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)

Best of Leman 2018 series #2: M. Schwartz – A systems approach to M. hyopneumoniae elimination

We launched a new series on the blog last year. Once a month, we are sharing with you a presentation given at the Allen D. Leman swine conference, on topics that the swine group found interesting, innovative or that lead to great discussions.

We can find all of the presentations selected from last year’s conference on the blog here.

Our second presentation for this year is from Mark Schwartz from Schwartz Farms sharing their experience trying to eliminate Mycoplasma hyopneumoniae from their system.

Click on the image below to see his presentation at the conference:

Science Page: African Swine Fever transmission and survivability

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.

There is no Science Page this week so we are sharing a favorite from this year, in which Dr. Carles Vilalta created a literature review on ASF virus transmission and survivability.

Keypoints

  • New introductions of ASF to free areas of the disease are usually by uncooked pork fed to pigs.
  • Virus can be inactivated with temperature and low pH.
  • Survivor animals may play a role in the transmission and persistence of the disease.

Further outbreaks of African Swine Fever virus (ASFV) were reported last week in China several miles away from what is thought to be the first outbreak. This geographic dispersal leads us to think about dissemination mechanisms within the country and between countries.

EPIDEMIOLOGY

Infected animals will go through a viremic phase and can shed the virus through nasal secretions, feces and urine. Therefore, the main transmission route is oral-nasal, as pigs can be exposed to ASF positive secretions or tissues (i.e. pork products). Indirect transmission can also occur by exposure to contaminated fomites. This virus can also be transmitted by ticks. This vector-borne route becomes relevant when the wild boar
population is present and moves across regions and countries. The common introduction route into ASF free regions is usually through positive pigs transported into the area, or contaminated pork products that are fed to other pigs. ASFV has also been detected in air samples; however, airborne transmission is considered a secondary route of transmission due to the high virus load needed.

VIRUS SURVIVABILITY

Inactivation and persistence

Although ASFV is highly resistant, the virus can be inactivated at pH < 4 and pH >11. Survivability outside the host is heavily related to temperature. For instance, the infectious half-life in urine and feces can range from 3 to 15 days and 4 to 8 days at 37°C and 4°C, respectively. The virus may persist for several weeks or months in frozen, fresh, or uncooked pork, as well as in salted dried pork products. In contrast, ASFV is inactivated at high temperatures (i.e. 70°C cooked or canned hams) and in cured or processed products such as Spanish cured pork products after day 122–140 of curing. Pigs can become persistently infected and the virus can stay viable in their carcasses for up to six months. Therefore, infected carcasses represent a risk to other pigs. More recently, an investigation simulating a trans-Atlantic shipping of ASFV contaminated feed ingredients from Europe proved that viable virus can be recovered after 30 days.

The role of survivor pigs

ASFV recovered and sub-clinically infected pigs become a source of virus to other pigs. This plays an important role in disease transmission and persistence in endemic areas as well as becoming one of the most important routes of transmission into disease-free zones. In-vivo experiments have revealed an infectious period of moderately virulent virus isolates ranging from 20 to 40 days. In another in-vivo transmission study, pigs that had been exposed to ASFV 90 days prior were commingled with naive pigs and the virus was transmitted to naive pigs.

Serological field studies performed in positive regions of Brazil, the Iberian Peninsula, East Africa, Kenya and Uganda revealed that the there was a very low percentage of seropositive animals one year after the outbreak. It was hypothesized that those few seropositive pigs were still carriers and could have been responsible of some of the newer outbreaks.

CONCLUSION

ASF has a complex epidemiology with different routes of transmission that can involve animals and ticks as direct transmission, and contaminated clothes, tools, and surfaces as indirect transmission. Thus, early detection and intervention of the diseases are key to containing disease spread in absence of an effective vaccine.

ASF threat: 3 swine vets share insights from the frontline

The rapid spread of African swine fever (ASF) throughout China and other regions of the world has raised concerns the disease will ultimately make its way to the US — a development that could cripple the nation’s pork industry if it doesn’t adequately prepare.

That was the ominous warning of three US swine veterinarians who came together for a roundtable discussion on ASF following their recent trip to China. Among them was Dr. John Deen from the University of Minnesota, coming back from the Leman China conference.

Follow the link to listen to their podcast.

The informative session was organized by the editors of Pig Health Today and sponsored by Zoetis.