Flea-borne transmission model to evaluate vaccine efficacy against naturally acquired bubonic plague

A flea-to-mouse transmission model was developed for use in testing new candidate vaccines for the ability to protect against flea-borne plague. The model was used to evaluate a recombinant fusion protein vaccine consisting of the Yersinia pestis F1 and V antigens. After one to three challenges with Y. pestis-infected fleas, 14 of 15 unvaccinated control mice developed plague, with an average septicemia level of 9.2 x 10(8) Y. pestis CFU/ml. None of 15 vaccinated mice developed the disease after similar challenges, and serological testing indicated that transmitted bacteria were eliminated by the immune system before extensive replication and systemic infection could occur. The transmission and development of disease in control mice correlated with the number of bites by blocked fleas but not with the total number of fleabites. The model provides a means to directly assess the efficacy of new vaccines to prevent naturally acquired bubonic plague and to study events at the vector-host interface that lead to dissemination and disease.     

Role of the Yersinia pestis Ail protein in preventing a protective polymorphonuclear leukocyte response during bubonic plague

The ability of Yersinia pestis to forestall the mammalian innate immune response is a fundamental aspect of plague pathogenesis. In this study, we examined the effect of Ail, a 17-kDa outer membrane protein that protects Y. pestis against complement-mediated lysis, on bubonic plague pathogenesis in mice and rats. The Y. pestis ail mutant was attenuated for virulence in both rodent models. The attenuation was greater in rats than in mice, which correlates with the ability of normal rat serum, but not mouse serum, to kill ail-negative Y. pestis in vitro. Intradermal infection with the ail mutant resulted in an atypical, subacute form of bubonic plague associated with extensive recruitment of polymorphonuclear leukocytes (PMN or neutrophils) to the site of infection in the draining lymph node and the formation of large purulent abscesses that contained the bacteria. Systemic spread and mortality were greatly attenuated, however, and a productive adaptive immune response was generated after high-dose challenge, as evidenced by high serum antibody levels against Y. pestis F1 antigen. The Y. pestis Ail protein is an important bubonic plague virulence factor that inhibits the innate immune response, in particular the recruitment of a protective PMN response to the infected lymph node.     

The rate of blood flow in experimental bubonic plague

Summary The circulation rate was studied during experimental plague in guinea pigs and rabbits by means of the isotope method. In guinea pigs with a rapid course of plague infection, with the animals' death occurring on the 4th–5th day, the circulation rate decreases 1–2 days after the infection. With a more torpid infectious process the circulation rate showed some rise during the initial period. Beginning from the 3rd–4th day after the infection in all the guinea pigs there was seen a 11/2–2 reduction of the circulation rate in comparison with the initial one. In rabbits suffering from typical plague with characteristic pathologico-anatomical changes and bacteriological isolation ofBacillus pestis from the organs, the circulation rate during the infectious process is almost halved in comparison with its initial value.(Presented by Active Member of the Akad. Med. Nauk SSSR N. N. Zhukov-Verezhnikov) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 53, No. 2, pp. 47–52, February, 1962     

yadBC of Yersinia pestis, a new virulence determinant for bubonic plague

In all Yersinia pestis strains examined, the adhesin/invasin yadA gene is a pseudogene, yet Y. pestis is invasive for epithelial cells. To identify potential surface proteins that are structurally and functionally similar to YadA, we searched the Y. pestis genome for open reading frames with homology to yadA and found three: the bicistronic operon yadBC (YPO1387 and YPO1388 of Y. pestis CO92; y2786 and y2785 of Y. pestis KIM5), which encodes two putative surface proteins, and YPO0902, which lacks a signal sequence and likely is nonfunctional. In this study we characterized yadBC regulation and tested the importance of this operon for Y. pestis adherence, invasion, and virulence. We found that loss of yadBC caused a modest loss of invasiveness for epithelioid cells and a large decrease in virulence for bubonic plague but not for pneumonic plague in mice.     

Oral vaccination against bubonic plague using a live avirulent Yersinia pseudotuberculosis strain

We evaluated the possibility of using Yersinia pseudotuberculosis as a live vaccine against plague because it shares high genetic identity with Y. pestis while being much less virulent, genetically much more stable, and deliverable orally. A total of 41 Y. pseudotuberculosis strains were screened by PCR for the absence of the high pathogenicity island, the superantigens YPM, and the type IV pilus and the presence of the pYV virulence plasmid. One strain (IP32680) fulfilled these criteria. This strain was avirulent in mice upon intragastric or subcutaneous inoculation and persisted for 2 months in the mouse intestine without clinical signs of disease. IP32680 reached the mesenteric lymph nodes, spleen, and liver without causing major histological lesions and was cleared after 13 days. The antibodies produced in vaccinated animals recognized both Y. pseudotuberculosis and Y. pestis antigens efficiently. After a subcutaneous challenge with Y. pestis CO92, bacteria were found in low amounts in the organs and rarely in the blood of vaccinated animals. One oral IP32680 inoculation protected 75% of the mice, and two inoculations induced much higher antibody titers and protected 88% of the mice. Our results thus validate the concept that an attenuated Y. pseudotuberculosis strain can be an efficient, inexpensive, safe, and easy-to-produce live vaccine for oral immunization against bubonic plague.     

Diagnosis of bubonic plague by PCR in Madagascar under field conditions

The diagnostic value of a PCR assay that amplifies a 501-bp fragment of the Yersinia pestis caf1 gene has been determined in a reference laboratory with 218 bubo aspirates collected from patients with clinically suspected plague managed in a regional hospital in Madagascar. The culture of Y. pestis and the detection of the F1 antigen (Ag) by enzyme-linked immunosorbent assay (ELISA) were used as reference diagnostic methods. The sensitivity of PCR was 89% (57 of 64) for the Y. pestis-positive patients, and 80.7% (63 of 78) for the F1 Ag-positive patients. The specificity of PCR for the culture-, F1 Ag-, and antibody-negative patients (n = 105) was 100%. Because in Madagascar most patients with plague are managed and their clinical samples are collected in remote villages, the usefulness of PCR was evaluated for routine diagnostic use in the operational conditions of the control program. The sensitivity of PCR was 50% (25 of 50) relative to the results of culture and 35.2% (19 of 54) relative to the results of the F1 Ag immunocapture ELISA. The specificity of PCR under these conditions was 96%. In conclusion, the PCR method was found to be very specific but not as sensitive as culture or the F1 Ag detection method. The limitation in sensitivity may have been due to suboptimal field conditions and the small volumes of samples used for DNA extraction. This technique is not recommended as a routine diagnostic test for plague in Madagascar.     

Identification of early disease progression in an ALS rat model

Transgenic rat models of amyotrophic lateral sclerosis (ALS) have recently been developed. Most assays of ALS-symptoms in these models monitor disease onset accurately, but do not identify individuals that will develop these symptoms before the motor deficits become apparent. Peak bodyweight has recently been shown to indicate affected individuals before motor deficits become apparent. However, it must be determined retrospectively due to weight fluctuation. Here, we report that exploratory activities detected by a photobeam activity system (PAS) and wire mesh ascending test can be used to detect slight motor deficits in the early phase of ALS. Thus, these tests may be used in addition to peak bodyweight to monitor early disease progression and to assay efficacy of new therapeutic interventions.     

雷帕霉素减缓大鼠被动Heymann肾炎的进展

 目的：观察雷帕霉素对大鼠被动Heymann肾炎（PHN）的影响,并探讨自噬在其中的作用。方法：雄性SD大鼠随机分为3组,即对照组、PHN模型组和雷帕霉素治疗组。以造模后第21天为观察结点,采用全自动生化分析仪测定24 h尿蛋白总量、血尿素氮和血清肌酐,过碘酸-六次甲基四胺银染色观察肾脏病变,Weibel-Gomez点计数方法计数足细胞数量,免疫荧光染色检测肾小球内C5b-9的沉积,免疫组化染色观察caspase-3的表达,Western blotting检测肾小球LC3的表达。结果：雷帕霉素明显减轻PHN模型大鼠的蛋白尿排出（P<0.05）,同时各组大鼠的肾功能均正常,其间无显著差异;雷帕霉素使PHN大鼠肾小球基底膜增厚的程度和范围有所减轻;雷帕霉素明显改善PHN大鼠足细胞缺失情况,减少足细胞凋亡;雷帕霉素可增强肾小球内固有细胞的自噬水平。结论：在PHN的病变过程中,适度增强自噬减少足细胞凋亡,减轻肾脏病变和缓解蛋白尿,可能是雷帕霉素减缓大鼠PHN进展的重要机制之一。     

Histopathologic host response to polypropylene-based surgical mesh materials in a rat abdominal wall defect model

Composite polypropylene-based surgical mesh materials including various synthetic polymers and naturally occurring biomaterials have been developed to ameliorate device-associated inflammatory response and associated reduced compliance of pure polypropylene meshes. This study evaluated the histomorphologic response of three composite polypropylene-based surgical meshes, Revive?, a polycarbonate polyurethane reinforced monofilamentous polypropylene scaffold, Assure?, a polycarbonate polyurethane reinforced monofilamentous polypropylene scaffold with a resorbable anti-adhesion layer of lactide caprolactone copolymer, and Proceed?, a polypropylene mesh modified with oxidized cellulose, in a soft tissue repair model in the rat. The host inflammatory response and neotissue formation were evaluated by semiquantitative histologic scoring including the amount of cellular infiltration, angiogenesis, presence of multinucleate giant cells, fibrous connective tissue formation, and host neo-extracellular matrix deposition for up to 26 weeks. All three composite surgical mesh materials showed good integration with host tissue as indicated by rapid cellular infiltration, abundant neo-vascularization, minimal shrinkage, and the lack of visible mesh degradation. The devices elicited a similar inflammatory response and the presence of a mild foreign body response in spite of the different composition and morphology of these composite mesh materials.     

Differences in host susceptibility to disease progression in the human challenge model of Haemophilus ducreyi infection

With human volunteers inoculated at two sites with Haemophilus ducreyi, outcomes for a subject were not independent. In a reinfection trial, 2 of 11 previous pustule formers and 6 of 10 previous resolvers resolved all sites of infection. There was no correlation between serum bactericidal or phagocytic activity and outcome in the trial. These data indicate that different hosts are differentially susceptible to disease progression versus resolution in the model.     

Immunogenicity and protective immunity against bubonic plague and pneumonic plague by immunization of mice with the recombinant V10 antigen, a variant of LcrV

In contrast to Yersinia pestis LcrV, the recombinant V10 (rV10) variant (lacking residues 271 to 300) does not suppress the release of proinflammatory cytokines by immune cells. Immunization with rV10 generates robust antibody responses that protect mice against bubonic plague and pneumonic plague, suggesting that rV10 may serve as an improved plague vaccine.     

Yersinia pestis YopJ suppresses tumor necrosis factor alpha induction and contributes to apoptosis of immune cells in the lymph node but is not required for virulence in a rat model of bubonic plague

The virulence of the pathogenic Yersinia species depends on a plasmid-encoded type III secretion system that transfers six Yop effector proteins into host cells. One of these proteins, YopJ, has been shown to disrupt host cell signaling pathways involved in proinflammatory cytokine production and to induce macrophage apoptosis in vitro. YopJ-dependent apoptosis in mesenteric lymph nodes has also been demonstrated in a mouse model of Yersinia pseudotuberculosis infection. These results suggest that YopJ attenuates the host innate and adaptive immune response during infection, but the role of YopJ during bubonic plague has not been completely established. We evaluated the role of Yersinia pestis YopJ in a rat model of bubonic plague following intradermal infection with a fully virulent Y. pestis strain and an isogenic yopJ mutant. Deletion of yopJ resulted in a twofold decrease in the number of apoptotic immune cells in the bubo and a threefold increase in serum tumor necrosis factor alpha levels but did not result in decreased virulence, systemic spread, or colonization levels in the spleen and blood. Our results indicate that YopJ is not essential for bubonic plague pathogenesis, even after peripheral inoculation of low doses of Y. pestis. Instead, the effects of YopJ appear to overlap and augment the immunomodulatory effects of other Y. pestis virulence factors.     

Yersinia outer proteins (YOPS) E, K and N are antigenic but non-protective compared to V antigen, in a murine model of bubonic plague

The pathogenic Yersiniae produce a range of virulence proteins, encoded by a 70 kb plasmid, which are essential for infection, and also form part of a contact-dependent virulence mechanism. One of these proteins, V antigen, has been shown to confer a high level of protection against parenteral infection with Y. pestis in murine models, and is considered to be a protective antigen. In this study, the protective efficacy of V antigen has been compared in the same model with that of other proteins (YopE, YopK and YopN), which are part of the contact-dependent virulence mechanism. Mice immunised with two intraperitoneal doses of V antigen or each of the Yops, administered with either Alhydrogel or interleukin-12, produced high antigen-specific serum IgG titres. As shown in previous studies, V+Alhydrogel was fully protective, and 5/5 mice survived a subcutaneous challenge with 90 or 9x10(3) LD50's of Y. pestis GB. In addition, these preliminary studies also showed that V+IL-12 was partially protective: 4/5 or 3/5 mice survived a challenge with 90 or 9x10(3) LD50's, respectively. In contrast, none of the mice immunised with the Yops survived the challenges, and there was no significant delay in the mean time to death compared to mice receiving a control protein. These results show that using two different vaccine regimens, Yops E, K and N, failed to elicit protective immune responses in a murine model of plague, whereas under the same conditions, V antigen was fully or partially protective.     

Early diagnosis of bubonic plague using F1 antigen capture ELISA assay and rapid immunogold dipstick

Plague is still prevalent in more than 20 countries. Two F1 antigen diagnostic assays (an immunocapture ELISA and an immunogold chromatography dipstick) were evaluated using bubo aspirates, serum and urine specimens from patients suspected with plague. The specificity of the two F1 assays was found 100%. Using bacteriology as a gold reference diagnostic assay, 52 patients were Yersinia pestis culture positive and 141 negative. The sensitivity of the F1 ELISA test was 100% in bubo, 52% in serum and 58% in urine specimens. In culture negative patients, the F1 antigen could be found in 10% bubo aspirates, 5% serum and 7% urine specimens of culture negative patients for whom a seroconversion for anti-F1 antibodies was also observed. The sensitivity of the dipstick assay was 98% on bubo aspirates specimens. Compared to the ELISA test, the agreement rate was 97.5% and the correlation coefficient tau = 0.90 (p < 10(-3)). In conclusion, the diagnosis of bubonic plague has to be performed on bubo fluid rather than on serum or urine specimens. Both the F1 ELISA and the dipstick assays are valuable tools for an early diagnosis and for the surveillance of plague.     

Combined effects of chemotherapy and host antitumor response in a murine histocompatible lymphoma model

Combined effects of 1,3,-bis-2-chloroethyl)-1-nitrosourea (BCNU) and host antitumor immune response were studied in mice inoculated intraperitoneally with histocompatible LSTRA leukemia cells carrying virus-induced transplantation antigens. Marked chemo-immune collaborative activity was found to occur when selected schedules of BCNU administration were employed. Moreover, synergist effects were also detected between chemotherapy and both specific and non-specific immunotherapy.     

Protective efficacy of recombinant Yersinia outer proteins against bubonic plague caused by encapsulated and nonencapsulated Yersinia pestis

To evaluate the role of Yersinia outer proteins (Yops) in conferring protective immunity against plague, six yop loci from Yersinia pestis were individually amplified by PCR, cloned, and expressed in Escherichia coli. The recombinant proteins were purified and injected into mice. Most Yop-vaccinated animals succumbed to infection with either wild-type encapsulated Y. pestis or a virulent, nonencapsulated isogenic variant. Vaccination with YpkA significantly prolonged mean survival time but did not increase overall survival of mice infected with the nonencapsulated strain. The only significant protection against death was observed in YopD-vaccinated mice challenged with the nonencapsulated strain.     

Modelling the black death. A historical case study and implications for the epidemiology of bubonic plague

We analysed a plague outbreak in the mining town of Freiberg in Saxony which started in May 1613 and ended in February 1614. This epidemic was selected for study because of the high quality of contemporary sources. It was possible to identify 1400 individual victims meaning that more than 10% of the population of the city perished. The outbreak was modelled by 9 differential equations describing flea, rat, and human populations. This resulted in a close fit to the historical records of this outbreak. An interesting implication of the model is that the introduction of even a small number of immune rats into an otherwise unchanged setting results in an abortive outbreak with very few human victims. Hence, the percentage of immune rats directly influences the magnitude of a human epidemic by diverting search activities of the fleas. Thus, we conclude that the spread of Rattus norvegicus, which might acquire partial herd immunity by exposure to soil- or water-borne Yersinia species due to its preference for wet habitats, contributed to the disappearance of Black Death epidemics from Europe in the 18th century. In order to prove whether or not the parameter values obtained by fitting a given outbreak are also applicable to other cases, we modelled the plague outbreak in Bombay 1905/06 using the same parameter values except for the number of humans as well as of immune and susceptible rats.