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Surveillance was conducted in South Korea to look for evidence of infection with Eastern equine encephalitis virus (EEEV), Western equine encephalitis virus (WEEV), Venezuelan equine encephalitis virus (VEEV), West Nile virus (WNV), St. Louis encephalitis virus (SLEV), and Japanese encephalitis virus (JEV) in horses from 2012 to 2013. The surveillance consisted of passive surveillance of testing horses with neurologic symptoms such as paralysis, incoordination, ataxia and circling, and active surveillance of testing for serologic evidence of infection in healthy horses. Passive surveillance was conducted for EEEV, WEEV, VEEV, WNV, SLEV, and JEV, and whole blood and/or brain samples received from 49 horses with neurologic signs were tested by polymerase chain reaction (PCR). Active surveillance was conducted for WNV and JEV, and 2,695 serum samples collected from horses across the country were tested for antibodies by IgM enzyme-linked immunosorbent assay (ELISA) and virus neutralization test (VNT), respectively. All samples tested by PCR were negative for EEEV, WEEV, VEEV, WNV, and SLEV, except for one whole blood sample (1/45) that was positive for JEV. All samples tested for WNV antibodies were shown to be negative. For JEV, because South Korea is endemic and horses in South Korea are vaccinated against JEV, various titers of antibodies for JEV were detected by VNT in 57.8% (737/1,274) and 58.9% (837/1,421) of the sera in 2012 and 2013, respectively. The surveillance provides evidence that supports the view that South Korea is free from EEEV, WEEV, VEEV, WNV, and SLEV. In addition, the surveillance scheme was shown to be able to identify JEV infections in the equine population and provide serologic data for JEV that could be used to improve the current vaccination program for JEV in horses.

Methicillin-resistant Staphylococcus aureus (MRSA) infections in humans can currently only be treated with vancomycin. Consequently, vancomycin-resistant Enterococcus spp. pose a serious public health hazard because MRSA can acquire their vancomycin resistance. While the microbiological and genetic characteristics of vancomycin-resistant enterococci (VRE) have been extensively studied, serological diagnostic tools for these organisms are lacking. The VanA and VanB classes of VRE show marked resistance. Here, we identified the VanA and VanB proteins that are immunogenic in mice. To do so, mice were orally infected with a VanA strain of E. faecium or a VanB strain of E. faecalis and the serologically immunogenic proteins were identified by SDS-PAGE and Western blot analysis. The mice reacted to the 27 and 65 kDa cell envelope (CE) proteins of VanA at 1 week post-infection (wpi) and then reacted to the 100 kDa cytoplasmic protein (CP) at 2-4 wpi. With regard to VanB, the mice responded at 1-4 wpi, 3-4 wpi, and 4 wpi to the 70 kDa, 25 and 35 kDa, and 79 kDa CE proteins, respectively, and at 3 wpi to the 39 kDa CP. The identification of these immunogenic proteins may be useful for diagnosing and for producing immunotherapeutic VRE antibodies.

Between 2011 and 2012, a total of 896 pig fecal samples were collected from nine provinces in Korea, and 50 salmonella enterica susp. enterica serovar Typhimurium (S. Typhimurium) was isolated. The characteristics of the 50 strains were analyzed, and 4 strains were identified as Salmonella enterica subsp. enterica serovar 4,[5],12:i:-. Salmonella 4,[5],12:i:- could not be distinguished from S. Typhimurium through phage typing, antimicrobial resistance testing or multiple-locus variable-number tandem repeat analysis (MLVA). However, among the four Salmonella 4,[5],12:i:- strains, one (KVCC-BA1400078) was identified as a Salmonella 4,[5],12:i:- clone isolated from humans in the United States, and another (KVCC-BA1400080) was identified as DT193, which has been primarily isolated from humans and animals in European countries. The presence of Salmonella 4,[5],12:i:- in Korea poses a significant threat of horizontal transfer between pigs and humans.

This study investigated the possible presence of the Bluetongue virus (BTV) in the Republic of Korea (ROK). Cell cultures were used to test blood samples collected from abattoirs throughout the country. Testing identified a single BTV isolate, which was characterized as BTV serotype 1 based on a nucleotide sequence analysis of the segment 2 gene. This report therefore indicates that BTV serotype 1 is present in the ROK. The potential importance of BTV in the ROK has been overlooked because cattle are mostly unaffected by the virus and because sheep, the most severely infected hosts, are uncommon in the ROK. However, as recent BTV serotype 8 outbreaks in Europe have demonstrated, certain BTV strains have the potential to cause severe disease in cattle. Additionally, with climate change continuously expanding the regions in which Culicoides vectors are able to survive, there is an increased need to study BTV in the Far East and ROK. To better prepare for future outbreaks of BTV, a sustained and effective level of surveillance for BTV in livestock will need to be established.

The purpose of this study was to investigate the prevalence of Brachyspira species and antimicrobial susceptibility of Brachyspira (B.) hyodysenteriae isolates in Korea. A total of fifty-five Brachyspira species were isolated; five (1.0%) beta-hemolytic Brachyspira species and 50 (10.4%) weak hemolytic Brachyspira species from 116 different diarrheic pig samples and 367 apparently normal pig samples. In farm level, beta hemolytic and weak hemolytic Brachyspira species were detected in 7.4% (5/68) and 19.1% (13/68) of tested pig farms, respectively. By phenotypic and genotypic characterization, all beta hemolytic Brachyspira isolates was classified as group I (B. hyodysenteriae), whereas weak hemolytic Brachyspira species isolates were group III (B. innocens or B. murdochii). B. hyodysenteriae isolates showed high level of minimum inhibition concentrations to macrolide antimicrobials. This study shows that the prevalence of pathogenic B. hyodysenteriae in pigs is low but antimicrobial resistance of the pathogens is high in Korea. This is the first report of the prevalence of Brachyspira group III and antimicrobial susceptibility of B. hyodysenteriae in pigs in Korea. Our results could provide basic data for the management and treatment guidelines of Brachyspira infection.

The objective of this study was to investigate the effects of different concentrations of WSP (wheat sprout powder) and ISP (isolated soy protein) on the quality of breakfast sausage. Treatments were formulated as follows: Control, T1 (2.0% ISP), T2 (1.5% ISP + 0.5% WSP), T3 (1.0% ISP + 1.0% WSP), T4 (0.5% ISP + 1.5% WSP) and T5 (2.0% WSP). The treatments were analyzed for color, pH, cooking loss, emulsion stability, protein solubility, viscosity, texture properties and sensory evaluation. Lightness and redness were reduced and yellowness was increased as increased level of WSP, due to the dark green color of WSP (p

Agroterrorism refers to attacks with any of a variety of biological or chemical agents against commercial crops or livestock populations, either as targets in their own right or as vehicles to attack humans. An agroterrorism incident would generally involve bioterrorism, and potential agents include pathogens such as viruses, bacteria, or fungi. Within the context of agroterrorism, livestock agroterrorism is described as the intentional introduction of an animal-borne infectious disease with the goal of spreading fear, producing economic losses, and/or threatening social stability. Causing human illness or human casualties is another potential goal of livestock agroterrorism. Livestock agroterrorism is considered to be attractive to terrorists because biological agents that affect livestock or poultry are more readily available and more difficult to monitor than are agents that infect humans. In addition, a terrorist attack on animal husbandry may have huge economic consequences with no human casualties. Therefore, a biological attack that targets the animal husbandry sector should be regarded as both a "high-consequence" event and a grave national security risk. This review addresses the use of biological weapons that may be used to target livestock or poultry rather than agricultural inputs or equipment. It first defines livestock agroterrorism. Then, the common priority disease agents that may be used to target livestock or poultry in an agroterrorist attack and that are attractive to terrorists are outlined.

Attacks against livestock and poultry using biological agents constitute a subtype of agroterrorism. These attacks are defined as the intentional introduction of an animal infectious disease to strike fear in people, damage a nation's economy and/or threaten social stability. Livestock bioterrorism is considered attractive to terrorists because biological agents for use against livestock or poultry are more readily available and difficult to monitor than biological agents for use against humans. In addition, an attack on animal husbandry can have enormous economic consequences, even without human casualties. Animal husbandry is vulnerable to livestock-targeted bioterrorism because it is nearly impossible to secure all livestock animals, and compared with humans, livestock are less well-guarded targets. Furthermore, anti-livestock biological weapons are relatively easy to employ, and a significant effect can be produced with only a small amount of infectious material. The livestock sector is presently very vulnerable to bioterrorism as a result of large-scale husbandry methods and weaknesses in the systems used to detect disease outbreaks, which could aggravate the consequences of livestock-targeted bioterrorism. Thus, terrorism against livestock and poultry cannot be thought of as either a 'low-probability' or 'low-consequence' incident. This review provides an overview of methods to prevent livestock-targeted bioterrorism and respond to terrorism involving the deliberate introduction of a pathogen-targeting livestock and poultry.

In vitro activities of 13 antibiotics were assessed against 85 Brucella abortus isolates from naturally infected cattle in the Republic of Korea during 1998-2006, using broth microdilution test. Tetracyclines showed the most excellent activity against B. abortus, displaying MIC values of 0.5 μg/ml or below. In particular, minocycline showed the lowest MIC 50/90 values (0.125/0.125 μg/ml) in this study. Among four fluoroquinolones tested, ciprofloxacin (MIC 50/90 , 0.5/1 μg/ml) and norfloxacin (MIC 50/90 , 8/8 μg/ml) had the most and the least activities, respectively. Gentamicin (MIC 50/90 , 1/1 μg/ml) was more effective than streptomycin, erythromycin, rifampin, and chloramphenicol (MIC 50/90 , 2/2 μg/ml).

Bovine tuberculosis (bTB), caused primarily by Mycobacterium bovis, continues to exert an economic loss, even in countries with active control measures, and is one of zoonotic diseases enable to be transmitted to human. The control and eradication of bTB are mainly based on a test and slaughter policy and/or abattoir surveillance. Various factors including limitation of diagnostic tests have been considered as major constraints to eradication. Single intradermal test (SIT) is the official diagnostic test. New diagnostic methods are needed to be developed, because of limitations of the test. In the present study SIT was compared with single intradermal comparative cervical test (SICCT) and interferon (IFN)-γ assay. There was very low correlation between SIT and SICCT. However, high correlation was shown between SIT and IFN-γ assay while no correlation was observed between SICCT and IFN-γ assay. Therefore, our results suggest the possibility of replacement of SIT with IFN-γ assay for the diagnosis of bovine tuberculosis.

The Brucella spp. are fastidious and relatively slow-growing organisms. The isolation of such strains in a variety of specimens often requires the use of a selective medium to reduce or eliminate the growth of unexpected microorganisms. The modified Brucella selective (MBS) medium, which contains improved antibiotic mixtures, erythritol as the only carbon source, and neutral red as a pH indicator, showed good selectivity for the Brucella abortus strains, including the RB51 vaccine strain. Erythritol in the MBS medium was able to promote and/or recover the delayed growth of the B. abortus strains through the antibiotic mixtures. The Brucella colonies, which assumed a pinkish color at their central part, were easily differentiated from other organisms. The MBS medium also allows the isolation of the Brucella strains even in contaminated specimens and/or in specimens containing small numbers of viable organisms. Moreover, this medium can be applied to environmental samples for the isolation of the Brucella strains, and it can thus offer epidemiologic traceback sources for the dissemination or transfer of diseases. Therefore, the MBS medium can be applied as a useful tool of important control measures in the eradication programs.

Summary Seven of 18 elk on a deer farm were found by the official Rose-Bengal agglutination test (RBT) and tube agglutination test to be brucellosis reactors/suspects. Evaluation with the competitive ELISA (C-ELISA) and the fluorescence polarization assay (FPA) tests revealed that six and five sera were positive respectively. The seven reactors/ suspects were slaughtered and their blood and tissues were collected. Brucella species could be isolated from three of the slaughtered animals, with nine isolates being obtained from the popliteal, supramammary and submandibular lymph nodes, vaginal discharge, mammary tissue and spleen. Brucella genus-specific PCR based on 16S rRNA and AMOS-PCR, which is specific for differential Brucella species, revealed that all nine isolates were Brucella abortus. These nine were further confirmed to be B. abortus biovar 1 by classical biotyping scheme assays. This is the first report of an outbreak of brucellosis in domestic elk in Korea. Our observations suggest that deer should be included in the routine Brucella surveillance programme for the effective control and prevention of brucellosis in Korea.

Enzyme-linked immunosorbent assay (ELISA) has been developed as the ancillary diagnosis of bovine tuberculosis at ante-mortem to overcome the disadvantages of intradermal skin test. In this study, the antigenic proteins were purified, applied to bTB ELISA, and identified through proteomic analysis. Culture filtrate protein of Mycobacterium bovis was fractionated by MonoQ® column chromatography, and examined the antigenicity by immunoblotting. The antigenic 20 kDa protein was in-gel digested and identified the antigenome by LTQ® mass spectrometer and peptide match fingerprinting, which were MPB64, MPB70, MPB83, Fas, Smc, Nrp, RpoC, Transposase, LeuA, and MtbE. The 20 kDa protein exhibited the highest antigenicity to bTB positive cattle in ELISA and would be useful for bTB serological diagnosis.

A total of 744 Escherichia coli strains isolated from 830 fecal samples of healthy cattle and pigs in all provinces of Korea were examined for resistance to 16 antimicrobials. The most frequently observed resistance in cattle isolates was to tetracycline (30.5%), followed by resistance to streptomycin (20.4%), ampicillin (12.0%) and chlorampenicol (6.9%). Prevalences of resistance to the same four antimicrobials in swine isolates were 96.3%, 66.8%, 66.1%, and 47.6%, respectively. The prevalence of resistance in pigs was much higher than that in cattle, with 98.3% of pig isolates and 37.1% of cattle isolates showing resistance to one or more of the antimicrobial agents tested.

Salmonella enterica subspecies enterica serovar 4,[5],12:i:-, a monophasic variant of Salmonella Typhimurium, has emerged as one of the most common serotypes related to human salmonellosis. In this study, the 22 isolates of S. 4,[5],12:i:- from food animals were identified by a specific multiplex polymerase chain reaction between 2009 and 2012. The isolation rate of S. 4,[5],12:i:- accounted for 1.7% (22/1271) of Salmonella spp. isolates from food animal origins: more specifically, 7.6% (18/235) from pigs and 0.6% (4/686) from chickens. The predominant S. 4,[5],12:i:- isolates in Korea belonged to phage type DT193 (12/22) with ampicillin-streptomycin-sulfonamide-tetracycline (ASSuT) resistance pattern (9/22). The XbaI-pulsed-field gel electrophoresis (PFGE) analysis revealed 11 different pulsotypes, and the major X-1 pattern was shared by 8 isolates. The isolates belonging to pattern X-1 were further subdivided into three BlnI-PFGE patterns and four variable-number tandem-repeat analysis (MLVA) allele combinations. The combining of MLVA and PFGE data could be valuable in characterizing highly clonal strains and discriminating their epidemiological relationship.

Bovine tuberculin purified protein derivative (bPPD) is used as an intradermal test (IT) reagent to detect bovine tuberculosis (bTB) in most countries. Identification of bPPD proteins is critical to understanding the immunological reaction of IT at the molecular level. While bPPD from the United Kingdom (UK) and Brazil (BR) have been recently defined at the proteomic level, bPPD from the Republic of Korea (KR) has not yet been analyzed. Here, bPPD KR proteome was examined for the first time. In total, 271 proteins were identified, including Mycobacterium bovis-specific proteins Mb0854c and Mb2898, and 42 known T cell antigens. On comparing with proteomes of bPPD UK and BR, 33 proteins were found to be common among all three bPPDs, of which 15 proteins were T cell antigens. M. bovis-specific antigens with T cell activity in bPPD may be novel candidates for use as alternatives to currently available bPPD in diagnostics.

Background: Schmallenberg virus (SBV), Akabane virus (AKAV) and Aino virus (AINV) are members of the Simbu serogroup within the genus Orthobunyavirus, family Bunyaviridae, which can cause reproductive disorders including abortion, stillbirth and congenital malformation in ruminants. Because, the clinical signs are similar, confirmatory diagnosis requires viral detection to differentiate infection between these three viruses. Methods: In this study, a one-step multiplex reverse-transcriptase quantitative PCR (one-step mRT-qPCR) was developed for the simultaneous detection and differentiation of SBV, AKAV and AINV. Results: The detection limit of the one-step mRT-qPCR for SBV, AKAV and AINV were 2.4 copies (10(0.6) TCID (50)/ml), 96.2 copies (10(1.5) TCID (50)/ml) and 52.3 copies (10(1.2) TCID (50)/ml), respectively. Various field samples such as bovine serum, bovine whole blood, bovine brain, goat serum and Culicoides were analyzed using the one-step mRT-qPCR and compared with previously published RT-qPCRs. The test results of the field samples were identical for the one-step mRT-qPCR and RT-qPCRs, which showed all samples to be negative for SBV, AKAV and AINV, except for one bovine brain sample (1/123) that was positive for AKAV. Conclusion: The one-step mRT-qPCR allows for the simultaneous detection of three viral pathogens (SBV, AKAV and AINV) that cause reproductive failure.

Japanese encephalitis virus (JEV), a mosquito-borne zoonotic pathogen, is one of the major causes of viral encephalitis. To reduce the impact of Japanese encephalitis among children in the Republic of Korea (ROK), the government established a mandatory vaccination program in 1967. Through the efforts of this program only 0-7 (mean 2.1) cases of Japanese encephalitis were reported annually in the ROK during the period of 1984-2009. However, in 2010 there was an outbreak of 26 confirmed cases of Japanese encephalitis, including 7 deaths. This represented a >12-fold increase in the number of confirmed cases of Japanese encephalitis in the ROK as compared to the mean number reported over the last 26 years and a 3.7-fold increase over the highest annual number of cases during this same period (7 cases). Surveillance of adult mosquitoes was conducted during the 2010 outbreak of Japanese encephalitis in the ROK. A total of 6,328 culicine mosquitoes belonging to 12 species from 5 genera were collected at 6 survey sites from June through October 2010 and assayed by reverse-transcription polymerase chain reaction (RT-PCR) for the presence of JEV. A total of 34/371 pooled samples tested positive for JEV (29/121 Culex tritaeniorhynchus, 4/64 Cx. pipiens, and 1/26 Cx. bitaeniorhynchus) as confirmed by sequencing of the pre-membrane and envelope protein coding genes. The maximum likelihood estimates of JEV positive individuals per 1,000 culicine vectors for Cx. tritaeniorhynchus, Cx. pipiens, and Cx. bitaeniorhynchus were 11.8, 5.6, and 2.8, respectively. Sequences of the JEV pre-membrane and envelope protein coding genes amplified from the culicine mosquitoes by RT-PCR were compared with those of JEV genotypes I-V. Phylogenetic analyses support the detection of a single genotype (I) among samples collected from the ROK in 2010.

Purified protein derivative (PPD) has served as a safe and effective diagnostic reagent for 60 years and is the only broadly available material to diagnose latent tuberculosis infections. This reagent is also used as a standard control for a number of in vitro immunological assays. Nevertheless, the molecular composition and specific products that contribute to the extraordinary immunological reactivity of PPD are poorly defined. Here, a proteomic approach was applied to elucidate the gene products in the U.S. Food and Drug Administration (FDA) standard PPD-S2. Many known Mycobacterium tuberculosis T-cell antigens were detected. Of significance, four heat shock proteins (HSPs) (GroES, GroEL2, HspX, and DnaK) dominated the composition of PPD. The chaperone activities and capacity of these proteins to influence immunological responses may explain the exquisite solubility and immunological potency of PPD. Spectral counting analysis of three separate PPD reagents revealed significant quantitative variances. Gross delayed-type hypersensitivity (DTH) responses in M. tuberculosis infected guinea pigs were comparable among these PPD preparations; however, detailed histopathology of the DTH lesions exposed unique differences, which may be explained by the variability observed in the presence and abundance of early secretory system (Esx) proteins. Variability in PPD reagents may explain differences in DTH responses reported among populations.

Purified protein derivative (PPD) is a widely used reagent for the diagnosis of Mycobacterium tuberculosis infection. Recently, the molecular composition of PPD was defined, with hundreds of mycobacterial protein representatives making up PPD. Which, if any, of these specific products drive the potency of PPD remains in question. In this study, two proteins (DnaK and GroEL2) previously identified as dominant proteins in PPD were tested for the capacity to induce delayed-type hypersensitivity (DTH) responses in H37Rv-infected or BCG-vaccinated guinea pigs. These two proteins were used in pull-down assays to identify interacting PPD products. Six proteins were identified as interacting partners with DnaK and GroEL2, i.e., Rv0009, Rv0475, Rv0569, Rv0685, Rv2626c, and Rv2632c. These six proteins were tested alone and in combination with DnaK and GroEL2 for the capacity to induce a DTH response in the guinea pig model. From these studies, two cocktails, DnaK/GroEL2/Rv0009 and DnaK/GroEL2/Rv0685, were found to induce DTH responses in H37Rv-infected or BCG-vaccinated guinea pigs that were indistinguishable from DTH responses driven by a PPD injection. The mechanism by which DTH responses were induced was elucidated by histologic examination, analysis of activated CD4 + /CD8 + T cells, and cytokine mRNA expression at the site of the DTH response. PPD and the protein cocktails tested induced strong DTH responses in H37Rv-infected guinea pigs. Ex vivo phenotyping of T cells at the DTH site indicated that this response is mediated by activated CD4 + and CD8 + T cells, with increases in gamma interferon and tumor necrosis factor alpha, but not interleukin-10, at the site of the DTH response. Our results demonstrate for the first time that the PPD response can be mimicked at the molecular level with defined protein cocktails. The use of this defined product will allow a more thorough understanding of the DTH response and may provide a platform for more rapid and sensitive second-generation skin test reagents for the diagnosis of M. tuberculosis infection.

The Rose Bengal Test (RBT) is the most widely used screening test for brucellosis in both humans and animals. Owing to its apparent simplicity of reading, however, interpretations of the RBT results can be affected by personal experience. This study describes a simple way to improve the accuracy and uniformity of reading the RBT reaction by counting the number of agglutinated particles using transparent OHP film with Quantity One, which was originally designed to count the bacterial colony numbers on agar plates. Using this system, the reactivities of three Rose Bengal antigens from different sources against international standard serum (1,000 units, VLA, UK) could be numerically measured: the intensity scale ranged from zero to around 1,600. This system enabled us to compare the antigenicity of Rose Bengal antigens from three different sources by using statistical analyses such as regression and mean intensity. Collectively, mathematical measuring of the reaction intensity used in this study may help interpret subtle test results by providing more reliable data and additional statistical information on the herd. In addition, the method would also be applicable to other agglutination test for other diseases.

MPB70 (mycobacterial protein of BCG 70) as T‐cell stimulator has been tried with an intradermal skin test (IST) and enzyme‐linked immunosorbent assay (ELISA) of bovine tuberculosis (BTB). In this study, crude mycobacterial protein 70 (CMP70) was prepared by anion exchange chromatography from the culture supernatant of Mycobacterium bovis AN5 and CMP70 ELISA was compared with purified protein derivative (PPD) ELISA. PPD and CMP70 ELISA have shown a positive reaction to the sera of M. bovis infected cattle and IST positive reactors. One of three IST negative cattle showed the nonspecific reaction in PPD ELISA, whereas all of the IST negative cattle (n=3) were did not show the nonspecific reaction in CMP70 ELISA. When each ELISA was applied to sixty‐two IST positive cattle, ELISA positive reactors were eighty four per cent to CMP70 antigen and fifty‐two per cent to PPD. CMP70 has been shown to be more specific and sensitive than PPD in ELISA.