Bacteriological and histopathological evaluation of guinea pigs after infection with Listeria monocytogenes.

Randomly bred guinea pigs were infected with Listeria monocytogenes using the intracardial, intravenous and intraperitoneal routes of infection. Doses of Listeria ranged from 5 to 1,000 x the 50% lethal dose based on the 50% lethal dose for intracardially injected Listeria. A complete necropsy was performed on all animals that died after infection. Gross and microscopic examination of tissues revealed major pathological features which include myocarditis, edema and congestion with interstitial pneumonitis present in the lungs, and fatty hepatic changes with focal necrosis. For all or a majority of the animals, large numbers of Listeria were likewise recovered from these organs and from lymph nodes, spleen, kidneys, and adrenal gland tissue. Of the three routes of infection used, guinea pigs were most susceptible to Listeria injected via the intracardial route. The relatively high lethal dose of listeric for the quinea pig, however, suggests that the organism is a low-grade pathogen for this species.     

Oral infection with signature-tagged Listeria monocytogenes reveals organ-specific growth and dissemination routes in guinea pigs

Listeria monocytogenes causes a serious food-borne disease due to its ability to spread from the intestine to other organs, a process that is poorly understood. In this study we used 20 signature-tagged wild-type clones of L. monocytogenes in guinea pigs in combination with extensive quantitative data analysis to gain insight into extraintestinal dissemination. We show that L. monocytogenes colonized the liver in all asymptomatic animals. Spread to the liver occurred as early as 4 h after ingestion via a direct pathway from the intestine to the liver. This direct pathway contributed significantly to the bacterial load in the liver and was followed by a second wave of dissemination via the mesenteric lymph nodes (indirect pathway). Furthermore, bacteria were eliminated in the liver, whereas small intestinal villi provided a niche for bacterial replication, indicating organ-specific differences in net bacterial growth. Bacteria were shed back from intestinal villi into the small intestinal lumen and reinfected the Peyer's patches. Together, these results support a novel dissemination model where L. monocytogenes replicates in intestinal villi, is shed into the lumen, and reinfects intestinal immune cells that traffic to liver and mesenteric lymph nodes, a process that occurs even during asymptomatic colonization.     

Effects of diet and genetics on Mycobacterium bovis BCG vaccine efficacy in inbred guinea pigs.

Strain 2 and strain 13 guinea pigs were vaccinated with Mycobacterium bovis BCG and placed on low-protein or protein-adequate diets. Five weeks later all animals were infected by the respiratory route with virulent Mycobacterium tuberculosis H37Rv organisms. Four weeks postchallenge, guinea pigs were skin tested with purified protein derivative and sacrificed. Protein deficiency resulted in significant reductions in body weight and thymus weight and in an impairment in the ability to control the M. bovis BCG vaccine organisms and to mount delayed hypersensitivity reactions. Protein deficiency also adversely affected the efficacy of the BCG vaccine as demonstrated by the numbers of virulent organisms recovered in spleens and lungs. Strain differences were observed in the number of leukocytes, thymus weight, and the responsiveness of blood lymphocytes to purified protein derivative stimulation. In general, strain 13 guinea pigs responded more dramatically to dietary insult than did their strain 2 counterparts. Protein deprivation completely abolished BCG vaccine protection in the lungs and spleens of strain 13 animals and significantly reduced the protection afforded to strain 2 animals. In both strains, the BCG vaccine protected normally nourished guinea pigs. There was no significant difference between strains with respect to susceptibility to pulmonary infection with virulent mycobacteria. Thus, diet and genetic pedigree each had a significant influence on BCG vaccine efficacy.     

Effect of myelopeptide 2 on the immunization of guinea pigs with live measles vaccine

A model is suggested to study the effects the myelopeptide 2 (MP2) immunomodulator on the results of live measles rash immunized by vaccine. Guinea pigs were found to respond to live measles vaccine L-16 the way man does, i.e. the antibodies' level in their sera was increasing until days 28-56 after a single intramuscular vaccine injection. An intramuscular injection of the MP2 immunomodular, made alongside with vaccination, was demonstrated to enhance the immune response to the measles vaccine and to induce its action. Finally, it was for the first time, that the possibility was proven to enhance the action of the live measles vaccine by an immunomodulator.     

Effect of immunosuppression on Rickettsia rickettsii infection in guinea pigs.

The role of the immune response in the pathogenesis of Rickettsia rickettsii infection in guinea pigs was investigated by immunosuppression, using antilymphocyte serum. Twenty guinea pigs were inoculated with R. rickettsii, Sheila Smith strain, on day 0. Fifteen animals received antilymphocyte serum on days --1, 0, 2, 4, and 6. Five animals received normal rabbit serum on the same schedule. At necropsy, specimens were collected for histological examination, rickettsial immunofluorescence, rickettsial titration, and antirickettsial antibody titration. All normal rabbit serum recipients and 12 of 15 antilymphocyte serum recipients developed typical disease. Comparison of animals in terminal stages of disease revealed the same clinical course and gross lesions, but differing rickettsial burden and cellular response. Immunosuppressed animals had higher titers of splenic rickettsiae and greater numbers of immunofluorescent rickettsiae. Thus, although antibody was undetectable in both groups, there appeared to be an inhibition of antirickettsial immunity. Microscopic vasculitis was similar quantitatively, but differed qualitatively, with immunocompetent animals having the typical monouclear/lymphocytic inflammation and immunosuppressed animals having neutrophilic predominance. This study demonstrates that immunopathological mechanisms are not necessary for the pathogenesis of experimental Rocky Mountain spotted fever. The rickettsiae themselves seem capable of causing cellular and tissue damage.     

Hepatic and pulmonary pathology of experimental Fascioloides magna infection in guinea pigs.

The guinea pig was used to study the pathology of Fascioloides magna, an important pathogen for sheep. Although flukes migrated freely through various tissues in infected guinea pigs, the most serious lesions occurred in the liver and lungs. The sequential development of lesions indicated that flukes first invaded the quadrate lobe of the liver and subsequently migrated to other liver lobes and tissues. Six weeks post-infection, there was a marked drop in the recovery of flukes from the liver along with a dramatic increase in pulmonary involvement. Much of the hepatic and pulmonary pathology in infected animals was secondary to extensive vascular lesions caused by migrating flukes. In the liver, vascular lesions predominantly involved the portal and hepatic veins. Thrombophlebitis and locally extensive necrosis, resembling infarction, were observed. Vascular lesions in the lungs occurred in the pulmonary arteries leading to thrombosis and haemorrhagic infarction. Discovery of a fluke in a pulmonary artery, along with the pattern of hepatic and pulmonary lesions, suggested that flukes probably used the cardiovascular system as a pathway for dissemination. Death in fluke-infested guinea pigs was most often associated with severe pulmonary lesions. The nature and distribution of fluke-induced lesions observed in this study demonstrate that the guinea pig is a suitable animal model for Fascioloides magna infection in sheep.     

Release of monokines by pulmonary macrophages following antigen challenge in sensitized guinea pigs

Guinea pigs were passively sensitized with immune serum to ovalbumin (OA), control serum, or saline. Twenty-four hours later, they inhaled aerosols of OA (2% in saline), saline, or lipopolysaccharide (LPS). Following anesthesia, bronchoalveolar lavage (BAL) was performed at 30, 60, 90 and 120 min postinhalation. Alveolar macrophages (AM) were isolated from the BAL fluid and incubated (18 h) in medium alone or with zymosan (1 mg/ml). Supernatants were collected and levels of interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-) determined by bioassays. Unstimulated AM from animals that inhaled OA, saline, or LPS secreted similar amounts of IL-1 at 30, 60, and 90 min postinhalation. Zymosan (1 mg/ml) significantly increased IL-1 secretion by AM collected at 60 and 90 min from OA-sensitized animals that inhaled OA or saline. AM from guinea pigs sensitized to OA that inhaled OA or LPS secreted significantly increased amounts of IL-6 at 30, 60, 90, and 120 min postchallenge compared to saline sensitized controls. In all groups, AM from LPS-treated animals secreted large amounts of TNF- at all sampling times postchallenge; AM from OA-sensitized and challenged animals secreted increasing amounts of TNF- with time postchallenge, spontaneously and in response to zymosan. By contrast, AM from saline sensitized and challenged guinea pigs did not release detectable amounts of TNF- spontaneously and secreted very low amounts in the presence of zymosan. These findings show that antigen challenge results in a rapid activation of AM isolated from BAL and suggest AM may initiate the development of inflammatory processes associated with antigen challenge.     

Effect of L-lysine-A-oxidase on the development of genital herpes infection in guinea pigs

Positive effect of local treatment with L-lysine-a-oxidase on the course of herpes genital infection was demonstrated on guinea pigs even after the appearance of infection symptoms. In animals treated with L-lysine-a-oxidase, the severity of the disease, virus reproduction and virus-induced changes in cells were significantly less pronounced than in untreated animals. The preparations exerted no toxic effects. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 128, No. 12, pp. 654–656, December, 1999.     

Congenital and perinatal infection with Junin virus in guinea pigs

Junin virus infection in guinea pigs is known to be similar to human Argentine hemorrhagic fever (AHF). The guinea pig was chosen as a model for transplacental transmission of Junin virus, as both guinea pig and man have a similar placental structure. Pregnant guinea pigs were infected with the pathogenic XJ strain of Junin virus intramuscularly route at different stages of pregnancy. The group infected during the last third of pregnancy produced 16 newborn, but mortality reached 100%: 18% were born with typical AHF hemorrhagic signs, 54% without signs, and the remainder were stillborn. Virus was recovered from organs of newborns, as well as placental tissues. A second group, infected in the second third of pregnancy, died with intrauterine fetuses, all of which showed hemorrhagic signs and virus present. In a last group, infected in the first third of pregnancy, fetuses were free from macroscopic lesions. In order to determine whether lactation may be an alternative infection route in guinea pigs, mother guinea pigs were infected with Junin virus at different times postparturition. The 84% noninfected newborn housed together with their infected mothers died during the suckling period, half with typical AHF signs. Junin virus transmission from mother to fetus was thus proved, and lactation may be considered as an alternative perinatal infection route.     

Effect of estradiol on chlamydial genital infection of female guinea pigs.

Female guinea pigs were treated daily with 1 mg of beta-estradiol-3-benzoate intramuscularly beginning 14 days before intravaginal inoculation with the chlamydial agent of guinea pig inclusion conjunctivitis and continuing during the course of the infection. Treatment with estradiol was found to markedly influence the course of genital infection with the chlamydial agent of guinea pig inclusion conjunctivitis, producing infections of greater intensity and longer duration than those in control animals. Moreover, pathogenesis was altered in that ascending infection was observed, resulting in endometritis, cystic salpingitis, and cystitis. Infection in the controls was limited to the cervix and vagina. Estradiol treatment increased the apparent number of infected cells in the cervix and vagina as detected by histopathology and immunofluorescent staining. Humoral and cell-mediated immune responses to the chlamydial agent of guinea pig inclusion conjunctivitis were comparable in estradiol-treated and untreated animals. These data indicate that hormonal manipulation may have profound effects on the course of chlamydial genital infections.     

Experimental infection with Treponema hyodysenteriae in guinea pigs.

Outbred and inbred (Hartley strain) guinea pigs (GP) were inoculated intragastrically with pathogenic and nonpathogenic Treponema hyodysenteriae. GP 3 to 16 weeks old received T. hyodysenteriae after a fasting period of 36 to 72 h. Infected GP with pathogenic T. hyodysenteriae developed a diarrheal and/or depressive condition, with mucus but not blood in the feces. Of 88 GP, 40 had gross lesions resembling those of swine dysentery. Lesions were limited mainly to the large intestine. TP used as controls or inoculated with nonpathogenic T. hyodysenteriae did not develop these lesions in the large intestine. These studies suggest that the GP may be used as an animal model for swine dysentery.     

Delayed hypersensitivity and acquired cellular resistance in guinea pigs infected with Listeria monocytogenes.

Randomly bred pigs of both sexes were injected intracardially with one-half of a 50% lethal dose of Listeria monocytogenes. When infected animals were skin tested with 30 mug of a water-soluble extract of sonically disrupted Listeria, both males and females had uniformly detectable levels of delayed hypersensitivity (DH) 4 days after infection. In males, cutaneous hypersensitivity to Listeria antigens reached a peak on day 5 or 6 of infection, and high levels of DH persisted through the 7th week. In females, DH reached a peak on day 6 or 7, remained at this level through the 4th week, and then dropped sharply. Cutaneous reactivity was usually higher for males than for females, and differences between the sexes were statistically significant 5, 6, and 7 weeks after infection. Low levels of DH were still present 41 weeks (females) or 46 weeks (males) after infection. Assays to determine the number of viable Listeria present in spleen homogenates indicated that bacterial multiplication occurred only during the first 24 hours of infection. The number of Listeria declined steadily thereafter, and by day 13 no bacteria could be recovered from the spleens of infected animals. Spleen assays indicated that Listeria-infected animals of both sexes were resistant to a small challenge dose of Listeria given 48 hours, 7 days, or 2 weeks after the primary infection. Resistance to re-infection was absent in females challenged at 41 weeks and in males challenged at 46 weeks.     

Vaccination of guinea pigs with DNA encoding the mycobacterial antigen MPB83 influences pulmonary pathology but not hematogenous spread following aerogenic infection with Mycobacterium bovis

Protection of cattle against bovine tuberculosis by vaccination could be an important control strategy in countries where there is persistent Mycobacterium bovis infection in wildlife and in developing countries where it is not economical to implement a tuberculin test and slaughter control program. The main aim of such a vaccination strategy would be to reduce transmission of infection by reducing the lung pathology caused by infection and preventing seeding of the organism to organs from which M. bovis could be excreted. Recent reports of successful DNA vaccination against Mycobacterium tuberculosis in small-animal models have suggested that DNA vaccines act by reducing lung pathology without sensitizing animals to tuberculin testing. We therefore evaluated the ability of vaccines consisting of DNA encoding the mycobacterial antigens MPB83 and 85A to reduce lung pathology and prevent hematogenous spread in guinea pigs challenged with a low dose of aerosolized M. bovis. Vaccination with MPB83 DNA reduced the severity of pulmonary lesions, as assessed by histopathology, and resembled M. bovis BCG vaccination in this respect. However, unlike BCG vaccination, MPB83 DNA vaccination did not protect challenged guinea pigs from hematogenous spread of organisms to the spleen. In contrast, vaccination with antigen 85A DNA, a promising DNA vaccine for human tuberculosis, had no measurable protective effect against infection with M. bovis.     

Effect of antithymocyte serum on the course of chlamydial genital infection in female guinea pigs

The treatment of female guinea pigs, infected in the genital tract with the chlamydial agent of guinea pig inclusion conjunctivitis, with rabbit anti-guinea pig thymocyte serum extended the course of the infection by 20 to 30 days. The rabbit anti-guinea pig thymocyte serum was shown to suppress delayed hypersensitivity responses to the guinea pig inclusion conjunctivitis agent and the contact allergen oxazolone. The appearance of antibody in genital secretions was delayed, but the infection persisted at low levels even when normal serum and secretory antibody titers were attained, indicating that cell-mediated immunity may play a role in the resolution of chlamydial genital infections.     

Comparison of the protective efficacy of DNA and baculovirus-derived protein vaccines for EBOLA virus in guinea pigs

The filoviruses Ebola virus (EBOV) and Marburg virus (MARV) cause severe hemorrhagic fever in humans for which no vaccines are available. Previously, a priming dose of a DNA vaccine expressing the glycoprotein (GP) gene of MARV followed by boosting with recombinant baculovirus-derived GP protein was found to confer protective immunity to guinea pigs (Hevey et al., 2001. Vaccine 20, 568-593). To determine whether a similar prime-boost vaccine approach would be effective for EBOV, we generated and characterized recombinant baculoviruses expressing full-length EBOV GP (GP(1,2)) or a terminally-deleted GP (GPa-) and examined their immunogenicity in guinea pigs. As expected, cells infected with the GPa- recombinant secreted more GP(1) than those infected with the GP(1,2) recombinant. In lectin binding studies, the insect cell culture-derived GPs were found to differ from mammalian cell derived virion GP, in that they had no complex/hybrid N-linked glycans or glycans containing sialic acid. Despite these differences, the baculovirus-derived GPs were able to bind monoclonal antibodies to five distinct epitopes on EBOV GP, indicating that the antigenic structures of the proteins remain intact. As a measure of the ability of the baculovirus-derived proteins to elicit cell-mediated immune responses, we evaluated the T-cell stimulatory capacity of the GPa- protein in cultured human dendritic cells. Increases in cytotoxicity as compared to controls suggest that the baculovirus proteins have the capacity to evoke cell-mediated immune responses. Guinea pigs vaccinated with the baculovirus-derived GPs alone, or in a DNA prime-baculovirus protein boost regimen developed antibody responses as measured by ELISA and plaque reduction neutralization assays; however, incomplete protection was achieved when the proteins were given alone or in combination with DNA vaccines. These data indicate that a vaccine approach that was effective for MARV is not effective for EBOV in guinea pigs.     

Adjuvant activity of purified peptidoglycan of Listeria monocytogenes in mice and guinea pigs.

The immunological properties of peptidoglycan (L-PG) purified from the cell wall skeleton (L-CWS) of Listeria monocytogenes strain EGD were investigated and compared with the properties of L-CWS. L-PG consisted of alanine, glutamic acid, alpha, epsilon-diaminopimelic acid, muramic acid, and glucosamine. L-PG showed potent adjuvant activities for circulating antibody formation and development of delayed-type hypersensitivity to bacterial alpha-amylase in vivo and for the primary immune response to sheep erythrocytes in vitro, as well as L-CWS. Both L-PG and L-CWS enhanced the generation of cell-mediated cytotoxicity in allogeneic mice and activated thioglycolate-elicited peritoneal macrophages and macrophage cell line RAW 264 to kill tumor target cells in vitro. We also found that L-PG acted on normal spleen cells as a mitogen. Both L-PG and L-CWS had tumor (Meth A)-suppressive and -regressive activities in syngeneic mice. Our results suggest that the L-PG moiety retains the adjuvant and antitumor activities of L-CWS.     

Bronchial sensitization in guinea pigs following ingestion of ovalbumin

Local bronchial mucosal hypersensitivity following antigen feeding was studied in the guinea pig. Groups of 6 animals were fed 1% ovalbumin (OA) in tap water or tap water without antigen (control group) for different feeding periods (14, 28, 42, and 56 days). Inhalative provocations with increasing concentrations of OA (0.5-8% OA) were performed at the end of each feeding period followed by body plethysmographic measurement of airway obstruction. Specific bronchial hypersensitivity to inhaled OA was not found in the control group, whereas specific bronchial reactivity to OA, described as reactivity index, was significantly different from the control group after 14 (p less than 0.05), 28 (p less than 0.001) and 42 days (p less than 0.01) of feeding. No difference to the control group was found after 56 days of feeding. Anti-OA IgG total and IgG1 in serum and bronchoalveolar lavage fluid were increased in OA-fed animals reaching maximal concentrations at day 28 of the feeding period. We conclude that oral feeding of a 1% solution of OA can induce a transient state of local hypersensitivity to inhaled antigen in the guinea pig as manifested by bronchoconstriction on OA inhalation and increased concentrations of local and systemic specific antibodies. This period of local hypersensitivity is followed by tolerance.