TI: Dexamethasone and oxytetracycline reverse the potentiation of neurogenic inflammation in airways of rats with mycoplasma-pulmonis infection 
AUTHOR:BOWDEN_JJ, SCHOEB_TR, LINDSEY_JR, MCDONALD_DM 
JOURNAL: AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE, 1994, Vol.150, No.5, pp.1391-1401 
AB: Mycoplasma pulmonis infection in rats causes a chronic 
 inflammatory airway disease. Along with extensive remodeling of 
 the airway mucosa, lymphocytic infiltrates, angiogenesis, and 
 mucosal thickening, there is an abnormal sensitivity of the 
 blood vessels to mediators that evoke ''neurogenic 
 inflammation.'' As a result, substance P, a peptide released 
 from sensory nerves, produces an unusually large amount of 
 plasma leakage. These changes can be prevented or reduced by 
 prophylactic treatment with antibiotics, but it is unknown 
 whether the extensive remodeling of the airway mucosa and 
 potentiation of neurogenic inflammation can be reversed once 
 they are established. We addressed this issue in F344 rats that 
 were infected with M. pulmonis at 8 wk of age. Six weeks later, 
 the rats were treated daily with an antibiotic 
 (oxytetracycline, 20 mg/kg intramuscularly), to reduce the 
 number of infecting organisms, or with an antiinflammatory 
 steroid (dexamethasone, 0.5 mg/kg intraperitoneally), to reduce 
 the inflammatory and immunologic response to the infection. 
 Sham-treated infected rats received daily injections of 0.9% 
 NaCl. After 1, 2, or 4 wk of treatment the rats were 
 anesthetized and then challenged with substance P (5 mu g/kg 
 intravenously). The sham-treated rats had pathologic changes in 
 their airways typical of severe M. pulmonis infection, and had 
 as much as a threefold increase in substance P-induced plasma 
 leakage. By comparison, after 4 wk of treatment with 
 oxytetracycline or dexamethasone the chronic inflammation was 
 nearly resolved and the response to substance P was in the 
 normal range. Unexpectedly dexamethasone, like oxytetracycline, 
 reduced the number of infecting organisms. We conclude that the 
 potentiation of neurogenic inflammation and many of the other 
 changes associated with the chronic airway disease produced by 
 M. pulmonis infection can be reversed by antibiotics or 
 antiinflammatory steroids. 
KP: MURINE RESPIRATORY MYCOPLASMOSIS, SENDAI VIRUS-INFECTION, NECROSIS-FACTOR-ALPHA, RAT TRACHEA, NEUTRAL ENDOPEPTIDASE, PLASMA EXTRAVASATION, SUBSTANCE-P, LYMPHOCYTE-T, F344 RATS, LEW RATS 

Mycoplasma-pulmonis 46-kda trypsin-resistant protein adheres to rat tracheal epithelial-cells 
AUTHOR: BEYERS_TM, LAI_WC, READ_RW, PAKES_SP 
JOURNAL: LABORATORY ANIMAL SCIENCE, 1994, Vol.44, No.6, pp.573-578 
AB: Results of competitive binding studies with radiolabeled and 
unlabeled Mycoplasma pulmonis and rat tracheal explant cultures 
indicated no effect of trypsin treatment on the ability of M. 
pulmonis to bind to explants. A trypsin-resistant 46-kDa 
membrane protein, which binds isolated rat tracheal epithelial 
cells in culture, was purified and used to produce specific 
antibodies that block adhesion of mycoplasmas to tracheal 
explants. These results suggest that M. pulmonis adheres to rat 
tracheal epithelium via a trypsin-resistant 46-kDa protein. 
KP: RESPIRATORY EPITHELIUM, PNEUMONIAE, GALLISEPTICUM, ERYTHROCYTES, ATTACHMENT 

Effect of time of exposure to rat coronavirus and mycoplasma-pulmonis on respiratory-tract lesions in the wistar rat 
AUTHOR: SCHUNK_MK, PERCY_DH, ROSENDAL_S 
JOURNAL:  CANADIAN JOURNAL OF VETERINARY RESEARCH-REVUE CANADIENNE DE RECHERCHE VETERINAIRE, 1995, Vol.59, No.1, pp.60-66 
AB: The effects of time of exposure on the progression of pulmonary 
lesions in rats inoculated with Mycoplasma pulmonis and the rat 
coronavirus, sialodacryoadenitis virus (SDAV) were studied, 
using six groups of 18 SPF Wistar rats (n = 108). Rats were 
inoculated intranasally as follows: Group 1, sterile medium 
only; Group 2, sterile medium followed one week later by 150 
TCID50 SDAV; Group 3, sterile medium followed by 10(5.7) colony 
forming units of Ill. pulmonis; Group 4, SDAV followed one week 
later by ill. pulmonis; Group 5, Ill. pulmonis followed one 
week later by SDAV; Group 6, M. pulmonis followed two weeks 
later by SDAV. Six rats from each group were euthanized at one, 
two and three weeks after the final inoculation. In a separate 
experiment, six additional animals were inoculated in each of 
groups 3, 5 and 6 (n = 18) and were sampled at five weeks after 
they had received Ill. palmonis. Bronchoalveolar lavage and 
quantitative lung mycoplasma cultures were conducted on two- 
thirds of the rats. Histopathological examination and scoring 
of lesion severity were performed on all animals. Based on the 
prevalence and extent of histopathological lesions, 
bronchoalveolar lavage cell numbers, neutrophil differential 
cell counts and the isolation of M. pulmonis, the most severe 
disease occurred in the groups that received both agents. There 
was no significant difference in lesion severity between the 
groups receiving both agents other than in those examined 
during the acute stages of SDAV infection. Based on these 
results, it is evident that SDAV enhances lower respiratory 
tract disease in Wistar rats whether exposure occurs at one 
week prior to or at various intervals following M. pulmonis 
infections. 
KP: VIRUS-INFECTION, SENDAI VIRUS, SIALODACRYOADENITIS, EXACERBATION, CALVES 

TI: Effects of aging on antibody avidity to mycoplasma-pulmonis 
AUTHOR: STEFFEN_MJ, EBERSOLE_JL 
JOURNALMECHANISMS OF AGEING AND DEVELOPMENT, 1995, Vol.78, No.2, pp.123-144 
AB: Antibody avidity of serum and lung lavage responses was 
examined in rats to determine aging effects on functional 
differences in antibody to Mycoplasma pulmonis. Three age 
groups of animals (weanling, adult and senescent) were 
immunized with either of two doses of formalinized M. pulmonis 
as the antigen, or a placebo control. Total immunoglobulin 
levels and specific antibody responses were examined in serum 
and lung lavage fluids and subsequently avidity measurements of 
the same samples were made for the specific antibody to M. 
pulmonis. The concentration of NH4SCN that dissociated 50% of 
the antibody was used to determine the avidity index of the 
serum and lung lavage samples. Total serum IgG and IgA were 
decreased in the weanling animals when compared to the other 
two age groups of animals. In contrast, serum IgA levels were 
substantially increased in senescent animals. Significant 
increases in serum IgA levels were noted following immunization 
that was not observed for IgG levels. Substantial increases in 
both serum antibody and lung lavage antibody were observed in 
response to immunization with either dose of antigen, but only 
the lung lavage samples showed both IgG and IgA isotypes 
differences that were attributable to age. Serum IgG avidity 
indices gradually increased over time following immunization 
with higher indices being observed in the weanling animals 
immunized with the higher M. pulmonis dose. Serum IgA avidity 
indices also increased over time with no significant 
differences noted among the age groups. Lung lavage IgG avidity 
demonstrated slightly higher indices in the weanling animals, 
while lung lavage IgA avidity showed higher avidity indices in 
the senescent animals at the higher antigen dose. These data 
suggest that senescent animals are capable of producing an 
apparently functional antibody response and that differences 
noted in increased disease susceptibility in older animals may 
be attributed to mechanisms other than a dysfunctional humoral 
immune response at mucosal surfaces. 
KP: IMMUNOGLOBULIN-G ANTIBODY, AGED MICE, IMMUNE-RESPONSES, INTERLEUKIN-2 PRODUCTION, LYMPHOID-TISSUES, CHOLERA-TOXIN, IGG-AVIDITY, T-CELLS, INFECTION, VIRUS 
WA: MYCOPLASMA PULMONIS, ANTIBODY AVIDITY, SERUM IGG, SERUM IGA 

Protection against mycoplasma-pulmonis infection by genetic vaccination 
AUTHOR: LAI_WC, BENNETT_M, JOHNSTON_SA, BARRY_MA, PAKES_SP 
JOURNAL: DNA AND CELL BIOLOGY, 1995, Vol.14, No.7, pp.643-651 
AB: The induction of an immune response against a foreign protein 
usually requires purification of that protein, which is 
injected into animals. The isolation of a pure protein is time 
consuming and costly. Recently, a technique called biolistic 
transformation (biological ballistic system) microparticle 
injection, gene gun, or particle bombardment was developed. The 
basic idea is that the DNA or biological material coated onto 
heavy tungsten or gold particles is shot into target cells or 
animals. We have vaccinated mice by introducing the gene 
(Mycoplasma pulmonis DNA or a specific fragment) encoding a 
protein recognized by a protective monoclonal antibody directly 
into the skin or muscle of mice by two methods: (i) using a 
hand-held form of the biolistic system that can propel DNA- 
coated gold microprojectiles (2 mu g of DNA) directly into the 
skin; (ii) using a conventional intramuscular injection of the 
DNA (100 mu g) into quadricep muscles of transfected mice. HeLa 
cells were transfected in vitro by the gene gun or by the 
liposomal delivery system. Indirect immuno-fluorescent antibody 
(IFA) assay of culture cells indicated that both methods could 
be successful. Production of antibody and cell-mediated 
immunity against M. pulmonis were monitored by assaying serum 
IFA and enzyme-linked immunosorbent assay (ELISA), and delayed 
type hypersensitivity. In addition, macrophage migration 
inhibition and lymphocyte transformation to antigen in spleen 
cells were also tested. Both delivery systems induced humoral 
and cellular immunity, and vaccinated the mice against 
infection. Genetic immunization by using the gene gun saves 
time, money, and labor; moreover, this general method is also 
applicable to gene therapy. 
KP: IMMUNODEFICIENCY-VIRUS TYPE-1, PLASMID DNA, IMMUNE-RESPONSES, MUSCLE INVIVO, MOUSE MUSCLE, MICE, IMMUNIZATION, EXPRESSION,INJECTION, INFLUENZA 
 

Natural uterine mycoplasma-pulmonis infection in female rats 
AUTHOR: BUSCH_K, NAGLIC_T 
JOURNAL: VETERINARNI MEDICINA, 1995, Vol.40, No.8, pp.253-255 
AB: Uterine washings from 124 apparently healthy and non-pregnant 
female Wistar rats of different ages were cultured for 
mycoplasmas and bacteria. The animals originated from four 
conventional breeding colonies which were known to be 
chronically infected with M. pulmonis from the previous 
microbiological examination. Mycoplasmas were isolated from the 
uterus in 30.6% of examined females. All the isolates were 
biochemically and serologically identified as M. pulmonis. 
Uterine colonization with this organism was first evidenced in 
non-mated female rats at the age of three months. After mating 
the number of infected females rapidly increased. This 
observation points out the microbiologically uncontrolled 
mating as an important factor in the distribution of genital 
infection within the colony. Bacterial examination of uterine 
washings revealed only ubiquitous organisms in some animals. 
Gross lesions in the form of purulent salpingitis and mild 
endometritis were observed only in two animals. 
      WA: MYCOPLASMA PULMONIS, UTERINE INFECTION, RAT, WISTAR 

Changes in pulmonary calcitonin-gene-related peptide and protein-gene-product-9.5 innervation in rats infected with mycoplasma-pulmonis 
AUTHOR: NOHR_D, BUOB_A, GARTNER_K, WEIHE_E 
JOURNAL: CELL AND TISSUE RESEARCH, 1996, Vol.283, No.2, pp.215-219 
AB: Changes in the expression of calcitonin gene-related peptide 
(CGRP) and polyneural protein gene product 9.5 (PGP) in hilar 
peribronchial innervation was investigated by 
immunohistochemistry in specific pathogen-free rats chronically 
infected with Mycoplasma pulmonis. Image analysis of 
immunostained sections revealed a reduction of approximately 
62% in the amount of CGRP- and PGP-immunoreactive innervation 
of the peribronchial area in the infected animals. The portion 
of the total bronchial perimeter occupied by bronchus- 
associated lymphoid tissue was increased six-fold. The decrease 
in the CGRP-immunoreactive area could be the result either of 
an enhanced CGRP release or of a loss of nerve fibres. The 
decrease in the PGP-immunoreactive fibres indicates a 
degenerative loss of nerves. Increased bronchus-associated 
lymphoid tissue and decreased bronchial innervation by neurons 
releasing the immunomodulatory neuropeptide CGRP might both 
contribute to the pathophysiology and symptoms of mycoplasmosis 
in the rat. 
KP: SUBSTANCE-P, LYMPHOCYTES, CELLS, INFLAMMATION, ANTIBODIES, RECEPTORS, CGRP 
WA: LUNG, NEURO-IMMUNE LINK, CALCITONIN GENE-RELATED PEPTIDE, PROTEIN GENE PRODUCT 9.5, BRONCHUS-ASSOCIATED LYMPHOID TISSUE, IMMUNOHISTOCHEMISTRY, MYCOPLASMA PULMONIS, RAT (LEW/ZTM) 

Sensory denervation by neonatal capsaicin treatment exacerbates mycoplasma-pulmonis infection in rat airways 
AUTHOR:: BOWDEN_JJ, BALUK_P, LEFEVRE_PM, SCHOEB_TR, LINDSEY_JR, MCDONALD_DM 
JOURNAL:  AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY, 1996, Vol.14, No.3, pp.L 393-L 403 
AB: Mycoplasma pulmonis infection in rats results in life-long 
disease, characterized by chronic inflammation of the airway 
mucosa with widespread accumulation of lymphoid tissue, mucous 
cell hyperplasia, and mucosal thickening. In addition, there is 
angiogenesis and increased sensitivity of mucosal blood vessels 
to substance P (SP), so tachykinins released from sensory nerve 
fibers cause an abnormally large amount of plasma leakage. We 
sought to learn whether the sensory nerves influence the 
severity of the chronic inflammatory response of M. pulmonis 
infection. Our strategy was to destroy the nerves by capsaicin 
pretreatment at birth, infect the rats with M. pulmonis at 8 wk 
of age, and then study the animals 6 wk later. We found that 
capsaicin pretreatment increased the severity of the infection, 
exaggerated the pathological changes in the tracheal mucosa, 
and increased the amount of SP-induced plasma leakage, as 
quantified with Monastral blue. The thickness of the tracheal 
mucosa in these infected rats was 80% greater than in their 
vehicle-pretreated counterparts and 200% greater than in the 
pathogen-free controls. The area density of Monastral blue- 
labeled blood vessels averaged 20% in the infected rats 
pretreated with capsaicin, which represented a 40-fold increase 
over the leakage in the pathogen-free group. By comparison, the 
amount of Monastral blue labeling was only 13% in rats 
pretreated with vehicle (P < 0.05), which was a 22-fold 
increase over the corresponding pathogen-free group. The number 
of SP-immunoreactive nerves fibers was reduced both by neonatal 
capsaicin and by infection (87 and 63% reductions, 
respectively); but when the two conditions were combined, their 
effects were not additive (79% reduction), perhaps because of 
nerve regrowth. We conclude that destruction of sensory nerves 
increases the severity of infection-induced chronic 
inflammation in the airway mucosa, with exaggerated mucosal 
thickening, angiogenesis, plasma leakage, and nerve remodeling. 
KP: MURINE RESPIRATORY MYCOPLASMOSIS, SUBSTANCE-P, NEUROGENIC INFLAMMATION, TRACHEAL EPITHELIUM, IMMUNE-RESPONSE, VIRUS-INFECTION, NERVE-FIBERS, F344 RATS, TRACT, NEUROPEPTIDES 
WA: ANGIOGENESIS, CHRONIC INFLAMMATION, IMMUNOHISTOCHEMISTRY, NERVE REGROWTH, PGP 9.5, PLASMA LEAKAGE, SENSORY NERVES, SUBSTANCE P 

Experimental genital mycoplasmosis - time of infection influences pregnancy outcome 
 AUTHOR: BROWN_MB, STEINER_DA 
JOURNAL: INFECTION AND IMMUNITY, 1996, Vol.64, No.6, pp.2315-2321 
AB: Genital infection of rats with Mycoplasma pulmonis causes 
adverse pregnancy outcome and can result in in utero spread of 
infection to the fetus. The current study was designed to 
determine whether the stage of pregnancy when infection occurs 
influences pregnancy outcome. Rats were inoculated with 3 x 
10(7) CFU of M. pulmonis at 10 days prior to breeding (-10) or 
at gestational day (gd) 11 or 14 and were necropsied at gd 11, 
14, or 18 or within 24 h of parturition (term). Control rats 
received sterile broth, M. pulmonis was isolated from the 
placenta, amniotic fluid, or fetal tissues only from rats 
infected prior to breeding (P < 0.001), All infected rats had 
significantly more loss of pups than did control rats (P < 
0.006), but rats infected prior to breeding or at the beginning 
of the third trimester (gd 14) were much more likely to have 
fetal losses. Rats infected in the early second trimester after 
implantation (gd 11) did not experience severe losses, Litter 
sizes, total litter weight, and individual pup weight from all 
infected rats, regardless of gestational stage when infected, 
were significantly smaller than those of control rats (P < 
0.001). On the basis of the results of this study, we conclude 
that the time of infection plays a major role in determination 
of pregnancy outcome and spread of infection from the genital 
tract to the respiratory tract. 
KP: SPRAGUE-DAWLEY RATS, PULMONIS, TRACT, HOST 

Detection of Mycoplasma pulmonis in cilia-associated respiratory Bacillus isolates and in respiratory tracts of ratsby nested PCR 
AUTHOR: Schoeb_TR, Dybvig_K, Keisling_KF, Davidson_MK, Davis_JK 
JOURNAL:  JOURNAL OF CLINICAL MICROBIOLOGY, 1997, Vol.35, No.7, pp.1667-1670 
AB: To improve the detection of Mycoplasma pulmonis contamination 
of isolates of cilia-associated respiratory (CAR) bacillus, we 
developed a nested PCR method using primers for 16S rRNA gene 
sequences, Of 140 samples of 16 different CAR bacillus 
isolates, 73 (52%) were inhibitory in the first PCR as 
indicated by the absence of amplicons of the internal control, 
but only 11 of 140 (7.9%) were inhibitory in the second PCR Of 
27 samples known to contain M. pulmonis, only 12 (44%) were 
positive in the first PCR, but 25 of 27 (93%) were positive in 
the second PCR, Nested PCR also detected M. pulmonis in 21 of 
61 (34%) CAR bacillus samples from which M. pulmonis could not 
be cultured and identified 2 additional M. pulmonis- 
contaminated CAR bacillus isolates, Of 359 respiratory and 
reproductive tract lavage samples from rats and mice, 35 (9.8%) 
were inhibitory in the first PCR, but only 15 (4.2%) were 
inhibitory in the second PCR, Of 72 lavage specimens from rats 
inoculated with an avirulent, poorly infective M. pulmonis 
strain, 14 (19%) were positive by nested PCR, but only 2 of 72 
(2.8%) were positive by culture. Nested PCR also detected M. 
pulmonis in 14 of 20 (70%) paraffin sections of lung and 
trachea from rats and mice inoculated with CAR bacillus 
isolates known to contain M. pulmonis, whereas single PCR gave 
no positive results, We conclude that nested PCR is superior to 
single PCR or culture for detecting M. pulmonis, and that M. 
pulmonis is present in all but four CAR bacillus isolates in 
our collection that were from naturally infected rats; the four 
isolates that were exceptions were obtained from rats from a 
single colony. 
      KP: POLYMERASE CHAIN-REACTION 

Upregulation of substance P receptors in angiogenesis associated with chronic airway inflammation in rats 
AUTHOR: Baluk_P, Bowden_JJ, Lefevre_PM, McDonald_DM 
JOURNAL:  AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY, 1997, Vol.17, No.3, pp.L565-L571 
AB: In rat airways, substance P released from sensory nerves 
 induces plasma leakage via neurokinin-1 (NK1) receptors on 
 endothelial cells. In pathogen-free rats, both leakage and 
 endothelial NK1 receptors are most abundant in postcapillary 
 venules. In Mycoplasma pulmonis-infected rats, extensive 
 angiogenesis occurs in the tracheal mucosa. The capillary-sized 
 (<10 mu m in diameter) angiogenic blood vessels are abnormally 
 sensitive to substance P. The aim of this study was to 
 determine whether increased expression of NK1 receptors 
 contributes to this abnormal sensitivity. Fischer 344 rats were 
 infected with M. pulmonis and were challenged with substance P 
 (5 mu g/kg iv), and then plasma leakage in the tracheal mucosa 
 was measured by extravasation of Monastral blue (30 mg/kg iv). 
 NK1 receptors on endothelial cells were localized by 
 immunohistochemistry. Five minutes after substance P, NK1 
 receptor-immunoreactive endosomes were five times more abundant 
 in endothelial cells of angiogenic capillaries in M. pulmonis- 
 infected rats than in corresponding capillaries in pathogen- 
 free controls (17.1 +/- 2.3 vs. 3.5 +/- 0.4 endosomes/100 mu 
 m(2) of endothelial surface). Endosomes were slightly more 
 abundant in postcapillary venules 15-35 mu m in diameter in 
 infected rats (23.0 +/- 0.6 vs. 19.2 +/- 0.7 endosomes/100 mu 
 m(2)). Similarly, after substance P, angiogenic capillaries had 
 much more Monastral blue labeling (area density: 18.8 +/- 1.5 
 vs. 2.9 +/- 0.5% of vessel wall), whereas postcapillary venules 
 had about the same amount of labeling (36.0 +/- 3.7 vs. 34.1 
 +/- 1.8%). We conclude that increased expression of NK1 
 receptors, which are internalized into endosomes after ligand 
 binding, contributes to the abnormal sensitivity of endothelial 
 cells of angiogenic blood vessels to substance P in the airways 
 of M. pulmonis-infected rats. 
      KP: MURINE RESPIRATORY MYCOPLASMOSIS, NECROSIS-FACTOR-ALPHA, NEUROGENIC INFLAMMATION, GENE-EXPRESSION, VIRUS-INFECTION, MESSENGER-RNAS, NK1 RECEPTOR, SPINAL-CORD, PULMONIS, POTENTIATION 
WA: plasma leakage, vascular permeability, Mycoplasma pulmonis  infection, trachea, endosomes, tachykinins, bronchus-associated lymphoid tissue, neurokinin-1 receptors