English
Abstract
Porcine enteric coronaviruses (CoVs), including swine acute diarrhea syndrome coronavirus (SADS-CoV), porcine epidemic diarrhea virus (PEDV), porcine deltacoronavirus (PDCoV), and porcine transmissible gastroenteritis virus (TGEV), are major pathogens causing porcine viral diarrhea syndrome (VDS), which brings significant economic losses to the swine industry; distinguishing between these clinically similar viruses has become a serious challenge. We developed a highly specific and interference-resistant porcine CoV multiplex digital PCR (dPCR) assay. The assay exhibited robust anti-interference capabilities, as the concentrations of the four viruses did not affect their accurate quantification. The coefficients of variation (CV%) of intra-batch and inter-batch repeatability for all target viruses were less than 11%. The limit of quantification (LoQ) of this dPCR assay reached 7.5 copies/reaction for each target, and it was one order of magnitude more sensitive than qPCR. The limits of detection (LoD) for SADS-CoV, PEDV, PDCoV, and TGEV were 2.72, 3.00, 3.56, and 3.19 copies/reaction, respectively. A total of 408 known samples were used for validation tests, and the results were highly consistent with the known conditions, showing a compliance rate of 97–100%. The diagnostic specificity (Dsp) of the method was 99–100%. In conclusion, the developed multiplex dPCR assay is highly suitable for early detection and quarantine in four porcine CoVs. The results indicate that this dPCR method is characterized by high specificity, anti-interference capabilities, repeatability, and high sensitivity. It also demonstrates a high compliance rate and diagnostic specificity in sample detection. This multiplex dPCR will contribute to the control of porcine enteric CoV-caused VDS and provide clues for subsequent research.
Keywords:
SADS-CoV; PEDV; PDCoV; TGEV; dPCR; porcine enteric coronavirus
1. Introduction
Coronaviruses (CoVs) are single-stranded, positive-sense RNA viruses with capsids. Their surface spike proteins interact with cell receptors in susceptible animals, facilitating invasion [1,2]. They are the largest known RNA viruses, capable of infecting a diverse range of vertebrates [3]. In the swine industry, porcine coronaviruses are major pathogens causing viral diarrhea, leading to significant economic losses and challenges in swine farming. Common porcine enteric coronaviruses include swine acute diarrhea syndrome coronavirus (SADS-CoV), porcine epidemic diarrhea virus (PEDV), porcine deltacoronavirus (PDCoV), and porcine transmissible gastroenteritis virus (TGEV). SADS-CoV is a new coronavirus discovered in Guangdong, China in recent years, belonging to the genus α-coronavirus, featuring a typical coronavirus structure and a genome length of approximately 27 kb [4,5]. It causes acute diarrhea and acute vomiting in infected piglets and a rapid decline in the body mass of newborn piglets within 5 days of age, leading to acute death, with a lethality rate of more than 90% [6]. PEDV, also an α-coronavirus, is a non-segmented, positive-sense, single-stranded RNA virus [7]. It infects pigs of all ages, particularly neonatal piglets, with a high incidence rate of more than 90%, and the incidence rate in sows ranges from 20 to 90% [8,9,10]. PDCoV, a δ-coronavirus, induces severe intestinal lesions in neonatal piglets [11,12]. In studies, the lumen of the small intestine in the infected piglets accumulated large amounts of yellowish fluid, the intestinal wall was thin and transparent, the intestinal villi of the jejunum and the ileum were severely atrophic, and enterocytes showed mild cytoplasmic vacuolization [4,11], with symptoms such as vomiting, acute diarrhea, dehydration and even death in severe cases [13]. TGEV, another α-coronavirus, targets porcine intestinal epithelial cells via the fecal–oral route and spreads rapidly, affecting all age groups [14,15,16]. Usually, in piglets within 20 d of age, if infected with TGEV, the survival time is less than 4 d [16]. The clinical characteristics of the onset of the disease are lethargy, depression, a short period of rapid weight loss, severe watery diarrhea, vomiting, and dehydration; the mortality rate can be up to 100% [17].
Since 2023, SADS-CoV outbreaks have mostly occurred in Southern China and have affected Guangdong, as well as its adjacent provinces, Fujian, Guangxi, and Jiangxi [18,19,20]. Although SADS-CoV has not reached pandemic levels, its novel structural properties and strong cell invasion capabilities pose a risk of zoonotic evolution, warranting heightened quarantine and control measures [21]. PEDV was first identified in the United Kingdom in 1971 [22] and began spreading in Asia during the early 1980s [23]. Between 2013 and 2014, PEDV epidemics spread from the United States to neighboring North American countries and quickly extended globally [24,25,26,27]. Currently, PEDV is recognized as a significant global health concern. PDCoV was initially discovered in 2012 in pig fecal samples during a coronavirus surveillance study in Hong Kong [28]. PDCoV infections have been reported in the U.S. and China in 2014, with substantial economic and safety concerns for the pig industry [29]. Studies indicate that PDCoV can infect animal-derived cell lines and achieve cross-species transmission; potential infection hosts include chickens, cattle, and even human beings [30,31,32]. TGEV was first discovered in the U.S. in 1946 and subsequently reported in countries across Europe, Asia, Africa, and the Americas [33]. In recent years, TGEV has been observed to be recombining and evolving continuously, and some of these recombinant strains can spread across species, making the detection and prevention of TGEV more complex [34,35].
Source: MDPI, Link: https://www.mdpi.com/1422-0067/26/17/8731
The FAI climbed 5.9 percent year-on-year in the first 11 months of 2018, quickening from the 5.7-percent growth in Jan-Oct, the National Bureau of Statistics (NBS) said Friday in an online statement.
The key indicator of investment, dubbed a major growth driver, hit the bottom in August and has since started to rebound steadily.
In the face of emerging economic challenges home and abroad, China has stepped up efforts to stabilize investment, in particular rolling out measures to motivate private investors and channel funds into infrastructure.
Friday's data showed private investment, accounting for more than 60 percent of the total FAI, expanded by a brisk 8.7 percent.
NBS spokesperson Mao Shengyong said funds into weak economic links registered rapid increases as investment in environmental protection and agriculture jumped 42 percent and 12.5 percent respectively, much faster than the average.
In breakdown, investment in high-tech and equipment manufacturing remained vigorous with 16.1-percent and 11.6-percent increases respectively in the first 11 months. Infrastructure investment gained 3.7 percent, staying flat. Investment in property development rose 9.7 percent, also unchanged.