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Call Transcript: Cyclosporiasis: Detecting, Investigating, and Preventing Cases and Outbreaks of this Foodborne Parasitic Disease

Moderators:Loretta Jackson Brown

Presenters:Barbara L. Herwaldt, MD, MPH

Date/Time:June 18, 2015 2:00 pm ET

Welcome and thank you for standing by. At this time all participants will be in a listen only mode until the question and answer session of today’s call. At that time you may press star 1 to ask a question from the phone lines. I’d like to inform all parties that today’s call is being recorded. If you have any objections you may disconnect at this time. I’d now like to turn the call over to Ms. Loretta Jackson Brown. Thank you. You may begin.

Loretta Jackson Brown:
Thank you (Crystal). Good afternoon. I’m Loretta Jackson Brown, and I’m representing the Clinician Outreach and Communication Activity - COCA - with the Emergency Communication System at the Centers for Disease Control and Prevention. I’m delighted to welcome you to today’s COCA webinar – Cyclosporiasis: Detecting, Investigating, and Preventing Cases and Outbreaks of this Foodborne Parasitic Disease.

Free continuing education is available for individuals who are participating in today’s webinar. Information on how to complete the activity evaluation and earn continuing education will be provided at the end of the webinar.

CDC, our planners, presenters, and their spouses/partners wish to disclose they have no financial interest or other relationships with the manufacturers of commercial products, suppliers of commercial services, or commercial supporters. Planners have reviewed content to ensure there is no bias. This presentation will not include any discussion of the unlabeled use of a product or products under investigational use, with the exception of discussion on the unlabeled use of the antimicrobial combination trimethoprim-sulfamethoxazole to treat cyclosporiasis.

At the conclusion of today’s session, the participant will be able to discuss what is known about how and where Cyclospora cayetanensis is transmitted; describe the available approaches for diagnosing, treating, and preventing Cyclospora infection, including the limitations of the approaches; explain why timely reporting of Cyclospora cases to public health authorities is essential; and discuss the reported US foodborne outbreaks of cyclosporiasis, including the findings and constraints of the outbreak investigations.

Today’s presenter is Dr. Barbara Herwaldt. Dr. Herwaldt is a medical epidemiologist in the Division of Parasitic Diseases and Malaria, Center for Global Health, at CDC. She earned her doctorate in Medicine from Johns Hopkins University School of Medicine, and a master’s degree in Public Health from Johns Hopkins University School of Hygiene and Public health. She is board certified in Internal Medicine and Infectious Diseases. She completed her fellowship in Infectious Diseases. She joined CDC - excuse me, after she completed her fellowship in Infectious Diseases, she joined CDC’s Parasitic Diseases Branch, initially as an Epidemic Intelligence Service Officer, and thereafter as a medical epidemiologist. Dr. Herwaldt is recognized as a national subject matter expert about cyclosporiasis. She has had central roles in collaborative investigations of this enigmatic parasitic disease over the last two decades.

At the end of this presentation you will have the opportunity to ask the presenter questions on the phone. Dialing star 1 will put you into the queue for questions. You may submit questions through the webinar system at any time during this presentation by selecting the Q&A tab at the top of the webinar screen, and typing in your question.

At this time please welcome Dr. Herwaldt.

Barbara Herwaldt:
Thank you very much. It’s a real privilege to speak to you today.

Cyclosporiasis is an enteric illness caused by Cyclospora cayetanensis, an enigmatic emerging parasite, which was named as recently as the early 1990s and has repeatedly caused US foodborne outbreaks.

And I’d like to focus today on principles and perspective. And I’d like to start with some historical perspective.

At the top of the timeline, you could see that, in retrospect, the first three documented human cases of infection with this parasite are thought to have been diagnosed in the late 1970s by Richard Ashford, a British parasitologist, who was working in Papua New Guinea at that time. But he wasn’t sure how to classify the organism and, therefore, did not name it; and referred to it as an “undescribed coccidian.” And you can see in the left margin my question: “What type of microbe?” And, starting in the mid-1980s, the organism was periodically rediscovered by astute observers, who were actually unaware of Ashford’s publication in 1979. As you can imagine, it’s difficult to search the literature for articles about organisms that haven’t been named. And people variably referred to it as a “big Crypto”—or “big Cryptosporidium-like object”—or a “Cryptosporidium muris-like object”; thought it might resemble a fungal spore or even blue-green alga, like a cyanobacterium-like body (so it’s referred to as “CLB”).

It wasn’t until 1993 and 1994 that Inez Ortega and colleagues confirmed that this organism is indeed a coccidian parasite and proposed the name Cyclospora cayetanensis. The species name cayetanensis was derived from the name of a university in Peru where seminal studies about the parasite had been conducted. In 1995, the results of a placebo-controlled clinical trial in Nepal were published, demonstrating that trimethoprim-sulfamethoxazole is effective therapy.

In the second half of my presentation, I’m going to be focusing on the many foodborne outbreaks that have occurred and been documented in North America and have been linked to various types of fresh produce. Now I initially designed this timeline (or created this timeline) for a publication in the year 2000; and I’ve modified it for today’s purposes, to indicate that we’re now in the year 2015 and Cyclospora is not going away.

So what’s known now about this parasite, and what’s unknown? Well, first of all, again, as I just mentioned, it’s now known that it is indeed a parasite. And it’s a protozoan parasite, meaning, it’s a unicellular parasite. It’s a coccidian parasite; and, therefore, oocysts rather than cysts are excreted in stool.

But it’s still unknown whether other Cyclospora species besides Cyclospora cayetanensis infect humans. In terms of host: again, we know that humans serve as host of this parasite. It’s found in the gastrointestinal tract; both asexual and sexual stages have been documented. But it’s still unknown whether there are any nonhuman hosts of this particular species. There certainly are nonhuman hosts of other Cyclospora species. But, again, we don’t yet know for this particular species.

In the next row of this table: you see a very key biologic point about this organism, and this biologic point affects the epidemiology of the parasite and the parasitic disease. This parasite requires an extrinsic maturation period. The parasite is not infective when it’s shed in stool. It requires days to weeks outside the human host, in the environment, to sporulate and become infective. And the process of sporulation typically requires at least one week under favorable laboratory conditions. But it’s still unknown how quickly the parasite might be able to sporulate under so-called optimal environmental conditions.

In terms of transmission: it certainly can occur via fecally contaminated food or water. But there’s absolutely no risk whatsoever if the person ingests unsporulated, noninfective oocysts, even if the inoculum is huge.

It’s still unknown what accounts for the seasonality of cases. Cyclosporiasis is found in areas that have highly disparate climates, and it’s distinctly seasonal. But the seasonality differs in different areas of the world, and it defies simplistic explanations.

Well, what is sporulation? And why was it necessary to demonstrate sporulation to be able to classify this microbe as a coccidian parasite in the Cyclospora (or the Eimeria) genus? In the upper panel, you see graphic illustrations: on the left, an unsporulated oocyst, with undifferentiated cytoplasm. On the right, a fully sporulated oocyst, which has two internal sporocysts, each of which has two internal sporozoites, for a total of four sporozoites; and the sporozoites actually are the infective units. And you might think: “oh, this is very academic; what difference does this make?”

Well, Ashford correctly had deduced that the organism was a coccidian parasite. But he was not sure how to classify it because he wasn’t able to determine how many sporozoites were present in a sporulated oocyst. And then also notice, as I pointed out, that the sporozoites are within sporocysts and that these sporocysts are within an oocyst wall. And it could be that having the sporocyst walls as well as the oocyst wall contribute in part to the fact that this is a very hardy parasite.

In the lower panel, you see images—photographic images—taken by differential interference contrast microscopy. On the left: again, you see an unsporulated oocyst, with undifferentiated cytoplasm; and, on the right, a fully differentiated oocyst. Now again, a clinical microbiologist is not going to see a fully differentiated oocyst because that takes—that process will take—days to weeks under favorable conditions.

This slide provides some perspective about the relative sizes of microbes and some additional perspective about structural issues. Cyclospora is in the middle. And, of course, in the real world, the sporozoites aren’t nice and bright and pink the way they are here. You can see that the oocyst is about 8 to 10 micrometers in diameter. On the right, you see an oocyst of Cryptosporidium parvum or Cryptosporidium hominis. Notice that it’s a fair bit smaller. It’s also a coccidian parasite, and that’s why I refer to it as an oocyst. And it is fully infective, fully sporulated at the time of excretion in the stool. And it has four naked sporozoites. And, by naked, I mean they’re not within sporocysts. Now both of these coccidian parasites are quite hardy. But we don’t know, as I alluded to before, whether Cyclospora, for example, might be even hardier, given that it has the sporozoites within sporocyst walls, within the oocyst wall.

Now on the far left, you see a cyst—not an oocyst—a cyst of Giardia intestinalis. Giardia is not a coccidian parasite, and it’s shown here simply for perspective. It’s a bit bigger. And, in the lower panel, you see—just for perspective in terms of relative sizes—note the bacteria: quite a bit smaller than these parasites. And then a virus: of course, it’s not drawn to scale because a virus would be even smaller than shown here.

So what about transmission and risk factors? To reiterate, infected people shed an immature form of the parasite in their feces, which has to mature to become infective. Environmental conditions that are conducive to oocyst sporulation and survival are poorly understood—both the macro-environmental conditions and the micro-environmental conditions, for example, on various types of produce.

Direct (versus indirect) person-to-person transmission is highly unlikely because of this need for an extrinsic maturation period. There are two well-established risk factors for US cases of cyclosporiasis, one of which is international travel to cyclosporiasis-endemic regions—for example, in parts of what I’m loosely referring to here as the tropics and the subtropics.

Another well-established risk factor is domestic consumption of contaminated fresh produce imported from such regions. Now, to our knowledge, cyclosporiasis is not endemic in the United States. But we can’t exclude the possibility of focal, low-level endemicity; and we’re definitely keeping an open mind.

In terms of prevention: persons planning to travel to the tropics or the subtropics should be told that food and water precautions for Cyclospora are similar to those for other intestinal pathogens, except that Cyclospora is not likely to be killed by routine chemical disinfection or sanitizing methods—for example, with chlorine or with iodine. The same applies to Cryptosporidium; it’s also quite hardy in this respect. And, not surprisingly, there’s no Cyclospora vaccine available at this time.

So what about the clinical features of symptomatic Cyclospora infection? And I’m focusing now on the US setting, meaning previously unexposed persons—the people who are symptomatically infected. Now the incubation period typically is about a week. It can be a bit shorter, a bit longer. But the median is about a week. And this has been well established through lots of outbreak investigations in situations in which it’s well known when the exposure occurred—for example, a social event.

Now this relatively long incubation period adds to delays in detecting and investigating cases and outbreaks. So this is a biologic delay, which can’t be overcome. We’re hoping to be able to overcome some of the delays in detecting and investigating cases and outbreaks. It’s one of the purposes for this call, for example, this webinar, to heighten awareness, to consider this diagnosis.

And symptoms can begin gradually or abruptly. And the most common symptom is watery diarrhea, which can be profuse. Other common abdominal symptoms include anorexia, weight loss, bloating, gas, cramps, pain, nausea. Of course, all of these are nonspecific, as is watery diarrhea. But I wanted to emphasize that it’s typically watery diarrhea, to make the contrast with pathogens that cause bloody diarrhea. And people also can have body aches. And, if fever is present, it generally is low grade. And these sorts of nonspecific symptoms—the aches and the fever—can be either part of the diarrheal illness or can be noted as part of a prodromal illness before the diarrhea is noted.

And this is a key point: if untreated, the illness may last for days to weeks to months, with remitting-relapsing symptoms—in other words, not necessarily a brief, trivial illness, albeit, fortunately, it’s typically not life threatening. Substantial weight loss and prolonged fatigue commonly occur. Now, of course, there’s a spectrum of severity and duration of illness. And, as I indirectly alluded to, there’s also a spectrum in terms of a range from asymptomatic to symptomatic infection. But, again, among previously unexposed persons in a US setting, particularly in outbreak settings, it’s typically the norm for the people to be symptomatic and, often, to be quite substantially symptomatic—with the caveat, of course, that, typically, we don’t get stool specimens from asymptomatic persons to test, to be able to say anything definitive in that regard.

This slide reiterates some of the points about the symptoms of Cyclospora infection. And these data are from the 1996 outbreak investigation. Overall, although the outbreak was much larger, there were 760 case-patients with laboratory-confirmed infection for whom data were available. Now, actually, the denominator varied for the various symptoms in the list. I want to focus on the fact that the first four that are listed—diarrhea, anorexia, fatigue, and weight loss—all were present in more than 90% of the case-patients. And, in contrast, we go to the bottom of the list and see vomiting: it was present in only about 27% of patients. I don’t mean to trivialize vomiting; I just am highlighting the comparison.

Now, of course, there’s somewhat of a bias in data such as these because these are all symptomatic persons who sought medical attention, whose cases of infection were diagnosed, etc. But, again, it provides some overall perspective that I hope is useful.

And what about the diagnosis of infection? If indicated, if person has clinically compatible illness, consider the diagnosis and request testing. Now, of course, as I already alluded to, the manifestations of the infection are nonspecific. But when I say clinically compatible, you know, one of the features to consider is that, again, this often is not an infection or an illness that, you know, goes away in just a few days—so someone who has persistent or remitting-relapsing diarrheal illness. And diarrhea actually can alternate with constipation.

Another key point is that stool specimens that are examined for ova and parasites usually are not optimally examined for Cyclospora unless such testing is requested explicitly. So this is an important point for clinicians who may think that: “Oh, I’m requesting stool exam for ova and parasites; ah, that will detect Cyclospora if it’s present”—well, not necessarily.

Another key point is that even persons with profuse diarrhea might not shed enough Cyclospora oocysts to be readily detectable. Several specimens from different days may need to be examined, and the lab should use sensitive recovery and detection methods. And, by sensitive recovery methods, I’m referring, for example, to concentration methods. And I’ll be showing you some examples of ways of highlighting the organism—so sensitive detection methods.

And, certainly, CDC and other reference laboratories can provide diagnostic assistance and confirmatory testing using microscopic and molecular techniques—in other words, PCR. And I also want to put in a plug for DPDx. DPD stands for “Division of Parasitic Diseases” and Dx for “Diagnosis.” It’s a diagnostic service. It’s a telediagnostic service, in which clinicians and laboratorians can submit electronic images 24 hours a day, seven days a week, to my laboratory colleagues—for example, if you’re suspecting that you might be looking at Cyclospora oocysts. And they can let you know whether they agree. And, of course, this service is not limited to Cyclospora; it ranges throughout all of the parasites and parasitic infections. But I wanted to let you know about this service.

Another key point is what is not available: so the last bullet on this slide. What’s not available? There are no methods yet for culturing this parasite. In fact, humans are the only known amplifying hosts of this parasite; there’s no means to propagate or maintain the parasite. By maintain, I mean it’s not as if you can maintain the organism in a laboratory animal, with passage through a laboratory animal. Nor, for example, is there a cryopreservation method that will maintain the viability of the parasite. Nor are there molecular tools for strain differentiation, and nor are there serologic assays.

Now this is an example of a sensitive screening method for looking at stool specimens. Cyclospora oocysts are autofluorescent; so this is ultraviolet fluorescence microscopy. And the oocyst jumps out at you. And so it’s a good way of looking at a stool specimen. And I’m oversimplifying here because this is not a talk for laboratorians and nor am I a laboratorian. I do want to add the caveat that a special cube or special filter is needed to be able to demonstrate this blue-green autofluorescence. And so this type of examination is not necessarily routinely done or available in clinical laboratories, though, of course, it does vary by laboratory. Some laboratories do use UV fluorescence microscopy when they’re examining for Cyclospora.

So, this slide shows various means of detecting the parasite. But first I want to emphasize that it’s easy to miss on an unstained slide. Of course, it’s relatively easy if there’s a box around it; but that’s not the real world. So it blends in with the background on an unstained slide. Now it’s relatively easy to spot, as I already mentioned, with UV florescence microscopy. And, in contrast, by the way, Cryptosporidium is not autofluorescent.

Acid-fast staining can be used both for Cyclospora and for Cryptosporidium species. Cyclospora is variably acid fast; so is Cryptosporidium, but Cyclospora is even more variably acid fast. And by that I simply mean that not all oocysts retain the stain. There are some oocysts that are red, and some that are pink, but some that remain just sort of ghostlike.

Now what about treatment of Cyclospora infection? The treatment of choice is trimethoprim-sulfamethoxazole. I already mentioned that the effectiveness of this therapy has been demonstrated in a placebo-controlled clinical trial. This is off-label use: cyclosporiasis is not an FDA-approved indication for this medication. The typical regimen for immunocompetent adults is one double-strength tablet, twice a day, for seven to ten days—sometimes longer. Unfortunately, no highly effective alternatives to trimethoprim-sulfamethoxazole have been identified yet for persons who are allergic to or cannot tolerate trimethoprim-sulfamethoxazole. And, if you encounter such patients, by all means, let us know and we can advise about potential alternative approaches.

The fact that the therapy, trimethoprim-sulfamethoxazole, differs from the medications used for most other enteric protozoan infections—for example, giardiasis, amebiasis, cryptosporidiosis—and also for a fair number of the bacterial infections (though some, of course, respond to trimethoprim-sulfamethoxazole) highlights the importance of diagnosing infection rather than treating empirically.

And now I’m transitioning to the second half of my presentation today. And I want to reorient you in time. I’d like to remind you that these outbreaks—the ones that were detected and initially investigated—occurred right after the organism had been confirmed to be a coccidian parasite, had been named Cyclospora cayetanensis, and trimethoprim-sulfamethoxazole had been shown to be effective. That was great news: that effective therapy had been identified. But the point I’m trying to make here is that in real time—I mean it seems obvious now—but, in real time, it was like being bombarded with outbreaks, initially without a clue about many aspects of the biology of the parasite and the epidemiology of the parasitic disease.

Again, it seems obvious now: but who would have predicted that a protozoan parasite would repeatedly cause large foodborne outbreaks? And, by large, I mean not just the numbers of case-patients, though that also has been and continues to be an issue, but also I’m referring to multistate outbreaks—in other words, exposures occurring in more than one state. And this is not what one typically thinks of with protozoan parasites. Yes, giardiasis, and, yes, cryptosporidiosis have been associated with foodborne outbreaks; but, typically, not on this sort of scale and not typically repeatedly associated with multistate outbreaks.

And the series of spring outbreaks of cyclosporiasis linked to Guatemalan raspberries: this series underscores recurring themes and challenges. And I’m not going to be focusing on individual outbreaks and providing relative risks and P values, etc., etc. I want to underscore recurring themes and challenges, and I’ll do the same thing for some of the recently identified outbreaks.

Now: the beginning—by beginning, I mean from the US perspective. And, of course, we’re very concerned about cyclosporiasis-endemic regions as well. But this talk is focusing on the US perspective and what occurred here. And why we’re focusing on this is because of the need for heightened awareness, to facilitate case detection, outbreak detection, outbreak investigation—to prevent cases and outbreaks, ultimately, not just in the United States, but also in areas of the world where the parasite and the parasitic disease are endemic.

Now initially there were some what we now think were harbinger outbreaks—1995. Of course, I’m simply referring to what’s been recognized and reported and investigated. It occurred in two eastern states: New York and Florida. And then bam: in the spring of 1996, an outbreak in which there were 1,465 reported cases. This includes both laboratory-confirmed and epidemiologically linked cases—both cluster-associated cases (meaning associated with social events, for example) and non-cluster-associated cases (both types of cases were investigated). And cases that were defined as outbreak associated occurred in 20 states, Washington DC, and two provinces. Now the clusters that I referred to involving social events: there were 55 such clusters that were identified and investigated. So think of it as like having to investigate 55 mini-outbreaks. Now it turns out that that can be advantageous, as I’ll be underscoring in some of the slides momentarily, because you can look for commonalities when you’re trying to identify a vehicle of infection and when you’re trying to determine the source of that vehicle.

Now, despite some interim control measures—for example, to improve water quality, etc.—bam: in the spring of 1997, again, another multistate outbreak that was linked to Guatemalan raspberries. Over 1,000 reported cases. There were 41 clusters of cases. Each of those clusters, you know, had many - had a range of cases associated with them. My point being that again these - this total case count includes both cluster-associated cases and non-cluster-associated cases.

And I want to take a momentary aside here and underscore, as I’ve repeatedly been doing, that Cyclospora oocysts, cases, and outbreaks are easily missed. One oocyst, though, one lab tech, one case, one phone call can make a big difference.

Even large, multistate outbreaks can be missed. For example, in 1996, the index cases were detected in two states—New York and Florida—that had laboratory personnel with unusual expertise regarding Cyclospora. By unusual, I mean unusual for that time. It wasn’t just that they had experience. The harbinger outbreaks in 1995: that they had detected those. They truly had unusual levels of expertise. Now, of course, once there’s media attention, then case ascertainment can be facilitated. But in the mid-1990s, it was a big deal to have lab personnel who really already were very knowledgeable about the organism. And, in 1997, several of the index cases and the index clusters were detected because of unusual circumstances—for example, case-patients who were health professionals who knew to consider the diagnosis of Cyclospora infection. Not all health professionals know that even today. But these did, and it made a big difference.

Now even if an outbreak is detected and even if you know to focus on fresh produce, identifying the vehicle of infection can be very difficult. Fresh produce often is served in mixtures and in inconspicuous ways—for example, in garnishes. I mean: Would you even remember this garnish? Would the chef? Would the details be specified on the menu or other records?

And even if that wasn’t an issue—and, frankly, it was an issue for many of the cluster investigations: people didn’t even remember that something had been eaten; and it was only in retrospect, when asking and re-asking, that it came out. But, again, there’s this problem of mixtures; and what do you do? How do you determine what item in a mixture is the pertinent one? And, as I already alluded to, the blessing of having many clusters of cases was that we could look for commonalities among different case clusters and for epidemiologic associations with illness. And, by the way, when I said “we,” I always mean “the collective we.” All the outbreak investigations were collaborative, multiagency, multidisciplinary, and multifaceted.

It turned out, to cut to the chase, that raspberries were the common theme and also were epidemiologically associated with illness.

But even if you can identify a vehicle of infection, traceback investigations for fresh produce often are very challenging—to go all the way back from tables to farms. Turned out that there was a lack of commonalities in the distribution system—for example, US ports of entry. So, to cut to the chase, it was important to get back to the farm level. And, again, because of having so many clusters, it was compelling, aggregate, multi-cluster traceback data that linked the implicated raspberries to Guatemala.

Now I’m not going to talk much about intervention measures, either for Guatemalan raspberries or in general. But just again to sort of orient you in time and to orient you in terms of what I’m going to say on the next slide, I want to give you some perspective—that, on May 30, 1997 (remember: in 1997, there was the second in the series of multistate outbreaks)—so, at FDA’s request, the government of Guatemala and the Guatemalan Berry Commission announced their decision to voluntarily suspend exports of fresh raspberries to the United States. But shipments could resume in mid-August of that year. And, in 1998, as planned—so this was planned in advance—shipments of fresh Guatemalan raspberries were suspended from mid-March through mid-August. So this applied to all farms throughout Guatemala. But this applied only to the United States, not to Canada.

And, in the spring of 1998, there was evidence from an inadvertent intervention trial: Canada, not the United States, had imported Guatemalan raspberries that spring and had a multi-cluster outbreak that was linked to Guatemalan raspberries. So this strengthened the evidence for skeptics who, they’d wondered: “well, you know, the outbreaks had been investigated on the basis of epidemiologic data and traceback data.” This really served to strengthen the evidence.

But it’s very unusual to have implicated produce items still available for Cyclospora testing by the time a vehicle has been implicated.

But, in the spring of the year 2000, evidence regarding Guatemalan raspberries was strengthened even further by detection of Cyclospora DNA in frozen leftovers of an epidemiologically implicated food item. I like to say that brides are blessings: a bride froze some leftovers of her wedding case. This wedding cake had been epidemiologically implicated. It had raspberry pieces in a cream filling, and it tested positive in Cyclospora PCR analyses at CDC and FDA.

And I want to make several points here. One is that the year 2000 was the last time that Guatemalan raspberries per se were implicated in an outbreak investigation. Raspberries from other places have been suspected in other years. And, as I’ll allude to momentarily, Guatemalan snow peas were implicated in the year 2004.

And I just want to briefly mention, without going into details, because the intervention measures, etc., have been quite complex: but it turned out that very few Guatemalan farms had been allowed to export (or we didn’t take their, you know, we didn’t import, if taking it from that perspective) in 1999 and the year 2000. And there was a farm that was implicated, if you take the aggregate evidence from this cluster—the wedding-cake cluster in Pennsylvania—with another cluster of cases in another state: there was one Guatemalan farm that was in common to those two clusters. And that farm was no longer allowed to export (in other words, we did not accept the imports).

I also want to mention that the spring harvest season for Guatemalan raspberries corresponds to the seasonality of cases of cyclosporiasis in Guatemala. And cryptosporidiosis is much less seasonal in Guatemala.

Now the routes of contamination of the raspberries remain unknown: unproven hypothesis—it’s via contaminated water. Turns out that drip irrigation is used or was used on most of these farms—on the ground, not near the berries—whereas pesticides, fungicides, insecticides, etc., were mixed in water and were sprayed on the raspberries.

It turned out that a number of these farms were investigated after the 1996 and other outbreaks. And water sources were found to be vulnerable to contamination. Now this is an unproven hypothesis; but, for various reasons, it seems plausible. And, of note, no one farm could have accounted for the 1996 or the 1997 outbreak, or for both in aggregate. And no one shipment. And, for example, the 1996 outbreak lasted more than a month. And, by lasted, I mean, you know, exposure dates and shipment dates, etc.

Well: not just raspberries and not just Guatemala; not just springtime, and not just one vehicle per year, and not just one source per vehicle. There have been many permutations and combinations of types of produce and sources. And, when I say many, I don’t mean to exaggerate. But I want to underscore the point that it’s not just been a series of outbreaks linked to Guatemalan raspberries.

1997, for example, there was a series—a documented series of five outbreaks of cyclosporiasis—and they’re listed here by month of exposure (for example, of social events). And second bullet you’ll see relates to the raspberries from Guatemala, as I’ve already mentioned. But right before that—March and April—there was a multi-cluster outbreak in Florida. And if we take the evidence from that outbreak, as well as from the last bullet (you see: December Florida?), those outbreaks likely were linked to mesclun lettuce from Peru. And, then, if you look at the third bullet—June and July; the Washington DC, Baltimore metro area—a multi-cluster outbreak linked to fresh basil, which had multiple possible sources. And it wasn’t—we weren’t able to figure out which of the sources was pertinent.

In September—this is of interest—in Virginia, a fruit plate was implicated; but it wasn’t possible to determine which ingredient. There was only one cluster of cases identified; there were no opportunities to triangulate. But it definitely was not linked to Guatemalan raspberries.

Now the overall series of US foodborne outbreaks of cyclosporiasis includes outbreaks I’ve already mentioned: in the mid-90s, the 1996 and 1997 multistate outbreaks linked to Guatemalan raspberries. Also, already mentioned: outbreaks linked to mesclun lettuce and basil.

Outbreaks have been detected almost every year thereafter. Almost assuredly there have been outbreaks that have been missed. And, even for those that have been detected, food vehicles and sources are not always identified. But examples include: in the year 2000, as I already mentioned; Guatemalan raspberries; also, as I briefly alluded to, Guatemalan snow peas in the year 2004.

And basil: this is not the basil outbreak back in 1997. These are subsequent basil-associated outbreaks. Basil from Peru was implicated. But it turns out there was a connection with 1997 after all because the mesclun lettuce in 1997 that came from Peru came from the same firm. And then also more recently, as I’ll talk about momentarily, cilantro from Mexico.

Now, to date, no commercially frozen produce has been implicated. And the routes of contamination of the produce have not been definitively established, though some outbreak investigations have led to interventions and prevention measures.

So briefly skipping forward now to the very recent past: 2013 and 2014.

2013, US outbreaks of cyclosporiasis: in late June 2013, public health officials in Iowa and Nebraska began receiving reports of laboratory-confirmed cases not associated with international travel during the previous two weeks. Overall, a total of 631 such cases not associated with international travel, with onset dates during June to August, were reported by 25 states. But the bulk of the cases—79%—were reported from three states: Texas, Iowa, and Nebraska.

This map shows the distribution of these 631 cases by state of residence of the case-patients. And you can see in dark blue: Texas reported 270 cases; Iowa, 140. And you can see in royal blue that Nebraska reported 87. Again, these are all in patients with illness onset during June to August.

This epi curve shows: time along the X-axis from June to August; number of cases or case-patients along the Y-axis. And then there’s three line graphs. One is a composite for Iowa plus Nebraska. And you can see that the peak is in mid-to-late June. Texas: notice that this line graph is more dispersed, and the peak is somewhat later in time. And then you can see in red a line for the other states.

Well, it turns out that these recent outbreaks in 2013—and, as I’ll briefly allude to, in 2014—underscore, as did the earlier outbreaks I mentioned, recurring themes and challenges. So, in 2013, at least two distinct outbreaks occurred, linked by epidemiologic and traceback data to different types of fresh produce from different parts of Mexico.

First of all, for Iowa and Nebraska, the subset of restaurant associated cases—and, by restaurant associated, I mean multiple locations of two chain restaurants that had the same parent company: these cases were linked to a bagged salad mix—a prepackaged mix that had multiple ingredients—from Guanajuato, Mexico. But what was the pertinent ingredient in the mix? As for Texas: some case clusters were associated with cilantro from Puebla, Mexico. (Again, these are different parts of Mexico: one is north of Mexico City, and the other is south of Mexico City.) In 2014, cases in another cyclosporiasis outbreak in Texas were linked to cilantro from Puebla, which served to strengthen the evidence.

But the bulk of the reported domestically acquired cases of cyclosporiasis in 2013 and 2014 were not directly linked to food vehicles, which underscores the need for molecular tools. We don’t yet have molecular tools that can distinguish among strains of the parasite. Development of such tools is a high priority for CDC, for FDA, and other agencies. And availability of such tools in the future could help public health investigators determine in general—and determine more rapidly—if cases are linked to each other, and if cases are linked to particular food items, and if those food items are from particular sources.

I’d also like to put in a plug for surveillance. Cyclosporiasis is a nationally notifiable disease. It has been such since January 1999. It was designated a nationally notifiable disease in the context of the large foodborne outbreaks in the mid-1990s. Prompt reporting and investigations of cases and outbreaks are essential. The fastest surveillance system is the phone. I’d like to put in a plug for telephone calls for real-time updates. And, in general, I’d like to underscore the ongoing needs for vigilant surveillance, for molecular subtyping tools for strain differentiation, and for targeted prevention strategies.

So in summary, as discussed: consider the diagnosis of Cyclospora infection, particularly for persons with persistent or remitting-relapsing diarrheal illness—regardless of whether they have a history of international travel and regardless of the time of year, but especially during spring and summer months: in other words, now. As we speak, cases of cyclosporiasis are being detected and reported and investigated. So it’s really important to be detecting cases. This is important both for individual patients, so that they get appropriate therapy, and because of the public health implications.

So, if indicated, explicitly request laboratory testing for this parasite per se, and promptly report confirmed and suspected cases to public health authorities. Even seemingly isolated cases could be part of outbreaks. So the need for prompt reporting should be self-evident: but it’s to expedite identification of food vehicles that cause outbreaks and help prevent ongoing and future transmission.

So I’ll just close with this philosophical slide. And I don’t think it needs any commentary. And I’ll turn it back to Loretta now. So thank you very much for your attention.

Loretta Jackson Brown:
Thank you so much Dr. Herwaldt for providing the COCA audience with such a wealth of information. We’ll now open up the lines for the question and answer session. And remember you can submit questions through the webinar system at any time.


Thank you. To ask a question from the phones lines please press star 1. You will be prompted to unmute your phone and record your name. Again it’s star 1 to ask a question. One moment please for the first question.

Loretta Jackson Brown:
And so Dr. Herwaldt, while we’re waiting for the first question, I have a question for you. Has the infectious dose of this parasite been determined?

Barbara Herwaldt:
Good question. It’s not been confirmed yet by human volunteer studies. In fact, there have been attempts to infect human volunteers, and those attempts have failed. But the infectious dose is presumed to be relatively low. And that’s on the basis of data for other protozoa and also from outbreak investigations. For example, even seemingly trivial exposures, such as one berry or one bite, have resulted in infection. For example, there was a lemon tart garnished by one raspberry that was associated with a cluster of cases. And that suggests that either the infectious dose is low and/or that the number of oocysts on that berry was high or that both might have been true.

Loretta Jackson Brown:
Okay, thank you. Operator, do we have a question on the phone?

Yes. The first question comes from Dr. Robert Ball. Sir, your line is open.

Robert Ball:
Thank you very much. And Barbara, thank you for a splendid presentation. I recall working with you in ’96 when we investigated that raspberry-associated cluster here in Charleston reported in MMWR. My question is twofold. Number one—in the prolong traceback investigation back to Guatemala—number one: how many federal agencies were involved to convince Guatemala to allow us in? And number two: how long did it take? As I recall, it took quite a while for our team to get down there. Thank you.

Barbara Herwaldt:
Well it’s good to hear your voice; and I remember you very, very well. And you’ve always been a great collaborator and a great resource. Of course, there was some variability from investigation to investigation in terms of the details, the particulars. But the Food and Drug Administration established a very close working relationship with Guatemala, which lasted a number of years. They worked very closely with the Guatemalans to improve growing conditions and conditions for the workers on farms that had ramifications beyond Guatemalan raspberries. But you’re absolutely right that particularly in 1996 it took a while to figure out that Guatemalan raspberries—well, that Guatemala was the source—because at the same time there were Chilean raspberries and raspberries from Guatemala, etc. And I’m happy, though, to talk with you in more detail offline, if you’d like, because I realize your question, which is excellent, is multifaceted and complex.

Robert Ball:
Thank you again. Look forward to the slides and further communication. Peace and blessings.

Barbara Herwaldt:
Thanks so much.

Thank you. And our next question comes from Peggy Carter. Your line is open.

Peggy Carter:
Yes, hi. That was a fantastic presentation. I really enjoyed it. And I’d like to know whether or not there is any other parasite that commonly accompanies Cyclospora when you’re doing your exams to look for parasites.

Barbara Herwaldt:
That’s an excellent question, and, to my knowledge, no. And the reason I’m talking with this tone of voice is because it is such an intriguing issue. It’s also an intriguing issue like, you know, accompanying bacteria. Why haven’t we had outbreaks in which at the same time that particular produce is contaminated with Cyclospora, it’s contaminated with not only a different type of parasite, as you’re alluding to, but, you know, with bacteria. And I’m oversimplifying a bit here. Well, first of all, our knowledge is very limited; and our knowledge about particular pathogen-produce combinations is very limited. And, also, there could be limitations in what’s detected. But, to cut to the chase, your question is very good. And I wish that I could answer it. It’s an example of one of the many gaps in our knowledge.

Peggy Carter:
Thank you.

Barbara Herwaldt:
I also should allude to the fact that, you know, there could be variabilities in certain settings in terms of what’s endemic or, you know, what may be contaminating the water, if indeed the water is what is contaminating the produce. And then also there may be certain types of microbes that are killed off or otherwise removed by certain sorts of processing. So there would be those complexities as well.

Thank you. And our next question comes from Alex da Silva. Sir, your line is open.

Alex da Silva:
Oh hi Barbara - very nice presentation. It’s kind of a question but, you know, I’ll start with a comment very quick. When you said that there was no molecular testing, well there is some good news there. There is a new FilmArray called BioFire, which is - has been cleared by the FDA to be used in diagnostic laboratories. And it detects about 22 pathogens, including Cyclospora, Giardia, Entamoeba, and Cryptosporidium. So do you think that, you know, with that new trend – and actually, you know, became widespread; there was a lot of labs acquiring it. Do you think that, you know, with that new capabilities, we’re going to be seeing more cases? And then what do you think will be the impact of that?

Barbara Herwaldt:
Alex, it’s good to hear from you. And I’m glad you raised that point. I also want to clarify. I hope people did not misunderstand. I did not mean to imply that there are no molecular tools. Certainly, there’s PCR available. The distinction I was making was that there are no molecular tools available yet for strain differentiation. But your point is excellent because I did not raise the fact that there are these sorts of panels. And you’re an expert about this, as a molecular biologist—BioFire: that can detect multiple pathogens. And we have had, as you may know, the collective we, have had some cases detected in this way. And you’re absolutely right that it may end up being a way of facilitating detection. Now I can’t personally comment about this particular test—that, you’d be in a better position to do so. But the general principle certainly is a good one. Thank you.

Alex da Silva:
Thank you.

Thank you.

Loretta Jackson Brown:
Operator, do we have any more questions on the phone?


No ma’am. There are no other questions in queue at this time.

Loretta Jackson Brown:
Okay. Well Dr. Herwaldt, I have a question through our webinar system. And hopefully you can maybe tie two together. So when we think about Cyclospora and ways to remove it, you talked a lot about produce. So if produce is contaminated, can Cyclospora be removed, thoroughly, with washing? And also a participant is interested in knowing at what temperature is needed to kill the sporulated oocyst.

Barbara Herwaldt:
Good questions. As always, fresh produce should be thoroughly washed before it’s eaten, which should reduce, but may not eliminate, the risk for infection. And a fair number of the outbreaks have been associated with produce that reportedly had either been washed or rinsed. In the laboratory setting, lettuce has been experimentally inoculated, so to speak, with oocysts and washed and then oocysts could still be demonstrated by electron microscopy. So, by all means, wash produce. But it’s not a way for totally eliminating the parasite.

Now, in terms of temperature, we think that, for example, boiling—and, of course, boiling per se doesn’t apply to some types of food items—but thorough cooking or boiling should kill the parasite. But I do want to underscore that we don’t have ways to directly assess the viability of the parasite. Whether a parasite sporulates is an indirect way of assessing viability and whether excystation of the infective units (you know, sporozoites) occurs is an indirect way.

But, again, we don’t have a direct way of assessing viability. But the bottom line is that thorough cooking—particularly, for example, boiling—should, we think, should kill the parasite. And, in terms of freezing, for what it’s worth, none of the detected outbreaks have been associated with commercially frozen produce.

Loretta Jackson Brown:
Well thank you. It looks like we are out of time. So on behalf of COCA, I would like to thank everyone for joining us today, but a special thank you to our presenter, Dr. Herwaldt.

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