The CLASI-FISH method reveals the bacterial community in dental plaque, which Dr. Mark Welch nicknamed “the hedgehog” because of the clusters of bacterial filaments. The core of the filament structure is composed of Corynebacterium (tagged purple). What kind of communities do oral bacteria live in? In the 1970s, electron microscopy was used to make images of bacteria, including that of dental plaque. “There are beautiful electron micrographs of dental plaque structures that were described as looking like corn cobs, because there were little spherical bacteria called cocci and longer filamentous bacteria that got clumped together,” says Dr. Mark Welch. At the time, researchers could trace channels of liquid through the biofilm, and they noticed that filament rich areas of the biofilm produced more calculus. “Then the inquiry sort of got dropped because they’d done all they could with the limits of electron microscopy. That’s where we came in,” she says. To create useful microscopy images, Dr. Mark Welch and her colleague Dr. Gary Borisy needed different kinds of bacteria to appear as different colours. Since the 1980s, researchers have been using short pieces of synthesized DNA with a fluorescent molecule attached to it to tag bacteria for microscopy. “You drop a short piece of DNA, like 20 bases or so, and it finds the ribosomal RNA it matches and attaches to it and then your bacterium has a fluorescent label,” says Dr. Mark Welch. “A standard fluorescence microscope can see about 3 or 4 different colours at once: red, green, blue, maybe also far red.” As the research team began their work, cell biology was using advanced microscopy imaging that could discriminate between colours with very similar spectra. Drs. Mark Welch and Borisy decided to see what they could discover if they applied this advanced tool to microbial ecology. “With this new tool, we could use up to 16 different fluorescent labels and tell them apart,” says Dr. Mark Welch. “Then we thought: What if we use the different fluorescent labels in combination? Then we could visually differentiate more than a thousand different kinds of bacteria in a single image. Wouldn’t that be cool?” Dr. Dewhirst and his team are using this idea to try to culture formerly unculturable bacteria by growing two kinds of bacteria together that are proximal inside the microbiome. “The idea is that if one bacterium has given up the gene it needs to make the metabolite that its neighbour makes, then they need each other to culture,” says Dr. Mark Welch. Drs. Mark Welch and Borisy’s innovative imaging method revealed that dental plaque has highly organized, complex bacterial structures that they called “hedgehogs.” 26 | 2024 | Issue 1 Issues and People
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