Heterosexual man: Development of a phage cocktail to control the top 7 shiga -toxin E. coli serogroups on fresh produce

Sorenne is afraid of bugs.

I told her the other day how Surgeoner made his students (and kids) stick their arms into boxes filled with mosquitoes – mossies as they’re called here – to see what dose of DEET worked, while the kids got bitten.

She became more terrified.

That’s OK, I have lots of parenting failures.

As the consumption of fresh produce increases worldwide, Shiga-toxigenic Escherichia coli (STEC) outbreaks linked to contaminated produce are becoming more frequent. Biocontrol of STEC using phages can be a safe and effective way to reduce STEC on fruits and vegetables.

Purpose: 1) Evaluate the effectiveness of 7 bacteriophages to reduce viability of E. coli serogroups O26, O45, O103, O111, O121, O145, and O157 on lettuce and sprouts at 4, 10 or 25 °C and storage for 1, 24, 48, and 72 h; 2) to assess STEC phage effectiveness to reduce STEC compared to a conventional chlorinated water wash (150ppm for fresh produce, 1000 ppm for seeds); and 3) to determine if phage-insensitive mutants arise after phage treatments. Methods: 1) Phages either individually or as a cocktail (~108 log PFU/ml) were sprayed to control 105 log CFU/ml STEC spot-inoculated onto fresh lettuce and sprouts that were stored for 1, 24, 48, and 72 h at 2, 10, and 25°C. Samples were collected and bacteria enumerated on McConkey (STEC total numbers) and Rainbow agar (STEC individual serogroup numbers) after each storage period. STEC isolates were confirmed using latex agglutination tests specific for STEC, PCR, and Immunomagnetic Separation (IMS) to recover any surviving STEC serogroups. 2) A scale-up experiment with STEC phage cocktail and chlorinated water applied to lettuce, sprouts, and mung bean seeds (MB) was undertaken to further assess the effectiveness of STEC phage cocktail to reduce STEC on larger quantities of produce and to compare its effectiveness to chemical disinfection. Lettuce, sprouts, and MB were treated with: i) chlorinated water wash (150 ppm for lettuce/sprouts, 1000 ppm for MB); ii) STEC phage cocktail; and iii) a combination of chlorinated water and STEC phage cocktail. Phage cocktails were delivered by immersion. After treatment, lettuce and sprouts were stored for 1, 24, 48, and 72 h at 2, 10, and 25 °C. MB were stored in the dark for 24 h at 25 °C. Seeds were germinated in sterilized water and the survival of STEC isolates was assessed by plating on Rainbow agar. 3) To determine if phage resistant mutants developed during STEC phage treatment, isolated colonies surviving after exposure to phage were randomly picked and tested for phage sensitivity using a microplate virulence assay. OD was measured after microplate incubation for 5 h at 37 °C. Results: 1) In spot-inoculation experiments, the highest STEC reduction (3.7 log10 CFU/g) caused by the phage cocktail was observed at 2 °C after 72 h of storage on lettuce; whereas on sprouts the highest reduction (2.45 log10 CFU/g) occurred at 25 °C after samples were stored for 1 h. All STEC O157, O26, and O103 were killed by spraying phages on both lettuce and sprouts. Overall STEC phages reduced STEC by > 2 log10 CFU/g on fresh produce. 2) During scale-up experiments, the combination of STEC phage cocktail and chlorinated water achieved the highest STEC reductions on produce. On MB, the highest reduction in STEC (1.69 log10 CFU/g) was observed after treatment with the STEC phage cocktail alone. On MB germinated sprouts, the reduction in total STEC was < 1 log10 CFU/g in all treatments. However, none of the treatments eliminated all 7 STEC serogroups. 3) Regarding phage resistance experiments, no STEC mutants were recovered. Conclusion: Results showed that the phage cocktail was able to reduce STEC O26, O45, O103, O111, O121, O145, and O157 serogroups alone, or in combination with chlorinated water on lettuce, sprouts, and MB seeds without the development of phage resistant mutants. However, for MB, the STEC phage cocktail was not effective at controlling STEC populations during germination.

Raw is still risky: Say no to raw dough

My mother used to make and lot of cakes and brownies with her groovy 1960s hand mixer and I always got to lick the beaters.

No more.

And it’s not just the raw eggs, it’s the raw flour.

In June, 2009, an outbreak of shiga-toxin producing E. coli (STEC, primarily O157:H7) in Nestle Toll House cookie dough sickened at least 77 people in 30 American states. Thirty-five people were hospitalized – from cookie dough.

The researchers could not conclusively implicate flour as the E. coli source, but it remains the prime suspect. They pointed out that a single purchase of contaminated flour might have been used to manufacture multiple lots and varieties of dough over a period of time as suggested by the use-by dates on the contaminated product.

The study authors concluded that “foods containing raw flour should be considered as possible vehicles of infection of future outbreaks of STEC.”

So it wasn’t much of a surprise when 63 people fell sick from the outbreak strain of E. coli O121 from Dec. 2015 to Sept. 2016 linked to raw General Mills flour.

There have been about a dozen other flour-related outbreaks. STEC means people – and kids – get quite sick.

Flour is a raw commodity, crops the flour is derived from could be exposed to anything, and testing is so much better than it used to be.

There are some brands of pasteurized flour out there, but people seem to have gotten used to flour as a cheap source of play-dough-like stuff for kids and something to throw at people.

The U.S. Centres for Disease Control says, nope.

This is not a Christmas conspiracy (although I prefer Solstice Season): it’s CDC providing information, like they are supposed to.

People can, and will, do what they want.

As Maggie Fox of NBC reports, “Do not taste or eat any raw dough or batter, whether for cookies, tortillas, pizza, biscuits, pancakes, or crafts made with raw flour, such as homemade play dough or holiday ornaments,” the CDC advises.

“Do not let children play with or eat raw dough, including dough for crafts.”

Handling food, including flour, requires care and hygiene.

“Keep raw foods such as flour or eggs separate from ready-to eat-foods. Because flour is a powder, it can spread easily,” the CDC notes. “Follow label directions to refrigerate products containing raw dough or eggs until they are cooked. Clean up thoroughly after handling flour, eggs, or raw dough.”

Shiga-toxin E. coli at UK nursery sparks NHS probe

PA reveals a Perthshire school was involved in an E.coli scare last week.

NHS Tayside launched an investigation after a suspected case of the bacteria in a child at Errol Primary School’s nursery.

The nursery will undergo three days of deep cleaning as a “precautionary measure”.

Parents at the school were issued with letters from the health board with information on the infection.

The child was being tested for a non-O157 strain.

Speaking on Friday a spokesperson at NHS Tayside confirmed: “NHS Tayside’s health protection team is aware of and currently investigating a single suspected case of E. coli non O157 infection in a child who attends a nursery in Perthshire.

“As a precaution, a letter has been issued to parents of children at the nursery for information and reassurance.

“The risk to the wider public is very low.”

Raw is risky, Mods vs. Rockers: Apple cider from Virginia farm market recalled

Over two decades later and food safety types still have to deal with this shit – literally, because it’s shit in the cider. Don’t eat poop, and if you do, cook it.

Mountain Man Market of Cana has recalled their half gallon containers of apple cider because it has the potential to be contaminated with shiga-toxin producing E. coli.

The apple cider was sold at Mountain Man Market on Fancy Gap Road on and before November 10.

Their cider hasn’t been pasteurized, which means it can contain harmful bacteria.

The Virginia Department of Agriculture and Consumer Services found the potential contamination after routine testing, and the Division of Consolidated Laboratories (DCLS) found shiga-toxin producing E. coli. in the cider.

VDCAS and Mountain Man Market say they will continue investigating into how the apple cider got contaminated in the first place.

(Quadrophenia is so much better than Tommy and a modern masterpiece)

5 sick raw is risky: Unpasteurized cheese made on Vancouver Island recalled for E. coli concerns

A voluntary recall has been issued for a Vancouver Island-produced cheese linked to an E. coli outbreak that infected five people.

Little Qualicum Cheeseworks’ Qualicum Spice cheese should be discarded or returned to the place of purchase, according to the BC Centre for Disease Control.

The E. coli outbreak occurred between August and October and was traced back to the unpasteurized product, which has a best before date up to and including April 24, 2019.

An investigation is ongoing “to determine the source and extent of contamination,” the BCCDC said.

Ya can’t stop what’s coming: E. coli happens in France too

Enterohemorrhagic Escherichia coli serogroup O80, involved in hemolytic uremic syndrome associated with extraintestinal infections, has emerged in France. We obtained circularized sequences of the O80 strain RDEx444, responsible for hemolytic uremic syndrome with bacteremia, and noncircularized sequences of 35 O80 E. coli isolated from humans and animals in Europe with or without Shiga toxin genes.

RDEx444 harbored a mosaic plasmid, pR444_A, combining extraintestinal virulence determinants and a multidrug resistance–encoding island. All strains belonged to clonal complex 165, which is distantly related to other major enterohemorrhagic E. coli lineages. All stx-positive strains contained eae-ξ, ehxA, and genes characteristic of pR444_A.

Among stx-negative strains, 1 produced extended-spectrum β-lactamase, 1 harbored the colistin-resistance gene mcr1, and 2 possessed genes characteristic of enteropathogenic and pyelonephritis E. coli. Because O80–clonal complex 165 strains can integrate intestinal and extraintestinal virulence factors in combination with diverse drug-resistance genes, they constitute dangerous and versatile multidrug-resistant pathogens.

Emerging Multidrug-Resistant Hybrid Pathotype Shiga Toxin–Producing Escherichia coli O80 and Related Strains of Clonal Complex 165, Europe

Cointe A, Birgy A, Mariani-Kurkdjian P, Liguori S, Courroux C, Blanco J, et al. Emerging multidrug-resistant hybrid pathotype Shiga toxin–producing Escherichia coli O80 and related strains of clonal complex 165, Europe. Emerg Infect Dis. 2018 Dec [date cited]. https://doi.org/10.3201/eid2412.180272

https://wwwnc.cdc.gov/eid/article/24/12/18-0272_article

EHEC in Germany: Cheese recalled

(Something may be lost in translation, which is where Pink Floyd comes in)

This was informed by the Federal office for consumer protection and food safety in Braunschweig, Germany, on Thursday.

It is the in several States product, marketed with the name “Petite Fleur herbs” (“Keiems bloempje met kruiden”) the date of minimum durability 08.11.2018 and the additional indication of “Lot 3603”, as a manufacturer Dischhof/Belgium has been specified.

In the worst case, kidney failure threatening, According to the Schwabacher company in an investigation of the image Verotoxin found E. coli in the cheeses, which are regarded as potential Ehec.

The editor recommends:

“From a consumption of the affected product is strictly not recommended”, the company said.

The Ehec can cause bloody diarrhoea and in severe cases lead to kidney failure, but there are also inconspicuous progressions. The severe course of a disease ends in about two percent of the cases fatal.

In the spring of 2011 had cost the largest German Ehec epidemic 53. In Germany, the disease occurs again and again, every year, about 1000 cases of Ehec are reported cases.

In the PCP Luke (15) died because no one recognized the symptoms – now, his mother warns all.

And this was the album of my grade 12.

Lettuce is overrated: STEC in Finland

Escherichia coli are Gram-negative rod-shaped bacteria and part of the normal bacterial flora in the gastrointestinal tract, while diarrhoeagenic E. colipathotypes such as Shiga toxin-producing E. coli (STEC) and enteropathogenic E. coli (EPEC) are able to cause gastrointestinal infections [1]. STEC can lead to a severe disease, such as haemolytic-uraemic syndrome (HUS) [2]. The risk of HUS has been related especially to children under 5 years and to elderly people. HUS is characterised by acute onset of microangiopathic haemolytic anaemia, renal injury and low platelet count.

More than 400 STEC serotypes have been recognised, of which the best-known serotype is O157:H7 [1]. The most common non-O157:H7 serotypes causing human infections are O26, O103, O111 and O145 [3]. The virulence of STEC is largely based on the production of Shiga toxin 1 or 2 and is identified by detecting the presence of stx1 or stx2 genes [1,4]. The virulence of EPEC is caused by its capability to form attaching and effacing (A/E) lesions in the small intestine. This capability requires the presence of virulence genes called the locus of enterocyte effacement (LEE) in a pathogenity island (PAI) that encodes intimin [4]. Unlike STEC, EPEC do not produce Shiga toxin. EPEC are divided into two distinct groups by the presence of EPEC adherence factor plasmid (pEAF) expressing bundle-forming pili (BFP), which is a virulence determinant of typical EPEC (tEPEC) [5]. Thus atypical EPEC (aEPEC) are defined as E. coli that produce A/E lesions but do not express BFP. Typical EPEC are best known as a cause of infantile diarrhoea, especially in developing countries [6]. Diarrhoea-causing aEPEC have been shown to be separate group without a close relation to tEPEC, but some serotypes are genetically related to STEC [5]. The pathogenity of aEPEC has been questioned but their involvement with diarrhoeal outbreaks supports the idea that certain strains are diarrhoeagenic [1,7].

Both STEC and EPEC are transmitted through the faecal-oral route, and outbreaks caused by STEC and aEPEC have been described after ingestion of contaminated food or water [7,8]. STEC is common in ruminants and can be found in foods contaminated by ruminant faeces [9]. Most studies on STEC have focused on the serotype O157:H7, but infections and outbreaks caused by non-O157 strains are increasingly reported in Europe and elsewhere [1013]. Atypical EPEC strains are found in animals used for food production, such as cattle, sheep, goat, pig and poultry, in contrast to tEPEC that has been found only in humans [1,14].

Since 1995, clinicians and clinical microbiology laboratories have been obliged to report culture-confirmed STEC infections to the Finnish Infectious Disease Registry (FIDR) maintained by the National Institute for Health and Welfare (THL) in Finland. EPEC infections are not reportable. Since PCR instead of culture became the standard for screening of diarrhoeal patients in 2013, the incidence of reported STEC infections has increased in Finland to 1.2–1.8 per 100,000 population between 2013 and 2015 compared with 0.2–0.6 per 100,000 between 2000 and 2012. From 1997 to 2015, six food- or waterborne STEC outbreaks were detected in Finland (Table 1).

Outbreak of multiple strains of non-O157 Shiga toxin-producing and enteropathogenic Escherichia coli associated with rocket salad, Finland, autumn 2016

15.may.18

Eurosurvelliance, Volume 23, Issue 35, https://doi.org/10.2807/1560-7917.ES.2018.23.35.1700666

Sohvi KinnulaKaisa HemminkiHannele KotilainenEeva RuotsalainenEveliina Tarkka,Saara SalmenlinnaSaija HallanvuoElina LeinonenOllgren JukkaRuska Rimhanen-Finne

https://www.eurosurveillance.org/content/10.2807/1560-7917.ES.2018.23.35.170066

UK disease detectives: Supermarket loyalty cards to trace an E coli outbreak

Kathie Grant and Lisa Byrne write in Public Health Matters that in November 2017, supermarket loyalty cards were used to trace the source of a large E coli outbreak affecting mainly men in England. Dr Lisa Byrne leads Public Health England’s surveillance of two key bacteria that lead to food poisoning – E Coli and Listeria. Dr Kathie Grant heads the PHE Gastrointestinal Bacteria Reference Laboratory. The two work together as part of a larger team dedicated to reducing foodborne illness and below tell us how they put the pieces of this puzzle together to find the source.

If you’ve ever had food poisoning you’ll know that feeling of mentally going through everything you ate recently, trying to pinpoint what it was that might have made you ill. It’s our job to do that at a national scale.  We bring together lots of different pieces of  information from the community and the lab to try to find  the source of a food poisoning outbreak and then, working alongside other government agencies,  ensure that more people don’t get sick.

We study and monitor many different stomach bugs – some of which you may never have heard of! While stomach bugs are a part of life, PHE works with organisations such as the Food Standards Agency and the Animal and Plant Health Agency to try and prevent them.

Every so often we see a spike in the number of cases.  When this happens it is important that we find the link between the cases and the cause of their illness.  To do this we need to identify the exact strain of a bug to understand if people have got ill from the exact same source.

Whole Genome Sequencing (WGS) ‒ a relatively new process for showing us the makeup of a bacterium or virus’s genes ‒ has changed the way we can find the cause of an outbreak and stop more people getting ill. You can learn more about the process and how it works in our explainer blog.

Before WGS it could take weeks to identify bacteria and sometimes the bacteria could be missed. This slowed down any investigations  as we could not be sure that all the case histories we were taking could be linked to an outbreak – there was a lot of ‘noise’ and false lines of enquiry. With WGS, we can rapidly and accurately identify if bacteria of cases are the same strain and rule out people from our investigation who just happened to be ill at the same time, but with a different illness.

It has also expanded what the word ‘outbreak’ means as we can link cases across several years and different countries, meaning we can more accurately piece together a picture of how something in the food supply chain impacts human health.

Scientists working in the Gastrointestinal Bacteria reference laboratory at Public Health England. The team are processing samples from people who have reported gastrointestinal symptoms, to understand the exact cause of their illness.

Identifying the source of an outbreak is a lot like putting together a jigsaw puzzle, combining multiple pieces of evidence to get the full picture. Sometimes, a common source is obvious, such as when a group of people get ill after eating the same meal, at the same restaurant, on the same day. But other times, we need to use an arsenal of investigative tools, as was the case in a recent E coli outbreak.

In November 2017 our surveillance system alerted us to 12 cases of E coli O157 – (a particular form of E coli), over a six week window. E coliO157 is a relatively rare cause of food poisoning, with only about 700 cases a year, but it can cause a very severe illness. Because of this, any case of E coli O157 identified by doctors and laboratories must be reported to Public Health England.  We monitor the number of cases with our surveillance systems to find any patterns.

Very quickly our reference laboratory used WGS which showed that the cases had the identical genetic “fingerprint” and the work began to trace the source of infection. The majority of people who became ill were men, which was unusual as E coli outbreaks are often linked to salad items ‒ traditionally more likely to be eaten by women.

It took a few rounds of interviews – carried out by colleagues in local authorities – to zero in on the potential source of food poisoning, and a picture started to emerge that implicated burgers from a particular retailer.

We asked the supermarket to analyse the loyalty card records of those who had become ill, to help identify the particular burger product the cases had eaten.  As you can imagine, there were many different types of burgers supplied by the supermarket and it’s often difficult for people to remember exactly what they ate.

Working with the Food Standards Agency we were able to identify that all the cases had bought a particular brand of burger, leading to a product recall to ensure others didn’t get sick. The recall involved removing all the suspected batches of burgers from the supermarket shelves. The supermarket also contacted people who had bought the burgers, advising them not to eat them and return them for a refund.

Sometimes, as in this case, we can rapidly find what is making people ill and quickly remove it from sale. It’s an exciting role and we get a real sense of satisfaction out of using our skills to help people in this way. Other times it can be more frustrating – some outbreaks remain unsolved and it’s a real worry that people will get sick because we can’t eliminate a threat from food distribution.

The role really keeps us on our toes. Our surveillance systems mean that we have a good sense of patterns of illness across the year and how we can intervene to stop people getting unwell – but changes to food habits can catch us by surprise. For instance, raw milk has become more popular recently, bringing with it all the disease risks you would expect from a product that has come straight from a cow without any treatment to kill off bacteria!

In another case, eight people in the UK were affected while on holiday in Germany that was related to seeds.  The seeds were decoratively used as a garnish on salads and were difficult for cases to remember eating. Nearly 1,000 people in Germany got ill in that outbreak and one of the approaches by authorities was to use tourist photos of food to try and identify the common item in meals that could be making people sick.

Solving food borne illness outbreaks can be a real challenge, but by using a variety of the different tools available to us we can quickly intervene to stop people getting ill.

UK toddler, 3, fighting for life after catching E. coli on holiday

Martin Coulter of the  Star reports that Harper Davis, 3, has been taking to intensive care and put on dialysis at Nottingham Children’s Hospital.

The youngster was described as being a “normal, healthy girl” before she picked up the lethal bug.

Harper is understood to have suffered acute kidney failure, seizures, peritonitis and developed diabetes as a result of her illness.

My 3 grandsons are pictured below, from a Canadian Thanksgiving feast put on by daughter Jaucelynn, on Monday.

My kids all know food safety, but sometimes knowing is never enough.