Microbiome yarns: the Global Phenotype–Genotype Survey

Dr. Noitall-Most : Yes: we already knew that our microbiomes seem to influence several health-related aspects of our biology, but the magnitude and range of human characteristics suggested by the survey to be influenced by the microbiome is just mind boggling. And now, one year later, we have the results of the trials carried out to test the predictions of the correlations found in the survey. The trials indeed established a number of really interesting human characteristics that are significantly influenced by our microbiome. A taste of this was given in the previous programme when I revealed that a new type of bacterium found in the oral cavity seems to influence diction.

Dr. Noitall-Most : Yes: we already knew that our microbiomes seem to influence several health-related aspects of our biology, but the magnitude and range of human characteristics suggested by the survey to be influenced by the microbiome is just mind boggling. And now, one year later, we have the results of the trials carried out to test the predictions of the correlations found in the survey. The trials indeed established a number of really interesting human characteristics that are significantly influenced by our microbiome. A taste of this was given in the previous programme when I revealed that a new type of bacterium found in the oral cavity seems to influence diction. 6 Ms. Repor-Tastory : Yes, Ani, that was absolutely amazing: please tell us more.
Dr. Noitall-Most : Right! This evening I should like to share with viewers new information on the influence of laryngeal microbes on vocal quality.
Firstly, as we know, smokers tend to have gravelly voices and habitually enjoy a recreational, somewhat phlegmy cough-athon after emerging from their slumber. This comforting, clearing-of-the-throat ritual was previously thought to be due to smoke-provoked irritation and inflammation of the larynx, and overnight accumulation of laryngeal mucous, but GLOPS and subsequent trials based on its findings have consigned this theory to the bin. Metagenome analysis revealed that smokers, but not non-smokers, have a particular bacterium in their laryngeal microbiota. This finding induced the Imaging Group of Mabriella Golinari and Ranfredy Mohde of the Walpur Gisnacht Institute for Cellular Pathology in Bad Hurzbarg in Northern Germany, 7 which was responsible for microbe isolation and characterization in GLOPS, to attempt to culture and characterise this bug.
Mab and Ran initially had no success using normal isolation media, but subsequently were able to tease it to grow on a special medium containing nicotine. It subsequently transpired that this microbe is nicotine-dependentyes, really: nicotine-addicted!and able to degrade and use nicotine as a food source. 8 They named it Seudomonas nicotinadicta, which was immediately re-named Snica by young smokers. Mab and Ran showed that Snica is distributed throughout the buccal cavity of smokers and is particularly dense on vocal chords.
Ms. Repor-Tastory : Gosh: imagine eating nicotine! Dr. Noitall-Most : Trueit does not bear thinking about! But of course microbes do not necessarily share our culinary tastes and, in reality, collectively they will eat most anything.
Nevertheless, nicotine is toxic for most bacteria, 9 as indeed it is to most types of cell, which is why it is so bad for our health. 10 Apparently, smoking toxicity produces changes in the laryngeal microbial community, 11 which becomes dominated by Snica. Snica continuously degrades and removes nicotine from the oral mucosa, and thereby reduces the exposure of other mucosal microbes to nicotine toxicitya sort of bioprotection service 12 it provides to the oral microbiota. Snica is also resistant to the toxic effects of nicotine, partly due to metabolic and oxidative stress responses, 13 and partly due to smoke-induction of the biofilm mode of growth, 13 which is the hunker-down mechanism microbes adopt when challenged by toxic insults, 13 such as antibiotics and immune defences, that enables them to avoid and resist antimicrobial activities. But, for the discussion this evening about oral microbiota and vocal quality, a key thing about Snica is that it is a prolific slime producer. 14,15 During the day, this slime is unconsciously swallowed, so is not noticeable, but during sleep it accumulates and ultimately triggers the resonant fruity bout of wake-up-to-the-joys-of Spring coughing that greets the smoker's daybreak. Importantly for our discussion today, the continual slime production also has the effect of changing the resonance of the vocal chords, such that the sound output has the gravelly timbre typical of the smoker.
Ms. Repor-Tastory : Well, that is an eye-opener: so a slimy bug in the throat is at the bottom of the deep voice! Dr. Noitall-Most : So it would seem. Anyway, staying on the subject of vocal harshness: whereas this is mostly a characteristic of smokers, it is also found in non-smokers and can be either inherited from a gruffy-voiced parent, or acquired, for example through perpetual shouting, as evidenced by non-smoking sergeant majors with typical parade ground timbres. In fact, GLOPS did suggest the presence of a non-Snica-type bacterium in the buccal cavity of non-smokers with harsh voices that was lacking in those blessed with mellow voices. Encouraged by their success with Snica, Mab and Ran wanted to complete their analysis of microbial involvement in vocal harshness and were, with some difficulty, able to isolate this bug. They named it Grufficus cockeri, which was immediately rechristened Gruffy by the music industry, and showed it to inhabit principally the surface of the epiglottis and, in some cases, also the vocal chords. It is present in large numbers in people with harsh voices and low numbers in those with euphonius voices. Its very obvious characteristic is that it forms macrofibres: when Gruffy is growing, newly-formed cells do not separate but remain attached to one another, forming a long chain which, in time, repeatedly folds back on itself. As cells continue to grow and form daughters, the force generated causes the filament to twist and form a socalled macrofibre. 16 Mab and Ran suggest that Gruffy macrofibres create a net-like structure that acts like a dishcloth in the voice box, collecting all manner of flotsam floating and flying around the buccal cavity, and thereby distorting the sounds emanating through the vocal chords, much like the electronic distortion of guitar notes produced by fuzz pedals 17 and the like.
Ms. Repor-Tastory : Well, on thinking about it, my throat also feels as if it contains a dishcloth, the morning after a long night of clubbing.
Dr. Noitall-Most : Yes, but that is something different and probably your state of dehydration after excessive alcohol consumption.
Anyway, to continue: a limited trial funded by a well-known music label involved the daily breakfast consumption of yoghurt containing G. cockeri by country singers with dulcet voices who were desperate to acquire gravelly voices so that they could sing their preferred musichard rock or heavy metalwithout needing to develop a smoking habit. In almost all cases, within one week, they were singing raucously like Joe Cocker 18 or Janis Joplin. 19 Ms. Repor-Tastory, amused enlightenment slowly spreading across her face : Oohhh, this explains the sharp deepening of my voice after a wild fling in Nashville with a hunky, husky bluegrass banjo player that took ages. . .... . .. . .(angry noises emanating from the in-ear headphone). . .Oh, sorry, I did not mean to interrupt. . .
Dr. Noitall-Most : Yes, wild flings can be followed by all sorts of surprises, not only deliveries of red roses.
Dr. Noitall-Most : But while the transfer of oral bugs between consenting adults is one thing, the unknown and perhaps unwanted transfer of such bugs is another. You will recall that a major focus of GLOPS was the transfer of microbiota among people and the phenotypic consequences of this? Well, one aspect of this related to tonight's topic was an assessment of the transfer of airway microbiota by pillows among travellers. You know the situation: those of us who sleep on our fronts or sides breathe, cough, sneeze and dribble into our hotel pillows for 8-h stretches, vigorously propelling our airway-oral bounties into their depths. Now while the pillowcases are regularly laundered and changed for every new guest, the pillows generally are not. And, those of us who sleep on our fronts or sides not only blow our stuff into pillows, we also inhale and, in the case of pillows used by others, inhale their stuff. Previously, it was thought that the post-travel respiratory issues we not infrequently experience were due to the drying out of the airway mucosa in low humidity aeroplane cabins or to infections gifted by other travellers in confined spaces, like elevators. But now we know that hotel pillows can also be a rich source of diverse microbes that enrich our airway microbiota, sometimes with unexpected, sometimes unwanted, results.
Ms. Repor-Tastory : Hhmmm! So: sleeping on our backs, or taking our own favourite pillow, populated with our own microbiota, to enjoy slumber comfort on our travels, also has the benefit of protecting our oral microbiota from invasion by those of others? I think I'll take my own pillow next time I am on the road! Oh, by the way, do microbes have anything to do with boys' voices breaking in puberty? Dr. Noitall-Most : Aahhh, that is a fascinating question, but we will leave it for another episode dealing with the Y-chromosome-specific microbiota. However, voice breaking is most dramatic when singing, and the survey did make a remarkable discovery about oral microbiota and singing voice purity. Although the samples in the stratification were small, there did seem to be a clear relationship between the presence of a new laryngeal bacterium and voice purity in professional singers, particularly opera singers. This bacterium was readily isolated by Mab and Ran from the buccal cavities of several stars, the identities of which have not been revealed, in anticipation of future commercialisation of this bug and the creation of new brands based on the names of famous singers. Amazingly, when cells of this bacterium were added to vocal chords excised from fresh cadavers, they bound tightly and spontaneously reorganised themselves in such a way that they increased the tension in and caused contraction of the chords. This was slightly disconcerting for Mab and Ran looking down the microscope at this happening in real time, given that the chords were nominally dead! The bug was given the name Harmonicoccus sublimiae. 20 Ms. Repor-Tastory : Gosh! Does Harmoniwhatshernamecontaining yoghurt improve singing voices? Most Fridays, a close girlfriend and I, both dressed to kill, go to a rather upmarket karaoke bar and it would be nice to have a bit of an edge over the competition. . ..
Dr. Noitall-Most : That is of course a key question, but the trial has not yet been concluded.
Ms. Repor-Tastory : Hmm -I'll have to get from you the coordinates of the trial leader during our post-transmission margarita.
Dr. Noitall-Most : Certainly. Another rather curious finding was that, rather unexpectedly, individuals involved in the business of persuasionspin doctors, politicians, lobbyists, ad(wo)men, salespersonsespecially those selling automobiles and real estate, industry consultants, fundraisers, gurus of all types, but especially lifestyle and diet gurus, vloggers, 21 and celebrity chefs, tend to have a somewhat atypical tongue microbiota containing significant numbers of a bacterium that was extremely difficult to isolate in pure culture, though it grew happily in a mixed culture containing other microbes from the buccal cavity. However, the highly experienced German pair finally obtained two isolates by extinction dilution in a rich medium containing traces of silver nitrate. When they characterised the bugs, they found that they have an exceptionally high affinity for silver ions in food and drink consumed, 22 which they convert to cell-bound silver nanoparticles, 23 giving the tongue a slightly silvery appearance. They named them Silvertonguium guruis, or Silg, and S. uberdentischzieha, or Sudz. 24 Curious to know if Silg and Sudz transplantation could improve the rhetorical ability of recipients, a small trial was carried out, but it was quickly aborted when it became apparent that Silg and Sudz failed to colonize and survive on the tongues of most donors.
Ms. Repor-Tastory : So, the adage that people with the power of persuasion have silver tongues is true after all! But why don't transplants of Silg and Sudz work?
Dr. Noitall-Most : Well, the explanation offered by the trial consultant, Professor Tim Kennis of the Queenton Institute of Advanced Studies is that exceptional rhetorical ability in certain individuals 25 appears to have a genetic component that predetermines not only development of oratorical skills but also predisposes such individuals to lifestyles involving persuading other members of society.
He considers it to be entirely plausible that this genetic trait simultaneously modifies normal tongue physiology such that it creates an attractive environment for Silg and Sudz. To explore this possibility, he carried out a small study on buccal fluids from Sudz-positive andnegative individuals and found that Sudz grew happily in fluid from Sudz-positive individuals but hardly at all in fluid from Sudz-negative individuals, which supports the view that the tongue physiology of persuaders is different from that normal folk. He has carried out bioinformatic analyses of the genomes of Sudz and its relatives and has shown that, in terms of human evolution, silver nanoparticle production by Sudz and its predecessors is a relatively ancient trait, but that it is only found in human-colonising strains, not in animal-colonising relatives. He suggests that humans have always valued the ability to persuade and direct the views of their fellow men and women, and that the genetic trait that favours this ability has co-evolved with Silg/Sudz ancestors over time to increase the efficacy of this ability. He suspects that if we could sample the tongues of the great orators, 25 like Aristotle, Cicero, Henry V, etc., we would be able to see this co-evolution of the human and Silg/Sudz ancestral genomes leading to current rhetorical abilities.
Ms. Repor-Tastory : Hmmso our silver tongue-based rhetorical genealogies have a silver thread! Ok, so finally, let me come to my traditional closing question: are there any applications of this part of the GLOPS investigation?
Dr. Noitall-Most : Yes, of course, Abi! Unexpectedly, there is not inconsiderable interest in Gruffy-like macrofibre producers for modulating vocal qualities. There is for example a significant, but previously unrecognised demand from people in the military, law enforcement agencies, team sport training, and the likeboth squeaky-voiced males and mellow-voiced femaleswho have all the necessary qualities for rapid advancement in these careers, but do not feel that they have the requisite harsh voice tone and throw needed on the parade ground, rioting streets, football field, etc.
Ms. Repor-Tastory, with a wicked look : How strange! I have always had a rather melodic voice and when I was a cheerleader of the SCU 5 Football team, I never had any problems getting the attention of the team.
Dr. Noitall-Most : Yeeeess, well. In addition to this niche market, there are a range of more general applications. First and foremost, nicotine-degrading bacteria and the enzymes of their catabolic pathways are being used in a variety of ways. These include the use of pathway enzymes for biocatalysisthe use of enzymes to carry out chemical reactionsgreen chemistry that is environmentally and energetically friendly, 26 and in the removal of nicotine in tobacco processing wastes, 27 which can cause environment pollution.
But perhaps most interesting to viewers are the health applications of this work.
As I mentioned earlier, smoking results in significant changes in the buccal flora. 12 Well, it turns out that it also has an impact on the composition and metabolic activities of the gut microbiota. 28 Whether or not these changes constitute dysbioses of airway and/or gut microbiota and, if so, whether or not they contribute to smoking-related diseases, is not presently known but is the subject of increasingly active research. For example, smokers have a greater susceptibility to respiratory infections. 29 While this may well be due to smoke-caused weakened resilience of the airway epithelium and associated immune defences, 29 it may instead or additionally be due to the functional consequences of airway microbiota dysbiosis.
And, given the physical and functional connectivity of the airway and alimentary microbiota, and the association of a number of health issues with gut microbiota dysbioses, it might well be that some of the other health prejudicial consequences of smoking result from air/gut microbiota dysbioses it causes. Thus, one type of investigation currently underway by new microbiome enterprises, and especially global food companies, which are increasingly interested in the new prophylaxis and therapy opportunities offered by functional foods, is to determine whether or not transplants of healthy oral microbiota might reduce smoking-associated diseases and, if so, which ones.
Ms. Repor-Tastory : Don't you find it amazing, and somewhat confusing, how blurred the business focus of many large companies has become in recent years? A food company used to satisfy our nutritional requirements, a phone company used to enable us communicate remotely with one another, an informatics company used to help us get on the digital highway, but now their activities are all mixed upthey would probably say strategically integratedand one way or another are all in the health or entertainment businesses.
Dr. Noitall-Most : Absolutely true Abi: the world is indeed changing.
Another topic receiving intense attention is nicotine addiction 30 which, as viewers may know, is the leading cause of preventable disease and premature death. 11 Its biological basis, involving inter alia the so-called dopamine reward pathway, is now well defined but so far attempts to find ways and means of aiding smokers to kick the habit have not yielded the desired outcome. 30 As a result of GLOPS, the Lorenzo von Syntech High Security Institute for Artificial Life in Madrid, headed by the world-renowned Professor Vic Torde, recently embarked on an ambitious synthetic microbiology programme to design an excellent throat colonising bacterium that will smooth the withdrawal process for those who finally decide that it is high time to quit. This bug, tentatively named by Vic Nicowithdrawia facilensis, will contain the following: (i) a highly efficient nicotine degradation pathway to reduce the amount of nicotine entering the lungs during puffing activity, which is intended to have the dual benefit of reducing nicotine-related tissue damage and of gently weaning smokers off their addiction, 31 (ii) a new metabolic route for high level production of vitamin C, to compensate for the increased need for vitamin C in smokers, 32 and (iii) novel biosynthetic pathways for the production of 2 neuroactive compounds that counteract the nicotine withdrawal symptoms of irritability, depressed mood, hedonic dysregulation, restlessness, anxiety, concentration difficulty, increased hunger, insomnia and tobacco craving 3233 . Experiments with some of the initial constructs given to addicted mice look promising, so smokers may soon have a golden bullet at their disposal.
Ms. Repor-Tastory : Well, I never: smoking mice! Dr. Noitall-Most : Vic is also very interested in the nanosilver production activities of Silg and Sudz because nanosilver has an amazing spectrum of applications in industry, food manufacture, medicine and consumer products 2334 . An important property of silver nanoparticles is their powerful antimicrobial activity, which is exploited in disinfectants, decontamination, air purification, and the coating of medical devices, like catheters, to prevent their colonisation by microbes, especially pathogens, and for impregnation of clinical clothing. Nanosilver is also used to improve diagnostic imaging and is showing promise as an enhancing agent in cancer therapies. Though he will not be very specific at this stage, Professor Torde is designing derivatives of Silg that not only produce silver nanoparticles at much greater efficiencies than natural microbes, but also derivatives that produce functionalised nanoparticles, that is, silver particles coated with molecules, including nanobodiesdesigned single chain camel antibiodies, 35 that endow them with high affinities for a diverse range of targets for the purpose of detection, measurement and, in some cases further metabolism or harvesting. I am sure we will learn of some very interesting advances from Vic in the near future.
And while on the topic of Silg and Sudz, an amazing number of people now routinely have their microbiomes sampled and composition determined, not only to upstage their friends at cocktail parties and on social media, but also to learn about their health-related genetic profiles. Interestingly, a not insignificant number of people have discovered that they have Silg or Sudz residing in their buccal mucosa. And not an insignificant number of such individuals pay genome companies small fortunes to carry out genealogy investigations to assess whether they might be descendants of one or other of the great Greek or Roman orators. Most of these do it out of curiosity, but others, especially budding spin doctors, politicians, sales personnel and lobbyists, do it in order to enhance their CVs with tenuous documentation of credentials and pedigrees of their persuasive abilities.
Ms. Repor-Tastory : well, viewers: on that rhetorical note, we end this edition of 'Discoveries that Change our Lives'. We will be back next week with a follow-up edition to reveal some more of the amazing findings of GLOPS and its experimental follow-up.
As the camera pulls away, viewers see Abi leaning towards Ani and hear her saying : to get back to my karaoke career aspirations. . .

Conflict of interest
None declared.
Explanatory Notes for Online Version at www. theabsurdmicrobe.com 1 To view previous issues of Microbiome Discoveries that Change our Lives, go to http://www.theabsurdmic robe.com/discoveries-oldstuff. 2 Health Warning. This article is a figment of the authors' imagination; any serious contemplation may unnecessarily challenge the mental harmony of the reader. 3 Any possible resemblance of the fictional characters and organisations in this fable to a real person, organisation or enterprise is coincidental and unintentional. 4 Dedicated to pioneering researchers of the microbiome, synthetic microbiology, and especially those revealing human phenotypes influenced by our microbial partners, to pioneers of microbial nicotine degradation, especially Rod Brandsch, of biocatalysis, especially to the memory of those heavyweights Andreas Kiener and Bernard Witholt, who have sadly passed on, of nanosilver production and application and, as always, to the memory of Dan Koshland, Jr., father of Dr. Noitall (e.g. see Editorial: A simple proposal for health care rationing. Science 264: 1515), who in turn is the father of Dr. Anastasia Noitall-Most, and David Jones, father of Daedalus, who invented Dreadco (see, e.g. https://www.newscientist. com/article/mg23531413-500-what-did-daedalus-talk-ab out-in-septembers-past/; https://www.improbable.com/ 2017/07/27/sad-news-daedalus-david-jones-is-gone/).