Bad Phage Terms

∞ generated and posted on 2017.03.20 ∞

Various phage terms exist which are ambiguous or improperly used, some of which generally should not find their way into publications or even conversations, especially without being precisely defined at the outset… Here a list and discussion of such terms is provided.

Please cite as:

Stephen T. Abedon
Bac Phage Terms.
www.phage.org/writings/bad_terms.html

Multiple Meanings (Ambiguous Terms):

• Latent Period: This first item actually is a little trivial but nevertheless is, I think, worth being aware of. The issue is when exactly the latent period ends, having begun at the point of phage adsorption. For a single single cell the answer is easy, it is at the point of lysis (for lytic phages at least). For a population of phages, such as during single-step growth experiments, the answer is less obvious. Thinking in population terms, the answer is the point of first release of virion particles since the population literally is "latent" – at least from an historical perspective of what is going on inside of bacteria during phage infections – prior to this point of virion release. Alternatively, however, one can think instead in terms of populations of individuals, in which case the latent period can be the average point of lysis for the population or, as an approximation, halfway, either temporally or in terms of the phage rise. Thinking in more sophisticated terms, we can consider a distribution of lysis times across the population. Getting even more sophisticated, burst sizes will not be the same across an infection population and in fact might negatively correlate with overall infection duration (the latter which, I should note, is the exact opposite of how many people think about how burst sizes should correlate with infection durations, with the issue being that it is important to not confuse genetic predispositions with physiologically variation). These are all things that can be important when considering the mathematical/computer modeling of phage population growth. Minimally, these points should serve as food for thought at least in terms of appreciating that things often can consist of at least a bit more complexity than at first they may seem.
  
  
• Lysis From Without: This term I list twice. This first instance is because two more or less distinct phage-related literatures use this term but with slightly different meanings. Historically the phage literature has defined lysis from without as lysis – yes, lysis, not cell death, though that happens as well – which is associated with high multiplicity adsorption (yes, actually, demonstrably adsorbing virions), and which occurs well prior to the normal end point of a single phage infection. By contrast is the endolysin/lysin/enzybiotics literature where lysis from without refers to bacterial lysis – again, lysis, note the consistency of lysis as a component of lysis from without – which is a consequence of application of purified lytic enzymes to target bacteria. Though I am fully aware of the desire/tendency for those familiar with only one of these literatures to declare the other literature's use of this term as historically invalid given precedence of usage (yes, I'm looking at you, phage people), all I can say is, just get over it. But also, for the sake of avoiding confusion, at least be aware that these two definitions are out there.
  
  
• Secondary Infection: Secondary in a temporal sense means "after", but it turns out that there are two distinct ways that infections can come after infections. As an aside, yes, to some degree infection is not necessarily being used correctly in both instances and generally I try to be more precise by stating "secondary adsorption" rather than infection but, alas, only with regard to one of the two meanings… In any case, the distinctions are between what happens to individual cells versus populations of cells. Thus, secondary infection of an already phage-infected bacterium (i.e., secondary adsorption) is one meaning of secondary infection, and perhaps the less prevalent meaning. The second meaning has to do with the infection of bacteria by phages released from other bacteria. Yes, all phage infections occur as a consequence of infection by phages released from other bacteria, but from an epidemiological perspective, the first phage entering a population of bacteria would give rise to what can be described as a primary infection whereas all subsequent phage infections within the same bacterial population can be described as secondary. Yes it gets more confusing since secondary infections by both meanings can occur simultaneously, i.e., secondary infection of an infection which itself is a secondary infection, the latter in an epidemiological sense, or indeed as involving secondary infections of already lysogenically infected bacteria. For the most part, however, such complications can be ignored or, at least, only paid attention to by those who really care about this sort of stuff. More importantly, it is important to realize that when one states "secondary infection", not everybody is going to appreciate just what precisely you mean by "secondary infection".
  
  
• Virulent: The two or three meanings of virulent for phages all seem to stem from the same source and that meaning is equivalent to the more general meaning of virulent, that is, the capacity by a pathogen to do harm to a host organism. Indeed, the original meaning of virulent for a phage probably is a capacity of a phage to do harm especially to a bacterial culture. As a consequence, that degree of harm can vary even for a strictly lytic phage. Thus, phages which are less able to lyse broth cultures, or produce non-turbid plaques, are less virulent than phages which can readily lyse broth cultures and produce clear plaques. It probably was this perspective on phage virulence which led to the second meaning, which is as associated with the term virulent mutant. These are mutants of otherwise temperate phages which have an ability not only to form clear plaques but to do so using their parental lysogens as indicator bacteria. I'm guessing that virulent mutant has come to more generally be thought of simply as a temperate phage which has lost an ability to display lysogenic cycles. From there it likely was a easy creep to considering all non-temperate phages a virulent, which is where we stand today as a third meaning of virulent for phages. The result, of course, is a mess, and one that most researchers, I'm guessing, are not entirely aware of. Still, there is no way that we are going to get rid of the use of "Virulent" as a synonym for non-temperate phages, so the best we can hope for, probably, is to be able to recognize what an author means from context. Within the context of phage therapy, there is real utility in the concept of virulent as a more quantitative term (degree of virulence towards a bacterial culture), with greater anti-bacterial virulence desired. Unfortunately, virulent phages (i.e., non-temperate phages) are desirable as well, and barring that, presumably virulent mutants will do in a pinch, though presumably virulent as an ability to lyse even parental lysogens is preferable to virulent mutants which simply are not or, indeed, less temperate. Phage virulence, in other words, is something of a semantic cesspool. BTW, if anyone out there is more familiar with this history than I am, please enlighten me!
  

Poorly Used Terms:

• Episomal: An episome is a genetic element which can exist either integrated into a host chromosome or instead as a plasmid. Yes, some or indeed perhaps many temperate phage are episomal, but not all plasmid prophages necessarily also can exist as chromosome-integrated prophages. Therefore, and as much as I would prefer otherwise, it is best to not describe temperate phages which exist during lysogenic cycles as plasmid prophages as episomal. See prophage for more on plasmid prophages.
  
  
• Lambdoid Phage: A lambdoid phage, traditionally, is a phage that is able to recombine with phage λ, resulting in a functional, hybrid phage. For such recombination to work, there needs to be significant sequence similarity and gene synteny. More loosely, this property may be inferred from phage whole genome sequences. By contrast, "Lambdoid" is not synonymous with syphovirus, though, yes, phage λ is indeed a syphovirus.
  
  
• Lysogenic: Lysogenic literally means lysis generating. If a phage, or more broadly a virus does not have lytic, productive infections (e.g., is not a lytic phage), then it really does not make logical sense to describe their latent infections as lysogenic. Are people going to change their ways with regard to this usage? My guess is no. Nevertheless, at least we can have a small chuckle at their expense every time they describe a host infected with a chronically releasing virus as lysogenic. ;-)
  
  
• Lysis From Without: As alluded to above, the key word in lysis from without is lysis. In short, if lysis is not observed, and particularly premature lysis, then it just is not reasonable to describe or even speculate that lysis from without is being observed. More to the point, generally the adsorption of a single virion will result in the death of a bacterium. Therefore, even if many phages are expected to have adsorbed to a single bacterium, the death of that bacterium should not be used to imply that lysis from without has occurred, though that has not stopped many from doing so. But, ask yourself, what is the point? That is, what is being gained by claiming that the death of a population of bacteria is due to lysis from without versus not? The answer to that question is that lysis from without should not (by definition!!!) result in increases in phage numbers due to productive infection. This may have a pharmacokinetic impact on phage-mediated biocontrol. Few authors, however, at least explicitly make that point. So, again, why bother making claims for lysis from without? One answer, of course, is that using the term sort of makes one sound more sophisticated: "Oh, lots of phages may be present, so that must be lysis from without…" ("Gosh", listeners declare, "He/she must really know their phage literature!") Four other points are worth mentioning. The first concerns multiplicity of infection, which may not be what you think it is; click here for further consideration of that point. The second point has to do with spot testing, which some like to speak of in terms of lysis from without. To a degree they may be correct, but they are also possibly missing the point as spots are zones of inhibition rather than necessarily zones of bacterial lysis or even bacterial death. But of even greater relevance, it just can't be repeated too often than if one phage adsorption kills a bacterium, then so what if multiple phage adsorptions are killing a bacterium? Three, it is important to recognize that as a consequence of Poisson distributions it actually does require somewhat more than one adsorbing phage per bacterium to kill off a population of bacteria. Still, this is not lysis from without. Indeed, it is statistics! Lastly, phages which are known to be subject to lysis from without also, perhaps not surprisingly, tend to display a resistance to lysis from without which has the effect of making lysis from without less likely. That is, even if one is working with a phage for which lysis from without is possible and applying high multiplicities of phages, you still may not end up inducing lysis from without. Again, one cannot just claim that lysis from without is occurring on the basis of there being lots of phages and subsequent bacterial death. It is simply a lot more complicated than all of that.
  
  
• Lytic Phage: The problem here is that there are not all that many phages that are not lytic, so how is the qualifier "Lytic" useful? The answer, unfortunately, I think, is that people use "Lytic" as a qualifier for "Phage" to distinguish non-temperate phages (a.k.a., lytic?!?) from temperate ones (a.k.a., not lytic?!?!?). My guess is that this usage stems from introductory biology or microbiology where an absolutely correct distinction is made between "Lytic cycles" and "Lysogenic cycles", and this is somewhere in the back of people's minds when they become confused, somehow imagining that (most) temperate phages don't also display lytic cycles. That inference – what can I say? – is just plain wrong.
  
  
• Polyvalent: The trouble with polyvalent is that everyone knows exactly what it means, and at the same time nobody does. Mostly the term is used to indicate that a phage possesses a broad host range, thereby begging, of course, the question of why "broad host range" isn't a good enough descriptor… The problem that arises, however, is just what broad host range means. Is it a large number of strains within the same species? An ability to infect multiple species? An ability to infect multiple genera? Indeed, what exactly does host range mean anyway as this is dependent on the host range-determining assay used? In the end, it would be more helpful to just describe, in some details, a phage's host range. But as with a number of the terms discussed here, this is one which people seem to like, despite its imprecision, so I guess we're stuck with it.
  
  
• Superinfection Immunity/Superinfection Exclusion: Superinfection itself often is a misnomer in the phage literature since what people can mean instead is secondary adsorption, i.e., the attachment of a virion to an already phage-infected bacterium. Superinfection, by contrast, involves infection by that secondarily adsorbing phages, where infection is more than just attachment or adsorption. That being said, superinfection exclusion literally is prevention of superinfection, that is, cell-envelope/post-attachment mechanisms of phage-infected bacterium resistance to phage infection. Superinfeciton immunity, by contrast, is immunity given superinfection, i.e., just as most vaccines don't actually prevent infection but instead prevent disease, superinfection immunity, well, allows infection by prevents "disease" (i.e., phage infection of an already phage-infected bacterium). Some authors use these terms – superinfection exclusion and superinfection immunithy – interchangeable, thus I suppose indicating a lack of appreciation of the literature. Others use one term where they ought to be using the other (ditto!). So, if you are talking about temperate phage repressors and their impact on homoimmune phages, that's superinfection immunity. If you are not, and the secondarily adsorbing phage is being blocked at the cell envelope, then that's superinfection exclusion. Just think about what "exclusion" and "immunity" actually mean and there should be no issues as to which term applies. As an aside, I'm actually corresponding author of two encyclopedia articles on superinfection immunity presumably because I know a thing or two about superinfection exclusion but, alas, not so much about superinfection immunity.
  

Terms Used Incorrectly:

• Multiplicity of Infection: Phage multiplicity of infection (MOI), perhaps along with lysis from without, represents a case study in how information, even when written down, can degrade over time. Particularly, what we have with multiplicity of infection is a real-life game of telephone. The "original" meaning of MOI – quotes because it still is the meaning!!! – is literally what it says, though with a slight modification owing to increased understanding phage biology. That is, it is "Multiplicity" of "Infection", i.e., the number of phages which infect individual bacteria. The slight modification has to do with the meaning of "Infection", which more precisely might read "Adsorption", as in "Multiplicity of Adsorption". But that is a very small quibble. The bigger problem is that multiplicity of "Infection" is only multiplicity of "Addition" under a relatively narrow sent of circumstances. These are where bacterial densities are high (e.g., 10⁸ bacteria/ml), phage adsorption is highly efficient (i.e., not << 100% per encounter with a bacterium), and phage encounter with bacteria is highly likely (e.g., bacteria are not found in biofilms or food matrices). In the laboratory, while performing, for example, genetic experiments, one might literally cheat by assuming that "Infection" and "Addition" are the same thing, though the early literature explicitly warns that this assumption is not necessarily valid. Unfortunately, people took this cheat and ran with it, ultimately resulting in an important publication making a claim that the multiplicity of "Addition" was the original meaning of MOI, while they provided a "new" (it wasn't) means of calculating MOI that in actuality was the original, Phage Course meaning of MOI. The result is a huge mess where people imagine that they are adding sufficient numbers of phages to bacteria to result in substantial phage adsorpion ("MOIs of 1000!!!!") but in reality they don't have a clue. But this story gets worse. Somehow MOI was exalted to a status so great than even mention of phage titer turned out to be more than could be bothered with. So we are left with a bunch of people using MOI incorrectly and then not providing enough information for readers to even guess at what the real MOI might be. As I write this, my mind drifts to the thought that if science had comedy writers, could they possible come up with a skit more absurd? And yet, here we are, acting this out on a daily basis. It has to stop, really, as it's embarrassing…
  
  
• Phage Rise: The phage rise is the time either following or during the phage latent period that extracellular virions "rise" in numbers, such as during single-step growth experiments. It is not the period that spans from the end of the phage eclipse period to the point of bacterial lysis. Yes, phage numbers do rise there as well, but it's really confusing (to those who care about these things) to use the same term to describe two completely separate but nonetheless similar phenomena. Confusion in science really is not a good thing. The phage rise historically has one meaning and we should not use the fact that the post-eclipse period does not really have a legitimate name to cause us to substitute in the one term which unquestionably is not its name.
  
  
• Prophage (as 100% synonymous with integrated virus): Time and again people state with seemingly strong conviction that lysogenic cycles involve phage integration into host chromosomes, forming a prophage. As with a number of issues listed here, this is probably another example of repeating what was learned in introductory biology as though it were fact. Actually, in fact, temperate phages exist which do not integrate into the host chromosome but instead exist as plasmid prophages. Some of these prophages even exist as linear plasmids! The distinction between plasmids and prophages, in other words, can be a bit blurry, and certainly it just isn't reasonable to equate prophages with integration into host chromosomes.
  

Terms That Probably Should Not be Used:

• Carrier State: Carrier state is used by many different authors to mean different things. Even authors who seem as though they agree on what they mean by this term don't always seem to agree. The term also has been used synonymously with pseudolysogeny, which in my book puts it in company with disaster…
  
  
• Pseudolysogeny: Seriously, don't even get me started on pseudolysogeny. Just suffice it to say that if you use the term without first precisely defining it then almost nobody is going to know what you are talking about. Indeed, even if you do precisely define it you probably are going to have trouble. I have no idea what to repeace the term with, except to say that multiple terms are probably the answer, and please please please don't just make up new terms without first consulting the literature. Bottom line: think really really hard before tossing out this term. Each time I see it my first thought is… Well, you don't really want to know that, but suffice it to say that we're not talking about dopamine surges here. Exceptional, I should point out, is the work of Robert V. Miller which, in my opinion, should be reserved as the sole legitimate meaning of this term.
  

Terms That Seriously Should Not be Used:

• Adolescent Period: This term has been used as a synonym for the phage eclipse period. The idea is that prior to the completion of the first phage virion a phage is in some sort of juvenile stage whereas afterward they are, perhaps, now adults. These adults are still trapped inside of the infected bacterium that produced them (teenagers?). They then progress to that more adulty business of making a living, i.e., adsorption, whereupon they are turned into what? Adolescent infections? Preadolescent infections? Metaphorically, sure, but maybe not. But it is really difficult to see how useful this terminology may be and we certainly don't need new words for eclipse period.
  
  
• Lysogenic Phage: Bacteria are lysogens. Phages which can display lysogenic cycles are temperate. This usage, "Lysogenic phage", often is simply a mistake, one which is not worth repeating, though occasionally phrasing may be legitimate. But if you see "Lysogenic phage" being used all by itself, then you should be thinking "Temperate phage", and certainly not follow that author's lead.
  

Other:

• Bacterial Half Lives (in presence of phages): Yes, the phage impact on bacteria really is a function of phage titer, and the phage adsorption rate constant. It is not a function of bacterial density except to the extent that bacteria can deplete (or add to) phage densities, and it certainly is a problem to think of bacterial half lives such as during phage therapy in terms of a poorly defined multiplicity of infection. Click here for additional consideration.
  
  
• Bacteriophage Therapy (to the exclusion of "phage therapy"): Why not at least mention "Phage therapy" as a keyword even if you prefer to call it "Bacteriophage therapy"? I suppose the same thing can be said for "Bacteriophage therapy" as perhaps that should be included as a keyword as well if "Phage therapy" is your preference. Hey, just do what you can do so that people can find your papers! Click on the above or following two links to see the current PubMed hits for "Phage therapy" versus "Bacteriophage therapy", respectively. The following two icons provide links to the equivalent Google Scholar searches:
  
  
• Infection (when what you really mean is adsorption or virion attachment): It's just imprecise to state that a phage infected a bacterium when you have no evidence that infection actually occurred, but instead only that a phage has adsorbed/attached. Yes, this is a fine point, but has the potential for confusion. For example, see superinfection exclusion/superinfection immunity.
  
  
• Lytic Cycle (as 100% equivalent to productive infection): Yes, let's not forget that the productive cycles of some phages, i.e., chronically released/filamentous ones, is not lytic but instead virions are released continuously. I make this point because way too many authors make authoritative statements which sound suspiciously as though all phage productive cycles are lytic cycles. That's just something you learned in introductory biology! Also, don't forget that not even all temperate phages have lytic cycles, since some produce phages chronically. All temperate phages, however, do have productive cycles, else they aren't phages!! (Not functional ones, anyway…)
  
  
• Poisson Distributions (and probabilities of multiple adsorption): If you don't understand Poisson distributions, then you can't understand the likelihoods of multiple phage adsorptions, period. If you don't want to understand Poisson distributions, I've created an online calculator which will do the calculating for you, at least once it's calculated multiplicity of infection: moicalculator.phage.org.
  
  
• Spot Testing: Spot testing is a great technique, but is both limited in its utility and often misunderstood. First of all, unless effected directly by endolysins, it is not lysis from without. Second, a spot is a zone of inhibition, just like in antibiotic testing. What you are seeing for the most part with a spot, given 'confluent lysis', is the killing of bacteria prior to their forming a lawn (phages are, after all, added before the lawn has formed, so why shouldn't killing occur then as well?!?). Third, while a great technique, spot testing is only a preliminary means of phage host-range determination. Especially false positives are a concern. If publications claim to have determined phage host range but only present spotting data – or worse, do not describe how host range is determined at all!!! – then you should be somewhat disappointed. That is, you can't determine phage host range by spotting alone. At best you can present a first approximation of what possibly that host range might consist of.
  
  
• Virus (to the exclusion of "phage"): It's obvious why this is done as it often is sexier, or more fundable, to work on "viruses" versus "bacteriophages". Nevertheless, to publish an article on "bacterial viruses" that never mentions the word "phage" and/or "bacteriophage" represents an almost intentional effort to dissociate one's work from that of the rest of the phage community. If nothing else, this has the effect of potentially seriously reducing the size of one's audience. Why do it?