Advanced Effect of Moringa oleifera Bioconversion by Rhizopus oligosporus on the Treatment of Atopic Dermatitis: Preliminary Study.

This study was conducted to determine if topical application of Moringa oleifera extracts and its bioconversion product fermented by Rhizopus oligosporus has therapeutic properties enhancement for treatment of atopic dermatitis. Rhizopus oligosporus (KCCM 11232P) was used to ferment Moringa leaves' extracts in this study. Comparison of organic acids and flavonols in Moringa simple extracts and their fermented product by HPLC analysis revealed that concentration of organic acids and flavonols of bioconversion product was lower than that of hot water extracts. The fermentation process is used as a nutrient for isolation of each component by microorganisms and growth of microorganisms. The results demonstrated that MF extracts effectively reduced clinical features based on macrography, scratching count, and severity scores, as well as model's serum IgE level, including histopathological analyses.

Identification of VIM-2 metallo-ß-lactamase-producing Pseudomonas aeruginosa isolated from dogs with pyoderma and otitis in Korea.

BACKGROUND: Pseudomonas aeruginosa is a challenging pathogen cultured from cases of acute and chronic canine otitis and sometimes in cases of deep pyoderma. The spread of antimicrobial resistance, especially carbapenem resistance, is a serious therapeutic challenge worldwide. HYPOTHESIS/OBJECTIVES: To investigate the identification and characterization of resistant P. aeruginosa clinical canine isolates. MATERIALS: Clinical isolates (n = 80) were collected from dogs with pyoderma (n = 18) and otitis (n = 62) in Korea. METHODS: Antimicrobial susceptibility was determined using agar dilution and using Clinical and Laboratory Standards Institute guidelines for recording susceptibility for human Pseudomonas isolates; genetic relatedness of isolates was investigated by multilocus sequence typing (MLST) and SpeI macrorestriction analysis. The class 1 integrons were amplified and sequenced using primer walking. RESULTS: Most isolates were susceptible to colistin (97.5%), polymyxin B (96.3%), ciprofloxacin (81.3%) and meropenem (80.0%); whereas resistance to aztreonam (80%), piperacillin (52.5%), piperacillin/tazobactam (41.3%) and cefepime (37.5%) was high; 12 carbapenem-nonsusceptible isolates (15%) were detected. MLST revealed 45 different sequence types (STs) and macrorestriction analysis detected 55 distinct pulsotypes (PTs), which were divided into 25 clonal groups. Among carbapenem-nonsusceptible isolates, 10 (83.3%) were VIM-2-producing strains. Nine VIM-2-producing isolates were identified as ST1047 and harboured the same 2.8 kb class 1 integron. One remaining isolate was ST1203 with 2.1 kb class 1 integron. CONCLUSIONS AND CLINICAL IMPORTANCE: This study demonstrated the diversity of the phenotype and genotype of clinical P. aeruginosa isolates from dogs with pyoderma and otitis. The identification of VIM-2-producing P. aeruginosa in dogs is alarming and warrants further surveillance.

A Study on Growth Performance of Ducks Fed Diets with Different Types of Sipjeondaebo-Tang Byproduct Meal and Red Ginseng Marc with Fermented Red Koji and Ammonia Fluxes in Duck Litter using Alum or Aluminum Chloride.

The aims of the present study were to investigate the growth performance of ducks fed diets with different types of Sipjeondaebo-tang (ST) byproduct meal and red ginseng marc with fermented red koji (RGMK), and to investigate ammonia (NH3) fluxes from duck litter treated with alum or aluminum chloride (AlCl3). A total of 270 1-d-old ducks (180 males and 90 females) were allotted in a completely randomized design with 6 treatments and 3 replicates of 15 birds per pen. The six diet treatments were: basal diet, pelleted 1% ST byproduct powder, pelleted 1% RGMK, 1% blends (a mixture of ST byproduct and RGMK) powder, 1% pelleted blends, and coated pellets of 1% blends. The six litter treatments with 6 diet treatments were: no treatment, 50, 100, or 200 g alum/kg duck litter, and 100 g or 200 g AlCl3/kg duck litter (treatments T1, T2, T3, T4, and T5, respectively). During days 10 to 40, ducks fed the 5 experimental diets had significantly different (p<0.05) weight gains and feed conversion ratios compared with those fed the control diet, but initial body weight, final body weight, feed intake, and mortality were not affected. There were significant differences (p<0.05) in NH3 fluxes among treatments over the 6 weeks of the study, except for week 0. The relative NH3 losses at week 6 were lower by 25.6, 45.3, 45.6, 46.7, and 48.6% than those in the controls in T1, T2, T3, T4, and T5 respectively. In conclusion, feeding pellets or coated pellets of ST and RGMK and using alum or AlCl3 in the litter at the same time improves weight gain and feed conversion ratio performance and reduces mortality and NH3 losses in ducks.

Clonal distribution of methicillin-resistant Staphylococcus pseudintermedius isolates from skin infection of dogs in Korea.

Bacterial infection by methicillin-resistant Staphylococcus pseudintermedius (MRSP) is challenging in a small animal practice. Zoonotic transmission may occur. The aim of this study was to investigate the genotypic profiles of MRSP isolated from bacterial infections of canine skin in Korea and to compare their molecular lineages with dominant strains from other countries. Sixty MRSP isolates were obtained from the lesions of canine pyoderma and otitis externa. Their genetic diversity was assessed by multilocus sequence typing (MLST), staphylococcal protein A (spa) typing and direct-repeat unit (dru) typing. Staphylococcal cassette chromosome mec (SCCmec) elements were characterized by multiplex PCR. Thirty-nine different sequence types (STs) were detected. Among them, 21 STs were identified as internationally new sequence types. Fourteen dru types (dts) were detected, and the major types were dt11a and dt11y. spa typing characterised 21 isolates (35%, 21/60), including spa types t02 (n=8), t05 (n=5), t06 (n=6), and t15 (n=2). Two clonal complexes, CC568 and CC677, were revealed by MLST; this result differed from the dominant STs detected in MRSP isolates from Europe, North America, and other Asian countries. SCCmec type V was the major type (27/60. 45%), and 30 (50%) isolates were non-typeable by conventional classifying method. This is the first report about the clonal lineage of MRSP isolated from Korea. MRSP isolated from dogs in Korea displays independent lineage from other countries. Surveillance is needed to confirm cross-national disseminating patterns.

Sorafenib potentiates ABT-737-induced apoptosis in human oral cancer cells.

OBJECTIVE: The mimetic BH3 ABT-737, a potent inhibitor of anti-apoptotic Bcl-2 family proteins, has potential as anti-cancer drug in many cancers. Recently, patients treated with ABT-737 have developed drug tolerance during cancer therapy. Therefore, we examined whether ABT-737 is effective in killing MC-3 and HSC-3 human oral cancer cells either alone or in combination with the oncogenic kinase inhibitor, sorafenib. DESIGN: The potentiating activities of sorafenib in ABT-737-induced apoptosis were determined using trypan blue exclusion assay, DAPI staining, cell viability assay and Western blot analysis. RESULTS: Combined use of ABT-737 and sorafenib synergistically suppressed cell viability and induced apoptosis compared with either compound individually. The combination of ABT-737 and sorafenib altered only Bax and Bak proteins and their activations, resulting in mitochondrial translocation of Bax from the cytosol. Additionally, combination treatment-mediated apoptosis may be correlated with ERK and STAT3 pathways. CONCLUSIONS: These results suggest that sorafenib may effectively overcome ABT-737 resistance to apoptotic cell death, which can be a new potential chemotherapeutic strategy against human oral cancer.

Topical allogeneic platelet-rich plasma treatment for a massive cutaneous lesion induced by disseminated intravascular coagulation in a toy breed dog.

A 2-year-old intact female miniature Pinscher weighing 1.7 kg with a body condition score of 2/5 was presented for acute vomiting, lethargy for 2 days, and large petechial skin lesions on the hip region including the tail. Acute pancreatitis was diagnosed by clinical signs, strong positive cPLI test, laboratory test and ultrasound appearance. While the clinical signs associated with acute pancreatitis had improved in 3-5 days, lesion of petechial appeared on the left hip region 7 days after the presentation, with a fast progression into a necrotic tissue along the left side hip. Allogenic platelet rich plasma (PRP) with Weibrich and Kleis method was administered to promote skin healing and regeneration. Gradual and complete improvement in the dog's wound lesions was noted approximately 1 month after applying allogeneic topical PRP. In this case report, allogeneic PRP was applied to a large regional cutaneous defect caused by coagulopathy induced by acute pancreatitis. Topical application of PRP in this case was unique in that allogeneic PRP was used instead of autologous PRP for the first time in cutaneous soft-tissue wound management in the veterinary medical field.

Change of calbindin D-28k protein expression in the mice hippocampus after lipopolysaccharide treatment.

A previous study showed that 1 mg/kg lipopolysaccharide (LPS) treatment did not lead to the any neuronal death/degeneration in the mouse hippocampus. In the present study, we examined the time-dependent changes of calbindin D-28k (CB) protein expression in the mouse hippocampus after a systemic administration of 1 mg/kg LPS. CB immunoreactivity was markedly increased in pyramidal cells of the hippocampal CA1/2 regions and in granule cells of the dentate gyrus from 3 hr to 48 hr after LPS treatment. At this point in time, CB protein level was also significantly increased in the mouse hippocampus. Thereafter, CB protein expression was decreased at 96 hr after LPS treatment. These results indicate that changes of CB protein expression may be associated with no neuronal death in the model of neuroinflammation with systemic administration of 1 mg/kg LPS.

A rare case of hepatic T-cell rich B-cell lymphoma (TCRBCL) in a juvenile dog.

A 7-month-old castrated male French Bull dog was presented with vomiting, lethargy, anorexia and weight loss of 2 weeks duration. The patient's history and clinical manifestations of suspected hepatopathy were subjected to ultrasonography, radiography, biochemical investigations and cytology of hepatic lesion. The cytologic impression was hepatic lymphoma, which was later confirmed by histopathology. The neoplastic cells were strongly diffusely immunoreactive for PAX5, but not immunoreactive for CD3, and B lymphocyte specific clonal proliferation was detected using by assay of antigen receptor rearrangement. Large numbers of immunoreactive mature non-neoplastic lymphocytes were admixed with the neoplastic cell population. Therefore, the immunohistochemical results were definitively consistent with a T-cell rich B-cell lymphoma (TCRBCL). This is the first description of a hepatic TCRBCL in a juvenile dog showing a poor response to aggressive chemotherapy.

Multicentric epitheliotropic T-cell lymphoma in an African hedgehog (Atelerix albiventris).

A 2-year-old female African hedgehog was presented with a 5-month history of pruritus, and diffuse spine and hair loss. A dermatologic examination revealed erythema, excoriation, scales, and crusting affecting the face, flanks, forelimbs, hindlimbs, and dorsal and ventral abdomen. Fine-needle aspiration was performed and skin biopsies were taken from several lesions for cytologic and histologic evaluation. The aspirates yielded smears characterized by a monomorphic population of medium-sized to large lymphocytes with scant to moderate amounts of clear to moderately basophilic cytoplasm and distinct nucleoli along with a low number of cytoplasmic fragments. On histopathologic examination, there were dense dermal lymphoid infiltrates invading the dermis and a monomorphic population of round cells that had infiltrated the overlying epidermis. Epitheliotropic cutaneous lymphoma was diagnosed based on morphologic features. Additional immunochemical analysis using anti-CD3 and anti-CD79a antibodies revealed strong CD3 expression by the tumor cells, which confirmed epitheliotropic cutaneous T-cell lymphoma. This is the first description of a multicentric pattern of epitheliotropic cutaneous T-cell lymphoma in an African hedgehog.

Comparison of N-methyl-D-aspartate receptor subunit 1 and 4-hydroxynonenal in the hippocampus of natural and chemical-induced aging accelerated mice.

In this study, we compared N-methyl-D-aspartate receptor type 1 (NMDAR1) and 4-hydroxynonenal (4-HNE) in the hippocampus of D-galactose (D-gal)-induced and naturally aging models of mice. These markers represent general phenotypes in aging, and they allowed us to examine the possibility of D-gal as a chemical model agent for aging. We observed an age-dependent reduction of NMDAR1 and an increase in 4-HNE in the dentate gyrus, CA1, and CA3 regions of the hippocampus via immunohistochemistry and western blot analyses. In the D-gal-induced chemical aging model, we observed similar changes in NMDAR1 and 4-HNE although the degree of reduction/increase in NMDAR1/4-HNE was not as severe as that in the naturally aged mice. These results suggest that the D-gal-induced aging model is comparable to naturally aged mice and may be useful for studies of the aging hippocampus.

Multiorgan fungal infection caused by Microsporum canis in a green iguana (Iguana iguana).

Multiple organ invasion by keratinophilic fungi in the green iguana (Iguana iguana) has not been previously reported. In this case, a 1-yr-old female green iguana presented with a nodular, darkly discolored skin lesion surrounded by necrosis in the right ventral abdominal region. A cytologic examination of the fine needle aspiration of the lesion revealed an exuberant proliferation of fibroblasts, macrophages, and multinucleated cells along with frequent filamentous structures consistent with hyphal elements. The necropsy revealed diffuse infiltration of the liver, lung, and cardiac apex with white nodules. A histopathologic examination of the lesions also confirmed a fungal infection associated with granulomatous inflammation. Rapid polymerase chain reaction (PCR) analysis of the chitin synthase 1 gene was conducted for rapid direct detection, and inter-simple sequence repeat fingerprinting was conducted to classify the infectious origin. The PCR analysis definitively demonstrated representative Microsporum canis fungus. The present report is the first case of disseminated M. canis infection with multiorgan involvement in a green iguana.

High prevalence of blaCTX-M group genes in Aeromonas dhakensis isolated from aquaculture fish species in South Korea.

The prevalence of resistant genes against ß-lactams in 119 Aeromonas strains was determined. A large number (99.2%) of the present fish strains were resistant to one or more ß- lactams including ceftiofur, amoxicillin-clavulanic acid, ampicillin, piperacillin and cefpodoxime. Among antibiotic resistance phenotypes, the simultaneous resistance to all ß-lactams occurred in 25.2% (n=30) of all strains, which consisted of 18 strains of A. dhakensis, 8 strains of A. caviae, 2 strains of A. hydrophila and only one strain of A. veronii. For exploring genetic background of the antibiotic resistances, multiple PCR assays were subjected to detect ß-lactamase-encoding genes, bla(TEM), bla(OXA-B) and bla(CTX-M). In the results, the bla(TEM-1) gene was harbored in all strains, whereas only 3 strains harbored bla(OXA) gene. In the case of bla(CTX-M) gene, the gene was detected in 21.0% (25 out of 119) of all strains, which countered with 80% (20 out of 25) of A. dhakensis, 8% (2 out of 25) of A. caviae and 12% (3 out of 25) of A. hydrophila. In addition, most of the bla(CTX-M) positive strains showed simultaneous resistance to all ß-lactams (18 out of 30 strains). In sequence analysis for bla(CTX-M) genes detected, they were CTX-M group 1-encoding genes including bla(CTX-M-33) from 3 eel strains of A. dhakensis. Therefore, A. dhakensis obtained from cultured fish could represent a reservoir for spreading genes encoding CTX-M group 1 enzymes and hence should be carefully monitored, especially for its potential risk to public health.

Determination of the prevalence of Helicobacter heilmannii-like organisms type 2 (HHLO-2) infection in humans and dogs using non-invasive genus/species-specific PCR in Korea.

Helicobacter spp. may have multiple routes of transmission. It is unclear, however, whether the agent is zoonotic and therefore transmitted from an animal reservoir, including dogs. The aim of this population-based study was to assess the relationship between pet ownership or frequent exposure to dogs and Helicobacter spp. infection, especially focusing on HHLO-2 (Helicobacter heilmannii-like organisms type 2) in saliva and feces samples in Korea, using non-invasive genus/species-specific PCR. One hundred twenty-four eligible human subjects and 39 dogs participated in this study. Relativity of contact with dogs and Helicobacter spp. infection diagnosed by genus-specific PCR showed a statistically significant result (P<0.01), but in the relativity analyses between contact with dogs and H. pylori, H. felis and H. bizzozeronii infections diagnosed using species-specific PCR, only Helicobacter felis showed a statistically significant result. Although H. pylori infection showed a statistically significant relativity, no statistically significant association was found between veterinarian subjects and Helicobacter. spp., H. felis and H. bizzozeronii infections. On performing risk factor analyses of HHLO-2 infection by transmission, using matching species, between HHLO-2-positive dog owners and HHLO-2-positive dogs, Helicobacter felis infection showed an extremely significant relativity (P<0.0001), and Helicobacter bizzozeronii may also be a possible significant risk factor (P<0.01). These results suggest that HHLO-2 infection might be a zoonotic infection, because continuous contact with dogs was proved to be correlated with human H. felis and H. bizzozeronii infections in this study.

Mithramycin A induces apoptosis by regulating the mTOR/Mcl-1/tBid pathway in androgen-independent prostate cancer cells.

Mithramycin A (Mith) is an aureolic acid-type polyketide produced by various soil bacteria of the genus Streptomyces. Mith inhibits myeloid cell leukemia-1 (Mcl-1) to induce apoptosis in prostate cancer, but the molecular mechanism underlying this process has not been fully elucidated. The aim of this study was therefore to investigate the detailed molecular mechanism related to Mith-induced apoptosis in prostate cancer cells. Mith decreased the phosphorylation of mammalian target of rapamycin (mTOR) in both cell lines overexpressing phospho-mTOR compared to RWPE-1 human normal prostate epithelial cells. Mith significantly induced truncated Bid (tBid) and siRNA-mediated knock-down of Mcl-1 increased tBid protein levels. Moreover, Mith also inhibited the phosphorylation of mTOR on serine 2448 and Mcl-1, and increased tBid protein in prostate tumors in athymic nude mice bearing DU145 cells as xenografts. Thus, Mith acts as an effective tumor growth inhibitor in prostate cancer cells through the mTOR/Mcl-1/tBid signaling pathway.

Molecular analysis of Malassezia pachydermatis isolated from canine skin and ear in Korea.

We investigated Malassezia species and genotypes colonizing dogs, comparing those recovered from the ear canal with those from other anatomical body sites, as well as from diseased and healthy skin. The Malassezia isolates were obtained from four types of skin samples, i.e., diseased ear, diseased skin, healthy ear, and healthy skin. Sequences of the 26S ribosomal DNA region, the intergenic spacer 1 (IGS-1) and the internal transcribed spacer 1 (ITS-1) DNA region were analyzed. These confirmed the presence of Malassezia pachydermatis, which could be separated into three main sequence genotype groups (A, B, C). Genotype A was the most common, only two genotype B isolates were recovered from diseased skin lesion and genotype C was more likely to be isolated from ear samples than from other healthy or diseased-skin sites. The present findings provide the basis for further studies of genotypic diversity in M. pachydermatis, as well as their pathogenic potential.

Elevated serum levels of S100 calcium binding protein A8 (S100A8) reflect disease severity in canine atopic dermatitis.

A monoclonal antibody to canine S100 calcium binding protein A8 (S100A8) was developed to determine the association between S100A8 and the disease severity of canine atopic dermatitis. Serum S100A8 concentrations were studied in dogs with canine atopic dermatitis (n=213) and healthy dogs (n=213). Statistical correlations between these indices and atopic dermatitis activity were established, and dermatitis severity was assessed according to the CADESI score. Serum S100A8 concentrations were measured with an enzyme-linked immunosorbent assay (ELISA). S100A8 serum levels were significantly higher in canine atopic dermatitis patients than in healthy dogs. A strong positive correlation was identified between S100A8 levels and canine atopic dermatitis patients. Our findings suggested that S100A8 is actively involved in the pathogenesis and clinical picture of canine atopic dermatitis.

Survey of Helicobacter infection in domestic and feral cats in Korea.

Discovery of Helicobacter (H.) pylori has led to a fundamental change in our understanding of gastric diseases in humans. Previous studies have found various Helicobacter spp. in dogs and cats, and pets have been questioned as a zoonotic carrier. The present study surveyed the Helicobacter infections and investigated the presence of H. felis and H. pylori infections in domestic and feral cats in Korea. Sixty-four domestic cats and 101 feral cats were selected from an animal shelter. Saliva and feces were evaluated by Helicobacter genus-specific polymerase chain reaction (PCR). Genus-specific PCR positive samples were further evaluated for H. felis and H. pylori using specific primer pairs. Thirty-six of 64 (56.3%) samples from domestic cats and 92 of 101 (91.1%) samples from feral cats were PCR positive; the positive rate of feces samples was higher than that of saliva samples in both groups. H. felis and H. pylori species-specific PCR was uniformly negative. The prevalence of Helicobacter spp. in feral cats was approximately two-fold higher than that of domestic cats. The fecal-oral route may be more a common transmission route not only between cats but also in humans.

Elevated expression of bisecting N-acetylglucosaminyltransferase-III gene in a human fetal hepatocyte cell line by hepatitis B virus.

BACKGROUND AND AIM: UDP-N-acetylglucosamine: alpha-D-mannoside beta-1,4 N-acetylglucosaminyltransferase III (GnT-III) is a key enzyme in N-glycan biosynthesis. Human GnT-III enzyme activity was found to be elevated in the serum of patients with hepatomas and liver cirrhosis and in hepatocellular carcinoma tissues. Therefore, to understand the relationship between the elevation in GnT-III activity and hepatitis B viral (HBV) hepartocarcinogenesis, we investigated GnT-III gene expression in the HBV-infected cells. METHODS: A cell line, HFH-T1, producing HBV was produced by natural infection of human fetal hepatocytes. A 170-bp band corresponding to the pre-S1 region of HBV was detected in the culture medium by polymerase chain reaction. Virions were also isolated from the culture medium by sucrose density gradient centrifugation. The synthesis of both alpha-fetoprotein and albumin as an indicator that these cells were functional hepatocytes and the extent of differentiation was examined. Polymerase chain reaction and Western blot analysis using a monoclonal antibody, GT273, which was prepared using human aglycosyl recombinant GnT-III were used for HBV DNA and GnT-III detection. RESULTS: Two types of HBV-related particles were secreted into the culture medium; one was a Dane particle (40 nm in size) containing HBV DNA and the other was a subviral hepatitis B surface antigen particle (20 nm in size) that did not contain the viral genome. The secretion from the cell line was diminished by the number of passages and, thus, this cell was renamed as HFH-T2. A decreased level of the HBV was secreted from the cells after a rest period. HFH-T2 cells showed a weak staining for alpha-fetoprotein and a moderate staining for albumin in the cytoplasm around the nucleus. High levels of a 0.7 kb DNA fragment originating from GnT-III DNA were detected in HFH-T2 cells. Western blot analysis using a monoclonal antibody, GT273, which was prepared using human aglycosyl recombinant GnT-III showed a single band, corresponding to Mr 63 kDa, whereas aglycosyl GnT-III showed a band at Mr 53 kDa, with a molecular weight difference of about 10 kDa. This indicates that HFH-T2 cells express glycosylated GnT-III. GnT-III activities were 347.2 +/- 53.6 pmol/mg of protein/h in HFH-T2, 276 +/- 26.3 in Hep3B, 252.5 +/- 23.3 in HepG2 and 30.7 +/- 3.4 in NIH-3T3. GnT-III activity was higher in HFH-T2 cells than in the hepatoma cell lines, Hep3B and HepG2. CONCLUSION: A human fetal hepatocyte cell line was transformed by infection with HBV and the cell line expressed high levels of GnT-III as the levels of secretion of HBV decreased. The decrease in HBV secretion from HFH-T2 cells could be due to a high level of expression of GnT-III. Such a cell line could be used to investigate relationships between HBV infection and glycosyltransferase gene expression. Furthermore, this cell line will be useful in future studies on the effect of the expression of GnT-III on other glycosyltransferase.