Immunohistochemical study of experimental malignant catarrhal fever in rabbits

Malignant catarrhal fever (MCF) is an often-fatal lymphoproliferative disease of a variety of ungulates that occurs worldwide. It is caused by either of the highly related but distinct gammaherpesviruses alcelaphine herpesvirus-1 (AlHV-1, wildebeest reservoir) or ovine herpesvirus-2 (OvHV-2, sheep reservoir). MCF in rabbits is an excellent model as it closely resembles the disease in susceptible ungulates that include cattle, deer and bison. In this study, newly available and previously characterized monoclonal antibodies specific for rabbit leucocyte differentiation molecules were used to perform a detailed immunohistochemical examination of both AlHV-1 MCF and OvHV-2 MCF in rabbits. Differences in the MCF caused by the two viruses included: less tissue necrosis and more lymphoid cell accumulations in AlHV-1 MCF compared with OvHV-2 MCF, and in particular marked tissue necrosis in the mesenteric lymph node, appendix and liver of OvHV-2-infected animals when compared with either other tissues in OvHV-2 MCF or AlHV-1 MCF lesions in any tissue. In both AlHV-1 MCF and OvHV-2 MCF, lymphoid cell accumulations in lymphoid and non-lymphoid tissues consisted mainly of T-cells with a corresponding absence of B-cells. CD8(+) T-cells accounted for a proportion of these in the non-lymphoid tissues, but there was evidence for the accumulation of an unidentified T-cell subset/subsets as well. This study extends our understanding of the mechanisms of immuno-pathogenesis of MCF.     

Field Validation of Laboratory Tests for Clinical Diagnosis of Sheep-Associated Malignant Catarrhal Fever

Until recently, sheep-associated malignant catarrhal fever (SA-MCF) was diagnosed mainly on the basis of clinical presentation and histopathological changes. Using clinically diagnosed field cases, we have evaluated a seminested PCR and a competitive inhibition enzyme-linked immunosorbent assay (CI-ELISA) and compared these assays in the diagnosis of SA-MCF in cattle with histopathology as a provisional “gold standard.” Samples from 44 cattle with clinical signs suggestive of SA-MCF were examined by histopathology, PCR, and CI-ELISA. In addition, samples from healthy cattle were evaluated by PCR (n = 96) and CI-ELISA (n = 75). Based on histopathology, 38 of the 44 clinical cases were classified as SA-MCF positive, 3 were classified as inconclusive, and 3 were classified as SA-MCF negative. The sensitivity of PCR was 95 to 97%, whereas the specificity ranged between 94 and 100%. The CI-ELISA showed a sensitivity of 56 to 87% and a specificity between 91 and 100%. In the field, there is good correlation between the diagnoses of SA-MCF by histopathology, PCR, and CI-ELISA. These data also confirm the close association of ovine herpesvirus 2 with SA-MCF in Switzerland.Malignant catarrhal fever (MCF) is a mostly fatal, although sporadic, disease of cattle and other ruminant species which is characterized by lymphoid proliferation and is often difficult to diagnose. Clinically, the most important differential diagnoses in cattle are mucosal disease, infectious bovine rhinotracheitis, foot-and-mouth disease, and rinderpest. There are two etiologically distinct forms of MCF: (i) a wildebeest-associated form, caused by alcelaphine herpesvirus 1 (AlHV-1 [10], previously AHV-1), and (ii) a sheep-associated form (SA-MCF), occurring worldwide and implicated with the putative ovine herpesvirus 2 (OvHV-2 [10], previously OHV-2).Histopathological examination is the most widely used diagnostic procedure to confirm clinical suspicion of SA-MCF. Lymphocytic infiltration and vasculitis in the brain and other organs are the most significant lesions. In 1990, a DNA sequence with high homology to AlHV-1 was discovered in lymphoblastoid cells from SA-MCF-diseased ruminants and strongly implicated the corresponding virus, the putative OvHV-2, with the etiology of SA-MCF (2). Subsequently, Baxter et al. (1) developed a PCR protocol to demonstrate OvHV-2 DNA.Using a monoclonal antibody (3) against a cross-reacting epitope shared by AlHV-1 and OvHV-2 but not by the highly prevalent bovine herpesvirus 4 and other common viruses in cattle, Li et al. established a competitive inhibition enzyme-linked immunosorbent assay (CI-ELISA) (4) for the detection of antibodies to MCF viruses in ruminant species.A number of clinical MCF cases in cattle have been examined by these laboratory methods (5, 9). However, a comparative evaluation of these tests using a larger number of independent field cases has not yet been published. We have assessed PCR and CI-ELISA for the intravitam laboratory diagnosis of SA-MCF in comparison to classical histopathological examination.     

Evaluation of milk haptoglobin and amyloid A in high producing dairy cattle with clinical and subclinical mastitis in Shiraz

Mastitis is one of the most common diseases in dairy cattle and results in considerable loss of animals. This study was designed to evaluate milk haptoglobin (Hp) and milk amyloid A (MAA) as an inflammatory indicator for clinical and subclinical mastitis of cattle in dairy farms in Shiraz, Iran. Forty-three subclinical mastitic cows with a positive California Mastitis Test (CMT) and no clinical signs of mastitis, 28 clinical mastitic cows, and 10 healthy cows with negative CMT were selected. After confirmation of clinical and subclinical mastitis by bacterial identification, milk samples were taken from four quarters of each cow and mixed, and one sample was taken from the pooled milk. The most dominant isolated bacterium from clinical and subclinical samples was Staphylococcus aureus (n = 25; 35.2%). The most dominant isolated bacterium from clinical (19/28) and subclinical (11/43) samples was Staphylococcus spp. Of isolated bacteria of milk in cattle with clinical mastitis, 67.8% (n = 19) was S. aureus. There was no bacterial growth in 37.1% (n = 16) of cattle with subclinical mastitis. Of isolated bacteria of milk in cattle with subclinical mastitis, 13.9% (n = 6) and 11.6% (n = 5) was S. aureus and Staphylococcus epidermidis, respectively. There were significant differences (P < 0.05) in concentrations of milk Hp, MAA, and somatic cell count between clinically healthy cattle and cows with clinical and subclinical mastitis. The concentrations of milk Hp, MAA, and somatic cell count in clinical mastitic cows were significantly higher than those in subclinical mastitic cows and control group. The optimal cutoff point was set, using the receiver operating characteristic curve analysis method, to >13.43 μg/ml for MAA, >9.71 ng/ml for milk Hp, and >14 × 10⁴ cell per millilitre for somatic cell count with corresponding 100% sensitivity and 100% specificity for MAA, 83.72% sensitivity and 100% specificity for milk Hp, and 88.37% sensitivity and 100% specificity for somatic cell count. The results of this study reveal that MAA is a sensitive factor for diagnosis of subclinical mastitis in cattle.     

Investigation of sheep-associated malignant catarrhal fever virus infection in ruminants by PCR and competitive inhibition enzyme-linked immunosorbent assay.

Development of control measures for the gammaherpesviral disease of cattle known as sheep-associated malignant catarrhal fever (SA-MCF) has been hampered by a lack of accurate diagnostic tests either for the causative virus or for antibody against that virus. A recently developed competitive-inhibition enzyme-linked immunosorbent assay (CI-ELISA) for the detection of antibody to malignant catarrhal fever (MCF) virus (MCFV) in ruminants based on a monoclonal antibody to a widely conserved epitope of MCFV (H. Li, D. T. Shen, D. P. Knowles, J. R. Gorham, and T. B. Crawford, J. Clin. Microbiol. 32:1674-1679, 1994) and a PCR assay based on previously reported primers (S. I. F. Baxter, I. Pow, A. Bridgen, and H. W. Reid, Arch. Virol. 132:145-159, 1993) were used to detect anti-MCFV antibody and SA-MCFV DNA in sheep and other ruminants. The PCR amplified a specific 238-bp SA-MCFV genomic DNA fragment from peripheral blood lymphocytes of adult sheep and other ruminants with clinical MCF. Of 144 samples from randomly selected healthy adult sheep, 143 (99%) were positive by PCR and 136 (94%) were positive by CI-ELISA. The agreement between the two assays exceeded 95%. Of nine samples collected from cattle and deer with clinical MCF of apparent sheep origin, seven were CI-ELISA positive and all 9 were PCR positive. Among 59 serum samples from presuckling lambs, none contained antibody detectable by CI-ELISA. After suckling, maternal anti-MCFV antibody was detectable for about 10 +/- 3 weeks.(ABSTRACT TRUNCATED AT 250 WORDS)     

The role of cattle in the epidemiology of <Emphasis Type="Italic">Echinococcus granulosus</Emphasis> in an endemic area of southern Italy

An epidemiological and molecular survey was conducted to investigate the role of cattle in the transmission chain of cystic echinococcosis (CE) in the Campania region of southern Italy. Out of a total of 434 cattle examined for CE, 45 (10.4%) were found infected. A total of 363 cysts were collected from the infected animals: 239 in the liver and 124 in the lungs. The cysts were either sterile (42.7%) or calcified/caseous (57.3%); no fertile cysts were found. Most of the cysts had sizes <3 cm (77.1%) and were unilocular (78.8%). The results of the linear regression model did not show any significant correlation between the age of infected cattle and the number of cysts. The sequencing of the mitochondrial cytochrome C oxidase subunit 1 (CO1) gene of 40 hydatid cysts produced sequences of 419 bp for each sample analyzed. Alignment of the obtained sequences with those present in GenBank showed 100% identity with the common sheep G1 (n = 21 cysts), the Tasmanian sheep G2 (n = 2 cysts), and the buffalo G3 (n = 17 cysts) strains, which constitute the species Echinococcus granulosus sensu stricto. The findings reported in the present study show that CE is widespread in cattle bred in the Campania region of southern Italy. However, the absence of fertile cysts and of the cattle strain (G5, E. ortleppi) suggests that cattle would not have any role in the persistence of this important zoonosis but rather a role as indicators of CE infection in this endemic area.     

Feeding patterns of biting midges of the <Emphasis Type="Italic">Culicoides obsoletus</Emphasis> and <Emphasis Type="Italic">Culicoides pulicaris</Emphasis> groups on selected farms in Brandenburg,Germany

Host feeding patterns of engorged sibling species of the Culicoides obsoletus and Culicoides pulicaris groups captured during three nights on two selected farms maintaining either cattle, sheep, horses, and pigs (Seedorf, Brandenburg) or cattle, sheep, moufflons, and red and fallow deer (Paulinenaue, Brandenburg) were determined by polymerase chain reaction amplification using conserved primers and sets of species-specific primers derived from vertebrates mitochondrial cytochrome b. Out of a total of 177 blood meals analysed, 115 (65%) tested positive for a blood meal from vertebrates. 63.5% (n = 73) of the cyt b positive specimens could be further assigned down to the species level. Cattle appeared to be the most attractive hosts for Palaearctic biting midges (79.5%, n = 58) even if other large vertebrates were kept in their immediate vicinity. If pigs or horses were additionally maintained on a farm, they were likewise attacked by biting midges but at a distinctly smaller rate than cattle (pigs 13.7%, horses 2.7%). In this study, game animals appear to be less attractive than cattle since only a few engorged midges had taken a blood meal from red deer (4.1%). None of the blood meals analysed tested positive for sheep. Preliminary results reveal that biting midges of the C. pulicaris and C. obsoletus groups can feed on a range of vertebrate hosts but with a distinct preference for cattle even if other livestock are maintained in adjacent areas.     

Claudin-18 in biliary neoplasms. Its significance in the classification of intrahepatic cholangiocarcinoma

Claudin-18 (CLDN18), a tight junction protein specific to stomach and lung, is aberrantly expressed in preinvasive and invasive neoplasms of the pancreas. To investigate the significance of CLDN18 expression in biliary neoplasms, immunohistochemical analysis was performed. CLDN18 expression was frequently observed in the epithelial cells of extrahepatic bile duct carcinomas (90%, n = 99), intrahepatic intraductal papillary neoplasms of the bile duct (IPNBs, 100%, n = 11), and extrahepatic IPNBs (89%, n = 9), while it was less frequent in intrahepatic cholangiocarcinomas (ICCs, 43%, n = 83). Interestingly, CLDN18 expression was also frequently observed in precancerous lesions such as biliary intraepithelial neoplasias (78%, n = 18). Among ICCs, CLDN18-positive cases showed higher frequencies of periductal infiltrative growth, perineural invasion, and lymph node metastasis. Multivariable analysis demonstrated that positive CLDN18 expression was an independent risk factor for lymph node metastasis in ICCs. Furthermore, CLDN18 expression was associated with poor overall survival by univariable analysis, as well as lymph node metastasis. These results suggest that CLDN18 may play an important role in biliary carcinogenesis, and especially in ICCs, it is associated with aggressive behavior and serves as a useful marker for the classification of ICC.     

Production and utilization of interleukin-15 in malignant catarrhal fever

Malignant catarrhal fever (MCF) is an often fatal lymphoproliferative disease of ungulates caused by either alcelaphine herpesvirus-1 (AlHV-1) or ovine herpesvirus-2 (OvHV-2). The pathogenesis of MCF is poorly understood, but appears to involve an auto-destructive pathology whereby cytotoxic lymphocytes destroy areas of a variety of tissues. The cytokine interleukin-15 (IL-15) is involved in the development and maintenance of cytotoxic lymphocytes and may therefore have a role in the pathogenesis of MCF. Virus-infected large granular lymphocytes (LGLs) were obtained from the tissues of rabbits infected with AlHV-1 or OvHV-2. These cells exhibited a similar proliferative response to IL-15 and to IL-2 in culture, but their content of the activated cytotoxic enzyme (BLT-esterase) was maintained at higher levels in the presence of IL-15 compared with IL-2. The LGLs did not express IL-15 mRNA or produce IL-15 protein. By contrast, there was abundant expression of IL-15 mRNA and protein in affected tissues. IL-15 production was associated with necrotic lesions of the mesenteric lymph node and appendix of OvHV-2-infected rabbits, but was not found in the same tissues of rabbits infected with AlHV-1 in which there were no necrotic lesions. The cellular source of the IL-15 was predominantly lymphoid cells that did not express B cell or monocyte-macrophage markers. Only a few IL-15+ cells (<10%) co-localized with pan-T cells or CD8+ T cells. The abundance of IL-15 in tissue with lesions of MCF suggests that this cytokine may have a role in the pathogenesis of MCF.     

Serologic study of anti-Neospora caninum antibodies in household dogs and dogs living in dairy and beef cattle farms in Tehran, Iran

A few studies have been done on the seroepidemiology of anti-Neospora caninum antibodies in dairy and beef cattle farms in Iran, which suggested the presence of N. caninum in these areas, but there is no published information directed on the presence or epidemiology of this organism in the dogs in Iran. To investigate anti-N. caninum antibodies in household dogs and dogs living in cattle farms, 100 blood samples were collected: 50 from dogs living in dairy and beef cattle farms and 50 from household dogs. Serum samples were screened for detection of anti-N. caninum IgG antibodies using indirect fluorescent antibody test (IFAT; ≥50). Antibodies were seen in 10 (20%) of 50 household dogs and in 23 (46%) of 50 farm dogs. There were significant statistical differences in seropositivity between these two groups (P = 0.005). The IFAT antibody titers were as follows: 1:50 in seven dogs, 1:100 in eight dogs, 1:200 in six dogs, 1:400 in seven dogs, 1:800 in three dogs, 1:1,600 in one dog, and 1:12,800 in one dog. There were no significant differences in seropositivity between males and females. The positive results were increasing with age, and positive results were significantly different in the age group of older than 2 years compared to the dogs of age group under 1 year (P = 0.000) and 1–2 years (P = 0.007). The results confirm the exposure of household and farm dogs to N. caninum in Tehran and the higher rate of exposure for the dogs of dairy and cattle farms around Tehran.     

Downregulated parafibromin expression is a promising marker for pathogenesis, invasion, metastasis and prognosis of gastric carcinomas

Parafibromin is a protein encoded by the hyperparathyroidism 2 oncosuppressor gene and its downregulated expression is involved in pathogenesis of parathyroid carcinomas. To clarify the roles of parafibromin expression in tumourigenesis and progression of gastric carcinomas, it was examined by immunohistochemistry (IHC) on tissue microarray containing gastric carcinomas (n = 508), adenomas (n = 45) and gastritis (n = 49) with a comparison of its expression with clinicopathological parametres of carcinomas. Gastric carcinoma cell lines (MKN28, AGS, MKN45, KATO-III and HGC-27) were studied for parafibromin expression by IHC and western blot. Parafibromin expression was localised in the nucleus of gastric epithelial cells, adenoma, carcinoma cells and cell lines. Its expression was gradually decreased from gastritis to gastric carcinoma, through gastric adenomas (p < 0.05) and inversely correlated with tumour size, depth of invasion, lymphatic invasion, lymph node metastasis and Union Internationale Contre le Cancer (UICC) staging (p < 0.05) but not with sex or venous invasion (p > 0.05). Parafibromin was strongly expressed in older carcinoma patients compared with younger ones (p < 0.05). There was stronger positivity of parafibromin in intestinal-type than diffuse-type carcinomas (p < 0.05). Univariate analysis indicated cumulative survival rate of patients with positive parafibromin expression to be higher than without its expression (p < 0.05). Multivariate analysis showed that age, tumour size, depth of invasion, lymphatic invasion, lymph node metastasis, UICC staging and Lauren’s classification but not sex, venous invasion or parafibromin expression were independent prognostic factors for carcinomas(p < 0.05). Downregulated parafibromin expression possibly contributed to pathogenesis, growth, invasion and metastasis of gastric carcinomas. It was considered as a promising marker to indicate the aggressive behaviours and prognosis of gastric carcinomas.     

Serum thyroid hormones and trace element concentrations in crossbred holstein cattle naturally infected with Theileria annulata

This research was carried out to study thyroid function in view of serum trace element and possible serum cortisol concentration changes in crossbred Holstein cattle naturally infected with Theileria annulata. Twenty cattle of different age and sex which had been naturally infected with T. annulata were examined. A control group were selected among the clinically healthy crossbred Holstein cattle with the same age, sex, and production stage similar to the naturally infected group. Serum T3, T4 concentrations were significantly lower in cattle suffering from theileriosis than in the healthy controls (p < 0.05). The cattle suffering from theileriosis had significantly lower concentrations of zinc and selenium in their sera as compared with the healthy control subjects (p < 0.05). Lower packed cell volume (PCV) theileriosis-affected cattle had a lower serum selenium concentration than higher PCV-affected group (p < 0.05). Significant differences were also found in PCV, total protein, albumin and total bilirubin concentrations and alkaline phosphatase and gama-gluthamyl transferase activities between theileriosis-affected and healthy cattle (p < 0.05). Total protein and albumin concentrations were statistically different in age (less and more than two years) and PCV (lower and higher) groups of theileriosis-affected cattle (p < 0.05). No significant changes were found in free T3, free T4, cortisol, copper, cobalt, and manganese levels of theileriosis-affected cattle.     

Evaluation of the resistance of indigenous Iranian cattle to Theileria annulata compared with Holstein cattle by measurement of acute phase proteins

This study was conducted to evaluate the resistance of indigenous cattle against Theileria annulata in comparison with that of Holsteins, through assessment of changes in acute phase proteins. Blood samples were collected from 24 indigenous and 26 Holstein dairy cattle, 2–3 years old, which had become naturally infected with T. annulata. Twenty-five healthy cattle, ten indigenous and 15 Holsteins were selected as a control group. The Theileria-infected group were divided into four subgroups according to their parasitemia rates (<1%, 1–3%, 3–5% and >5%). Measurement of red blood cells (RBCs), packed cell volume (PCV), haemoglobin (Hb), haptoglobin (Hp), serum amyloid A (SAA), ceruloplasmin and fibrinogen were done for all animals using validated methods. Results showed significant differences in RBCs, PCV, Hb and concentrations of Hp, SAA, ceruloplasmin and fibrinogen between healthy cattle and those infected with T. annulata with different parasitemia rates in both breeds (P < 0.05). In both breeds, there was significant negative correlation between parasitemia and RBCs, PCV and Hb (P < 0.05). In contrast, with increasing parasitemia rate, a significant increase in MCV, Hp, SAA, ceruloplasmin and fibrinogen was evident. Iranian indigenous cattle in comparison with Holsteins showed lower parasitemia rate, milder clinical manifestations and significantly lower levels of acute phase proteins including Hp, SAA, ceruloplasmin and fibrinogen (P < 0.05).     

High resolution analysis of DNA copy-number aberrations of chromosomes 8, 13, and 20 in gastric cancers

DNA copy-number gains of chromosomes 8q, 13q, and 20q are frequently observed in gastric cancers. Moreover gain of chromosome 20q has been associated with lymph node metastasis. The aim of this study was to correlate DNA copy-number changes of individual genes on chromosomes 8q, 13q, and 20q in gastric adenocarcinomas to clinicopathological data. DNA isolated from 63 formalin-fixed and paraffin-embedded gastric adenocarcinoma tissue samples was analyzed by whole-genome microarray comparative genomic hybridization and by multiplex ligation-dependent probe amplification (MLPA), targeting 58 individual genes on chromosomes 8, 13, and 20. Using array comparative genomic hybridization, gains on 8q, 13q, and 20q were observed in 49 (77.8%), 25 (39.7%), and 49 (77.8%) gastric adenocarcinomas, respectively. Gain of chromosome 20q was significantly correlated with lymph node metastases (p = 0.05) and histological type (p = 0.02). MLPA revealed several genes to be frequently gained in DNA copy number. The oncogene c-myc on 8q was gained in 73% of the cancers, while FOXO1A and ATP7B on 13q were both gained in 28.6% of the cases. Multiple genes on chromosome 20q showed gains in more than 60% of the cancers. DNA copy-number gains of TNFRSF6B (20q13.3) and ZNF217 (20q13.2) were significantly associated with lymph node metastasis (p = 0.02) and histological type (p = 0.02), respectively. In summary, gains of chromosomes 8q, 13q, and 20q in gastric adenocarcinomas harbor DNA copy-number gains of known and putative oncogenes. ZNF217 and TNFRSF6B are associated with important clinicopathological variables, including lymph node status.     

Status of HER1 and HER2 in peritoneal,ovarian and colorectal endometriosis and ovarian endometrioid adenocarcinoma

A role for the EGF system, in particular HER1 and 2, in growth of the endometrium has been suggested but HER1 and 2 have not been studied in all locations of endometriosis and in ovarian endometrioid adenocarcinoma (OEC) which is a rare form of malignant transformation of endometriosis. Immunohistochemistry (IHC) was used for studying HER1 and HER2 in ovarian (n = 10), peritoneal (n = 10), colorectal endometriosis (n = 20) and OEC (n = 10). Fluorescent in situ hybridisation (FISH) was used for analysing the status of HER2 gene in colorectal endometriosis and OEC. All samples were negative for HER2 in both glandular and stromal cells and in glandular cells for HER1 by IHC. In 15 out of 20 colorectal endometriosis, there was a weak expression in stromal cells. Following FISH, two colorectal samples had a partial 17 aneusomy and three OEC, a 17 polysomy. The other samples were 17 disomic without HER2 amplification; HER1 and 2 do not seem to have a role in endometriosis physiopathology.     

Demonstration of sheep-associated malignant catarrhal fever virions in sheep nasal secretions

Ovine herpesvirus-2 (OvHV-2) is the causative agent for sheep-associated malignant catarrhal fever, which has never been propagated in vitro. Previous studies from this laboratory demonstrated significantly high levels of OvHV-2 DNA in sheep nasal secretions, suggesting a likely avenue of transmission. In the present study, real-time PCR was used to identify sheep experiencing an episode of intense OvHV-2 DNA shedding in their nasal secretions. A nuclease-resistance assay was used to examine the secretions for the presence of intact cell-free enveloped OvHV-2 virions. The results revealed that all nasal secretion samples from five selected individuals experiencing intensive shedding events contained cell-free OvHV-2 virions. Virions could not be identified in secretion samples from 11 OvHV-2 infected sheep that were not experiencing a shedding event. This is the first unequivocal demonstration of cell-free OvHV-2 virions. These results suggest that OvHV-2 lytic infection occurs in the epithelium of certain tissues in the upper respiratory tract of the natural host.     

Real-time PCR strategy and detection of bacterial agents of lymphadenitis

The aim of this study was to compare 16 S rRNA gene amplification and sequencing with a systematic real-time PCR assay screening strategy that includes all common known pathogens recovered from lymph node biopsy specimens. Lymph node biopsy samples sent to our laboratory from January 2007 to December 2008 were tested in the study. Lymph nodes were screened for the presence of any bacteria by PCR amplification and sequencing targeting the 16 S rRNA gene and also by a specific real-time PCR strategy that includes Bartonella henselae, mycobacteria, Francisella tularensis, and Tropheryma whipplei. By testing 491 lymph nodes, we found that the sensitivity of our specific real-time PCR assay strategy was significantly higher than 16 S rRNA PCR amplification and sequencing for the detection of Bartonella henselae (142 vs 98; p < 10⁻⁴), Francisella tularensis (16 vs 10, p < 10⁻⁴), and mycobacteria (8 versus 3, p < 10⁻⁴). None of the samples was positive for Tropheryma whipplei. Our study demonstrates the usefulness and specificity of a systematic real-time PCR strategy for molecular analysis of lymph node biopsy specimens and the higher sensitivity compared with standard 16 S rRNA gene amplification and sequencing.     

Chromosomal evolution in bovids: a comparison of cattle,sheep and goat G- and R-banded chromosomes and cytogenetic divergences among cattle,goat and river buffalo sex chromosomes

A G- and R-banding comparison of cattle (Bos taurus, 2n=60), goat (Capra hircus, 2n=60) and sheep (Ovis aries, 2n=54) chromosomes at the 450 band level was made. The study revealed a large number of banding homologies among the autosomes of the three species and resolved some ambiguities in arranging some of their small disputed acrocentrics by direct and indirect comparisons with some bovid marker chromosomes. A loss of the subcentromeric G-positive band in sheep chromosome 2q was observed when the G-banding patterns of sheep 2q and homologous cattle and goat chromosome 2 were compared. The chromosomal divergences among cattle, goat and river buffalo (Bubalus bubalis, 2n=50) sex chromosomes are shown to have occurred by pericentric and paracentric inversions with a loss (or acquisition) of constitutive heterochromatin.     

Effect of Splenectomy and Autologous Spleen Transplantation on the Serum Platelet-Activating Factor Acetylhydrolase (PAF-AH) Activity and Acute Phase Response (APR) in a Porcine Model

The aim of this study was to evaluate the inflammatory response after total splenectomy and spleen autotransplantation in a porcine model by measuring serum platelet-activating factor acetylhydrolase activity, C-reactive protein and albumin concentrations. Nineteen piglets were used in the experiment. After induction of anesthesia, animals were randomly divided into three groups: sham-operation with spleens intact (n = 6), total splenectomy (n = 6), and splenic autotransplantation (n = 7) with small fragments of the spleen autotransplanted into the greater omentum. The blood samples were taken just before surgery and on day 1st, 5th, 12th, 26th and 40th postoperatively. PAF-AH activity, CRP and albumin concentrations were assayed in the sera. After total splenectomy, PAF-AH activity was significantly increased on day 5th, while there was no significant increase after spleen autotransplantation or the sham-operation. CRP was significantly increased after surgery in all experimental groups. Albumin was significantly decreased after surgery from day 5th until day 40th in splenectomized and autotransplanted pigs. Increased PAF-AH activity after splenectomy and spleen autotransplantation might be attributed to inflammatory conditions due to the loss of splenic tissue and trauma. Time-course increase of CRP, in all groups after surgery suggests post-injury inflammatory response due to tissue lesion during operation.     

Expression of Krűppel-like factor 5 in gastric cancer and its clinical correlation in Taiwan

Krűppel-like factors (KLFs), highly conserved zinc-finger proteins, play either anti- or pro-proliferation roles in different human cancers through regulating a wide range of genes’ expression. Here, we investigated the expression of KLF5 in gastric cancers and its correlation with clinicopathological parameters and overall survival rates. In this study, KLF5 expression was measured by an immunohistochemical microarray assay of tissue taken from 76 surgical specimens. Higher KLF5 expression was significantly associated with lower tumor grade (P < 0.001). Nuclear staining of the KLF5 expression was significantly associated with a higher tumor grade (P = 0.000), higher clinical stage (P = 0.019), lymph node status (P = 0.016), and 2-year survival (P = 0.017). Patients with nuclear staining of KLF5 had a significantly lower disease-free survival rate compared to patients with negative nuclear staining, as defined by a log-rank test (P = 0.041). Our results revealed that KLF5 may play an oncogenetic role in gastric carcinogenesis.