Before continuing, you may want to read our previous post.
We are happy to announce the bacterial 16S rDNA sequencing results from our two Chobani Greek yogurt samples: a strawberry-flavored cup of normal appearance, and a recalled lime cup. You can find a non-technical summary at the bottom of this post.
We have determined unambiguously that the lime cup used in our study was involved in the recall because the IMS number matches exactly with that posted on the FDA recall site (and the yogurt showed all of the characteristics). No such IMS number could be located on the strawberry cup, whose contents appeared normal, so we used it to compare with the recalled lime yogurt. Top photos of each cup are shown below, while profile views are shown in the previous post.
Before we share the results, we want to point out that we do not contest anything Chobani has reported about the cause of the recall, including the mold contaminant and its non-hazardous nature. Although the FDA is investigating complaints of illness related to the recalled yogurt, no determination has been made whether or not such a link exists. Importantly, the data we present below do not in any way show that the recalled Chobani yogurt was capable of causing illness or caused illness of any kind.
Our study was conducted purely to satisfy curiosity, and our experiment has several shortcomings, including: (a) a very small samples size (n = 2), (b) the comparison of 0% fat strawberry and 2% fat lime flavors, (c) inhomogeneity in the yogurt cup, which could lead to an unrepresentative sample being collected, (d) PCR bias, (e) the possibility of misidentification of organisms sharing a common 16S rDNA sequence, (f) sequencing errors resulting in mischaracterization of bacterial taxonomy, (g) incorrect alignment databases, and (h) possibility of DNA contamination. There may be other shortcomings of which we are not aware.
Below are pie charts showing the taxonomic order associated with the most abundant 16S rDNA reads from the normal-appearing strawberry cup (top) and the recalled lime cup (bottom).
Chobani indicates that they have three probiotic cultures in their yogurt: Lactobacillus acidophilus, Lactobacillus casei, and Bifidus. The first two belong to taxonomic orders Lactobacillales and the latter belongs to Bifidobacteriales, which is consistent with what we found by sequencing bacterial DNA in the normal strawberry cup (see top chart).
However, the recalled lime yogurt shows a considerably different distribution. Out of a total of 1,950,616 reads for this sample, 190,331 (9.8%) belonged to the order Oceanospirillales (see bottom chart). Fortunately, the sequence data contain more information than the order; they also give insight into the taxonomic family of the bacterium (Oceanrickettsiaceae) and the genus (Oceanrickettsia). This is where things get extremely obscure, however, because aside from a couple of sequence entries in Genbank on an organism sharing the same genus (Oceanrickettsia ariakensis), we know almost nothing about these bacteria. So little, in fact, that the taxonomic family and genus have not been formally accepted and the organism has been relegated to a list of “unclassified Gammaproteobacteria.”
From what little we can glean, two Chinese scientists affiliated with the South China Sea Institute of Oceanology, Drs. Xiangyun Wu and Yang Zhang, deposited the sequence data for Oceanrickettsia ariakensis into the database in 2005 and indicated that they isolated the bacteria from an oyster (Crassostrea ariakensis). They believed that the bacteria infected the oyster and contributed to its massive die off. It is very important to note that there is no evidence that this bacteria would be pathogenic in humans. Furthermore, the work of Wu and Zhang does not appear to have been independently verified or subjected to peer review (to the best of our knowledge).
Does this mean that the Chobani yogurt recall had anything to do with oyster contamination? Absolutely not! It’s possible that the bacteria whose DNA we detected may have already been present in tiny quantities in one of the yogurt’s ingredients, and when the mold took over the yogurt culture, it created conditions that allowed it to thrive. In support of this theory, we observed 198 reads out of 1,713,939 total that corresponded to order Oceanospirillales (0.012%) in the strawberry yogurt. Or perhaps our findings do provide some insight into the contamination source. It’s too early to say.
But what we can say is that we have a lot more to learn about bacteria. We can only culture (and therefore, thoroughly study) about 1% of the bacterial species on the planet. With the latest generation of sequencing technologies, we can sequence everything. Wonder what else is out there? So do we.
And in case you were wondering, I haven’t stopped eating Chobani yogurt and am still a big fan! Many trusted brands have faced a recall at some point during their history. Microbial contamination is a constant concern among food manufacturers and is not uncommon. The overwhelming majority of the time, the extra passengers we call bacteria simply go along for the ride, completely unnoticed.
Bacterial DNA was extracted from a normal-looking Chobani strawberry yogurt cup and from a recalled lime yogurt cup. The bacterial DNA from each sample was sequenced to gain insight into the kinds of bacteria that may have been present in the original yogurt cups. The strawberry yogurt showed a profile consistent with the probiotic cultures expected to be in the yogurt according to Chobani’s FAQ. In contrast, the recalled lime yogurt showed a somewhat different profile, and suggested the presence of a bacteria belonging to the class Gammaproteobacteria. The original discoverer of the bacterial DNA we sequenced has proposed that it comes from bacteria belonging to the genus Oceanrickettsia. Very little reliable information exists about this bacteria, and it appears not to be formally classified. There is no evidence that it causes disease in humans.
We had just extracted bacterial DNA from a set of facial microbiome samples when I decided to reward myself with a cup of lime-flavored Chobani Greek yogurt. Oddly, the foil on the cup was bulging, the yogurt was thin, and it was extremely carbonated. A photo comparing it to a “normal” strawberry flavored cup is shown below.
A quick google search brought me to Chobani’s Facebook page, which showed that many people were reporting the same issue. Since there are lots of bacteria capable of doing lactic fermentation, we decided to extract the bacterial DNA from this funky lime cup and a “normal” strawberry cup and do 16S rDNA sequencing to identify the bacteria present.
Since the time we commenced our study, Chobani announced a voluntary recall and has since reported that it believes the contamination is from a mold (Mucor circinelloides).
Our technique will only give us information about the bacteria present, not about molds, which are a type of fungus. Nevertheless, we are very excited to see if there are any unexpected bacteria in the culture and to understand how the mold contamination changes the composition of the original bacteria in the yogurt culture. We will report our results within a few days.
Below is a pic of our collaborator, Danny, in front of Illumina’s MiSeq instrument.
And in case you were wondering, yes, I ate the lime yogurt and I was fine. The carbonation was exciting, generating a kefir-like (mojito-like?) yogurt. While Greek yogurt purists might take issue (would they really be eating lime-flavored yogurt anyway?), I think Chobani should try to achieve the carbonation with a lactic fermenting bacteria and launch it as new product.