Effects of irradiation and leukoreduction on down-regulation of CXCL-8 and storage lesion in stored canine whole blood.

White blood cells (WBCs) and storage period are the main factors of transfusion reactions. In the present study, cytokine/chemokine concentrations after leukoreduction (LR) and irradiation (IR) in stored canine whole blood were measured. Red blood cell storage lesion caused by IR and LR were also compared. Blood samples from 10 healthy Beagles were divided into four groups (no treatment, LR-, IR-, and LR + IR-treated). Leukocytes were removed by filtration in the LR group and gamma radiation (25 Gy) was applied in the IR group. Immunologic factors (WBCs, interleukin-6 [IL-6], C-X-C motif chemokine ligand 8 [CXCL-8], and tumor necrosis factor-alpha) and storage lesion factors (blood pH, potassium, and hemolysis) were evaluated on storage days 0, 7, 14, 21, and 28. Compared to the treated groups, IL-6 and CXCL-8 concentrations during storage were significantly higher in the control (no treatment) group. LR did not show changes in cytokine/chemokine concentrations, and storage lesion presence was relatively mild. IR significantly increased CXCL-8 after 14 days of storage, but IR of leukoreduced blood did not increase CXCL-8 during 28 days of storage. Storage lesions such as hemolysis, increased potassium, and low pH were observed 7 days after IR and storage of blood, regardless of LR. IR of leukoreduced blood is beneficial to avoid immune reactions; however, storage lesions should be considered upon storage.

Improvement of bacterial clearance and relief of clinical signs of Salmonella enterica serovar Typhimurium infection in pigs through upregulation of Th 1-specific responses by administration of a combination of two silicate minerals, biotite and bentonite.

Biotite and bentonite are phyllosilicate minerals that were originally used in industrial applications. Several beneficial activities of them have recently been reported, especially regulation of the immune system and antimicrobial effects. Therefore, we investigated the immune-enhancing and bacterial clearance effects of a biotite and bentonite mixture (BBM) on experimental infection of Salmonella enterica serovar Typhimurium (S. Typhimurium) to determine whether the BBM could be used as an alternative antibiotic. We administered 1% or 2% BBM as a feed supplement. We then evaluated the bacterial clearance effects of the BBM against S. Typhimurium. We also evaluated the immune-enhancing effect of the BBM through several immunological experiments that included examination of the lysozyme activity, CD4(+)/CD8(+) T lymphocyte ratio and the T-helper type 1 (Th 1) cytokine profile. The clinical signs of S. Typhimurium and the number of viable bacteria in feces and tissues were significantly decreased in both BBM groups, especially in the 2% BBM group. The BBM also markedly enhanced the lysozyme activity, CD4(+)/CD8(+) T lymphocyte ratio and expression levels of IFN-γ and IL-12 in S. Typhimurium-challenged pigs. Therefore, the BBM could be a good candidate as an alternative antibiotic that improves Th 1-specific immune responses and the bacterial clearance effect.

Seroprevalence of Toxoplasma gondii and Neospora caninum in dogs from Korea.

Toxoplasma gondii and Neospora caninum are closely related protozoan parasites, they share many common hosts, and can cause neurological diseases in dogs. Dogs can have close contacts with humans and livestock and therefore they can act as reservoirs of these parasites. The aim of this study was to survey the seroprevalence of antibodies against T. gondii and N. caninum and their co-infection rate in dogs in Korea. In total, sera from 553 domestic dogs were collected from different breeds, sexes, and ages of dogs from nine provinces across the country of Korea during 2006 and 2007. The presence of antibodies against T. gondii and N. caninum was analyzed using the latex agglutination test (LAT) with a cut-off value of 1:32, and the indirect fluorescent antibody test (IFAT) using a serum titer of 1:100. In the total dog population, 71 (12.8%) dogs were positive for anti-T. gondii antibodies and only 20 (3.6%) were positive for anti-N. caninum antibodies. Relatively higher seropositive frequencies of antibodies against T. gondii (20.1%) and N. caninum (4.9%) were detected in the dog population from the Gyeonggi. A higher proportion of animals seropositive for anti-T. gondii antibodies was found in stray dog populations as compared to household dog populations: 18.5% (59/319) vs 5.1% (12/234), respectively. The Chi-square tests revealed significant differences in the seropositive frequencies of antibodies against T. gondii between stray and household dogs in the total population (p<0.0001), and in dogs from the Gyeonggi (p<0.01). No significant differences were observed for the presence of antibodies against T. gondii or N. caninum when compared across the sex or age (p>0.05). The first serological survey on antibodies against both T. gondii and N. caninum parasites across the entire country showed that co-infection was not common in these canine populations with a seropositive level of 0.72%. The significantly higher positive frequency of T. gondii antibodies in stray dogs in both, Gyeonggi and in the total dog populations suggests that further investigation on the seroprevalence of parasites should focus on stray dogs.

Immune-enhancing effect of fermented Maesil (Prunus mume Siebold & Zucc.) with probiotics against Bordetella bronchiseptica in mice.

Maesil (Prunus mume) has long been used as a traditional drug and healthy food in East Asian countries. It possesses a number of beneficial biological activities including potential antimicrobial effects against pathogens. Probiotics also have antibacterial effects. Moreover, some probiotics have an important role in regulating the immune system. The present study evaluated the immune enhancing effects of fermented Maesil with probiotics (Saccharomyces cerevisiae, Bacillus subtilis and Lactobacillus acidophilus) in mice, especially against Bordetella bronchiseptica, as an initial step towards the development of feed supplements for the promotion of immune activity and prevention of disease, especially in pigs. Continuous ingestion of fermented Maesil with probiotics markedly increased the macrophage ratio in peripheral blood and the T lymphocyte ratio in the spleen. In addition, antibody production against formalin-killed B. bronchiseptica significantly increased in the mice fed fermented Maesil compared with the control group. The number of leukocytes was significantly higher in the bronchio-alveolar lavage obtained from the fermented Maesil-fed animals compared to it in the control group at day 3 (maximal peak time) after experimental B. bronchiseptica infection. Moreover, at 7 day post-infection, relative messenger RNA expression levels of tumor necrosis factor- α and interferon-γ were significantly increased in splenocytes of mice fed fermented Maesil compared with those in the control group. Taken together, these findings suggest that feed containing fermented Maesil with probiotics enhances immune activity in mice, especially against B. bronchiseptica, via the potent stimulation of non-specific immune responses.

Fermented Maesil (Prunus mume) with probiotics inhibits development of atopic dermatitis-like skin lesions in NC/Nga mice.

Maesil (Prunus mume Siebold & Zucc.), a potential source of free radical scavengers and inhibitor of pro-inflammatory mediators, is used in traditional Korean medical preparations as a remedy for skin disorders as have probiotics. The action of a probiotic fermented Maesil preparation on the development of atopic dermatitis (AD)-like skin lesions was determined in a NC/Nga mouse model as an initial step towards the development of a therapeutic feed supplement for use in dogs. Continuous ingestion of the experimental feed markedly inhibited the development of the AD-like skin lesions, as evidenced by a marked decrease in skin signs and reduced inflammation within the skin lesions. Efficacy was confirmed by significant decreases in eosinophil ratio and serum IgE concentration, and a reduction in the number of Staphylococcus aureus recovered from the ear. Relative mRNA expression levels of IL-4, interferon-gamma and tumour necrosis factor-alpha in the spleens of the experimental animals were also decreased and there was an increased serum concentration of IL-10 with a concurrent decreased IL-4 concentration in comparison to a control group. Taken together, the results indicate that some component(s) of fermented Maesil have the ability to suppress the development of AD-like skin lesions, possibly by stimulation of IL-10. Beneficial effects of fermented Maesil may thus be expected in dogs with AD, although this and the nature of the active pathway remain to be explored.

Antimicrobial resistance and phage types of Salmonella isolates from healthy and diarrheic pigs in Korea.

OBJECTIVE: To investigate the serotypes, antimicrobial resistance, and phage types of Salmonella spp. isolated from healthy and diarrheic pigs in Korea. MATERIALS AND METHODS: A total of 1072 fecal samples and mesenteric lymph nodes were collected from healthy (n = 641) and diarrheic (n = 431) pigs throughout Korea during 2006-2007. Salmonella isolates were tested for antimicrobial susceptibility and phage types. RESULTS: A total of 204 Salmonella were isolated, and 3 and 9 serotypes were identified in the isolates from healthy and diarrheic pigs, respectively. Salmonella Typhimurium (38.9%), Salmonella Rissen (25.3%), and Salmonella Schwarzengrund (17.9%) were the most commonly observed serotypes in healthy pigs, while Salmonella Typhimurium (89.7%) in diarrheic pigs. A high frequency of resistance to antimicrobials, including tetracycline, sulfamethoxazole, and streptomycin, was observed in Salmonella isolates. In particular, Salmonella Typhimurium showed much higher rates of resistance than those of other serotypes, and most of Salmonella Typhimurium had resistance to more than four antimicrobials tested. Twelve definitive phage types (DTs) of Salmonella Typhimurium were identified, and the most commonly observed types were PT194 (15%) and PT203 (14%). Only 3% (4/133) of Salmonella Typhimurium were identified as DT104, and they all showed resistance to multiple drugs (> or = 4). CONCLUSION AND APPLICATIONS: This is the first report on the prevalence of Salmonella status in Korean pig production system obtained from a nationwide survey. Our results show alarming rates of resistance and multiple resistances. This information can help select appropriate drug agents for empirical therapy of Salmonella Typhimurium infection. It may also be helpful to set the guidelines on prevention and control of this pathogen in swine production system.

Inert matrix and Na 4 EDTA improve the supercritical fluid extraction efficiency of fluoroquinolones for HPLC determination in pig tissues.

A supercritical fluid extraction method combined with high-performance liquid chromatography-fluorescence detection was developed for the determination of enrofloxacin, danofloxacin, and ciprofloxacin in pig muscle, lung, and kidney samples. The optimal SFE conditions were 80 degrees C, 300 kg/cm(2), 30% methanol for 40 min as a dynamic extraction time, in addition to 0.2g Na(4)EDTA and 7.0 g sea sand in the extraction vessel. The use of Na(4)EDTA and sea sand on SFE extraction resulted in improvement of the recoveries of ciprofloxacin, a polar and hydrophilic compound, as well as enrofloxacin and danofloxacin. Overall, the recoveries ranged from 86.7 to 113.1% using the Na(4)EDTA/sea sand-assisted SFE extraction method. The Na(4)EDTA/sea sand-assisted SFE-HPLC-FLD validated method was successfully carried out in pig tissues, and proved to be specific, sensitive, reliable, and accurate. The method was also applied satisfactorily for accurate quantitative residue analysis in incurred pig tissues.

Persistence of vanA-type Enterococcus faecium in Korean livestock after ban on avoparcin.

Prevalence of vancomycin-resistant enterococci (VRE) was investigated in Korean livestock 4 years after the ban of avoparcin in feed additives. VRE were isolated from approximately 16.7% of the chicken samples (57 strains from 342 meat samples) and 1.9% of the pig samples (4 from 214 fecal samples). No VRE, however, was isolated from 110 bovine fecal samples. All the 61 VRE isolates were vanA-type Enterococcus faecium expressing a high-level resistance to vancomycin, and showed resistance to teicoplanin as well except two poultry isolates. In addition, the VRE isolates had heterogeneous pulsed-field gel electrophoresis (PFGE) patterns of SmaI-digested DNA, although identical or closely related profiles were observed among strains isolated from the same farm. Although the chicken isolates were all poultry type with G at position 8,234 of the vanX gene, the pig isolates were all swine type with T at position 8,234 of the vanX gene.

Detection and isolation of winter dysentery bovine coronavirus circulated in Korea during 2002-2004.

Although winter dysentery (WD) has been suspected to occur frequently in Korea, to date the exact epidemiology of WD has remained unknown. Therefore, we investigated the causative agents of WD by using electron microscopy, ELISA, RT-PCR, and nested PCR from 97 fecal samples of 32 WD-affected herds collected from 8 provinces during 2002-2004. The bovine coronavirus (BoCV) was consistently detected in all herds with WD. Of other pathogens, only coccidian oocyts were inconsistently but concurrently detected with BoCV. Ten isolates were identified as BoCV by immune electron microscope, immunofluorescent test and ELISA with antiserum to BoCV, and RT-PCR. From these results, it is concluded that WD caused by BoCV occurred in relatively high frequency and was widespread in Korea. The results provide important epidemiological data for the control and establishment of a surveillance system for WD in Korea.

Molecular analysis of S gene of spike glycoprotein of winter dysentery bovine coronavirus circulated in Korea during 2002-2003.

Since the molecular analysis of spike (S) glycoprotein gene of bovine coronavirus (BCoV) has been conducted and compared mainly among American and Canadian isolates and/or strains, it is unclear whether BCoV circulated in the other countries are distinctive in genetic characteristics. In the present study, we analyzed the S glycoprotein gene to characterize 10 winter dysentery (WD) coronavirus strains circulated in Korea during 2002-2003 and compared the nucleotide (nt) and deduced amino acid (aa) sequences with the other known BCoV. The phylogenetic analysis of the entire S glycoprotein gene revealed that the aa sequences of all Korean WD strains were more homologous to each other and were very closely related to respiratory bovine coronavirus (RBCV) strain OK and enteric bovine coronavirus (EBCV) strain LY-138, but were distinct from the other known BCoVs. Based on the phylogenetic analysis of the hypervariable region of the S1 subunit, all Korean WD strains clustered with the respiratory strain OK, BCQ3994 and the enteric strain LY-138, while the Canadian BCQ calf diarrhea and WD strains, and the American RBCV LSU, French EBCV F15 and avirulent VACC, L9, and Mebus strains clustered on a separate major branch. These data suggest that the WD strains circulated in Korea had a genetic property of both RBCV and EBCV and were significantly distinct from the ancestral enteric strain.

Prevalence and characteristics of Escherichia coli O157 from major food animals in Korea.

Escherichia coli O157:H7/NM (E. coli O157) is now recognized as an important cause of diarrhea, hemorrhagic colitis and hemolytic-uremic syndrome worldwide. There have been several cases of human E. coli O157 infection in Korea since it was first isolated from a patient with hemolytic-uremic syndrome in 1998. Meat, other foods, and recreational and drinking water contaminated with animal feces are probably the major sources of the E. coli O157 infection. In this study, we investigated the prevalence of E. coli O157 in fecal and meat samples of cattle, pigs and chicken in Korea from April 2000 to July 2002. Eighty-six (3.03%) of 2843 samples were positive for E. coli O157. Most of the E. coli O157 strains were isolated from fecal samples of beef and dairy cattle from May to October of each year. Of 86 E. coli O157 isolates, 73 were serotype O157:H7 and 13 were serotype O157:NM. Polymerase chain reaction (PCR) analysis of E. coli O157 virulence markers revealed that all O157:H7/NM isolates were positive for EhlyA, eaeA and rfb(O157), and 77 isolates were positive for stx1 and/or stx2. Cytotoxicity analysis revealed that many of the E. coli O157 isolates showed high cytotoxicity on Vero cells. Our data suggest that the majority of Korean E. coli O157 isolates from food animals can cause serious diseases in humans.

Molecular typing of enterotoxigenic Staphylococcus aureus isolated from bovine mastitis in Korea.

One hundred and sixty-six Staphylococcus aureus isolates from mastitic milk samples from different cows on 26 farms were investigated for staphylococcal enterotoxins(SEs) and toxic shock syndrome toxin-1(TSST-1) by polymerase chain reaction(PCR) and reverse passive latex agglutination assay(RPLA). SEs and the TSST-1 gene were detected in thirty-seven isolates based on a multiplex PCR; SEA was detected in 32 isolates, SEB in 3 isolates, SEC in 1 isolate, and SEA and the TSST-1 gene in 1 isolate. Of the 37 enterotoxigenic isolates, thirty-three isolates were enterotoxigenic according to RPLA, where 29 isolates produced SEA, 3 isolates produced SEB, and 1 isolate produced SEC. The enterotoxin-producing S. aureus isolates were further characterized by pulsed-field gel electrophoresis(PFGE). A macrorestriction analysis revealed 11 PFGE patterns. Among the 33 enterotoxigenic S. aureus isolates, 45.4% exhibited the same PFGE pattern I. Accordingly, although the enterotoxin-producing S. aureus isolates from bovine mastitis were genetically diverse, 1 common genotype prevailed on the farms, indicating that PFGE pattern I isolates may be the most disseminated in Korea.