Population structure of rat-derived Pneumocystis carinii in Danish wild rats.

The rat model of Pneumocystis carinii pneumonia is frequently used to study human P. carinii infection, but there are many differences between the rat and human infections. We studied naturally acquired P. carinii in wild rats to examine the relevance of the rat model for human infection. P. carinii DNA was detected in 47 of 51 wild rats and in 10 of 12 nonimmunosuppressed laboratory rats. Evidence for three novel formae speciales of rat-derived P. carinii was found, and these were provisionally named Pneumocystis carinii f. sp. rattus-secundi, Pneumocystis carinii f. sp. rattus-tertii, and Pneumocystis carinii f. sp. rattus-quarti. Our data suggest that low-level carriage of P. carinii in wild rats and nonimmunosuppressed laboratory rats is common and that wild rats are frequently coinfected with more than one forma specialis of P. carinii. We also examined the diversity in the internally transcribed spacer (ITS) regions of the nuclear rRNA operon of Pneumocystis carinii f. sp. carinii by using samples from wild rats and laboratory rats and spore trap samples. We report a lack of variation in the ITS1 and ITS2 regions that is consistent with an evolutionary bottleneck in the P. carinii f. sp. carinii population. This study shows that human- and rat-derived P. carinii organisms are very different, not only in genetic composition but also in population structure and natural history.

Experimental corticosteroid induction of Pneumocystis carinii pneumonia in piglets.

Animal models of Pneumocystis carinii (Pc) pneumonia (PCP) play a central role in research on the Pc microorganism itself and the disease, especially the pathogenesis and the host defence. The classic rat model with corticosteroid-induced reactivation of a latent infection has been most widely used. In our search for alternative non-rodent models, six 31/2-week-old piglets were injected intramuscularly with methylprednisolone acetate, at 18 mg/kg body weight, once a week for 6 weeks. Six littermate piglets constituted the control group. The principals showed a markedly lower growth rate than the controls. Furthermore, they developed "moon face" and "pot belly", snoring sounds while eating, and pronounced respiratory distress during handling. Significant changes in haematological parameters, including lymphopenia, were observed in the principal group. The Pc antibody titres of the controls increased to high levels, whereas the principals were all low-titred or seronegative for Pc at the last blood sampling. At necropsy, the mean body weight of the principals was about half that of the controls. In addition, they had an extreme reduction of the thymus together with dark red consolidations of the frontal lung lobes and/or atelectatic looking diaphragmatic lobes. Histopathologically, there was a focal interstitial pneumonia. Alveolar walls and interstitia had mononuclear cell infiltrations and the alveolar lumina were occluded by foamy acidophilic honeycomb material with a varying number of Pc cysts. The reduced body weight, the thymus involution, and the lymphopenia, together with the reduced levels of specific Pc antibodies and the histomorphology of the PCP, were consistent parameters of the principal group and comparable to the findings of the classic rat model. Thus, the present study is the first to describe that prolonged administration of high doses of methylprednisolone acetate can induce PCP in piglets.

Cultured rat and purified human Pneumocystis carinii stimulate intra- but not extracellular free radical production in human neutrophils.

The production of free radicals in human neutrophils was studied in both Pneumocystis carinii derived from cultures of L2 rat lung epithelial-like cells and Pneumocystis carinii purified from human lung. Using the cytochrome C technique, which selectively measured extracellular superoxide generation, hardly any free radical production was observed after stimulation with cultured rat-derived P. carinii. A chemiluminescence technique, which separately measured intra- and extracellular free radical production, was subsequently employed to differentiate the free radical generation. It was established that 1) P. carinii stimulated intra- but not extracellular free radical production in human neutrophils, 2) opsonized cultured rat-derived P. carinii stimulated human neutrophils to a strong intracellular response of superoxide production, and 3) opsonized P. carinii, purified from human lung also stimulated human neutrophils to produce intracellular free radicals.

Different ultrastructural morphology of Pneumocystis carinii derived from mice, rats, and rabbits.

Pneumocystis carinii (PC) is a fungus present in the lungs of many mammal species. Even though studies of the genome, the isoenzymes, and the antigens have proved some host-species-linked heterogeneity, the existence of distinct Pneumocystis species or subspecies has still not been accepted. Comparative studies of the ultrastructural morphology of pneumocysts derived from several host species may support evidence of host-species-linked heterogeneity. We have compared the ultrastructural morphology of pneumocysts derived from mice, rats, and rabbits. The density of membrane-limited electron-dense cytoplasmic granules was found to be higher in mouse-derived pneumocysts than in rabbit-derived pneumocysts, and furthermore the average diameter of the granules from mouse pneumocysts was larger than that of granules from rabbit-derived pneumocysts. The average diameter of the filopodia of mouse-derived pneumocysts was smaller than that of filopodia from rat-derived pneumocysts, which was smaller than that of filopodia from rabbit-derived pneumocysts. Globular electron-dense bulbous dilatations at the tip of the filopodia were described for the first time and they were only found on filopodia of mouse-derived pneumocysts. These distinct host-species-linked morphological differences of pneumocysts from mouse, rat, and rabbit may support previous biochemical data indicating the existence of different Pneumocystis species or subspecies.

Identification of porcine Pneumocystis carinii as a genetically distinct organism by DNA amplification.

DNA was amplified from lung samples from three piglets infected with Pneumocystis carinii, using oligonucleotide primers designed to the P. carinii mitochondrial large subunit ribosomal RNA gene. The nucleotide sequence of the amplification product was determined and indicated lack of sequence variation among these pig-derived P. carinii samples at this locus. The data showed that porcine P. carinii was genetically distinct from P. carinii isolated from other mammalian host species.

Pneumocystis carinii from pigs and humans are antigenically distinct.

The antigens of Pneumocystis carinii cysts isolated from pigs and humans were compared by the Western immunoblotting technique. Convalescent pig serum reacted with two antigens (approximately 78 kDa and 32.5 kDa) of porcine P. carinii cysts, whereas convalescent serum from humans did not react with porcine P. carinii cyst antigens. The results indicate that porcine and human P. carinii cysts are antigenically distinct.

A case of cerebral paragonimiasis in Denmark. Case report.

A case of cerebral paragonimiasis with severe neurological symptoms is presented. The patient, a 45-year-old woman, recovered completely after resection of a large cyst at the C3 level. The pathogenesis is discussed.

A survey of birds in Denmark for the presence of Pneumocystis carinii.

One hundred eighty-three toluidine blue O-stained necropsy lung imprint smears from different avian species were examined microscopically for Pneumocystis carinii. No cyst forms of the organism could be identified. Seventy-eight serum samples from a total of 155 chickens were examined by a competition enzyme-linked immunosorbent assay (ELISA) for antibodies to P. carinii; 53 serum samples were from individual chickens, and 25 samples were pools of sera from two to five chickens. Diluted 1:50, the 78 serum samples showed a specific ELISA-inhibition of 4% to 56% (the 95% confidence limit being 25% to 30% inhibition). Diluted 1:50, nine serum pools representing 34 chickens and 17 of the 53 individual serum samples (32.1%) showed an inhibition greater than 30%. No specific pneumocyst DNA could be detected in serum from 13 of the 53 chickens using polymerase chain reaction and dihydrofolate reductase gene as a specific probe. Specific antibodies to a 116,000-molecular-weight antigen of rat pneumocysts were shown in two (13.3%) of 15 individual chicken serum samples. The results indicate that P. carinii organisms do not commonly reside in the lungs of birds, although some birds may be exposed to external sources of organisms.

Antibodies to Pneumocystis carinii in Danish blood donors and AIDS patients with and without Pneumocystis carinii pneumonia.

Sera from 1) 8 AIDS patients with Pneumocystitis carinii pneumonia, 2) 13 HIV antibody-positive patients, 9 with AIDS and 4 with AIDS-related symptoms, all 13 without Pneumocystis carinii pneumonia, and 3) 36 healthy male blood donors, all HIV antibody negative, were examined for antibodies against Pneumocystis carinii using the indirect immunofluorescence antibody technique. Seven (87%) out of eight sera from group 1, 12 (92%) out of 13 sera from group 2, and 35 (97%) out of 36 sera from group 3 were positive. There was no significant difference between group 1 and 2, and group 2 and 3. The difference between group 1 and group 3 was significant (p = 0.03).

The piglet as a potential model of Pneumocystis carinii pneumonia.

Spontaneous Pneumocystis carinii infections occur in piglets. In this report we describe the symptoms, pathology and predisposing conditions of P. carinii pneumonia in the pig. We also discuss the advantages and disadvantages of the pig as an experimental system to study P. carinii pneumonia.

Host-parasite relationship in Pneumocystis carinii infection: activation of the plasmalemmal vesicular system in type I alveolar epithelial cells.

Ultrastructural studies of the attachment zone between Pneumocystis carinii (Pc) and type I alveolar epithelial cells showed a new aspect of the host-parasite relationship, i.e. an activation of the plasmalemmal vesicular system in the alveolar cells associated with Pc trophozoites in close apposition. This phenomenon may be involved in the nutrition of the trophozoite.

Human subcutaneous dirofilariasis caused by Dirofilaria repens. Report of a case in Norway and review of recent literature.

A case of dirofilariasis in a Norwegian man is reported. The causal agent was identified as Dirofilaria repens Railliet et Henry, 1911. To the best of our knowledge, this is the first report from a Scandinavian country on dirofilariasis caused by Dirofilaria repens. The infection was probably acquired in Majorca four years before it was diagnosed in Molde, Norway.

Morphology of Pneumocystis carinii and activation of the plasmalemmal vesicular system in alveolar epithelial cells of the host. An ultrastructural study.

Lungs from rats with dexamethasone-induced Pneumocystis carinii pneumonia were examined. The ultrastructure of Pneumocystis carinii and their zone of attachment on type I alveolar epithelial cells are described. An activation of the plasmalemmal vesicular system of type I alveolar epithelial cells was observed and is described here for the first time. The significance of this observation is discussed.

Pathological characteristics for the diagnosis of Pneumocystis carinii pneumonia. A retrospective autopsy study.

The clinical records of 63 patients who during the period 1980-1983 showed pneumocysts in lung specimen imprints obtained at autopsy have been reviewed in order to establish possible characteristics for the diagnosis of Pc pneumonia. Autopsies were performed at 2 Copenhagen hospitals, Rigshospitalet and the Finsen Institute. The 63 Pc-positive cases included 9 with blood diseases, 27 with solid tumors, 5 with immunological disorders and 22 with other diseases. The characteristic extensive changes in the lungs in Pc pneumonia included increased firmness, dark red/brown or black/grey colour of sectioned areas, stasis and edema sometimes excessive, and foci of white/grey or red/brown areas giving the lung a marbled or speckled appearance. The content of air was always decreased. The microscopic changes reported included intra-alveolar eosinophilic material, an intra-alveolar transudate containing macrophages and very few neutrophilic granulocytes, dilated capillary tubes, stasis and edema, thickened alveolar septae and peri-alveolar or diffuse fibrosis. The chest X-rays showed no specific features, and the infection had been subclinical in all cases; the only evidence of Pc infection was the demonstration of pneumocysts in imprints of lung specimens stained with toluidine blue in combination with observations during pathology. The diagnosis depends to a great extent upon a keen alertness in addition to the information given in the clinical record.

Survival periods in male and female Wistar rats with steroid-induced Pneumocystis carinii pneumonia.

Rats treated with corticosteroids developed Pneumocystis carinii pneumonia. Male rats showed the same mortality (100%) as female rats, but the female rats survived on average 11.7 days longer than the male Wistar rats.

Pneumocystis carinii in large domestic animals in Denmark. A preliminary report.

By microscopic examination of toluidine blue O stained autopsy lung imprints Pneumocystis carinii was shown in 6 calves out of 160 (3.8%), in 1 sheep out of 28 (3.6%) and in 8 pigs out of 119 (6.7%). This is the first report of Pneumocystis carinii in calves, sheep and pigs in Denmark.

Pneumocystis carinii in human lungs at autopsy.

To get some information about the prevalence of Pneumocystis carinii in the Danish population we have examined autopsy lung biopsies from patients at 2 large hospitals in Copenhagen, Righospitalet (RH) and Finseninstituttet (F), in a 5-year period from 1979 to 1984. Imprints made from the biopsies were stained with the toluidine blue O method and examined microscopically for pneumocysts. A total of 1762 patients were examined and pneumocysts found in 83 (4.7%); at RH 54 (4.9%) of 1106 patients and at F29 (4.4%) of 656 patients. A possible influence of certain climatic factors on the prevalence of pneumocysts in humans was shown: low temperature, low vapour pressure and low relative humidity seem to be correlated with low prevalence of pneumocysts. The general use of trimethoprim-sulfamethoxazole in the hospitals may possibly influence the prevalence of pneumocysts: the amount requested in the period of the investigation is negatively correlated with the prevalence of pneumocysts.

Pneumocystis carinii Delanoë & Delanoë, 1912 found in lungs of freeliving animals in Denmark at autopsy.

Stained imprints made from autopsy lung biopsies from hares, foxes, badgers, stone martens, one hedgehog, roe deer from Denmark and from caribou from Greenland were examined microscopically for Pneumocystis carinii. Pneumocysts were found in the lungs of 3 out of 28 hares, 3 out of 18 foxes and 1 out of 6 roe deer. This is the first report of Pneumocystis carinii in hares, foxes, and roe deer in Denmark. It seems to be the first report of Pneumocystis carinii in roe deer.