectious droplets. However, little is known about the historical spread of plague in Europe during the Second Pandemic (14-19th centuries), including the Black Death, which led to high mortality and recurrent epidemics for hundreds of years. Several studies have suggested that human ectoparasite vectors, such as human fleas (Pulex irritans) or body lice (Pediculus humanus humanus), caused the rapidly spreading epidemics. Here, we describe a compartmental model for plague transmission by a human ectoparasite vector. Using Bayesian inference, we found that this model fits mortality curves from nine outbreaks in Europe better than models for pneumonic or rodent transmission. Our results support that human ectoparasites were primary vectors for plague during the Second Pandemic, including the Black Death (1346-1353), ultimately challenging the assumption that plague in Europe was predominantly spread by rats.

Survival analysis of the Black Death: Social inequality of women and the perils of life and death in Medieval London.

OBJECTIVES: Described as an indiscriminate killer by many chroniclers, the Black Death descended on London during the 14th century. To best understand the pattern of transmission among demographic groups, models should include multiple demographic and health covariates concurrently, something rarely done when examining Black Death, but implemented in this study to identify which demographic groups had a higher susceptibility. Female predisposition to the Black Death was also explored, focusing on whether social inequality added to vulnerability. MATERIALS AND METHODS: Three attritional cemeteries from the Wellcome Osteological Research Database were compared with the Black Death cemetery, East Smithfield. A Cox proportional hazards regression estimated hazards ratios of dying of the Black Death, using transition analysis ages-at-death as the time variable, and sex and frailty as covariates. Additionally, a binomial logistic regression generated odds ratios for age-at-death, sex, and frailty. RESULTS: The Cox model produced a significant hazards ratio for individuals deemed frail. Similarly, the logit model calculated significantly increased odds ratios for frail individuals, and decreased odds for individuals aged 65+. DISCUSSION: The older individuals were not undergoing growth during times of great stress in London pre-dating the Black Death epidemic, which may explain the decreased odds of contracting the Black Death. Further, although women dealt with social inequality, which partially led to the demographic puzzle of the Medieval "missing" women, women's susceptibility to infection by the Black Death was not increased. The phenomenon of the missing women may be due to a combination of factors, including infant and child mortality and preservation.

Sex differences in adult famine mortality in medieval London.

OBJECTIVES: Recurrent famine events during the medieval period might have contributed to excess mortality during the Black Death in London, England (c. 1349-1350). Previous research using conventional methods of age estimation revealed that adult males experienced lower risks of mortality under "normal" (attritional) but not famine mortality conditions following the Black Death. However, given the biases inherent in conventional age estimation methods, this study reassesses sex differences in risks of medieval adult famine mortality using ages estimated via transition analysis, which avoids some of the limitations of conventional age estimation methods. MATERIALS AND METHODS: We apply hazards analysis (the Gompertz model of adult mortality) to ages estimated for human skeletal remains (n = 1245) from London cemeteries dated to the pre-Black Death (c. 1000-1250 CE) and post-Black Death (c. 1350-1540 CE) periods. RESULTS: The results reveal no sex differences in risks of mortality before the Black Death but indicate that adult males faced lower risks of mortality after the Black Death during conditions of normal and famine mortality. CONCLUSIONS: These findings largely support those of our previous research, which suggested that selective mortality during the Black Death or sex-biased improvements in standard of living following the epidemic reduced risk of mortality for adult males in the post-Black Death period under normal mortality conditions. However, the use of transition analysis age estimates also revealed a reduced risk of mortality for post-Black Death adult males under famine conditions.

A Recombinant Attenuated Yersinia pseudotuberculosis Vaccine Delivering a Y. pestis YopENt138-LcrV Fusion Elicits Broad Protection against Plague and Yersiniosis in Mice.

In this study, a novel recombinant attenuated Yersinia pseudotuberculosis PB1+ strain (χ10069) engineered with ΔyopK ΔyopJ Δasd triple mutations was used to deliver a Y. pestis fusion protein, YopE amino acid 1 to 138-LcrV (YopENt138-LcrV), to Swiss Webster mice as a protective antigen against infections by yersiniae. χ10069 bacteria harboring the pYA5199 plasmid constitutively synthesized the YopENt138-LcrV fusion protein and secreted it via the type 3 secretion system (T3SS) at 37°C under calcium-deprived conditions. The attenuated strain χ10069(pYA5199) was manifested by the establishment of controlled infection in different tissues without developing conspicuous signs of disease in histopathological analysis of microtome sections. A single-dose oral immunization of χ10069(pYA5199) induced strong serum antibody titers (log10 mean value, 4.2), secretory IgA in bronchoalveolar lavage (BAL) fluid from immunized mice, and Yersinia-specific CD4+ and CD8+ T cells producing high levels of tumor necrosis factor alpha (TNF-α), gamma interferon (IFN-γ), and interleukin 2 (IL-2), as well as IL-17, in both lungs and spleens of immunized mice, conferring comprehensive Th1- and Th2-mediated immune responses and protection against bubonic and pneumonic plague challenges, with 80% and 90% survival, respectively. Mice immunized with χ10069(pYA5199) also exhibited complete protection against lethal oral infections by Yersinia enterocolitica WA and Y. pseudotuberculosis PB1+. These findings indicated that χ10069(pYA5199) as an oral vaccine induces protective immunity to prevent bubonic and pneumonic plague, as well as yersiniosis, in mice and would be a promising oral vaccine candidate for protection against plague and yersiniosis for human and veterinary applications.

A survival analysis of the last great European plagues: The case of Nonantola (Northern Italy) in 1630.

This paper develops the first survival analysis of a large-scale mortality crisis caused by plague. For the time-to-event analyses we used the Cox proportional hazards regression model. Our case study is the town of Nonantola during the 1630 plague, which was probably the worst to affect Italy since the Black Death. Individual risk of death did not depend on sex, grew with age (peaking at ages 40-60 and then declining), was not affected by socio-economic status, and was positively associated with household size. We discuss these findings in light of the historical-demographic and palaeo-demographic literature on medieval and early modern plagues. Our results are compatible with the debated idea that ancient plague was able to spread directly from human to human. Our methods could be replicated in other studies of European plagues to nuance and integrate the findings of recent palaeo-biological and palaeo-demographic research on plague.

Induction of Type I Interferon through a Noncanonical Toll-Like Receptor 7 Pathway during Yersinia pestis Infection.

Yersinia pestis causes bubonic, pneumonic, and septicemic plague, diseases that are rapidly lethal to most mammals, including humans. Plague develops as a consequence of bacterial neutralization of the host's innate immune response, which permits uncontrolled growth and causes the systemic hyperactivation of the inflammatory response. We previously found that host type I interferon (IFN) signaling is induced during Y. pestis infection and contributes to neutrophil depletion and disease. In this work, we show that type I IFN expression is derived from the recognition of intracellular Y. pestis by host Toll-like receptor 7 (TLR7). Type I IFN expression proceeded independent of myeloid differentiation factor 88 (MyD88), which is the only known signaling adaptor for TLR7, suggesting that a noncanonical mechanism occurs in Y. pestis-infected macrophages. In the murine plague model, TLR7 was a significant contributor to the expression of serum IFN-ß, whereas MyD88 was not. Furthermore, like the type I IFN response, TLR7 contributed to the lethality of septicemic plague and was associated with the suppression of neutrophilic inflammation. In contrast, TLR7 was important for defense against disease in the lungs. Together, these data demonstrate that an atypical TLR7 signaling pathway contributes to type I IFN expression during Y. pestis infection and suggest that the TLR7-driven type I IFN response plays an important role in determining the outcome of plague.

Pneumonic Plague Transmission, Moramanga, Madagascar, 2015.

During a pneumonic plague outbreak in Moramanga, Madagascar, we identified 4 confirmed, 1 presumptive, and 9 suspected plague case-patients. Human-to-human transmission among close contacts was high (reproductive number 1.44) and the case fatality rate was 71%. Phylogenetic analysis showed that the Yersinia pestis isolates belonged to group q3, different from the previous outbreak.

Patterns of Human Plague in Uganda, 2008-2016.

Plague is a highly virulent fleaborne zoonosis that occurs throughout many parts of the world; most suspected human cases are reported from resource-poor settings in sub-Saharan Africa. During 2008-2016, a combination of active surveillance and laboratory testing in the plague-endemic West Nile region of Uganda yielded 255 suspected human plague cases; approximately one third were laboratory confirmed by bacterial culture or serology. Although the mortality rate was 7% among suspected cases, it was 26% among persons with laboratory-confirmed plague. Reports of an unusual number of dead rats in a patient's village around the time of illness onset was significantly associated with laboratory confirmation of plague. This descriptive summary of human plague in Uganda highlights the episodic nature of the disease, as well as the potential that, even in endemic areas, illnesses of other etiologies might be being mistaken for plague.

The sex-selective impact of the Black Death and recurring plagues in the Southern Netherlands, 1349-1450.

Although recent work has begun to establish that early modern plagues had selective mortality effects, it was generally accepted that the initial outbreak of Black Death in 1347-52 was a "universal killer." Recent bioarchaeological work, however, has argued that the Black Death was also selective with regard to age and pre-plague health status. The issue of the Black Death's potential sex selectivity is less clear. Bioarchaeological research hypothesizes that sex-selection in mortality was possible during the initial Black Death outbreak, and we present evidence from historical sources to test this notion. OBJECTIVE: To determine whether the Black Death and recurring plagues in the period 1349-1450 had a sex-selective mortality effect. MATERIALS AND METHODS: We present a newly compiled database of mortality information taken from mortmain records in Hainaut, Belgium, in the period 1349-1450, which not only is an important new source of information on medieval mortality, but also allows for sex-disaggregation. RESULTS: We find that the Black Death period of 1349-51, as well as recurring plagues in the 100 years up to 1450, often had a sex-selective effect-killing more women than in "non-plague years." DISCUSSION: Although much research tends to suggest that men are more susceptible to a variety of diseases caused by bacteria, viruses and parasites, we cannot assume that the same direction of sex-selection in mortality applied to diseases in the distant past such as Second Pandemic plagues. While the exact reasons for the sex-selective effect of late-medieval plague are unclear in the absence of further data, we suggest that simple inequities between the sexes in exposure to the disease may not have been a key driver.

Dynamics of the pneumonic plague epidemic in Madagascar, August to October 2017.

Transmission potential and severity of pneumonic plague in Madagascar were assessed. Accounting for reporting delay, the reproduction number was estimated at 1.73. The case fatality risk was estimated as 5.5%. Expected numbers of exported cases from Madagascar were estimated across the world and all estimates were below 1 person from August to October, 2017.

New Role for FDA-Approved Drugs in Combating Antibiotic-Resistant Bacteria.

Antibiotic resistance in medically relevant bacterial pathogens, coupled with a paucity of novel antimicrobial discoveries, represents a pressing global crisis. Traditional drug discovery is an inefficient and costly process; however, systematic screening of Food and Drug Administration (FDA)-approved therapeutics for other indications in humans offers a rapid alternative approach. In this study, we screened a library of 780 FDA-approved drugs to identify molecules that rendered RAW 264.7 murine macrophages resistant to cytotoxicity induced by the highly virulent Yersinia pestis CO92 strain. Of these compounds, we identified 94 not classified as antibiotics as being effective at preventing Y. pestis-induced cytotoxicity. A total of 17 prioritized drugs, based on efficacy in in vitro screens, were chosen for further evaluation in a murine model of pneumonic plague to delineate if in vitro efficacy could be translated in vivo Three drugs, doxapram (DXP), amoxapine (AXPN), and trifluoperazine (TFP), increased animal survivability despite not exhibiting any direct bacteriostatic or bactericidal effect on Y. pestis and having no modulating effect on crucial Y. pestis virulence factors. These findings suggested that DXP, AXPN, and TFP may modulate host cell pathways necessary for disease pathogenesis. Finally, to further assess the broad applicability of drugs identified from in vitro screens, the therapeutic potential of TFP, the most efficacious drug in vivo, was evaluated in murine models of Salmonella enterica serovar Typhimurium and Clostridium difficile infections. In both models, TFP treatment resulted in increased survivability of infected animals. Taken together, these results demonstrate the broad applicability and potential use of nonantibiotic FDA-approved drugs to combat respiratory and gastrointestinal bacterial pathogens.

Survival protein A is essential for virulence in Yersinia pestis.

Plague is a highly pathogenic disease caused by the bacterium Yersinia pestis. There is currently no vaccine available for prophylaxis and antibiotic resistant strains have been isolated, thus there is a need for the development of new countermeasures to treat this disease. Survival protein A (SurA) is a chaperone that has been linked to virulence in several species of bacteria, including the close relative Yersinia pseudotuberculosis. In this study, we aimed to evaluate the role of SurA in virulence of the highly pathogenic Y. pestis by creating an unmarked surA deletion mutant. The Y. pestis ΔsurA mutant was found to be more susceptible to membrane perturbing agents and was completely avirulent in a mouse infection model when delivered up to 2.1 × 10(5) CFU by the subcutaneous route. This provides strong evidence that SurA would make a promising antimicrobial target.

The Experience of Plague in East Kent, 1636­38.

On 2 July 1637 43-year-old Elizabeth Burgess, from Canterbury, was buried at Birchington, Kent, having died of the plague in the hamlet of Wood in that parish. She was the first victim of an outbreak of plague in 1637. This article firstly examines that outbreak in Birchington in detail; then it considers how far the parish's experience matched that in nearby Faversham and Sandwich. It finally discusses the extent to which the infection spread to the surrounding area.

Impact of the Pla protease substrate α2-antiplasmin on the progression of primary pneumonic plague.

Many pathogens usurp the host hemostatic system during infection to promote pathogenesis. Yersinia pestis, the causative agent of plague, expresses the plasminogen activator protease Pla, which has been shown in vitro to target and cleave multiple proteins within the fibrinolytic pathway, including the plasmin inhibitor α2-antiplasmin (A2AP). It is not known, however, if Pla inactivates A2AP in vivo; the role of A2AP during respiratory Y. pestis infection is not known either. Here, we show that Y. pestis does not appreciably cleave A2AP in a Pla-dependent manner in the lungs during experimental pneumonic plague. Furthermore, following intranasal infection with Y. pestis, A2AP-deficient mice exhibit no difference in survival time, bacterial burden in the lungs, or dissemination from wild-type mice. Instead, we found that in the absence of Pla, A2AP contributes to the control of the pulmonary inflammatory response during infection by reducing neutrophil recruitment and cytokine production, resulting in altered immunopathology of the lungs compared to A2AP-deficient mice. Thus, our data demonstrate that A2AP is not significantly affected by the Pla protease during pneumonic plague, and although A2AP participates in immune modulation in the lungs, it has limited impact on the course or ultimate outcome of the infection.

Pneumonic plague outbreak, Northern Madagascar, 2011.

Yersinia pestis, the causative agent of plague, is endemic to Madagascar, particularly to the central highlands. Although plague has not been previously reported in northern Madagascar, an outbreak of pneumonic plague occurred in this remote area in 2011. Over a 27-day period, 17 suspected, 2 presumptive, and 3 confirmed human cases were identified, and all 15 untreated 20 patients died. Molecular typing of Y. pestis isolated from 2 survivors and 5 Rattus rattus rat samples identified the Madagascar-specific 1.ORI3-k single-nucleotide polymorphism genotype and 4 clustered regularly interspaced short palindromic repeat patterns. This outbreak had a case-fatality rate of 100% for nontreated patients. The Y. pestis 1.ORI3-k single-nucleotide polymorphism genotype might cause larger epidemics. Multidrug-resistant strains and persistence of the pathogen in natural foci near human settlements pose severe risks to populations in plague-endemic regions and require outbreak response strategies.