Medicinal herb extracts ameliorate impaired growth performance and intestinal lesion of newborn piglets challenged with the virulent porcine epidemic diarrhea virus
© Kim et al. 2015
Received: 26 May 2015
Accepted: 28 August 2015
Published: 8 October 2015
The objective of this study was to evaluate effects of a combined use of extracts of medicinal herbs Taraxaumi mongolicum, Viola yedoensis Makino, Rhizoma coptidis, and Radix isatidis (MYCI) on porcine epidemic diarrhea (PED). Twenty-two 3-day-old piglets received an oral challenge with 3 × 103.5 TCID50 of the virulent PED virus (PEDV) in PBS or PBS only and daily oral administration of 60 mg of the MYCI mixture suspended in milk replacer or the vehicle for 7 days in a 2 × 2 factorial arrangement of treatments. Average daily gain (ADG) increased (p < 0.05) in response to the MYCI treatment in the PEDV-challenged piglets (−18 vs. 7 g for the vehicle- vs. MYCI-administered group), but not in unchallenged animals (27 vs. 28 g). Diarrhea score and fecal PEDV shedding, however, were not influenced by the MYCI treatment. The PEDV challenge caused severe intestinal villus atrophy and crypt hyperplasia, both of which were alleviated by administration of the MYCI mixture as indicated by an increase in the villus height and a decrease in the crypt depth due to the treatment. Overall, medicinal herb extracts used in this study ameliorated impaired growth performance and intestinal lesion of newborn piglets challenged with the virulent PEDV. Therefore, our results suggest that the MYCI mixture could be used as a prophylactic or therapeutic agent against PED.
The porcine epidemic diarrhea virus (PEDV) or genus Alphacoronavirus causes an acute or sub-acute gastrointestinal disorder in pigs of all ages resulting in serious economic losses across Europe and Asia [1–3]. Primary clinical symptoms of the PEDV infection include acute diarrhea, vomiting, weight loss, and dehydration [4, 5]. Moreover, an increased mortality as well as higher morbidity in suckling piglets vs. older ones has also been reported as to the damage of PEDV infection . Many attempts have been made to find effective prophylactic or therapeutic agents against PEDV infection, but no satisfactory agent has been reported to date [7, 8].
Medicinal herbs have been used as antimicrobial agents for many years in Asian countries [9–12]. For example, extracts of Houttuynia cordata are known to be effective against herpes simplex virus type 1 (HSV-1), influenza virus, human immunodeficiency virus type 1 (HIV-1) , and coronavirus . Moreover, Taraxacum mongolicum, Viola yedoensis Makino, Rhizoma coptidis, and Radix isatidis, which have been selected for the present study, are also known to have antimicrobial and antiviral properties . Taraxacum mongolicum has anti-inflammatory, anti-carcinogenic, anti-allergic and anti-viral properties [16–18] whereas extracts of Viola yedoensis Makino have antiviral and antibacterial activities against HSV-1 and Bacillus subtilis [19, 20]. Rhizoma coptidis, which contains berberine as one of its active components, has a potent antiviral activity against herpes virus as well as an antibacterial activity [21, 22]. Radix isatidis, roots and leaves of which have been used in traditional Eastern medicine for centuries to treat upper respiratory infections and encephalitis, has been reported to have anti-inflammatory  and antiviral effects against hepatitis, severe acute respiratory syndrome (SARS), influenza, and epidemic encephalitis  as well as a stimulatory action on macrophage phagocytosis .
Taraxacum mongolicum, Radix isatidis, Viola yedoensis Makino, Rhizoma coptidis, and Radix isatidis are potentially useful medicinal herbs for PED as could be extrapolated from their antimicrobial activities described above. To our knowledge, however, such a possibility has not been examined to date. The present study was therefore conducted to investigate the effects of a combined mixture of extracts of the four herbs (MYCI) on growth performance, diarrhea, and intestinal morphology in newborn piglets challenged with PEDV.
Animals and treatments
Calculated chemical composition of the milk replacer
Crude protein (%)
Crede fat (%)
Crude fiber (%)
Crude ash (%)
Vitamin A (U/kg)
Detection of fecal shedding of PEDV
RNA contained in the feces was extracted using RNasy Mini Kit® (QIAGN, Germany) after removal of contaminants by centrifugation for 10 min at 4000 × g. Presence of PEDV in the sample was identified by the reverse transcription-polymerase chain reaction (RT-PCR) targeting a coding region of the membrane protein of the PEDV . The base sequences of the forward and reverse primers were 5’-GGGCGCCTGTATAGAGTTTA-3’ and 5’-AGACCACCAAGAATGTGTCC-3’, respectively. The PCR conditions were initial denaturation at 95 °C for 2 min, 35 cycles of 95 °C for 20 s, 56 °C for 40 s, and 72 °C for 1 min, and final extension at 72 °C for 3 min. A 412-bp PCR product was identified by the agarose gel electrophoresis.
The intestinal tissue was fixed in a 10 % neutral formalin solution. The fixed tissue was embedded in paraffin, sectioned to a 4-μm thickness, stained with hematoxylin and eosin, and subjected to measurement of the villus height (VH) and crypt depth (CD) under a microscopic field as previously described .
Data were analyzed using the GLM procedure of SAS (SAS Inst., USA) in all variables except for fecal PEDV shedding. The statistical model included the PEDV challenge and administration of the MYCI mixture as main effects as well as their interaction. In the frequency of fecal PEDV shedding, which was analyzed using the chi-squared test of SAS, effects of the PEDV challenge and administration of the MYCI mixture were analyzed separately.
Effects of oral administration of the MYCI mixture on growth performance of newborn piglets after an oral administration of PEDVa)
CONb) (n = 5)
MYCI (n = 5)
CON (n = 6)
MYCI (n = 6)
C × M
Initial wt, kg
Final wt, kg
Diarrhea and detection of fecal shedding of PEDV
Effects of oral administration of the MYCI mixture on fecal PEDV shedding of newborn piglets after an oral administration of the virusa)
Control (n = 5)
MYCI (n = 5)
Control (n = 6)
MYCI (n = 6)
Effects of oral administration of the MYCI mixture on intestinal mucosal morphology of newborn piglets after an oral administration of PEDVa)
CONb) (n = 5)
MYCI (n = 5)
CON (n = 6)
MYCI (n = 6)
C × M
While only sporadic PED outbreaks were reported in Europe during the past few decades, PEDV infection has caused enormous economic losses in the pig industry in Asian countries including China, South Korea and Vietnam during the same period. Furthermore, swine farmers in North America have also been experiencing devastating economic losses resulting from prevalence of PED since its outbreak in April of 2013 in the USA . Because the efficacy of PEDV vaccines is not good enough to prevent PEDV infection, it has been a research focus to search for safe, inexpensive and effective measures to prevent or treat PED. Accordingly, a variety of studies including the present one have been conducted in Europe, Asia, and America to find antiviral agents in medicinal herbs as a means of alternative medicine [6, 28].
Diarrhea and vomiting are the main clinical signs of PEDV infection resulting in suboptimal growth performance in suckling piglets . The decreased ADG as well as diarrhea of the newborn piglets following the PEDV challenge in the present study was thus consistent with the known clinical signs of the virus infection [29, 30]. More importantly, administration of the MYCI mixture alleviated the growth check of the piglets due to the PEDV infection.
The PEDV infection causes gross and microscopic lesions such as severe atrophic enteritis with villous fusion similar to those found in the infection of the transmissible gastroenteritis virus [29, 31]. The atrophied villus structure as well as crypt hyperplasia indicated by the decreased VH and increased CD observed in the PEDV-challenged piglets was therefore consistent with the known histopathological signs of the PEDV infection. Furthermore, the increase in VH in the duodenum, jejunum, and ileum as well as the decrease in CD in the jejunum and ileum in response to administration of the MYCI mixture indicates that the MYCI recipe has a beneficial effect on the integrity of the intestinal mucosal structure of the piglets infected with PEDV.
The ultimate goal of PEDV vaccination is to prevent PEDV infection. However, currently available PEDV vaccines are not effective enough to prevent PED, although some of the vaccines are effective for alleviating fecal virus shedding and the severity of the clinical signs of PED . In this regard, administration of the MYCI mixture was not effective for reducing the frequency of fecal PEDV shedding in the challenged piglets with all its positive effects on the integrity of the intestinal mucosal structure. Nevertheless, it is thought to be worth trying to examine if a combined use of a PEDV vaccine and the MYCI mixture has any synergistic prophylactic or therapeutic effect in PED.
A number of phytochemicals contained in medicinal plants, which include limonoids, alkaloids, lignana, organosulfur, furyl, thiophenes, polylines, terpenoids, flavonoids, polyphenolics, sulphides, saponins, coumarins, and chlorophyllins, are known to exhibit the antiviral effects by blocking viral entry or RNA/DNA replication or by virtue of their anti-oxidant activity . Likewise, active antiviral phytochemicals, i.e. 11β,13-dihydrotaraxinic acid, 6,7-dimethoxycoumarin, berberine, and indirubin, have been identified in Taraxaumi mongolicum , 6,7- Viola yedoensis Makino , Rhizoma coptidis , and Radix isatidis , respectively, extracts of which were used in the present study. It also seems possible that there exist more active phytochemicals other than those described above in those herb extracts used in the present study, because the list of active components contained in medicinal herb extracts is rapidly expanding . Overall, the present results suggest that administration of the MYCI mixture is a promising recipe which could be used as a prophylactic or therapeutic measure for PED.
We evaluated effects of a combined use of extracts of medicinal herbs Taraxaumi mongolicum, Viola yedoensis Makino, Rhizoma coptidis, and Radix isatidis (MYCI) on PED. In summary, medicinal herb extracts used in this study ameliorated impaired growth performance and intestinal lesion of newborn piglets challenged with the virulent PEDV. Therefore, our results suggest that the MYCI mixture could be used as a prophylactic or therapeutic agent in PED.
The present study was supported by 2014 Research Grant from Kangwon National University (No. 120141471) and the Regional Animal Industry Center at Gyeongnam National University of Science and Technology. The authors thank Jin Song (School of Medicine, Boston University) for critical reading of the manuscript.
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