What Do Food Microbiology Students Know About Campylobacter?

We asked a class of university students taking Food Microbiology at North Carolina State University to fill out a survey regarding knowledge of Campylobacter. The class was composed of students from the Food Science department (denoted FS) and Microbiology department (MB). There were both undergraduate and graduate students in the course.

The first question asked the students if they had ever heard of Campylobacter. Of 57 students who participated in the survey, 34% responded “no” and 66% responded “yes”. The data was further divided and Fisher’s Exact tests were performed to see if the class (FS405, FS505, MB405 or MB505), major (Food Science or Microbiology) or rank (undergraduate or graduate) affected the outcome of having heard of this pathogen. There was no statistically significant effect of class on the outcome of whether or not students had heard of Campylobacter (p=0.298). Additionally, there was no effect from major ( food science or microbiology) or status (graduate vs. undergraduate) (p= .750 and .097 respectively).

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If participants answered yes to the first question, we then asked them if Campylobacter was a virus, a bacterium, or chemical. The vast majority (97%) of participants who had heard of Campylobacter knew that it is a bacterium.

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Next, the participants were asked what percentage of foodborne illnesses caused by 31 known pathogens Campylobacter is responsible for? The most commonly reported answer was that Campylobacter is responsible for 25% of foodborne illnesses. The correct answer of 9% was selected by 30% of participants, and no participants selected chemical as a response.

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Next, the participants were asked where one might catch Campylobacter. The most common and a correct response (34%) is that Campylobacter is associated with poultry. However, 18% of respondents mistakenly thought that this might include eggs. Unpasteurized milk and dairy products was another common response (23%) and is also a correct answer. No respondents chose a sick friend as a mode of infection, and 18% of respondents chose produce.

Finally, participants were asked where they get information about foodborne outbreaks. The results were nearly equally split between social media (30%) and TV (29%). Word of mouth was also popular (15%). Print news (9%), radio (4%), government (5%), and other (8%) made up the rest. The “other” category commonly included other websites such as FoodSafetyNews.com which the students wrote in.

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It is not surprising that so many of the students taking Food Microbiology have heard of Campylobacter. These students take general microbiology, and might have learned about foodborne pathogens then. Students are over estimating the burden of reported foodborne diseases from Campylobacter in the U.S . They also do not have a clear understanding of the routes of transmission.

 This survey was administered at the beginning of the semester. It may help professors identify gaps in students’ knowledge regarding this organism. For example, students should understand that Norovirus and Salmonella are responsible for many more illnesses than Campylobacter. However, the possibility of serious consequences from campylobacteriosis makes this a serious organism to consider. Additionally, since Campylobacter is rarely responsible for large outbreaks, it may not make headlines. Survey participants get much of their news on social media and TV and may miss these reports.

2014 NARMS data released!

On November 18, 2016 the 2014 NARMS report was released. The report includes a brand new interactive tool that lets users choose which combination of pathogen, source, and antibiotic results they would like to see. It’s especially useful because it makes comparing current data to past results incredibly easy. The 2014 NARMS report tested a  total of 4,122 Campylobacter isolates from human clinical cases, turkeys, cattle, and swine.  33% of retail chickens carried Campylobacter, the lowest level since 2003. Additionally, 12% of chicken ceca was Campylobacter positive and 6.1% of turkey.

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Macrolide resistance is especially interesting to Campylobacter researchers because they are the first line therapies for human cases and are approved for use in all food-producing animals. Since testing began in 1997, macrolide resistance in C. jejuni has remained below 4%. In 2014, resistance to macrolides fell below 2% in all sources except market hogs. Macrolide resistance in C. coli is  much higher than in C. jejuni. In the 2012-2013 NARMS report the incidence had more than doubled in human, retail, and PR/HACCP chicken C. coli isolates. This led NARMS to consider whole genome sequencing (WGS) to test for the presence of the recently identified erm(B) gene. Noneof the 12 tested isolates carried erm(B). These resistant isolates carried previously identified mutations in the 23 S ribosomal gene, and were not genetically similar indicating that there has not been clonal expansion of this resistance in Campylobacter.

Fluoroquinolones (e.g. ciprofloxacin and nalidixic acid) were banned for use in poultry in 2005. NARMS data indicates that resistance to fluoroquinolones in C. jejuni isolated from humans has reached the highest level since NARMS testing began in 1998. This is important as the fluoroquinolones are an alternative treatment for campylobacteriosis. Retail chicken and PR/HACCP chicken isolates also experienced increased incidence of resistance. Fluoroquinolone resistance could not be determined because of the number of turkey-derived C. jejuni  isolates (n=1). Of the 15 C. coli isolates from turkeys, approximately 40% were resistant to fluoroquinolones.

1.4% of the 1,251 C. jejuni isolates from humans were resistant to gentamicin. None of the eight C. jejuni from chicken ceca or the 369 retail chicken isolates were resistant to gentamicin. To the best of the author’s knowledge, no C. jejuni from turkeys were tested for resistance to gentamicin. This may be due to there only being one C. jejuni isolated from turkeys. However, 13.3% of the 15 C. coli isolates taken from turkey ceca were found to be gentamicin resistant. The Kathariou lab utilizes the yearly NARMS reports for much of their research. Check out more of our website to see what the Kathariou lab is doing to monitor antibiotic resistance in Campylobacter.

 

 

 

The Economic Burden of Foodborne Illnesses- How Much Does Campylobacter Contribute?

A 2015 USDA publication estimated the yearly economic burden of 15 foodborne illnesses on the U.S. population at $15.5 billion. Right off the bat it should be clarified that economic burden is NOT the same thing as out-of-pocket expenses. The economic burden is a result of factors such as the illness’ “frequency, severity, and health impact”1. The report states that “Conceptually, economists measure the economic burden of a disease as the sum of the willingness to pay by all individuals in society to reduce its incidence or likelihood”1. Campylobacter spp. ranks 5th in this list for the greatest contributor to economic burden at $1.9 billion.

Campylobacter accounts for 9% of illnesses where a biological agent can be identified. Illnesses are self-limiting, but ~1% of illnesses will require hospitalization. 15% of all foodborne illnesses that require hospitalization are caused by Campylobacter. Mild cases may go away in two to five days. However, severe cases are assumed to take six days in the hospital and three days of at-home recuperation. Guillain-Barre Syndrome (GBS) is a rare but severe autoimmune disease that may follow illnesses with certain pathogens. And, an estimated 40% of GBS cases are triggered by Campylobacter. Approximately 56% of the $1.9 billion economic burden from Campylobacter is actually a result of GBS. GBS occurs in less than 0.25% of Campylobacter cases, but those cases account for over 50% of the economic burden. Campylobacter deaths result in 34% of the burden, and 10% is from both non-hospitalized and hospitalized illnesses.

While Campylobacter may not be as well-known as other foodborne pathogens, the costs of the illness are staggering. Handling or eating raw or undercooked poultry is still a major risk-factor for developing campylobacteriosis. Handwashing and avoiding cross-contamination are the best ways to keep oneself from developing this illness.

 

 

  1. Hoffmann, Sandra, Bryan Maculloch, and Michael Batz. Economic Burden of Major Foodborne Illnesses Acquired in the United States, EIB-140, U.S. Department of Agriculture, Economic Research Service, May 2015.

Campylobacter and WGS: The Review We’ve Been Waiting For

There’s an amazing article out now that describes how whole genome shotgun sequencing (WGS) has contributed to the epidemiology of Campylobacter jejuni, leading cause of gastroenteritis in Europe.  Our friends at barfblog (check them out here) sent this out the other day and it an example of how this revolutionary technology is contributing to the increased understanding and research into C. jejuni.  Even GenomeTrackr (WGS FDA facility- we talk about them in our other post here), is looking into adding some C. jejuni sequences into their database.

The article is a review of how WGS has helped researchers gain a better understanding of the evolution and epidemiology of C. jejuni.  As these high throughput methods become better understood, they are rapidly replacing older molecular methods due to their greater specificity and how easily they they can be shared via open access databases (like MG-RAST, QIIME, GenomeTrackr, etc).  C. jejuni WGS, when used in conjunction with epidemiological methods, has helped identify sources of outbreaks and transmission pathways. WGS has served to majorly advance detection and drastically improve surveillance of C. jejuni infections.

However, Campylobacter is not as well-known as other pathogens such as Listeria and Salmonella (see here and here).  This factor makes it hard to interpret the data that WGS outputs for C. jejuni. We do not currently have a comprehensive understanding or picture of the genetic lineages of C. jejuni, which makes it difficult to determine genetic relationships between strains.  There is also not yet a consensus on what analysis pipeline is best to use or what cut off values for quality should be used within a pipeline.

But these problems are seen in all high throughput methodology (16S, functional metagenomics, etc), and are not necessarily exclusive to C. jejuni WGS.  Despite these obstacles it is exciting to see WGS being applied to C. jejuni outbreak epidemiology and research in general.  It will allow new resolution of C. jejuni genetics and certainly revolutionize the detection and response time for outbreaks.

Read the pre-pub: http://biorxiv.org/content/early/2016/10/01/078550

IFT Reflections: Understanding FSMA and How it Relates to Campylobacter

With the implementation of the Food Safety Modernization Act of 2011 (FSMA), food safety practices are undergoing their largest re-haul since the Food, Drug and Cosmetic Act of 19381.  FSMA has provided much needed updates to food safety practices, an emphasis on traceability, and a large focus on produce.  However, FSMA will change the farming and food industry as a whole.  The poultry industry, where most of the Kathariou Lab’s Campylobacter isolates come from, may also be effected.  To understand how FSMA will affect the poultry industry one has to understand FSMA.  It is also important to keep in mind that meat, poultry, and eggs are covered under the USDA so FSMA (which effects the FDA) will not effect them directly.  However, the effects of FSMA will ripple into all parts of the food industry so these kinds of changes might take place in the USDA several years down the road.

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Why Isn’t Campylobacter Better Known? – By Hannah Bolinger

If you have read What is Campylobacter, you will know that it is a top five foodborne pathogen for illness, hospitalization, and death. With that being the case it is remarkable that Campylobacter is still an relatively unknown pathogen. However, there are a few reasons for this including the self-limiting and sporadic nature of the disease, and under reporting of illnesses.

Campylobacteriosis most often occurs as a sporadic illness i.e. a common source for an outbreak is not noted. This tends to draw less attention from both the media and government agencies than the large multi-state outbreaks that we sometimes hear about.  However, larger outbreaks of Campylobacter can occur and are often the result of unpasteurized milk or untreated water.

Campylobacteriosis is also known as being a relatively minor illness. This is true in the sense that most people will recover from their gastrointestinal discomfort without needing to see a doctor. However, illness can also be extremely acute with the pains sometimes mimicking appendicitis. Campylobacter also has the potential of causing more serious infections in the very young, old, pregnant, or immunocompromised. After a Campylobacter infection there is the potential for autoimmune complications such as Guillain-Barré syndrome, a form of reversible paralysis. Those suffering from Guillain-Barré may take months to recover and may require respiratory support. Another autoimmune complication that can follow a Campylobacter infection is reactive arthritis, characterized by painful joints, eye, and urinary tract problems.

Finally, under reporting is another contributor as to why this disease is not discussed more. Because most healthy adults will recover on their own, it is estimated that only about 1 in 31 cases is reported. Many patients do not seek medical care. Even if a patient does seek medical attention, it is not guaranteed that the doctor will perform a culture-based diagnosis. So, there may not be a definitive conclusion as to what caused the illness. Doctors may treat a patient based on one’s symptoms, and because the symptoms of Campylobacter are nearly indistinguishable from other agents that may cause gastroenteritis e.g., diarrhea and/or cramping, doctors may prescribe antibiotics that may not be optimal for treating campylobacteriosis.

It is likely that Campylobacter infections occur much more often than reported. Because the illness can be mild and of relatively short duration patients do not always seek medical care. However, even if they do it is not guaranteed that a specific etiologic agent will be identified. And, because large outbreaks are rare the media does not give this pathogen the same coverage as others which may cause more severe illness or larger outbreaks. But, because of the high number of illnesses, and the potential for severe infections and autoimmune sequelae Campylobacter should be treated as a much more important foodborne pathogen.

My Experience as an Undergraduate Lab Researcher – By Paige Franek

My name is Paige and I am a senior in Microbiology at North Carolina State University. I have been doing research in Dr. Kathariou’s lab since February of 2016. So far, my experience has been invaluable for my future career, and has made me much more excited about my future in microbiology. It is one thing to take classes and occasionally work in a lab environment, but having my own experiments to work on makes the results that much more rewarding. I enjoy lab classes, but they do not give any freedom to be creative or alter experiments if they are not working. The instructors and students already know what will happen. This is not always the case in a real lab setting.

Doing research in a lab outside of the classroom has given me the opportunity to learn how to make various types of agar, how to use an autoclave, and the correct techniques to keep everything sterile. In lab classes we always have the agar plates already made for us. Other skills I have learned include spot plating, DNA purification, how to use and interpret NanoDrop results, perform PCR, and how to work with Campylobacter and Listeria.

Although working with Campylobacter has been very fascinating, I find it much trickier than other bacteria I have worked with. Experiments do not always go as planned. Because of this I feel much more prepared for a real lab environment in terms of expectations of results, problem solving, and creative thinking. For example, we have performed various transformation experiments with Campylobacter. The goal was to transform 800 base pairs of PCR amplicon from erythromycin resistant strains to susceptible strains. We did this by adding PCR product or genomic DNA from resistant strains directly to susceptible strains. This experiment has been especially interesting because we have been able to transform using genomic DNA, but the PCR fragments have not worked yet.

One of my favorite experiments to date is a competitive fitness assay. This involves the  inoculation of chicken skin fragments with various Campylobacter strains. After counting the resulting plates I then make graphs to quantify the results. This has been one of the first times I have had to make such detailed reports in Excel. So far, we have found that there is not a measurable difference between strain survival over the time points we are measuring. So, our next attempt will be to lengthen the time between measurements.

I would like to thank the amazing people that work in the lab with me, especially Kshipra who has been a fantastic mentor. I came to the lab with minimal experience and not much knowledge on Campylobacter, and was a bit nervous starting out. However, I have learned so many techniques. The lab members happily help whenever I have questions or need some guidance on experiments. I am very happy with my choice to do research in the Kathariou lab and am excited to see what my future holds.

 

What Are Poultry Producers Doing About Campylobacter? – By: Hannah Bolinger, Donna Carver, Sophia Kathariou

Recently, USDA-FSIS released performance standards for Campylobacter and Salmonella requiring producers to limit the presence of these bacteria on raw poultry products1. One of the best ways to meet the Campylobacter standards is to raise flocks free of this organism. However, there are a number of reasons why it is difficult to produce Campylobacter-free birds. Campylobacter can be naturally found in the intestinal tract of birds and is easily spread via fecal contamination. As birds move around a poultry house, they may sample other birds’ droppings. Thus ingesting Campylobacter and becoming carriers. Usually, once one bird becomes colonized with Campylobacter it does not take very long for the rest of the flock to become colonized as well. The litter, feces, communal water and insects (e.g. flies) in a poultry house are all possible routes of transfer among the birds.

Growers use a number of strategies to keep the birds free of Campylobacter and other human pathogens, as well as microbes that could make the birds sick. Rodents are known carriers of Campylobacter so growers keep the areas around the poultry houses free of tall grass and debris.  Doing so eliminates areas where rodents can hide or build nests. Growers have additional pest control measures in place within the poultry houses.   Additionally, flies and other insects can also carry Campylobacter as well as other undesirable microbes so fly bait, insecticides, and litter treatments are used to limit their populations. Growers also wear overalls and plastic overboots so they do not track Campylobacter into the houses from the outside.

As previously mentioned, communal water and litter may play a role in spreading Campylobacter through a flock. To prevent this, the water is generally sanitized via chlorination or acidification to reduce Campylobacter or other pathogens. Campylobacter is known to be a rather fragile organism that does not tolerate oxygen or dry conditions well. Growers till the litter in the poultry houses, which helps to both oxygenate and dry out the litter making it less hospitable to Campylobacter.

At slaughter, there are a number of steps to prevent Campylobacter from being present on the raw, finished product. Externalizing the internal organs for FSIS inspection can result in microbes from the intestine contaminating the carcass. Removal of the intestines can also result in feces (and the bacteria in the feces) to leak onto the carcass.  Any intestinal microbes, including Campylobacter can contaminate the carcass at this point.  Because bacteria are invisible to the naked eye, all birds are potentially contaminated with Campylobacter and from this point on go through a number of decontamination procedures.  To ensure the safety of the product, the carcasses get washed and may be sprayed with organic acids or other compounds that have antimicrobial action. Next, the carcasses are chilled quickly either in ice water or cold air to prevent remaining bacteria from growing and increasing in number.  Chlorine may be added to the chill water to reduce microbial levels,   including Campylobacter.  After this step, the birds are sealed in packages and are shipped to retail.

Even though the government and food industries work together to provide safe food, poultry is still one of the more common vehicles of foodborne illness2. This makes the correct handling of raw poultry a critical step in preventing foodborne illness. You can visit check out the USDA’s tips on preventing cross contamination. By using correct thawing procedures, keeping raw poultry separate from ready-to-eat foods, and washing hands we all  can help decrease the chances of becoming ill.

 

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  1. https://www.federalregister.gov/articles/2016/02/11/2016-02586/new-performance-standards-for-salmonella-and-campylobacter-in-not-ready-to-eat-comminuted-chicken#h-12
  2. Robertson, Kis, et al. “Foodborne Outbreaks Reported to the US Food Safety and Inspection Service, Fiscal Years 2007 through 2012.” Journal of Food Protection®3 (2016): 442-447.

 

 

Campylobacter Biofilms: A Super Resistant Army

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By Kshipra Chandrashekhar

C. jejuni doesn’t show up so much in the headlines due to the self-limiting nature of infection. In developed countries like the United States, incidences of campylobacteriosis are more of an economic burden than a medical liability. Campylobacter is commonly found in the gastrointestinal tract of birds, cattle and the agricultural environment, acting as reservoirs of infection. Although Campylobacter is a fastidious organism with stringent growth requirements such as decreased tolerance to atmospheric oxygen, their ability to survive and thrive in the environment gives rise to the ‘Campylobacter conundrum’ which the poultry industry is trying to tackle. There are several ways by which Campylobacter infection can be controlled. Firstly at the farm level, by preventing the colonization of Campylobacter in poultry through the implementation of strict biosecurity measures. Secondly, by decreasing the bacterial load during handing of chicken in the abattoir and the production facility. To plan effective intervention strategies for Campylobacter control, a better understanding of Campylobacter survival under nutrient poor conditions outside the poultry gastrointestinal tract is crucial.

Biofilm is a community of single or multiple species of bacteria, held together in a slimy complex sugar matrix attached to a surface. Bacterial biofilms are extremely resistant to antimicrobials and disinfectants thus offering a continuous source of infection. Recent studies have shown that C. jejuni were able to persist in poultry slaughterhouses and food manufacturing units. Campylobacter gets attached to surfaces such as the chicken skin and plastic surfaces acting as a source of persistent contamination, thus resisting routine cleaning procedures. The formation of biofilms by Campylobacter has been identified to play a role in its ability to withstand environmental stresses, such as heating, chilling and exposure to chemicals, commonly followed during carcass cleaning. Campylobacter jejuni can form biofilms on abiotic surfaces and is also capable of colonizing pre-existing biofilms. There is increasing evidence that biofilms play an imminent role in transmission of Campylobacter through the food chain. Studies indicate that Campylobacter can better survive atmospheric conditions in biofilms. Surfaces at the meat processing facility and at farms that are prone to biofilm formation include dressing, washing and packaging tables, facility floor, cutting utensils, plastic surfaces of the feeders and watering distribution systems; therefore acting as a source of Campylobacter contamination. Food processing facilities can have the perfect environment for Campylobacter biofilm formation because of organic material build up on surfaces. These organic layers are rich in nutrients containing carbohydrates, lipids and sugars, creating a good surface for bacterial attachment and survival. Studies have also shown that soiling of surfaces with chicken juice positively impacts Campylobacter biofilm formation.

Seen below is a Campylobacter biofilm formed on a plastic surface (Pic courtesy: Dr Louise Salt, IFR, UK)

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Most bacterial biofilms in nature are made up of multiple species. Studies have shown that Campylobacter can form single species (Campylobacter only) as well as mixed species biofilm, which provide a good environment for their survival due to the reduced oxygen levels in such biofilms. Campylobacter is able to form mixed species biofilms with Pseudomonas, which is a chicken carcass spoilage bacteria. Mixed species biofilms also act as a source of nutrient exchange, aiding in Campylobacter survival in such an environment.

These findings from Campylobacter studies epitomize the challenges faced by the poultry industry and food producers. The resilience of Campylobacter biofilms gives them the ability to thrive under adverse conditions. A better understanding of the mechanisms involved in biofilm formation is essential in developing cleaning programs, subsequently decreasing the persistence and transmission of Campylobacter. Strategies to eliminate such reservoirs of Campylobacter during food manufacture will certainly bring a positive impact on our efforts to reduce the incidence and transmission of Campylobacteriosis and other food borne infections throughout the food chain.