Organization, barrier function and antimicrobial lipids of the oral mucosa.

As one moves from the skin across the vermilion region of the lip and into the oral cavity, the oral mucosa is encountered. The oral mucosa consists of connective tissue known as the lamina propria covered by a stratified squamous epithelium. In the regions of the hard palate and gingiva, the epithelium is keratinized like the epidermis. In the buccal region, the floor of the mouth and the underside of the tongue, the epithelium is non-keratinized. The epithelium on the dorsum of the tongue is a specialized epithelium, but can be approximated as a mosaic of keratinized and non-keratinized epithelia. The non-keratinized epithelial regions do not produce a stratum corneum. Nuclei with intact DNA are retained in the superficial cells. In all regions, the outer portions of the epithelium provide a protective permeability barrier, which varies regionally. Antimicrobial lipids at the surfaces of the oral mucosa are an integral part of innate immunity.

Sphingoid bases are taken up by Escherichia coli and Staphylococcus aureus and induce ultrastructural damage.

Sphingoid bases found in the outer layers of the skin exhibit antimicrobial activity against gram-positive and gram-negative bacteria. We investigated the uptake of several sphingoid bases by Escherichia coli and Staphylococcus aureus, and assessed subsequent ultrastructural damage. E. coli and S. aureus were incubated with D-sphingosine, dihydrosphingosine, or phytosphingosine at ten times their MIC for 0.5 and 4 h, respectively, to kill 50% of viable bacteria. Treated bacterial cells were immediately prepared for SEM, TEM, and analyzed for lipid content by QTLC. E. coli and S. aureus treated with sphingoid bases were distorted and their surfaces were concave and rugate. Significant differences were observed in the visual surface area relative to controls for both E. coli and S. aureus when treated with dihydrosphingosine and sphingosine (p < 0.0001) but not phytosphingosine. While sphingoid base-treated S. aureus exhibited disruption and loss of cell wall and membrane, E. coli cytoplasmic membranes appeared intact and the outer envelope uncompromised. Both E. coli and S. aureus cells contained unique internal inclusion bodies, likely associated with cell death. QTLC demonstrated extensive uptake of sphingoid bases by the bacteria. Hence, sphingoid bases induce both extracellular and intracellular damage and cause intracellular inclusions that may reflect lipid uptake.

The emerging role of peptides and lipids as antimicrobial epidermal barriers and modulators of local inflammation.

Skin is complex and comprised of distinct layers, each layer with unique architecture and immunologic functions. Cells within these layers produce differing amounts of antimicrobial peptides and lipids (sphingoid bases and sebaceous fatty acids) that limit colonization of commensal and opportunistic microorganisms. Furthermore, antimicrobial peptides and lipids have distinct, concentration-dependent ancillary innate and adaptive immune functions. At 0.1-2.0 µM, antimicrobial peptides induce cell migration and adaptive immune responses to coadministered antigens. At 2.0-6.0 µM, they induce cell proliferation and enhance wound healing. At 6.0-12.0 µM, they can regulate chemokine and cytokine production and at their highest concentrations of 15.0-30.0 µM, antimicrobial peptides can be cytotoxic. At 1-100 nM, lipids enhance cell migration induced by chemokines, suppress apoptosis, and optimize T cell cytotoxicity, and at 0.3-1.0 µM they inhibit cell migration and attenuate chemokine and pro-inflammatory cytokine responses. Recently, many antimicrobial peptides and lipids at 0.1-2.0 µM have been found to attenuate the production of chemokines and pro-inflammatory cytokines to microbial antigens. Together, both the antimicrobial and the anti-inflammatory activities of these peptides and lipids may serve to create a strong, overlapping immunologic barrier that not only controls the concentrations of cutaneous commensal flora but also the extent to which they induce a localized inflammatory response.

Streptococcus mutans strains recovered from caries-active or caries-free individuals differ in sensitivity to host antimicrobial peptides.

Antimicrobial peptides (AMPs) are among the repertoire of host innate immune defenses. In the oral cavity, several AMPs are present in saliva and have antimicrobial activities against oral bacteria, including Streptococcus mutans, a primary etiological agent of dental caries. In this study, we hypothesized that unique S. mutans strains, as determined by DNA fingerprinting from sixty 13-year-old subjects with or without experience of caries, would have different susceptibilities to α-defensins-1-3 (HNP-1-3), ß-defensins-2-3 (HBD-2-3) and LL-37. The salivary levels of these peptides in subjects were also measured by enzyme-linked immunosorbent assays. We found that S. mutans strains from children with active caries showed greater resistance to salivary HNP-1-2, HBD-2-3 and LL-37 at varying concentrations than those from caries-free subjects. In addition, combinations of these peptides increased their antimicrobial activity against S. mutans either additively or synergistically. The salivary levels of these peptides were highly variable among subjects with no correlation to host caries experience. However, the levels of a number of these peptides in saliva appeared to be positively correlated within an individual. Our findings suggest that the relative ability of S. mutans to resist host salivary AMPs may be considered a potential virulence factor for this species such that S. mutans strains that are more resistant to these peptides may have an ecological advantage to preferentially colonize within dental plaque and increase the risk of dental caries.

A multiplex immunoassay demonstrates reductions in gingival crevicular fluid cytokines following initial periodontal therapy.

BACKGROUND AND OBJECTIVE: Cytokines and chemokines play an important role in the pathogenesis of periodontal diseases. The objective of this study was to quantitatively assess the effect of initial periodontal therapy on gingival crevicular fluid levels of a comprehensive panel of cytokines and chemokines, including several less extensively studied mediators. MATERIAL AND METHODS: Clinical examinations were performed and gingival crevicular fluid samples obtained from six subjects with generalized severe chronic periodontitis prior to initial periodontal therapy and at re-evaluation (6-8 weeks). Four diseased and two healthy sites were sampled in each subject. Twenty-two gingival crevicular fluid mediators were examined using a multiplex antibody capture and detection platform. Statistical analyses were performed by fitting mixed effects linear models to log-transformed gingival crevicular fluid values. RESULTS: Gingival crevicular fluid interleukin (IL)-1alpha and IL-1beta were the only cytokines to differ in initially diseased vs. initially healthy sites. Following initial therapy, 13 of the 16 detectable cytokines and chemokines decreased significantly in diseased sites, including IL-1alpha, IL-1beta, IL-2, IL-3, IL-6, IL-7, IL-8, IL-12 (p40), CCL5/regulated on activation, normally T cell expressed and secreted (RANTES), eotaxin, macrophage chemotactic protein-1, macrophage inflammatory protein-1alpha and interferon-gamma. At healthy sites, only three of the 16 mediators were significantly altered following therapy. CONCLUSION: This is the first study, to our knowledge, to evaluate such an extensive panel of gingival crevicular fluid mediators within the same sample prior to and following initial therapy. The results confirm that periodontal therapy effectively reduces pro-inflammatory cytokines and chemokines, including less well-described mediators that may be important in initiation and progression of periodontitis. The multiplex assay will prove useful for future gingival crevicular fluid studies.

Degradation of human alpha- and beta-defensins by culture supernatants of Porphyromonas gingivalis strain 381.

Porphyromonas gingivalis produces proteases capable of degrading cytokines, host heme proteins and some antimicrobial peptides. In this study, we show that P. gingivalis culture supernatants fully or partially degrade human neutrophil peptide alpha-defensins and human beta-defensins after 30 min. This observation suggests that proteases from P. gingivalis degrade defensins and this activity could abrogate defensin-related innate immune functions.

SMAP29 congeners demonstrate activity against oral bacteria and reduced toxicity against oral keratinocytes.

INTRODUCTION: Cathelicidins are antimicrobial peptides found in epithelial and mucosal tissues as well as the secondary granules of neutrophils. SMAP29, a sheep cathelicidin, has differential antimicrobial properties against various pathogens, including periodontal organisms. The purpose of this study was to evaluate the antimicrobial properties and cytotoxicity of SMAP29, SMAP28, and three congeners (SMAP18A, SMAP18D, and SMAP14A). METHODS: The peptides at concentrations ranging from 0.25 to 250 microg/ml were tested for their activity against multiple strains of Streptococcus mutans, Streptococcus sanguis, Actinomyces israelii, Actinomyces naeslundii, Actinobacillus actinomycetemcomitans, Fusobacterium nucleatum, Peptostreptococcus micros, and Porphyromonas gingivalis using a radial diffusion assay. Cytotoxicity of keratinocytes was evaluated by measuring lactate dehydrogenase release after incubation with the individual peptides. RESULTS: SMAP28, thought to be the biologically active peptide, was the most potent antimicrobial (range of minimum inhibitory concentrations 0.06-7.03 microg/ml, P < 0.05); however, the activity of SMAP28 and SMAP29 was strongly associated (r = 0.933). The congeners also demonstrated antimicrobial activity against the bacteria tested (range of minimum inhibitory concnetrations 0.21-79 microg/ml). Overall, F. nucleatum was the most susceptible organism, while P. gingivalis was the least susceptible. Keratinocyte cytotoxicity was dependent on peptide length and dose. SMAP28 was the most cytotoxic, while SMAP14A was the least cytotoxic. CONCLUSION: The antimicrobial activities against oral microorganisms and the minimal toxicity seen in this study suggest that the congeners of SMAP29 may serve as an alternative to traditional antibiotics in the prevention and treatment of periodontal and other oral diseases.

The effects of danofloxacin and tilmicosin on neutrophil function and lung consolidation in beef heifer calves with induced Pasteurella (Mannheimia) haemolytica pneumonia.

Pneumonia caused by Pasteurella (Mannheimia) haemolytica was induced in weaned beef heifer calves, approximately 6 months of age. Calves were treated at 20 h after challenge with therapeutic doses of danofloxacin or tilmicosin. Peripheral blood neutrophils were collected at 3, 24 and 48 h after treatment. The ex vivo effects on neutrophil function, neutrophil apoptosis, and hematological parameters were examined, as was the effect on percentage lung consolidation. Neutrophil function assays included random migration under agarose, cytochrome C reduction, iodination, Staphylococcus aureus ingestion, chemotaxis, and antibody-dependent and antibody-independent cell-mediated cytotoxicity. Apoptosis was determined using a cell death detection kit. Killing was performed at 72 h after treatment. Statistical comparisons were made among the three groups of challenged-treated animals: saline, danofloxacin, and tilmicosin. Comparisons were also made between nonchallenged nontreated animals (NCH) and challenged saline-treated animals. There were no significant differences for any of the neutrophil function assays or neutrophil apoptosis among the challenged-treated groups. This suggests that danofloxacin and tilmicosin have no clinically significant effects on neutrophil function or apoptosis. There were also no significant differences in percentage lung consolidation among the challenged-treated groups. Significant differences were found between the NCH calves and the challenged saline-treated calves in several neutrophil assays, which were attributed to effects of P. haemolytica infection.

Defensin-induced adaptive immunity in mice and its potential in preventing periodontal disease.

The severity of periodontal disease is dependent on a combination of host, microbial agent and environmental factors. One strong correlate related to periodontal disease pathogenesis is the immune status of the host. Here we show that human neutrophil peptide (HNP) defensins or human beta-defensins (HBD), co-administered intranasally with the antigen ovalbumin (OVA), induce unique immune responses that if used with microbial antigens may have the potential to hinder the pathogenesis of periodontal disease. C57BL/6 mice were immunized intranasally with phosphate buffered saline (PBS) containing 1 micro g HNP-1, HNP-2, HBD1 or HBD2 with and without 50 microg OVA. At 21 days, isotypes and subclasses of OVA-specific antibodies were determined in saliva, serum, nasal wash, bronchoalveolar lavage fluid, and fecal extracts. OVA-stimulated splenic lymphoid cell cultures from immunized mice were assessed for interferon (IFN)-gamma, Interleukin (IL)-4 and IL-10. In comparison with mice immunized with only OVA, HNP-1 and HBD2 induced significantly higher (P < 0.05) OVA-specific serum IgG, lower, but not significant, serum IgM and significantly lower (P < 0.05) IFN-gamma. In contrast, HNP-2 induced low OVA-specific serum IgG and higher, but not significant, serum IgM. HBD1 induced significantly higher (P < 0.05) OVA-specific serum IgG, higher, but not significant, serum IgM, and significantly higher (P < 0.05) IL-10. The elevated serum IgG subclasses contained IgG1 and IgG2b.

A selectin inhibitor decreases neutrophil infiltration during acute Mannheimia haemolytica pneumonia.

The degree to which the selectin inhibitor TBC1269 reduces neutrophil infiltration in specific microscopic locations of the lung during acute pneumonia of neonates was determined. Neonatal calves were inoculated intrabronchially with Mannheimia (Pasteurella) haemolytica or saline, and lung tissue was collected at 2 and 6 hours postinoculation (PI). One 6-hour group inoculated with M. haemolytica received TBC1269 intravenously before and after inoculation with M. haemolytica. Infiltrates of neutrophils were significantly higher in the alveolar lumen and septae but lower in the bronchial lumen and epithelium at 6 hours PI than at 2 hours PI. Significantly fewer neutrophils (P < 0.05) were present in the alveolar lumen and septae, and the bronchiolar lumen and lamina propria in the lungs of TBC1269-treated calves compared with untreated calves at 6 hours PI. TBC1269 did not alter the infiltration into bronchi and blood vessels or the expression of the selectin-independent adhesion molecule, ICAM-1. This work suggests that during acute pneumonia of neonates 1) neutrophil infiltrates progressively increase in the alveolar lumens and septae but decrease in the bronchial lumen and epithelium with time, 2) TBC1269 reduces neutrophil infiltration into specific regions of alveoli and bronchioles rather than uniformly throughout the lung, and 3) selectin inhibition does not affect the location and intensity of ICAM-1 expression.

Cellular distribution of anionic antimicrobial peptide in normal lung and during acute pulmonary inflammation.

Anionic peptides (APs) are small antimicrobial peptides present in human and ovine lung. In this study APs were also detected in bovine lung, and production of APs in lungs with acute inflammation induced by various stimuli was determined. The distribution and intensity of APs were determined by immunohistochemistry in lungs of 1) neonatal calves (1-3 days of age) inoculated with Mannheimia (Pasteurella) haemolytica, a known inducer of the bovine beta-defensin lingual antimicrobial peptide (LAP) or pyrogen-free saline (PFS), and 2) growing calves (3 months of age) similarly inoculated with M. haemolytica, a lipopolysaccharide (LPS) from M. haemolytica, an LPS-associated protein from M. haemolytica, or PFS. APs were also detected by western blots with the same antibody in lungs of the calves above, as well as in calves inoculated with Pseudomonas aeruginosa, and an adult cow. Anionic peptide (AP) immunoreactivity was detected in bands (approximate weights) in the western blots of lung at 28-30 (strongest signal), 31, 45, and 52-60 kd regardless of inoculum. The adult cow lacked bands at 45 kd, but it had additional bands at 64 (inconsistently) and 35-38 kd. All these band sizes are consistent with those of the western blots of human and ovine lung. The cellular distribution of APs in lung of neonatal and growing cattle was similar to that in lung of human and sheep. In lungs with acute inflammation induced by live bacteria, LPS, or protein, AP distribution and intensity were similar to those in control (PFS-inoculated) lungs and slightly decreased in bronchioles. This work demonstrates that AP is present in lung of cattle and is thereby conserved among two ruminant species and man. Distribution and intensity of AP production are not enhanced by infection or acute inflammation and are decreased in bronchioles, which suggests that AP is not induced like beta-defensins such as LAP, but, instead, is produced constitutively.

Congeners of SMAP29 kill ovine pathogens and induce ultrastructural damage in bacterial cells.

SMAP29, an ovine cathelicidin, was systematically altered to create a family of 23 related peptides for MIC and minimum bactericidal concentration determinations. SMAP28, SMAP29, and a derivative of SMAP29 called ovispirin were all antimicrobial. However, many congeners of SMAP29 and ovispirin were not as active as the parent molecules. With immunoelectron microscopy, SMAP29 was seen on membranes and within the cytoplasm of Pseudomonas aeruginosa PAO1.

Mast cell density and substance P-like immunoreactivity during the initiation and progression of lung lesions in ovine Mannheimia (Pasteurella) haemolytica pneumonia.

To determine the density of mast cells (MCs) and the extent of substance P (SP) immunoreactivity during initiation and progression of pneumonic pasteurellosis (PP), 18 lambs were inoculated intrabronchially with Mannheimia (Pasteurella) haemolytica or saline, and lung tissue was collected at 1, 15 and 45 days post-inoculation (n=3, each group). Additionally, the left (non-inoculated) contralateral lungs in bacteria-inoculated animals were collected as controls. At 1 day after bacterial inoculation the lungs had typical M. haemolytica lesions. These pneumonic lesions had fewer numbers of MCs and reduced histamine content. Macrophages infiltrating some of the inflamed areas were strongly immunoreactive for SP. At 15 days, MCs remained scarce at sites where lung damage persisted, i.e. pyogranulomatous foci, but were increased in number in areas of interstitial damage. Pulmonary ganglion neurons were strongly immunoreactive for SP. By 45 days the fibrosing changes became more defined as pleural fibrosis, fibrosing alveolitis, alveolar epithelial hyperplasia and bronchiolitis obliterans. These lungs had increased numbers of MCs, but histamine content was not different from saline- and non-inoculated left lungs. Substance P immunoreactivity occurred only in nerves and was scarce and mild. This work demonstrates that MC density decreases initially with PP, but increases with progression of PP. SP fibres tend to be decreased during the initiation and at 45 days of PP, but other cells, such as macrophages and neuronal ganglion cells, produce substance P during progression of PP and thereby constitute an additional source of substance P.

Suppression of Mannheimia (Pasteurella) haemolytica serovar 1 infection in lambs by intrapulmonary administration of ovine antimicrobial anionic peptide.

In this study, the efficacy of ovine antimicrobial anionic peptide (AP) was assessed in a lamb model of acute pneumonia. A single intratracheal dose of the peptide, H-DDDDDDD-OH (0.5 mg) reduced pulmonary inflammation and the concentration of Mannheimia (Pasteurella) haemolytica in infected lung tissue. Administration of H-DDDDDDD-OH after infection was more effective in reducing the consolidation and lesion scores at the deposition site than its administration prior to infection. Hence, the in vivo effectiveness of AP suggests that it may have applications in the treatment of pulmonary infections. Further studies are needed to confirm these findings and also to determine the optimal doses and intervals of H-DDDDDDD-OH therapy.

Comparison of bronchoalveolar lavage fluid obtained from Mannheimia haemolytica-inoculated calves with and without prior treatment with the selectin inhibitor TBC1269.

OBJECTIVES: To determine effects of selectin inhibitor TBC1269 on neutrophil infiltration, and neutrophil-associated injury during pneumonia induced by Mannheimia haemolytica and concentration of antimicrobial anionic peptide (AAP) in bronchoalveolar lavage fluid (BALF) as well as antimicrobial activity of BALF from healthy (control) neonatal calves, neonatal calves with M haemolytica-induced pneumonia, neonatal calves with prior treatment with TBC1269, and adult cattle. ANIMALS: Eighteen 1- to 3-day-old calves and 9 adult cattle. PROCEDURE: Calves were inoculated with M haemolytica or pyrogen-free saline (0.14M NaCl) solution into the right cranial lung lobe, and BALF was collected 2 or 6 hours after inoculation. Thirty minutes before and 2 hours after inoculation, 4 calves received TBC1269. The BALF collected from 9 adult cattle was used for comparison of BALF AAP concentration and antimicrobial activity. Protein concentration and neutrophil differential percentage and degeneration in BALF were determined. An ELISA and killing assay were used to determine BALF AAP concentration and antimicrobial activity, respectively. RESULTS: Total protein concentration was significantly decreased in BALF from calves receiving TBC1269. Similar concentrations of AAP were detected in BALF from all calves, which were 3-fold higher than those in BALF from adult cattle. However, BALF from neonates had little or no anti-M haemolytica activity. CONCLUSIONS AND CLINICAL RELEVANCE: These results suggest that TBC1269 decreases pulmonary tissue injury in neonatal calves infected with M haemolytica. Although AAP is detectable in neonatal BALF at 3 times the concentration detected in adult BALF, neonatal BALF lacks antimicrobial activity for M haemolytica.

Gallinacin-3, an inducible epithelial beta-defensin in the chicken.

Gallinacin-3 and gallopavin-1 (GPV-1) are newly characterized, epithelial beta-defensins of the chicken (Gallus gallus) and turkey (Meleagris gallopavo), respectively. In normal chickens, the expression of gallinacin-3 was especially prominent in the tongue, bursa of Fabricius, and trachea. It also occurred in other organs, including the skin, esophagus, air sacs, large intestine, and kidney. Tracheal expression of gallinacin-3 increased significantly after experimental infection of chickens with Haemophilus paragallinarum, whereas its expression in the tongue, esophagus, and bursa of Fabricius was unaffected. The precursor of gallinacin-3 contained a long C-terminal extension not present in the prepropeptide. By comparing the cDNA sequences of gallinacin-3 and GPV-1, we concluded that a 2-nucleotide insertion into the gallinacin-3 gene had induced a frameshift that read through the original stop codon and allowed the chicken propeptide to lengthen. The striking structural resemblance of the precursors of beta-defensins to those of crotamines (highly toxic peptides found in rattlesnake venom) supports their homology, even though defensins are specialized to kill microorganisms and crotamines are specialized to kill much larger prey.

Effects of the synthetic selectin inhibitor TBC1269 on tissue damage during acute Mannheimia haemolytica-induced pneumonia in neonatal calves.

OBJECTIVE: To determine effects of the selectin inhibitor TBC1269 on neutrophil-mediated pulmonary damage during acute Mannheimia haemolytica-induced pneumonia in newborn calves. ANIMALS: Eighteen 1- to 3-day-old colostrum-deprived calves. PROCEDURE: Mannheimia haemolytica or saline (0.9% NaCl) solution was inoculated in both cranial lung lobes of 12 and 6 calves, respectively. Calves were euthanatized 2 (saline, n = 3; M haemolytica, n = 4) or 6 hours (saline, n = 3; M haemolytica, n = 8) after inoculation. Four M haemolytica-inoculated calves euthanatized at 6 hours also received TBC1269 (25 mg/kg, IV) 30 minutes before and 2 hours after inoculation. Conjugated diene (CD) concentrations, inducible nitric oxide synthase (iNOS) expression, and apoptotic cell counts were determined in lung specimens collected during necropsy. RESULTS: Conjugated diene concentrations were significantly increased in all M haemolytica-inoculated groups, compared with saline-inoculated groups. Calves treated with TBC1269 had decreased concentrations of CD, compared with untreated calves, although the difference was not significant. Number of apoptotic neutrophils and macrophages increased significantly inTBC1269-treated calves, compared with untreated calves. Inducible nitric oxide synthase was expressed by epithelial cells and leukocytes. However, iNOS was less abundant in airway epithelial cells associated with inflammatory exudates. Degree of iNOS expression was similar between TBC1269-treated and untreated calves. CONCLUSIONS: Mannheimia haemolytica infection in neonatal calves resulted in pulmonary tissue damage and decreased epithelial cell iNOS expression. The selectin inhibitor TCB1269 altered, but did not completely inhibit, neutrophil-mediated pulmonary damage.

The ovine cathelicidin SMAP29 kills ovine respiratory pathogens in vitro and in an ovine model of pulmonary infection.

Cathelicidins are antimicrobial peptides from sheep (SMAP29 and SMAP34), rabbits (CAP11 and CAP18), rodents (CRAMP), and humans (FALL39, LL37, and h/CAP18). In a broth microdilution assay against nine ovine pathogens, SMAP29, SMAP34, mouse CRAMP, CAP18, CAP18(31), CAP18(28), CAP18(22), and CAP18(21a) were the most active, with MICs as low as 0.6 microg/ml. Other cathelicidins were less active. In lambs with pneumonia, 0.5 mg of SMAP29 reduced the concentration of bacteria in both bronchoalveolar lavage fluid and consolidated pulmonary tissues. Hence, the antimicrobial activity of SMAP29 suggests that it has applications in the treatment of respiratory tract infections.

Ultrastructural location of the cross-protection factor on in vivo and in vitro grown Pasteurella multocida.

Pasteurella multocida from infected turkey tissues expresses a unique immunogen called cross-protection factor (CPF) that induces immunity to challenge by both homologous and heterologous serotypes. In this study, we used a monoclonal antibody (AMP MAb) to CPF and protein A-colloidal gold (PACG) to locate CPF on P. multocida. After incubation with AMP MAb and PACG, CPF was detected at the bacterial surface and cell periphery of P. multocida in infected turkey liver and P. multocida isolated from infected turkey blood. CPF was not detected on P. multocida incubated with control monoclonal antibody. Pasteurella multocida isolated from infected turkey blood and cultivated in the peptone-based medium did not express CPF consistently, and some cells contained more CPF than others. The location of CPF also varied, and CPF was detected both intracellular and extracellular on the cell surface. In the latter cells, CPF was heavily concentrated to a specific lateral site or detected sloughing from the cell surface. These results correlate with laboratory observations that CPF detected on P. multocida from infected turkey tissues, P. multocida isolated from infected turkey blood, and P. multocida cultivated in peptone-based medium is associated with outer membrane fractions.

Response of the ruminant respiratory tract to Mannheimia (Pasteurella) haemolytica.

Pneumonia is a leading cause of loss to the sheep and cattle industry throughout the world. Mannheimia (Pasteurella) haemolytica is one of the most important respiratory pathogens of domestic ruminants and causes serious outbreaks of acute pneumonia in neonatal, weaned and growing lambs, calves, and goats. M. haemolytica is also an important cause of pneumonia in adult animals. Transportation, viral infections with agents such as infectious bovine rhinotracheitis virus, parainfluenza-3 virus or bovine respiratory syncytial virus, overcrowding, housing of neonates and weaned animals together and other stressful conditions predispose animals to M. haemolytica infection [1, 2]. This review assimilates some of the findings key to cellular and molecular responses of the lung from a pathologist's perspective. It includes some of what is known and underscores areas that are not fully understood.