Previous studies have shown that proline mutations in the heptad repeat region stabilize the coronavirus spike (S) protein in a pre-fusion state. To understand the impact of proline substitutions on the fusogenicity of the S protein, we engineered the swine acute diarrhea syndrome coronavirus (SADS-CoV) S protein with two proline substitutions (S-PP) and examined its fusogenicity using dual-split-protein based cell-cell fusion assay. Unlike the wild-type S (S-WT), S-PP rarely formed syncytia. Additionally, protein expression of S-PP was impaired compared to S-WT, as previously reported. Our results indicate that pre-fusion stabilized S protein is unable to induce membrane fusion and provide a better understanding of SADS-CoV S and vaccine antigen design.

This study aimed to investigate the effects of probiotic treatment with IgY and Bacillus subtilis complex on diarrhea and weight gain in Hanwoo calves. Bacillus and E. coli levels were measured in the feces of calves post probiotic administration. Subsequently, calves with diarrhea were divided into two groups. The total and daily weight gain was measured in 23 calves after three months of suckling and divided into three groups. In Test 1, Bacillus was detected of the control group constantly at 104 CFU/g; a statistically significant difference was observed between the treatment and the control groups on the 1st and 3rd day (p<0.05) but not on the 7th day, confirming that the Bacillus subtilis survived in the intestine only until the 7th day. The treatment and control groups in Test 2 did not show any significant difference in the E. coli levels. However, comparison of changes in the E. coli levels showed a decreased of 6.39×108±9.80×108 CFU/g in the control group increase by 1.54×109±2.16×109 CFU/g, and, a decrease by 1.47×109±2.20×109 CFU/g; a decrease of by 4.20×107±3.90×108 CFU/g was observed in the treated group, an increase by 7.19×108±2.04×109 CFU/g, anda a decrease by 9.22×108±1.79×109 CFU/g. The changes in, control group were more rapid than in the treatment group. Therefore we hypothesize that probiotics inhibit E. coli growth. A concurrent improvement in weight gain was observed following probiotic administration. The IgY and Bacillus subtilis probiotics complex used in this study can be effectively used for the prevention and treatment of calf diarrhea.

Porcine epidemic diarrhea virus (PEDV) is a porcine coronavirus that causes enteric diseases characterized by watery diarrhea and dehydration in suckling piglets. Concentrated and highly purified viruses are required for the preparation of vaccines, diagnostics, and virus research. Currently, most protocols for virus purification require ultracentrifugation, which can be an instrumental barrier to routine operations in a laboratory. In this study, the efficacy of low-speed centrifugation for virus concentration was examined. The SM98 strain of PEDV was propagated in Vero cells and pelleted by centrifugation for 3 h at high speed (100,000 × g) or for 18 h at low speed (10,000 × g). The efficacy of virus concentration was analyzed by virus titration and western blotting. The amounts of infectious viruses and viral proteins in the pelleted samples obtained by low-speed centrifugation were comparable to those obtained by high-speed centrifugation. Interestingly, the pelleted sample impurity level was lower in low-speed than in high-speed centrifugation. In summary, we describe an efficient, easy-to-perform protocol for the preparation of purified and concentrated PEDV.

Porcine epidemic diarrhea virus (PEDV) infects all-age pigs and causes enteric diseases. Genetic diversities in isolates been reported from each country, and those diversities highlighted in pathogenicity and vaccine. In this manuscript, we are reporting of new PEDV isolation in Korea, and with genetic characteristics. Our new isolate belongs to G2b and put the name as CNUP6-2018.

Various viral and bacterial pathogens interact with environmental factors to cause diarrhea in piglets. Enterococcus spp. are Gram-positive anaerobic bacteria that are commonly found in the gastrointestinal tract of several animal species, including pigs. Enterococcus spp. have been reported to infect several animal species as a pathogen. However, gastrointestinal infection by Enterococcus hirae is rare in pigs; only a few cases have been reported worldwide. Four piglets with diarrhea were examined in the diagnostic laboratory of Optipharm Inc. (Cheongju, Korea). During the initial post-mortem examination, no disease lesions were observed. Upon microscopic examination, we found numerous Gram-positive cocci that were adhered to epithelial villi in the jejunum and ileum. However, the villi did not exhibit significant structural damage. Cultured bacteria were identified as E. hirae using the VITEK 2 system and polymerase chain reaction (PCR). Using PCR, we also confirmed that viruses and protozoa that can potentially infect piglet intestines were absent. In antibiotic susceptibility test, the bacteria were resistant to most types of antibiotics. This study presents rare cases of E. hirae infection of the piglet small intestine, which can occur in association with diarrhea possibly by the continuous use of antibiotics.

Viral particles of Porcine epidemic diarrhea virus (PEDV) consist of a four structural proteins. Among them Spikeprotein mediated responsible for receptor binding and membrane fusion during viral infection and therefore the main targetof neutralizing antibodies. Virus-like particles (VLPs) are consisted of one or more viral structural proteins, and theirmorphologies closely resemble those of the native virus. VLPs have no virulence and can elicit robust immune responsesas compared with inactivated or live-attenuated virus vaccines. Thus, in this study, we tried two methods for VLP constructionin Bombyx mori, one is traditional method and the other is chimeric VLP method using the influenza matrix protein.Both methods could produce successfully PEDV VLPs.

이유자돈 시기에 자돈의 성장성적을 개선하기 위하여 자돈의 사료섭취량을 극대화하는 것은 중요 하다. 본 연구는 사료섭취량을 증대시키기 위한 방법으로써의 향미제의 사용이 이유자돈의 성장성적 및 혈액성상에 미치는 영향을 규명하기 위해 수행되었다. 평균 체중 6.43±0.955kg의 3원 교잡종 ([Yorkshire×Landrace])×Duroc) 160두를 공시하였으며, 4처리 5반복 펜당 8마리씩 성별과 체중에 따 라 난괴법으로 배치하였다. 처리구는 1)향미제를 첨가하지 않은 처리구, 2)밀크-바닐라향 0.3%를 첨가 한 사료 3)버터향 0.3%를 첨가한 사료 4)어분향 0.3%를 첨가한 사료이다. 실험결과 성장성적, 혈액성 상 및 설사지수에서 처리구간 유의적인 차이를 발견하지 못하였다. 따라서 향미제의 종류에 상관없이 향미제를 0.3% 첨가한 사료는 그렇지 않은 사료에 비해 이유자돈의 성장성적이나 혈액성상에 있어서 아무런 긍정적인 영향을 끼치지 않을 것으로 판단된다.

Bovine viral diarrhea virus (BVDV) is a major pathogen that may be one of the main reasons for economic losses in the livestock industry. BVDV is a well-characterized member of Flaviviridae family with plus-stranded RNA viruses. Non-structural NS5B protein is RNA-dependent RNA polymerase, which is responsible for viral RNA synthesis and genome replication of BVDV. Therefore, the NS5B polymerase is a key target for the discovery of anti-BVDV drugs. A number of small-molecule inhibitors against the NS5B polymerase have been reported in literature of which we collected series molecules having various scaffold with their biological data determined by evident experimental conditions, methods and procedures. Then, we constructed database of 655 small-molecule NS5B inhibitors having definitive activity values, structural parameters, and physicochemical properties (such as molecular hydrophobicity, hydrophilicity, polarity, Hbond donors and H-bond acceptors) associated with their absorption and permeability through a cheminformatics approach. The database was opened to provide insight for allosteric NS5B inhibitors of BVDV with an accessible platform on the web (http://nabic.rda.go.kr/chemical genomic database/BVDV RNA dependent RNA polymerase inhibitors). This molecular information in the database would be useful in attempting to identify features and decision factors that enhance anti-BVDV activity or increase selectivity of the allosteric inhibitor. These anti-BVDV molecules could also be screening for the purpose of exploiting potent NS5B inhibitors in the same family (e.g., HCV, CSFV, YFV, WNV, and DENV).

The coronavirus porcine epidemic diarrhea virus (PEDV) infects the cells lining the small intestine of a pig and, causes porcine epidemic diarrhea (PED). Owing to its highly infectious nature, PEDV has a substantial economic burden, which results in significant morbidity and mortality in piglets. In this study, the virucidal efficacy of a powder disinfectant containing a phosphate compound against PEDV was investigated. Virucidal efficacy was assessed as the infectivity of PEDV toward Vero cells after exposure of the virus to the disinfectant. PEDV was exposed to the disinfectant in the presence of either hard water (HW) or an organic matter suspension (OM). In the HW condition, PEDV was inactivated by 4-fold dilution of the disinfectant. In the presence of OM, the disinfectant showed virucidal activity with a 2-fold dilution. As the disinfectant possessed virucidal activity against PEDV, it should be an effective reagent for limiting the spread of animal viral diseases.

본 연구는 이유 후 돈군이동지연이 자돈의 성장, 혈액성상에 미치는 영향을 규명하기 위해 수행되었다. 평균 체중 6.77 ± 1.395kg의 3원 교잡종([Yorkshire × Landrace]) × Duroc) 180두를 공시하였으며, 3처리 6반복 펜 당 10마리씩 성별과 체중에 따라 난괴법으로 배치하였다. 처리구는 1) D0: 이유 직후 분만사에서 자돈사로 이동한 처리구, 2) D3: 이유 후 분만사에서 3일 대기한 후 자돈사로 이동한 처리구 3) D5: 이유 후 분만사에서 5일 대기한 후 자돈사로 이동한 처리구이다. 실험결과 성장성적과 설사지수에서 처리구간 유의적인 차이를 발견하지 못하였다. 혈액성상에서는 이유 직전의 BUN (blood urea nitrogen) 수치가 분만틀에서 대기하는 기간이 늘어남에 따라 감소하였으며, 이유 후 cortisol 농도에서는 대기하는 기간이 늘어남에 따라 증가하였다. 하지만 면역성상에서는 IgG와 IgA 모두 다 유의적인 차이를 보이지 않았다. 따라서 이유직후에 돈군이동(D0)을 하는 것이 분만틀에서 대기 하는 것보다 자돈의 체내 질소의 이용률이나 이유 후의 동물들의 복지 측면에서 농가에게 더 효율적인 사양시스템인 것으로 사료된다.

Porcine epidemic diarrhea virus (PEDV) causes acute diarrhea and dehydration in pigs with high mortality. PEDV is belong to Coronavirus, enveloped, single-stranded, positive-sense RNA virus. PEDV particles were composed of four structure proteins such as a glycosylated peplomer (spike, S) protein, envelope (E), glycosylated membrane (M) protein, and unglycosylated RNA-binding nucleocapsid (N) protein. Many of previous studies talk about this four structure proteins have a great potential to diagnosis and prevent PEDV. In this study we investigated expression of these structure proteins using the bacterial and baculovirus expression system. In bacterial expression system, our results showed that structure proteins fused polyhedrin and intein gene were expressed higher than non-fusion structure proteins. The expressed fusion proteins were used to immune mice for generating a polyclonal antibodies. In baculovirus expression system, co-infection of insect cells with these four recombinant baculoviruses led to self-assembly of virus-like particles as demonstrated by Transmission electron microscopy. They were confirmed by western blot analysis using pre-made polyclonal antibodies. Finding in this study may provide important information for vaccine and diagnostic development.

Porcine epidemic diarrhea virus (PEDV) causes porcine epidemic diarrhea (PED) and a considerable economic loss in the swine industry. In this study, the virucidal efficacy of a disinfectant composed of citric acid, benzalkonium chloride and phosphoric acid against PEDV was investigated. Virucidal efficacy was assessed as the infectivity of PEDV toward Vero cells after exposure of the virus to the disinfectant. PEDV was exposed to the disinfectant in the presence of either hard water (HW) or an organic matter suspension (OM). In HW condition, PEDV was inactivated by 600-fold dilutions of the disinfectant. In the presence of OM, the disinfectant showed virucidal activity after a 200-fold dilution. As the disinfectant possesses virucidal activity against PEDV, it should be an effective reagent to use to limit the spread of animal viral diseases.

The objectives of the current study were to evaluate the therapeutic effect of dioctahedral smectite (smectite) against calf diarrhea caused by pathogenic E. coli and/or Salmonella typhimurium. In this study, 20 calves (aged 2~3 months) with diarrhea were used for evaluation of the efficacy of smectite on calf diarrhea with 20% smectite suspension in PBS. Calves received 10 ml smectite suspension three times per day after feeding, and fecal samples were collected at the gate of treatment and on the first, second, third, fourth, and fifth day after administration. On the fifth day after treatment with smectite suspension, the diarrhea index showed a significant decrease in the treated group, compared to the control group (P<0.001). The number of pathogenic E. coli in feces of the treated group was significantly decreased, compared to each control group from the second day after treatment (P<0.001), and that of Salmonella typhimurium was significantly decreased from the first day after treatment (P<0.05). According to the results of the current study, 20% smectite suspension had a therapeutic effect on diarrhea caused by E. coli and/or Salmonella in calves.

Hemorrhagic Escherichia coli (E. coli) is one of the most important agent of diarrhea in piglet. The aim of this study was to investigate the characteristics of virulence genes and genetic diversity in hemorrhagic E. coli isolated from piglets with diarrhea. Among 122 hemorrhagic E. coli, 62 isolates carried single toxin gene like heat-stable toxin (ST), heatlabile toxin (LT), verotoxin 1 (VT1), verotoxin 2 (VT2), spa and eae. The most prevalent toxin gene carried by isolates was ST gene (23 isolates), while the most common association was ST/LT (14 isolates) and ST/LT/VT2 (13 isolates). In pulsed field gel electrophoresis, isolates were not classified as one cluster by epidemiological information including isolated area, isolated year and possessed toxins.

During 2008 2010, 943 swine sera were collected from 45 farms located nationwide. Antibodies against porcine epidemic diarrhea virus (PEDV) were tested via serum neutralization antibody test (SNT) using PEDV-SN, which was adapted and propagated on the Vero cell monolayer with trypsin-free culture media supplemented with more than 10% fetal bovine serum (FBS). All 45 farms were shown to have at least one or more seropositive pig. Of the 943 swine sera that were tested, 931 sera were neutralizing antibody positive against PEDV. These high seroprevalence rates seemed to be due to vaccination or natural infection of PEDV. In a plaque reduction neutralization test (PRNT) using a swine serum showing SN titer of 1:32, a greater than 50% plaque reduction was observed in up to 160 times serum dilution.

Ruminant pestiviruses of bovine viral diarrhea virus (BVDV) and border disease virus (BDV) are closely related to classical swine fever virus (CSFV) and all belong to the genus of Pestiviruses. BVDV is one of the most important viral pathogen of cattle and has been recorded in most countries where cattle are raised. Natural host for BVDV is cattle, but BVDV is able to infect pigs as well. The purpose of this study was to investigate the prevalence for antibodies against BVDV in domestic pig farms in South Korea from 2009 to 2011. In this study, 2,755 pigs in 239 farms in South Korea's inland and 5,293 pigs in 613 farms in Jeju province (CSF free region) were investigated for antibodies against two pestiviruses, BVDV and CSFV by a virus neutralization test (VNT). The seroprevalences on the individual level and on herd level against BVDV were 5.3 % and 21.2 % in South Korea's inland, 5.2 % and 6.5 % in Jeju province, respectively. Based on the ratio of respective antibody titers by the comparative VNT, 273 pigs in Jeju province with BVDV infection were detected and they were distinctly negative to CSF. It is recognized that porcine infections with BVDV naturally occurred in Jeju province. Whereas, antibody titers against BVDV of South Korea's inland were cross-reactivity with CSFV.

Although several enteric viral pathogens including the porcine groups A, B and C rotaviruses (PGARV, PGBRV, and PGCRV), sapovirus (PSaV), and torovirus (PToV) are known to cause endemic diarrheas in weaning and post-weaning piglets, their precise prevalence in Korea is not clear. Therefore, we examined 1,222 diarrhea stools obtained from 627 farms during 2004～2009 by RT-PCR and/or nested PCR for evaluating their precise prevalence in Korea. PGARV was the predominant pathogen during 2004～2007 but its prevalence was markedly reduced during 2008～2009. PGBRV infections caused endemic diarrhea during 2004～2007, but was hardly detected during 2008-2009. PGCRV was detected at 27.0%, 14.5%, 42.4%, 28.8%, 7.3%, and 54.2% during each year of 2004～2009, respectively, indicating its high prevalence in Korea throughout the years. PSaV induced with high prevalence (32.4-39.2%) during 2004～2005 but its detection rate was markedly decreased during 2006～2009. PToV caused sporadic infections only during 2006 (1.0%) and 2007 (6.9%). These enteric viruses were detected in diarrhea specimens in piglets usually in combination with each other and/or together with bacterial pathogens including the Escherichia coli, Salmonella spp., Brachispira hyodysenteriae, and Lawsonia intracellularis. Infections with PGARV, PGCRV, PSaV, and PToV were more prevalent in fecal samples collected in cold seasons. These results provide important epidemiological data for the control and establishment of a surveillance system for the prevailing enteric viruses in Korea.