Enhancement of natural and experimental respiratory mycoplasmosis in rats by hexamethylphosphoramide.

Hexamethylphosphoramide (HMPA) was given orally (100 mg/kg/day) to: a) conventional rats of Sprague-Dawley and Long-Evans substrains known to have indigenous Mycoplasma pulmonis infection, b) uninfected pathogen-free (PF) Fischer rats, and c) PF and axenic Fischer rats inoculated intranasally with M. pulmonis strains having a wide range of virulence. Treated rats infected with virulent M. pulmonis, either naturally or experimentally, developed severe clinical signs of murine respiratory mycoplasmosis (MRM) with mortalities of 25 to 60% compared to relatively mild MRM and no deaths in untreated, infected controls. Deaths were attributed to unusually severe lung lesions of MRM (extreme neutrophilic exudation into major bronchi and bronchiectasis) with ulceration of respiratory mucosa and hemorrhage. Rhinitis also was increased in severity by HMPA in conventional rats, but not in experimentally infected PF or axenic rats. Severity of otitis media and tracheitis was not affected by HMPA. Incidence of lesions of MRM was unchanged except for increased frequency of gross lung lesions. In the absence of M. pulmonis infection, HMPA treatment of rats caused thinning and microulceration of respiratory epithelium in major bronchi without inflammatory lung disease. Other effects induced by HMPA, with or without the infection, were destruction and fibrous replacement of olfactory epithelium, atrophy of testes, and reduced weight gains. It was concluded that HMPA markedly enhances both rate of progression and severity of the pneumonia while inconsistently potentiating the rhinitis of MRM in rats. Previous studies of HMPA are emphasized as an additional example in which the synergistic effects of an experimental chemical and an indigenous pathogen of laboratory rats have given misleading experimental results.     

Development of an aerosol model of murine respiratory mycoplasmosis in mice.

Animal models of murine respiratory mycoplasmosis due to Mycoplasma pulmonis provide excellent opportunities to study respiratory disease due to an infectious agent. The purpose of the present study was to develop and characterize an aerosol model for the production of murine respiratory mycoplasmosis in mice. The exposure of mice for 30 min to aerosols generated with a DeVilbiss 45 nebulizer in a nose-only inhalation chamber consistently reproduced typical lesions. The chamber was operated with a nebulizer air flow of 5.3 liters/min at 5.0 lb/in and a diluting air flow of 20 liters/min, with the nebulizer containing 5 ml of a suspension of viable M. pulmonis organisms (a concentration between 6 X 10(5) to 6 X 10(10) CFU/ml). Infective aerosol particles of less than a 4.0-micron median aerodynamic diameter with a geometric standard deviation of approximately 2.0 reached the lungs and were evenly distributed among the different lung lobes. A minimum 1.5-log loss of viability in the M. pulmonis suspension was demonstrated. With the exception of the 50% lethal dose, all of the parameters previously established by intranasal inoculation could be examined with the aerosol model. The major advantages of the aerosol model were excellent reproducibility of exposure (both between different experiments and between animals in a given experiment), the avoidance of anesthetization, and the ability to immediately deposit the majority of the organisms in the lung. The only disadvantage was the requirement for large volumes of mycoplasmal cultures.     

Murine respiratory mycoplasmosis in F344 and LEW rats: evolution of lesions and lung lymphoid cell populations.

By comparison of two strains, LEW and F344, which are known to differ in susceptibility to Mycoplasma pulmonis respiratory disease, it was shown that differences in lesion severity and progression were associated with changes in lung lymphocyte populations. Lung lesions in LEW rats developed earlier after infection, became more severe, and were characterized by continued proliferation of all classes of lymphoid cells, T lymphocytes, B lymphocytes, and plasma cells, throughout the 120-day observation period. In contrast, lymphoid proliferation in F344 rats reached a plateau at 28 days and was restricted to an increase in T lymphocytes, immunoglobulin A (IgA)-bearing B lymphocytes, and IgA and IgG plasma cells. Although approximately 10 times as many IgG B cells and 4 times as many IgG plasma cells were found in infected LEW rats as compared with F344 rats, the specific anti-M. pulmonis IgG response in the two strains was roughly parallel. The same relationships held true, although to a lesser extent, for specific IgA antibody responses and cellular responses. Whereas lung lesions showed a tendency to resolve in F344 rats by 120 days, severe lesions persisted in LEW rats. The disparity between the cellular response and specific antibody response, the seemingly uncontrolled lymphocyte proliferation in LEW rats, and the mitogenic potential of M. pulmonis suggest that differences between LEW and F344 rats in lung lesion severity and progression are related to differences in the degree of nonspecific lymphocyte activation in the two strains, an imbalance in regulation of lymphocyte proliferation in LEW rats, or both.     

Intracage ammonia promotes growth of Mycoplasma pulmonis in the respiratory tract of rats.

Ammonia (NH3) from soiled cage bedding is known to enhance the progression and severity of murine respiratory mycoplasmosis in rats. To test the hypothesis that NH3 directly or indirectly enhances the growth of Mycoplasma pulmonis in vivo, pathogen-free F344 rats were inoculated intranasally with 1 x 10(4) to 4 x 10(4) or 4 x 10(6) to 5 x 10(6) colony-forming units of M. pulmonis and exposed to less than or equal to 1.5 or 76 microgram of NH3 per liter (less than or equal to 2 or 100 ppm, respectively). Nasal passages, larynges, tracheas, and lungs from rats killed at intervals up to 28 days after inoculation were quantitatively cultured. Growth of M. pulmonis was much greater in NH3-exposed rats than in controls, particularly in those inoculated with the lower dose. Increases in M. pulmonis populations were more rapid in proximal airways than in distal airways. Serum immunoglobulin G and M antibody responses to M. pulmonis as measured by an enzyme-linked immunosorbent assay were greater in NH3-exposed rats. In other experiments, the nasal passages absorbed virtually all NH3 when the rats were exposed to less than 380 micrograms of NH3 per liter (500 ppm), indicating that NH3 induced increases in the numbers of organisms in the distal respiratory tract, probably by a secondary, rather than a direct, effect. Also, NH3 exposure did not inhibit pulmonary antibacterial activity as measured by clearance of radiolabeled Staphylococcus epidermidis. The growth of M. pulmonis in vitro was inhibited by 1 mM NH4+ added to the medium as NH4OH but not by NH4+ concentrations of 0.5, 0.1, or 0.01 mM, suggesting that NH3 increases growth indirectly through effects on the host.     

Impact of experimental genital mycoplasmosis on pregnancy outcome in Sprague-Dawley rats.

Specific-pathogen-free (SPF) female Sprague-Dawley rats were infected by intravaginal inoculation with 3 x 10(7) CFU of Mycoplasma pulmonis X1048 in 0.1 ml of Frey's broth or with an equal volume of sterile Frey's broth. A minimum of 10 days postinfection, rats were bred to noninfected males. Rats were necropsied at days 11, 14, and 18 of gestation and within 24 h of parturition. Throughout pregnancy, at least 50% of rats remained infected in the lower genital tract. At parturition, the major site of colonization was the respiratory tract (P = 0.02). M. pulmonis was not isolated from any site of any control rat. Pregnancy outcome was adversely affected by infection with M. pulmonis. Infected rats had significantly smaller litter sizes at day 18 of gestation (P < or = 0.01) and at term (P < or = 0.004). No statistically significant differences among the gestational stages in infected rats were noted for litter size. Total litter weight is a reflection of individual pup weight and of the number of pups born. Therefore, it was obvious that infected rats would have a significantly lower (P < or = 0.008) total litter weight than noninfected controls. However, when individual pup weights were considered, infected pups (n = 49) also had significantly lower (P < or = 0.0001) birth weights than did noninfected controls (n = 68). The incidence of an adverse pregnancy outcome at term (stillbirths, macerated fetuses, or resorptions) was higher (P < or = 0.01) in infected rats than in noninfected control rats. No stillborn pups or macerated fetuses were observed in any control term rats (n = 5). All control rats had live-born pups. Three infected rats had no live-born offspring. Resorptions were more common in infected rats than in control rats (P < or = 0.01). The mean number of resorptions per rat was greater in rats which went to term than in rats necropsied during gestation, indicating that the severity of disease was progressive. The rat is frequently the laboratory animal of choice for a wide variety of reproductive studies, and the experimental parameters that are most often measured (litter size, pup weight, and neonatal survival) were all adversely affected by genital mycoplasmosis. Genital mycoplasmosis is important as an animal model for the interaction of infectious agents and the host during pregnancy as well as in its own right as a confounding variable affecting research projects which use the rat as a model to study reproductive function and physiology.     

Morphologic changes in the kidneys of calves with respiratory mycoplasmosis]

The kidneys of 144 naturally diseased calves and those of 47 experimentally infected with Mycoplasma bovirhinis calves were investigated by histological and immunomorphological methods. The kidneys of calves with respiratory mycoplasmosis were often found to be involved in the pathological process, and at the initial stage of the disease and at its height prolifirative, membranous-prolifirative glomerulonephritis, and at later stages proliferative-fibroplastic glomerulonephritis developed. The secondary glanular kidney is a most often outcome of glomerulonephritis. The authors hold that lesions of glomerules were caused by immunopathological mechanisms.     

Nonspecific lymphocyte responses in F344 and LEW rats: susceptibility to murine respiratory mycoplasmosis and examination of cellular basis for strain differences.

Mycoplasma pulmonis produces a mitogen which may play a role in the pathogenesis of murine respiratory mycoplasmosis in rats. Since LEW rats are more susceptible to this disease than F344 rats are, these two strains were used to examine a possible association between disease severity and the level of nonspecific lymphocyte stimulation by mitogens, including M. pulmonis membrane preparations. F344 and LEW spleen, lung, blood, and lymph node lymphocytes were exposed to various mitogens. LEW lymphocytes gave a significantly higher response to mitogenic stimulation, regardless of their anatomical source. These differences in lymphocyte responsiveness were primarily due to differences within the nonadherent cell population. Significantly higher numbers of W3/25+ (T helper) cells were found in LEW lymphoid populations, whereas no difference was found in MRC OX-8+ (T suppressor/cytotoxic) cells. These data suggest an association between disease severity and host responsiveness to nonspecific stimuli.     

Genital mycoplasmosis in rats: a model for intrauterine infection

PROBLEM: Microbial infections of the chorioamnion and amniotic fluid have devastating effects on pregnancy outcome and neonatal morbidity and mortality. The mechanisms by which bacterial pathogens cause adverse effects are best addressed by an animal model of the disease with a naturally-occurring pathogen. METHOD OF STUDY: Intrauterine infection in humans as well as genital mycoplasmosis in humans and rodents is reviewed. We describe a genital infection in rats, which provides a model for the role of infection in pregnancy, pregnancy wastage, low birth weight, and fetal infection. RESULTS: Infection of Sprague-Dawley rats with Mycoplasma pulmonis either vaginally or intravenously resulted in decreased litter size, increased adverse pregnancy outcome, and in utero transmission of the microorganism to the fetus. CONCLUSION: Mycoplasma pulmonis is an ideal model to study maternal genital infection during pregnancy, the impact of infections on pregnancy outcome, fetal infection, and maternal-fetal immune interactions.     

The respiratory response in sensitized rats to aerosol challenge.

Hooded Lister rats were sensitized to DNP-ovalbumin. These rats showed a significant alteration in respiratory pattern when challenged with DNP-ovalbumin aerosol. The respiratory response was not produced by challenging sensitized rats with intragastric DNP-ovalbumin. The magnitude of the respiratory response did not correlate with the specific IgE concentration in the serum. A similar pattern of respiratory response was inducible in unsensitized rats by intravenous administration of IgE-rich sera.     

The role of the medical profession in the environmental arena.

Environmental agencies at both federal and state levels have enormous powers to control economic activities, yet these agencies must use these powers knowing very little about the actual effects of pollutants on human health or the environment. The author describes how this situation came about by reviewing (1) the history of the 19th-century sanitarians and how their traditions (especially of taking preventive action in the absence of definitive data, in order to ensure a margin of safety) later influenced the policies of the U.S. Public Health Service and, more recently, those of the Environmental Protection Agency (EPA); (2) the tradition of recovering damages from someone who harms your property or person; (3) the tradition of engineers to eliminate pollutants without concern for their effects; and (4) the value system of conservationists and ecologists. He then outlines four difficulties that these sometimes-conflicting traditions and values create for the EPA and other similar bodies. After reviewing the progress that has been made despite these difficulties, the author states the global nature of the environmental challenge that is upon us and how the public health tradition of prudent action will compel us to gather more data about global problems, which in turn will lead to more fine-tuned and appropriate actions. Finally, he states how important it is for health professionals to use their technical and scientific knowledge--especially their "habits of mind"--to help develop more intelligent and prudent environmental policies, and describes the crucial role of "citizen-health professionals" in the environmental arena of the future.     

The pathogenesis of respiratory syncytial virus infection in cotton rats.

The cotton rat is susceptible to respiratory synctial virus infection in both the upper and lower portions of the respiratory tract. Virus replicates to high titer in the nose and lungs and to relatively low titer in the trachea. Immunofluorescence studies demonstrated viral antigen in the nasal epithelium and the bronchial and bronchiolar epithelium but not in the trachea or the alveolar cells of the lungs. Histopathologic changes included a desquamative, exudative rhinitis of moderate severity and a mild proliferative bronchiolitis. Serum neutralizing antibody developed in all animals by the ninth day after infection, reaching extremely high titer in several instances. Unlike the previously described response of experimentally infected infant ferrets, cotton rats are uniformly susceptible to pulmonary infection throughout life, thereby offering a model for long-term pulmonary studies heretofore not available.     

Role of phenotypic diversity in pathogenesis of avian mycoplasmosis.

The interactions between avian mycoplasmas and their host cells are far more complex than might be anticipated from their apparent structural and functional simplicity. Phenotypic diversity in the form of reversible phase variation, antigenic variation or size variation is an adaptive mechanism that enables avian mycoplasmas to survive in a hostile and highly evolved host. Despite significant similarities between major membrane antigens of Mycoplasma gallisepticum and Mycoplasma synoviae, the molecular mechanisms that mediate phenotypic variation in these two pathogens are completely different. Throughout the years, these mechanisms have evolved side by side with their host immune system and provided mycoplasmas the capacity to colonize, invade and persist in an intricate host. In this article, recent advances in the understanding of the molecular mechanisms of phenotypic variation are reviewed, and implications of such variation in pathogenesis of the disease and development of vaccines and diagnostic assays are outlined.     

Depletion of alveolar macrophages exacerbates respiratory mycoplasmosis in mycoplasma-resistant C57BL mice but not mycoplasma-susceptible C3H mice.

Indirect evidence suggests that innate immune mechanisms involving alveolar macrophages (AMs) are of major importance in antimycoplasmal defense. We compared the effects of AM depletion on intrapulmonary killing of Mycoplasma pulmonis during the early phase of infection in mycoplasma-resistant C57BL/6NCr (C57BL) and mycoplasma-susceptible C3H/HeNCr (C3H) mice. More than 80% of AMs were depleted in both strains of mice by intratracheal insufflation of liposome-encapsulated dichloromethylene bisphosphonate (L-Cl2MBP), compared to no significant AM depletion in either strain following insufflation of liposome-encapsulated phosphate-buffered saline (L-PBS), PBS alone, or no treatment. AM-depleted (L-Cl2MBP) and control (L-PBS) mice were infected intranasally with 10(5) CFU of M. pulmonis UAB CT, and their lungs were quantitatively cultured to assess intrapulmonary killing at 0, 8, 12, and 48 h postinfection. AM depletion exacerbated the infection in C57BL mice by reducing killing of the organism to a level comparable to that in C3H mice without AM depletion. In contrast, AM depletion did not alter killing in C3H mice. These results directly identify the AM as the main effector cell in early pulmonary antimycoplasmal defense and suggest that differences in mycoplasmal killing by AMs may explain the resistance of C57BL mice and the susceptibility of C3H mice to mycoplasmal infection.     

The role of the pathologist in environmental medicine and public health.

The present paper summarizes the role of the morbid anatomist and clinical pathologist in environmental carcinogenesis. It points out that in the past he has contributed considerably to the identification of rare tumors and their etiology. He has an important role to play in the future in providing more accurate data on which epidemiology studies can be developed. The view is also expressed that it is highly important that modern pathologists have an understanding of toxicologic and pharmacologic techniques and their potential application to biologic material in order that they may be in a position to correlate and develop multidisciplinary approaches to the identification of environmental hazards. Some of these approaches are illustrated and their potential developments outlined.     

The role of eicosanoids in respiratory mucus hypersecretion

The role of eicosanoids in the excessive secretion of respiratory mucous glycoproteins (MGP) accompanying immediate hypersensitivity and inflammatory pulmonary states has only been addressed in the last few years. Three lines of evidence suggest that eicosanoids may participate in the physiologic regulation of MGP secretion as well as being capable of stimulating increased MGP production. First, inhibition of eicosanoid generation with corticosteroids or eicosatetraynoic acid (ETYA) reduces ongoing baseline MGP secretion while selective inhibition of prostaglandin production with nonsteroidal anti-inflammatory agents increases MGP release. Second, arachidonic acid and a variety of eicosanoids stimulate MGP secretion in vitro. In fact, several lipoxygenase pathway metabolites including hydroxyeicosatetraenoic acids (HETEs) and leukotrienes (LTC4 and LTD4) significantly increase MGP secretion in nanogram quantities. Third, a variety of pulmonary cells including airway epithelium, endothelial cells, mast cells, alveolar macrophages and neutrophilic leukocytes generate eicosanoids under conditions that would be encountered in clinical states in which mucus secretion occurs. Thus, mucus secretion during both normal and stimulated states would be influenced by eicosanoids. It seems likely that this eicosanoid-mucus secretion relationship is very important in all forms of asthma, but most particularly in aspirin-related asthma.     

The role of respiratory control disorders in SIDS

Although sudden death in infants resulting from cardiac arrhythmias are well documented these appear to account for no more than 5-10% of SIDS cases. Sudden respiratory failure currently is viewed as the most likely cause of death in the remainder. Accidental asphyxiation appears to have a causal role in less then 50% of deaths diagnosed as SIDS. The rest are most likely do to some form of acute respiratory failure. Although failure of autoresuscitation or failure to arouse from sleep likely contribute to the final sequence of events leading to at least some SIDS deaths, these cannot be regarded as causes of the primary respiratory failure initiating the fatal sequence. Past and current studies provide strong circumstantial evidence that obstructive sleep apnea and/or apnea of prematurity likely account for respiratory failure leading to SIDS in some or many deaths. In drawing conclusions it is well to recognize that mechanisms leading to death in SIDS are heterogeneous and therefore there is room for several plausible theories for respiratory or circulatory abnormalities contributing to SIDS.     

The role of allergy in manifestations of respiratory disease in adult cystic fibrosis.

BACKGROUND: Variability is present in the expression of the clinical phenotype in cystic fibrosis (CF). Part of this variability may be explained by the coexistence of allergy in CF. OBJECTIVE: To determine the rate of allergy in adult CF and evaluate the association between allergy and the manifestations of upper and lower airway disease. METHODS: We performed a cross-sectional study of consecutive patients enrolled in a university hospital adult CF clinic. Allergen specific IgE was determined by radioallergosorbent and skin prick tests to common aeroallergens. We characterized features of upper and lower airway disease by clinical evaluation of rhinitis and spirometry before allergy testing. RESULTS: The study population consisted of 55 patients. Allergen specific IgE was present to at least 1 aeroallergen in 67% by skin prick testing and 80% by radioallergosorbent testing. Rhinitis occurred in 50% of the population and was associated with immediate-type hypersensitivity to aeroallergens other than molds. The frequency of rhinitis increased when there was sensitization to a greater number of aeroallergens and rarely occurred in the absence of allergic sensitization. There was no detectable difference in lung function between those with and without allergic sensitization. CONCLUSIONS: Immediate-type hypersensitivity to aeroallergens commonly occurs in adult CF. The coexistence of allergy in CF is associated with clinical features of rhinitis. Because allergic manifestations of CF warrant appropriate therapy, individuals with CF should be evaluated for coexistent allergy.     

Strain differences in susceptibility to murine respiratory mycoplasmosis in C57BL/6 and C3H/HeN mice.

Not only is murine respiratory mycoplasmosis, due to Mycoplasma pulmonis, a complication of biomedical research, it provides excellent animal models to study the development of a naturally occurring respiratory disease induced by an infectious agent. The understanding of pathogenic mechanisms of disease can be greatly facilitated by studying genetic differences in susceptibility. Five strains of mice with various H-2K haplotypes were examined for their susceptibility to murine respiratory mycoplasmosis; of these, C57BL/6 and C3H/HeN mice were chosen for additional study. There were no significant differences in the incidence of infection in either the upper or lower respiratory tract or in the severity of upper respiratory tract lesions in the two strains as determined at 14 days postinfection. In striking contrast, the C57BL/6 mice were significantly more resistant to the development of gross and microscopic lung lesions and to death due to pneumonia as shown by an almost 100-fold difference in the 50% lethal dose, 50% gross pneumonia dose, and 50% microscopic lesion dose. The most apparent differences in lung lesions between the two strains were in the severity of acute lesions of the bronchial epithelium, the amount of mixed inflammatory response in the alveoli, and the amount of lymphoid infiltrates. All were significantly more severe in C3H/HeN mice. In addition, more C3H/HeN mice developed antibody responses to M. pulmonis. The amount of antibody correlated with lesion severity in both strains.