1 00:00:00,790 --> 00:00:08,320 [Music] 2 00:00:12,740 --> 00:00:11,360 this is kind of a side project that I 3 00:00:14,749 --> 00:00:12,750 work on 4 00:00:16,790 --> 00:00:14,759 I don't normally study salty systems 5 00:00:20,000 --> 00:00:16,800 this is kind of like a side pet project 6 00:00:24,800 --> 00:00:20,010 that we have going on until my other 7 00:00:26,900 --> 00:00:24,810 samples come in next year so why do we 8 00:00:28,880 --> 00:00:26,910 care about salt flats and astrobiology 9 00:00:32,359 --> 00:00:28,890 because we know there are salt flats on 10 00:00:37,340 --> 00:00:32,369 Mars these are the salt flats that were 11 00:00:39,619 --> 00:00:37,350 studied in 2005 and they're thought to 12 00:00:41,990 --> 00:00:39,629 be a remnant of a large body of water 13 00:00:45,020 --> 00:00:42,000 that used to be in that area a long time 14 00:00:47,840 --> 00:00:45,030 ago and these are the Salt Flats in Utah 15 00:00:52,459 --> 00:00:47,850 that I'm currently working on that were 16 00:00:54,680 --> 00:00:52,469 also where a Lake Bonneville used to be 17 00:00:57,770 --> 00:00:54,690 and then when Lake Bonneville evaporated 18 00:01:00,049 --> 00:00:57,780 we are left with the Bonneville Salt 19 00:01:02,060 --> 00:01:00,059 Flats and so there's a couple very key 20 00:01:04,340 --> 00:01:02,070 difference things between these two 21 00:01:06,770 --> 00:01:04,350 places one is that the Salt Flats in 22 00:01:09,710 --> 00:01:06,780 Utah undergo these ephemeral wet/dry 23 00:01:11,810 --> 00:01:09,720 cycles and so right now and all winter 24 00:01:16,700 --> 00:01:11,820 long they were covered in a layer water 25 00:01:19,429 --> 00:01:16,710 and then as it heats up and warms up the 26 00:01:24,039 --> 00:01:19,439 water evaporates off and dries out and 27 00:01:26,270 --> 00:01:24,049 then you get the salt crust formation so 28 00:01:28,940 --> 00:01:26,280 there haven't been too many studies on 29 00:01:31,730 --> 00:01:28,950 the salt crust formation and what are 30 00:01:34,310 --> 00:01:31,740 the biophysical chemical properties that 31 00:01:36,140 --> 00:01:34,320 lead to it um there have been some 32 00:01:38,630 --> 00:01:36,150 geology studies where they've gone and 33 00:01:40,340 --> 00:01:38,640 drilled cores and looked at like kind of 34 00:01:42,679 --> 00:01:40,350 what are the mineral components on these 35 00:01:44,539 --> 00:01:42,689 salt flats um but the only microbiology 36 00:01:46,880 --> 00:01:44,549 studies that have been done to date our 37 00:01:49,249 --> 00:01:46,890 culture dependent one of our 38 00:01:51,530 --> 00:01:49,259 collaborators has gone out and tried to 39 00:01:53,870 --> 00:01:51,540 grow things from them and isolated your 40 00:01:55,969 --> 00:01:53,880 standard halo archaea some Salona back 41 00:01:57,289 --> 00:01:55,979 there things like that that we know are 42 00:02:02,810 --> 00:01:57,299 insult these systems and aren't very 43 00:02:05,030 --> 00:02:02,820 interesting from my perspective so when 44 00:02:07,280 --> 00:02:05,040 we culture things we only get like 1% of 45 00:02:09,790 --> 00:02:07,290 the microbial community so we're taking 46 00:02:13,070 --> 00:02:09,800 a culture independent approach and 47 00:02:13,760 --> 00:02:13,080 looking at what are the total anaerobic 48 00:02:16,460 --> 00:02:13,770 and aerobic 49 00:02:18,530 --> 00:02:16,470 microbe communities out there and then I 50 00:02:20,300 --> 00:02:18,540 want to dig a little bit deeper and sort 51 00:02:22,820 --> 00:02:20,310 of ask this question what are they doing 52 00:02:24,590 --> 00:02:22,830 and what are the species that are really 53 00:02:27,410 --> 00:02:24,600 responsible for the primary carbon 54 00:02:30,380 --> 00:02:27,420 fixation in the system and do they need 55 00:02:33,730 --> 00:02:30,390 hydrogen as the electron source and so 56 00:02:36,770 --> 00:02:33,740 this second question sort of comes more 57 00:02:38,750 --> 00:02:36,780 into play when we think about the 58 00:02:41,300 --> 00:02:38,760 anaerobic community members that could 59 00:02:43,610 --> 00:02:41,310 be fixing carbon in there and more 60 00:02:46,070 --> 00:02:43,620 specifically the methanogens and the 61 00:02:48,770 --> 00:02:46,080 asita jhin's that um might possibly be 62 00:02:52,010 --> 00:02:48,780 there because they both need hydrogen as 63 00:02:57,010 --> 00:02:52,020 electron donor and so the Bonneville 64 00:02:59,300 --> 00:02:57,020 Salt Flats are also an interesting space 65 00:03:00,650 --> 00:02:59,310 aside from other salt flats because you 66 00:03:04,190 --> 00:03:00,660 have a lot of human activities that go 67 00:03:05,960 --> 00:03:04,200 on out there so these high speed land 68 00:03:09,020 --> 00:03:05,970 races take place every year that the 69 00:03:10,640 --> 00:03:09,030 salt crust thickness um is actually 70 00:03:13,250 --> 00:03:10,650 thick enough for them to occur and so 71 00:03:14,870 --> 00:03:13,260 you have like the highest speeds that 72 00:03:16,310 --> 00:03:14,880 have occurred on land have been out here 73 00:03:20,690 --> 00:03:16,320 and also the highest speeds on a 74 00:03:22,580 --> 00:03:20,700 barstool have taken place there and so 75 00:03:24,620 --> 00:03:22,590 we also have mining processes that are 76 00:03:27,530 --> 00:03:24,630 going on there they pump out the 77 00:03:31,040 --> 00:03:27,540 underground water and pump it into 78 00:03:33,020 --> 00:03:31,050 evaporating ponds and then sell that as 79 00:03:35,680 --> 00:03:33,030 table salt so you might be eating salt 80 00:03:40,400 --> 00:03:35,690 from Utah with your dinner tonight um 81 00:03:41,990 --> 00:03:40,410 okay so we went out and sampled some 82 00:03:44,600 --> 00:03:42,000 things as we do as environmental 83 00:03:46,699 --> 00:03:44,610 microbiologist so we paired up with our 84 00:03:49,430 --> 00:03:46,709 geology collaborators and the geology 85 00:03:52,640 --> 00:03:49,440 took samples all over here from every 86 00:03:55,280 --> 00:03:52,650 single site labeled they actually use 87 00:03:57,830 --> 00:03:55,290 this really cool drill and drilled them 88 00:03:59,630 --> 00:03:57,840 out and they got down there fairly deep 89 00:04:01,040 --> 00:03:59,640 however all of these samples were 90 00:04:03,410 --> 00:04:01,050 useless to us because they collected 91 00:04:06,440 --> 00:04:03,420 them in a way that was not aseptic there 92 00:04:10,820 --> 00:04:06,450 was no way to drill them cleanly um so 93 00:04:13,940 --> 00:04:10,830 we went out and instead dug pits um so 94 00:04:16,550 --> 00:04:13,950 we we sterilized our tools between sites 95 00:04:22,460 --> 00:04:16,560 as best as we could with ethanol and 96 00:04:24,440 --> 00:04:22,470 flame and these okay these are the sites 97 00:04:27,230 --> 00:04:24,450 that we sampled in red right here and so 98 00:04:29,480 --> 00:04:27,240 um this black line right here is the 99 00:04:31,400 --> 00:04:29,490 racetrack and so we tried to get a 100 00:04:33,950 --> 00:04:31,410 sample on and off the racetrack and sort 101 00:04:37,610 --> 00:04:33,960 of in this transect across the entire 102 00:04:39,680 --> 00:04:37,620 area and these are what our pits look 103 00:04:43,370 --> 00:04:39,690 like at each one of those sites um so 104 00:04:44,570 --> 00:04:43,380 you can see in in this site right here 105 00:04:44,839 --> 00:04:44,580 oh these aren't really coming up too 106 00:04:49,969 --> 00:04:44,849 well 107 00:04:51,770 --> 00:04:49,979 textural layers as you got down and so 108 00:04:55,070 --> 00:04:51,780 they looked different and we split them 109 00:04:57,409 --> 00:04:55,080 up according to those like visual and 110 00:04:59,629 --> 00:04:57,419 textural layers some of them had only 111 00:05:04,059 --> 00:04:59,639 two layers and some of them had three 112 00:05:07,370 --> 00:05:04,069 layers and one of them had four layers 113 00:05:09,469 --> 00:05:07,380 and so we got back to the lab with all 114 00:05:11,870 --> 00:05:09,479 of these samples homogenized them 115 00:05:14,689 --> 00:05:11,880 extracted DNA from them sent a subset 116 00:05:18,710 --> 00:05:14,699 off to geology to do mineral analysis 117 00:05:21,589 --> 00:05:18,720 and then we sequence them using 16s 118 00:05:24,920 --> 00:05:21,599 sequencing and we will be doing 119 00:05:26,600 --> 00:05:24,930 metagenomic sequencing on a select few 120 00:05:28,370 --> 00:05:26,610 of these hits we haven't decided which 121 00:05:31,909 --> 00:05:28,380 ones to do that yet that's kind of the 122 00:05:34,459 --> 00:05:31,919 next step and I also set up these 123 00:05:35,450 --> 00:05:34,469 enriched carbon incubations try to get 124 00:05:37,219 --> 00:05:35,460 at this question of what are the 125 00:05:39,320 --> 00:05:37,229 microbes doing there so I wanted to dig 126 00:05:40,999 --> 00:05:39,330 a little bit deeper beyond who is there 127 00:05:45,350 --> 00:05:41,009 and try to answer this what are they 128 00:05:50,659 --> 00:05:45,360 doing and so I use stable isotope 129 00:05:53,240 --> 00:05:50,669 tracing with 13c carbon so these are all 130 00:05:55,999 --> 00:05:53,250 of the media that I set up incubations 131 00:05:57,499 --> 00:05:56,009 with I chose glucose to sort of get an 132 00:06:01,010 --> 00:05:57,509 idea of what the heterotrophic community 133 00:06:03,379 --> 00:06:01,020 is like so what are the organisms that 134 00:06:04,760 --> 00:06:03,389 are consuming carbon produced by other 135 00:06:06,950 --> 00:06:04,770 species there 136 00:06:10,300 --> 00:06:06,960 I chose acetate because I'm interested 137 00:06:12,670 --> 00:06:10,310 in the interplay between asita jhin's 138 00:06:16,520 --> 00:06:12,680 organisms that make acetate methanogens 139 00:06:18,260 --> 00:06:16,530 organisms to make methane and that have 140 00:06:19,580 --> 00:06:18,270 trophic community so can the 141 00:06:23,420 --> 00:06:19,590 heterotrophic community consume the 142 00:06:27,439 --> 00:06:23,430 acetate made by the citizens and I set 143 00:06:29,180 --> 00:06:27,449 up these bicarbonate incubations also to 144 00:06:31,010 --> 00:06:29,190 sort of get at this play between 145 00:06:35,120 --> 00:06:31,020 methanogens and its citizens they both 146 00:06:37,519 --> 00:06:35,130 usually compete for co2 as well as even 147 00:06:39,589 --> 00:06:37,529 sometimes you have sulfate reducers that 148 00:06:42,019 --> 00:06:39,599 can use all of these compounds 149 00:06:43,429 --> 00:06:42,029 and so sulfate readers will generally 150 00:06:45,709 --> 00:06:43,439 out-compete both the citizens and 151 00:06:49,100 --> 00:06:45,719 methanogens for things there's much 152 00:06:52,519 --> 00:06:49,110 better at grabbing these things and they 153 00:06:53,839 --> 00:06:52,529 it takes some much less energy to do 154 00:06:58,010 --> 00:06:53,849 what they're doing then the methanogens 155 00:06:59,419 --> 00:06:58,020 and these heated ins um so so that's why 156 00:07:02,559 --> 00:06:59,429 I did all of these and then we had one 157 00:07:05,419 --> 00:07:02,569 with hydrogen and one without hydrogen 158 00:07:07,339 --> 00:07:05,429 again this is because the citizens and 159 00:07:09,469 --> 00:07:07,349 the methanogens specifically need the 160 00:07:11,179 --> 00:07:09,479 hydrogen as electron donor as well as 161 00:07:14,269 --> 00:07:11,189 bicarbonate to grow um 162 00:07:16,189 --> 00:07:14,279 and so these are the incubations right 163 00:07:19,939 --> 00:07:16,199 here they hung out on my bench for about 164 00:07:23,059 --> 00:07:19,949 sixty days and then I extracted all of 165 00:07:27,230 --> 00:07:23,069 the headspace um inside of these EXA 166 00:07:29,149 --> 00:07:27,240 Taner tubes and split that up into two 167 00:07:31,999 --> 00:07:29,159 samples for the glucose and acetate 168 00:07:33,920 --> 00:07:32,009 samples one to be sent off for thirteen 169 00:07:38,839 --> 00:07:33,930 co2 analysis one to be sent off for 170 00:07:40,429 --> 00:07:38,849 thirteen ch4 analysis um and so the 171 00:07:43,159 --> 00:07:40,439 reason we did this is because if you 172 00:07:46,309 --> 00:07:43,169 have something consuming glucose that is 173 00:07:48,769 --> 00:07:46,319 labeled with this 13 C they'll respire 174 00:07:50,779 --> 00:07:48,779 it out as 13 co2 and so you can catch 175 00:07:52,609 --> 00:07:50,789 that in the headspace and likewise with 176 00:07:54,049 --> 00:07:52,619 methane if you have active methanogens 177 00:07:57,319 --> 00:07:54,059 in your community that are using any of 178 00:07:59,959 --> 00:07:57,329 these urgency car pot compounds um they 179 00:08:04,399 --> 00:07:59,969 will you can see that as a tracer in the 180 00:08:05,749 --> 00:08:04,409 headspace gases so this kind of answers 181 00:08:07,309 --> 00:08:05,759 the question of what is the total 182 00:08:09,889 --> 00:08:07,319 microbial community doing but doesn't 183 00:08:12,290 --> 00:08:09,899 get at the individuals that are doing it 184 00:08:15,949 --> 00:08:12,300 and so I'm also doing these single cell 185 00:08:17,480 --> 00:08:15,959 Raman sorting experiments um using a 186 00:08:20,329 --> 00:08:17,490 microfluidic device that I'm currently 187 00:08:22,429 --> 00:08:20,339 in the process of designing and I'm not 188 00:08:24,259 --> 00:08:22,439 going to talk too much about this um 189 00:08:26,119 --> 00:08:24,269 because I don't actually have any data 190 00:08:28,489 --> 00:08:26,129 or results to show you guys or even a 191 00:08:31,759 --> 00:08:28,499 device to do them with but the idea is 192 00:08:34,279 --> 00:08:31,769 basically you can shine a Raman laser on 193 00:08:36,800 --> 00:08:34,289 a single cell and if that single cell 194 00:08:40,759 --> 00:08:36,810 has incorporated 13c into its biomass 195 00:08:43,040 --> 00:08:40,769 you'll see a shift in the spectra so you 196 00:08:45,829 --> 00:08:43,050 can then take that single cell sort it 197 00:08:48,079 --> 00:08:45,839 off to the side sequence it and answer 198 00:08:50,350 --> 00:08:48,089 the question of what cell was using 199 00:08:54,770 --> 00:08:50,360 these 13 C carbon compounds 200 00:08:56,450 --> 00:08:54,780 okay um so I will talk to you about the 201 00:09:00,740 --> 00:08:56,460 headspace data because I have some of 202 00:09:02,750 --> 00:09:00,750 that back so this is all of the data 203 00:09:05,000 --> 00:09:02,760 that I have back in a really complicated 204 00:09:09,020 --> 00:09:05,010 chart um I will just kind of go through 205 00:09:11,480 --> 00:09:09,030 it step by step so um we'll look at the 206 00:09:16,220 --> 00:09:11,490 methane data first so as I told you 207 00:09:19,120 --> 00:09:16,230 methanogens need hydrogen to grow um so 208 00:09:21,500 --> 00:09:19,130 we would expect methanogens to be only 209 00:09:24,020 --> 00:09:21,510 producing methane in this control right 210 00:09:26,630 --> 00:09:24,030 here with hydrogen however we can see 211 00:09:29,720 --> 00:09:26,640 that there is some methane production in 212 00:09:35,180 --> 00:09:29,730 these other incue bation x' without 213 00:09:37,580 --> 00:09:35,190 hydrogen so this kind of indicates that 214 00:09:38,960 --> 00:09:37,590 methanogens might be using hydrogen but 215 00:09:43,820 --> 00:09:38,970 they could also be using something else 216 00:09:46,670 --> 00:09:43,830 for their electron donor and so we did 217 00:09:49,940 --> 00:09:46,680 aerobic and anaerobic incubations were 218 00:09:51,920 --> 00:09:49,950 not really expecting any sort of methane 219 00:09:53,420 --> 00:09:51,930 production in these aerobic incubations 220 00:09:56,120 --> 00:09:53,430 because methanogens or anaerobic 221 00:09:58,430 --> 00:09:56,130 organisms and oxygen generally kills 222 00:10:01,250 --> 00:09:58,440 them if they're exposed to it for a 223 00:10:02,600 --> 00:10:01,260 really long time so this is kind of what 224 00:10:07,430 --> 00:10:02,610 we expected to see across the board 225 00:10:09,290 --> 00:10:07,440 either in one of these two however in 226 00:10:11,630 --> 00:10:09,300 this one pit fifty-six 227 00:10:14,030 --> 00:10:11,640 we did get some methane production in 228 00:10:18,590 --> 00:10:14,040 the aerobic incubations and not the 229 00:10:21,200 --> 00:10:18,600 anaerobic incubations um so this is very 230 00:10:22,970 --> 00:10:21,210 strange and I don't really have too many 231 00:10:26,000 --> 00:10:22,980 ideas of what's going on here um 232 00:10:29,900 --> 00:10:26,010 especially because you can see in layer 233 00:10:32,900 --> 00:10:29,910 four you have a lot of it and maybe they 234 00:10:35,030 --> 00:10:32,910 were consuming all of the oxygen that 235 00:10:36,530 --> 00:10:35,040 was in the - or maybe other my grew new 236 00:10:38,420 --> 00:10:36,540 members were consuming all oxygen in the 237 00:10:40,490 --> 00:10:38,430 tube really quickly and so you get to 238 00:10:43,100 --> 00:10:40,500 that anaerobic state fairly fast 239 00:10:45,890 --> 00:10:43,110 um I didn't set up anaerobic incubations 240 00:10:48,130 --> 00:10:45,900 for that layer because our koi chamber 241 00:10:51,310 --> 00:10:48,140 sort of crapped out on us that night and 242 00:10:54,140 --> 00:10:51,320 refused to work because it had no gas um 243 00:10:56,810 --> 00:10:54,150 so I wasn't able to set up once for that 244 00:10:59,360 --> 00:10:56,820 layer unfortunately but also I had a 245 00:11:03,620 --> 00:10:59,370 little bit of methane production right 246 00:11:06,140 --> 00:11:03,630 here in this layer too 247 00:11:08,480 --> 00:11:06,150 as you'll note up here we also had 248 00:11:10,850 --> 00:11:08,490 methane production from glucose which is 249 00:11:13,130 --> 00:11:10,860 very strange because methanogens can't 250 00:11:16,520 --> 00:11:13,140 use glucose they can't even transport it 251 00:11:19,250 --> 00:11:16,530 inside of their cell and so because this 252 00:11:21,920 --> 00:11:19,260 occurred in aerobic incubations and not 253 00:11:23,300 --> 00:11:21,930 the anaerobic incubations this kind of 254 00:11:25,610 --> 00:11:23,310 suggests that there's some sort of 255 00:11:27,380 --> 00:11:25,620 partnering going on with the aerobic 256 00:11:34,940 --> 00:11:27,390 microbial community members and the 257 00:11:37,910 --> 00:11:34,950 methanogens and so we can kind of extend 258 00:11:39,250 --> 00:11:37,920 this into pit 12v where I also saw the 259 00:11:41,660 --> 00:11:39,260 same thing 260 00:11:43,700 --> 00:11:41,670 there was methane production from 261 00:11:46,790 --> 00:11:43,710 glucose going on under aerobic 262 00:11:49,400 --> 00:11:46,800 conditions but not anaerobic conditions 263 00:11:52,580 --> 00:11:49,410 down here and so this was kind of a 264 00:11:54,500 --> 00:11:52,590 strange result but it also kind of makes 265 00:11:58,370 --> 00:11:54,510 sense when you look at the co2 data so 266 00:12:02,270 --> 00:11:58,380 in these same tubes right here where I 267 00:12:04,700 --> 00:12:02,280 have methane production from glucose we 268 00:12:06,410 --> 00:12:04,710 know that there's labeled co2 in there 269 00:12:09,430 --> 00:12:06,420 so I kind of have even more evidence 270 00:12:12,140 --> 00:12:09,440 that these aerobic community members are 271 00:12:14,090 --> 00:12:12,150 aerobic lee respiring glucose turning it 272 00:12:16,130 --> 00:12:14,100 into 13 co2 and then the most antigens 273 00:12:18,950 --> 00:12:16,140 are able to use that 13 co2 and making 274 00:12:22,610 --> 00:12:18,960 methane and I think a similar story is 275 00:12:27,290 --> 00:12:22,620 happening with acetate right here in pit 276 00:12:29,720 --> 00:12:27,300 12b and so some methanogens can directly 277 00:12:31,520 --> 00:12:29,730 use acetate to make methane however I 278 00:12:37,820 --> 00:12:31,530 don't think that's going on because if 279 00:12:40,940 --> 00:12:37,830 we look at the whoops okay um so if we 280 00:12:47,420 --> 00:12:40,950 look at the anaerobic ones down here 281 00:12:49,670 --> 00:12:47,430 with acetate and uh yeah so if we look 282 00:12:51,470 --> 00:12:49,680 at the the ones down here that are 283 00:12:55,310 --> 00:12:51,480 anaerobic with acetate there is no 284 00:12:57,020 --> 00:12:55,320 methane production so methanogens are 285 00:12:58,580 --> 00:12:57,030 anaerobic if they were able to use 286 00:13:00,130 --> 00:12:58,590 acetate directly we would expect to see 287 00:13:04,300 --> 00:13:00,140 methane production in the anaerobic 288 00:13:07,280 --> 00:13:04,310 controls also but we only see it in the 289 00:13:11,480 --> 00:13:07,290 aerobic ones so this is further evidence 290 00:13:13,370 --> 00:13:11,490 that there is in fact some sort of 291 00:13:15,040 --> 00:13:13,380 interaction between the aerobic 292 00:13:16,790 --> 00:13:15,050 community members and the methanogens 293 00:13:25,840 --> 00:13:16,800 especially in 294 00:13:31,250 --> 00:13:25,850 layer two right here okay so um whoops 295 00:13:32,509 --> 00:13:31,260 let me go on to what's going on Oh okay 296 00:13:37,730 --> 00:13:32,519 so that was what I just mentioned where 297 00:13:39,290 --> 00:13:37,740 you have um no so you have no methane 298 00:13:44,660 --> 00:13:39,300 production right here from bicarbonate 299 00:13:46,670 --> 00:13:44,670 also uh in layer one all right right 300 00:13:48,920 --> 00:13:46,680 here yeah so in layer one you have no 301 00:13:52,519 --> 00:13:48,930 methane production in the anaerobic 302 00:13:54,410 --> 00:13:52,529 controls from bicarbonate but we kind of 303 00:13:57,500 --> 00:13:54,420 see it as we go deeper in the pits and 304 00:13:59,630 --> 00:13:57,510 so this is a result that sort of makes 305 00:14:01,430 --> 00:13:59,640 sense because you would expect 306 00:14:03,590 --> 00:14:01,440 methanogens to be deeper down in the 307 00:14:07,910 --> 00:14:03,600 soil where they're kind of isolated from 308 00:14:10,460 --> 00:14:07,920 that oxygen that's getting to them okay 309 00:14:13,940 --> 00:14:10,470 and then lastly we'll kind of look up 310 00:14:16,310 --> 00:14:13,950 here at what was going on in the aerobic 311 00:14:18,410 --> 00:14:16,320 bicarbonate incubations and so you can 312 00:14:23,030 --> 00:14:18,420 see there's a little bit of methane 313 00:14:24,079 --> 00:14:23,040 production in that first layer um so for 314 00:14:26,150 --> 00:14:24,089 some reason there were a little bit of 315 00:14:28,400 --> 00:14:26,160 methanogens that maybe we're hanging out 316 00:14:30,170 --> 00:14:28,410 in here but they can't really do 317 00:14:31,430 --> 00:14:30,180 anything because they're not they're not 318 00:14:32,510 --> 00:14:31,440 doing anything right here when it's in 319 00:14:34,069 --> 00:14:32,520 aerobic they can't really do anything 320 00:14:38,840 --> 00:14:34,079 unless they have their aerobic partners 321 00:14:40,610 --> 00:14:38,850 um so this pit right here definitely 322 00:14:43,670 --> 00:14:40,620 there's some interesting things going on 323 00:14:45,079 --> 00:14:43,680 and I don't really have too much else to 324 00:14:47,870 --> 00:14:45,089 say about that other than there's some 325 00:14:50,000 --> 00:14:47,880 evidence that it's happening um but I 326 00:14:51,440 --> 00:14:50,010 don't really know too much more until I 327 00:14:55,940 --> 00:14:51,450 dig a little bit deeper into this story 328 00:14:59,210 --> 00:14:55,950 um okay so who are the microbial 329 00:15:03,620 --> 00:14:59,220 community members so this is our total 330 00:15:06,350 --> 00:15:03,630 16s sequencing of all of the archaea 331 00:15:08,750 --> 00:15:06,360 unity members and from sample to sample 332 00:15:12,350 --> 00:15:08,760 layer to layer it all pretty much looked 333 00:15:15,530 --> 00:15:12,360 like this and so it's mainly dominated 334 00:15:18,350 --> 00:15:15,540 by hey Leo Beck bye halo bacteria AC II 335 00:15:21,290 --> 00:15:18,360 which isn't too surprising they're from 336 00:15:24,410 --> 00:15:21,300 Salt Flats so layer 1 to layer 3 you 337 00:15:26,900 --> 00:15:24,420 just get a bunch of halo back t racy and 338 00:15:29,600 --> 00:15:26,910 these are the heterotrophic halo archaea 339 00:15:32,069 --> 00:15:29,610 that we just heard to talk about 340 00:15:33,600 --> 00:15:32,079 except they're not cold-adapted they're 341 00:15:37,350 --> 00:15:33,610 kind of hanging out in normal 342 00:15:40,980 --> 00:15:37,360 temperatures okay so I looked at the 343 00:15:42,059 --> 00:15:40,990 similarity between these samples um even 344 00:15:43,319 --> 00:15:42,069 though they kind of looked all the same 345 00:15:45,989 --> 00:15:43,329 when I just look at who's there you can 346 00:15:47,819 --> 00:15:45,999 this is an MDS plot and so the Axius 347 00:15:49,379 --> 00:15:47,829 don't really matter too much they're 348 00:15:51,110 --> 00:15:49,389 largely arbitrary it's sort of like a 349 00:15:53,730 --> 00:15:51,120 PCA plot if you're familiar with that 350 00:15:55,590 --> 00:15:53,740 and it's really just the distance 351 00:15:57,780 --> 00:15:55,600 between each of these points that have 352 00:15:59,429 --> 00:15:57,790 any meaning on here and so some 353 00:16:02,129 --> 00:15:59,439 interesting trends sort of come out when 354 00:16:04,590 --> 00:16:02,139 you look at the Archaea unity in this 355 00:16:09,090 --> 00:16:04,600 way and you can see all of these circles 356 00:16:11,009 --> 00:16:09,100 on here are layer 1 all of the triangles 357 00:16:13,470 --> 00:16:11,019 on here are layer 2 and all of the 358 00:16:15,660 --> 00:16:13,480 squares are layer 3 so you kind of have 359 00:16:18,019 --> 00:16:15,670 like this general shift in your 360 00:16:21,299 --> 00:16:18,029 microbial community as you go down and 361 00:16:23,040 --> 00:16:21,309 just to kind of show you I don't have 362 00:16:25,259 --> 00:16:23,050 these labeled I just told you what was 363 00:16:27,150 --> 00:16:25,269 in there these are all just down to you 364 00:16:30,179 --> 00:16:27,160 the genus level different halo bacteria 365 00:16:32,040 --> 00:16:30,189 a seee um but it's not really shifting 366 00:16:33,629 --> 00:16:32,050 in who's there it's just the abundances 367 00:16:34,910 --> 00:16:33,639 that are shifting as we go down in 368 00:16:41,160 --> 00:16:34,920 layers 369 00:16:43,829 --> 00:16:41,170 okay so bacteria so bacteria mostly 370 00:16:45,720 --> 00:16:43,839 Salona bacter and some unclassified 371 00:16:47,249 --> 00:16:45,730 bacteria where the 16s sequences didn't 372 00:16:50,160 --> 00:16:47,259 really line up to anything in the 373 00:16:52,079 --> 00:16:50,170 database but it's pretty much dominated 374 00:16:54,030 --> 00:16:52,089 by cilona vector again not too 375 00:16:56,699 --> 00:16:54,040 surprising we would completely expect 376 00:16:58,199 --> 00:16:56,709 this um you can't see it too well but we 377 00:17:00,360 --> 00:16:58,209 have some sulfate reducers right here 378 00:17:03,480 --> 00:17:00,370 and we do actually find some sea urchins 379 00:17:05,059 --> 00:17:03,490 the acetyl Darma right here um oh I 380 00:17:07,860 --> 00:17:05,069 forgot to point it out on the air keel 381 00:17:10,559 --> 00:17:07,870 graph but we did find a tiny amount of 382 00:17:13,199 --> 00:17:10,569 methanogens um so this is really 383 00:17:15,569 --> 00:17:13,209 interesting because as I was hoping we 384 00:17:18,569 --> 00:17:15,579 do have an environment where a citizen's 385 00:17:22,079 --> 00:17:18,579 methanogens and sulfate reducers are all 386 00:17:24,569 --> 00:17:22,089 coexisting and so I want to dig into a 387 00:17:26,460 --> 00:17:24,579 little bit deeper why they're not out 388 00:17:29,130 --> 00:17:26,470 competing each other and why they're all 389 00:17:33,060 --> 00:17:29,140 able to coexist in these sort of lower 390 00:17:34,350 --> 00:17:33,070 abundances in this community but I don't 391 00:17:36,299 --> 00:17:34,360 really have an answer to that yet that's 392 00:17:40,260 --> 00:17:36,309 future work we'll be looking into that a 393 00:17:41,860 --> 00:17:40,270 little bit more and so we can look at if 394 00:17:43,870 --> 00:17:41,870 we have this same 395 00:17:45,070 --> 00:17:43,880 shift that's going on in the bacterial 396 00:17:47,590 --> 00:17:45,080 community like we did with the archaea 397 00:17:51,970 --> 00:17:47,600 unity and again we have circles our 398 00:17:56,019 --> 00:17:51,980 layer one and the triangles or layer two 399 00:17:58,210 --> 00:17:56,029 and the squares are layer three and so 400 00:17:59,830 --> 00:17:58,220 again we see sort of a little bit more 401 00:18:01,870 --> 00:17:59,840 muddled than the layer two and layer 3 402 00:18:04,120 --> 00:18:01,880 with the bacterial sequences but there 403 00:18:07,690 --> 00:18:04,130 is still like a general shift sort of a 404 00:18:10,080 --> 00:18:07,700 way from layer 1 2 as you go deeper into 405 00:18:15,639 --> 00:18:10,090 the more anaerobic community members and 406 00:18:19,090 --> 00:18:15,649 here's another plot and so this is genus 407 00:18:20,950 --> 00:18:19,100 level again and it's just kind of 408 00:18:23,560 --> 00:18:20,960 showing that it doesn't really change 409 00:18:26,639 --> 00:18:23,570 the same colors are between all of the 410 00:18:29,260 --> 00:18:26,649 sequences it's really just the sizes of 411 00:18:30,880 --> 00:18:29,270 what's there so the abundances of the 412 00:18:33,070 --> 00:18:30,890 bacterial community are changing not 413 00:18:35,500 --> 00:18:33,080 exactly the individual sequences 414 00:18:37,930 --> 00:18:35,510 themself ok 415 00:18:40,360 --> 00:18:37,940 so lastly I'm going to talk a little bit 416 00:18:42,760 --> 00:18:40,370 at how I looked at the human impact on 417 00:18:47,049 --> 00:18:42,770 these microbial communities so I 418 00:18:50,200 --> 00:18:47,059 classified all of the all of the pits as 419 00:18:54,430 --> 00:18:50,210 such these ones are on the racetrack and 420 00:18:58,240 --> 00:18:54,440 these ones were off the racetrack and so 421 00:19:00,250 --> 00:18:58,250 then you can look at this you look at 422 00:19:03,399 --> 00:19:00,260 this all using a multivariate analysis 423 00:19:05,769 --> 00:19:03,409 and the everything so if you cut it off 424 00:19:09,730 --> 00:19:05,779 at zero right here every little dot 425 00:19:14,409 --> 00:19:09,740 represents a sequence and all of these 426 00:19:17,649 --> 00:19:14,419 ones that are negative belong to one of 427 00:19:20,909 --> 00:19:17,659 those pits that were on the racetrack 428 00:19:26,139 --> 00:19:20,919 whereas all of these ones up here are 429 00:19:29,970 --> 00:19:26,149 sequences from those pits that were off 430 00:19:33,240 --> 00:19:29,980 the racetrack and so I did this using 431 00:19:35,860 --> 00:19:33,250 sequences from each layer so I split I 432 00:19:37,450 --> 00:19:35,870 go back one so I split all of the 433 00:19:39,580 --> 00:19:37,460 sequences from these pits up into layer 434 00:19:43,240 --> 00:19:39,590 1 layer 2 and layer 3 and did the same 435 00:19:45,610 --> 00:19:43,250 thing layer 1 and 3 didn't have anything 436 00:19:48,850 --> 00:19:45,620 that was significant so these ones in 437 00:19:52,000 --> 00:19:48,860 red right here are the only sequences 438 00:19:54,120 --> 00:19:52,010 that are significantly different from 439 00:19:57,990 --> 00:19:54,130 any sequences up here 440 00:20:01,770 --> 00:19:58,000 um so then all of these red dots right 441 00:20:06,180 --> 00:20:01,780 here indicate a sequence from some 442 00:20:08,400 --> 00:20:06,190 microbe that is similar to everything 443 00:20:13,830 --> 00:20:08,410 that is off the racetrack that isn't 444 00:20:17,550 --> 00:20:13,840 really um on the racetrack so then you 445 00:20:20,640 --> 00:20:17,560 can say what are those sequences and um 446 00:20:24,210 --> 00:20:20,650 this isn't really that exciting it's 447 00:20:37,680 --> 00:20:24,220 just halo back to here AC again um which 448 00:20:39,300 --> 00:20:37,690 is an archaeological primers aren't 449 00:20:41,910 --> 00:20:39,310 really acting in a domain-specific way 450 00:20:43,320 --> 00:20:41,920 there's known issues with that I have to 451 00:20:45,030 --> 00:20:43,330 look into this a little bit more and 452 00:20:47,520 --> 00:20:45,040 actually go in and pull out these exact 453 00:20:48,960 --> 00:20:47,530 sequences and see if they where they 454 00:20:51,300 --> 00:20:48,970 line up to the primers and see if 455 00:20:53,400 --> 00:20:51,310 they're actually were hitting the primer 456 00:20:56,610 --> 00:20:53,410 of it's some like weird sequencing error 457 00:20:57,480 --> 00:20:56,620 um okay so anyways I don't have too much 458 00:21:02,520 --> 00:20:57,490 to say about that 459 00:21:03,840 --> 00:21:02,530 other than that we found a little bit of 460 00:21:05,520 --> 00:21:03,850 differences that might indicate human 461 00:21:08,250 --> 00:21:05,530 impact and I'm going to look into it a 462 00:21:09,750 --> 00:21:08,260 little bit further as well as do some of 463 00:21:12,030 --> 00:21:09,760 these other things like make some more 464 00:21:14,970 --> 00:21:12,040 comparisons to are there differences in 465 00:21:16,950 --> 00:21:14,980 these pits in mineralogy elemental 466 00:21:19,590 --> 00:21:16,960 concentrations like heavy metals and 467 00:21:21,390 --> 00:21:19,600 sulfur as I mentioned before we're going 468 00:21:24,930 --> 00:21:21,400 to metagenomic Li sequence some of these 469 00:21:26,580 --> 00:21:24,940 samples to sort of get an idea of what 470 00:21:29,640 --> 00:21:26,590 genes are present and link that back to 471 00:21:31,590 --> 00:21:29,650 the incubation data and then I have some 472 00:21:33,330 --> 00:21:31,600 additional headspace results that are 473 00:21:36,810 --> 00:21:33,340 going to come in from all of those 474 00:21:38,460 --> 00:21:36,820 incubations and I'm going to do the 475 00:21:42,480 --> 00:21:38,470 Raman single cell sorting of these 476 00:21:45,480 --> 00:21:42,490 samples and then right now or earlier 477 00:21:47,160 --> 00:21:45,490 this week I guess um my lab was going 478 00:21:49,650 --> 00:21:47,170 out and sampling wells that are located 479 00:21:51,450 --> 00:21:49,660 on the Salt Flats so we have shallow 480 00:21:53,040 --> 00:21:51,460 wells and deep wells and we'll be 481 00:21:54,780 --> 00:21:53,050 sequencing those and sort of comparing 482 00:21:57,450 --> 00:21:54,790 them to the surface communities to see 483 00:21:59,670 --> 00:21:57,460 if we can see if some of the shallow 484 00:22:03,420 --> 00:21:59,680 wells in the surface communities share 485 00:22:05,130 --> 00:22:03,430 similarities um and with that I would 486 00:22:07,800 --> 00:22:05,140 like to acknowledge my lab the browser 487 00:22:10,440 --> 00:22:07,810 sin lab um as well as our clabber 488 00:22:14,220 --> 00:22:10,450 readers in the geology department Brenda 489 00:22:16,800 --> 00:22:14,230 Bowen and Betsy Club WA who was the lead 490 00:22:20,010 --> 00:22:16,810 microbiologist who was at the University 491 00:22:30,110 --> 00:22:20,020 of Westminster but has since changed to 492 00:22:36,960 --> 00:22:34,290 what kind Oh what kind of salt is on the 493 00:22:39,000 --> 00:22:36,970 on this salt flat is it just mostly 494 00:22:40,800 --> 00:22:39,010 sodium chloride or I think it's got more 495 00:22:43,830 --> 00:22:40,810 magnesium than sodium in it 496 00:22:50,850 --> 00:22:43,840 I think magnesium is more than potassium 497 00:22:53,340 --> 00:22:50,860 magnesium chloride or okay um real quick 498 00:22:55,470 --> 00:22:53,350 I looked at um it looked like the soil 499 00:22:57,030 --> 00:22:55,480 below the salt was really dark is with 500 00:22:59,130 --> 00:22:57,040 is that true when you're there is it 501 00:23:01,080 --> 00:22:59,140 really dark yeah there's a lot of 502 00:23:03,180 --> 00:23:01,090 organics I guess that are below the salt 503 00:23:04,920 --> 00:23:03,190 yes so I'm really excited to get some of 504 00:23:07,500 --> 00:23:04,930 the mineral data back from the 505 00:23:08,910 --> 00:23:07,510 geologists um they have a undergrad 506 00:23:11,220 --> 00:23:08,920 who's gonna process all those all summer 507 00:23:14,430 --> 00:23:11,230 long for me um but yeah some of these 508 00:23:16,440 --> 00:23:14,440 pits you would dig down and you you 509 00:23:18,900 --> 00:23:16,450 would hit this like weird interface 510 00:23:21,750 --> 00:23:18,910 between two salt layers that was just 511 00:23:23,370 --> 00:23:21,760 really black and sulfur II smelling like 512 00:23:25,430 --> 00:23:23,380 all you just as soon as you hit it 513 00:23:31,050 --> 00:23:25,440 you're just like oh god that's sulfur 514 00:23:35,340 --> 00:23:31,060 cool all right thank you all right we 515 00:23:37,610 --> 00:23:35,350 have time for one last question so were 516 00:23:40,230 --> 00:23:37,620 you able to remove the community members 517 00:23:44,520 --> 00:23:40,240 that were predominantly present on the 518 00:23:47,130 --> 00:23:44,530 racetrack from the rest of the data um I 519 00:23:50,760 --> 00:23:47,140 don't know if I understand let me go 520 00:23:54,360 --> 00:23:50,770 back here okay so from all of these guys 521 00:23:57,420 --> 00:23:54,370 right cuz you would assume that those 522 00:23:59,850 --> 00:23:57,430 are due to human impacts well the 523 00:24:04,230 --> 00:23:59,860 problem with this is that none of these 524 00:24:06,390 --> 00:24:04,240 were really significantly different from 525 00:24:09,330 --> 00:24:06,400 ones that were off the racetrack so it 526 00:24:12,060 --> 00:24:09,340 seemed like we had sequences that were 527 00:24:13,500 --> 00:24:12,070 different off the racetrack that weren't 528 00:24:18,270 --> 00:24:13,510 found on the racetrack so is the other 529 00:24:21,160 --> 00:24:18,280 way around got it okay thanks 530 00:24:28,140 --> 00:24:21,170 all right thank you very much