Putative progressive and abortive feline leukemia virus infection outcomes in captive jaguarundis (Puma yagouaroundi).

BACKGROUND: Feline leukemia virus (FeLV) is an exogenous gammaretrovirus of domestic cats (Felis catus) and some wild felids. The outcomes of FeLV infection in domestic cats vary according to host susceptibility, virus strain, and infectious challenge dose. Jaguarundis (Puma yagouaroundi) are small wild felids from South and Central America. We previously reported on FeLV infections in jaguarundis. We hypothesized here that the outcomes of FeLV infection in P. yagouaroundi mimic those observed in domestic cats. The aim of this study was to investigate the population of jaguarundis at Fundação Parque Zoológico de São Paulo for natural FeLV infection and resulting outcomes. METHODS: We investigated the jaguarundis using serological and molecular methods and monitored them for FeLV-related diseases for 5 years. We retrieved relevant biological and clinical information for the entire population of 23 jaguarundis held at zoo. Post-mortem findings from necropsies were recorded and histopathological and immunohistopathological analyses were performed. Sequencing and phylogenetic analyses were performed for FeLV-positive samples. For sample prevalence, 95% confidence intervals (CI) were calculated. Fisher's exact test was used to compare frequencies between infected and uninfected animals. P-values <0.05 were considered significant. RESULTS: In total, we detected evidence of FeLV exposure in four out of 23 animals (17%; 95% CI 5-39%). No endogenous FeLV (enFeLV) sequences were detected. An intestinal B-cell lymphoma in one jaguarundi was not associated with FeLV. Two jaguarundis presented FeLV test results consistent with an abortive FeLV infection with seroconversion, and two other jaguarundis had results consistent with a progressive infection and potentially FeLV-associated clinical disorders and post-mortem changes. Phylogenetic analysis of env revealed the presence of FeLV-A, a common origin of the virus in both animals (100% identity) and the closest similarity to FeLV-FAIDS and FeLV-3281 (98.4% identity), originally isolated from cats in the USA. CONCLUSIONS: We found evidence of progressive and abortive FeLV infection outcomes in jaguarundis, and domestic cats were probably the source of infection in these jaguarundis.

Experimental infection of Boa constrictor with an orthoreovirus isolated from a snake with inclusion body disease.

Orthoreoviruses have been associated with disease in reptiles, but have not previously been isolated from snakes with inclusion body disease (IBD). An orthoreovirus was isolated from a Boa constrictor diagnosed with IBD and then used to conduct a transmission study to determine the clinical importance of this virus. For the transmission study, 10 juvenile boas were experimentally infected with the isolated orthoreovirus and compared to 5 sham-infected control animals. Orthoreovirus was reisolated for a period of 18 wk after infection and weight gain was reduced in infected snakes. Histological examination showed a mild hepatitis in three of four virologically positive snakes up to 12 wk after infection. Results indicated that the orthoreovirus was moderately pathogenic, but, no evidence was found to indicate that it was the causal agent of IBD. In the light of the discovery of Arenaviruses in some snakes with IBD, it was proposed that orthoreoviruses may play a role in synergistic infection.

Nonuraemic nonfatal idiopathic calciphylaxis in a kitten.

BACKGROUND: Calciphylaxis is a rare cutaneous disorder, characterized by vascular calcification and progressive skin necrosis, not yet described in cats. It is scarcely reported in animals, mostly due to iatrogenic or uraemic disturbances of the calcium-phosphate balance. In human patients, it is most commonly seen with end-stage renal disease, but several nonuraemic disorders, including inherited dysfunctions of tissue calcification inhibitors, have also been described. HYPOTHESIS/OBJECTIVES: To describe a case of nonuraemic calciphylaxis in a cat. ANIMALS: A 10-week-old male domestic short hair kitten was presented with hyperacute skin lesions. Initial dermatological signs were characterized by sharp demarcated erosions and ulcerations on the face, including the nasal planum and lips. Cutaneous lesions rapidly progressed into thick crusts with ulcerations, involving parts of the face and pinna as well as abdominal skin. METHODS: Complete blood count, serum chemistry profile, urinalysis, parathyroid hormone measurement and histopathological examination of skin biopsies. RESULTS: Histopathology from newly developed abdominal skin lesions revealed severe epidermal necrosis and calcification, multifocal pannicular calcification and calcified subcutaneous vessels, supporting a diagnosis of calciphylaxis. Treatment consisted of systemic and topical antimicrobials, analgesics, pentoxifylline, Lantharenol(®) , sodium thiosulfate and vitamin K. After initiation of therapy, no further progression was noticed; all medications could be discontinued eventually, and no relapse was seen in the following 2 years. CONCLUSIONS AND CLINICAL IMPORTANCE: Calciphylaxis should be considered as a differential diagnosis for ulcerative cutaneous disorders in young cats. More information on this disease is needed to elucidate the pathomechanism.

Prevalence and spectrum of Johne's disease lesions in cattle slaughtered at two abattoirs in Kampala, Uganda.

This study was conducted to determine the prevalence and characteristics of Johne's disease (JD) lesions in Ugandan cattle slaughtered at two of the main abattoirs in Kampala. Ileocaecal junction and the associated lymph nodes of 1,022 cattle were examined for gross and microscopic lesions, followed by Ziehl Neelsen staining of the tissues bearing lesions. Confirmation of Mycobacterium avium subsp. paratuberculosis infection was done in some of the tissues using culture and IS900 PCR. The lesions were then described, characterised and tabulated. Characteristic Johne's disease granulomas were found in 4.7% of the samples examined, derived from Zebu, Ankole longhorn, Friesian breeds of cattle and their crosses. Lesions were found both in the lymph nodes and ileocaecal junction mucosa. The lesions tended to be more severe in the lymph node than in the mucosa. There were also some unique and atypical lesions found in association with Johne's disease granulomas. The diagnostic values of various gross lesions and criteria of lesion classifications and diagnosis are revisited and discussed based on the findings of this study. The prevalence of Johne's disease lesions among slaughtered cattle in Kampala's two abattoirs indicates that the disease is well established in the cattle population in the country. The diverse manifestations in lesions of JD need to be considered when making histological diagnosis in tissues where the disease is suspected.

Atherosclerosis aggravates ischemia/reperfusion injury in the gut and remote damage in the liver and the lung.

OBJECTIVE: We investigated whether mesenteric ischemia/reperfusion (I/R)-associated gut injury and remote liver and lung damage are affected by prevalent atherosclerosis. METHODS: Mesenteric ischemia was induced in atherosclerotic ApoE-deficient (ApoE(-/-)) and control C57BL/6 mice by clamping the superior mesenteric artery for 30 min. Mesenteric microcirculatory dysfunction and leukocytic inflammation were studied in the terminal ileum by intravital fluorescence microscopy (IVM). Histological analyses included quantitative assessment of parenchymal injury in the terminal ileum, liver and lung. RESULTS: In the gut, IVM of the terminal ileum revealed aggravated postischemic microcirculatory dysfunction and absence of reactive hyperemia-induced vasodilation in atherosclerotic mice compared to controls. In addition, leukocyte-endothelial cell adhesive interactions, i.e. rolling and firm adhesion, were significantly increased in atherosclerotic animals. This was associated with enhanced mucosal tissue damage in ApoE(-/-) mice. Moreover, mesenteric I/R-provoked remote parenchymal injury in the liver was found to be significantly aggravated in atherosclerotic mice. This was accompanied by enhanced neutrophilic lung inflammation in ApoE(-/-) mice. CONCLUSION: Prevalent generalized atherosclerosis not only aggravates splanchnic microcirculatory dysfunction and leukocytic inflammation in response to mesenteric I/R, but also exacerbates mucosal tissue damage and remote injury in the liver and the lung.

Dominance of highly divergent feline leukemia virus A progeny variants in a cat with recurrent viremia and fatal lymphoma.

BACKGROUND: In a cat that had ostensibly recovered from feline leukemia virus (FeLV) infection, we observed the reappearance of the virus and the development of fatal lymphoma 8.5 years after the initial experimental exposure to FeLV-A/Glasgow-1. The goals of the present study were to investigate this FeLV reoccurrence and molecularly characterize the progeny viruses. RESULTS: The FeLV reoccurrence was detected by the presence of FeLV antigen and RNA in the blood and saliva. The cat was feline immunodeficiency virus positive and showed CD4+ T-cell depletion, severe leukopenia, anemia and a multicentric monoclonal B-cell lymphoma. FeLV-A, but not -B or -C, was detectable. Sequencing of the envelope gene revealed three FeLV variants that were highly divergent from the virus that was originally inoculated (89-91% identity to FeLV-A/Glasgow-1). In the long terminal repeat 31 point mutations, some previously described in cats with lymphomas, were detected. The FeLV variant tissue provirus and viral RNA loads were significantly higher than the FeLV-A/Glasgow-1 loads. Moreover, the variant loads were significantly higher in lymphoma positive compared to lymphoma negative tissues. An increase in the variant provirus blood load was observed at the time of FeLV reoccurrence. CONCLUSIONS: Our results demonstrate that ostensibly recovered FeLV provirus-positive cats may act as a source of infection following FeLV reactivation. The virus variants that had largely replaced the inoculation strain had unusually heavily mutated envelopes. The mutations may have led to increased viral fitness and/or changed the mutagenic characteristics of the virus.

Congenital infiltrative lipomas in a calf.

The current report describes a case of congenital subcutaneous and intramuscular tumors of the neck and tail base in a 4-week-old female Angus-Charolais crossbred calf. Results of clinical and ultrasound examination are summarized. Biopsy and necropsy findings indicated an infiltrative lipoma. Congenital lipomas are uncommon tumors in bovids. Clinical and morphologic differentials, as well as classification and the possible pathogenesis of congenital neoplasms, are discussed.

Lack of Bcl-2 expression in feline follicular lymphomas.

Bcl-2, an anti-apoptotic protein, is commonly overexpressed in follicular lymphomas in humans. This is usually the result of a chromosomal translocation that transposes the Bcl-2 gene into the immunoglobulin gene locus. The immunohistochemical assessment of this overexpression can be used as a tool for the differentiation of follicular lymphoma and follicular hyperplasia. In cats, little information about the expression of Bcl-2 in follicular lymphoma exists. We investigated 18 follicular lymphomas histologically and immunohistochemically for the expression of Bcl-2, CD3, CD45R, and feline leukemia virus. Clonality was assessed by PCR for antigen receptor gene rearrangements. Although the histology resembled that of their human counterparts, diffuse expression of Bcl-2 within the follicles of the feline lymphomas, as seen in human cases, was not present. Only single cells within the follicles, comparable to the reactive controls, were positive for Bcl-2 expression. The mean survival time of 4.6 y confirmed the indolent character of the tumor. None of the clinical parameters assessed were statistically significant predictors of survival. Furthermore, a statistically significant difference in survival of animals with or without anti-neoplastic therapy was also not demonstrable.

Efficacy of a continuous, multiagent chemotherapeutic protocol versus a short-term single-agent protocol in dogs with lymphoma.

OBJECTIVE: To compare response rates and remission and survival times in dogs with lymphoma treated with a continuous, multiagent, doxorubicin-based chemotherapeutic protocol or with a short-term single-agent protocol incorporating doxorubicin. DESIGN: Nonrandomized controlled clinical trial. ANIMALS: 114 dogs with lymphoma. PROCEDURES: Dogs were treated with a chemotherapeutic protocol consisting of L-asparaginase, vincristine, cyclophosphamide, doxorubicin, methotrexate, and prednisolone (n=87) or doxorubicin alone (27). RESULTS: 63 of 86 (73%) dogs treated with the multiagent protocol (data on response was unavailable for 1 dog) and 14 of 27 (52%) dogs treated with the single-agent protocol had a complete remission. Dogs with lymphoma classified as substage

High resolution melting analysis (HRM) for the assessment of clonality in feline B-cell lymphomas.

Analysis of clonality is gaining importance in diagnosing lymphomas in veterinary medicine. Usually, PCR for the analysis of antigen receptor rearrangement (PARR) is followed by electrophoretic separation of the PCR products. Aim of this study was to test the feasibility of HRM for the assessment of clonality in B-cell lymphomas of cats. High resolution melting analysis differentiates PCR products by their different melting point using the decrease in fluorescence of an intercalating dye during melting of the PCR product. Additionally, the method is easy to use with no post-PCR manipulation of the samples. Forty-seven feline B-cell lymphomas and 31 reactive lymphatic proliferations of cats were investigated by PARR followed either by capillary electrophoresis or an HRM assay. To objectify the interpretation of the HRM results a recently published mathematical approach was applied to the melting curve. To overcome discrepancies between the visual interpretation and the mathematical approach, the latter was modified to include testing of reproducibility and recognition of pseudoclonality. In 11 of 47 lymphoma cases clonal populations were detectable by HRM assay compared to 14 of 47 lymphomas in which clonal populations were detected by capillary electrophoresis assay. Neither of the methods showed a clonal pattern in any of the reactive samples. However, the HRM assay showed a unique pattern in cases of follicular lymphatic hyperplasia that had no corresponding pattern in capillary electrophoresis. CONCLUSION: The capillary electrophoresis assay could identify 3 lymphomas that were not detected by the HRM assay and is therefore regarded superior to the HRM assay. The comparison however, was hampered by the overall bad performance of the PARR, that might be the consequence of insufficient primer binding due to somatic hypermutation of the binding sites during antigen stimulated proliferation of the B lymphocytes.

MHC class II expression by follicular keratinocytes in canine demodicosis--an immunohistochemical study.

MHC class II proteins present fragments of extra cellular antigen to stimulate CD4(+) T lymphocytes. Aim of this study was the detection of MHC class II antigens on different cutaneous cells in canine demodicosis. Histopathological and immunohistochemical examination of skin biopsies from 44 dogs with demodicosis is reported. The control group consisted of skin biopsies taken from 10 necropsied dogs without obvious skin lesions. The immunohistological assessment of the MHC class II expression revealed MHC class II proteins on different cell types of infiltrating inflammatory cells, i.e. APCs (antigen-presenting cells), macrophages, T lymphocytes and B lymphocytes. The plasma cells, however, only showed expression in 32 (73%) of 44 cases. Generally it was noticeable that most plasma cells but never all of them expressed MHC class II. Neutrophils, mast cells and eosinophils were MHC class II negative. Furthermore, in 39 biopsies (89%) from dogs with demodicosis MHC class II positive follicular keratinocytes were found. The control group did not show MHC class II expression on epithelial cells. Concerning the endothelial cells, a total of 25 biopsies (57%) showed MHC class II expression in which different vascular plexuses were affected by staining. This examination shows that MHC class II expression in the skin of dogs suffering form demodicosis is elevated. Especially the MHC class II expression by follicular keratinocytes seems to be conspicuous. We hypothesize that this is in association with the development and the maintenance of follicular inflammation.

Malignant renal schwannoma in a cat.

A nine-year-old male European shorthair cat with rapidly enlarging mass at the left kidney doubted to be malignant was presented. The purpose of this study is to present the clinical, radiological and pathological findings of a primary renal tumor in the cat. Grossly, the mass mostly encapsulated the kidney. Histologically, excisional biopsy showed worrying histological features. A sarcoma-like tumor composed mainly of neoplastic spindle-shaped cells. Neoplastic nodules of aggregations of fusiform cells arranged in multidirectional bundles. Immunohistochemically, several immunohistochemical satins (melan-A, S-100, vimentin, actin, desmin, cytokeratin, neurofilament, melan-A, NSE, synaptophysin, chromogranin, Glial Fibrillary Acidic Protein GFAP, Collagen IV and CD99) were used to differentially diagnose the mass. The stained neoplastic sections positively tested to S-100, but negative to the other aforementioned immunohistochemical stains. Immunohistochemistry with S-100 antibody staining showed an unusually strong positive reaction throughout the tumor cells. Based on our comparative diagnosis relative to other tumors, in addition to the progressive clinical signs, histopathological and immunohistochemical results, this case was presumptively diagnosis as a malignant schwannoma. According to our investigation of the relevant literature, this study of malignant renal Schwannoma (malignant peripheral nerve sheath tumor) is a highly rare case not previously characterized in a cat.

Natural feline coronavirus infection: differences in cytokine patterns in association with the outcome of infection.

Natural and experimental feline coronavirus (FCoV) infection leads to systemic viral spread via monocyte-associated viraemia and induces systemic proliferation of monocytes/macrophages. In the majority of naturally infected animals, FCoV infection remains subclinical and is associated with generalised B and T cell hyperplasia, but no other pathological findings. A minority of cats, however, develop feline infectious peritonitis (FIP), a fatal systemic granulomatous disease. This is generally accompanied by B and T cell depletion. The obvious functional differences of lymphatic tissues in FCoV-infected cats with and without FIP suggest that they contribute to the outcome of FCoV infection. This study attempted to evaluate the functional changes in haemolymphatic tissues after natural FCoV infection, with special emphasis on the magnitude, phenotype and function of the monocyte/macrophage population. The spleen, mesenteric lymph nodes and bone marrow from naturally FCoV-infected cats with and without FIP and specific pathogen-free (SPF) control cats were examined for the quantity and activation state of monocytes/macrophages both by immunohistology and by quantitative real time PCR for the transcription of interleukin (IL)-1beta, IL-6, IL-10, IL-12 p40, tumour necrosis factor (TNF), granulocyte colony stimulating factor (G-CSF), macrophage-CSF (M-CSF) and GM-CSF. Compared to cats with FIP, FCoV-infected cats without FIP exhibited significantly higher IL-10 levels in the spleen and significantly lower levels of IL-6, G- and M-CSF in mesenteric lymph nodes. In cats with FIP, however, IL-12 p40 levels were significantly lower in lymphatic tissues in comparison to both SPF cats and FCoV-infected cats without FIP. In comparison to SPF cats, FIP cats had significantly higher IL-1beta levels and lower TNF levels in mesenteric lymph nodes and lower M-CSF levels in the spleen. Findings indicate that FCoV-infected cats which do not develop FIP are able to mount an effective FCoV-specific immune response and can avoid excessive macrophage activation and FIP, possibly by upregulation of IL-10 production. Development of FIP, however, might be due to a lack of IL-12 which inhibits an effective cellular immune response and allows for monocyte/macrophage activation and the development of FIP.

Natural FCoV infection: cats with FIP exhibit significantly higher viral loads than healthy infected cats.

Natural feline coronavirus (FCoV) infection has been shown to not only induce intestinal infection with viral shedding, but also systemic infection which either remains without clinical signs or leads to feline infectious peritonitis (FIP). As systemic infection is not the key event in the development of FIP, the question arises as to whether a potential difference in viral load might be of importance. Therefore, the purpose of this study was to quantitatively assess feline coronavirus (FCoV) RNA loads in haemolymphatic tissues of healthy, long-term FCoV-infected cats and cats with FIP. In cats that died from FIP, viral loads were significantly higher, indicating a higher rate of viral replication or a reduced capacity for viral clearance in cats developing and/or suffering from FIP.

Dysuria due to discospondylitis and intervertebral disc herniation in a male alpaca (Vicugna pacos).

BACKGROUND: Dysuria in camelids is usually associated with the presence of lower urinary tract disease such as urolithiasis. As another differential diagnosis, urine retention may be caused by neurological disturbances resulting from infections of the spinal cord, discospondylitis or trauma. CASE PRESENTATION: A 2.5-year-old male Huacaya alpaca (Vicugna pacos) presented with dysuria due to damage of the lumbosacral intumescence of the spinal cord. On presentation the alpaca was recumbent. Clinical examination revealed abdominal pain, oliguria, leucopenia with neutrophilia, and slightly elevated creatinine kinase. Ultrasonography of the abdomen showed an irregularly shaped, dilated urinary bladder with hyperechoic serosa. Magnetic resonance imaging revealed discospondylitis of the fourth and fifth lumbar vertebrae and herniation of the intervertebral disc between these vertebrae and the spinal cord. Postmortem examination confirmed severe chronic purulent discospondylitis with ventral spondylosis and narrowing of the spinal canal. Urolithiasis could not be verified. CONCLUSION: Although rare, diseases of the spinal cord should be considered as a differential diagnosis for impaired micturition in camelids.

Identification of a novel feline large granular lymphoma cell line (S87) as non-MHC-restricted cytotoxic T-cell line and assessment of its genetic instability.

Feline large granular lymphocyte lymphomas are rare but very aggressive tumors with a poor prognosis. In this study, a cell line from an abdominal effusion of a cat with large granular lymphoma was characterized. Immunophenotype staining was positive for CD3 and CD45R, and negative for CD4, CD8, CD56, CD79α, BLA.36 and NK1. A TCR γ gene rearrangement was detectable by PARR. Neither FeLV antigen nor exogenous FeLV provirus could be detected. A chromosomal instability associated with a centrosome hyperamplification could also be determined. The cell line is able to lyse target cells without antigen presentation or interaction with antigen presenting cells. Therefore, these cells were classified as genetically instable non-MHC-restricted cytotoxic T cells with large granular lymphocyte morphology.

Detection of porcine circovirus type 2 and its association with PMWS in wild boars and domestic pigs in Germany: a histopathological, immunohistochemical and molecular biological study.

Beside domestic pigs wild boars can also be affected by postweaning multisystemic wasting syndrome (PMWS). For the first time a nationwide survey of wild boars (n = 356) and domestic pigs (n = 340) was carried out in Germany by histopathology, immunohistochemistry (IHC) and quantitative PCR (qPCR). Whereas 102/340 domestic pigs were immunoreactive for PCV2 antigen in at least one examined tissue, only 8/356 wild boars reacted positively. Similar findings could be found in qPCR: all domestic pigs showed viral DNA in at least one tissue, while in the examined tissues of 170 wild boars PCV2-DNA was not detectable. The specimens were examined histologically for histiocytosis and depletion of lymphocytes, both typical for PMWS. Based on these findings, six wild boars and 69 domestic pigs were assumed to be affected by PMWS.

Comparison of different tests to diagnose feline infectious peritonitis.

Clinical data from 488 cats (1979-2000) with histopathologically confirmed feline infectious peritonitis (FIP) and 620 comparable controls were evaluated retrospectively to assess the value of several diagnostic tests frequently used in the evaluation of cats with suspected FIP. Diagnostic utility of serum albumin to globulin ratio for the diagnosis of FIP was greater than of the utility of serum total protein and gamma-globulin concentrations. Diagnostic utility of these variables was higher when performed on effusion. On effusion, positive and negative predictive values of Rivalta's test, a test that distinguishes between exudates and transudates (0.86 and 0.97), anti-coronavirus antibody detection (0.90 and 0.79), and immunofluorescence staining of coronavirus antigen in macrophages (1.00 and 0.57) were investigated. The positive and negative predictive values of presence of anti-coronavirus antibodies were 0.44 and 0.90, respectively, antibody concentrations (1:1,600) were 0.94 and 0.88. presence of immune complexes measured by a competitive enzyme-linked immunosorbent assay were 0.67 and 0.84, and detection of viral RNA by serum reverse-transcriptase polymerase chain reaction (RT-PCR) were 0.90 and 0.47. Effusion RT-PCR was performed in 6 cats; it was positive in all 5 cats with FIP and negative in the cat with another disease. Diagnostic assays on the fluid in cats with body effusion had good predictive values. Definitive diagnosis of FIP on the basis of measurement of various variables in serum was not possible. Serum tests can only be used to facilitate the decision for more invasive diagnostic methods.

Identification of T cell receptor signaling pathway proteins in a feline large granular lymphoma cell line by liquid chromatography tandem mass spectrometry.

Tryptic peptides of a feline large granular lymphoma cell line were analyzed by liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS). Seventeen proteins of the T cell receptor signaling pathway could be identified by this approach. So far the existence of these proteins has only been postulated in the protein databases while experimental proof of their expression is predominantly pending. This article suggests where these proteins are located within the T cell receptor signaling pathway, thereby giving a short overview of the structure and function of this cascade.

Occurrence and severity of lung lesions in slaughter pigs vaccinated against Mycoplasma hyopneumoniae with different strategies.

Different vaccination strategies against Mycoplasma hyopneumoniae have been adopted worldwide. Reports from the field indicate varying levels of protection among currently available vaccines. The goal of the present study was to compare the efficacies of three widespread commercial vaccination strategies against M. hyopneumoniae under field conditions. 20 farms were included. 14 farms used different single dose vaccines (vaccine 1 [V1], 8 herds; vaccine 2 [V2], 6 herds); another 6 farms (V3) used a two dose vaccination strategy. Gross lesions of 854 lungs and histopathology from 140 lungs were quantified, and a quantitative PCR was applied to detect M. hyopneumoniae and porcine circovirus 2 (PCV2) DNA in lung tissue (n=140). In addition, porcine reproductive and respiratory disease virus (PRRSV), swine influenza virus (SIV), Actinobacillus pleuropneumoniae, Haemophilus parasuis and Pasteurella multocida were tested by qualitative PCR. 53% of lungs were positive for M. hyopneumoniae. 55.9% of lungs showed macroscopic enzootic pneumonia (EP)-like lesions. Lung lesion scores (P<0.001) and M. hyopneumoniae-loads (P<0.008) differed significantly among the vaccination groups, with the most severe cases and highest amounts occurring in V1. Histological alterations differed (P<0.001) between V1 and V3. Lung lesion scores and histopathological changes were significantly correlated, with prevalence and load of M. hyopneumoniae indicating that the applied diagnostic tools are valuable in confirming the prevalence and severity of M. hyopneumoniae infections. Comparing different vaccination strategies against M. hyopneumoniae indicates varying levels of protection. M. hyopneumoniae is still a major problem despite the widely applied vaccination.