Survival of porcine coronaviruses in feed ingredients and impact of feed additives

A lot of research has been done at the University of Minnesota regarding the survival of porcine coronaviruses in the feed and how to impact their survival. We are presenting today two papers published this spring looking at this important topic. First, Trudeau et al. showed that the feed ingredient which lead to the longest porcine coronaviruses’ survivability was soybean meal. Then, Cottingim et al. showed that some feed additives could inactivate PDCoV.

Importance of porcine coronaviruses and their relationship to swine feed

Porcine coronaviruses of importance in the swine industry nowadays are Porcine Epidemic Diarrhea virus (PEDV), Transmissible Gastroentiritis virus (TGEV), and Porcine Delta Coronavirus (PDCoV). All cause enteric issues in swine and some can lead to up to 100% mortality in nursing piglets. The role of feed ingredients in spreading PEDV and causing outbreaks in Northern America in 2013 has been questioned since then.

Survival of PEDV, TGEV, and PDCoV in complete feed and feed ingredients

The first research project evaluated the persistence of PEDV, TGEV, and PDCoV in porcine feed and feed ingredients. To do so, complete feed and major feed ingredients samples (spray dried porcine plasma, meat meal, meat and bone meal, blood meal, corn,
soybean meal, and corn dried distillers grains with solubles) where inoculated with PEDV, TGEV, or PDCoV and kept for up to 56 days. Aliquots were tested 11 times between the inoculation day and the end of the trial. Time necessary to reduce the viral concentration by 1 log was recorded.

Soybean meal took the longest time to attain the reduction in concentration for all of the coronaviruses, reaching 7.5 days for PEDV, and 42 days for both PDCoV and TGEV. This study also demonstrated that there was a modest positive correlation between moisture content and persistence of TGEV and PDCoV. On the other end, there was a moderate negative correlation between ether extract content and TGEV survival, not observed with the other two viruses.

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Trudeau coronavirus feed swine survival PED

Feed additives and PDCoV survival

In this second project, the survival of PDCoV was evaluated after being put in contact with nursery feed samples containing one of six different commercial feed acids (UltraAcid P, Activate DA, KEMGEST, Acid Booster, Luprosil, and Amasil), salt, or sugar. Acids were added following the recommended concentrations in the first part of the experiment and then, were double-dosed. Feed samples were inoculated with PDCoV and kept for up to 35 days. Like in the previous article, days to achieve a reduction of virus concentration by 1 log were recorded.

At recommended values, there was no difference between viral load reduction in feed samples with or without additives. When acids were added to the feed at a double concentration, the time period to attain the reduction in viral load was decreased to 0.28 days or less for all acids except for Amasil which increased it to 4.95 days (control: 0.35 days). The difference between acidifiers may be explained by the active ingredients used in the products. Furthermore, the addition of salt decreased PDCoV survival whereas sugar increased it.

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Cottingim feed additives survival PDCoV coronavirus swine

 

Detection of Mycoplasma hyopneumoniae in naturally infected gilts over time

A longitudinal study to assess Mycoplasma hyopneumoniae natural infection in gilts

This study was conducted by Dr. Karine Takeuti under the supervision of Dr. Maria Pieters from the University of Minnesota, College of Veterinary Medicine. The objective was to sample replacement gilts from 20 days of age until the day before weaning to detect Mycoplasma hyopneumoniae . Laryngeal swabs, tested by PCR, were taken every 30 days at a Mycoplasma positive sow farm. Therefore, the animals were naturally infected.

Gilts were found positive at 110 days, no detection in piglets

11.4% of the gilts were found positive at 110 days whereas all the previous samples came back negative. Positive results peaked at 140 days when 36.4% of the samples were positive for Mycoplasma hyopneumoniae. 27.3% of the gilts got positive results twice or more during the sampling period but 18.2% of the animals remained negative for the duration of the study.  All of the 220 piglets samples were also negative.

Takeuti longitudinal gilt mycoplasma hyopneumoniae 2017
Abstract

Mycoplasma hyopneumoniae causes a chronic respiratory infection in pigs and its transmission occurs mainly by direct contact and by vertical transmission (sow-to-piglet). The objective of this study was to assess the detection dynamics and persistence of M. hyopneumoniae natural infection in replacement gilts. Forty-four twenty-day-old gilts were selected from a M. hyopneumoniae positive farm and followed up to one day prior to their first weaning. Laryngeal swabs were collected every 30 days, starting at day 20, for M. hyopneumoniae detection by real-time PCR, resulting in 12 samplings. Piglets born to selected females were sampled via laryngeal swabs one day prior to weaning to evaluate sow-to-piglet transmission. The M. hyopneumoniae prevalence was estimated at each one of the 12 samplings in gilts and a multiple comparison test and Bonferroni correction were performed. Bacterial detection in gilts started at 110 days of age (doa) and a significant increase (p < 0.05) occurred at 140 doa. The M. hyopneumoniae prevalence remained above 20% from 140 to 230 doa, decreasing thereafter. However, it did not reach 0% at any sampling after 110 doa. In this study, M. hyopneumoniae was not detected in piglets sampled prior to weaning. The M. hyopneumoniae detection pattern showed that in natural infections, gilts were positive for M. hyopneumoniae for one to three months, but occasionally long-term detection may occur. Moreover, the lack of M. hyopneumoniae detection throughout the study in 18.2% of gilts indicated the existence of negative subpopulations in positive herds.

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Mycoplasma hyorhinis prevalence varies based on pigs’ age

Summary

  • Mycoplasma hyorhinis can cause polyserositis and arthritis in post-weaning pigs.
  • To study M.hyorhinis‘ prevalence based on age, nasal swabs were taken from pigs at 1, 7, 14, 21, 28, 35, 42, 49, 56, 63, 70 and 77 days as well as from sows, in 3 different Minnesotan herds (A, B, and C).
  • 8.8% of the sows were positive for M.hyorhinis in herds A and B whereas 3.3% of the sows were positive in herd C.
  • The percentage of positive piglets (<21 days of age) was low: between 0 and 10% depending on the herds.
  • At 28 days of age, the prevalence of M.hyorhinis in pigs increased dramatically to around 50% in herd A and 100% in herd B. After 42 days of age, the prevalence in those herds stayed above 95%.
  • The prevalence in herd C stayed close to 0% until the pigs reached the age of 77 days, time at which the prevalence increased to 100%.

Did you see our Science page on Mycoplasma hyorhinis and swine conjunctivitis?

Mhyorhinis prevalence baed on age Rovira 2017

Abstract

Mycoplasma hyorhinis is one of the causative agents of polyserositis and arthritis in postweaning pigs. Knowledge regarding colonization frequency and age distribution in modern pig production is lacking. The objective of this study was to estimate the prevalence of M hyorhinis colonization in different age groups across three commercial pig populations. Nasal swabs were collected from sows, piglets and nursery pigs of different ages. Oral fluids were collected from nursery pigs. Necropsies were performed to assess the presence of M hyorhinis-associated disease. M hyorhinis was detected in 5/60 sows in herd A, 3/60 in herd B and none in herd C. In herd A and B, the prevalence was low in preweaning piglets (∼8 per cent) and high in postweaning pigs (∼98 per cent). A total of 7/8 oral fluids tested PCR positive in herds A and B, while 1/8 tested positive in herd C. In herd C, the preweaning and postweaning prevalence was low. In herds A and B, necropsied pigs had polyserositis lesions where M hyorhinis was detected by PCR. This study showed that prevalence of M hyorhinis colonization varies with pig age and across farms. Information generated will aid in the design and implementation of control and prevention strategies.

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Detecting Senecavirus A in tissues: development of a new diagnostic test at the University of Minnesota

Summary

In Canada and the USA alike, Senecavirus A is a challenge for producers and veterinarians because of its clinical similarity to Food and Mouth Disease (FMD). Indeed, Senecavirus A, is a causative agent of swine vesicular disease with lesions developing on the snout, around the mouth and on the coronary band of the feet. Therefore, being able to differentiate Senecavirus A infections from FMD rapidly is of utmost importance to be able to take the appropriate measures.

In the past months, several diagnostic tests have been developed at the University of Minnesota to detect antibodies against Senecavirus A. The difference between those tests and the in situ hybridization (ISH) described here is that ISH targets the genetic material included in the viral particle and marks it as a red spot as can be seen on the figure below. This advantage of this method is to be able to locate the virus and gives additional information to researcher wanting to study the behavior of Senecavirus A in the body of the pig.

in situ hybridization senecavirus A pigs
Red dots and clusters represent the presence of SVV mRNA within an erosive lesion on the tongue of a pig © 2017 Resende et al.

Abstract

Seneca Valley virus (SVV) is the causative agent of an emerging vesicular disease in swine, which is clinically indistinguishable from other vesicular diseases such as foot-and-mouth disease. In addition, SVV has been associated with neonatal mortality in piglets. While a commercial SVV qRT-PCR is available, commercial antibodies are lacking to diagnose SVV infections by immunohistochemistry (IHC). Thus, a novel in situ hybridization technique—RNAscope (ISH) was developed to detect SVVRNA in infected tissues. From a total of 78 samples evaluated, 30 were positive by qRT-PCR and ISH-RNA, including vesicular lesions of affected sows, ulcerative lesions in the tongue of piglets and various other tissues with no evidence of histological lesions. Nineteen samples were negative for SVV by qRT-PCR and ISH-RNA. The Ct values of the qRT-PCR from ISH-RNA positive tissues varied from 12.0 to 32.6 (5.12 x 106 to 5.31 RNA copies/g, respectively). The ISH-RNA technique is an important tool in diagnosing and investigating the pathogenesis of SVV and other emerging pathogens.

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Can biosecurity measures prevent PEDV transmission?

Summary:

Porcine Epidemic Diarrhea virus is highly contagious.

The 2013 Porcine Epidemic Diarrhea virus’ (PEDV) outbreak in the USA taught the swine industry that the virus is highly contagious. This event forced producers and veterinarians to review and upgrade their biosecurity procedures.

Drs. Torremorell, Cheeran, and Goyal from the University of Minnesota evaluated some of these measures and how they can prevent PEDV transmission.

Changing Personal Protective Equipment (PPE) and showering before entering a new room prevented contamination.

Among the measures included in this study were the use and change of PPE as well as showering in and out of a facility. In the low biosecurity setting, personnel went from a room with PEDV positive pig straight to a room with naive pigs, contaminating them after the very first movement. In the medium biosecurity setting, personnel washed their hands and face and change their PPE before being in contact with the naive pigs. In this situation, pigs stayed negative for PEDV but  two personnel hair/face swabs came back positive for viral genetic material. On the contrary, personnel showered before getting in contact with the high biosecurity group. Those pigs as well as all personnel tests remained negative for PEDV during the study.

 

Torremorell PEDV biosecurity 2017

Abstract

Background:

The effectiveness of biosecurity methods to mitigate the transmission of porcine epidemic diarrhea virus (PEDV) via farm personnel or contaminated fomites is poorly understood. This study was undertaken to evaluate the effectiveness of biosecurity procedures directed at minimizing transmission via personnel following different biosecurity protocols using a controlled experimental setting.

Results:
PEDV RNA was detected from rectal swabs of experimentally infected (INF) and sentinel pigs by real-time reverse transcription polymerase chain reactio n (rRT-PCR). Virus shedding in INF pigs peaked at 1 day post infection (dpi) and viral RNA levels remained elevated through 19 dpi. Sentinel pigs in the low biosecurity group (LB) became PEDV positive after the first movement of study personnel from the INF group. However, rectal swabs from pigs in the medium biosecurity (MB) and high biosecurity (HB) groups were negative during the 10 consecutive days of movements and remained negative through 24 days post movement (dpm) when the first trial was terminated. Viral RNA was detected at 1 dpm through 3 dpm from the personal protective equipment (PPE) of LB personnel. In addition, at 1 dpm, 2 hair/face swabs from MB personnel were positive; however, transmission of virus was not detected. All swabs of fomite from the HB study personnel were negative.
Conclusions:
These results indicate that indirect PEDV transmission through contaminated PPE occurs rapidly (within 24 h) under modeled conditions. Biosecurity procedures such as changing PPE, washing expose d skin areas, or taking a shower are recommended for pig production systems and appear to be an effective option for lowering the risk of PEDV transmission between groups of pigs.

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Characterizing Canadian rotavirus A strains and their similarity to a commercial vaccine

Summary:

Rotaviruses A are genetically diverse.

Rotaviruses are responsible for increased mortality in neonatal swine populations. They are different genetically and more studies are needed to characterize their diversity. This is the objective of this study coordinated by Dr. Marthaler’s lab focusing on rotaviruses strains found in Canada.

Viral proteins 7 and 4 are used for rotavirus A classification.

Rotaviruses are classified based on two viral proteins (VP) found on their outer capsid called respectively VP7 and VP4. Those two proteins are also essential to induce an efficient immune response against the virus. This project characterized VP7 and VP4 sequences in 136 Canadian samples and compared them with the strains used in a rotavirus commercial vaccine.

The VP7 (n=32) and partial VP4 (n=25) were analyzed, identifying the G3P[13], G5P[7], G5P[x], G9P[7], G9P[13], G9P[19], and G9P[x] genotypes.
Minimal differences in the antigenic epitopes for the G5, G9, and P[7] strains were identified.
Major differences in the antigenic epitopes of the G3, P[13], and P[19] may question the effectiveness of the ProSystems RCE RVA.

Marthaler rotavirus A Canada 2017

Abstract

Surveillance of Rotavirus A (RVA) infections in North America swine populations are limited and not performed over a significant time period to properly assess the diversity of RVA strains in swine. The VP7 (G) and VP4 (P) genes of 32 Canadian RVA strains, circulating between 2009 and 2015 were sequenced, identifying the G3P[13], G5P[7], G9P[7], G9[13], and G9[19] genotype combinations. The Canadian RVA strains were compared to the RVA strains present in the swine ProSystems RCE rotavirus vaccine. The comparison revealed multiple amino acid differences in the G and P antigenic epitopes, regardless of the G and P genotypes but specifically in the Canadian G3, P[13] and P[19] genotypes. Our study further contributes to the characterization of RVA’s evolution and disease mitigation among swine, which may optimize target vaccine design, thereby minimizing RVA disease in this economically important animal population.

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Sample and diagnostic types for early detection of Mycoplasma hyopneumoniae

Summary:

Mycoplasma hyopneumoniae is the causative agent enzootic pneumonia, an economically significant disease in pigs. In this study published by Drs. Pieters and Rovira from the University of Minnesota, pigs experimentally inoculated with M.hyopneumoniae were sampled 0, 2, 5, 9, 14, 21, and 28 post-inoculation.

Different sample types were compared:

  • Nasal swabs
  • Laryngeal swabs
  • Tracheobronchal lavages
  • Oral fluids
  • Serum samples

Using different diagnostic tests:

  • PCR
  • ELISA IgG anti M.hyopneumoniae
  • ELISA Ig M anti M.hyopneumoniae
  • ELISA C-reactive protein

Laryngeal swab samples tested by PCR were highly sensitive for detection of Mycoplasma hyopneumoniae in live pigs.
Various commercial ELISA kits for detection of Mycoplasma hyopneumoniae antibodies showed similar sensitivity.
Oral fluids showed a low sensitivity for detection of Mycoplasma hyopneumoniae in experimentally infected pigs.

Pieters Mhyopneumoniae early detection test sample 2017

Abstract

Detection of Mycoplasma hyopneumoniae in live pigs during the early stages of infection is critical for timely implementation of control measures, but is technically challenging. This study compared the sensitivity of various sample types and diagnostic methods for detection of M. hyopneumoniae during the first 28 days after experimental exposure. Twenty-one 8-week old pigs were intra-tracheally inoculated on day 0 with M. hyopneumoniae strain 232. Two age matched pigs were mock inoculated and maintained as negative controls. On post-inoculation days 0, 2, 5, 9, 14, 21 and 28, nasal swabs, laryngeal swabs, tracheobronchial lavage fluid, and blood samples were obtained from each pig and oral fluid samples were obtained from each room in which pigs were housed. Serum samples were assayed by ELISA for IgM and IgG M. hyopneumoniae antibodies and C-reactive protein. All other samples were tested for M. hyopneumoniae DNA by species-specific real-time PCR. Serum antibodies (IgG) to M. hyopneumoniae were detected in challenge-inoculated pigs on days 21 and 28. M. hyopneumoniae DNA was detected in samples from experimentally inoculated pigs beginning at 5 days post-inoculation. Laryngeal swabs at all samplings beginning on day 5 showed the highest sensitivity for M. hyopneumoniae DNA Detection, while oral fluids showed the lowest sensitivity. Although laryngeal swabs are not considered the typical M. hyopneumoniae diagnostic sample, under the conditions of this study laryngeal swabs tested by PCR proved to be a practical and reliable diagnostic sample for M. hyopneumoniae detection in vivo during early-stage infection.

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