Lab II-1.pdf

Buildingand Observing

Microcosms

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MATERIALS YOU PROVIDE

• Gloves

* Bag, brown paper

• Camera (opbonal)

• Eggshell and yolk (see text)

• Funnel (or aluminum toil to make your own)

• Jars, widemouth (see text)

• Mixing howl or similar container

• Newspaper

* Pondwater‘edn1ent/v’egetabon (see text)

Shredder and/or scissors

• Sell drink bottles, 500 mL or 1 L

• Trowel, ladle, or other large scoop

• Water. spring or boiled tap

C

EQUIPMENT AND MATERIALS

You It need the

following Items

to Complete

this

_{lab sesslon}

_(The

_{standard kIt for}

_this

_book

available from wwrv,[hehomescientist.coiTl,

_{includes the items listed}

_in

_the

first

group.)

MATERIALS FROM KIT

Goggles

BACKGROUND

‘dust of the other oh sessions in this hook ireorriariized ‘rito related groups. may be per1nr ned in any order you ,‘iish, and many (not all) can be completed over the

of onelab perrodtoat most ‘rVi’ridlab eririds over a seek or two. Unfortunately—unlike chemistry. physics, rednlnst oth’ rscences—’—hio lnesi ohs a an allh0_{h oi}or down a to t-’cctanedsinge oh05’OOPSu iiin[ ‘i

awn scrieduies.hue cyiles ire wiiot Pay ‘a.. rodira a powerlessto speed themrip orslow hear down.

derIding and observing mri’rocosros a. .in ‘-‘cafent oxanipie

of one of theseon-its ownschedule .ircP’/tlrrs. rhire isnoji

o be learned by doing dioarti’iity,but nuroulrrg t properly inquires frequent and detailed ohserv.itioris 0/er a per ndat ,seeks to months. Accordingly, ratherthan attempt to diotrihute these activities outovernany lh sessIon rnups. we decided

toconsolidate them in this group ofSibsessions, which we’ll begun at the start of the semcstcr and contunue throughout the

soniester.

Someof the irrdividral sessionsinthisgroupwillhe repeated periodically throughout thesemester. hegirinii’rg earlyand

continuing.For example, we’llobserve microscopically the populations of our pond water microcosmsthroughoutthe samester, rioting changes that occur in tire types and mix of dutferent organisms as the microcosm life cycle progresses

rumjuvenile to mature to senescent. Our mlcroco’;rns will also

.orveasresources for some later topics. For example, we’ll

ubservesuccession in the microcosms, which properly belongs ii rcnOgy. hut requires extensive and repeated observations warthecourse of many weeks. Similarly, rNe can use our urricrocosms asa source of live protistswhen we tothe lab cession on protists.

InIbislab session we’ll buildtwotypes of microcosms based on water, sediment, and plant life gathered from a peed or stream. We’ll then observe these rnicrocosms periodically over the course of several weeks, noting changes that occur throLigh the life cycles of the various organisms present.

Fbifirst_{type t microc.osnr resembles a}standard .nluarum. We llusewide-mouth lars tilled with a relatively thin layer of sediment, with pond water arid iquatic plamm[’u aboveit

Fhese aquaria will provide the opportunity to observe a wide diversity of microlite, especially protists. We’ll build several of thosemicrocosms_{initially, some ot which we’ll nmodity later to} observe the effects of environment changes on the populations

oresent.

The second type of microcosm, called a Winogradsky column, isa tall column that contains a thick layer of pond serf irnent arid pond water. We’ll build several Winogradsky columns Lrsing 500 niL or 1 L soft drink bottles, with different columns containing different nutrients, and observe the changes in these columns over the course of several months. Althougti many types of lifeforms will he present initially, the real purpose of the Winogradsky column is to produce a mixed bacterial culture and observe the different types of bacteria that thrive in different rnrcroenvironments within the columns.

Fhe Winogradsky column has been used for decades in biology classes to demonstrate the metabolic diversity of prokarvotes (bacteria and archaea). All organisms require a carbon source and an energy source to live and grow. Some organisms, called

.‘mutotrophs,obtain carbon from atmnspheric carbon dioxide,

Microcosms

are simplified artificial ecosystems that are used to simulate and observe the

behavior of natural ecosystems under controlled conditions, Open or closed microcosms provide

an experimental area for ecologists to study natural ecological processes. Microcosm studies

can

he very useful to study the effects of disturbance

or

to determine the ecological role of key

species. A

Winogradsky column

is an example of a microbial microcosm.

•‘•‘••l’ -ui-. ,t.c u’iach’.c h’’

.1 . in-i c ‘.urs c-ut

Sr

;Jiile otlinr’. cited/rr’tentrnpfra, nbtiri ;.jrbori frcxii tire

.eganic c-omponnds suas carbohydrates that are produced by autotropi a Sim arly. come orgariisiiis, c.afled phototrophs,

obtain eneryv directly tro in sunhght vO photosynthesis, whe fhers, called i’iierrinfiopfi’ obtain enniryfrom hro:iknrg down

I iemical in ip .rmdc

‘ifferirrit crr,rii’ciirc usc all four pusibl” niritimn:if inn’, of these

ir’thodc f nfit,imir Sir lxii arid r’nrp’.’

Photniiitohophs obtas carbon fi nm carbon decode and energy from sunlight

Chemnaulotropfrs obtain carbon frnrn carbon dioxide, hut canirnt ,ht,iin encu” mm sunlight. md somud nhtainif

by bre.ikiii’: mio.vr cflririicum, coiripnund,.

PLif obi:io iiiwfiw.,rii,c,miij.,in:isonm,rlt.iisf from car bohydratos or other organic compounds.

Chemn/rictemotropfrs obtain both carbon and energy by breakinp down cheirricalcompounds.

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ri’rIsimie’’’ ‘cc,.

‘i’:.’irr’’cmt’yl. orrt’,l’’citfciri.b’,..

dl tour of th’’o air jteio’i urn represented ri a typical

‘Iiriogradsky coirririn. whir h also providescivisible example of

nwdifferent organisms occupy different ecoingical niches.

cording to how they obtain carbon and energy

A Winogradoky column, from top to bottom. may include the following ecological mmcrconches:

Aerobic water layer

the cellulose rnsenf iratmafly caumaa microbial bloom, which quickly exfaausts fhe oxygeniiiftc. miedimert and most of t hewatercot rican, leaving only the fop centimeter

orso ofthe wafer cci Irmicin aerated. Aerobic specie; survive

and fl:; ‘h ,cnivmu fbi’. tot th ‘iclnn:n.

[fincur ..‘oi this ,rvci is pa prii;ifeil by strrafhr’d li,-rctercr [he remainder ot the am nhc water aver mc populated by cyanobiiforii(tui nOriv jlld bin r’grer’n at ml. wfnm.n are

theonly photosynthetic bacteriaspecies. In some columns, cyannbci.terma may blnon’i. oxygenating tfieentire water

column and even the fop portion of the sediment columrr.

Anaerobicwater layer

because mygoci diffuses very slowly in water, most nf the wafer columnisanaem ohio (oxygen depleted). hper:ies snot .rc[i’,mi aIm nainanifir L-’,ctcnia fbrmve mcthis10cr

Anaerobic boundary layer

This boundary layer between [Ice anaerobic sediment layer beneath it and the anaerobic water layer above itis

populated by green sulfur bacteria lust above the sediment and purple sulfur bacteria ust above the greerr sultum bacteria. Ihese tactera consume the hydrogen sUlfide gas produced in the anaerobic sedin’rent layer beneath, converting sulfide irns to sulfate ions. The bacteria are vishleascithin purplisfi layer lying just above a thin

greenish layer, which ri turn lies lust above the serhiment.

Anaerobic sediment layer

This layer, populated by anaerobic sulfatereducing species such as Clostridium and Desu/fovihrio. consumes the sulfate ions produced bythesulfur bacteria in the lawn aboveitand releases hydrogen sultide gas. which in turn feeds the sLilfur bacteria

Each Winograd’iky column is unique, even ititstarted with the same combination of water, sediment, arid nutrients, so we’ll make several Winogradsky columns and observe and compare them as they mature.

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PROCEDURE 114-1: GATHERING MATERIALS

FIELD TRIP!

The raw materials we need for

our microcosms

are simple. and can all

be

obtained from the

nearest

pond

or stream: water. sediment. and aquatic plant Ufe. You can

obtain

these specimens

at any time of year, although it

is

best [the source

is

not frozen oVer.

The amounts needed depend on theornnher and size of mic ocosois you infond to0uSd For oxample if you intend to build six Iquoririm rnicroco,rno ri 25 F poanut hurter irsnod four Winner .idsky iii inns ri500 nI soft drink hofflls. you :o(l neer about10 liters ofwnter (cleanSL sotf drink bottles mike convenient collection vessels), perh:rs three liters of sediment. and enough aquatic plant life to provide substantial and diverse plant popi lotions ri eachofthe aquarium rnicrncosnis. It’s better to cot co rub raw rri.rtercrl than not i.r;oiglr.

Whon colluoting the plants nod sediment, try to avoid collecting (s sticks, ,ind offrer large i.rhiects (Well, actually, a snail or

ri fruit. hutit ,ou ricindo sriiil inoreutyour aquaria ni.ike :urr .1 on to iocluJ its ufxrwfOtat(on to make sure the

poor snail doesn’t suffocate.)

5nrxiy typos :1 )lnnts os ps’ otiie 00111mg Ito(t Ouftom, iio.itir(r(iiithew.rtor, andlyingon

furhottorni If (bore is v’;sihle pond scum (01000) uresenf,obtain

unionofif i’ well. possifilo, keep tImplants segregotedby

type, using small jars or zip seal plastic bags to contain them. Colloct sufficient enrinples of each type of plant to provide

hif n onormot your aqnarilTr nicrocosrns. On each

ol(oc[ion ronfa ru_i. note ftr,rtpl.inf5mDrivironirnenf (for uxample.

troc flnafiirp, rooted ri (tie seitirnent, redsonm

Build your microcosms as soon as possible otter you collect the

specimens. lIvenif yoiIalluw the plant and sediment specirriemrs ii fry cut, roust01 the typos of orgarisrns present will survive,

lint orcoil is toi’iprio(rico is closelyas possible tIre erginai

ocusystoiri presentin (ho pondur stream,so the less delay the hotter.

Fit.iii sortiein.dpond or sfre.’rrl sClliiT00rit and vegetation hi use. Sprout (ho irraterial irr a thin layer on newspaper and Slow it to fry thoroughwy in the sh1rrte. (Avnui.t direct sunlight.) Shirethe material mr a labeled paper tag.

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Ion05( b othei’zsaso no..1 (I. nix

0. spoc!n(’( 15015001 (5(05dii for.Tb(DD. (Dcipul Plo os Sb 5.’i ‘ ,

0000 5 .0. ‘0 01

Use o rtrorrie caut ion when you cbtain and handle mond or -1trear’n water, sediment. arrd pl.nrit life. Must of the

‘(arrisrnsferind in typical ponris and streams are (armless to humans. hut mere than afeware hunron pathegens.Always wearglovesandgoggleswhen obtainingor handling specimens, and always wash ihorcugtity with soap and water otterdoing ‘so. if’salso

rgood.ctea to spray or rinse the corrfaniin...rted00terrors

(f sealeri coilecticn containers with disinfectant before

PROCEDURE 11-1-2: BUILDING AQUARIUM MICROCOSMS

ofar as s

possible, our goal s to make each

of

our aquarium

microcosms ienucaI, using the

same

amount

of

sediment.

the

same amount

of water, and the same amount

and

mix of aquatic

plant life.

Arrauce allof you cado 0 dli iison you work surfu’.n di- t555

3. Transfer sofficicrrt pondor stream sediment toeach ar to

fillit_{afoot oonqoartei full Try to keep thu levels in Il jars} the cain-.

0 iisw ‘ or ‘.t.-.s ‘‘-i-cl o.’anD1er-, luar con

foci oct oct00 to iyinp on the hnl torn, piano those p loot specunens accordingly in each of your containers, trying to keep thu quauhhes and row the same in ach of your con to e s

H. _{transfer sufhcirnt pondorstroamwaterto each ar to hil}

abouf 5 cm from fhe top Agarn, try to keep the wafer Insets as similar as possihle,

Ii _{examples of any freedlo,iting plants alp.io and} other ionirnchored flora to the confainers, ,‘ir’airiIi

to ki’op the popoiaton’. and n von the en of r nrc,

7, Replace the lids, seal them tightly, and then spray or drerioh the containers thoroughly with cfisuifecfant I’tilt any orrrai’iisrrison the externr’; of thee-outer

h enter al-icithu oontainer 15. iianc,ioh-is

remain ondislurhed and will be nuposod ail layton ‘lif daylight. hLit not to rtirect sunlight. If necessary, you can put the containers Lrndor a plaid prow light to onsuiPthey recnice adequate light fer oneper dc ev’ ‘lay It

101101 turf thattheconfinnis Ire maritairiid it ,ihiit ‘‘our temperature,soavoid areaswherett’ieymignt hem po’ixI tolaigo temperature variations,

9. Remove and discard your gloves and wash your ‘rinds thoroughlywithsoap and water.

PROCEDURE 11-1-3: BUILDING WINOGRADSKY COLUMN MICROCOSMS

Just as with the aquarium microcosms, our goal is to

make each of our Winogradsky column

it

crocosms

as

similar as possible, other than differences in added nutrients. With the exception

ri

one control Winogradsky column, which contains only unenriched sediment, each of the other

Jinogradsky columns begins with sufficient enriched sediment to fill the column roughly half

ill, To that, we’ll add add about half as much unenriched sediment to fill the column to a total of

Pout three quarters. Finally, we’ll fill the column with pond or stream water, leaving a small air

i’J.

We’555cr nr.r1W’ ‘ccn’rr-c’ madicc’ toni “h”rnoprahnk.y

enrLir’iinrnic.roc’ as, vi ci1k.sccta hottla which ri’qrr rica Pilotiii .“it’rOtit !... c,rctin”s’ct

1, Putonyour gloves and goggles.

2. All but the first Winogradsky colur’nn will contain sediment

supplemented with shredded newspaper (cellutose) toprovide a carbon source. One doubie-width sheet of newsprint will provide sufficient cellulose for those three columns. Shred or cut the newspaper into small pieces.

in ncwspape threucis ,r coritetti isreddieg office

bredder. butyoucan nsteaduse a strip shredder and of the sth rite snss er Hound with ‘;cisshid. If

.ervca’rt. ,se.in ‘e’Ci5Oi.

.3 Transfer enough sediment to half tilt three bottles to a mixini’ owl or simrtar container iti:r n the paper hIs to

mrx them throughout the sediment. Retain the remaining

sediment for use later.

4 Label one bottle “paper only.” Transfer sufficient cellulose’ nor x’hedsediryrintto that bottle to fill it about half full and

set the bottle a’ade.

.1r-zrw nor’. ycli’ Ic the ‘;e’imert i’ernarnot 0 tii rT’’O5

I i iI r Ii I

‘‘.lk provides a suitor source

Ii, Label a second bottle “paperxyolk. Trarrsfer enough ot trio cellulose arid egg yolk enriched sediment to fill that LottIetothe same levelasthe first bottle and set the bottle

mle.

7 Crush the ogg stroll into small pieces and mix thom into tIre ‘Hmpntrerrrainirrg in the mixing, bowl Eggshell provides a

,alcium source.

8. Libel a third bottle “paperC_{yolk ÷ shell.” Transfer enough} the cellulose, egg yolk. and egg shell enriched serlimr’rrt toirS that bottletothe same level as thefirstand secorrd H Ito sand rut the bottle aside. Drscard any nemarnrrrrg

cOo] erlirr’rerrt.

9. Carefully transfer enough urrenniched sediment to the trrst bottle to fill about half the menrr;irnrirrgs.sece. 130 so gently to avoid mixing the sediment layers. Your goal is to have

distinct layers of sediment, with the enriched sedirireret the bottom. cosru od by a layer of rinenrichod sediment

for the rer’rr.-rirrne t:o hottlos. nringrrrg the .,ohiinnnt10the ‘.,rrno level in .‘rll three.

fl _{label a I:irttr hurtle ‘ucerrricl’i’i} ‘ ar’sfor wifticr.ent onsririched sediment to that bottle to bring the level to tho same as the other three bottles,

11. Disturbing the sorliment layers as little as possible,

crrnlnrlly transfer soffrcient poutor stream water tocacti hurtle to fillif, leaving a’ small arm cap at the top ‘at the bottle,

t2. Ppl.’rce the Sits ‘in II for ir Portion coil ttionr tiphtty .rnil

or H H’ ricO rn ‘or s

dsirrtectar’it tonIl artyel.irII”rri5 oilthe enerrors of (tie

containers.

13. Fr..rnsfer all ot the bottles toan area where they canremain ornilisforhedcurtwill he exposed all day to hrigbt daylight,

but rot to direct crirrlght lf necessary. ,‘ou can out the Pulfle’; under a planhgrnw lrgtrttoensure Ihey receive

..irlerluiite lrgl’rtforlong periods every day. It’s important that the hu.tttrrs be r’rr,nrrtorned it .rbort room tnrrnper ‘Pure, so avoid areas whore they night ire exposed to iarge

temperature varrations.

14 Remove and discard your gloves and wash your hands ftiororighly with ‘map arid water.

PROCEDURE 11-1-4: OBSER’J1NG WINOGRADSKY COLUMN

MICROCOSMS

Observin.g a Winogradsky column microcosm is a ongterm project. Some visible changes occur

soon after you build the microcosm, but the major changes take place gradually over rNeeks and

months.

I. Err the first week afterYOL hiGh your Wincgradsky column rnicrocosms, observe eachofthem daily, Note any change

in the appearance of the layers, Ifyouhave a camera, shoot imagesofeach of the micmocosms and file those images by

date and microcosm makeup.

REVIEW QUESTIONS

Q

I

:We cc fct’ in coicu rdnwridlinci, ri ewm specirncnnitr the pOsbiO [)]t••.)

U5inp the Intc ret or otherresou rcce. irt mutt twofliiiirr _{nothogens in each of I he three casses: err} ies.bacteria,ann protozotr.

Q2:

Ot’ .‘i . i rIO’ till 5.with rudimentcurrchnd by carhon. carttonsulfur. end carhon÷sultur +colcurir Using the

.1 rvr,rI mitre c’ierriuntr wr rrrrrht rave used tociii iiithe sediment. The eictrIleof these mtic‘roteis nknnwn. ito

possiblefi en pathogersc: rr r’nc When you hnished oheetieyourVVinogradeky column m:crocesnrc.

disposeof thu hy incinerahon or w submerging them in

bucket of chfonne bleach c;oiution before rn rn onraig, the cap

andmixing II conten softhe n’cr ‘s”m with the bleach sokahen.

Advancedtrid’rrtecon usethese n ire‘‘ore assourcer

of different bach ‘ia species to cultu rr’cample. the

Uhodospiriliumiithrum we use in a later oh sessron’but it’s much less risky simply to buy pure cultures of the species

you need rather then attemptrng to culture them from wild sources Do not open any of these mcrocosmsunless

you are confident that you are competent to work with possibly pathogenic organisms.