Search

The origin and behaviour of keratinizing stratified squamous epithelium, an essential component of cholesteatoma occurring in the middle-ear cavity, has puzzled otologists for decades. In this experimental study in 16 cats, central (n = 23) and peripheral (n = 9) tympanic membrane perforations were observed for up to 63 days before sacrifice. The tympanic membranes with bony rim were excised, decalcified and embedded in Epon 812. Sections were stained with toluidine blue and examined using a light microscope. The perforation had been sealed by meatal epithelium exhibiting pronounced hyperplasia and keratin formation, lying on a bed of granulation tissue. Subtotal central perforations healed within 14 days, forming a bowl-shaped tympanic membrane and leaving parts of the handle of the malleus (with meatal epithelium) protruding freely into the middle-ear cavity. Stratified squamous epithelium, morphologically identical with that of external ear canal epidermis, could be observed on the malleus even 63 days after operation. This meatal epithelium was non-keratinizing, non-invasive, and showed no destructive properties typical of acquired cholesteatoma. During certain circumstances, the cell cycle of hyperplastic epidermal epithelium within the middle-ear cavity can evidently be arrested and inactivated by a local defence mechanism.

The function of Shrapnell's membrane (pars flaccida; PF) in middle-ear mechanics is still an enigma, though numerous proposals have been put forward, e.g. protection of pars tensa, equalizing of middle-ear pressure, sound transmission, and the site of origin of otitis media. In this study the PF was studied in a mammal (the hooded seal) which exposes itself to extreme pressure differences (from 1 to 100 atmospheres) when diving. Formaldehyde-fixed temporal bones obtained from newborn, one-year-old, and adult seals (three of each) were cleansed and decalcified in 10 per cent EDTA. The lateral wall of the middle-ear cavity, including the whole tympanic membrane with its bony surroundings, was then excised and photodocumented. Thin sections were cut parallel with, and perpendicular to, the handle of the malleus, stained with haematoxylin-eosin, toluidine blue or Giemsa stain and examined under a light microscope. One seal head was subjected to high resolution computerized tomography (HRCT) before sectioning. The PF was observed to be a narrow fissure measuring a maximum of 0.8 mm between processus brevis of the malleus and the notch of Rivinus in pars squamosa (pars tensa diameter 10-12 mm). It seems unlikely that the PF of the hooded seal participates in pressure equalization in the middle ear. The main function of the lateral wall of the attic, including the minimal PF, appears to be to protect the middle-ear ossicles and allow movement of the malleus.

Objective: Oral and pharyngeal cavities harbor a commensal bacterial flora which is kept in check by several innate and acquired agents. In this study, we focused on the proportions in which some antibacterial moderators (lysozyme, lactoferrin, IgG and S-IgA) coat the tonsillar surface bacteria in healthy individuals, in patients with acute tonsillitis (AT) culture-positive for Streptococcus pyogenes, and in patients with infectious mononucleosis (IM) caused by Epstein-Barr virus (EBV)., Methods: Bacterial samples were collected for aerobic culturing and immunocytochemical evaluation from the tonsillar surfaces of eight healthy individuals (four males, four females; age range 16-22 years), eight patients with current AT (two males, six females; age range 16-29 years) and seven patients with IM (four males, three females; age range 15-21 years). The immunocytochemical assay was based on gold-labeled antiserum to human lysozyme, lactoferrin, IgG and S-IgA followed by gold particle tracing in the transmission electron microscope., Results: During AT, a significant increase in lysozyme coating (P<0.05) and lactoferrin coating (P<0.0005) of the bacteria was noted, whereas the S-IgA coating was significantly reduced (P<0.0005). During IM infection, a significant increase in lactoferrin coating was noted (P<0.0005) whereas immunoglobulin coating was significantly reduced (IgG P<0.025; S-IgA P<0.0005) compared with healthy controls. During IM, all antibacterial moderators evaluated were significantly reduced compared with the situation during AT., Conclusions: Noticeable changes in the local innate and acquired bacterial defence system were observed during tonsillar infections, particularly during IM.

Objective: the human external auditory canal (EAC) hosts a commensal bacterial flora of mainly non-pathogens, but bacterial pathogens may also be present. The latter are important in the aetiopathogenesis of external otitis and middle ear cholesteatoma with discharge. The purpose of this study was to quantify the bacteria normally harboured in the healthy EAC and on a well-defined region of the cavum conchae (CC) of children., Methods: bacterial samples were collected from the CC and bony portion of the EAC of 32 children (18 boys, 14 girls; median age 5 years). Prior to sampling, the EACs were either left untreated, anaesthetized with Bonain's solution, or washed with 70% alcoholic solution. The samples were incubated at 37 degrees C and evaluated regarding bacterial species and number., Results: the predominant bacterial non-pathogens were coagulase-negative Staphylococci and coryneforms (diphtheroids) and pathogens Staphylococcus aureus and Pseudomonas aeruginosa. Total bacterial counts of the CC ranged between 2 x 10(3) and 4.6 x 10(4) CFU/cm(2) (median value 7 x 10(3); n = 14). Total bacterial counts from the bony portion of the EAC ranged between 0 and 5.7 x 10(7) CFU/EAC (median value 8 x 10(3); n = 32). Pre-treatment of the EACs with Bonain's solution containing the highly bactericidal substance phenol or with 70% alcoholic solution did not sterilize the EAC., Conclusions: small numbers of bacterial non-pathogens (and sometimes pathogens) are found in the EAC of children. Neither phenol nor 70% alcoholic solution can inhibit or eradicate all these microorganisms., (Copyright 2002 Elsevier Science Ireland Ltd.)

Objective: A common complication of virus-induced upper respiratory tract infections is acute otitis media caused by bacterial pathogens. Simultaneously, increased bacterial colonization in the nasopharynx occurs. Our intention in this study was to identify the causes of this increased colonization of bacteria by evaluating their coating with the antibacterial substances lysozyme, lactoferrin and immunoglobulins IgG, S-IgA and IgM and their ability to penetrate epithelial cells during infectious mononucleosis (IM) caused by Epstein-Barr virus., Methods: Cellular samples were collected from the oropharynx of 21 patients (16 males, five females; age range 10-21 years) with current IM. An immunocytochemical assay using gold-labelled antiserum to human lysozyme, lactoferrin, IgG, S-IgA and IgM followed by gold particle and epithelial cell tracing in the transmission electron microscope., Results: A significant reduction in bacterial coating with IgG (P<0.05) and S-IgA (P<0.01) was noted, whereas there was a significant increase in coating with lactoferrin (P<0.01) and IgM (P<0.01). No significant change in lysozyme coating of the bacteria was noted, compared with healthy controls. Bacterial penetration into epithelial cells was seen particularly in patients culture-positive for beta-haemolytic streptococci., Conclusions: Reduced bacterial coating with IgG and S-IgA immunoglobulins, combined with bacterial penetration into epithelial cells, may exacerbate the bacterial colonization on oropharyngeal mucosal membranes observed during IM.

Bacterial samples were obtained from the tonsillar surfaces of seven patients (four males, three females; median age 18 years, range 15 to 21 years) suffering from acute infectious mononucleosis with concomitant pharyngotonsillitis, and from five healthy controls. By using gold-labelled antiserum to human lysozyme and lactoferrin, micro-organisms on the tonsillar surfaces coated with these antibacterial substances could be identified by tracing the gold particles in the transmission electron microscope. In healthy individuals, most of the bacteria were coated with lysozyme and significantly more bacteria were coated with lysozyme than with lactoferrin (p < 0.01). In patients there was a non-significant reduction in lysozyme-coating of the bacteria, whereas lactoferrin-coating was significantly increased (p < 0.01). Changes in the lysozyme and/or lactoferrin coating of the tonsillar surface bacteria on the palatine tonsils during infectious mononucleosis cannot explain the tendency to immense local bacterial colonization with commensals and proneness to bacterial penetration into the epithelial cells.

Recent studies using a satellite-linked dive recorder have shown that the hooded seal (Cystophora cristata), a common Arctic pinniped, can dive to a depth of > 1000 m and stay submerged for close to 1 h. At these depths the water pressure reaches 100 atm, entailing obvious risk of serious damage to the hearing apparatus, mainly the tympanic membrane (TM) and middle ear (ME). We dissected and photodocumented the temporal bones of five newborn and three adult hooded seals in order to study the temporal bone structure and reveal its protective mechanisms for extreme pressure changes. Specimens were sectioned and stained for light microscopy. The thicknesses of the pars tensa and pars flaccida were found to average 60 and 180 microm, respectively. The ME cavity hosts a cavernous tissue of thin-walled vessels beneath the modified respiratory epithelium. The ME and external ear canal (EAC) volumes can be altered appreciably by filling/emptying the cavernous tissue with blood. The ossicles were fixed by contracting the tensor tympani and stapedius muscles simultaneously with complete occlusion of the EAC. According to Boyle's law, the volume of the gas-filled ME cavity at a depth of 1000 m is only 1% of its volume at the surface of the sea. Ascent from such a depth allows the gas in the ME cavity to expand, causing the TM to bulge laterally. This movement is counteracted by a reduction in the blood volume inside the cavernous sinuses, action in the tensor tympani and stapedius muscles and discharge of gas through the Eustachian tube. The presence of a firm, broad-based exostosis in the floor of the EAC lateral to the TM helps to obstruct the EAC.

The purpose of this study was to establish whether lactoferrin (hLf) attached to Streptococcus pyogenes, one causative agent of acute pharyngotonsillitis (AT), during the course of the disease. Bacterial samples were obtained from the tonsillar surfaces of 7 patients (6 females, 1 male; median age 26 years; range 16-50 years) suffering from AT who were culture-positive for S. pyogenes and from 5 healthy adult controls who were culture-negative for this pathogen. Using gold-labelled antiserum against S. pyogenes and hLf, this pathogen and other bacteria on the tonsillar surfaces coated with hLf could be identified by tracing the gold particles in a transmission electron microscope. In healthy adults, 8% (median value; range 6-12%) of the surface tonsillar bacteria were coated with hLf. In AT patients, 59% (median value; range 42 67%) of S. pyogenes were coated with hLf, in contrast to 9% (median value; range 0-26%) of all other bacteria (p < 0.01). This study hints that hLf might participate in recovery from AT in several ways, e.g. by binding to the S. pyogenes pathogens, in addition to its well-known virtue of iron-binding capacity.

Epithelial cells and bacteria were sampled from the tonsillar surfaces of seven patients (six males, one female; median age 16 years, range 10 to 21 years) suffering from acute infectious mononucleosis with concomitant pharyngotonsillitis. By using gold-labelled antiserum to human IgG and secretory IgA (sIgA), micro-organisms on the tonsillar surfaces coated with these immunoglobulins could be identified by tracing the gold particles in the transmission electron microscope. The patients harboured significantly fewer bacteria coated with immunoglobulins than did healthy controls. More bacteria were coated with IgG immunoglobulins than with sIgA. Reduced immunoglobulin-coating of the bacteria on the tonsillar surfaces during infectious mononucleosis can explain their tendency to immense local colonization and proneness to penetrate into the epithelial cells.

Bacterial penetration into epithelial cells, scraped from the palatine tonsils of 14 patients (10 males, four females; median age 16 years) with current infectious mononucleosis and concomitant membranous tonsillitis, was studied using the transmission electron microscopic (TEM) technique. Bacteria were seen to adhere to and penetrate the epithelial cells, some of which were completely filled with bacteria. This finding suggests intracellular proliferation of bacteria. Epstein-Barr virus, the causative agent of infectious mononucleosis, especially when associated with growth of beta-haemolytic streptococci on the palatine tonsils, induces bacterial penetration into tonsillar tissue, that in turn might be a causative mechanism in the development of peritonsillar abscess.

Exostoses of the external auditory canal (EAC) develop after protracted mechanical, chemical or thermal irritation in particular. This is a common disorder among aquatic sportsmen and has been considered unique to Man. We dissected and photodocumented the EACs of 5 newborn and 3 adult Hooded Seals (Cystophora cristata). Serial sections of the EACs were prepared for light microscopic evaluation after staining with haematoxylin-eosin or toluidine blue. All EACs exhibited a firm, broad-based. mountain peak-shaped exostosis on the floor of the meatus, lateral to the eardrum. In addition, the meatal skin of the bony EAC harboured large venous sinuses. The exostosis and venous sinuses of the seal EAC participate in the protection of the sensitive hearing apparatus, particularly the pars tensa portion of the drum, during divine.

Streptococcus pyogenes is the most frequent causative agent of acute pharyngotonsillitis (AT). The first events in the etiopathogenesis of an AT infection caused by these bacterial pathogens are their penetration through the mucus film covering the oropharyngeal mucosa, and their attachment to the surface epithelium. Adherence of S. pyogenes to tonsillar epithelial cells is a precondition for bacterial colonisation, for triggering off cell activation, internalising of bacteria into the epithelial cells and cytokine release from the epithelial cells with subsequent induction of an inflammatory reaction in underlying tissues. Scanning and transmission electron microscopic studies revealed that the surface epithelium of the human palatine tonsils consisted of a weakly keratinized, stratified squamous epithelium built up of pentangular cells where the apical cell surface formed an irregular pattern of microridges. The distance between two adjacent microridges was roughly one-third of the diameter of a S. pyogenes bacterium. By using gold-labelled antiserum to S. pyogenes, we showed that the target region for these pathogens on the epithelial cells during an on-going AT infection was located on the crests of the microridges where bacterial pili made adhesin-receptor contact with the tonsillar surface epithelium.

The role of the non-specific (mucociliary clearance, mucus, lactoferrin, lysozyme, fibronectin, lactoperoxidase, complement, phagocytosis) and specific (immunoglobulins) immune systems in combating bacterial pathogens invading the middle ear cavity is discussed. The non-specific system is present from birth, acts promptly against a broad spectrum of microorganisms, has no memory, and is present for life. In contrast, immunoglobulins act specifically. Secretory IgA antibody prevents bacteria from attaching to the epithelium, has no pro-inflammatory effect and does not activate complement. IgG antibodies can opsonize the bacteria for phagocytosis and eventually prevent the bacteria from penetrating the epithelium. IgG is pro-inflammatory and can activate a complement cascade.

The external auditory canal is less susceptible to infections than the sensitive middle-ear cavity. Since recent research has provided insight to the production of potent antimicrobial peptides from various surface epithelia, we wanted to investigate whether protection of the external auditory canal in part could be explained by the production of human beta-defensin-1 (HBD-1). This particular peptide is known to be constitutively expressed in various surface epithelia, such as airway, skin, and urogenital tissues. By reverse transcriptase PCR we demonstrate HBD-1 mRNA in the pars tensa and pars flaccida of the tympanic membrane and in the meatal skin. In situ hybridization studies localized the HBD-1 mRNA to the epidermal layer of these tissues. The HBD-1 transcripts were also evident in the sebaceous glands and in hair follicles of the meatal skin. In contrast, HBD-1 mRNA was not detected in the tympanal epithelium of the eardrum. The widespread presence of mRNA encoding for this broad-spectrum antimicrobial peptide in the meatal skin and tympanic membrane suggests that HBD-1 participates in the innate antimicrobial defense of the external auditory canal and middle-ear cavity.

Bacterial swabs were collected from the tonsillar surfaces of eight patients with current acute tonsillitis, culture-positive for Streptococcus pyogenes. Using gold-labelled antiserum to S. pyogenes, these micro-organisms could be localized in the samples by transmission electron microscopy. S. pyogenes pathogens were further characterized with gold-labelled antiserum to human IgG and SIgA antibodies. Roughly 90% of the pathogens were found coated with IgG antibodies, irrespective of the duration of the disease, whereas the proportion of SIgA-coated pathogens increased with disease duration. Insufficient IgA coating of pathogens might well be a contributory cause of the induction of tonsillar infection, probably due to inadequate prevention of the attachment of the S. pyogenes bacteria to the tonsillar surface epithelium.

Bacterial and epithelial cell samples were obtained, within 24 h of onset of pharyngeal symptoms, from the palatine tonsils of nine patients (four female and five male; age range 10-40 years, median age 23) with acute tonsillitis, culture-positive for Streptococcus pyogenes. The specimens were examined using fluorescein isothiocyanate- (FITC) and gold-labelled antiserum to S. pyogenes and fluorescence, scanning electron and transmission electron microscopy. S. pyogenes could be identified both in the mucous layer covering the tonsils and attached to the surface epithelial cells. Long chains of coccus-shaped bacteria could be seen encroaching on the epithelial cell borders. S. pyogenes can apparently penetrate the mucous barrier, attach to the epithelial cells, spread from cell to cell and possibly penetrate into the outermost layer of the epithelial cells. These events in turn provoke cytokine production and/or complement activation, which induce inflammatory reaction in the tonsillar tissue.

Fifty-five samples of pus were collected from 51 acute, non-perforated, two spontaneously ruptured and two recurrent peritonsillar abscesses (35 males and 18 females; median age 18 years) and analysed regarding (i) aerobic and anaerobic bacteria (standard culturing), (ii) morphology of bacteria and inflammatory cells (direct microscopy of acridine orange-stained material), and (iii) the percentage of bacteria coated with immunoglobulins IgG, secretory IgA (SIgA) and IgM and complement cleavage product C3b (immunofluorescence assay). Seventy-one per cent of the abscesses harboured a mixed bacterial flora of various aerobes and anaerobes. In none of the cases with a single bacterial species (27 per cent) could immunoglobulin- or complement-coated bacteria be found. In abscesses with a mixed flora, 18 per cent harboured IgG-coated, 15 per cent SIgA-coated, five per cent IgM-coated and five per cent C3b-coated bacteria, respectively. All pus samples contained inflammatory cells in abundance but they were mostly deformed and only occasionally could intracellular bacteria be recognized. Insufficient immunoglobulin-coating of bacteria might be an important aetiopathogenic factor in the development of a peritonsillar abscess. Bactericide in the abscesses is accomplished chiefly by protective mechanisms not dependent on antigen recognition by antibodies.

Sixty consecutive patients (48 males, 12 females; age range 16 to 57, mean age 35 years) underwent nasal septal surgery at our out-patient department for symptoms of nasal obstruction. The operations were performed under local anaesthesia supplemented with 2.5-5 mg intravenous midazolam. Only one surgeon and one nurse were needed for the procedure. The total time spent at the hospital averaged three hours. The preliminary short-term results of surgery are comparable to those at our day-care unit. The complication rate was negligible. Ninety-five per cent of these patients regarded the arrangements for their operation as safe. Our results show that nasal septal surgery can be performed with good cost-effectiveness, satisfactory quality and with good safety and patient satisfaction as an out-patient procedure.

Localization and semiquantification of beta-haemolytic streptococci, Group A (GABHS), GABHS attachment and general bacterial attachment to epithelial cells (bacterial number and morphology) were studied during GABHS-positive acute tonsillitis and pharyngitis infections and among healthy GABHS carriers. Samples were collected from various areas of the oropharynx (palatine tonsils, posterior oropharyngeal wall, palatoglossal arch and buccal mucosa). During acute tonsillitis and pharyngitis, GABHS grew in samples obtained from the palatine tonsils and posterior oropharyngeal wall. The ratio of GABHS colonies to other aerobic colonies increased, and GABHS became attached to epithelial cells of both palatine tonsils and posterior oropharyngeal wall. In GABHS carriers, GABHS were found mainly on the palatine tonsils, but these microorganisms were not attached to the epithelium. Overall bacterial attachment to tonsillar and oropharyngeal epithelial cells increased during active tonsillitis and pharyngitis.

Effusion material was aspirated from 51 consecutive peritonsillar abscesses (34 male, 17 female; age range eight to 46 years) and subjected to direct microscopy after staining with acridine orange. Bacteria were counted per ml effusion material and their morphology was analyzed. In addition, aerobic and anaerobic bacterial culturing was performed. Effusions containing beta-haemolytic streptococci Group A, which appeared as a single species contained fewer bacteria (8 x 10(6) per ml, median value) than effusions harbouring a mixed flora (7 x 10(8) bacteria per ml, median value). Direct microscopy of effusions obtained from peritonsillar abscesses makes possible rapid identification of a single or mixed flora, which is of importance for the antibiotic treatment of the disease.

Local protection of the maxillary sinuses against bacterial invasion takes both specific and non-specific forms. The present study is intended to evaluate the participation of the specific protective factors, immunoglobulins IgG, secretory IgA, IgM and complement, in protecting the maxillary sinuses during chronic maxillary sinusitis (CMS). We collected 47 sinus effusion samples from 37 patients (17 male, 20 female) with current CMS of at least 3 months' duration. Patients' ages ranged from 3 to 80 years. The effusion material was subjected to qualitative and quantitative bacteriological analyses, while bacterial coating with IgG, SIgA, IgM and C3b was determined using an immunofluorescence technique. Detectable bacteria were harboured by 55% of the samples, the most common species being Moraxella catarrhalis, Streptococcus pneumoniae and Staphylococcus aureus. The bacterial counts ranged from 0 to 10(9) per ml effusion. Half of the samples hosting detectable bacteria showed microorganisms coated with protective immunoglobulins. Antibacterial factors had completely eradicated the microorganisms in 45% of the CMS cases and coated the organisms with specific immunoglobulins in a further 28%.

The occurrence of otitis media (OM) was examined retrospectively in files at Longyearbyen Hospital, Svalbard, Norway during 1991-1994. Acute OM (AOM) with and without spontaneous drum perforation, secretory OM (SOM) and chronic OM (COM) with and without suppuration were registered, as were patient's sex, age, and climatic conditions. These findings were compared with weather observations obtained from the Norwegian Meteorological Institute, Tromsø in the same period. Sixty-four percent of the OM patients registered were classified as AOM, 17% having a spontaneous drum perforation. Thirty percent suffered from SOM, whereas 6% had COM. Forty percent of the patients were younger than 6 years. The distribution of OM cases during the year showed a peak during the spring, especially in May. Another minor peak was noted in September. Average daily temperature in the period was -5.6 degrees C, range -14 degrees C to +6 degrees C. Humidity and precipitation were fairly stable throughout the observation period 74% and 240 mm/year, respectively. OM is a common disease at Svalbard and is apparently related to seasonal shifts in temperature.

Epithelial cells were swabbed from the tonsillar surfaces of 5 patients with acute tonsillitis culture-positive for Streptococcus pyogenes. By using 10 nm gold particles conjugated to antiserum to S. pyogenes it was possible to trace the actual microorganisms when examined in a transmission electron microscope. The S. pyogenes bacteria, usually in pairs, were attached to the epithelial surface by their pili. The bacteria often formed a hollow in the epithelial cell surface. Coccus-shaped bacteria expressing positive affinity to immunogold-labelled antiserum were intermingled with bacteria, often rods, having no affinity whatsoever to the antiserum. With the immunocytological technique outlined in this study it is possible to study more closely cellular/bacterial adhesion mechanisms.

Epithelial cells were harvested from the surface of the palatine tonsils of seven patients with current acute tonsillitis, proven culture-positive for Streptococcus pyogenes. The epithelial cells harboured attached bacteria, which expressed positive affinity to gold-labelled antiserum to S. pyogenes. The gold particles adhered selectively to the bacterial capsules. The microorganisms were held in place by projections protruding from the epithelial cells, which were in close contact with the pili of the bacteria. In some areas, positive immunogold-labelled bacteria intermingled with bacteria lacking such labelling. None of the culture-negative controls harboured epithelial cells with positive immunogold-labelled bacteria. Orally administered phenoxymethylpenicillin caused a significant reduction in both culture-positive S. pyogenes and bacteria displaying positive coating with specific gold-labelled antiserum to S. pyogenes.

On day 2 post partum, swab samples were obtained from the oropharyngx of 20 healthy, breast-fed babies. The samples were examined for aerobes (culture on agar plates), for bacterial coating with the immunoglobulins SIgA, IgG and IgM (immunofluorescence assay), for bacterial attachment to epithelial cells (fluorescence microscopy of acridine orange stained material, and scanning electron microscopy). alpha-Hemolytic streptococci grew almost exclusively in the oropharynx, while 32% (median value) of the epithelial cells showed bacterial attachment in abundance, viz. > 50 attached bacteria per cell, 66% (median value) of the bacteria showed positive reactivity when treated with anti-human SIgA serum. 72% (median value) of the bacteria were coated with IgG, but no IgM coating of the bacteria could be detected. Newborns do not possess IgA antibodies, as only IgG is able to pass the placental barrier. The SIgA-rich colostrum is capable of coating the oropharyngeal bacteria of breast-fed babies during their first days after birth. However, despite abundant SIgA-coating, bacteria still succeed in attaching to the epithelial cells of the oropharynx. This finding hints that factors other than SIgA impede the bacterial/cellular adhesion mechanism.

Bacterial adherence to the oropharyngeal epithelium is a significant factor in normal microecology, etiopathogenesis of diseases (tonsillitis, gingivitis) and possibly also induction of immune response. Bacterial adhesion to human tonsillar epithelium of whole tonsils and swabs was studied by fluorescence, scanning, and transmission electron microscopy. The epithelial cell borders were well demarcated. On the apical surface of the cells there were irregular microridges. All forms of microscopy visualized epithelial cells with attached bacteria, often forming microcolonies on the free surface. Some bacteria formed excavations on the cell surface. Most attached bacteria were coccoid, but variously sized rods were also visible. In transmission electron microscopy, epithelial cells with intracellular bacteria were regularly observed.

Fifty samples of maxillary sinus effusion from 40 patients (median age 29 years; 26 females and 14 males) with acute maxillary sinusitis (AMS) were subjected to quantitative and qualitative bacteriological analysis. Furthermore, bacteria coated with immunoglobulin (IgG, secretory IgA, IgM) or complement (C3b) were evaluated using an immunofluorescence assay. 72% of the samples harboured detectable bacteria, the most common pathogens being Streptococcus pneumoniae and non-typable Haemophilus influenzae. Of the bacteria-positive samples, 28% harboured immunoglobulin-coated bacteria. During the first weeks of an AMS infection, the present immunoglobulins are insufficient to protect the maxillary sinuses.

Sequential bacterial samples were obtained from the tonsillar surface of 19 consecutive patients (12 females, seven males; mean age 16.1 years, range four to 24 years) suffering from infectious mononucleosis with membranous tonsillitis. The specimens were examined with respect to aerobes (culture on blood and chocolate agar plates) and proportions of bacteria coated with immunoglobulins (secretory IgA, IgG, IgM) by using an immunofluorescence assay. In the early stage of the membranous tonsillitis phase, attachment of secretory IgA (SIgA) and IgG to the bacteria was greatly suppressed, as compared with healthy controls. Coating with IgM was evident only late in the membranous tonsillitis phase but was contracted and still evident even after the clinical throat symptoms had abated. The findings suggest that the B-lymphotropic Epstein-Barr virus, causative agent of infectious mononucleosis, exerts a transient suppression of immunoglobulin-coating of bacteria harboured on the tonsillar surfaces, with consequent abundant bacterial attachment to the epithelial cells and massive bacterial colonization on the palatine tonsils.

Known numbers (CFU/ml) of middle ear pathogens (S. pneumoniae, H. influenzae, M. catarrhalis, S. aureus and beta-haemolytic streptococci group A) and non-pathogens (alpha-haemolytic S. mitis) were inoculated into one end of a micropipet containing a 35 mm long pillar of culture-negative middle ear effusion. The micropipets with the effusion/bacterial suspension were incubated at 37 degrees C and kept at various inclinations to the horizontal plane (15 degrees, 30 degrees and -75 degrees). All S. aureus, M. catarrhalis, alpha- and beta-haemolytic streptococci isolates examined survived in this medium for 18 h. The majority of alpha- and beta-haemolytic streptococci isolates penetrated the MEE, irrespective of the inclination of the micropipet, whereas the number of S. pneumoniae (p<0.01) and H. influenzae (p = NS) isolates penetrating the substrate increased when the micropipets were inclined at -75 degrees to the horizontal. None of the S. aureus and M. catarrhalis isolates penetrated the MEE pillar during the incubation. The present in vitro study demonstrated that MEE possesses an antibacterial property and is able to selectively hinder bacterial penetration.

Swabs of buccal and gingival epithelial cells from healthy young adult donors were washed in physiological saline solution, smeared on glass slides and stained with acridine orange. The presence of bacteria attached onto epithelial cells was examined under a fluorescence microscope. Four hours after a chlorhexidine rinse, the number of cells with > 50 attached bacteria had almost completely vanished. The degree of bacterial colonization seemed to re-establish at a level exceeding the baseline. One week after chlorhexidine treatment the degree of colonization was still over the control level. Toothbrushing with a conventional toothpaste reduced immediately the number of cells with > 50 bacteria. The colonization had re-established to the pre-washing levels at the buccal sites at 1 h and at the gingival sites at 4 h after toothbrushing. A method for the evaluating of the antimicrobial power of oral hygiene products is presented.

The effects of keeping rat tympanic membranes with an artificially made pars tensa perforation in tissue culture were observed under a scanning electron microscope. After one day and onwards, spreading and thickening of the keratinizing, outer squamous epithelium (OE) was noted. In addition, ballooning of the innermost cells of the outer epithelium apposing the inner tympanic epithelium (IE) was seen. No appreciable reaction was noted in the connective tissue layer of the drum. The inner tympanic epithelium appeared to be swollen, containing spherical structures in the cytoplasm, especially close to the area of contact with the outer meatal epithelium. No complete cover of the drum defect was seen after 14 days of tissue culture. Hyperplasia and spreading of the keratinizing, outer squamous epithelium of the drum is not sufficient to achieve covering of a drum perforation and complete healing cannot take place unless supported by granulation tissue formation.

Samples of middle ear effusions (MEE) obtained from 50 children suffering from acute purulent otitis media (AOM; 23 ears), otitis media with effusion (OME; 15 ears) or chronic suppurative otitis media (COM; 23 ears) were subjected to cytological (cellular differentiation, degree of phagocytosis) and bacteriological (species quantification and identification, degree of opsonization) analyses. Methods used were direct microscopy of acridine orange stained material, immunofluorescence assay using fluorescein-labelled antibodies to immunoglobulin G (IgG) and complement cleavage product C3b, and standard bacterial culturing on agar plates. The most intense opsonization and phagocytosis was evident in COM effusions culture-positive for Staphylococcus aureus. AOM and OME effusions showed rather poor opsonization and roughly 1% of the phagocytes harboured intracellular bacteria. COM effusions, culture-positive for Pseudomonas aeruginosa, showed no bacterial opsonization or phagocytosis whatsoever. Opsonization and phagocytosis of bacteria in the middle ear cleft is highly species-specific.

Sixteen patients with biochemically proven primary hyperparathyroidism (PHPT) underwent ultrasonography (US), fine-needle aspiration (FNA) for cytologic sampling (n = 9), or intact parathormone assay (n = 3) before operation (n = 15) in order to determine the accuracy of the methods. Pre-operative US was found sensitive (100 per cent), but two thyroid lesions were initially diagnosed as parathyroid tumours by US (i.e. false positives). Parathyroid cells were detected in six cytologic specimens, one sample was insufficient and another inconclusive, while one was diagnosed as thyroid tissue. Parathormone assay revealed a high hormone content in all three patients who underwent the procedure. We conclude that US is sufficiently sensitive to detect enlarged parathyroid tumours. Specificity can be improved by US-guided FNA for cytology or parathormone assay prior to neck exploration.

The proportions of secretory IgA (SIgA)-, IgG- and C3b-coated bacteria obtained from a well-defined area on the posterior wall of the nasopharynx (NPH) close to the Eustachian tube were determined. Samples taken from 25 otitis-prone (OP) and 25 non-otitis-prone (NOP) children with normal serum levels of IgA and IgG were evaluated using an immunofluorescence assay. Both groups harboured significantly more nasopharyngeal bacteria coated with IgG than with SIgA (p < 0.001). The OP children had significantly fewer SIgA-coated bacteria (p < 0.05) but more C3b-coated bacteria (p < 0.01) in the NPH than the NOP children had. No significant difference was noted between the two groups regarding IgG coating. The occurrence of Branhamella catarrhalis in the NHP was more pronounced in the OP group (p < 0.05). No significant differences in the occurrence of other middle ear pathogens (Streptococcus pneumoniae, Haemophilus influenzae, Staphylococcus aureus) or quantitative dominance of pathogens were noted between the two groups. Deficiency in SIgA coating of the nasopharyngeal bacteria may contribute to the otitis-prone condition.

Samples from the fur covering the palatine tonsils of 10 patients with infectious mononucleosis (IM) were subjected to fluorescence microscopy after staining with acridine orange. Standard aerobic cultures from swabs were set up simultaneously. The fur constituted a mass of variously sized cocci and rods intermingled with granulocytes and cellular detritus. Five patients had growth of beta-haemolytic Streptococci on the tonsils. Bacteria in abundance were found attached to the tonsillar epithelial cells. The B-lymphotrophic Epstein-Barr virus, responsible for the IM infection, causes an immense increase in the bacterial colonization of the tonsillar surfaces, probably due to an increased attachment of bacteria to the tonsillar epithelial cells.

Epithelial cells obtained by gentle scraping with a brush tip from the gingival margin of d 11 or 51, adjacent labial mucosa and buccal mucosa of children, adolescents and adults were stained with acridine orange and examined under a fluorescence microscope. The amount of bacteria appearing on the epithelial cells was assessed and the form of the adhered bacteria was evaluated. An age-related descending trend (negative Pearson's r in all sites) could be seen in the number of colonized cells (> 50 bacteria present) in gingival, labial and buccal areas. Adolescents and adults had significantly fewer epithelial cells with colonization of bacteria in the labial and buccal areas compared with children under 6 years. Only cocciformed bacteria were seen associated with the epithelial cells. By studying bacterial adhesion according to methods outlined in the present study, it should be possible to more closely investigate the bacteria-host relationships as they appear in nature.

Eighty mucoid effusion samples obtained from 56 patients with otitis media with effusion (OME) were subjected to quantitative and qualitative bacteriological analysis using standard culturing methods, direct microscopy and immunofluorescent assay. 30% of the samples contained culture-positive pathogens (H. influenzae, S. pneumoniae, B. catarrhalis), with counts never exceeding 5 x 10(5) per ml. In addition, 19% of the samples had dormant H. influenzae and S. pneumoniae, which did not grow on standard agar plates. Viable and dormant bacteria, as well as bacterial remnants, play a crucial role in the pathogenesis of OME and similarities between OME and reactive arthritis, i.e. Lyme arthritis, Reiter's syndrome and rheumatic fever, are evident.

Attachment of bacteria to nonciliated cells of the nasopharyngeal epithelium was evaluated by immunofluorescence assay in 25 otitis-prone and 25 non-otitis-prone children undergoing ear, nose, or throat surgery under general anesthesia. The bacterial findings were analyzed simultaneously, both qualitatively and quantitatively. In otitis-prone children, there was a significant preponderance of epithelial cells having greater than 50 attached bacteria (P less than .001) and of epithelial cells with attached Streptococcus pneumoniae and Haemophilus influenzae (P less than .05). Also, the occurrence of Branhamella catarrhalis in the nasopharynx was more pronounced in the otitis-prone group (P less than .05). No significant differences in the occurrence of other middle ear pathogens or quantitative dominance of pathogens were noted between the two groups. Abundant attachment of pathogens to the epithelial cells close to the nasopharyngeal orifice of the eustachian tube is of a significant factor for the development of the otitis-prone condition.

Quantification of bacteria in various types of middle ear effusion (MEE) obtained during current acute otitis media (AOM), otitis media with effusion (OME) and chronic suppurative otitis media (COM) was performed. The bacteria were stained with acridine orange and their number per ml effusion evaluated under the fluorescence microscope according to a method described in detail elsewhere. During AOM, 53% of the MEE samples were culture-positive and contained 10(6)-10(8) bacteria per ml (median value 10(7) per ml). During OME, serous effusion and 78% of the mucoid effusions contained no bacteria whatsoever, whereas the remaining mucoid effusions contained 10(4)-5 x 10(5) bacteria per ml (median value 10(4) per ml). Mucopurulent effusions contained 6 x 10(5)-10(8) bacteria per ml (median value 5 x 10(6) per ml). During COM, purulent MEE had 6 x 10(6)-10(9) bacteria per ml (median value 10(8) per ml). Quantification of bacteria involved in middle ear diseases provides further information about the etiopathogenesis and appropriate management of various pathological conditions of the middle ear.

Epithelial cells were obtained by swabbing the posterior wall of the nasopharynx (NPH) of 15 patients (age one to 6 8/12 years; eight males, seven females) undergoing ENT-surgery under general anaesthesia for otitis media with effusion (OME). Individually matched, ear healthy children served as controls. Bacteria attached to the non-ciliated cells were calculated according to a method described. Furthermore, quantitative and qualitative bacteriological analyses were performed on samples obtained from mucoid middle ear effusion material as well as from the NPH. All patients and controls harboured middle ear pathogens (S. pneumoniae, H. influenzae, B. catarrhalis, or S. aureus) in the NPH. Only 33 per cent of the patients harboured middle ear pathogens in the middle ear effusion and the same pathogen was invariably found in the corresponding NPH. Attachment of bacteria to the non-ciliated cells of the NPH diminished significantly with growing age in the ear healthy control group but not in the OME group. OME is closely correlated to the presence of middle ear pathogens in the NPH and to attachment of bacteria to the epithelial cells in the NPH.

Otitis media develops when certain bacterial pathogens gain access to the middle ear cavity from the nasopharynx through the eustachian tube. Adhesion of bacteria, in particular Streptococcus pneumoniae and Haemophilus influenzae, to the non-ciliated epithelial cells of the nasopharynx, close to the opening of the eustachian tube, is significantly correlated to the otitis-prone condition in children. Otitis-prone children have significantly fewer bacteria in the nasopharynx coated with the immunoglobulin secretory IgA (SigA) then healthy children have. Adhesion and occurrence of middle ear pathogens in the nasopharynx decreases with advancing age. Epstein-Barr virus, causative agent of infectious mononucleosis, causes a remarkable increase in bacterial adhesion to epithelial cells.

We used an immunofluorescence assay to investigate the content of secretory IgA- (SIgA), IgG-, IgM- and C3b-coated bacteria in middle ear effusions obtained within 12-72 h after the onset of acute symptoms of purulent otitis in 28 patients (37 ears). Simultaneously we analyzed the bacteria, both qualitatively and quantitatively, using standard culturing methods and fluorescein conjugated (FITC) antibodies to Haemophilus influenzae and Streptococcus pneumoniae. The ages of the patients were in the range of 5 months to 17 years; 18 were males and 10 females. 73% of the samples harboured no antibody- or C3b-coated bacteria, and particularly those of young patients (< 20 months). 92% of the samples were culture-positive, while 8% showed dormant bacteria. The predominant species were S. pneumoniae and H. influenzae. In 8% of the samples there was heavy and in 11% slight peripheral SIgA-coating of the 5% showed heavy and 19% slight peripheral IgG-coating. Only 3 samples were intensely opsonized, i.e. the bacteria were coated with IgG and C3b simultaneously. In most cases of acute purulent otitis media, the middle ear cavity of young individuals is not able to coat pathogens with SIgA, IgG, IgM and C3b during the early course of infection.

Attachment of Streptococcus pneumoniae and Haemophilus influenzae to epithelial cells on the posterior wall of the nasopharynx (NPH) was determined in 10 healthy children, culture-positive for either of these microorganisms. By using immunofluorescence technique and specific fluorescein-labelled antisera against these microorganisms, it was shown that in only 2 of the children studied were these pathogens firmly attached to the non-ciliated cells of the NPH. Attachment of S. pneumoniae and H. influenzae to the epithelial cells close to the nasopharyngeal opening of the Eustachian tube is of the utmost importance for the development of invasive disease, especially acute otitis media. Attachment of these pathogens to the epithelial cells covering the adenoid tissue may naturally be of significance for the induction of specific antibody body production against these microorganisms.

Samples of middle ear effusion were obtained from 16 patients with chronic suppurative otitis media (COM) (20 ears, age range from 1 1/12 to 69 years) and 13 patients with secretory otitis media (SOM) (15 years, age range from 8 months to 13 8/12 years). Qualitative and quantitative bacteriological analyses of the effusions were performed and the intensity of immunoglobulin G (IgG) and complement component C3b adhering to the bacteria was estimated using immunofluorescence technique. In COM effusions, S. aureus in particular was intensely opsonized, whereas Ps. aeruginosa showed minimal, if any, coating with IgG and C3b. SOM effusions exhibited extremely sparse IgG- and C3b-coated bacteria. The difference in IgG and C3b coated bacteria between COM and SOM effusions was significant (p less than 0.01). Specific (IgG) as well as non-specific (C3b) coating of the bacteria that have gained access to the middle ear cavity is of importance for the host defence.

Epithelial cells from the nasopharynx of 42 young healthy individuals were obtained by swabbing. The filtered cellular mixture was stained with acridine orange and bacteria attached to the epithelial cells were counted using a fluorescence microscope. Simultaneously qualitative and quantitative bacteriological analyses were performed. Young individuals (less than 2 years) had significantly more attached bacteria than older ones (11-15 years) (P = 0.0001). The presence of middle ear pathogens in the nasopharynx of young children was significantly more prominent than in the older age groups (P less than 0.001). Attachment of middle ear pathogens to nasopharyngeal epithelial cells of young individuals may predispose to otitis media.

SIgA- and IgG-coated bacteria obtained from 17 discharging middle ears (14 patients, 9 male, 5 female, age range from 1 to 79 years) were evaluated using an immunofluorescence assay. Simultaneously, quantitative and qualitative bacteriological analyses of the middle ear effusions (MEEs) were performed. MEEs containing Staphylococcus aureus harboured bacteria which were intensely coated with both SIgA and IgG antibodies. In contrast, MEEs containing Pseudomonas aeruginosa displayed minimal, if any, SIgA- and IgG-coated bacteria. Two young patients harboured bacteria (Haemophilus influenzae and S. aureus/Streptococcus pneumoniae, respectively) which were heavily coated with IgG, but not with SIgA. Immunoglobulin-coating of bacteria involved in otitis media is of the utmost importance in eradication of the infection.