Bac­te­r­ial Blues

by Janie

The color blue is an odd­ity. Of all pig­ments in na­ture – chem­icals that se­lec­tively ab­sorb and re­flect cer­tain wave­­lengths of vis­i­ble light – blue ones are among the rarest. Pro­ducing blue dyes was once such a costly busi­ness that me­dieval Eu­ro­peans consid­ered it to be as pre­cious as gold. Blue was a marker of wealth and sta­tus, and artists reserv­ed its use only for the most impor­tant occa­sions, as in the strik­ing ultra­marine blue of the head­scarf in Vermeer's Girl with a Pearl Ear­ring.

Fig. 1. Girl with a Pearl Ear­ring, Jo­hannes Ver­meer c. 1665. Source

Nat­ural true blue pig­ments are excep­tional. The only one known so far to be pro­duced by an­i­mals is found in the wings of the ob­rina olivew­ing but­ter­fly. Flow­ers such as hy­drangeas and fruits such as blue­berries par­tially owe their col­oration to a shifty fam­ily of com­pounds called antho­cyanins – a condi­tional blue. The mole­cules are red in acidic condi­tions and blue (or even black) in alka­line condi­tions, with a slid­ing scale of pur­ple in the mid­dle.

Then what about all the other appar­ent blues in na­ture? In liv­ing organ­isms, the color­ation we see is due ei­ther to pig­ments or to the phys­ical proper­ties of the organ­ism's sur­face that per­form a bit of sleight of hand with op­tics. Mem­bers of the lat­ter, which owe their appear­ance to micro­scopic struc­tures that scat­ter and re­fract light, are called struc­tural col­ors. The vast ma­jor­ity of "blues" in na­ture are struc­tural, in­clud­ing bird feath­ers, poi­son dart frog skin, and people's eyes. In par­rots, for ex­am­ple, reds and yel­lows are due to psittaco­fulvin pig­ments (which, interest­ingly, con­fer re­sistance to feather degrad­ation by Bacil­lus species) but all bird blues are struc­tural. Any kind of irides­cence or metal­lic lus­ter, as in pea­cock feath­ers or the cara­paces of jewel bee­tles, owe their glit­ter to struc­tural col­ors (see a pre­vi­ous STC post on irides­cent bac­te­r­ial colonies here). Color-chang­ing camou­flage, as in chameleons and oc­to­puses, is also struc­tural, thanks to manip­ulation of the spac­ing be­tween pig­ment crys­tals. "True blue" is in­deed rare.

Mi­crobes, how­ever, seem to be unusual­ly tal­ented prod­ucers of blue pig­ments. To find varia­tions on the blue pig­ment theme, look no fur­ther than the sec­ondary meta­bolites of the tini­est organ­isms.

Fig. 2. Strep­to­myces coeli­color colonies. Source

First up, some re­dox clas­sics from some soil-dwellers. Py­o­cyanin from Pseudo­monas aeru­ginosa is a blue-green pig­ment that can kill com­pet­ing mi­crobes by caus­ing re­­dox-stress. It's also in­volved in a fasci­nating mole­cular fer­ry sys­tem for shut­tling elec­trons through­out a bio­film. Phena­zine pig­ments like pyo­cyanin from sev­eral bacter­ial gen­era can pro­duce every color in the vis­i­ble spec­trum, and blue is no excep­tion. Yet an­other re­­dox-ac­tive anti­bi­otic is actino­rhodin, a blue poly­ketide hail­ing from Strep­to­­­myces coeli­color. Here's a wonder­ful bac­terial name: coelus is Latin for "sky" and is a nod at the color of the celes­tial ex­panse, again encap­sulated in the Span­ish word for light blue, ce­leste.

Fig. 3. Left: Glaukothalin iso­lated from Rhein­heimera sp. Strain HP1. Frontis­piece. Source. Right: Glaukothalin chem­i­cal struc­ture. Source

Glaukothalin (from the Greek glaukos  for "blue" and tha­latta for "sea") is aptly named: this deep-blue pig­ment is from Rheinheim­era baltica, a blue ma­rine bacter­ium that was first iso­lated from the Baltic Sea, as its name hints. Glaukothalin in­hibits the growth of some other ma­rine bac­te­ria and is cy­to­toxic to brine shrimp, but its ecolo­gi­cal role re­mains mys­te­ri­ous.

Fig. 4. Color changes of in­oc­u­lated G plates, num­bers show­ing the hours of in­cu­ba­tion at 16 °C. Source

A beaut­iful deep blue pig­ment from the plant path­ogen Pan­toea agglo­merans (for­merly Er­winia herbi­cola) is pro­duced only at temp­era­tures above 10°C at high cell den­sity. Un­like R. baltica above, the Pan­toea cells them­selves are not blue; in­stead, the wa­ter-sol­u­ble pig­ment they pro­duce seeps out into their agar sur­round­ings. This dye also turns pink in acidic condi­tions, like the plant pig­ment antho­cyanin.

Fig. 5. The pro­posed path­way for the di­ver­gent biosyn­the­sis of MIN and in­digoi­dine. Source

Next up is indigo­idine – note that this blue pig­ment is chemic­ally dis­tinct from in­digo, as the for­mer is a bi­pyridine and the lat­ter is a bi­in­dole (more on in­digo shortly!). A num­ber of species natur­ally synthe­size this blue, in­clud­ing the plant path­ogen Dick­eya dadan­tii (for­merly Er­winia chrysan­themi) and var­i­ous strains of Strepto­myces. Roseo­bacter species are known to ex­ploit the an­timi­cro­bial prop­erties of indigo­idine to kill com­pet­ing species, as does one species that coats the eggs of Hawai­ian bob­tail squids to pro­tect from vib­rios. In Strepto­myces hygro­scop­icus, it turns out that a sin­gle non­ribo­somal pep­tide synthe­tase is respons­ible for coordin­ating switches be­tween synthe­sizing indigo­idine and the anti­biotic mini­mycin; it can synthe­size indigo­idine all by it­self and also ini­tiates the bio­synth­esis of mini­mycin. NRPSs have been en­gi­neered in fungi to scale up produc­tion of the com­pound.

Fig. 6. Dif­fer­ent en­zy­matic routes to­wards in­digo, ei­ther via dioxy­gena­tion (A), di­rect hy­drox­y­la­tion to in­doxyl (B), or via epox­i­da­tion ©. Source

Fi­nally, in­digo, the quint­essen­tial blue pig­ment used to dye tex­tiles as far back as 6000 years ago in an­cient Peru. The histor­ical means of produc­tion was process­ing leaves from the plant genus In­digofera, which con­tain the pre­cur­sor mole­cules in­di­can and isa­tan B. But chem­ical synth­esis took over in 1870 – a process that unfor­tunately calls for harsh chem­icals and gener­ates harm­ful waste. It turns out that mi­crobes pos­sess an ar­ray of re­dox en­zymes that are ca­pa­ble of produc­ing in­digo sans the toxic byprod­ucts and environ­mental pollu­tion. (Note: A mole­cule re­lated to in­digo is respon­sible for the blue pig­ment formed when X‑gal is bro­ken down by b‑galacto­sidase in X‑gal plates!)

Mi­crobe-de­rived pig­ments like the ones men­tioned here are eco-friendly dyes with myr­iad applic­ations be­yond tex­tiles. Blue pig­ment prod­uction by P. agglo­merans is tem­per­a­ture-de­pen­­dent, so per­haps there is poten­tial for use as a temper­ature in­di­ca­tor for foods and medi­cines. Voge­sella indigo­fera only pro­duces indigo­idine un­der a thresh­old concen­tration of Cr⁶⁺, so per­haps there is poten­tial here for a heavy metal bio­sensor. The cyano­bacterial acces­sory pig­ment phyco­cyano­bilin has even been ap­proved by the FDA as a food color­ing.

Mi­cro­bial pig­ments are also in the works as immuno­suppres­sive and anti­cancer drugs – a fit­ting throw­back to the meth­yl­ene-blue ori­gins of drug discov­ery. (As a med­ical stu­dent, Paul Ehrlich was fascin­ated by methyl­ene blue, which stains only nerve cells. His insist­ence on pursu­ing his new­found obses­sion with dyes and their "chem­ical affin­ity" for cer­tain cells – to the dis­may of his med­ical school profes­sors who thought it a use­less distrac­tion – led him to de­velop impor­tant drugs like sulfa anti­biotics.)

But it's not only pig­ments that pro­duce both beaut­iful and func­tional color­ation in mi­crobes. We'll fol­low up on this post with an­other on unexpec­ted func­tional bonuses of struc­tural color­ation in bac­te­ria. In the mean­time, see this prev­ious post here for an­other look at microb­ial pigment­ation.