Attacking Biofilms: Another Quote, Plus Some Discussion

Stephen T. Abedon

Department of Microbiology – The Ohio State University

phage.org – phage-therapy.org – biologyaspoetry.org


 

This quote is from Lee Watkins and J. W. Costerton (1984). Growth and biocide resistance of bacterial biofilms in industrial systems. Chemical Times and Trends (October):35-40.

The article has nothing to do with viruses. The quote is interesting, however, since it speaks to the question of how exactly to employ viruses in the biocontrol of microorganisms, specifically in the biocontrol and indeed elimination of biofilms from surfaces (quotation marks in the original):

lt is important to be able to answer that old question “Shall we slug with a biocide, shall we continuously treat with a biocide or shall we soak with a biocide—what is the best deal for the situation?”

To me these three alternatives are distinguishable in terms of how we might think about treatment of bacteria or biofilms with phages, that is, phage-mediated biocontrol of bacteria, or phage therapy.

The first alternative I interpret as the application of large amounts of biocide over short periods, perhaps in a single dose, i.e., slugging, or what we might describe as passive treatment in the case of phage therapy. Keep in mind that passive treatment should mean that for every bacterium targeted not only should at least 10 phages be added but at least 10 phages should be adsorbing, per adsorbable bacterium.

Continuous application, to me, would imply the application of lower but still minimally effective concentrations of biocide over longer periods. Continuous application represents an extreme of multiple dosing, i.e., where the time gap between applications is reduced to zero. Key here is that something other than overwhelming amounts of biocide is being applied, what many (unfortunately) would describe as something other than high multiplicities of infection (MOI) in the case of phage application. One can view such continuous application a preventive, or prophylactic.

Lastly there is soaking, which could also be viewed as continuous application, though this is an application that takes place over a relatively short period, i.e., days or weeks rather than months or years. This would be equivalent to the application of phages by soaking bandages, soaking various absorbent material (one sees mention of “tampons” in various places in the phage therapy literature, though it’s important to realize that the word has a medical definition), or instead via the application of, e.g., Phage Bioderm (for example, as discussed here).

What’s missing, of course, are any assumptions that the biocide will replicate in situ, i.e., so-called active treatment, which is typically considered to be a hallmark of phage-mediated biocontrol/phage therapy. That absence, though, is not unexpected given that this is from a discussion of chemical and physical anti-biofilm biocides rather than of phages.

Still, it once again is nice to see that there really is little that is new under the sun. When dealing with bacterial infections, particularly chronic ones which are associated with biofilms, it can be important to keep in mind these ideas:

  1. We can hit them very hard (literally overkill) over short periods,
  2. We can hit them less hard (minimally adequate biocide concentrations) but over long periods, perhaps particularly towards prevention, or
  3. We can soak the infections over intermediate periods, presumably with periodic re-invigoration of dosing, using antibacterial levels which, also presumably, are somewhat in excess of what might be viewed as minimally adequate.

Any other approach, unless backed by hard data, should be considered to represent mostly wishful thinking.

Some additional reading:

Abedon, S.T. 2016. Bacteriophage exploitation of bacterial biofilms: phage preference for less mature targets?  FEMS Microbiol. Lett. 363:fnv246. [PubMed]

Abedon, S.T. 2016. Commentary: phage therapy of staphylococcal chronic osteomyelitis in experimental animal model.  Front. Microbiol. 7:1251. [PubMed]

Abedon, S.T. 2016. Phage therapy dosing: The problem(s) with multiplicity of infection (MOI).  Bacteriophage 6:e1220348. [PubMed]

Abedon, S.T. 2014. Bacteriophages as drugs: the pharmacology of phage therapy., p. 69-100. In J. Borysowski, R. Miedzybrodzki, and A. Górski (eds.), Phage Therapy: Current Research and Applications. Caister Academic Press, Norfolk, UK.

Abedon, S. 2011. Phage therapy pharmacology: calculating phage dosing.  Adv. Appl. Microbiol. 77:1-40. [PubMed]

Abedon, S.T., S.J. Kuhl, B.G. Blasdel, and E.M. Kutter. 2011. Phage treatment of human infections.  Bacteriophage 1:66-85. [PubMed]

Abedon, S.T. and C. Thomas-Abedon. 2010. Phage therapy pharmacology.  Curr. Pharm. Biotechnol. 11:28-47. [PubMed]

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Bacterial Lawns as Biofilm-Like Environments: A New Old Quotation

Stephen T. Abedon

Department of Microbiology – The Ohio State University

phage.org – phage-therapy.org – biologyaspoetry.org


 

Way back in 2010 we (Abedon and Thomas-Abedon) suggested that the growth of phage plaques within bacterial lawns could serve as mimics of bacteriophage interaction with bacterial biofiolms. In fact, we made a rather extensive argument with six Roman numeraled points: (i) constraint of bacterial movement, (ii) bacterial growth within lawns as microcolonies, (iii) inhibition of phage movement, (iv) plaque-like phage growth within actual biofilms, (v) possible temporary shielding of bacteria within lawn microcolonies from phage attack, and (vi) variation in bacterial physiologies again as found within microcolonies within lawns and as potentially equivalent to bacterial microcolonies within biofilms. We concluded that, “Given these similarities, phage plaques as a facile laboratory model therefore could enrich our understanding of phage-bacterial interrelations as they may occur during the phage therapy of biofilm-producing bacterial infections.”

Indeed, we noted as well that Gallet et al. (2009) described phage formation of plaques also as phage growth within a “biofilm-like environment”.

Here I provide a quote from an earlier publication which serves to further these arguments. From Gilbert and Brown (1995) [Mechanisms of the protection of bacterial biofilms from antibacterial agents, p. 118-130. In J. W. Costerton and H. Lappin-Scott (ed.), Microbial biofilms. Cambridge University Press, Cambridge, UK.], p. 119:

The most simple in vitro method of generating biofilms to study antimicrobial sensitivity is to inoculate the surface of an agar plate to produce a confluent growth. Such cultures, whilst not fully duplicating the in vivo situation, have been suggested to model the close proximity of individual cells to one another and the various gradients found in biofilms. In this respect, colonies grown on agar may bе representative of biofilms at solid-air interfaces.

Of course, one cannot claim that bacteria growing within soft agar overlays are perfect representations of naturally occurring biofilm structures. Nonetheless, as we’ve noted previously, e.g., Abedon and Yin (2009), plaque formation within them is a lot more complex than people otherwise may realize.

Phage Tails as Polymeric Substance Probes: Arguments For and Against

Stephen T. Abedon

Department of Microbiology – The Ohio State University

phage.org – phage-therapy.org – biologyaspoetry.org


 

Recently I suggested that:

Phage tails, to the extent that they display smaller diameters than phage virions as a whole, might contribute to nonenzymatic virion translocation into EPS [Extracellular Polymeric Substance, i.e. as associated with biofilm matrix], perhaps with longer, narrower tails permitting deeper or faster local penetration to biofilm-surface located bacteria.

Perhaps not unexpectedly, I now find that this was not an entirely original thought. From Wilkinson (1958), p. 68:

Physical blocking of the surface receptor. Can the presence of a capsule protect the cell simply because of its physical properties? Presumably an infective phage must be able to inject its DNA through the cytoplasmic membrane and, therefore, the main body of the phage must be at a distance from the cytoplasmic membrane smaller than the length of the phage tail (rarely longer than 150 mµ [meaning 150 nm]). Therefore, any layer outside the cytoplasmic membrane which is greater in thickness than 150 mµ and is impermeable to phages will act as a nonspecific phage inhibitor. It has already been shown that a capsule is by definition greater than 150 mµ in thickness and that
it is probably impermeable to particles of the size of a phage head (about 100 mµ). An additional barrier to the phage might be the high negative charge of the polysaccharide capsular surface. In confirmation of this role, capsulate bacteria have been reported to be generally phage resistant…

In my defense, my suggestion pointed specifically to longer phage tails, and the general thrust of my article was that it is especially less mature aspects of biofilms which may be more vulnerable to phages. Less maturity, in other words, might be associated with less thick biofilm matrix, e.g., as perhaps associated with new growth on biofilm surfaces.

Indeed, Wilkinson discusses further an article, unfortunately which is not in English, suggesting that when polymeric substance material is thinner then successful phage adsorption may be more likely (as continuing directly from the previous quote):

Thus, Kauffmann and Vahlne (82) found that most capsulate strains of E. coli were resistant to phage and that when capsulate strains were attacked, there was a proportionality between the thickness of the capsule and the resistance.

One could speculate therefore that it could be, conversely, that shorter tails would be less able to penetrate thicker “capsulate”.

It should be noted that I am making no claims that phage tails exist solely for the sake of penetrating extracellular polymers towards adsorption of bacteria, though it is entirely possible that such penetration could serve as a benefit of possessing especially “longer, narrower tails”.

Freezing Selects for Phage T7 Deletion Mutations… Not!

Stephen T. Abedon

Department of Microbiology – The Ohio State University

phage.org – phage-therapy.org – biologyaspoetry.org


 

Myth: Freezing phage T7 can select for deletion mutations…

From Clark and Geary (1973), with emphasis added:

ATCC freeze-dries phages for distribution (Clark and Geary, 1969). Davis and Hyman (1971) have reported the existence of at least two authentically different T7 phage strains, T7M (Meselsohn) and T7L (Luria) (ATCC 11303-B7). They also stated that T7L lyophilized stock from ATCC contained a high percentage of deletions in the DNA molecule, and they attributed this to lyophilization selecting for DNA deletions. However, since this publication, ATCC prevailed upon these authors to test the ATCC broth stock of T7L, which had been maintained at 4° C unlyophilized or unfrozen since its deposit in the Collection by Luria in 1952. They reported in a letter to W. A. Clark that the T7L stock before freeze-drying contained the same deletions as the freeze-dried product.

In other words, the evidence is not consistent with freezing selecting for deletion mutations in phage T7…

REFERENCES

Clark, W. A., and D. Geary. 1969. The collection of bacteriophages at the American Type Culture Collection, p. 179-187. In T. Nei (ed.), Freezing and Drying Microorganisms. University Park Press, Baltimore.

Clark, W. A., and D. Geary. 1973. Preservation of bacteriophages by freezing and freeze-drying. Cryobiology 10:351-360.

Davis, R. W., and R. W. Hyman. 1971. A study of evolution: The DNA base sequence homology between coliphages T3 and T7. J. Mol. Biol. 62:287-301.

d’Hérelle, F. (1918). Sur le rôle du microbe filtrant bactériophage dans la dysentérie bacillaire. Compt. rend. Acad. Sci. 167:970-972.

Stephen T. Abedon

Department of Microbiology – The Ohio State University

phage.org – phage-therapy.org – biologyaspoetry.org


 

I’ve been meaning to post machine-translated articles for some time now. Here is what you are seeing:

  • The posts are broken up by paragraphs with three sections per
  • The first section is the paragraph more or less retaining original page breaks (for easier reconciliation with the original document)
  • The second section is the paragraphs minus those page breaks
  • The third section, in bold, is the translation generated by Google with occasional help from a human being (e.g., moi)

Please feel free to make suggestions as to how to improve transcriptions and/or translations.

Oh, yes, and feel free to skip to the last “sentence” (actually, fragment), which, I think, is historically the most important point of the article.

BACTÉRIOLOGIE. – Sur le rôle du microbe filtrant bactériophage dans la

dysenterie bacillaire. Note de M. F. D’HÉRELLE, présentée par M. Roux.

 

BACTÉRIOLOGIE. – Sur le rôle du microbe filtrant bactériophage dans la dysenterie bacillaire. Note de M. F. D’HÉRELLE, présentée par M. Roux.

BACTERIOLOGY. – On the role of the filter microbe bacteriophage in bacillary dysentery. Note to Mr. F. D’HÉRELLE, by Mr Roux.

 

Dans une Note préliminaire (1) j’ai décrit un microbe filtrant trouvé

dans les déjections des convalèscents de dysenterie bacillaire. L’emploi

d’une technique moins imparfaite que celle dont je m’étais servi tout

d’abord (2) joint à l’examen systématique des selles de trente-quatre

malades, tous atteints de dysenterie à bacilles de Shiga, et dont plusieurs

ont pu être suivis journellement depuis le début de la maladie jusqu’à la fin

de la convalescence, m’ont permis d’etudier d’une manière plus complète

le mode d’action du microbe bactériophage et de préciser son rôle dans

l’évolution de la maladie.

 

Dans une Note préliminaire (1) j’ai décrit un microbe filtrant trouvé dans les déjections des convalèscents de dysenterie bacillaire. L’emploi d’une technique moins imparfaite que celle dont je m’étais servi tout d’abord (2) joint à l’examen systématique des selles de trente-quatre malades, tous atteints de dysenterie à bacilles de Shiga, et dont plusieurs ont pu être suivis journellement depuis le début de la maladie jusqu’à la fin de la convalescence, m’ont permis d’etudier d’une manière plus complète le mode d’action du microbe bactériophage et de préciser son rôle dans l’évolution de la maladie.

In a preliminary Note (1) I wrote [of] a filter[able] microbe found in excreta of dysentery bacillary [from] convalescents. The use of a technique less imperfect than the one I had used first (2) attached to the systematic examination of stools of thirty-four patients, all dysentery bacilli Shiga, and several of were followed daily from the beginning of the disease until the end of convalescence, allowed me to study in a more complete way the mode of action of the microbe bacteriophage and clarify its role in evolution of the disease.

 

(1) Comptes rendus, t. 165, 1917, p. 373.

(2) Comptes rendus de la Société de Biologie, séance du 7 décembre rg18.

 

Dans les cas de dysenterie bacillaire, même très graves, mais dans lesquels

l’état du patient s’améliore ‘rapidement, le microbe ,bactériophage

manifeste sa présence d’une· manière très active d’emblée, tant sur les cultures

du bacille isolé des déjections du patient que sur les souches du Shiga

du laboratoire, à partir du moment où les symptômes commencent à

s’amender. Le pouvoir bactériophage vis-à-vis du bacille dysentérique cesse

brusquement d’être décelable au début de la convalescence. A· partir de ce

moment, des examens répétés montrent également l’absence de bacilles

pathogènes.

 

Dans les cas de dysenterie bacillaire, même très graves, mais dans lesquels l’état du patient s’améliore ‘rapidement, le microbe ,bactériophage manifeste sa présence d’une manière très active d’emblée, tant sur les cultures du bacille isolé des déjections du patient que sur les souches du Shiga du laboratoire, à partir du moment où les symptômes commencent à s’amender. Le pouvoir bactériophage vis-à-vis du bacille dysentérique cesse brusquement d’être décelable au début de la convalescence. A partir de ce moment, des examens répétés montrent également l’absence de bacilles pathogènes.

In cases of bacillary dysentery, even very serious but in which the patient’s condition is improving quickly, the microbe, bacteriophage demonstrates its presence in a very active from the outset, both cultures of isolated bacillus excreta of the patient as the Shiga strains of laboratory, from the time symptoms begin to mend. Bacteriophage power vis-à-vis the dysentery bacillus suddenly ceases to be detectable in early convalescence. From that moment, repeated examinations also show the absence of pathogenic bacilli.

 

Dans les cas où la maladie se prolonge, le microbe bactériophage ne

manifeste qu’une action mille ou peu marquee, tant que l’état du patient

reste stationnaire. Si, dans quelques cas, l’action bactéricide est reiativement élevée sur les souches ayant subi de nombreux passages sur les

milieux de culture, par contre, elle est toujours inappréciable ou très faible

sur les cultures du bacille provenant du malade en observation. L’amélioration

se manifeste dès que l’action bactériophage devient énergique vis-à-vis de ce dernier.

 

Dans les cas où la maladie se prolonge, le microbe bactériophage ne manifeste qu’une action mille ou peu marquee, tant que l’état du patient reste stationnaire. Si, dans quelques cas, l’action bactéricide est relativement élevée sur les souches ayant subi de nombreux passages sur les milieux de culture, par contre, elle est toujours inappréciable ou très faible sur les cultures du bacille provenant du malade en observation. L’amélioration se manifeste dès que l’action bactériophage devient énergique vis-à-vis de ce dernier.

In cases where the disease is prolonged, the microbe bacteriophage manifesto that an action or a thousand little marquee as long as the patient remains stationary. If, in some cases, the bactericidal action is relatively high stem having undergone many passages on culture media, by cons, it is always invaluable and very low on the cultures of the bacillus from the observation sick. The improvement was evident as soon as the bacteriophage action becomes strong vis-à-vis the latter.

 

Dans les formes de longue durée et à rechutes, le pouvoir bactériophage

du microbe filtrant peut, à certains moments, être très énergique vis-à-vis

des bacilles de culture et variable d’un jour à l’autre, quoique toujours

relativement faible, vis-à-vis du bacille du malade. La guérison suit de près

le moment où l’action du microbe bactériophage se manifeste d’une manière

aussi intense pour l’une comme pour l’autre souche. Cette action persiste;

avec des fluctuations dans l’activité, aussi longtemps que le patient reste

porteur de germes. Ce dernier fait serait même de nature à faciliter le dépistage

des porteurs de germes, la mise en évidence du microbe bactériophage

étant plus simple et plus sûre que la recherche du bacille pathogène dans

les selles.

 

Dans les formes de longue durée et à rechutes, le pouvoir bactériophage du microbe filtrant peut, à certains moments, être très énergique vis-à-vis des bacilles de culture et variable d’un jour à l’autre, quoique toujours relativement faible, vis-à-vis du bacille du malade. La guérison suit de près le moment où l’action du microbe bactériophage se manifeste d’une manière aussi intense pour l’une comme pour l’autre souche. Cette action persiste; avec des fluctuations dans l’activité, aussi longtemps que le patient reste porteur de germes. Ce dernier fait serait même de nature à faciliter le dépistage des porteurs de germes, la mise en évidence du microbe bactériophage étant plus simple et plus sûre que la recherche du bacille pathogène dans les selles.

In the forms of long-term and relapsing, the bacteriophage of the filter microbe authority may, at times, be very aggressive vis-à-vis the bacilli culture and varies from one day to another, though still relatively low, vis-à-vis the bacillus of the patient. Healing closely when the action of the microbe bacteriophage manifests an intense way for the one as for the other strain. This action persists; with fluctuations in activity, as long as the patient remains buoyant germs. The latter would be able to facilitate the screening of carriers, the detection of the bacteriophage microbe is simpler and safer than the search for the pathogenic bacterium in the stool.

 

J’ai pu vérifier que , l’action du microbe bactériophage était prépondérante,

non pas seulement en ce qui touche à la disparition du bacille dysentérique

de l’intestin une fois la maladie déclarée, mais encore lors de son

éclosion. Au cours de la récente épidémie, j’ai eu l’occasion d’observer

plusieurs cas extrêmement bénins dans lesquels les symptômes se limitèrent

à quelques épreintes et à deux ou trois selles diarrhéiques: or, dans

tous ces cas, le microbe bactériophage fut, dès le début, présent et doué

d’un pouvoir antagoniste élevé. Malgré la bénignité de l’affection, il s’agissait

bien de dysenterie car, dans trois de ces cas, je pus isoler de la

première selle diarrhéique émise un bacille de Shiga typique.

 

J’ai pu vérifier que , l’action du microbe bactériophage était prépondérante, non pas seulement en ce qui touche à la disparition du bacille dysentérique de l’intestin une fois la maladie déclarée, mais encore lors de son éclosion. Au cours de la récente épidémie, j’ai eu l’occasion d’observer plusieurs cas extrêmement bénins dans lesquels les symptômes se limitèrent à quelques épreintes et à deux ou trois selles diarrhéiques: or, dans tous ces cas, le microbe bactériophage fut, dès le début, présent et doué d’un pouvoir antagoniste élevé. Malgré la bénignité de l’affection, il s’agissait bien de dysenterie car, dans trois de ces cas, je pus isoler de la première selle diarrhéique émise un bacille de Shiga typique.

I could verify that the action of the bacteriophage was dominant microbe, not only as it relates to the disappearance of dysentery bacillus bowel disease once declared, but during its outbreak. In the recent outbreak, I had the opportunity to observe several extremely mild cases in which symptoms were limited to a few tenesmus and two or three loose stools: gold, in all these cases, the microbe was bacteriophage, from the beginning, and now endowed with a high antagonistic power. Despite the mildness of the disease, it was good of dysentery because in three of these cases, I could isolate the first loose stool issued a typical Shiga bacillus.

 

Le microbe bactériophage préexiste dans l’intestin ou il vit narmalement

aux dépens du B. coli. Dans les selles normales, son pouvoir antagoniste

vis-à-vis de ce dernier bacille est toujours très faible; il peut devenir

considérable dans divers états morbides, dans certaines formes d’entérites

et de diarrhée banales, par exemple. La présence de bacilles dysentériques

dans l’intestin détermine en premier lieu une exaltation considérable de

la virulence du microbe bactériophage vis-à-vis du B. coli, puis, par une

accoutumance plus ou moins rapide, cette virulence s’exalte vis-à-vis du

bacille dysentérique; elle atteint d’emblée ou graduellement une puissance

considérable amenant la disparition rapide ou graduelle du bacille pathogène. Si la virulence du microbe bactériophage s’exalte d’emblée, les

bacilles dysentériques sont détruits dès le début de leur culture dans le

contenu intestinal, la maladie avorte avant tout symptôme ou se borne à

quelques troubles passagers. Si, pour une cause qui reste à déterminer, la

virulence du microbe bactériophage vis-à-vis du microbe pathogène ne se

manifeste pas d’emblée ou ne se manifeste que faiblement, une lutte s’établit

entre les deux organismes, les bacilles dysentériques se multiplient

dans le contenu intestinal, infiltrent la’ muqueuse, la maladie èclate et

l’état du patient enregistre ensuite fidèlement les fluctuations de la lutte.

 

Le microbe bactériophage préexiste dans l’intestin ou il vit narmalement aux dépens du B. coli. Dans les selles normales, son pouvoir antagoniste vis-à-vis de ce dernier bacille est toujours très faible; il peut devenir considérable dans divers états morbides, dans certaines formes d’entérites et de diarrhée banales, par exemple. La présence de bacilles dysentériques dans l’intestin détermine en premier lieu une exaltation considérable de la virulence du microbe bactériophage vis-à-vis du B. coli, puis, par une accoutumance plus ou moins rapide, cette virulence s’exalte vis-à-vis du bacille dysentérique; elle atteint d’emblée ou graduellement une puissance considérable amenant la disparition rapide ou graduelle du bacille pathogène. Si la virulence du microbe bactériophage s’exalte d’emblée, les bacilles dysentériques sont détruits dès le début de leur culture dans le contenu intestinal, la maladie avorte avant tout symptôme ou se borne à quelques troubles passagers. Si, pour une cause qui reste à déterminer, la virulence du microbe bactériophage vis-à-vis du microbe pathogène ne se manifeste pas d’emblée ou ne se manifeste que faiblement, une lutte s’établit entre les deux organismes, les bacilles dysentériques se multiplient dans le contenu intestinal, infiltrent la’ muqueuse, la maladie èclate et l’état du patient enregistre ensuite fidèlement les fluctuations de la lutte.

The microbe bacteriophage pre-exists in the intestine where it normally lives at the expense of B. coli. In normal stool, his antagonist power vis-à-vis the latter bacillus is still very low; it can become considerable in various disease states, in some forms of enteritis and diarrhea mundane, for example. The presence of dysentery bacilli in the intestine first determines considerable exaltation of the virulence of the microbe bacteriophage vis-à-vis the B. coli, and by a more or less rapid habituation, this virulence exalts vis-à -vis the dysentery bacillus; she reached immediately or gradually considerable power causing the rapid or gradual disappearance of pathogenic bacillus. If the virulence of the microbe bacteriophage exalts the outset, the dysentery bacilli are destroyed at the beginning of their culture in the intestinal contents, disease aborted before any symptoms or merely some temporary disturbance. If, for reasons yet to be determined, the virulence of the bacteriophage vis-à-vis the pathogen microbe does not manifest itself immediately or appears only faintly, a struggle takes place between the two organizations, dysentery bacilli multiply in the intestinal content, infiltrate the mucosal, the disease breaks out and the patient then records faithfully the fluctuations of the struggle.

 

En résumé, la pathogénie et la pathologie de la dysenterie pacillaire sont

dominées par deux facteurs agissant en sens contraire: le bacille dysentérique,

agent pathogène, et le microbe filtrant bactériophage, agent d’immunité.

 

En résumé, la pathogénie et la pathologie de la dysenterie pacillaire sont dominées par deux facteurs agissant en sens contraire: le bacille dysentérique, agent pathogène, et le microbe filtrant bactériophage, agent d’immunité.

In summary, the pathogenesis and pathology of dysentery pacillaire are dominated by two factors working in opposite directions: the dysentery bacillus, pathogen, and the filter microbe bacteriophage immunity agent.

 

Comme corollaire, l’expérimentation sur le lapin montre que les cultures

du microbe bactériophage jouissent d’un pouvoir préventif et curatif

dans la maladie expérimentale; d’autre part, le microbe bactériophage se

trouve invariablement présent dans l’intestin des malades dès que les symptômes s’amendent; il semble donc logique de proposer comme traitement

de la dysenterie bacillaire l’administration, dès l’apparition des premiers

symptômes, de cultures actives du microbe bactériophage.

 

Comme corollaire, l’expérimentation sur le lapin montre que les cultures du microbe bactériophage jouissent d’un pouvoir préventif et curatif dans la maladie expérimentale; d’autre part, le microbe bactériophage se trouve invariablement présent dans l’intestin des malades dès que les symptômes s’amendent; il semble donc logique de proposer comme traitement de la dysenterie bacillaire l’administration, dès l’apparition des premiers symptômes, de cultures actives du microbe bactériophage.

As a corollary, testing on rabbits showed that cultures of bacteriophage microbe enjoy a preventive and curative power in the experimental disease; secondly, the microbe bacteriophage is consistently present in the intestine of patients as soon as symptoms make amends; so it seems logical to propose as a treatment for shigellosis administration, from the onset of symptoms, active cultures of the microbe bacteriophage.

 

 

 

Cages of phages: improved production of hydrogen by enzymes encapsulated in phage capsids

A guest post by Dr. Paul Hyman,
Ashland University, Ashland, OH, USA
phyman@ashland.edu

Bacteriophage capsids have been studied as frameworks for the development of new materials. In part this is an outgrowth of phage display which allows the precise placement of binding elements on the phage surface. But another approach is to use the native capsids and to nonspecifically attach conductive metals to create wire-like structures or to capture and encapsulate molecules in the capsids as the capsids assemble. In a recent paper in Nature Chemistry, Trevor Douglas’ group at Indiana University and colleagues have shown that the efficiency of an enzyme that reduces protons to form hydrogen gas is more efficient and stable when enclosed in a bacteriophage capsid.

The bacteriophage they used is the Salmonella phage P22.  P22’s capsid is an icosahedron composed of a major coat protein assembled onto a scaffold protein. Coat protein and scaffold protein self-assemble into a procapsid that during a normal infection is then packaged with the phage DNA.  The researchers fused the two subunits of a hydrogenase protein to separate scaffold protein genes.  When expressed together with coat protein, procapsid self-assembled with the heterodimeric hydrogenase protein inside as outlined in this figure.

Hyman blog post

(modified from Jordan 2015)

When they tested for hydrogenase activity, the highest efficiency was found if the scaffold protein/hydrogenase subunits were expressed several hours before the coat protein (line one in the table).  This pre-encapsulation period presumably allowed the two hydrogenase subunits to assemble into the active heterodimer before being constrained by the coat protein.

pH 5 pH 8
P22 encapsulated with sequential expression 6118 nmol H2/mg min 3218 nmol H2/mg min
P22 encapsulated with simultaneous expression 757 nmol H2/mg min
Unencapsulated hydrogenase + scaffold without coat protein 46 nmol H2/mg min 12.6 nmol H2/mg min
Free hydrogenase 12-38 nmol H2/mg min (pH not specified)

Data from Jordan 2015

Additional experiments showed that the encapsulation also partially protected the hydrogenase against trypsin, heat denaturation (60°C for 45 min.) and air exposure.

The reason for the increased enzymatic activity is not entirely clear.  The increased activity and protection results suggest that the enzyme’s quaternary structure is stabilized in some way in the capsid.  It may also be that enzyme efficiency is higher when several hundred copies of the enzyme are in close proximity to each other in some sort of synergistic effect.

Overall, this paper demonstrates another way that phages can be used in non-biological technologies as well as biological.  Independent of the phage aspect, an improved catalyst for production of hydrogen gas could prove quite valuable as alternative fuels, such as hydrogen, are increasingly sought after.

Reference: Paul C. Jordan, Dustin P. Patterson, Kendall N. Saboda, Ethan J. Edwards, Heini M. Miettinen, Gautam Basu, Megan C. Thielges, and Trevor Douglas, “Self-Assembling Biomolecular Catalysts for Hydrogen Production”, Nature Chemistry doi:10.1038/nchem.2416, published on-line December 21, 2015.

Archaeal Virus as Cat Toy

Stephen T. Abedon

Department of Microbiology – The Ohio State University

phage.org – phage-therapy.org – biologyaspoetry.org


 

This post is/was inspired by Stedman, K. M., M. DeYoung, M. Saha, M. B. Sherman, and M. C. Morais. 2015. Structural insights into the architecture of the hyperthermophilic fusellovirus SSV1. Virology 474:105-109. (ncbi.nlm.nih.gov/pubmed/25463608)

Turns out this spindle-shaped virus of Sulfolobus, potentially resembling “two fused fullerene cones”, is an almost perfect prolate spheroid, which (apparently) is the shape of a pig’s bladder, which in turn is the shape of the American football.

The virus itself, however, possesses a “hexameric tail”, which sort of looks like streamers emanating from one end of the “football”. And that, in turn, sure looks an awful lot like this, particularly were the tail trimmed:

http://www.entirelypets.com/jw-pet-cataction-football-streamers.html

“Art” imitates life belongs to cats!

For my renderings on this subject – archaeal virus as football – see:

https://www.facebook.com/pages/Bacteriophage-Ecology-Group/111721928901953

 

 

The ecology of viruses that infect eukaryotic algae

The world of algal viruses has only gotten more interesting in recent years as it has become increasingly clear just the sheer quantity of mortality they cause and how much that drives global nutrient cycling in ways we’ve been largely blind to.

Chlorovirus

The ecology of viruses that infect eukaryotic algae

Because viruses of eukaryotic algae are incredibly diverse, sweeping generalizations about their ecology are rare. These obligate parasites infect a range of algae and their diversity can be illustrated by considering that isolates range from small particles with ssRNA genomes to much larger particles with 560 kb dsDNA genomes. Molecular research has also provided clues about the extent of their diversity especially considering that genetic signatures of algal viruses in the environment rarely match cultivated viruses. One general concept in algal virus ecology that has emerged is that algal viruses are very host specific and most infect only certain strains of their hosts; with the exception of viruses of brown algae, evidence for interspecies infectivity is lacking. Although some host–virus systems behave with boom-bust oscillations, complex patterns of intraspecies infectivity can lead to host–virus coexistence obfuscating the role of viruses in host population dynamics. Within the framework of population dynamics, host density dependence is an important phenomenon that influences virus abundances in nature. Variable burst sizes of different viruses also influence their abundances and permit speculations about different life strategies, but as exceptions are common in algal virus ecology, life strategy generalizations may not be broadly applicable. Gaps in knowledge of virus seasonality and persistence are beginning to close and investigations of environmental reservoirs and virus resilience may answer questions about virus inter-annual recurrences. Studies of algal mortality have shown that viruses are often important agents of mortality reinforcing notions about their ecological relevance, while observations of the surprising ways viruses interact with their hosts highlight the immaturity of our understanding. Considering that just two decades ago algal viruses were hardly acknowledged, recent progress affords the optimistic perspective that future studies will provide keys to unlocking our understanding of algal virus ecology specifically, and aquatic ecosystems generally.

Data Storage and Standard Parts

Genetic data storage, scaleable cell-cell communication, and still-better gene expression, all thanks to phage!

Cross-talk between Diverse Serine Integrases

Abstract
Phage-encoded serine integrases are large serine recombinases that mediate integrative and excisive site-specific recombination of temperate phage genomes. They are well suited for use in heterologous systems and for synthetic genetic circuits as the attP and attB attachment sites are small (< 50 bp), there are no host factor or DNA supercoiling requirements, and they are strongly directional, doing only excisive recombination in the presence of a recombination directionality factor. Combining different recombinases that function independently and without cross-talk to construct complex synthetic circuits is desirable, and several different serine integrases are available. However, we show here that these functions are not reliably predictable, and we describe a pair of serine integrases encoded by mycobacteriophages Bxz2 and Peaches with unusual and unpredictable specificities. The integrases share only 59% amino acid sequence identity and the attP sites have fewer than 50% shared bases, but they use the same attB site and there is non-reciprocal cross-talk between the two systems. The DNA binding specificities do not result from differences in specific DNA contacts but from the constraints imposed by the configuration of the component half-sites within each of the attachment site DNAs.

New Applications for Phage Integrases

Within the last 25 years, bacteriophage integrases have rapidly risen to prominence as genetic tools for a wide range of applications from basic cloning to genome engineering. Serine integrases such as that from ϕC31 and its relatives have found an especially wide range of applications within diverse micro-organisms right through to multi-cellular eukaryotes. Here, we review the mechanisms of the two major families of integrases, the tyrosine and serine integrases, and the advantages and disadvantages of each type as they are applied in genome engineering and synthetic biology. In particular, we focus on the new areas of metabolic pathway construction and optimization, biocomputing, heterologous expression and multiplexed assembly techniques. Integrases are versatile and efficient tools that can be used in conjunction with the various extant molecular biology tools to streamline the synthetic biology production line.