Transgenic microalgae

Why transform microalgae?

Basic research
‐ Photosynthesis (Chlamydomonas reinhardtii)
‐ Cilia/flagella (Chlamydomonas reinhardtii)
‐ Development of multicellularity (Volvox carteri / Chlamydomonas reinhardtii)
‐ Primary and secondary endosymbiosis (Rhodophytes, Diatoms, Cryptophytes)
‐ Extremophiles Extremophiles (Cyanidioschyzon Cyanidioschyzon merolae merolae, Chlamydomonas Chlamydomonas nivalis/acidophila nivalis/acidophila)

Ecological importance
‐ ~50% of annual carbon fixation by microalgae
‐ Algal blooms (Aureococcus anophagefferens)

High‐end commercial products
‐ Carotenoids (Haematococcus, Dunaliella)
‐ Feed stock for aquaculture (Nannochloropsis) ‐ Food supplementals (Chlorella)
‐ Hydrogen Hydrogen (Chlamydomonas Chlamydomonas reinhardtii reinhardtii)
‐ Biodiesel (Botryococcus braunii)

Practical 6 : Propagation of seagrass in artificial seawater

INTRODUCTION

Seagrasses are the only flowering plants that can live underwater. They grow in sediment on the sea floor with erect, elongate leaves and a buried root-like structure (rhizome). Seagrasses live in the coastal waters of most of the worlds’ continents. They are the main diet of dugongs and green turtles and provide a habitat for many, smaller marine animals, some of which, like prawns and fish, are commercially important. They also absorb nutrients from coastal run-off and stabilise sediment, helping to keep the water clear. Therefore, it is important for us to know the techniques of seagrass culture to maintain their biodiversity and for other uses.

METHODOLOGY

Every group cultured their own seagrass species that has been taken during last sampling procedure at Teluk  Pelanduk.

o   A culture system was filled using two aquarium tank consist of artificial seawater with 15 ppt of salinity.

o   A white cloth was placed inside the tray to act as a substrate. Plant seagrass above the surface of the sandy substrate. Each tanks contain three plastic trays; arranged at different sides: left,middle and right side.

o   The plastic trays were labelled.

o   Air stones were fixed to mainatin water circulation in the aquarium tank.

o   The aquaculture system was keep under natural light (16L:14D)

o   Aquarium tanks were observed on a weekly basis; salinity,temperature and the growth rate of seagrass.

Figure 1 1

Figure 1 2

Figure 1 3

Figure 1 4

o   Rainwater was added if there is evaporation.

o   The leaf length,width and petiole length or any changes that can be observed of seagrass were recorded.

RESULTS

Date : 22 Oct 2015

Water parameter

pH                                               : 8.04

Ammonia                                   : 0

Salinity                                        : 37 ppt

Dissolved oxygen (DO)            : 52.6%

Temperature                             : 25◦c

Conductivity                              : 59 ms

Total Dissolved Solid (TDS)     : 39.3 g/l

Our seagrass cultured was failed. They didnt grow new leaf neither petiole lenght stay the same as previously.

Field Trip to Aquascape Paradise

On 29 October 2015, class of Aquatic Flora Culture went for a trip to Aquascape Paradise,Shah Alam.

The objectives of this trip :

• To gain knowledges about various types of freshwater macrophytes
• To get ideas on our mini project

There are many concept that are being used in aquacaping such as :

• Iwagumi concept - rock formation
• Biotope concept - waterfront plants
• Dutch concept - identity

Examples of aquacape that availables there :

About 114 species of freshwater macrophytes are availables. 

For examples :

• Java fern
• Anubias sp.
• Cabomba sp.

We were also demonstrated on techniques of aquascaping which is shown below :

Before

After

At the end of this trip, we were managed to know :

• Various species of freshwater macrophytes that are marketable
• Various concept and techniques of aquascape
• Materials and equiptments needed for aquatic plants maintenance

Issues/problems in aquatic plants

Submerged Plants

An excessive amount of submerged plants can cause problems for the pond owner. The exception might be for the dedicated wildlife pond in which no other use is desired. When submerged plants are too abundant, they can cause problems similar to those described for filamentous algae.

Excessive amounts of submerged plants can present a problem for the pond owner who values his/her fishery. Once plant levels exceed 20% of the surface area, largemouth bass predation on bluegill becomes less effective. Small bluegill can effectively avoid being eaten by darting into the excessive vegetation. Thus, too many bluegill survive, and their growth declines due to increased competition from overcrowding. Bass growth also decreases, as they are unable to find prey effectively and consume enough to grow well. The classic symptoms of this scenario are populations of small, thin largemouth bass and bluegill.

Excessive submerged plants also pose a problem in ponds where water is being pumped out for irrigation, livestock watering, or might be pumped out in case of a fire. These plants can clog a pump intake, lessening the amount of water being pumped and shortening the lifespan of the pump due to excessive wear and tear.

Floating Plants

An excess of nutrients can cause an overabundance of watermeal and duckweed. It is not uncommon for these small plants to completely cover a ponds surface in a few short weeks. This type of cover will substantially reduce light penetration so that oxygen-producing photosynthesis in the water underneath the floating plants ceases. Warm water temperatures during summer months keep oxygen-consuming animal respiration and decomposition rates high, possibly resulting in a fish kill from lack of oxygen.

Water lilies are notorious for overspreading a pond in short order, especially if the pond is shallow and the water is clear. Clear water allows these lilies to send up stems and leaves from a greater depth. This severely inhibits recreational activities. Many pond owners have planted water lilies for aesthetics, only to be frustrated with their quick coverage of the pond.

Emergent Plants

An overabundance of emergent plants can create a problem for some pond owners. This is particularly true for cattails, which account for nearly all pond owner complaints about emergent plants. Cattails have the ability to completely surround a pond and extend several feet into the water if allowed to do so. Even for a pond managed for wildlife, this poses a problem. Habitat diversity is good for wildlife, and a monoculture of cattails does not provide diversity. Excessive emergent plants can create problems for anglers who will find it difficult to effectively fish from shoreline areas.

Cattails are highly attractive to muskrats, a mammal that can damage a pond in some circumstances. Dams are vulnerable to their burrowing activities, and muskrat burrows can compromise their integrity. Muskrat use cattails for a variety of reasons, including food, den material, and as an escape from predators.

One emergent plant species deserves special mention. Purple loosestrife is an invader to Ohio ponds and wetlands and should be controlled, either by pulling the plant and roots or by spraying a herbicide. If left uncontrolled, purple loosestrife quickly spreads and crowds out desirable native emergent plants. A monoculture stand of loosestrife is the result, providing no benefits to the pond owner or wildlife.

                                                                                                                   (http://ohioline.osu.edu/)

Other issues :

Non invasive, native plants are much more desirable than non native, invasive varieties. Non native plant species out compete native species for nutrients because the native species are in better balance with the entire system. The non native species that overtake a pond are rapid growers and do not have the organisms to keep them in check because they are not native to the area. Native plants can also grow rapidly if not monitored and held in check. Therefore, the key is balance and preventing one specific aquatic plant from overtaking your entire pond. A nice mix of species will provide a healthy pond ecosystem and also a beautiful setting.

Nutrients are the main cause for excessive aquatic plant growth. Only a small amount of nutrients added to a pond can cause a lot of problems with aquatic plant growth. It is important to look at the source of your ponds nutrients and try to get a handle on that while you are working on the immediate problem with aquatic plants.

Treatments :

Physical/Mechanical Treatments

Chemical Treatments

Copper

Diquat Herbicide

Fluridone Herbicide

Biological Treatments

Aquatic Plants

Plant Eating Fish

Bacteria

Pond Environment Treatments

Water Level

Flushing

Shading

Water patterns

Dredging

                                                                                                      (http://www.dunnsfishfarm.com/)

Practical 5 : Propagation of seaweed in enrichment media

     INTRODUCTION

Each group was given the same species of seaweed, Glacilaria changii but different group need to use different media which is each of them consists of 3 replicates.

Seaweed

Glacilaria changii

Media

Sterile seawater,VSE,PES

Bacterial cultures

17 different bacteria

Aquaflora : SS,VSE,PES,B 

Phytoteam : B1,B2,B3,B4 (SS)

Floratic :  B5,B6,B7,B8

Greendot :  B9,B10,B11,B12

Floria :  B13,B14,B15,B16

*SS = sterile seawater

SW = seaweed

B = bacteria

 METHODOLOGY

o   The seaweed was placed in a container filled with seawater.

o   Seaweed was rinsed in a series of containers by submerging and shaking vigorously underwater several times.

o   The wet weight of seaweed was recorded.

o   Seaweed was introduced in beaker containing Von Stosch’s Enrichment (VSE) and Provasoli Enrichment Seawater (PES) media.

o   Seaweed was incubated for a photoperiod of 12hr light and 12hr dark with aeration.

o   The growth was compared by determining the gain / loss of the seaweed wet weight.

o   Any changes  that can be observed were recorded.

1)Wash with tap water

2)Cut 1cm-1cm

3)Weight

4)Wash with sterile seawater (30 s)

5)Fill well-plate with sterile seawater

6)Add 10^4 cfu/ml of selected bacteria

7)Put seaweed

8)Observe 7 days-7 days

RESULTS

Sterile seawater + seaweed
Before After
1 0.02 g
2 0.04 g
3 0.03 g
VSE + seaweed
Before After 
4 0.02 g
5 0.02 g
6 0.02 g
PES + seaweed
Before After 
7 0.03 g
8 0.05 g
9 0.04 g
Sterile seawater + bacteria + seaweed
Before After 
10 0.04 g
11 0.05 g

12 0.04 g

It was shown that almost all of the seaweed grow bud on it. 

DISCUSSION

. Low PES strength was also found to be more suitable for Gracilaria species in the previous study as reported by Ursi et al. (2008). This work indicated that growth promotion effects were seen using lower concentration of PES. Ferreira et al. (2006) reported that higher growth rate was observed for low strength media such as PES.However, G. changii is an indigenous seaweed species in Malaysia and adapted to tropical oliogotrophic water which contains lower nutrient concentration compared to temperate sea (Eppley, 1972; Phang, 2006). Dilution of PES media demonstrated to be ideal for achieving better growth rate for G. changii.

Teluk Pelanduk and Teluk Kemang

On 29 September 2015, class of Aquatic Flora Culture went for sampling of seagrass and seaweed at Teluk Pelanduk and Teluk Kemang.

The aims for this sampling procedure is :

• to gain knowledges about various kinds and species of seagrass and seaweed available there
• to gain experiences on how to do sampling of seagrass and seaweed
• to take seagrass sample as an adopted seagrass to be cultured

What kind of species have been reported there ?

Seaweed

Amphiroa fragilisima

What kind of studies have been done?

As these places are known as one of the place that have richness of seagrass and seaweed in Malaysia, there are many studies and researches have been done here.

For example :

Macrophytes diversity and distribution at Teluk Pelanduk and Teluk Kemang,Negeri Sembilan,Malaysia by Muta Harah et al.This study examined the diversity and distribution pattern of macrophytes by using transect and quadrat methods at two inter-tidal shores with distinct land form characteristics.

Practical 7 : Propagation of freshwater macrophytes

INTRODUCTION

We were provided  with 3 types of freshwater macrophytes. Every group had to choose only one type which is  4 replicates each.

METHOD

Before proceed to the procedure, agar must be transfered first to allow them harden in time.

•         Excise root and leaf

•         Wash with sterile dH²O + tween 20 (3mins)

•         Rinse with sterile dH²O (3mins)

•         Wash with 70%,50%,30%,10% chlorox (1 min each)

•         Rinse with sterile dH²O  (3 mins)

•         In laminar flow,wash explant with 70% ethanol (3 mins)

•         Rinse with sterile dH²O  (3 mins)

•         Plant explants into agar

•         Observe and record