What’s in a name? Why it’s important to classify bacteria accurately.

There are thousands, perhaps millions, of bacterial species living on Earth. Most operate below the radar until some event grabs our attention and forces us to get to know them better. Sometimes the event is a major outbreak. Nearly 40 years ago, a fatal respiratory disease led scientists to a new family of pathogenic bacteria, the Legionellaceae that was named for the American Legion convention in Philadelphia where the lethal cases occurred. In other situations, the trigger is an environmental survey. In 2012, a global public health and safety group that set out to learn more about microbial hot spots in Michigan homes isolated a new member of the Gram-negative Klebsiella genus from a bathroom toothbrush holder1. Yuck, is right.

While the novelty of uncovering a new germ captivates the imagination—who can resist Michael Crichton, after all —there are also examples of bacteria ending up on the wrong biological library shelf, so to speak. There are many instances where species have ended up with one name but investigations show that they should be classified as a different species.

So does knowing the precise lineage of a bacterial species—otherwise known as bacterial taxonomy—really matter to anyone other than the researcher who discovers it?

Of course it does. Getting the nomenclature right means appropriate actions can be taken after discovering a product contamination or appropriate drugs can be prescribed for an infection. Misclassifying bacteria leads to inappropriate remediation or patient care and distorts the organism’s clinical significance. It can taint the work of future studies.

Finding the Family Tree

Scientists have been naming bacteria since the late 1800s. But even with 21st century tools, classifying bacteria properly can be a maddening process. In taxonomy, organisms are usually classified into subspecies, species, genera, families, and higher orders. For eukaryotes (plants and animals), the defining feature in species classification is the ability to sexually reproduce and produce fertile offspring, but bacteria do not undergo sexual reproduction so other criteria are used to classify them. This is where the work gets tricky, and why misidentifications likely occur.

Since the mid-1990s, the standard tool used by scientists to identify bacteria has been to sequence the small subunit (16S) ribosomal RNA gene (the 16S rDNA). Sequencing the 16S rDNA allows for comparative analysis of published sequences in microbial databases; you can determine if you have uncovered something that has been seen before or something that looks to be entirely new. While this powerful technique has revolutionized the classification of bacteria, it has also elucidated the misclassification of numerous bacteria that were named before the advent of DNA sequencing. Historically, species would be classified as different based on phenotypic or clinical properties. These properties can vary based on how a bug is grown or what host it has been exposed to, but the DNA stays the same. Once the 16S rDNA sequences were derived, the story got muddled and different species had the same name or the same species had different names. The challenge is to unravel the confusion that is bacterial taxonomy.

Take the investigation of the bacterium isolated from a toothbrush holder as an example. The research team from NSF International’s Applied Research Center in Ann Arbor, which funded the work, knew the capsulated microbe with a slimy surface belonged to the family of enteric bacteria, which also contains Escherichia coli. When the 16S rDNA sequence was analyzed, it was unique. Different tests, MALDI-TOF and phenotypic analysis, pointed at different species, but detailed phylogenetic analysis provided a new identity—Klebsiella michiganensis.

NSF International, who launched the household survey and stumbled on the new microbe, had also looked at the aerosolization of microbes when you flush the toilet—what gets in the air and where does it settle. But this is hardly the only example of a modern-day germ hunter. Industrial biologists are also looking for organisms with enzymatic pathways that might be useful for manufacturing or bioremediation, and university teams are looking at the impact of extreme environments on the survival of different species or the original bio warfare where bacteria produce potentially new antibiotics to take out the competition.

Cataloguing Bacteria

While finding new microbes can get a bacterial taxonomist excited, attention is also spent reviewing organisms that have simply ended up in the wrong place. Here at Accugenix, we have developed and maintain multiple identification libraries as references for different kingdoms of organisms (bacteria and fungi) so that our customers—pharmaceutical, medical device, dietary supplement and personal care product manufacturers—know what bugs might be lurking in their facilities or products and how to respond appropriately. We comb the scientific literature for examples of new and reclassified species that might be good candidates to add to our library. We vet each organism to make sure its nomenclature is right. We may order the type strain from one of the international culture collections, sequence the 16S rRNA gene and evaluate its phylogeny or taxonomy.

But it’s the samples we get from customers that often help us spot discrepancies. We get tens of thousands of “unknown” microbial samples annually. We sequence the target DNA from the ribosomal region and use that data to construct phylogenetic trees to determine the closest related species. When we are evaluating the trees, we sometimes uncover glitches in the taxonomy. For instance, we discovered that one organism initially classified as a member of the genus Caulobacter—which is an excellent model microorganism for analysis of the regulation of the cell cycle—was more likely Sphingomonas. We also found a strain of Bradyrhizobium—the most common source of nitrogen fixation in plants—misclassified as the Agromonas genus.

When we encounter what we think is a mismatch, we annotate our library entry with what we believe is the actual name. When a microbiologist publishes a reclassification and it matches what we had been denoting, we can then provide the history. We also occasionally find different species published in the scientific literature that are actually the same species. Those, however, are harder to correct because you are essentially saying someone’s initial paper was wrong.

The work of bacterial taxonomists may seem academic, but it really isn’t. The federal government requires that companies monitor their environment and keep track of what is in that environment.  If you can’t get an accurate name on an organism, it makes it difficult to be confident in the results obtained from routine microbial surveillance.

This is the reason why we maintain such extensive microbial reference libraries—which includes a MALDI-TOF bacterial database, a sequencing library database for fungal identification and one for bacterial identification.

Which brings us back to the case of Klebsiella michiganensis. Scientists can predict by observing the microbe that its sticky surface could help it to attach to mucus membranes, evade immune responses, and invite other germs to build biofilm communities that can lead to persistent infections. Not surprisingly, perhaps, they also discovered that the microbe is resistant to three common antibiotics.

So by properly classifying the microbe, scientists and manufacturers now have a better shot of at least tracking it and designing ways to eliminate or control it.

As the saying goes, a microbe by any other name…

References:

  1. Curr. Microbiol  72 2013.