Polymi­cro­bial In­fec­tions

Per­haps The Rule, Not The Ex­cep­tion

by Katy Pa­tras
 

The re­cent film World War Z de­scribes an apoc­a­lyp­tic event where a ra­bies-like virus spreads via bite wounds to the ma­jor­ity of Earth's pop­u­la­tion and turns them into zom­bies. Since the world's lead­ing vi­rol­o­gist dies early in the film, it is up to Brad Pitt's char­ac­ter to find the source and cure for this zom­bie virus. Spoiler alert: He dis­cov­ers that virus-in­fected in­di­vid­u­als will not bite hu­mans with other in­fec­tions or ter­mi­nal ill­nesses, and pro­ceeds to in­fect him­self with a BSL4 pathogen and walk among the zom­bies un­scathed. Al­though every sci­en­tist read­ing has first cringed and then come up with at least a half dozen rea­sons why this is not pos­si­ble, the point is that even the film in­dus­try is rec­og­niz­ing that path­o­genic (and non-path­o­genic) mi­crobes are ca­pa­ble of com­mu­ni­cat­ing with one an­other within the host to al­ter the course of dis­ease.

Fig­ure 1. Crit­i­cally ill pa­tients in the in­ten­sive-care unit are good ex­am­ples of the di­ver­sity of sites at which bac­te­ria and fungi can in­ter­act and cause dis­ease. The boxes de­scribe the or­gan­isms that are most com­monly found at each site. Source.

In­ter­mi­cro­bial com­mu­ni­ca­tion must in fact a com­mon event, since symp­to­matic polymi­cro­bial in­fec­tions arise at con­sid­er­able fre­quency, al­though ex­act epi­demi­o­log­i­cal oc­cur­rences are un­known at this point. This is hardly sur­pris­ing given that we are in con­stant con­tact with nu­mer­ous op­por­tunis­tic mi­crobes re­sid­ing on our body's ep­ithe­lial bar­ri­ers (skin, mu­cous mem­branes). Click here for a re­view of this topic. How­ever, in typ­i­cal cur­rent clin­i­cal prac­tice, one pathogen re­spon­si­ble for the dis­ease is iden­ti­fied and tar­geted for treat­ment with more or less spe­cific an­timi­cro­bial drugs. Be­cause of this nar­row scope of treat­ment, other col­o­niz­ing mi­crobes, not nor­mally vir­u­lent, may be given a com­pet­i­tive edge, lead­ing to chronic in­fec­tions or se­vere com­pli­ca­tions. An old Aesop's adage comes to mind: A doubt­ful (op­por­tunis­tic) friend is worse than a cer­tain (path­o­genic) en­emy.

Al­though we have been aware of polymi­cro­bial in­fec­tions, ar­guably for cen­turies, only re­cently has biotech­nol­ogy al­lowed the del­i­cate dis­sec­tion of the role of in­di­vid­ual mi­cro­bial species in in­fec­tions.  This in­cludes meta­bolic and chem­i­cal in­ter­ac­tions be­tween mi­crobes as well as joint at­tacks on the host im­mune sys­tem. I will fo­cus on a few re­cent dis­cov­er­ies that il­lus­trate types of mi­cro­bial col­lab­o­ra­tions dur­ing in­fec­tion.

Mol­e­c­u­lar In­ter­play Be­tween Co-In­fect­ing Mi­crobes

How do mi­crobes es­tab­lish com­plex in­ter­ac­tions when present to­gether in a given tis­sue?  The an­swer is: every which way! Some am­plify vir­u­lence of one or more species; oth­ers pro­mote long-term per­sis­tence. For ex­am­ple, in chronic skin wounds, Pseudomonas aerug­i­nosa vir­u­lence is ac­ti­vated by Gram-pos­i­tive or­gan­isms, such as Staphy­lo­coc­cus au­reus. Here, Pseudomonas senses the pres­ence of pep­ti­do­gly­can shed by Staphy­lo­coc­cus, which re­sults in en­hanced pro­duc­tion of the toxin py­ocyanin. The re­sult is greater host killing. Clearly, in­ter­species bac­te­r­ial com­mu­ni­ca­tion in­flu­ences the reg­u­la­tion of gene ex­pres­sion. Here is an in vitro ex­am­ple:  co-cul­ture of two op­por­tunis­tic pathogens of the hu­man res­pi­ra­tory tract, Strep­to­coc­cus pyo­genes (Group A Strep­to­coc­cus, GAS) and Moraxella ca­tarrhalis, in­creases tran­scrip­tion of sev­eral GAS vir­u­lence fac­tors in­clud­ing hyaluro­nan syn­thase and strep­to­coc­cal mi­to­genic ex­o­toxin Z. At the same time, the ex­pres­sion of 12 GAS two-com­po­nent reg­u­la­tory sys­tems (con­tain­ing the well-stud­ied con­trol of vir­u­lence (covR/S) reg­u­la­tory sys­tem, which down reg­u­lates vir­u­lence fac­tors) de­creases, thus mak­ing the point that in­ter­ac­tion be­tween two mi­crobes can turn an op­por­tunist such as GAS into a se­ri­ous pathogen.

Fig­ure 2. Polymi­cro­bial biofilm of C. al­bi­cans and S. au­reus. C. al­bi­cans (10⁶ CFU/ml) and S. au­reus to (10⁷ CFU/ml) were con­cur­rently added to 96-well tis­sue cul­ture-treated cham­ber slides and in­cu­bated for 24 h at 37°C. Biofilm for­ma­tion was mon­i­tored by flu­o­res­cence mi­croscopy and scan­ning elec­tron mi­croscopy. Biofilms were stained with SYTO 9 (green, live bac­te­ria), FUN‑1 (in­tra­cel­lu­lar red, live fun­gal cells), cal­co­fluor white (blue, fun­gal cell wall). Ca, C. al­bi­cans hy­phae; Sa, S. au­reus. Source

In­ter­ac­tion be­tween species also con­tributes to suc­cess­ful mi­cro­bial per­sis­tence.  For ex­am­ple, S. au­reus, which is a poor for­mer of mono­cul­ture biofilms, and Can­dida al­bi­cans to­gether form un­usual, but ef­fec­tive, mixed biofilms, where the yeast cre­ates a scaf­fold for the bac­terium. Within this mi­croen­vi­ron­ment, S. au­reus pro­duces two vir­u­lence fac­tors: co­ag­u­lase and the pro­tein Efb. Co­ag­u­lase binds fib­rino­gen and ef­fec­tively coats Can­dida species with a layer of fib­rin, pre­vent­ing phago­cy­to­sis of Can­dida by the host. Efb also binds fib­rino­gen and pre­vents com­ple­ment-me­di­ated op­soniza­tion by a dif­fer­ent mech­a­nism. So, vir­u­lence syn­ergy hap­pens not only be­tween Gram-neg­a­tives and Gram-pos­i­tives, but also be­tween bac­te­ria and yeast. Pathogens may be bad tax­on­o­mists, but they ex­cel at patho­gen­e­sis.

Con­certed Ma­nip­u­la­tion Of Host Im­mune Func­tion

As is well known, per­sons with a pathogen-in­duced com­pro­mised im­mune sys­tem, such as that caused by HIV-me­di­ated de­struc­tion of CD4⁺ T cells, die of sec­ondary in­fec­tions if un­treated.  Here, in­fec­tion by one pathogen ma­nip­u­lates host im­mu­nity to the ben­e­fit of other pathogens. Many di­verse in­ter­ac­tions have re­cently been dis­cov­ered that di­rectly in­hibit in­nate or adap­tive im­mu­nity. For in­stance, dur­ing pul­monary co-in­fec­tion, in­fluenza A virus seems to pro­tect My­cobac­terium bo­vis by re­duc­ing the num­bers of my­cobac­te­r­ial-spe­cific CD8⁺ T cells and in­hibit­ing my­cobac­te­r­ial clear­ance.

In­ter­est­ingly, the abil­ity to mod­u­late the host im­mune re­sponse also de­pends on the host dis­ease state. For ex­am­ple, in cys­tic fi­bro­sis (CF) pa­tients, bron­chi­o­lar ep­ithe­lial cells first in­fected with P. aerug­i­nosa and then with rhi­novirus pro­duce lower lev­els of in­ter­fer­ons, whereas in nor­mal bron­chi­o­lar ep­ithe­lial cells, in­ter­feron pro­duc­tion in­creases. Thus, in CF pa­tients, not only is in­nate im­mu­nity mod­u­lated by chronic P. aerug­i­nosa in­fec­tion, the dis­ease is si­mul­ta­ne­ously ex­ac­er­bated by dis­ease-in­duced ox­ida­tive stress. Aren't you cap­ti­vated by the pre­cise and ef­fec­tive mea­sures pathogens have jointly de­vel­oped to evade host im­mu­nity and pro­mote trans­mis­sion?

Amal­ga­mated Vir­u­lence Lead­ing To Host Dis­ease Pro­gres­sion

Epi­demi­o­log­i­cal stud­ies have re­cently un­cov­ered cor­re­la­tions be­tween seem­ingly un­re­lated in­fec­tious agents that cause un­re­lated-sound­ing dis­eases. For ex­am­ple, co-in­fec­tion with Le­gionella pneu­mophila and in­fluenza A virus de­creases host sur­vival, not, as could be an­tic­i­pated, by height­en­ing the in­flam­ma­tory re­sponse, but rather by im­pair­ing the host's abil­ity to tol­er­ate tis­sue dam­age. This takes place in part by re­press­ing the ep­ithe­lial growth fac­tor path­ways that are crit­i­cal for main­tain­ing tis­sue home­osta­sis and re­pair. The in­volve­ment of pathogens in dis­ease pro­gres­sion may be even more ob­scure. In a large-scale com­mu­nity study, sep­a­rate in­fec­tion of two in­de­pen­dent or­gans, by Chlamy­dia pneu­mo­niae in the lungs, and by He­li­cobac­ter py­lori in the stom­ach, was sig­nif­i­cantly as­so­ci­ated with hy­per­ten­sion. Al­though how this works re­mains to be elu­ci­dated, it is pos­si­ble that in­fec­tions of these two agents sep­a­rately in­crease risk of in­fec­tion by in­duc­ing au­toim­mu­nity, per­turb­ing cho­les­terol me­tab­o­lism, and sus­tain­ing proin­flam­ma­tory sig­nal­ing.

Sur­pris­ingly, co-in­fec­tions are not al­ways detri­men­tal to the host. A re­cent study re­ported that in pa­tients co-in­fected with two strains of hu­man pa­pil­lo­mavirus (HPV strains 16 and 34), the in­ci­dence of lymph node metas­ta­sis was re­duced com­pared to those in­fected with HPV 16 alone. And, just as un­ex­pected, in­fec­tion with H. py­lori may be ben­e­fi­cial in re­duc­ing al­ler­gic asthma through height­en­ing the ac­ti­va­tion and ac­cu­mu­la­tion of T_reg cells in the lungs. The span of mi­cro­bial in­ter­ac­tions within the hu­man body is cer­tainly un­pre­dicted and may even turn out to be un­pre­dictable.

Fu­ture Di­rec­tions

Pub­li­ca­tions per­tain­ing to polymi­cro­bial in­fec­tions have dou­bled from 2010 to 2011 and in­creased again by 50% in 2012. Ad­di­tion­ally, the list of known polymi­cro­bial dis­eases, and the na­ture of mi­cro­bial com­mu­ni­ca­tion, is grow­ing.

There has def­i­nitely been a height­ened aware­ness both in the sci­en­tific and med­ical com­mu­ni­ties as we progress be­yond the nar­row but use­ful de­f­i­n­i­tion of a pathogen as de­fined by Koch's pos­tu­lates. But where is this field go­ing to be in five years? Most likely, there will be bet­ter di­ag­nos­tics to more read­ily as­sess mi­cro­bial com­mu­ni­ties in in­fected tis­sue. Al­ready, com­pa­nies have been striv­ing for rapid, ac­cu­rate, and cost-ef­fec­tive next-gen­er­a­tion se­quenc­ing of clin­i­cal sam­ples to bet­ter di­ag­nose com­plex bac­te­r­ial in­fec­tions. Just as im­por­tantly, we may be­gin al­ter­ing treat­ment reg­i­mens for in­di­vid­u­als with mul­ti­ple or­gan­is­mal in­fec­tions that are caused by an ar­ray of bac­te­ria, virus, fungi, and par­a­sites. Treat­ment with de­worm­ing med­ica­tions in helminth-in­fected pop­u­la­tions has un­for­tu­nately led to in­creased mor­bid­ity and spread of other pathogens, such as HIV and malaria. Con­se­quently, in­te­grated ap­proaches must be de­vel­oped based on the na­ture and sever­ity of spe­cific co-in­fec­tions. Al­though this is by no means a new field, the in­creased mind­ful­ness of the na­ture of polymi­cro­bial in­fec­tions should aid in staving off treat­ment-dri­ven an­timi­cro­bial re­sis­tance and strengthen the over­all global com­bat of in­fec­tious dis­ease.

And per­son­ally, if I were to at­tempt to cure the world of a zom­bie virus, I would tar­get the host im­mune sys­tem as op­posed to vol­un­tar­ily re­ceiv­ing a lethal in­fec­tion. What would you do?
 

 

Katy is a grad­u­ate stu­dent in Kelly Doran's lab in the San Diego State University/UCSD joint doc­toral pro­gram in mi­cro­bi­ol­ogy and a par­tic­i­pant in the 2013 UCSD/SDSU In­te­grated Mi­cro­bi­ol­ogy course.