Microbes and UV
So, do microbes grow under ultraviolet (black) light?
Well, some bacteria do – but not in the way many seem to think based on those ever-reliable barometers, TV and the Internet. Just point your Google-Fu at any combination of ‘bacteria’, ‘UV’, ‘black light’ and you’ll see what I mean.
A disturbing trend is the promotion of fluorescence under UV as a diagnostic tool for unsanitary areas, doubtless fuelled by TV crime dramas where natural fluids or organic matter are detected with UV, as well as in other TV formats in the ‘How Minging Is Your…’ genre. I’m now seeing systems being promoted in food and healthcare claiming you can expose bacteria on surfaces with UV ‘because bacteria fluoresce’. Some even suggest such systems are suitable for validation of cleaning effectiveness in food prep and healthcare. I’m calling Bollocks on that notion for some very simple reasons – but not Utter Utter Bollocks (µ²B) on the entire concept because the detail is a lot more interesting. To geeks like me, at least.
So what’s the deal?
Many natural substances do glow under UV: semen, urine, mineral oil, earwax, perspiration – and pest control companies use UV light to detect rat faeces and urine which also fluoresce under UV – more on that later. But what about the bugs?
Our Microbial Overlords as Illuminati
It is true that some bacteria do glow in and of themselves and many more will glow under UV light to a greater or lesser degree, so the plausibility factor is there. And UV light is used industrially in food production and other processes to detect certain specific bacteria. But we’ve all seen those TV programmes where a ‘clean’ washroom is plunged into darkness, the UV switched on and – euwwwww! But what does it really tell us?
Fluorescence is a form of luminescence. It occurs when a substance absorbs light at one wavelength and re-emits it at a longer wavelength. This property is used in optical brighteners in laundry products – they absorb (invisible) UV light and re-emit (visible) light in the blue part of the spectrum. Many natural molecules exhibit this property including rocks and gems, some vitamins and even your G&T – tonic water fluoresces blue due to the presence of quinine. Many, many molecules can do this but one class of chemical that fluoresces intensely when exposed to ultraviolet light are flavins. A key natural source of flavins is Vitamin B. Uric acid and its salts and soap scum all contain Vitamin B.
Many claim certain species of bacteria accumulate on surfaces where high concentrations of flavins are found so the claim is made by many sellers of hand-held UV systems that UV ‘exposes’ bacteria because bacteria fluoresce. This is not true.
As a rule bacteria do not fluoresce. Some strains of Salmonella and Shigella relevant in foodborne illness can, and many contain materials which fluoresce – but that fluorescence not always visible to the naked eye, either because it’s at the wrong wavelength or there just isn’t enough of it.
Washroom fixtures including toilets, urinals, walls, partitions, mirrors, floors and counter tops will commonly have deposits of compounds derived from urine and soap scum. When you shine ultraviolet light on surfaces with deposits of urea salts or soap scum they absorb it and emit visible light to the naked eye (if viewed in total darkness). Surfaces can be inspected with the use of an appropriate ultraviolet light but this is NOT an accurate indicator of presence of bacteria. But what it can do is indicate areas where bacteria might be found. This is where Rule 1 of hygiene and infection control comes in: You Can’t Disinfect Dirt.
A surface must be visibly clean before you can disinfect it and one that fluoresces under UV most likely isn’t.
Is UV an Indicator of Bacterial Contamination?
Yes and no. There is a serious scientific flaw in the use of UV for detection of bacteria using the method described above. A recent study swabbed a number of surfaces in critical healthcare situations and looked at levels of bacterial contamination of surfaces that fluoresced under UV versus those that didn’t. It found that colony densities were significantly higher in non-fluorescing areas versus fluorescing areas. It also found that the colour of fluorescence may determine the level of sanitation necessary to properly clean a room. While this is interesting one cannot ignore the fact that surfaces that do not fluoresce under UV cannot be considered either clean or free of bacterial contamination – presence of fluorescence can indicate a surface is not clean and may well have a few good bugs on it but lack of fluorescence is not indicative of lack of bacterial contamination. It’s a great bit of science ‘theatre’ and very effective when used in hand washing etc training but (very big but) surfaces that do not fluoresce cannot be considered hygienic.
So, next time you are told somewhere hangs their hat on UV for microbiological safety – eat somewhere else. In healthcare UV is used for decontamination and trials are taking place to see if it can be used to see if a surface (like a bed rail) is free of Our Microbial Overlords – but this involves some serious lab kit and pukka swabbing technique – it’s not the sort of thing that’s works in situ especially when used by a TV presenter waving a UV lamp.
Real Uses of UV and Fluorescence
Fluorescence is used as a diagnostic technique – bacteria can be stained or genetically engineered so they will fluoresce but this is in the realm of proper grown-up labs with specialist techniques and equipment, not some idiot waving a 20W UV lamp bought on the Internet.
Fluorescent biomolecules such as flavins and the amino acid tryptophan do exhibit fluorescence and these are present in bacteria and other living cells but fluorescence is very specific: a particular molecule will absorb at a particular wavelength and re-emit at another, specific wavelength – and the emitted wave needs to be within the visible spectrum for us to see it without specialised equipment. For example tryptophan emits at 280nm and re-emits at 330nm – and the visible spectrum is about 380 to 750nm so you can’t see it. And molecules like tryptophan will often be buried in a cell or its membrane so you still might not spot it even using the really trick microscopic techniques we have in the lab.
BUT Riboflavin (vitamin B2) reliably emits at a bright green 560nm when exposed to a broad spectrum of UV and bacteria are rather partial to flavins – as are we all: they are essential for metabolism. This is the molecule most UV systems are detecting. It is true that you do tend to find bacteria on surfaces where flavins are present but this does not mean that flavin-free or non-fluorescing surfaces are free of bacteria – far from it. This is why the sorts of UV emitters you see on TV programmes are about as much use as tits on a nun as a pukka diagnostic technique.
Clinical Uses of UV
I’m not a clinician (for which the world should be eternally grateful) but the ability of biological molecules to fluoresce is and has been used as a useful diagnostic tool; Corynebacterium minutissimum will glow coral red, Pseudomonas a yellow-green, Propionibacterium acnes orange. And molecules like porphyrins will glow too to give physicians signs of other diseases.
But What About Forensics?
Forensic Scientists don’t use the sort of UV lights you can buy off the Interwebs. They use very trick forensic lights that can emit a range of specific wavelengths. This is important because you need to differentiate what you’re looking for from what it’s on, and often the surface will fluoresce as well as your target substance – but if you vary the wavelength you can often ‘tune out’ the background noise. And many substances will just absorb UV and so will look darker – for example blood and semen at 415nm. Forensic lights (where you can vary the wavelength) can be used in latent fingerprint identification, crime scene searches, identifying counterfeit documents as well as in biology and serology. Some substances will happily emit all by themselves, some need help with an identifying spray such as fluorescein.
Does UV Kill Bugs?
UV is pretty good at killing bugs. Many hospitals have UV-sterilised air and water systems. UV is used during depuration of shellfish, water purification and many other processes. My lab – like pretty much any other – vents to the open air because the combination of massive dilution of any of our Microbial Overlords and UV in sunlight provides very effective decontamination of anything we blast out.
It has long been proven that UV-C light is antimicrobial when bacteria are exposed to it for various lengths of time. UV- C light extends from 180nm to 280 nm and exposes the bacteria to rays which mutate bacterial DNA by the formation of pyrimidine diamers which lead to an inaccurate DNA template, which prevents them from reproducing. At 254 nm, which offers 99.9% destruction of sundry microorganisms the UV energy in microwatt-seconds per cm² necessary to cause cell death ranges from 2,500 to 36,000. You will not get this from a hand held UV light.
What Microbiologists Use It For
Not for enumerating Our Microbial Overlords in sodding kitchens or hotel rooms using a shitty £5 UV lamp!
UV is used by microbiologists to spot Our Microbial Overlords – but we need a hand to identify the little bastards. We can use fluorescent dyes for counting the number of bacterial cells such as acridine orange – which stains both living and dead cells by shagging both the DNA and protein components of cells – which then fluoresce Guantanamo Orange when exposed to UV. This is really useful in – for example – testing soil or water where you can have very diverse bug populations – situations where you can’t just whack them on a plate and see what grows as a TVC (or total viable count in the argot). Many marine microbiologist pals use this because bug levels are low and a TVC will return a massive underestimate of total number of bacteria – a TVC can show less than 1% of a direct count for marine samples.
More useful for my purposes are reagents like cyanoditolyl tetrazolium chloride (CTC) – it binds to proteins involved in respiration and so can flag live cells – which is all I’m interested in (apart from C. diff spores and the noroviruses but that’s another post or ten). There are other reagents such as auramine and rhodamine which bind to the cell walls of mycobacteria and glow under the right UV. This is important because the mycobacteria don’t Gram stain well – which is a capital offence to anyone with a copy of Bergey’s and a sample from a really ill patient. These are replacing the old acid-fast stains I was taught to use as a cub microbiologist in the 1980s and it’s a Good Thing: the non-TB mycobacteria are on the increase principally due to shit tattoos and lipo-tourism. I kid you not…
So yes, you can make bacteria glow and surfaces that re-emit at visible wavelengths might well be dirty. But that doesn’t mean surfaces that don’t are clean or free of Our Microbial Overlords. Like everything in biology, it’s complicated….