Science Page: To filter or not to filter, that’s not the question anymore!

This is our Friday rubric: every week a new Science Page from the Bob Morrison’s Swine Health Monitoring Project. The previous editions of the science page are available on our website.

This week, Drs. Torremorell and Janni explain what is new in the world of air filtration.

Key Points

  • There are multiple components that affect the effectiveness of filtration for a particular farm.
  • Virus concentrations, particle size, and prevalence of particle size impact virus concentration within the barn.
  • Filtration type and functionality over season can also affect virus concentration within the barn.

Long gone are the days when we debated whether it’s beneficial to install air filtration in a farm. If you are in a high dense area and you have new breaks often enough that filtration pays off, then filtrate! Whether it is the removal of virus from the air (which is what filters do), or the enhancement and enforcement of basic biosecurity measures which are part of the “filtration package”, air filtration has been shown to reduce the number of PRRSV (porcine reproductive and respiratory syndrome virus) breaks.

However, filtration has not always met farm owner expectations. Retrospective data analysis from the Morrison Swine Health Monitoring Project suggests that PRRSV incidence hasn’t been reduced as much as we had hoped for, although it’s still better than no filtration.

So, what can be going on?

To help producers and veterinarians to contrast and compare filtration options and to help understand the components that affect filtration, we created a model that estimates the theoretical number of airborne viruses that would enter a barn through either filters or through leakage given certain assumptions.

The model takes into account various inputs considered important to affect filtration and provides a look at the interaction between ventilation rate, building leakage, filter particle removal efficiency and particle size distribution where viruses may attach.

We have learned a few things already.

First, and the obvious one, is that filtering ventilation air does decrease virus concentration inside filtered barns. Second, ambient virus concentrations and their size distributions, which are largely unknown, have a large impact on virus concentration inside the barn. Although we have done some measurements of virus distribution based on particle size, the relative distribution is likely affected by various factors such as environmental conditions, type of virus, source of virus aerosol, etc.

In general we can say that if viruses are found mostly in the smallest particles (<1 microns), MERV 14 filters will do quite poorly since their lowest removal efficiency is for particles less than 1 micron (~ 76% removal efficiency). In contrast, if the largest amount of virus is found in medium and large particles (>1 micron), as our studies suggest, then MERV 14 filters should do quite well and the more viruses there are in the largest particles (>3 microns), the better MERV 14 filters do and in this case performance is similar to MERV 16 filters.

However, since virus particle size distribution is likely to change throughout the day and season, the only way to minimize the impact of virus particle size distribution is by using higher MERV rating filters with removal efficiencies above 95 % if not more. But, the overall importance of filter collection efficiencies is uncertain because the ambient virus concentrations and their size distributions are not really known.

The other point that is important to recognize is that there are higher barn virus concentrations with lower mechanical ventilating rates and higher barn leakage rates. In other words, during low ventilation rates (i.e. winter) the number of virus particles per cubic feet per minute is higher than during high ventilation rates (i.e. summer). However, when we consider total amount of virus particles that may enter the barn per minute, then higher ventilation rates result in higher in-barn virus concentration compared to lower ventilation rates. This observation is important also when considering positive ventilated barns since it is not uncommon that in the winter, they operate at higher flow rates than negative ventilated barns resulting in the potential introduction of more viruses through the filter, even though leakage is nearly eliminated.

Lastly, as it is well known reducing barn static pressure drop by increasing filter area helps reduce leakage and virus concentrations in the barn. So, it is not a question whether air filtration helps, but rather knowing which factors to consider when making the best of air filtration. Our model does not measure risk of PRRSV infection into a farm but it shows a fairly complex, not always obvious, interaction between ventilation rate, building leakage, filter particle removal efficiency and viral particle size distribution that knowing it, may be useful to producers and veterinarians when evaluating air filtration systems for sow farms.

For more information about the model, contact Kevin Janni (kjanni-at-umn.edu) or Montse Torremorell (torr0033-at-umn.edu) at the University of Minnesota.

UMN swine groups meet with the Minnesota Pork Board members

On December 17th, swine-focused faculty members from the College of Veterinary Medicine and the College of Food, Agricultural and Natural Resource Sciences met with the members of the Minnesota Pork Board research committee. 

At the Christensen Farms’ truck wash

This year was the third iteration of this meeting, hosted for the first time by Christensen Farms. Guests were invited to tour either the feed mill or the truck wash located in Sleepy Eye before sitting down for a day of productive exchange. 

After a short presentation by the host company, Deans Brian Buhr and Trevor Ames gave an update on the main initiatives from their respective Colleges. Dean Ames introduced the latest AGREETT faculty hires, Noelle Noyes and Declan Schroeder, as well as the anticipated arrival of Dr. Matheus Costa in the new year. The 2+2 program in collaboration with South Dakota State University was received with enthusiasm by the swine producers.

After lunch, attendees were divided into groups to brainstorm research ideas in four main areas: swine nutrition, swine health, manure management, and swine production and housing. After two hours of spirited discussion, 15 researchable questions were identified as the most pressing problematics faced by the Minnesotan pork industry. The meeting left the researchers inspired to keep offering science-driven solutions, and all agreed to meet again in 2019.

NHF: What does livestock-associated MRSA mean for the neighborhood?

Our monthly collaboration with the National Hog Farmer continues; this month Dr Peter Davies from the University of Minnesota, College of Veterinary Medicine,  explains what is livestock-associated MRSA and if it affects people living near pig farms.

“Livestock-associated” MRSA first isolated in 2004

Methicillin Resistant Staphylococcus aureus (MRSA) Source: Wikimedia Commons

Most people are likely familiar with MRSA (methicillin resistant Staphylococcus aureus), a flagship “superbug” that is a major concern to human medicine. And just about everybody in the pig industry has heard that certain variants of MRSA are very common in some livestock populations (including pigs), and these are referred to as “livestock-associated” MRSA (LA-MRSA).

A novel variant of MRSA (labelled ST398 using a DNA typing method) was first found in pigs in the Netherlands in 2004. Subsequently, ST398 MRSA and several other types (e.g., ST9 in Asia, ST5 in North America) were reported in pigs in numerous countries, and often in their caretakers as well. The discovery that pigs may be a large MRSA reservoir created some justifiable panic and confusion, raising questions about the implications for human health, particularly for industry workers (e.g., farmers, veterinarians, processing plant workers), pork consumers and, last but not least, people living in the neighborhood of pig farms.

Generally, LA-MRSA  lack most of the key “virulence factors” that enable the bacteria to cause clinical infections in people.

Human clinical infections by “livestock-associated” MRSA are rare

Although workers on MRSA-positive farms often harbor LA-MRSA in their nose, significant clinical infections in healthy workers have been rare. Human clinical infections with LA-MRSA do occur, but most cases tend to be of relatively mild disease (such as skin infections), with more severe infections typically limited to elderly and medically compromised patients.

Remembering that about 2% of healthy U.S. citizens carry human adapted variants of MRSA, the relative clinical importance of LA-MRSA appears to be minimal in most countries. Globally since 2004, there have been around 10 fatal cases of LA-MRSA infections reported, compared with about 50 fatal MRSA cases per day (18,650 per year in 2005) in the United States alone.

2016 study of Iowa hospitals found probable livestock variants in only 0.24% of MRSA cases, and 1% of S. aureus infections. In North Carolina, another leading swine-producing state, there were no LA-MRSA variants among more than 1,200 MRSA isolated from human bloodstream infections between 1995 and 2015 (Dr. Vance Fowler, Duke University, personal communication).

Living next to pig farms does not increase the risk of exposure

Although MRSA can be isolated from meat products, there is little evidence to suggest cause for concern about food-borne transmission. In contrast, conclusions of studies looking at the neighborhood risk of exposure to LA-MRSA from pig farms are conflicting. We will focus on the findings of studies that have compared pig workers and neighbors directly, measured the distance from pig barns to residences directly; and used laboratory testing to confirm the presence of LA-MRSA in the study populations. 

Across three early studies in Europe, LA-MRSA prevalence (nasal carriage) was greater than 180 times higher in 352 pig industry workers (44%) than in 2,094 rural residents without farm exposure (0.24%).

  • A similar study in Holland in 2017 showed similar prevalence of nasal colonization (0.56%) in people without livestock contact, but also found the positive people on average lived closer to farms (of any type). Importantly, the authors noted that routes of transmission underlying this were not known.
  • A very detailed study of cases of MRSA infection in Denmark showed that overall MRSA risk did not differ between pig-dense regions versus other regions. However, the likelihood that a MRSA infection would be a LA-MRSA type was higher in the pig-dense regions, confirming some “spillover” from the industry to the community.
  • Notably, a follow-up study published in 2018 found that, within pig-dense areas, the patients with LA-MRSA infections did not live closer to pig farms than population controls. The authors conclude that direct environmental spread from neighboring pig farms was unlikely and suggested that community spread through contact with people working with livestock,  might be the predominant mechanism.

In summary, the overall impact of LA-MRSA relative to human variants remains very small in most countries including the United States. There is no evidence that residence in rural areas increases overall MRSA risk.

Science Page: Biosecurity screening tool; Benchmarking PRRSv biosecurity vulnerability using a short survey

This is our Friday rubric: every week a new Science Page from the Bob Morrison’s Swine Health Monitoring Project. The previous editions of the science page are available on our website.

This week we are sharing a report by Dr. Linhares’ lab at Iowa State University. In this Science Page are the results of a study looking at biosecurity aspects associated with PRRS frequency.

Key Points

  • New methods allow estimation of the overall PRRS-vulnerability risk score by asking 20 or less questions.
  • This can help producers and veterinarians to (a) measure and benchmark key biosecurity aspects, and (b) toidentify sites at relatively higher (or lower) risk of PRRSv introduction.

Study Summary: This study aimed to identify a small set of biosecurity aspects that, when combined, have a strong association with the frequency of PRRSv introduction into swine breeding herds.

Parameters included in the 2 models (A and B) to predict the number of PRRS outbreaks in farms for the past 5 years.

Preliminary Results: A cross-sectional study assessed biosecurity aspects in 84 breeding herds from 14 production systems in 2017. Models were trained to predict whether a farm had or not reported a PRRS outbreak in the past 5 years, given a set of biosecurity aspects. Two methods were used, and both models were able to classify the herds with a great overall performance based on few biosecurity aspects (See figure). The variables used by both methods were related to the frequency of risk events in the farm, swine density around the farm, farm characteristics/ requirements to visitors, and operational connections to other sites.

Note: The Gini coefficient (or index) is a single number aimed at measuring the degree of inequality in a distribution. (Source: Wikipedia) The higher the number, the less equally distributed the farms will be.

When comparing the predicted positive value obtained by the models, they showed a strong positive correlation (0.7 and 0.76, respectively) with the frequency of past outbreaks.

Enroll on our follow-up study: Study farms will be asked to fill a short survey. Using the methods above, the PRRS-vulnerability risk score will be generated for each farm enrolled. The information will be collected via an Excel file and the name of the farms and production systems will be kept confidential.

To enroll or request additional information please contact: Gustavo Silva (gustavos-at-iastate.edu) or Daniel Linhares (linhares-at-iastate.edu) at Iowa State University.

Effects of Lawsonia intracellularis infection in the proliferation of different mammalian cell lines

Today, we are sharing a publication by Dr. Talita Resende, a phD candidate working with Drs. Gebhart and Vannucci. Dr. Resende’s research focuses on the mechanisms enabling Lawsonia intracellularis’ infectivity and pathogenesis. Her latest paper, available in open access from Veterinary Microbiology, looks at the effects of Lawsonia intracellularis on different cell lines.

Highlights

  • Effects of L. intracellularis on intestinal cell lines in vitro is unknown.
  • Impact of nutrient deprivation on cell proliferation was cell line dependent.
  • L. intracellularis did not lead to proliferation of the cell lines tested.
  • L. intracellularis and Ki-67 were co-localized in all cell lines tested.
  • Single cell cultures are not a suitable model for L. intracellularis pathogenesis.

Material and Methods

4 different intestinal epithelial cells lines were compared in this study: IPEC-J2 , IEC-18, Caco-2, and  McCoy cells. McCoy were used as a reference since previous publications have shown that Lawsonia intracellularis can grow in this cell type.

Each cell line was infected with 2 types of Lawsonia intracellularis: low and high passage. Infected cell lines were used as control during the experiment. At days 1, 4, and 7 post-infection, the number of cells highly infected by Lawsonia (i.e. that had more than 30 organisms in their cytoplasm) was counted. To estimate cell proliferation, the amount of DNA in each cell line was evaluated. Additionally, a fluoerescence marker called Ki-67 was used to identified eukaryotic cells undergoing division. Lastly, a wound closure assay was done by scraping infected cell lines with a pipette and measure the width of the “wound” over time.

Results and Discussion

Arrows point towards cells highly infected by Lawsonia intracellularis.
Credit: Veterinary Mivrobiology

All cell lines tested were susceptible to L. intracellularis infection with typical intracellular bacterial growth of about 30–100 per cell in the cytoplasm of infected cells. 

There was no statistical difference in cellular proliferation within or among groups at 0 and 1 dpi. Additionally, no increased proliferation in any cell line infected by L. intracellularis was noted, regardless of the bacterial passage status.

To verify whether cells infected by L. intracellularis would proliferate and migrate faster than non-infected cells through a scratched monolayer, a wound closure assay was executed. There were no differences among treatment groups for wound closure at any time point (0 to 24h and 24h to 48h)

It is suggested that L. intracellularis preferentially infects actively proliferating cells in intestinal crypts. By looking at both Lawsonia and Ki-67 markers, it was noted that in the majority of treatment groups and with the exception of the IPEC-J2 cell line, the proportion of cells that were double positive (L. intracellularis was co-localized with Ki-67) was higher than cells that were L. intracellularisinfected, but negative for Ki-67.

Taken together, these findings have decisively shown that two-dimensional intestinal epithelial in vitro cultures do not reproduce the characteristic proliferative effect of L. intracellularis infection in vivo.

Access to the entire paper

Abstract

Lawsonia intracellularis is an obligate intracellular bacterium that causes proliferative enteropathy in various animal species. While cellular proliferation of intestinal cells is recognized as the hallmark of L. intracellularis infection in vivo, it has not been demonstrated in in vitromodels. In order to assay the effect of L. intracellularis, various cell lines were infected with pathogenic and non-pathogenic passages of the bacterium. Because of the high proliferative rate of these cell lines, serum deprivation, which is known to reduce proliferation, was applied to each of the cell lines to allow the observation of proliferation induced by L. intracellularis. Using antibodies for Ki-67 and L. intracellularis in dual immunofluorescence staining, we observed that L. intracellularis was more frequently observed in proliferating cells. Based on wound closure assays and on the amount of eukaryotic DNA content measured over time, we found no indication that cell lines infected with L. intracellularis increased proliferation and migration when compared to non-infected cells (p > 0.05). Cell arrest due to decreased serum in the culture media was cell-line dependent. Taken together, our findings provide data to support and expand previous subjective observations of the absence of in vitro proliferation caused by L. intracellularis in cell cultures and confirm that cell lines infected by L. intracellularis fail to serve as adequate models for understanding the cellular changes observed in proliferative enteropathy-affected intestines.

Science Page: Assessing the relative vulnerability of swine breeding herds to the introduction of PRRS virus

This is our Friday rubric: every week a new Science Page from the Bob Morrison’s Swine Health Monitoring Project. The previous editions of the science page are available on our website.

This week we are sharing a report by Dr. Daniel Linhares’ lab at Iowa State University. The report summarizes the findings of his study regarding the factors making a sow farm vulnerable for PPRS introduction. 

Key Points:

  • A model to quantify and identify biosecurity vulnerability in breeding herds is now available.
  • Events related to swine movements, transmission by air and water, and people movements were the variables most associated with PRRS outbreak.
  • Biosecurity vulnerability scores may help producers/veterinarians prioritize biosecurity investments.

Study Summary:

Herd-specific biosecurity assessments are needed to determine herd-specific risk for PRRS outbreaks. Thus, we developed and validated a biosecurity vulnerability score (BVS) that measures the relative vulnerability of swine breeding herds to PRRSv introduction. The BVS was based on a multi-criteria decision algorithm that ranked risk events associated with outbreaks. A comprehensive biosecurity assessment was used to obtain information of the biosecurity practices from each participating herd. The practices performed in each herd were weighted by the relative importance of each event obtained from an expert opinion panel resulting in a score that identifies the events that should be prioritized. In two independent data sets, the scores consistently revealed that farms with higher scores had a higher frequency of PRRS outbreaks. In addition, results suggest that events related to swine movements,transmission by air and water, and people movements should be prioritized.

Follow-up study:

We are developing a new screening tool to validate the minimum number of questions associated with frequency of PRRS outbreak. Study farms will be asked to fill out a short survey. This can help producers and veterinarians to identify sites at relatively higher risk of PRRSv introduction.

To enroll or to request additional clarification please contact: Gustavo Silva at Iowa State University (gustavos-at-iastate.edu)