An untapped potential for imaging of peripheral osteomyelitis in paediatrics using [18F]FDG PET/CT -the inference from a juvenile porcine model.

PURPOSE: To examine parameters affecting the detection of osteomyelitis (OM) by [18F]FDG PET/CT and to reduce tracer activity in a pig model. BACKGROUND: [18F]FDG PET/CT is recommended for the diagnosis of OM in the axial skeleton of adults. In children, OM has a tendency to become chronic or recurrent, especially in low-income countries. Early diagnosis and initiation of therapy are therefore essential. We have previously demonstrated that [18F]FDG PET/CT is promising in juvenile Staphylococcus aureus (S. aureus) OM of peripheral bones in a pig model, not failing even small lesions. When using imaging in children, radiation exposure should be balanced against fast diagnostics in the individual case. METHODS: Twenty juvenile pigs were inoculated with S. aureus. One week after inoculation, the pigs were [18F]FDG PET/CT scanned. PET list-mode acquired data of a subgroup were retrospectively processed in order to simulate and examine the image quality obtainable with an injected activity of 132 MBq, 44 MBq, 13.2 MBq, and 4.4 MBq, respectively. RESULTS: All lesions were detected by [18F]FDG PET and CT. Some lesions were very small (0.01 cm3), and others were larger (4.18 cm3). SUVmax was higher when sequesters (p = 0.023) and fistulas were formed (p < 0.0001). The simulated data demonstrated that it was possible to reduce the activity to 4.4 MBq without compromising image quality in pigs. CONCLUSIONS: [18F]FDG PET/CT localized even small OM lesions in peripheral bones. It was possible to reduce the injected activity considerably without compromising image quality, impacting the applicability of PET/CT in peripheral OM in children.

A clinical and histopathological comparison of the effectiveness of salicylic acid to a compound of inorganic acids for the treatment of digital dermatitis in cattle.

Bovine digital dermatitis (DD) is a painful infectious disease, causing lameness, reduced animal welfare, and production losses in dairy herds. The main factors contributing to DD are an infection with Treponema spp. and poor hygiene. Topical treatment has primarily consisted of antibiotics; however, the demand for effective nonantibiotic alternatives is increasing. The objective was to evaluate the performance of 3 nonantibiotic topical treatments (salicylic acid and a compound of inorganic acids in a 20% solution and in a dry form) on DD in a commercial dairy herd. Within the 30-d test period, 42 DD lesions on 33 Holstein cows were assigned to receive 1 of the 3 treatments. Lesions were biopsied before and after treatment and were clinically evaluated 5 times. Improved lesions were clinically defined as either healed (regeneration of the skin) or healing (dry lesions covered by a scab). Unhealed lesions were defined as either active [with a raw, moist, strawberry-like (granulating) surface] or mature (with a raised papillomatous appearance). The effectiveness of treatment was evaluated histopathologically using the following scores: 0 (no spirochetes present), 1 (small number of spirochetes present in the epidermis), 2 (moderate number of spirochetes present and reaching an intermediary level in the epidermis), and 3 (large number of spirochetes present and reaching the deepest part of the epidermis or the superficial dermis). The improvement rate was 10/14 (71%) for salicylic acid, 11/15 (73%) for the inorganic acid solution, and 8/13 (62%) for the inorganic acid powder. The analysis showed no difference among treatments. The association between clinical score and histopathological score was determined by an odds ratio. The odds ratio of a healed lesion having spirochetes in the epidermis was 0.58 and that of an active DD lesion having spirochetes in the epidermis was 26.5.

Expression of tissue factor in canine mammary tumours and correlation with grade, stage and markers of haemostasis and inflammation.

Tissue factor (TF) expression in human cancers has been associated with a procoagulant state and facilitation of metastasis. This study was conducted in order to evaluate if TF was expressed in canine mammary tumours. Forty epithelial mammary tumours from 28 dogs were included. TF expression of the tumours was evaluated by immunohistochemistry using a polyclonal antibody against recombinant canine TF. In addition, thromboelastography, haemostatic and inflammatory parameters were evaluated in the patients. TF was recognized in 44% of benign and 58% of malignant tumours. TF localized to the cytoplasmic membrane of neoplastic luminal epithelial cells and/or diffusely in the cytoplasm. No association was found between TF expression and stage or grade of disease. A significant association between TF expression and antithrombin and plasminogen was found, and extensive TF expression was seen in a lymph node metastasis classified as anaplastic mammary carcinoma from a dog with concomitant disseminated intravascular coagulation (DIC).

Disseminated intravascular coagulation in a novel porcine model of severe Staphylococcus aureus sepsis fulfills human clinical criteria.

Sepsis is a common and often fatal complication in human patients in intensive care units. Relevant and well characterized animal models of sepsis may provide valuable information on pathophysiological mechanisms and be a mean of testing new therapeutic strategies. Large animal models of Staphylococcus aureus sepsis are rare, even though S. aureus increasingly affects human patients. Sepsis changes the haemostatic balance and leads to endothelial cell (EC) activation, coagulopathy and, in severe cases, disseminated intravascular coagulation (DIC). The aim of this study was to characterize the haemostatic and vascular alterations in a novel porcine model of severe S. aureus sepsis, investigating whether the changes fulfill the human clinical criteria for DIC. Five pigs were inoculated intravenously with S. aureus and two control animals were sham-inoculated. Blood samples were collected for thromboelastography (TEG) and assessment of plasma-based haemostatic parameters. Tissue was collected for histopathology and reverse transcriptase quantitative real-time polymerase chain reaction for measurement of mRNA encoding EC markers. All infected animals developed DIC; including procoagulant activation represented by hypercoagulable TEG profiles and prolonged clotting time. Histologically, numerous pulmonary thrombi were present in one pig. Inhibitor consumption was represented by decreasing antithrombin levels in infected pigs. Hyaline globules were found in three infected pigs, confirming fibrinolytic activation. EC activation was identified by expression of von Willebrand factor in small vessels together with elevated mRNA encoding activated EC markers. Severe haemostatic and vascular changes fulfilling the human criteria for DIC were therefore seen in all infected pigs. A tendency towards uncompensated DIC was seen in two animals.

Detection of porcine circovirus type 2 and viral replication by in situ hybridization in primary lymphoid organs from naturally and experimentally infected pigs.

Porcine circovirus type 2 (PCV2) infection is the cause of postweaning multisystemic wasting syndrome (PMWS). It has been speculated whether cell types permissive of replication are found in the primary lymphoid organs and whether infection of these tissues has an important role in the pathogenesis of PMWS. The aim of this study was to determine if primary lymphoid organ cells support viral replication during PCV2 infection. This was done by histopathological examination of thymus and bone marrow from pigs experimentally inoculated with PCV2 (n = 24), mock-infected pigs (n = 12), pigs naturally affected by PMWS (n = 33), and age-matched healthy control animals (n = 29). In situ hybridization (ISH) techniques were used to detect PCV2 nucleic acid irrespective of replicative status (complementary probe, CP) or to detect only the replicative form of the virus (replicative form probe, RFP). PCV2 was not detected in the experimentally PCV2-inoculated pigs or the control animals. Among the PMWS-affected pigs, 19 of 20 (95%) thymuses were positive for PCV2 by CP ISH, and 7 of 19 (37%) of these also supported viral replication. By CP ISH, PCV2 was detected in 16 of 33 (48%) bone marrow samples, and 5 of 16 (31%) of these also supported replication. The 2 ISH probes labeled the same cell types, which were histiocytes in both organs and lymphocytes in thymus. The RFP labeled fewer cells than the CP. Thus, PCV2 nucleic acids and replication were found in bone marrow and thymus of PMWS-affected pigs, but there was no evidence that primary lymphoid organ cells are major supporters of PCV2 replication.

Pathology and biofilm formation in a porcine model of staphylococcal osteomyelitis.

A porcine model was used to examine the potential of human and porcine Staphylococcus aureus isolates to induce haematogenously spread osteomyelitis. Pigs were inoculated in the right femoral artery with one of the following S. aureus strains: S54F9 (from a porcine lung abscess; n = 3 animals), NCTC-8325-4 (a laboratory strain of human origin; n = 3 animals) and UAMS-1 (a human osteomyelitis isolate; n = 3 animals). Two pigs were sham inoculated with saline. At 11 or 15 days post infection the animals were scanned by computed tomography before being killed and subjected to necropsy examination. Osteomyelitis lesions were present in the right hind limb of all pigs inoculated with strain S54F9 and in one pig inoculated with strain NCTC-8325-4. Microscopically, there was extensive loss of bone tissue with surrounding granulation tissue. Sequestrated bone trabeculae were intermingled with colonies of S. aureus as demonstrated immunohistochemically. By peptide nucleic acid fluorescence in situ hybridization bacterial aggregates were demonstrated to be embedded in an opaque matrix, indicating that the bacteria had formed a biofilm. Development of experimental osteomyelitis was therefore dependent on the strain of bacteria inoculated and on the formation of a biofilm.

The impact of Staphylococcus aureus concentration on the development of pulmonary lesions and cytokine expression after intravenous inoculation of pigs.

Acute respiratory distress syndrome is a common complication in severe sepsis. In pigs, the lungs play an important role in clearing systemic bacterial infections due to pulmonary intravascular macrophages found specifically in pigs. However, this increases the exposure of the porcine lungs to pathogens and potential injury. The authors propose that increasing the concentration of the inoculum without changing the bacterial dose will lead to severe sepsis with pronounced pulmonary lesions. This could potentially create a risk of cytokine spillover to the circulation, leading to an increased systemic response. Eight Danish Landrace pigs, approximately 10 weeks old, were inoculated twice with a low or once with a high concentration of Staphylococcus aureus. Three pigs were sham-inoculated. The animals were grouped based on macro- and microscopic lung lesions. The mRNA expression of local pulmonary inflammatory markers was compared to protein levels of systemic inflammatory markers. The most severe pulmonary lesions were observed in animals receiving the high S. aureus concentration, indicating that severity of lesions is dependent on inoculum concentration rather than total numbers of bacteria. Furthermore, local mRNA expression of inflammatory cytokines appeared to be dependent on the magnitude and severity of tissue destruction, including the ability to confine the lesions. Increasing mRNA levels of serum amyloid A could be a confident marker of severity of pulmonary lesions. Since no correlation was observed between local and systemic levels of inflammatory cytokines, this finding could indicate an ability of the porcine lung to compartmentalize the local inflammatory response and thus restrict systemic contribution.

Exonization of a LINE1 fragment implicated in X-linked hypohidrotic ectodermal dysplasia in cattle.

A case of X-linked hypohidrotic ectodermal dysplasia (XHED) was identified in a family of Danish Red Holstein cattle. The ectodysplasin-signalling protein (EDA) is known to be central in the normal development of ectodermal structures, and mutations in the ectodysplasin A (EDA) gene have been reported to cause XHED. In this study, we analysed different EDA transcript variants in affected and unaffected cattle and identified a new transcript variant including a LINE1-derived pseudoexon between EDA exons 1 and 2. The 161-bp-long pseudoexon introduces a shift in reading frame and a premature stop codon early in EDA exon 2 and is probably the cause of XHED in this Danish Red Holstein family.

An investigation of the pathology and pathogens associated with porcine respiratory disease complex in Denmark.

Respiratory infections are among the most important diseases of growing pigs. In order to elucidate the multifactorial aetiology of porcine respiratory disease complex (PRDC) in Denmark, lungs from 148 finishing pigs with cranioventral bronchopneumonia (case group) and 60 pigs without lung lesions (control group) were collected from abattoirs. The pathogens involved in PRDC and their interactions were identified and linked to the histopathological diagnosis. The lung samples were cultured for bacteria and tested by multiplex polymerase chain reaction for presence of swine influenza virus (type A), porcine reproductive and respiratory syndrome virus (both European and US type), porcine circovirus type 2 (PCV2), porcine respiratory coronavirus, porcine cytomegalovirus, Mycoplasma hyopneumoniae and Mycoplasma hyorhinis. All cases had cranioventral lobular bronchopneumonia consistent with PRDC. There was a broad range of microscopical lesions and the cases were characterized as acute (n=10), subacute (n=24) or chronic (n=114) bronchopneumonia. Five bacterial species, five viruses and two Mycoplasma spp. were detected in different combinations. PCV2, M. hyopneumoniae, M. hyorhinis and Pasteurella multocida were detected most frequently among the PRDC affected swine and the diversity and number of pathogens were higher in these animals compared with controls. No clear-cut associations were detected between pathogens and histological lesions or histopathological diagnoses. PRDC occurs more frequently than enzootic pneumonia among Danish finishing pigs and has complex and varied histopathology.

Occurrence and tissue distribution of porcine circovirus type 2 identified by immunohistochemistry in Danish finishing pigs at slaughter.

Infection with porcine circovirus type 2 (PCV2) may be subclinical or lead to the development of porcine circovirus disease (PCVD), which includes the entities of post-weaning multisystemic wasting syndrome (PMWS) and the porcine respiratory disease complex (PRDC). PCV2 infection and PMWS occur in the early post-weaning period and are also recognized in finishing pigs of 12-19 weeks of age. The aim of the present study was to assess the role of PCV2 infection in disease of finishing pigs. Accordingly, the occurrence and tissue distribution of PCV2 was examined in Danish finishing pigs at the time of slaughter. Multiple lymph nodes and the spleen, lungs and kidneys from 136 pigs with PRDC (case group) and 36 pigs without lung lesions (control group) were examined by immunolabelling for the presence of PCV2. Additionally, follicular dendritic cells (FDC) were identified immunohistochemically. One or more tissues of 61% of the pigs were positive for PCV2 antigen. Up to 78% of the pigs had mild lymphoid depletion, indistinct lymphoid follicles and/or histiocytic infiltration of the lymph nodes, but these lesions were not associated with PCV2. No association was found between the presence of lung or kidney lesions and detection of PCV2. Three distinct patterns of cellular PCV2 antigen labelling were recognized: (1) labelling of cells with stellate morphology and reticular distribution, (2) labelling of isolated non-epithelial, cells, and (3) epithelial labelling. The reticular pattern was most common and localized to the centres of lymphoid follicles, corresponding to the morphology and distribution of FDCs. This observation may be interpreted to suggest that PCV2 may interact with FDCs to cause depletion of B lymphocytes. Alternatively, the FDCs may be a reservoir of infective PCV2 in subclinically infected animals or represent a simple storage site for PCV2 antigen in pigs that have recovered from infection.

Polysynovitis after oligofructose overload in dairy cattle.

Acute bovine laminitis is a systemic disease with local manifestations primarily affecting the claws. However, distension of the tarsocrural joints has been observed after experimental oligofructose overload in dairy heifers as a part of the complex interpreted as acute, clinical laminitis. Therefore, the aim of the present study was to study bovine synovial joints and tendon sheaths after oligofructose overload. Ten dairy heifers received oral oligofructose overload (17 g/kg body weight); four were killed 24h after overload and six after 72 h. Six control heifers received tap water and were killed after 72 or 96 h. Clinical examination included locomotion scoring and palpation of the tarsocrural joints. Ruminal fluid and blood was collected for measurements of pH and hydration status. Total protein concentrations and white blood cell (WBC) counts were determined in synovial fluid collected from tarsocrural joints after death. Synovial joints and tendon sheaths were examined and synovial membranes were studied microscopically. Swabs taken from the synovial cavities were subject to bacteriological culture. Heifers with oligofructose overload developed signs of ruminal and systemic acidosis. Lameness was observed in three of ten heifers 24h after overload and in all remaining heifers after 72 h. Distension of tarsocrural joints was observed from 18 h after overload and peaked at 30 h when all examined joints were moderately or severely distended. The synovial fluid was turbid and protein content and WBC counts were increased at both 24 and 72 h compared with controls. Bacterial culture was negative. Synovial membranes 24 and 72 h after overload had a fibrinous and neutrophil inflammatory reaction that regressed in severity between 24 and 72 h after overload. Heifers subjected to oligofructose overload therefore developed generalized sterile neutrophilic polysynovitis. Focus on this aspect of bovine laminitis may shed new light on the pathogenesis of this complex disease.

Investigation of chicken intestinal bacterial communities by 16S rRNA targeted fluorescence in situ hybridization.

The aim of the investigation was to quantify selected dominant bacterial groups in the chicken intestinal tract. Conventional production was used as model and the effect of the supplement with Salinomycin was evaluated. Hybridization conditions were optimized for published probes with respect to a panel of reference bacteria. In chicken intestinal samples bacteria were quantified by fluorescence in situ hybridization with 16S rRNA oligonucleotides directed towards bacteria related to Lactobacillus, Bacillus, Enterococcus-Streptococcus-Lactococcus, Enterobacteriaceae, Bacteroides, Clostridium and the domain Bacteria in lumen of ileum and cecum as well as on the intestinal wall including mucus of four individuals. Salinomycin in feed reduced counts of the Lactobacillus-, Enterobacteriaceae- and Clostridium-like bacteria in lumen of ileum compared to the conventional control. Increased or decreased bacterial counts were registered by Salinomycin in the ceca compared to the control. Relatively higher counts of Bacteroides- and Clostridium-like bacteria were found on the intestinal wall including mucus compared to lumen. The increase in numbers of some bacterial groups as well as the expected reduction by Salinomycin and the observed difference in the relative distribution of bacteria between lumen and intestinal wall are new observations that will need further investigation.

Immunohistochemical identification of amyloid, using an anti-human serum amyloid P component (SAP) antibody, is possible in ruminants but not in dogs and cats.

Amyloidosis represents a heterogenous group of diseases that have in common the deposition of fibrils composed of proteins of beta-pleated sheet structure, a structure which can be specifically identified by histochemistry using the Congo red or similar stains. Amyloid consists primarily of the amyloid fibrils but also of the amyloid P component (AP). This component, which is identical with the serum counterpart (SAP), is found in all types of human amyloid, and immunohistochemical identification of AP has been proposed as an adjunct to the universal, type-independent diagnosis of human amyloidosis. In the present study of animal amyloidosis, we compared the amyloid-specific Congo red stain with an immunohistochemical protocol using an anti-human SAP antibody for the identification of amyloid in formalin fixed tissue samples. The species and types of amyloidoses investigated were: (i) seven cows, one yak (Bos grunniens), and one sheep affected with amyloidosis of presumed AA type, (ii) one dog with a pancreatic endocrine tumour producing amyloid of presumed AIAPP type, (iii) two cats with presumed AIAPP-amyloidosis of the islets of Langerhans, one cat with presumed AA-amyloidosis, and one cat with an amyloid-producing odontogenic tumour. Intense immunostaining co-localized with amyloid, identified by its congophilia and green birefringence, using a protocol without any antigen retrieval in each of the seven cows, the yak and the sheep. The method seemed more sensitive in the ruminants than the Congo red stain, but was unable to detect amyloid in the dog and the cats regardless of the application of various antigen retrieval protocols. However, specific identification of amyloid still rests on the Congo red method or similar histochemical techniques.

Purification, characterization and immunolocalization of porcine surfactant protein D.

Surfactant protein D (SP-D) is a collectin believed to play an important role in innate immunity. SP-D is characterized by having a collagen-like domain and a carbohydrate recognition domain (CRD), which has a specific Ca(2+)-dependent specificity for saccharides and thus the ability to bind complex glycoconjugates on micro-organisms. This paper describes the tissue immunolocalization of porcine SP-D (pSP-D) in normal slaughter pigs using a monoclonal antibody raised against purified pSP-D. Porcine SP-D was purified from porcine bronchoalveolar lavage (BAL) by maltose-agarose and immunoglobulin M affinity chromatography. The purified protein appeared on sodium dodecyl sulphate-polyacrylamide gel electrophoresis as a band of approximately 53,000 MW in the reduced state and approximately 138,000 MW in the unreduced state. Porcine SP-D was sensitive to collagenase digestion and N-deglycosylation, which reduced the molecular mass to approximately 24,000 MW and approximately 48,000 MW respectively, in the reduced state. N-deglycosylation of the collagen-resistant fragment, reduced the molecular mass to approximately 21,000 MW showing the presence of an N-glycosylation site located in the CRD. Porcine SP-D bound to solid-phase mannan in a dose and Ca(2+)-dependent manner with a saccharide specificity similar to rat and human SP-D. The purified protein was used for the production of a monoclonal anti-pSP-D antibody. The antibody reacted specifically with pSP-D in the reduced and unreduced state when analysed by Western blotting. Immunohistochemical evaluation of normal porcine tissues showed pSP-D immunoreactivity predominantly in Clara cells and serous cells of the bronchial submucosal glands, and to a lesser extent in alveolar type II cells, epithelial cells of the intestinal glands (crypts of Lieberkuhn) in the duodenum, jejunum and ileum and serous cells of the dorsolateral lacrimal gland.

Actinomyces hyovaginalis associated with disseminated necrotic lung lesions in slaughter pigs.

Lungs from 26 slaughter pigs with a diagnosis of pyaemic lung lesions (disseminated necrotic lesions and abscesses), as determined at post-mortem meat inspection, were subjected to a thorough examination, including re-evaluation of gross pathology and histopathological and microbiological examination of samples from multiple lesions. The pulmonary lesions, which appeared identical on gross inspection, could be divided into three histopathological types, namely (1) abscesses, (2) circumscribed (contained) necrotic lesions, and (3) bronchiolar ectasias. Some characteristic relations between bacterial species and histopathological type were demonstrated. Thus, abscesses were dominated by Staphylococcus aureus infections, and circumscribed (contained) necrotic lesions were dominated by infections with an Actinomyces species, identified as Actinomyces hyovaginalis by 16S rRNA gene sequencing. Actinomyces hyovaginalis was demonstrated in 23% of all cases, pointing to this organism as an important agent of disseminated lung lesions of pigs. Furthermore, a characteristic pyogranulomatous reaction with a central area of necrosis was found to be associated with this infection.

Major histocompatibility complex-linked immune response of young chickens vaccinated with an attenuated live infectious bursal disease virus vaccine followed by an infection.

The influence of the MHC on infectious bursal disease virus (IBDV) vaccine response in chickens was investigated in three different chicken lines containing four different MHC haplotypes. Two MHC haplotypes were present in all three lines with one haplotype (B19) shared between the lines. Line 1 further contains the BW1 haplotype isolated from a Red Jungle Fowl. Line 131 further contains the B131 haplotype isolated from a meat-type chicken. Finally, Line 21 further contains the international B21 haplotype. The chickens were vaccinated with live attenuated commercial IBDV vaccine at 3 wk of age, followed by a challenge with virulent IBDV at 6 wk of age. In this study, we found a notable MHC haplotype effect on the specific antibody response against IBDV, as measured by ELISA. The BW1 haplotype was found to have a significantly higher serum antibody titer against IBDV (7,872) than haplotypes B19 (mean 5,243), B21 (5,570), and B131 (5,333) at 8 d postinfection. However, a virus-neutralizing antibody test did not reflect this result. Nevertheless, the MHC haplotype-associated protective immunity was further supported by the bursa of Fabricius (bursa) recovery from the disease, as measured by histological scorings of the bursa. Chickens carrying the BW1 haplotype had a significantly lower bursa lesion score (1.7) than the haplotypes B19 (mean 3.8), B21 (3.6), and B131 (4.3) 8 d postinfection. Furthermore, multiple line effects were found in other variables when comparing Day 6 with Day 8. Body weight, relative weights of the bursa and the spleen, percentage and relative number of MHC II molecules on MHC II-positive lymphocytes, percentage and relative number of CD4 molecules on CD4-positive lymphocytes, and the specific antibody response all differed significantly among lines. Line 1, with Red Jungle Fowl genes, was clearly differentiated from the other two investigated lines. These results suggest an MHC II restricted T-cell dependent secondary antibody response against IBDV.

Aerogenous infection of microbiologically defined minipigs with Streptococcus suis serotype 2. A new model.

Streptococcus suis serotype 2 is the cause of serious infections in animals and humans, but certain aspects of the infection pathogenesis still remain unclear. In this study an experimental model of aerogenous infection and induction of septicemia with S. suis serotype 2 was established in microbiologically defined Göttingen minipigs. Ten animals were exposed to aerosolized S. suis after previous exposure to mild acetic acid in aerosol. Six of the animals were immunosuppressed with prednisolone acetate on different days. All the animals were monitored clinically until euthanasia on days 6 to 13 after exposure. Necropsy was performed and samples were taken for microbiology, histopathology, and immunohistochemistry Three out of four animals immunosuppressed on days 5 to 7 after exposure developed S. suis septicemia, and S. suis could be detected in the tonsil of the soft palate and/or the nasal cavity of all exposed animals. Thus, using the presented model, local as well as systemic infection with S. suis serotype 2 was established in the Göttingen minipig. Since this breed is defined as free of S. suis and a range of other endemic porcine pathogens, the experimental model could prove useful in the study of this infection.

The use of serotype 1- and serotype 3-specific polymerase chain reaction for the detection of Marek's disease virus in chickens.

A serotype 1- and serotype 3-specific detection of Marek's disease virus (MDV) by polymerase chain reaction (PCR) was developed. The sensitivity of the method when applied to cell culture grown virus was comparable with that of cultivation. The method was applied to various tissue samples from chickens experimentally inoculated with serotype 1 or serotype 3 MDV.The serotype 1 strains CVI988 and RB-1B could be detected in feather follicle epithelium up to 56 and 84 days post-inoculation (p.i.), respectively, while the MDV-3 serotype was detected until 42 days p.i. The purpose of this study was to develop and evaluate a reliable and easy-to-handle method for surveillance of the occurrence of MDV in chicken flocks. We emphasize the development of a method, which can be applied to types of samples conveniently collected in the field, e.g. feather tips and blood samples. In addition, the PCR was applied to samples collected from four commercial table egg layer flocks of young stock or pullets vaccinated with either serotype 1 (CVI988) or serotype 3 (HVT) vaccine. These flocks had various clinical signs of Marek's disease. MDV-1 was detected in buffy-coat cells, spleen, liver, skin, feather tips and ovaries. The detection of MDV in feather tips appeared to be as sensitive as co-cultivation of buffy-coat cells, although an inhibiting factor was observed in extracts from feather tips of non-white chickens. This inhibition could be overcome in most extracts by applying a bovine serum albumen pretreatment. The PCR proved to be a convenient tool for the monitoring of MDV in the poultry population, and feather tips were the most convenient and sensitive samples.

Mannan-binding lectin (MBL) in chickens: molecular and functional aspects.

Mannan-binding lectin (MBL) is a serum collectin (i.e. mosaic protein with collagenous and lectin domains) involved in the innate immune defence against various microbes. In vitro studies indicate that MBL exerts its function by binding to the microbial surface through its carbohydrate recognition domains followed by direct opsonization or complement activation via the MBL associated serine proteases MASP-1 and MASP-2. In Aves (i.e. chickens), as in man, only one MBL form has been found, while traditional laboratory animals (i.e. mouse and rat) have two MBL forms in serum. MBL has been extensively studied in mammals but recently also in Aves. This review summarizes the present knowledge of MBL in chickens and compares it to the situation in mammals.

Serum levels of chicken mannan-binding lectin (MBL) during virus infections; indication that chicken MBL is an acute phase reactant.

Mannan-binding lectin (MBL) is a serum collectin which is believed to be an opsonin of the innate immune defence against various microorganisms. MBL is a minor acute phase reactant in man. We investigated the concentration of serum MBL in chickens infected with infectious bronchitis virus (IBV) and infectious laryngotracheitis virus (ILTV). The concentration of serum MBL increased about twofold (from approximately 6 to 12 microg/ml) due to these viral infections. The concentration peaked 3-7 days after infection with IBV, and 3-5 days after ILTV infection, depending on the ILTV strain used. The increased levels returned to normal values 6-10 days after infection. The results indicated that MBL is a minor acute phase reactant in chickens.