Production of defective interfering virus in the brains of mice by an avirulent, in contrast with a virulent, strain of Semliki forest virus.

An avirulent strain (A7) of Semliki Forest virus formed nearly as much haemagglutinating and complement fixing antigen in the brains of adult mice as a virulent (V13) strain, yet the infectivities of the brain tissues were different by about 100-fold. It appeared therefore that defective virus particles were fomed by A7 but these were not demonstrated by fluorescent antibody studies. In short-term organ cultures of adult mouse brain, A7 derived from mouse brain showed a typical interference pattern in inoculum infectivity response curves. Furthermore, when mixed suspensions of brain-grown V13 and A7 with equal infectivities were inoculated the inoculum infectivity response patterns showed significant depressions of the V13 response at higher inocula. Such interference was not detected if chick cell grown A7 and V13 were substituted for the mouse grown virus. The avirulence of A7 in adult mice and its rapid protective effect against lethal V13 infection could be due to the production of defective interfering virus particles in the brain.     

Investigations of reasons for the avirulence of the A7 strain of Semliki Forest virus in adult mice.

A strain of Semliki Forest virus (A7) which is avirulent in adult mice killed baby mice in a similar manner to strain V13 which was also virulent for adult mice. In the muscle and brains of baby mice A7 and V13 replicated and produced haemagglutinating activity similarly. Our previous suggestion that defective interfering particles were present in the brains of A7-infected adult mice appears not to be so. The interference formerly detected was due to an inhibitor present in brain tissue of adult and baby mice both normal and infected. Homogenates of A7-infected adult brain produced normal RNA species in BHK cells and not those characteristic of defective interfering particles. Organ culture experiments indicated that avirulence of A7 was not due to lack of release of virus from infected adult brain cells. Also, A7 as well as V13 was detected and was probably replicating in all parts of the brain and spinal cord that were sectioned and examined. Evidence is presented that suggests that the reason for the avirulence of A7 for adult mice compared with its virulence for baby mice may relate to a lower ability to replicate in brain tissue per se rather than to interaction with host defence mechanisms.     

Site-specific immunity to Leishmania major in SWR mice: the site of infection influences susceptibility and expression of the antileishmanial immune response.

Inbred strains of mice usually develop either of two divergent patterns of infection in response to Leishmania major. Resistant mice, which develop self-limiting infections, respond immunologically with the activation of gamma interferon-secreting Th1 helper T cells, while nonhealing infections in susceptible mice are characterized by the proliferation of interleukin-4-secreting Th2 cells. Development of these divergent responses is dependent primarily on the strain of mouse infected, although factors such as the infective dose, species, and strain of parasite can also influence the degree of resistance. In this study, we show that a single mouse strain, SWR, can develop totally divergent patterns of infection depending on the site of parasite inoculation. Both SWR mice and highly susceptible BALB/c mice developed progressive, ultimately fatal disease when inoculated in the dorsal skin over the base of the tail. However, SWR mice infected in the hind footpad developed far less severe infections, which were for the most part controlled, whereas BALB/c mice infected in this site developed severe, nonhealing lesions. Production of gamma interferon and interleukin-4 and measurement of immunoglobulin E levels in serum were used to assess the degree of Th1 and Th2 cell activation in infected mice. Cytokine profiles early in infection had characteristics of a mixed Th1-Th2 response and were similar in SWR mice infected at either site. These early cytokine responses were not predictive of the ultimate disease outcome, since lymph node cells from healing mice eventually produced higher levels of gamma interferon than did those from nonhealing mice, and healing mice had lower levels of immunoglobulin E in serum, suggesting a functional bias toward Th1 cell activity in these animals. The differential ability of SWR mice to heal infections at different cutaneous sites provides a new model for the study of resistance to cutaneous leishmaniasis. Unlike traditional models of infection in which resistant and susceptible strains of mice are compared, this model allows for the study of factors that contribute to healing and nonhealing infections in a genetically identical strain of mouse.     

Differential susceptibility and resistance of immunocompetent and immunodeficient mice to fatal Hantaan virus infection

Summary Susceptibility and resistance to fatal Hantaan virus meningoencephalitis were studied in immunocompetent (nu/+) and congenitally T-cell deficient (nu/nu) CD-1 mice of different ages. Susceptibility of nu/+ mice to fatal infection was age-dependent, as evidenced by 100 percent mortality in mice inoculated intracerebrally with Hantaan virus (strain 76–118) during the first week of life, 60 percent mortality in mice inoculated at 9 days of age, and no disease or death in mice inoculated at 14 to 42 days of age. Deaths occurred significantly earlier in nu/+ mice inoculated at 5 and 7 days of age than in nu/+ mice less than 24 hours old. Nu/nu littermates of the same age did not exhibit a similar inverse relationship between age and survival times. Moreover, nu/nu mice 14 days or older remained susceptible, albeit with delayed onset of illness and time of death. Virus titers in brain tissues of nu/+ mice inoculated at 7 days and of nu/nu nice inoculated at 7, 9 and 14 days of age were nearly identical. In older nu/+ mice, peak virus titers were comparatively lower, but they did not seem to be influenced by the magnitude of the neutralizing antibody response. The more rapidly fatal course in nu/+ mice inoculated at a week of age than at birth, and the differential resistance between weanling nu/+ and nu/nu mice to Hantaan virus meningoencephalitis suggest that cell-mediated immunity may be responsible for both enhancement of disease and recovery from infection.With 5 Figures     

Host defenses in murine malaria: nonspecific resistance to Plasmodium berghei generated in response to Mycobacterium bovis infection or Corynebacterium parvum stimulation.

Infection with Mycobacterium bovis (BCG) or injection of killed Corynebacterium parvum protected some strain B6D2 F1 (C57BL/6xDBA/2) mice but did not protect strain ICR or A mice from lethal challenge with Plasmodium berghei strain NYU-2. B6D2 mice were not protected against challenges delivered immediately after intravenous injection of these materials, but rather protection developed by day 7 and persisted through at least day 84. Infections in protected mice progressed to about 10% parasitemia in parallel with infections initiated with the same inoculum in untreated controls. However, infections in most of the protected mice were cleared subsequently, whereas infections in untreated controls were uniformly fatal. A small number of treated mice developed protracted high-level erythrocytic infections, which led to markedly delayed death. BCG-infected mice which survived P. berghei infections had a factor in their sera which protected passively immunized recipients from P. berghei. BCG-infected mice passively immunized with protective serum controlled P. berghei infections better than normal mice given the same amount of the same serum and challenged with the same P. berghei inoculum. The capacity of BCG-infected B6D2 mice to resist P. berghei infection was not directly related to the pattern of growth of BCG, to the degree of splenomegaly, or to the level of activation of macrophages (measured as microbicidal capacity) caused by BCG infection. Therefore, I concluded that (i) BCG infection or injection of killed C. parvum altered the immunological potential of B6D2 mice in such a way as to allow the production of measurable levels of a protective humoral factor in response to infection with P. berghei; (ii) BCG infection caused the generation of a capacity which, when expressed in the presence of immune serum, provided an anti-P. berghei capacity which was superior to that provided by BCG infection alone or immune serum in the absence of BCG infection; and (iii) not all strains of mice could be protected from P. berghei by BCG or C. parvum injection.     

Dissemination and proliferation of Salmonella typhimurium in genetically resistant and susceptible mice.

Genetically resistant A/J and CBA mice were inoculated intraperitoneally with either 10(3) or 10(4) organisms of a virulent strain of Salmonella typhimurium; susceptible C57BL/6J and BALB/c mice were inoculated with either 10(2) or 10(3) organisms. Except with the smaller dose in resistant mice, fatal infection ensued. Bacteraemia occurred within 1 h after inoculation, except that it was not detectable during the first 6 h in the susceptible mice inoculated with 10(2) organisms. From day 2, the circulating bacterial population continued to increase in all infected mice, except that it remained under control in the resistant mice inoculated with the lower dose (10(3) organisms). The pathogen proliferated logarithmically in the liver from day 2, and a bacterial count of c. 10(8) cfu/g of tissue was reached when the animals died at 5-7 days; again, the resistant mice inoculated with 10(3) organisms were an exception in which the hepatic bacterial population was kept under control and the mice survived.     

Age Dependence of Viral Expression: Comparative Pathogenesis of Two Rodent-Adapted Strains of Measles Virus in Mice

The pathogenesis of two rodent-adapted strains of measles virus was studied in 1- to 2-day-old suckling and 4-week-old weanling BALB/c mice. Both the mouse-adapted Edmonston (MAEd) strain and the hamster-neurotropic (HNT) strain caused necrotizing giant-cell encephalitis with a 90 to 100% mortality after intracerebral inoculation into suckling mice. After intracerebral inoculation into weanling mice, MAEd virus caused fatal disease in 20% of the mice; HNT virus caused fatal disease in 30%, but an additional 35% of these mice developed disease and then recovered. Even when mice were moribund there was little histological evidence of disease in weanling mice inoculated intracerebrally with either strain of virus. Fluorescent-antibody staining showed extensive measles virus antigen in the suckling mouse brain and focal areas of measles virus antigen in the weanling mouse brain. Infectious virus was recovered easily from the brains of suckling mice by plaquing on Vero cells, but no infectious virus could be recovered similarly from weanling mice. However, virus could be recovered by intracerebral inoculation of weanling mouse tissue homogenates into suckling animals. The immune response appeared to play no role in the recovery from infection or in these age-related differences in disease. It appears that maturation of the cells of the mouse central nervous system converted the production of measles virus from the infectious form in the suckling mouse to a primarily defective infection in the weanling mouse.     

Cerebral malaria in mice: interleukin-2 treatment induces accumulation of gammadelta T cells in the brain and alters resistant mice to susceptible-like phenotype

In this study, we report that infection with Plasmodium yoelii 17XL, a lethal strain of rodent malaria, does not result in death in the DBA/2 strain of mice. In contrast to BALB/c mice, DBA/2 mice developed significantly less parasitemia and never manifested symptoms of cerebral malaria (CM) on infection with this parasite. Moreover, the histological changes evident in the brain of susceptible BALB/c were absent in DBA/2 mice. Interestingly, the resistant DBA/2 mice when treated with recombinant interleukin (IL)-2, were found to develop CM symptoms and the infection became fatal by 6 to 8 days after infection. This condition was associated with an augmented interferon-gamma and nitric oxide production. Unexpectedly, IL-10 levels were also elevated in IL-2-treated DBA/2 mice during late stage of infection (at day 6 of infection) whereas the inverse relationship between IL-10 and interferon-gamma or nitric oxide was maintained in the early stage of infection (at day 3 after infection). The level of tumor necrosis factor-alpha production was moderately increased in the late phase of infection in these mice. Histology of brain from IL-2-treated mice demonstrated the presence of parasitized erythrocytes and infiltration of lymphocytes in cerebral vessels, and also displayed some signs of endothelial degeneration. Confocal microscopical studies demonstrated preferential accumulation of gammadelta T cells in the cerebral vessels of IL-2-treated and -infected mice but not in mice treated with IL-2 alone. The cells recruited in the brain were activated because they demonstrated expression of CD25 (IL-2R) and CD54 (intercellular adhesion molecule 1) molecules. Administration of anti-gammadelta mAb prevented development of CM in IL-2-treated mice until day 18 after infection whereas mice treated with control antibody showed CM symptoms by day 6 after infection. The information concerning creating pathological sequelae and death in an otherwise resistant mouse strain provides an interesting focus for the burden of pathological attributes on death in an infectious disease.     

Susceptible adult murine model for Junin virus

The adult mouse model had been considered resistant to Junin virus (JV) infection. However, we found that C3H/HeJ murine strain proved highly susceptible up to 5 months of age to intracerebral inoculation with the prototype XJ JV strain, showing neurological signs and 80-90% mortality within 13 days. Neutralizing antibodies (Nt Ab) were absent, but low immunofluorescent Ab levels (1:5) were detected as from day +7. The virus could only be rescued by coculture of brain samples with Vero cells. Histopathological findings were consistent with the suckling mouse model and with a delayed-type hypersensitivity reaction. XJ inoculation by extraneural routes failed to cause disease, however, it induced Nt Abs. Ic infection with XJCL3 strain of JV attenuated for man and guinea pig, but not for mouse, induced high Nt Ab levels but not mortality. In either case, mice resisted ic XJ challenge. Thus, C3H/HeJ is the first adult mouse model susceptible to JV.     

Encephalopathy in Mice Following Inapparent Semliki Forest Virus (SFV) Infection

The avirulent “A8” strain of S.F.V. gave rise to acute lesions of encephalitis in mice within 7 days of infection. After 6 weeks, however, all signs of disease disappeared from the brain, but when such animals were destroyed 2 years later they had advanced lesions of hydrocephalus, spongy degeneration and above all a severe astrocytic reaction in many centres of the brain. No such lesions were observed either in animals inoculated with formolinactivated virus, or in uninoculated controls of the same age.     

Recombination of influenza A strains with fowl plague virus can change pneumotropism for mice to a generalized infection with involvement of the central nervous system

Recombination between fowl plague virus FPV/Rostock (Hav1N1), an influenza A strain which produces a generalized fatal infection with involvement of the central nervous system in chickens, and mouse lung-adapted human strains England/1/61 (H2N2) or PR/8/34 (H0N1 results in recombinants which produce a generalized infection in suckling mice with involvement of the central nervous system. A combination of the hemagglutinin and M protein from FPV/Rostock (Hav1N1) with certain polymerase proteins from the mouse lung-adapted strains produces generalized infections in mice. Infectious virus was present in lung, brain, and blood even after intranasal infection. A correlation between the ability to induce a generalized infection of suckling mice in vivo and the replication in mouse embryo fibroblasts in vitro could be demonstrated.     

Etiologic role of lactic dehydrogenase virus infection in an age-dependent neuroparalytic disease in C58 mice

Lactic dehydrogenase virus (LDV) associated with transplantable line Ib lymphocytic leukemia in C58/Wm mice, K36 lymphocytic leukemia in AKR/J mice, and the Gardner lymphosarcoma in C3H/HeJ mice elicited a fatal neuroparalytic disease when injected ip into 7- to 9-month-old X-irradiated indicator C58 mice. LDV associated with the WEHI-3B line of transplantable myelomonocytic leukemia or the Harding-Passey transplantable myeloma in BALB/c mice failed to elicit the disease. Recipients of such tumor extracts were immune to rechallenge by line Ib-associated LDV. Tumor lines free of LDV failed to elicit the disease or immunize recipient mice to line Ib LDV challenge. The Plagemann (P-LDV), Riley (R-LDV), and Notkins (N-LDV) strains of LDV were less neuropathogenic than the line Ib-derived strain (Ib-LDV). Indicator C58 mice that survived infection by the P-LDV, R-LDV, and N-LDV strains were immune to rechallenge by Ib-LDV. Antiserum prepared in young C58 mice to Ib-LDV or R-LDV protected indicator C58 mice from Ib-LDV challenge. These results show that a common viral contaminant of transplantable tumors and virus stocks that ordinarily is not pathogenic elicits a fatal neurologic disease in genetically susceptible, immunosuppressed, C58 mice.     

Pathogenicity of the skalica strain (from the tick-borne encephalitis complex) for white mice

Outbred white mice of different age were inoculated by subcutaneous (s. c.) route with the Skalica strain of tick-borne encephalitis virus (TBE). In the CNS of 3-days-old mice diffuse necrotizing encephalitis, abundant cytoplasmic fluorescence of viral antigen in nearly each neuron and high levels of virus (8.8 log LD50/mg tissue) were found. In 10-days-old mice, the extent of encephalitis and that of immunofluorescence in neurons were less widespread; the peak titre of the virus did not exceed 5.5 log LD50/mg brain tissue. In the CNS of 21-days-old mice the infectivity titre was either very low (1.5 log50/mg on day 3 post infection) or the virus was not detected at all. A few neurons revealed positive fluorescence of viral antigen in the basal ganglia in 1 out of 2 mice examined by day 3 post infection (p. i.). No virus was isolated from the CNS of 2-months-old mice observed for 53 days. In the CNS of 3 out of 10 juvenile mice examined histologically within 8 days post infection, minimal inflammatory changes were seen; foci of neurons showing positive immunofluorescence were not found. The failure to recover infectious virus from cultured brain tissue fragments coming from these mice confirmed the negative outcome of direct virus isolations. It is concluded that the Skalica strain was not pathogenic for juvenile mice when administered by s. c. route.     

Effect of deficiency of tumor necrosis factor alpha or both of its receptors on Streptococcus pneumoniae central nervous system infection and peritonitis

Tumor necrosis factor alpha (TNF-alpha) and TNF-beta are key mediators in bacterial inflammation. We therefore examined the role of TNF-alpha and its two receptors in murine pneumococcal central nervous system infection. TNF-alpha knockout mice and age- and sex-matched controls and TNF receptor (p55 and p75)-deficient mice and heterozygous littermates were infected intracerebrally with a Streptococcus pneumoniae type 3 strain. Mice were monitored until death or were killed 36 h after infection. Bacterial titers in blood, spleen, and brain homogenates were determined. Leukocyte infiltration and neuronal damage were assessed by histological scores. TNF-alpha-deficient mice died earlier than the controls after intracerebral infection although overall survival was similar. TNF-alpha deficiency did not inhibit leukocyte recruitment into the subarachnoid space and did not lead to an increased density of bacteria in brain homogenates. However, it caused a substantial rise of the concentration of S. pneumoniae cells in blood and spleen. Spleen bacterial titers were also increased in p55- and p75-deficient mice. TNF receptor-deficient mice showed decreased meningeal inflammation. Neuronal damage was not affected by either TNF-alpha or TNF receptor deficiency. In a murine model of pneumococcal peritonitis, 10(2) CFU of S. pneumoniae produced fatal peritonitis in TNF-alpha-deficient, but not wild-type, mice. Early leukocyte influx into the peritoneum was impaired in TNF-alpha-deficient mice. The lack of TNF-alpha or its receptors renders mice more susceptible to S. pneumoniae infections.     

Different chemokine expression in lethal and non-lethal murine West Nile virus infection

West Nile (WN) virus is a mosquito-borne flavivirus that can cause lethal encephalitis in humans and horses. The WN virus endemic in New York City (NY) in 1999 caused large-scale mortality of wild birds that was not evident in endemic areas in other parts of the world, and the pathogenesis of the WN virus strain isolated in NY (NY strain) appears to differ from that of previously isolated strains. However, the pathogenesis of NY strain infection remains unclear. This study examined CC (RANTES/CCL5, MIP-1 alpha/CCL3, MIP-1 beta/CCL4) and CXC (IP-10/CXCL10, B lymphocyte chemoattractant (BLC/CXCL13), and B cell- and monocyte-activating chemokine (BMAC/CXCL14)) chemokine expression during lethal NY strain and non-lethal Eg101 strain infection in mice. We found that the mRNA of the CC chemokines, RANTES, MIP-1 alpha, MIP-1 beta, and IP-10 was highly up-regulated in the brain of NY strain-infected mice. By contrast, BLC mRNA was not detected in either group of mice, and BMAC mRNA was highly up-regulated in late stage of infection with the non-lethal Eg101 strain relative to levels in NY strain-infected mice.     

Protection of mice against Vibrio vulnificus disease by vaccination with surface antigen preparations and anti-surface antigen antisera.

Vaccination of mice with either Formalin-killed cells or cell extracts of a virulent strain and a weakly virulent strain of Vibrio vulnificus or with rabbit antisera (AS) against the Formalin-killed cells and cell extracts protected against the virulent strain of V. vulnificus. However, the virulent strain vaccines and AS elicited a significantly stronger immune response than the weakly virulent strain vaccines and AS. Adsorption of the AS with either the homologous or heterologous Formalin-killed cells significantly reduced the ability of the AS to protect mice. The major protective antigen(s) in the cell extracts migrated in the void volume of Sephacryl S-400 superfine, was not effectively sedimented by centrifugation at 100,000 X g for 2 h, had an isoelectric point of 3.8 to 4.2, and was sensitive to boiling or autoclaving for 15 min, periodate oxidation, and exposure to pH 12 but was resistant to 56 degrees C, trypsin, pronase, RNase, neuraminidase, and pH 4.5. Electron microscopy revealed that the virulent strain possessed a more dense ruthenium red-staining layer on its outer membrane and had a much smoother surface than the weakly virulent strain. Our results provide evidence that a major protective antigen and virulence factor of V. vulnificus is a heat-labile, acidic polysaccharide located on the bacterial surface.     

Activation of macrophages by Entamoeba histolytica extracts in mice

The effect of Entamoeba histolytica extracts on the production of inflammatory macrophages and the release of hydrogen peroxide (H2O2) and superoxide (O2-) from these cells was examined in C57BL/6 mice. Two different strains of E. histolytica, either virulent (IP:0682:1) or nonvirulent (DKB), were used in this study. The number of macrophages recovered from the peritoneal cavity of mice treated 5 days previously with 150 micrograms of either strain of amoebic extracts was significantly higher than in the saline-treated groups. Macrophages from mice treated with 150 micrograms of the IP:0682:1 strain of amoebic extracts exhibited a powerful burst of oxidative metabolis when triggered with phorbol myristate acetate (PMA). Extract from the non-virulent strain was much less effective in activating macrophages for the production of oxidative metabolites. Thus, a crude extract preparation from E. histolytica, particularly the virulent strain, induces a strong macrophage inflammatory response in which the cells also produce high levels of H2O2 and O2-.     

Natural killer cells appear to play no role in the recovery of mice from Sindbis virus infection.

Previous studies have suggested that non-specific defence mechanisms may be important in the development of age-dependent resistance to fatal Sindbis-virus infection and in the recovery of adult mice from non-fatal infection. In these studies, natural killer (NK) cell induction was studied in 7-day-old susceptible mice and 28-35-day-old resistant mice. It was found that Sindbis virus infection induced NK cells in both the young and older mice, suggesting that NK cells were not important in the acquisition of resistance to fatal Sindbis-virus infection. Transfer of 10(8) lymph node cells from adult, mice, at the peak of NK cell activity, did not protect young mice from fatal infections, supporting the in vitro findings. The pathogenesis of Sindbis virus infection in C57BL/6J bg/bg (NK-cell deficient) and bg/+ (NK-cell normal) mice was also studied. Despite a defect in the induction of NK cells by Sindbis virus infection in the bg/bg mice, there were no significant differences in the pathogenesis of either peripheral or intracerebral infection in these strains of mice. These studies suggest that although NK cells are induced, they may not be important in the recovery of mice from Sindbis virus infection.     

Experimental Yersinia enterocolitica infection in mice: Kinetics of growth

Infection of several strains of laboratory mice with a virulent strain of Yersinia enterocolitica was followed by performing viable bacterial counts on homogenates of selected tissues at intervals after intragastric, aerogenic, or intravenous infection. It is observed that CD-1 mice are more susceptible to Y. enterocolitica infection than either the C(57)B1/6 or B6D2 strains. Development of an enteric infection is dose dependent; less than 5 x 10(7) organisms by mouth yields sporadic, low levels of systemic infection, with many of the animals showing no apparent infection. Increasing the challenge inoculum by a factor of 10 eliminates the variability among the animals, giving rise to an enteric infection in all of the mice that moves quickly to the mesenteric lymph node. The bacterial population in the lymph node multiplies rapidly, and the infection is disseminated to the spleen, liver, and lungs, ultimately killing most of the animals. Exposure to an aerogenic challenge of less than 1,000 organisms resulted in a fulminating pneumonitis with an invariably fatal outcome. Intravenous challenge with 500 organisms caused a rapidly fatal, systemic infection. The growth of the bacteria in the intravenously infected mouse depends upon the temperature at which the challenge inoculum had been grown in vitro. At temperatures below 26 C, the bacteria are cleared from the blood at a slower rate and are more resistant to intracellular killing, as compared to organisms grown at 37 C. This effect results in the inoculum increasing to greater numbers in the tissues in a shorter period of time.