 The end of the day, with Marco Mare. Thank you. Welcome back after lunch. I'm Marco Mare, post drug in Rosalind Alen's lab in Ena, before we were in Edimura, so we move recently to Ena. And today I'm going to present something regarding algae and bacteria interaction. So dynamics of algae and bacteria interaction in artificial plant in communities. First of all, something regarding Ena, Kaj je ta, da vlastnji srpa češč, kaj je je Viena. So Viena je srpa v medelj v Nere, v zelo. Zelo je srpa, da pričeš, da čekaj ne. Vesel je, da je srpa, da je vnit, da je več integrate. V Viena je informacija za mikrover. Zelo je vzelo in v tudi vzelo. Viena je poslednja včeli in izgledo vzelo. Vzelo je prinsipa, in interakciju delaj. v mikrobejo komunitivnih. Prezitno udelijo v mikrobejo komunitivnih z drožnega, v uvršče, v sojelji, v platnih. Kaj je inga inga inga inga inga inga inga inga inga inga inga. Kaj je inga inga inga inga inga. Všeč odlično vključno je vsev tez, kaj bi se na vsev tez po včelji. Prezitno uključno je uvrščen, zelo, da smo prišli v anti-microbiali resistenje, zelo, da smo modeljali nekaj nekaj mikroskajov, zelo, da smo prišli v mikrobiali komunitivnih, zelo, da smo prišli v mibah, zelo, da smo prišli v e-coli, zelo, da smo prišli v nekaj mikroskajov, zelo biofilms. V zelo, da smo prišli v nekaj mikroskajov, ki mali polenjkni drug, še vse dostajevaj, brutalni drug zelo pod van purposes www.dh.t draw. Prad ose… Zaronek Breve bou z remarkablenima po nekaj metrag say Artagy. Tukaj imam tukaj o tukaj mali algi, tudi unicellular algi, ki smo tukaj pravili, in tukaj je diatoms. Tukaj ima vse exoskelektron, kaj ima vse algi, ki je unicellular in tukaj ima fotozyntesis in tukaj biomas z tukaj. Tukaj, kaj je vse vse vse? Tukaj je tukaj pravili, tukaj ima vse vse mikroorganizm, kaj je tukaj phytoplankton. Tukaj phytoplankton je unicellular algi in tukaj ima vse bakterije. Tukaj ima vse vse, kaj je vse vse, kaj je vse vse tukaj vse 30% glodnjih CO2 in tukaj ima vse 20% in tukaj ima vse tukaj vse. Tukaj vse vse tukaj ima vse marin fudweb. Tukaj ima vse marin envaromen. Algi ima vse vse vse karakteristiki, ker je vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse vse kaj bal, vse vse in po meters, ta pesadla in invade behaviors and then decayin세� hatred. When they age, they change the color to brown. This is because the chlorine field changes the functionality in service s. And we're farmed at the most in m that depends on what the next things we see. Upon measuring this cannotという kontakt. leza prokso in biomusove agii. Klorofaj kontent tako, se vilete v imu vsem vougni, ki vam sem z tukaj našel. In tega motivación vsevene. Zelo, da sem dajbeno, všetak zvali sample v zelo prevod, zame v Jeremii vsi in fron mene in kako je pašla. Zelo je vsega tukajbrenost. in sem jaziv kratil je izgovoril that there are obviously algae in bacteria interacting living together, in we are interested in the composition of this environment that is actually complex, because there are many players together, and then the interaction that defines this composition, which are interaction. Interaction can be interaction within the same species, vseh vseh vseh nekakrati, zato je zelo prišljena na vseh nekakrati, in vseh nekako je vseh nekakrati, naša gačna vseh. Tudi je ima za vseh nekakrati, začnega, ko da je pozitivo, negativno, tudi, kako vseh nekakrati je zelo prišljena na vseh nekakrati, tako pa vseh nekakrati, da je vseh nekakrati, in micronutrajov tema, da je Algi, v sveče v metabolice. V nekaj nekaj veči nekaj reprešen, in kako, če je Vakstira, ne zvedaj, da je Algi, vse nekaj nekaj nekaj nekaj nekaj regojajo v halje. V tudi, da se priče, kaj je pa vse injeračne? Proč, jo in tudi ni. Ne, kaj neko vse, in nekaj nekaj nekaj nekaj neutralne, the carbon sources are always flowing from the phytoplankton to the bacteria. No, that's not completely true. Ok, most of the time they have a common source of carbon, but it really depends on the species that you look at. Well, bacteria prefer obviously glucose species or all these stuff that contain a lot of carbon. And while algae are able also to rely on light, obviously. V tem stvari, da osrednje. Še nekaj so odliči in nekaj ne musimo zapovnači. Ej, kaj je tako pobodil, da je za več nešta objevaja. Tako i kompleksne, vštiri, z našem našte malo zval decipherovanih materijami. Per zelo stavaj mora pri procesu kompetitiv unloadov. Spravujem izgledo izima. Media was, let's get simple. Samp'icales, a smelvrim in mu enchじゃ, extract what we need, then we produce it in the lab. It's not so easy because these kind of stuff grow in a very complex media, and they are not able to grow in well established media that we have in the lab, so usually you do an extraction of marine water, you purify it, use it in dependetive what's inside to make them grow, In s pr basically, it is to the simple system to understand what are the kind of influences on each other. We have a couple of questions to understand that we would like to answer, and obviously is very regarding theمل interaction, secondly, the dynamics, but is something also not so easy in this case because communities are quite complex. And the first question that was moving this kind of research in je to vse drugi bilj, ki se všim vse je vse in zači se nekaj dolgovi roh. Z namelynev, nekaj dolgovi roh, tako, da bomo načinjeni unačnja diatom, bi začal, da bo vse nekaj negativ nekaj dolgov na tebe, in vse dočal vse načinje vse. Zobah, da si tudi se boši vse načinje, je to veči nekaj dolgov, da se boši vse načinje, in da se pričovalo, da je rečno in nekako, je nalegil način na prejmeni. Zato sem da tegači, da je ne univeresan. To je vse vrtega, ker smo predstavili letov. Protoželjamo, da smo vrtegači, da je vrtegači, in ki smo počeljali z tegačnih metod in tegačnih modelov, tako, da sem tukaj površen, nekaj všeč. Kaj je reserčen metod, ko smo vse zelo, da sem ljudi, počekaj mi, čekaj. Površen metod je površen. Prvno, je tukaj skrin, vsepljenje v marinu voda za relevantno-baktyračne interakcije, kaj je tukaj skrin, ker nekaj je tukaj površen, da je tudi početne nedev. Tudi je zašličen, da je dovolj tačne, da je način način način, in zelo, da ne vršajo zelo. In tukaj se predstavila, da ne vštarno od vršča. Tukaj je vzvek, da je včin način, da ne vršča, da je dneziv, in tukaj se vzvek na Ževa. Ako Ževa je zelo, bo vzvek je velik. Zelo si videli, nisem v mikroskopu. z 10-100 mikromov. Zelo, naredite nekaj, da se predstavili pravne procese, zato je zelo načinjeljno nekaj komplek. Zato smo zelo načinjeljno, da smo načinjeljno, in da smo načinjeljno, in da smo načinjeljno, zelo načinjeljno, kultučno. Zelo načinjeljno, načinjeljno, načinjeljno, vsešli, ki je vse medim. In potem smo se zelo začelično vsega kultura. Tudi so možemo da se počelično vsega, kako je vsega kvalitera ideja. Nekaj, ideja je zelo začelično vsega matematika začelično vsega komunitivnega. A z tvojom je bilo, da se vsega načinati, ki se vsega vsega vsega načinati, ki se vsega vsega načinati, kako se vsega vsega načinati, kako se vsega vsega vsega vsega, kaj je vsega o časno cega z svetom komunitvu. So we are creating artificial community inferring some properties by the single growth code and per groß growth codes. How we did this stuff, so it is actually a huge work. So something like six years of work on these things, because growing in left these bugs are complex. Vizon je napravil na postdoc Joondang, v georgaponer labu. To, da je bilo, je to, da je bilo, da je bilo, da je bilo, da je bilo, da je bilo, da je bilo, da je bilo, da je bilo, da je bilo. But, you are taught in the one, the other one are fixed time, independently, whether they are dying or not. But, yeah, you are, you are right, so it's difficult to, to infer same A general condition. They just took a single time point to be, to have a four screen understanding which bacteriaうわ, which algae they should decide to take in the experiment later. So, was a four screen. in reliferujemo, da je lahko velika. Jaz umoževamo, da je namoze to nebezražena, kdaj jaz ne zazmamo večne coulge, zelo zelo poten, ko je to začugal s joinede in ja. Tistim, da vidi lahko ne bo začugalerska vzela in ne bo začugal jel in ljub in kol. Zelo to zelo tudi, da no se bil v vzelo, spri, iz kaj je, je in prišlam še stide je ovrednico pri vsem in v carsi vse prišlj mnogo. Z WHATäljč Future, da je vse vse prišlj tačnja, zelo je to parfaitne. Bez da vienne ljubov, da je zelo če, da je imel zelo, zelo če, da je perega, nekaj, perega nekaj, nekaj je čaša vse, nekaj je ni zaštrufna. Se nan, depenja danemiko in efektu na vsej vsej izgledanje. So, after vision, they choose the most representative. Actually, they are not the most representative, but they are the one that are easy to collect and make it grow in the lab, that are the algae cosimodiscos radiatus, that I will call diatom, because I am not able to... Yeah, there is a question. Is... Sorry, that it becomes more positive, generally, the interaction. Do you think that's just because there's more algae dying and there's more bursts, or am I completely missing something? We don't know what is actually. We don't know whether it is dying or decreasing for other reasons, especially algae have a kind of exoskeletron, so you don't know whether... Even if a dye stays bare algae, because it's kind of empty, you just have to measure the chlorophyll to see whether it's dead or not. We don't know why it happens. We don't know only that in the culture, if you sample and you measure it, you see less chlorophyll. We don't know what's going on. But if you do a life that's stained of the algae, or a count, I don't know. We counted in the microscope. That's how we did it. Basically, you see where the algae is dead or alive from the color, and you can already infer how many cells are there, and yeah, that's what is done. The question was whether they are dying, so removing, or maybe they are sedimenting, or maybe they are keeping a number of cells. We don't know because this stuff is happening in a flask, and after we sample out at two time points, we are not monitoring everything over time completely, but only these two time points, and so it's difficult to infer what is going on in between. And I think also reason is the same question that we had before, so I can't tell you connected to. Yeah, another word? Is the media buffered? Okay, the media is very complex. Actually, I don't know the answer precisely, but I don't think it's buffered. It's just an extraction of marine water, whatever it is inside, because whenever you modify it, they tend not to grow. And regarding what's going on with bacteria and algae is what we are doing now. So we are actually taking also extra amount of media and looking at the metabolites, the properties to understand what's going on. So I cannot give you an answer for better moment. Another verb? Sorry, I don't think I understood. So you're using filter seawater or synthetic medium for these experiments? I think it's filter, yes. Okay. Synthetic, you would know what's going on inside, and you know all the components. Here we are not sure about everything. Yeah? So here are the names that we show, sorry, what's going on. So the diatom is Cosinodiscus radiatus, just to give you a name. And the four bacteria are called for short CS4, Rosae1, Rosae1, and CS1, okay? So I won't give you the name for long because I don't remember them. So and it's also quite exciting to do in Jena because the first guy, I mean, one of the first guys that systematically inspect and describe the shape of the diatom was N-stakel, and he lived in Jena, studied there, so it's quite connected also to the environment inside Jena. And so how Jung did this kind of experiment, basically already told you more or less what's going on, so first of all he grows separately, the algae and the bacteria in the same medium. Here you see on the X axis the days of inoculation inside the medium, so basically in the medium we sample out, we count under the microscope the number of algae alive and also dead, looking at the color of the algae by eye. And here you can see the number of cells, so the concentration, and this curve in orange that is the same curve in all these four plots is the case when algae are growing alone, okay? So these four plots with the same curve, just to give a guide for the eye. And then you do the same by adding bacteria and counting again the number of cells over time of the diatoms, and for example when you add CS4 you see that this is the count of algae in green, this one in yellow is again the count of algae, this one in red and this one in violet, I think, with the presence of a different for bacteria. You can count also bacteria obviously, so this is done by QPCR, so it's quite complex, was difficult to make a standard, so that's why you see up and down in the curves, and here you measure the amount of bacteria, actually the concentration, and over time in gray again the bacteria alone, so CS4 cross A1, Rose 1 and CS1 alone in gray, and with the presence of algae you can measure again bacteria, and you see that you obtain this green, yellow, red and violet counting. What you can notice already from this kind of plot is quite clear, so here when you add bacteria to the algae, algae are growing better, so they are rescue in the death phase. Here instead when you add algae this guy is basically killing, Rose 1 is killing partially the growth of the algae. Vision again is increasing, vision is doing something very crazy, but we are not really able to understand of the moment, we have some ins, is shifting the starting point of the growth, and also for bacteria vision happens, it's a little bit more difficult to understand what's going on, but vision is very clear, so basically this bacterium is not able to grow without algae, but in the presence of algae is able to grow, so probably algae are supplying something to the growth of bacteria. So they have different growth dynamics obviously, and depends also on the algae aging, because you see that, for example, this rescue doesn't happen here, but only on this late phase, and vision is something that we already knew from the very first inspection. Now, going to the model, very simple, so we discussed a lot about the last week, I don't go in detail why it's good or not, was the only option that we had obviously, because we don't know anything regarding the physiology of the stuff, so we know something regarding the algae, we don't know much about this kind of bacteria, about the specific physiology about the metabolite that are released, and so we use a lot of other model, just to give you an idea, again, there are the four different bacteria that interact with the diatom, X is the density of the cell, and then you have a growth rate, the interaction term that are divided between self-interaction and perverse interaction, and you can, I mean, you know already that if term is positive, means that there is a positive interaction, so one in S is the other one, negative there is a repression, and zero means that there is no interaction, so from the feat of axonic culture, so culture alone, growing alone, you obtain the growth rate and the self-interaction term, nothing special. From feat of perverse culture, you obtain the interaction terms, and here there is also kind of a trick, because we have to include the carrying capacity for the algae, apparently they have, they are using a lot of nutrients so we have to fix the amount of carrying capacity that we reached, and once you do that, you obtain this kind of interaction, so apparently CS4 and diatom are repressing each other, Rose1 is helping diatom to grow, diatom helps close A1, and CS1 interact negatively with the diatom, so these are the main interactions, and as I told you, there are two set of parameters, because we have conditions for the early growth phase and the late stationary phase that are different, especially in the late stationary phase is quite boring, nothing happens, so it is repressing the diatom, and now the question is, what happens if you put actually all together this stuff and the diatom, is the model representing well the data, so this was just the fitting of the model and the prediction of the model, but we have not tested yet, so you can test it, and here I represented again once more the metrics of interaction here, times after inoculation of the algae alone or bacteria and algae, in this case it is just algae alone, so just to guide the eye for you, and then you have here the number of algae, so this again is the fit of the diatom in absence of algae, sorry, in absence of bacteria, and when you put instead all the bacteria together with algae, the model gives this prediction in black and the measurements gave this dotted line, so actually we can see that they are quite in good agreement, so Lotka Volterra here works pretty nicely, and to be sure we also did ok, here is another stuff, first of all we have also to say that there are no interactions between bacteria-bacterias so no higher order interaction and since the model already fit very nicely the data we think that probably perverse interaction are negligible although the kind of error that you have are not really able to distinguish between the two cases probably even if the bacteria are very weak interaction, you cannot see them then we check that neutral case, so no interaction gave a completely different as well in the green one here and finally you can actually divide the system in excluding all the interactions but two, and are these CS1 and this cross A1 so these error and these error are already able to give this magenta line that is very nice very nicely next to the black line we have all the interactions, so only two are leading the dynamics, and actually are the faster glowers so you can do the same for bacteria we can give a prediction for the growth of bacteria because you can also measure them I don't know what's happening these are the prediction of the modeling solid line and the measurement with qPCR are the dotted line so we are more or less in the same hierarchy since we are out of time I will give you also the summary so here we studied the agent-bacteria interaction they don't present any universal effect the community dynamic depends on the growth phase of algae and the outlook I can also give you an intro about the outlook we would like to investigate the communities that are more similar to the natural one but to do that we have to understand better the physiology of algae the physiology of bacteria and how they interact so what they exchange and that's why the group of pornet is actually because they are able to measure all the metabolite inside the system and just to give you an idea that's the last slide the model works more or less in this way I mean we are already on the mathematical part but I won't present the result about this one so basically algae are using some nutrients nitrogen silicate especially and light and they release some substrate that is used by bacteria so bacteria is able to grow in this substrate but over time algae are dying so they age and they produce less and less nutrient they start dying so bacteria at this point are not receiving any substrate they are unhappy, they start dying too but bacteria are not stupid they release some metabolites that stimulate algae to produce some vesicle that shuttle out some actually are redox product of the redox stuff in the algae so basically you get rid of what make you old and they get back to be younger at this point we are able to produce more substrate and the bacteria are happy again and they stop in producing the metabolites so this one is kind of a loop obviously depend on the algae of bacteria that you have we are investigating the specific substrate and metabolite that are released so as I told you is a work in progress with this and I would like to acknowledge the people who have worked with me so in Jena, in Max Planck for chemical ecology in Jena our funding agency and I would like to advertise something so we have twitter, theoretical biophysics we have also a podcast so if people would like to listen to the podcast and participate and help us enlarging this community would be very nice and we have also a population dynamics virtual seminar every three weeks online open to everyone I know that already some people already had subscribed and other were speaker by us so this stuff is actually published in this paper and we would like also to save it next week, not in two weeks 90 anyway we have this small one day seminar in one day symposium in Jena usually in TRC in modern ecology and evolution of microbiomes you can come to Jena or it's for free or you can follow it online and for example Yakopa will be one of the invited speaker so we would like to thank you for the attention there are any more questions yeah it's very short I know nothing about algae but I was wondering in your natural samples do you see your microorganisms in like aglomerates in clusters or do they swim freely? I don't know better I've not done the experiment I've not seen it, I've seen just what happened after I know that actually in the the theory is that in marine water since nutrients are so diffused and so lighting concentration aglomeration of algae and bacteria because in the other case nutrients would be too much dispersed and not usable so for sure there are aglomeration and the basic idea of this phytoplankton is that one I don't know if you sample it and you put it in a microscope immediately you see it, they don't create clusters instead of biofilms or snowflakes or this stuff so the follow-up question is that you did not observe this forming in your artificial... actually everything is very well-shaked there and the reason is to keep it simple at the moment thank you just a couple of questions one is regarding the modeling so you do put bacteria-bacteria interactions and they turn out to be negligible ok, no, I've not put it I tried to put it it was not fit in any sense because it was over-fitting so I cannot see any given better that we have I cannot distinguish them it's already done and fine in that way we said no, maybe it's not useful to put it in ok, the other thing is since in these experiments I know you haven't done the experiment but since in these experiments you're using filter seawaters there's a possibility that this filter seawater contains lots of phages that might affect the bacteria they purified before in that case they get rid of the phages they get rid of it one more question I don't know I have no idea I have no idea I can ask and tell you but I don't know I know that there is a part of micro filtration several times several layers of micro filtration but I don't know how they get rid of all these living and interacting stuff I can ask thanks, the speaker again