Moina as an Epigenetic model organism

drishtantmkawale · April 23, 2020, 9:33am

CUBE Nerul, Navi Mumbai Home Locality Lab
Moina Model System

23 April 2020

Update of the Red-Moina Set-up prepared on 20 April 2020
Breaking News!

There were ~20-25 Moinas in all the Control Cups (200mL DC Water + 1 drop of milk) yesterday, but today there are around 50 in all except the cup C2!
The Test cups have Orangish Red coloured Moinas!

Control: C1- ~60 colourless C2- ~30 colourless C3- ~45 colourless C4- ~45 colourless
I expected there to be around ~40 colourless Moinas in the Control cups.
These expectations have been met more or less!

Test: T2- 10 Moinas (4-Pale Yellow, 3 Orangish Red, Rest colourless) T3- 8 Moinas (3 Pale Yellow, 2 Orangish-Red and Rest colourless)
Here, the number of Moinas have increased in the cups and the colour change of the Moinas from Yellow to Orangish-Red in the test cup is visible before two days i.e. based on my previous observations, I had seen that they turn into orangish-red on the 5th day but today is the 4th day, the colour has changed (not of all the Moinas).

But, the cups T1 and T4 do not have any Moinas so I haven’t discarded them but I have kept them to look for Moinas if there might have been some resting eggs (epiphium).

Expectations for 24 April 2020
Control Cups: ~60 colourless Moinas in each cup.
Test Cups: ~15 Moinas in total out of which ~5 should be Orangish Red and rest colourless and Yellow coloured in cup T2 and T3.

The Control cup C1 having ~60 Moinas today whereas yesterday’s count was ~30.

The Moina on the right side is a Colourless one, with an Orangish Red Moina on the left side.
The Orangish-Red Colour is usually visible on the 5th day but here it is visible on the 4th day itself.

drishtantmkawale · April 27, 2020, 11:14am

CUBE Nerul, Navi Mumbai Home Locality Lab
Moina Model System

27 April 2020

Update of the Red-Moina Set-up prepared on 20 April 2020

Breaking News!
NO RED MOINAS in the Test Cups even after 7 days!

Control: 200mL of Dechlorinated Water + ~10 Colourless Moinas + 1 drop of milk daily.
Control: C1- ~100 colourless C2- ~100 colourless C3- ~50 colourless C4- ~60 colourless
I expected there to be around ~70 colourless Moinas in the Control cups.
These expectations have been met more or less!

Test: 200mL of Dechlorinated Water + ~10 colourless Moinas + 6 drops of milk on alternate days.
Test: T2- 100 Moinas (60- Yellow, 10 Orangish Red, Rest colourless) T3- 80 Moinas (30 Yellow and Rest colourless)
Today is the 7th day of the set-up, I expected around 30 Red Moinas, in the test cups (200mL Dechlorinated Water + 6 drops of milk). But instead, I have been observing Orangish-Red and Yellow coloured Moinas.
My expectations were/are on the basis of my previous observations of the same set-up at CUBE Elphinstone as well as

Expectations for 28 April 2020
Control Cups: ~ 120 colourless Moinas in Cup C1 and C2, ~60 colourless Moinas in Cup C3, ~ 100 colourless Moinas in cup C4
Test Cups: ~120 Moinas in total out which 60 should be yellow 30 Orangish-Red.

Around ~100 colourless Moinas in the Control Cup

drishtantmkawale · April 29, 2020, 7:39pm

CUBE Nerul, Navi Mumbai Home Locality Lab
Moina Model System

29 April 2020

Update of the Red-Moina Set-up prepared on 20 April 2020

Breaking News!
NO RED MOINAS in the Test Cups even after 9 days!
The Moinas in both the test cups have turned colourless😳!

Control: 200mL of Dechlorinated Water + ~10 Colourless Moinas + 1 drop of milk daily.
There are ~100 colourless Moinas in all the Control cups!!

Test: 200mL of Dechlorinated Water + ~10 colourless Moinas + 6 drops of milk on alternate days.
All the Moinas in the test cups T2 and T3 have turned colourless!!

Today is the 9th day of the set-up, I expected around 60-70 Red Moinas, in the test cups (200mL Dechlorinated Water + 6 drops of milk). But instead, I have been observing colourless Moinas in both the cups.

Both Moinas looking colourless!
The Moinas in the left is from the Test cup and the Moina on the right is from the Control Cup.

I have observed this Red-coloured debris!
Why is it Red?

Normally, in the bottles kept in labs as well as the control cups here, the debris is whitish in colour and after a week of getting Red Moinas, the reddish debris appears.

I asked my 14-year-old brother that what does he think?
What must have turned this debris into red colour?
He added that maybe the Haemoglobin of Moina (I had told him about that😬) must have been dispersed in the water!

Also, I had kept all the culture cups in the window for sunlight to fall on them.

The whitish-yellow debris from the control cup.

My Research Question: How does the low-oxygen condition (hypoxia) change the colour of the Moinas and is it due to histone modifications?
Long-term Objective: To prove Histone modification due to low oxygen in Moina using Valproic Acid

My Short-term Objective: To prepare a Red-Moina Setup at home and get Red Moinas during the lockdown period.

Although as per my short-term objective, I had prepared the Red-Moina setup, I haven’t got Red Moinas till now (9 days)!

But I can confidently say that I had induced hypoxia in the test cups which led to the colour change of the Moinas!
I had observed a yellowish as well as Orangish Red Moinas on the 4th and 5th days respectively!

It means that the production of extra-haemoglobin was in process but complete Red colour was not seen.
The estimation of Haemoglobin protein in the Moina can be done by Sahli’s Method of Haemoglobin Estimation (I have this in my Semester II practicals😬)

Although I didn’t meet my expectations of getting ~50 Red Moinas in both the Test cups in 7 days, a colour change was observed (which goes along with my hypothesis).
So,
I WOULD NOT SEE THIS AS A FAILED EXPERIMENT!

jtd · May 1, 2020, 10:28am

There are no failed experiments. Merely experiments that generate data contrary to expectations. Which therefore raises the need for more experiments.

drishtantmkawale · May 2, 2020, 4:54pm

Yes exactly!
An experiment is meant to be a trial which is done to prove or disprove something which we tend to think might/might not happen.
In that, our work is to record the observations and collect data out of it.
The development and application of this data will help us in interpreting, analysing the results of the current experiment and designing the next experiment!

What say?

drishtantmkawale · May 2, 2020, 5:02pm

This is a graph showing the Growth Curve of the Red-Moina Setup which was set up on 20 April 2020.

My objective was to induce a low oxygen condition or hypoxia in the culture medium and observe the colour change of the Moinas over the days which (in simple words) will prove that due to less oxygen in the medium because I added 6 drops of milk in the Test cups consisting of 200mL of Dechlorinated water + ~10 colourless Moinas initially.

Milk has nutrients (carbohydrates-lactose, protein-casein, vitamins, minerals and fat) as well as bacteria. This will be a nutrient source for the DC water bacteria as well as the milk bacteria due to which they will reproduce and consume the oxygen!
Because of which less oxygen would be remaining for the Moinas and this condition will lead to the production of extra-haemoglobin protein due to which the Moinas will appear Orangish-Red or Red!

Talking about the Test cups, the Moinas were Orangish Red in both the test cups on the fifth day i.e. 25th April 2020.

If we jargonise this objective , I wanted to prove that due to hypoxia, the Moina starts producing extra-Hb which is a result of Histone modifications and as per my observations, the modifications happened due to which we were able to see the orangish-red Moinas and also, they turned back to colourless on the 8th day.

This graph follows more or less the same trend as the bacterial growth curve.
There is a bit of variation but I expected a graph like this based on my previous observations!

When I follow the procedure of taking 250mL of DC water + 1 drop of milk + ~10 arbitrary Moinas in a bottle, this is what I expect:
Day 1: 10 Moinas
Day 2: 15 Moinas
Day 3: 25 Moinas
Day 4: 40 Moinas
Day 5: 50 Moinas
Day 6: 70 Moinas
Which means, as the number of days increase, the Moinas increase too.
The peak stage starts from the 10th day onwards where the number ranges till ~200 Moinas on the 14th day!

And there onwards, their number starts decreasing to ~30 Moinas remaining till the 20th day.

This graph has also followed the same fashion.

Now, I will wait till the 14th day i.e. till 4th May 2020 and then reuse these Moina for stock culture.

drishtantmkawale · May 13, 2020, 8:29pm

CUBE Nerul, Navi Mumbai Home-Lab

So this is how we sub-culture Moinas!
Sub-culturing basically means making new culture media for the Moinas and then transferring the Moinas from the existing culture medium.

So here, I have a stock culture of Moinas which came from my previous cultures.
As very less number of Moinas (~20 in each bottle) were remaining in each bottle, I took those (the ones from the five bottles) Moinas and transferred them into a plastic cup which would be my stock culture.
Then, I washed the bottles with soap, water and then dried them.

I had prepared Dechlorinated Water a month ago.
How?
I took half-a-bucket of tap water and kept it open for a day.
The next day, I got dechlorinated water which was then transferred into the green and transparent bottles.

The simplest requirements for maintaining a CUBE Model-Organism which has relations with the 2019 Prize-Winning work in Physiology or Medicine :
Plastic Bottles, DC Water, Moinas, Milk, Dropper, Stickers (for labelling)
All of this except the dropper would be easily available in our soon-to-be homelabs

The above video tells us about the preparation of sub-cultures!
And we see replicates are of utmost importance!
Five - 500mL Plastic bottles (whose volume is not exactly 500mL but more than that) have been included in the design which sum up as replicates of each other.

Bottles: L1-L5 where L stands for Lockdown which means that these bottles are prepared during the lockdown period.
500mL Dechlorinated Water + 2 drops of milk daily + ~20 Arbitrary Moinas.
By Arbitrary Moinas I mean that the Moinas which I transferred are of small/big size, Gravid (pregnant) or Normal and also, I don’t know their age.

Expectations for today - 14 May 2020
~30 Moinas in each bottle.

drishtantmkawale · May 18, 2020, 6:43pm

CUBE Nerul, Navi Mumbai Home Lab
18 May 2020

Objective: To maintain Moina cultures fed with Yeast.

Prepared two Moina Culture Bottles - Y1 and Y2 where Y stands for Yeast.
These bottles have 500mL of Dechlorinated Water + ~10 Moinas and will be fed with 4 Yeast Granules suspended in 3mL of Dechlorinated Water on alternate days.

The Yeast pellets and granules which are being suspended are of Live Yeast or Baker’s Yeast which is used in Kitchens.

Why 4 Yeast granules suspended in 3mL of Dechlorinated Water?
I have no past experience of culturing the Moinas with Yeast. So, I took the help of CUBists from CUBE Kolenchery who have been culturing the Moinas and are feeding them with Yeast. So, @Akshitha and Abhijith Vinod suggested, 4 Yeast Granules would be enough for 500mL of DC Water. But, as I am having granules, they won’t suspend in the water readily, so I thought that I should suspend the granules in the water first and then add in the culture.

Expectations: ~15 Moinas in both the bottles Y1 and Y2.

Why Yeast suddenly?
Yeast is a unicellular organism, a fungus basically which is used in our kitchen for making the dough, bread, cakes fluffy or spongy. Yeast does that by fermentation. So basically, it is used as a fermenting agent. Fermentation is the breakdown of the Carbohydrates or Sugars into Alcohol (-OH) and Acid (-COOH) which also gives Carbon Dioxide (CO2) as a by-product. So, when we add live yeast in the form of granules, powder or pellets, we get a fluffy or spongy or uplifted cake, bread or dough due to the accumulation of gas inside.

When we suspend the Live-Yeast granules in the culture medium, the live yeast cells get dispersed which are then taken up by our crustaceans Moina.

Abhijith Vinod and @Akshitha from CUBE Kolenchery, Kerala have been maintaining the Moinas successfully from August 2019 and are feeding them by suspending the Yeast pellets in the culture medium.
The BREAKING NEWS is that they haven’t changed the water of the culture medium since August 2019 which sums up to 9 months and their culture is still going well!

Here, when I use milk as a feed to the Moinas, for example in 250mL of DC Water + 1 drop of milk daily + ~10 Moinas to start with, they start decreasing at the end of 4 weeks or 30 days or say a month and eventually there are no Moinas left (due to the accumulation of undigested milk, nitrogenous waste and probably low pH) in the bottle (however, I tend to sub-culture or transfer the Moinas in a new culture media).

But here, the culture is going on and on without any disturbance except gathering of some debris and condensation as mentioned by Abhijith Vinod.

So, here we can see that CUBists have developed another method of maintaining an organism which is a part of the CUBE lineage by finding out an alternative of Milk; YEAST both of which are a part of our Kitchen (Kitchen labs!!).
This is what we want!

Correct me if I am wrong anywhere.

The two Moina culture bottles Y1 and Y2.
The 4YG-Alter on the sticker means that 4 Yeast Granules to be fed on alternate days i.e. after a gap of one day.

A very slight movement of the Moinas detected in the bottle.

Here too, movement is seen. I tried taking a shot from the top, but the water is appearing to be turbid a bit so the Moinas won’t be visible. Although, I need improvements😬.

drishtantmkawale · May 28, 2020, 10:07pm

CUBE Nerul, Navi Mumbai Homelab
May 28, 2020

BREAKING NEWS!
Greenish colour observed in Moina bottles fed with Yeast!
Totally unexpected!!

The two bottles Y1 and Y2 (having 500mL of Dechlorinated Water + ~10 Moinas fed with 4 Yeast Granules suspended in the Water) which were prepared on 18th of May 2020 with an objective of culturing the Moinas by feeding them with Yeast [usually fed with Bacteria (Milk)], when I observed them today, showed some greenish colour at the base and on the walls of the bottle!

History of bottles Y1 and Y2 (same observations are seen in both the bottles):
18th May: ~10 Moinas fed with suspension of 4 Yeast Granules
19th: ~15 Moinas observed in both the bottles. (Not fed on this day)
20th: ~5 Moinas observed in both the bottles. (fed on this day)
21st May: No Moinas were observed.

So we saw that the Moinas died on the 4th day.

These bottles were kept near the WIndow so that they could get some light (not direct sunlight).

What could be the possible reason for the greenish colour?
Algae? Yes, that could be possible due to its appearance on the walls. In CUBE HBCSE, I had observed similar kind of greenish-growth in the Moina tanks and there, the culture was self-sustaining i.e. the Moinas were surviving without feeding them anything (like bacteria or anything externally) just like a Natural-Ecosystem!

If it is algae, where did it come from?
No goof-ups as such had taken place (well, I am no one to say that but some goof-ups may arise while we discuss!).

The water used was tap water which was dechlorinated (by keeping it overnight) and then used.
The 500mL plastic bottles were newly-opened Mineral water bottles.
The Moinas were taken from the Moina stock (currently having ~200 Moinas).
The feed was suspension of Yeast Granules (live Baker’s Yeast).
The Dropper/Pasteur Pipette used, was a new one.

After 21st May, till 28th May, I had not observed the bottles.
Yesterday I observed this and saw Greenish growth!

Further plan: To keep these bottles at the same place as earlier i.e. near the window, take photos and videos everyday and to see whether the greenish growth sustains and increases.

The photo of the bottle Y1 showing prominent greenish growth at the base of the bottle as well as on the walls which is speculated to be Algae.
Let’s see if this increases.
Date of Video: 28th May, 2020
Time: 11.30pm
Data Collector: Drishtant MK.

The photo of the other bottle Y2 showing very little yellowish-green growth at the base of the bottle and none on the walls which is speculated to be Algae.
Date of Video: 28th May, 2020
Time: 11.30pm
Data Collector: Drishtant MK.

Let’s discuss this!
Is this another Penicillin-Alexander Fleming kind of moment!!?

drishtantmkawale · May 29, 2020, 8:48pm

CUBE Nerul, Navi Mumbai Home lab
May 29, 2020

BREAKING NEWS!!
The Yeast fermented the Sugar!!!

From the past week, I was in a worry that why didn’t the Moinas which were fed with suspension of Yeast Granules survive!
Abhijith Vinod from CUBE Kolenchery had suggested that may be the Yeast Granules which I had used were not viable or usable.
Why?
I had used the yeast granules from the packet which was opened 9 months ago whereas the shelf-life of Yeast granules is 4 months.
So how could I check whether the Yeast which I have is viable or not?
I designed a quick experiment on the basis of the simple of fermentation by Yeast.

What is Yeast?
What is fermentation?

Yeast is an unicellular fungi which is used in our kitchens as a fermenting agent in baking the dough, making the breads, cakes fluffy/puffy. This puffiness or softness is due to the fermentation done by Yeast.

Fermentation is the breakdown of carbohydrates present in the substrate performed by Yeast giving out acid, alcohol and Carbon Dioxide (CO2) gas as by-product. As there is an evolution of gas, it fills inside the bread, cake and makes them soft and uplifted.

So, with the help of the ingredients and materials available at home, I performed an experiment to prove whether the Yeast which I am having is working or not.

The Hypothesis:
The Yeast granules which I am using has proteins, fats, vitamins and minerals along with the organism Yeast in inactive or dehydrated form. So, when I will suspend these granules in the water, they will get activated and then it will ferment the sugars present inside the water. Some bubbles/effervescence would be seen after some time.

The 9-month old Yeast granules won’t show effervescence whereas the 1-month old will show.
Here, I used 4 cups half-filled with warm water (warm, as suggested by my Mother), 9-month old Yeast granules, 1-month old Yeast granules, Sugar granules as a source of Sucrose which is a carbohydrate which will be broken down/hydrolysed.

As always, I used replicates along with test and control. Control is used for the comparison with the test.
Control: 2 cups half-filled with warm-water + 1 teaspoon or 5gms of sugar + half-a-teaspoon of 1-month old Yeast Granules.
Test: 2 cups half-filled with warm-water + 1 teaspoon or 5gms of sugar + half-a-teaspoon of 9-month old Yeast granules.

After this was done, I kept it for 15-minutes.

When observed after 15mins, I saw that the control cups were showing bubbles which showed Carbon Dioxide is evolving and the test cups were almost clear with very less bubbles.

This shows that the older Yeast is not viable hence did not work and the newer Yeast did show results!
Now planning to design an experiment in which the Moinas whill feed on Yeast and the experiment will include Control as well as replicates.

This was totally an out of the box experiment thanks to @Arunan, who suggested me this during the May 29 Webinar!

The cups before the incubation time of 15-minutes. Having warm water, yeast and sugar.
The right ones are control and the left ones are test.

Date and Time of Photo: May 29th, 2020 at 11.30pm.
Data Collector: Drishtant MK.

In these photos, we can clearly see the bubbles which are popping in the two cups above shows that the Yeast which was inactive in the Granular form, now had become active when suspended in the water and very less bubbles, a faint layer is seen the cups below.

Date and Time of Photo: May 29th, 2020 at 11.45pm.
Data Collector: Drishtant MK.

This shows that the complex processes which we study in our textbooks, are having such applications which can be done at home too!
A practical application of fermentation was seen here!
Just a bit interest needs to be cultivated. That’s it!!

drishtantmkawale · May 30, 2020, 7:58pm

CUBE Nerul, Navi Mumbai Home Lab

Plan of Work and the design of the experiment

Now that I have concluded by the experiment performed yesterday that the Yeast granules which I had used earlier was not able to sustain Moinas, the newer Yeast granules showed bubbles when suspended in warm water and sugar which showed that sugar was broken down. So, I will be using the 1-month old Yeast granules for feeding the Moinas in my new setup.

Objective: To maintain a culture of the Moinas by feeding them with Yeast instead of its traditional feed Milk.

Design of the experiment:
I will be using Moinas, 8 - ~200mL plastic cups, 2L of Dechlorinated water, Live Yeast (Baker’s Yeast in the form of inactive Yeast granules), Milk, and droppers/pasteur pipettes.

The 8 cups will be divided into 4-control and 4-test.
All will be having 200mL of Dechlorinated Water + ~10 Moinas in them.

The control, further would be divided into two and two;
The first two (replicates of each other) would consist of 200mL of Dechlorinated Water + ~10 Moinas and nothing will be fed to them externally.

Why?
As per my observation in the previous Moina-Yeast experiment, the Moinas died after 3 days. Despite feeding them with Yeast granules (which now I came to know that were not viable😬) suspended in the water twice. So, I suspect that the Moinas were just alive on bacteria which were there in the Dechlorinated Water.

I am starving them to death (serious crime ) so that I will come to know, how many days will they survive just on the bacteria present in the Dechlorinated Water and also some nutrients which came along with the Moinas in water in the form of water from the earlier culture medium from where they will be taken from (although it is a minute variable, it needs to be looked upon!).

The other two Control cups (these too, replicates of each other) would be having 200mL of Dechlorinated Water + ~10 Moinas + 1 drop of milk daily.

Why Milk suddenly?*
Milk is being used as my control as I know that the crustaceans will definitely survive when I am feeding them with Bacteria.
Milk = Carbohydrates (Lactose) + Proteins (Casein) + Vitamins + Minerals + Fats + bacteria.
DC Water = Bacteria + some dissolved minerals and nutrients.

The bacteria in both the cases will feed on the nutrients present in the milk (and will replicate) which then would be consumed by Moinas.

The 4 Test Cups would all be the same; having 200mL of DC Water + ~10 Moinas + 2 Yeast Granules suspended in 2mL of warm water (to hydrate, activate the dry yeast) and then adding it in the bottles on alternate days.

Note: The proportion of yeast is as per suggested by Abhijith Vinod and Akshitha from CUBE Kolenchery, Kerala as they have been culturing Moinas using Yeast since August 2019. So, 4 granules to be suspended in 500mL of Dechlorinated Water was suggested. Here, as I am using 200mL of Water, the number of granules are now halved.

Expectations:
Control 1 (starving): Moinas in both the cups should survive till 3 days (just an assumption which will be confirmed).
Control 2 (Milk): Moinas shall survive and will reproduce in both the cups.
Test (Yeast): Here too, they should survive and reproduce in all the cups.

Note: Plastic cups are being used because I don’t have enough plastic bottles.

Photos and Videos will be shared after the set-up is prepared!

Please contribute by improvising this design!

drishtantmkawale · June 5, 2020, 11:53am

CUBE Nerul, Navi Mumbai, Home lab

Update and Plan of Work 5th June 2020

Breaking News!!
The greenish growth is flourishing!!

The first instance of observing the greenish growth in the two bottles Y1 and Y2 was on 28th of May, 2020.
Since then till yesterday i.e. till 4th of June, 2020 the growth has been increasing.

I suspect this greenish growth to be algae.

Why?
I can provide the following information:

The greenish *growth has been increasing day by day.
As I am keeping the two bottles in the window to get sunlight (not direct sunlight), the growth is seen to be increasing, it shall be a photosynthetic algae which is sustaining in the water.
Here, I had taken Dechlorinated Water , which is freshwater . So, the algae surviving in that shall be a freshwater photosynthetic algae.

Further plan:
To introduce Moinas into the two bottles having suspected-to-be- algae.

Moinas are found in freshwater bodies like ponds, lakes and shallow streams where they feed on Algae, Bacteria, Fungi and decaying matter.

Keeping this is mind, when the Moinas will be put in these bottles having 500mL of dechlorinated water + greenish growth, I expect them to sustain/survive in it.

I have observed the same greenish growth in one of the Moinas tanks in CUBE HBCSE, Mumbai where Moinas were surviving without adding any external source of food such bacteria.

It will be mimicking their natural ecosystem!

As this too is an experiment, it shall have a defined control as well as test, both with replicas.

Control will show that the Moinas will survive definitely.
In Test, I will be checking (by adding Moinas) whether the survive or not.

I was planning to prepare 5 stock culture bottles with 500mL of dechlorinated water + 2 drops of milk + ~10 Moinas in each bottle.
And also, my control will consist of the same. So, I need not make a separate control for the experiment. The stock bottles will act as a control.

The test, two bottles Y1 and Y2 having 500mL of dechlorinated water + ~10 Moinas + greenish growth (assumed to be algae) inside the bottle as the feed.

Why only two bottles for test?
Actually, this was a spontaneous observation of greenish growth (I didn’t expect that). So, I thought that I should introduce some Moinas in them and see if they survive by feeding on algae. So, just two bottles have this greenish growth.

I do not know how do Moinas feed on algae (but they are alive). So everyone’s contribution is needed in finding that out.

Note: From the start, I have five Moina stock culture bottles fed with milk. So, when their growth curve starts coming to an end, I gather all the Moinas in a cup and then wash the bottles with tap water, then I prepare a new culture media for them. It consists of 500mL of DC Water + 2 drops of milk (in the bottles), later in which I add Moinas.

So right now, I have around 100 Moinas as my stock.

The above pictures show the increase in the greenish growth from 28th of May, 2020 to 4th of June, 2020, in the bottles Y1 and Y2, both having 500mL of dechlorinated water and are kept in the window to get sunlight (not direct sunlight).

Date of Pictures: 28th May 2020 to 4th June 2020
Data Collector: Drishtant MK.

drishtantmkawale · June 5, 2020, 6:14pm

The picture depicting bottles L1-L5 (having 500mL of dechlorinated water + ~10 Moinas + 2 drops of milk) prepared today as the stock culture. These bottles are also a Control for the Test bottles which are showing greenish growth.

Date of Picture: 5th June 2020
Data Collector: Drishtant MK.

Two Test bottles A1 and A2 (‘A’ stands for algae) having 500mL of dechlorinated water + ~10 Moinas + greenish growth as visible which is suspected to be algae. Moinas shall feed on the algae.
I was very curious to add Moinas in these bottles. So, I added ~10 Moinas (from the stock culture) in both the bottles before the Webinar started. Now to my astonishment, there are ~20 and ~15 Moinas including juveniles in bottles A1 and A2 respectively, which means that there would’ve been gravid (pregnant) females too in the Moinas which I added. Good to see.

Date of Picture: 5th June 2020
Data Collector: Drishtant MK.

Video showing the movement of around 20 Moinas in bottle A1 with flourished greenish growth. I expect that the Moinas shall survive as well as reproduce in here as this bottle is now like their natural ecosystem!

Date of Video: 5th June 2020
Data Collector: Drishtant MK.

drishtantmkawale · June 7, 2020, 8:07pm

CUBE Nerul, Navi Mumbai Home Lab
Update - 7th June 2020

Breaking News!!
The culture is flourishing!!!

Although this isn’t closer to my research objective of studying the colour change in Moinas, I saw this thing as interesting!

As updated earlier on 5th June 2020, ~10 Moinas were added in the bottles A1 and A2 having 500mL of dechlorinated water and greenish algal growth. Their number had become ~20 at the end of the day on 5th June 2020 (whereas the Moinas were initially added in the afternoon).
On the 6th of June 2020, there were around 50 Moinas in both the bottles. The astonishing thing here is that their number doubled in 24hours!
Yesterday, i.e. 7th June 2020, bottle A1 had ~70 Moinas and bottle A2 had ~60 Moinas.
These are good numbers according to me!

Such numbers (of Moinas) are obtained after a week of culturing them in 500mL of dechlorinated water + 2 drops of milk daily + ~10 Moinas to start with.

But🤔, there can also be a case that coincidentally, the ~10 Moinas which were added on 5th June, they must have been mature adults as well as gravid (pregnant) females due to which, such growth is seen.

I have five bottles L1-L5 having 500mL of DC water + 2 drops of milk daily in which the number of Moinas aggregate to ~100.

Further plan is to prepare a Moina-Yeast culture in which the Moinas will be fed with live Yeast. This will be done when there will be ~300 Moinas in aggregate so that I can take ~100 Moinas from it. Also, I need 8 cups in total for the experiment, currently, I am having 4. So, need to gather them too.

Video showing ~40 Moinas from bottle L4 (having 500mL of DC water + 2 drops of milk). We can see the movement of small/juvenile Moinas along with the adult ones.
Couldn’t take a video from the top as it is cloudy (due to milk) right now.

Date of Video: 7th June 2020
Data Collector: Drishtant MK.

The Star
Bottle A1 with ~70 Moinas including juveniles and adults feeding on the greenish algae; a similar ecosystem resembling the pond.
Rotifers, bacteria, Moina, algae, decaying matter.

Date of Video: 7th June 2020
Data Collector: Drishtant MK.

This the the design for the Moina-Yeast experiment.

drishtantmkawale · June 12, 2020, 6:59pm

CUBE Nerul, Navi Mumbai Home lab
Update-11th June 2020

Breaking News!!
Algae-The Killer?
The Moinas in the algal bottles have died!

Bottles A1 and A2 are the test bottles.
The bottles A1 and A2 (500mL of dechlorinated water + algal growth) had been transferred with ~10 Moinas on 5th June 2020.
6th June: 50 Moinas in both the bottles. This trend continued till 9th of June where A1 had ~100 Moinas and A2 had ~60. On 10th June, A1 had around 50 Moinas and A2 had 30 Moinas.

And yesterday i.e. 11th June 2020 when I saw, there were none in A1 and just 2 Moinas in A2!

What could be the reason?
How is this possible that all the Moina in both the bottles died!!?

If we compare the test bottles with the control bottles L1-L5 (having 500mL of dechlorinated water + 2 drops of milk daily) which were prepared on the same day i.e. 5th June, these bottles have ~70 Moinas in them in a week.

Further plan: I will keep the bottles as they are. We saw that the Moinas have died in both the bottles. We had concluded on the basis of the discussion a few days back that the Moinas die if there is an unfavourable condition.
And also, for their generations to continue, there is a possibility that Moinas must have left dormant eggs which shall hatch after the favourable conditions occurs for them.

So I will wait for a week at least to see if I can see the Moinas.

No movement of Moina is seen in the bottle A1 which too had ~50 Moinas two days back.

Date of video: 11th June 2020
Data Collector: Drishtant MK.

The movement of just a single Moina is visible in bottle A2 which had ~30 Moinas 2 days back!

Date of video: 11th June 2020
Data Collector: Drishtant MK.

One of the five Control bottles L3 showing the movement of around *100 Moinas including juveniles and adults both.

Date of Video: 12th June
Data Collector: Drishtant MK.

drishtantmkawale · June 15, 2020, 1:26pm

CUBE Nerul, Navi Mumbai Home lab

Moving ahead with the objective: to check the growth of Moina (again) in algal water.
For this, I will transfer ~10 Moina in both the bottles A1 and A2 having 500mL of dechlorinated water and algae along with the debris of the dead Moina along with yeast.

Before that, I will grow some algae in two plastic cups AG1 and AG2 having 200mL of dechlorinated water and in that I will add ~10mL of algal water from the bottles and will keep them like that to see whether algae will grow in the cups.
If it grows, then Moina will be transferred in them.

This is the graph showing the timeline of bottles A1 and A2 having Moina feeding on Yeast (earlier)and then Algae.
Prepared by: Drishtant MK

Lateral/Side view of the bottle A1 showing the algae on which the Moina were feeding earlier and now, there are no Moina.

Date of Video: 14th June 2020
Data Collector: Drishtant MK

Lateral/Side view of the bottle A2 showing comparatively less algae as compared to bottle A1 but still there are no Moina.

Date of Video: 14th June 2020
Data Collector: Drishtant MK

On the other side, the lateral/side view of one of the five control bottles (fed with 2 drops of milk in 500mL of dechlorinated water) showing the movement of around 150 Moina. These control bottles were made on the same day when the Moina were transferred in the algae water.

Date of Video: 14th June 2020
Data Collector: Drishtant MK

drishtantmkawale · June 17, 2020, 12:08pm

CUBE Nerul, Navi Mumbai Home lab
17th June 2020-Plan of work and Design of the experiment

As posted and discussed earlier, the Moina in the two bottles having algae died.
So, as this was a sudden event and I didn’t see this coming, I am not sure why/how did the Moina died on 11th June 2020. This has been done just once, so it would not be right to derive a conclusion.

So, I will be conducting an experiment here in my home lab in which the Moina will be feeding on the algae.

Design of the experiment
I will be having 6 cups and 2 bottles.
All the six cups will be having 200mL of dechlorinated water + ~10 Moina in them.
They (cups and bottles) are going to be divided into control and test (along with replicas).

Positive Control: This will ensure that the Moina will survive definitely in this.
2 plastic cups PC1 and PC2: 200mL of DC water + ~10 Moinas + 2 drops of milk daily.
Expectations: the Moina will survive and reproduce throughout the experiment as bacteria + nutrients (carbohydrates + fats + vitamins + proteins + minerals from milk) will be given to them

Control: This will tell us how many days will Moina survive without any external feed apart from the bacteria dissolved in the DC water.
2 plastic cups C1 and C2: 200mL of DC water + ~10 Moinas + No feed.
Expectations: I haven’t tried this earlier, so I think that Moina shall survive a couple of days (or more) and then die as there will be no more bacteria as a feed for them.

Test 1: This will have ~20mL (just an arbitrary volume) of algal water (which shall have algal spores) from the bottles A1 and A2 in which the greenish growth of algae was seen and the rest ~180mL will be dechlorinated water.
2 plastic cups AM1 and AM2 (AM= Algae Moina): 200mL of DC water + ~10 Moina feeding on algal spores.
Expectations: They shall survive until the algae, as well as bacteria, is there.

Test 2: Here, we come back to the original bottles A1 and A2 with algae. Everything will remain the same, just ~10 Moinas will be added in these in which they will feed on the remaining algae in the bottles (if no toxicity is involved here).
2 plastic bottles A1 and A2: 500mL of DC water + remaining algae + ~10 Moinas.
Expectations: Moina shall survive in these bottles by feeding on the algae.

Other than these, I have 4 stock culture bottles having an aggregate of ~300 Moinas.

Photos and videos of the set-up will be posted after their preparation.

drishtantmkawale · June 19, 2020, 1:03pm

CUBE Nerul, Navi Mumbai Home Lab
Update-17th June 2020

Breaking News
The culture set-up for the algae experiment has been prepared!

Apologies for retrieving back late.

So, as per the design of the experiment posted earlier, I have prepared the culture setup.
It includes 6 cups and 2 bottles.
Cups
Positive Control: PC1 and PC2 having 200mL of DC water + ~10 Moinas + 2 drops of milk daily
Expectations: ~20 Moina the next day
Control: C1 and C2 having 200mL of DC water + ~10 Moinas + No feed.
Expectations: ~20 Moina the next day
Test 1: AM1 and AM2 (AM= Algae Moina) having 200mL of DC water + ~10 Moina feeding on algal spores.
Expectations: ~20 Moina the next day.

Bottles
Test 2: A1 and A2 having 500mL of DC water + remaining algae + ~10 Moinas.
Expectations: ~20 Moina the next day.

The expectations in all the cups and bottles are the same as the number of Moina too are the same and the expectation ~20 Moina is on the basis of previous experience.

There is also a chance that the Moina in the bottle A1 and A2 had died earlier due to toxicity or unfavourable condition, so they must have left behind dormant eggs too, which may account for the sudden increase in number (just a possibility)

A top view of the cups C1 and C2 showing the movement of ~10 Moinas.

Date of Video: 17th June 2020
Data Collector: Drishtant MK

Top view of the cups PC1 and PC2 showing the movement of ~10 Moina.

Date of Video: 17th June 2020
Data Collector: Drishtant MK

Bottom view of the cups AM1 and AM2 showing the aggregation of ~10 Moina.
This was taken from the bottom because the Moina weren’t coming upwards when I flashed it up.
So I shook in front of them

Date of Video: 17th June 2020
Data Collector: Drishtant MK

Bottom view of bottle A1 showing a lush greenish base of algae on which the Moina are feeding.

Date of Video: 17th June 2020
Data Collector: Drishtant MK

Bottom view of bottle A2, here too showing a greenish base of algae on which the Moina are feeding.

Date of Video: 17th June 2020
Data Collector: Drishtant MK

drishtantmkawale · June 19, 2020, 1:20pm

CUBE Nerul, Navi Mumbai Home Lab
Update-18th June 2020

Breaking News
It’s the second day, and they are surviving!

The Moina-Algae setup (prepared on 17th June 2020), on the second day, still had Moina in it.
Which means, some unfavourable condition (as observed earlier) isn’t there yet.
Good!

The videos below are the evidences:

Lateral/side view of the control cups C1 and C2 showing ~15 Moina in both the cups. Here, nothing is added as an external feed to them.

Expectations have been met more or less.

Expectations: Two possibilities; Either ~25 Moina or less than 15 Moina the next day (as nothing is being fed)

Date of Video: 18th June 2020
Data Collector: Drishtant MK

Lateral/side view of the Positive control cups PC1 and PC2 showing the movement of ~20 Moina in both the cups.
The cups are behaving as expected.

Expectations: ~30 Moina on the next day.

Date of Video: 18th June 2020
Data Collector: Drishtant MK

Lateral/side view of the test cups AM1 and AM2 in which there are ~15 and ~10 Moina (negligible) feeding on the Algal water/spores.

AM1 has met the expectations more or less, but AM2 hasn’t.

Expectations: ~25 Moina in AM1 and ~20 in AM2 the next day.

Date of Video: 18th June 2020
Data Collector: Drishtant MK

Lateral/side view (with torchlight at the opposite end) of one of the test bottle A2 having ~15 Moina feeding on algae.

Here too, the expectations have been more or less met.

Expectations: ~25 Moina on the next day.

Date of Video: 18th June 2020
Data Collector: Drishtant MK

Lateral/side view (with torchlight at the opposite end) of one of the test bottle A1 having ~20 Moina feeding on algae.

Here too, the expectations have been more or less met.

Expectations: ~30 Moina on the next day.

Date of Video: 18th June 2020
Data Collector: Drishtant MK

drishtantmkawale · June 20, 2020, 7:59pm

CUBE Nerul, Navi Mumbai Home Lab
Update-19th June 2020

Breaking News
Third day, slow and steady!

The Moina-Algae setup (prepared on 17th June 2020), on the third day, Moina are still seen but a few fluctuations are there in their number. The amount of Algae in the bottles has also reduced which means Moina are feeding on it!

The videos of the control and test cups and bottles below are the evidence:

Lateral/side view (with torchlight at the opposite end) of one of the test bottles A1 having ~15 Moina feeding on algae. The amount of algae has been reduced.

Here the expectations have not met.
So we can say that, as seen earlier, due to onset of some unfavourable condition, Moina had died. Here, it has not happened yet but their number too, hasn’t increased and their movement is retarded too.

Expectations: ~25 Moina on the next day.