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Liz Milan, our Chief Trainer, has been known for many years as “The Bug Lady”. In her native New Zealand, Liz was the “go to gal” for anyone who suspected that there might be a little something off in their water or sewage treatment plant. Liz, armed with her trusty microscope, takes great glee in identifying little “critters” as they float by, invisible to the naked eye.
In our bi-monthly training newsletter, Liz provides us with her take on some of the bugs your treatment plant may encounter. This gallery is where we can revisit them. We’ve also included some of Liz’s microscope videos (some from back before she joined S&B) to help give you a glimpse as to what might be going on in your water….
Drepanomonas: A Shape-Shifter
This member of the Microthoracidae family can be quite tricky to identify. When you see these little jellybean shaped guys in your mixed liquor sample, take a closer look at them as they flit around the flocs. If you can see a very flat (thin like an oval button) shape when viewing from the ends, look for the side ridges and head cilia. This helps give away their identity. You can see in the 100X, 400X and 1000X photos below how difficult it would be to tell them apart from some other small free swimming protozoa you might find. They swim in an erratic, rotating way, making their shape look like it is constantly changing. Trying to get a decent side view to identify them can be tough!
My experience with these little guys is they will often be associated with high mixed liquor solids e.g. mixed liquor from MBR AS plants. Perhaps their thin shape helps them move between the thick flocs more easily! They were also typically found when anoxic conditions prevailed in significant zones of the plant. I have traditionally found them in activated sludge systems receiving dairy factory waste.
The Handsome Mr. Rotifer
Rotifers feed by extending and rotating the coronas of cilia (hairlike fibres) on their heads to create a vortex. The vortex of liquid and suspended particles passes through a chewing pharynx, called the mastax, into their gut …they are filter feeders. A few rotifers in your mixed liquor are great, they help to clean up (by eating them!) the non-flocced suspended particles that contribute to high final effluent suspended solids and poor colour or clarity.
If you see a lot of, or an increase of rotifers in your mixed liquor it can signal a few things are getting out of whack, especially if you have simultaneously seen an increase in staked ciliate numbers…remember good floccing usually equals good settleability and clean effluent.
The problem could be:
- Long sludge age. Rotifers are multicellular animals and take longer to grow and reproduce than bacteria or protozoa (lower lifeforms). Older sludge tends to be made up of hungry, stressed bugs who are less likely to have surplus food to make EPS (floc glue), hence more unattached cells in the inter-floc liquid. You will probably be seeing pin-floc coming over your clarifier weir in the effluent.
- Fluctuating food load, oxygen levels or nutrient deficiencies, causing the floc to grow then dissociate again
You’ve been slugged by something (chemical?) in the influent which has killed off some bacteria, allowing the floc to disintegrate or dissociate (toxin e.g. herbicide or emulsifier e.g. degreaser)
What should you do?
Investigate population changes and build a history of cause and effect for your plant.
Pinkie Tusacadaro!!! (Blepharisma)
This colourful little creature may appear from time to time in activated sludge mixed liquor, typically from extended aeration AS systems with reasonably high mixed liquor suspended solids (3000 mg/L plus).
Blepharisma is a unicellular free swimming ciliate protozoa,
typically 150 – 300 µm long.
It can be distinguished by its pale to bright pink colour. This pink colour is caused by a pigment called Blepharismin, a light sensitive pigment located just under cell wall (skin) and gives Blepharisma its name. This pigment can be quite toxic to other protozoa and rotifers so it helps protect the Blepharisma from being eaten !!
Blepharisma is quite a predator itself, eating bacteria, flagellate algae, small rotifers , other ciliate protozoa and members of its own species (cannibalism). Blepharisma is photophobic (light hating). When exposed to light or sometimes if stressed, specimens lose their pink colour and “bleach out” into colourless organisms. When in darker conditions (for example higher mixed liquor solids), the pink colour intensifies.
comparative population numbers can give the operator valuable insight into prevailing conditions in their system.
Vorticella provide a valuable service in two ways. Firstly, they help “clean up” any non-flocculated bacteria, preventing them from staying in suspension causing cloudy effluent after clarification. Secondly, their comparative population numbers can give the operator valuable insight into prevailing conditions in their system.
A big increase in Vorticella may indicate that the floc is getting too hungry (not enough EPS (extra cellular polymeric substances) to glue the floc together), or that a toxic shock has hit the plant resulting in deflocculation with an increase in free bacteria, or that the sludge age is getting a little too high and the flocs are pin floccing. A big increase in Vorticella, particularly if accompanied by an increase in Rotifers (also filter feeders) should make you suspicious and investigate.
Vorticella reproduce by growing a new daughter head out of the side of the parent head. It starts off looking like a large pimple and develops into a head nearly the size of the original. Once mature, it detaches and swims off in a form we call a telotroch, until it finds somewhere suitable to establish itself and grow its own new stalk. In the picture below at 1000X magnification (1 scale division = 1 µm) you can see the mature telotroch on the left of the parent head, nearly ready to detach and establish independently.
Check that out in one of Liz’s microscope videos (this one is narrated) below:
Vorticella can be identified by the fact they grow as a single colony (single head), unless they are reproducing. You can see the muscle fibre inside the stalk allowing the Vorticella to move it’s head away from trouble!
Rebellious Worm
This cheeky rascal is actually a rarely encountered (in activated sludge) oligochaete worm called Dero. Sometimes they are also known as Microfex Worms.
Despite looking like he was giving me the finger, this is actually his tail end! These amazing multicellular animals have a hand-like structure on their tail, with finger-like projections (gills). These are covered in tiny hairlike cilia to create a current and promote oxygen uptake in low dissolved oxygen conditions…makes a change from breathing through your head!
They can be an indication of highly polluted waters (for example in the sludge or slime from the waveband of a lagoon system). They are voracious feeders on detritus and decaying matter, making them useful for cleaning up “rubbish” e.g. plant material out of your system. Some aquarium keepers deliberately grow Dero to use as a food supply for their fish and other aquatic pets like frogs or salamanders. This specimen was seen in the mixed liquor of an STP which was returning supernatant from a sludge lagoon.
Opercularia! (Letting it all hang out!)
Here is a spectacular member of that category of protozoa.
You will notice this guy grows in a colony (multiple branches and heads), as opposed to the Vorticella, which is seen as a single head.
There are two other significant differences:
Firstly, the Opercularia stalk does not contain a muscle fibre (no myoneme), so the stalk is not able to contract; Secondly, the ring of cilia that the Opercularia rotates to draw in food is on an extended “process” which extends out of its head, rather than around the edge of the head like the Vorticella cilia has. It is likely that it is this extension that gives the Opercularia its name, from Operculum, a structure resembling a lid or a small door that opens and closes.
Opercularia may be found in systems with a medium to older sludge age, medium to higher food to mass loads and can tolerate periods of comparatively lower dissolved oxygen levels (perhaps due to low RAS rate and prolonged sludge retention in the secondary clarifier).
A truly spectacular microorganism to see through the microscope!
Suctoria – Hidden Assassin!
Suctorians are one of my favourite types of protozoans that might be found in activated sludge samples.
They are generally associated with biomass populations having lots of free swimming ciliate protozoa present. They lurk in wait until some unsuspecting free swimmer brushes against the end of their protruding tentacles (spikes). These spikes are hollow, the swimmer gets stuck to the end and the suctoria shoots little pellets of bug “curare” (poison) down the tube into the swimmer. This not only paralyses the swimmer but turns its insides into gel, which the suctoria then sucks up its tentacle like using a straw. The empty husk (cell wall) of the victim is then released.
Suctorians Tokophyra (on top) and Podophyra (on the bottom) both at 400X magnification. See the algae (Scenedesmus species) in the photos. The plant was returning supernatant from a lagoon the time.
Euplotes moebiusi – Santa!
This fascinating member of the group of ciliate protozoa known as spirotrichs was seen in the activated sludge of a WWTP receiving dairy trade waste. This group of ciliates is differentiated by having a flattened body shape, a large and obvious oral groove (running nearly the length of the body), and hardened cilia, called cirri. The Euplotes uses these cirri to crawl over the surface of flocs, swimming and to capture food. E. moebiusi is quite rare compared to the more frequently seen E. patella. Euplotes feed on aggregated bacteria, also on microalgae and some small flagellates.
In my experience they are found in systems with large dense flocs, which provide a good base for them to crawl over. I have typically seen them appear following a “bloom” of the smaller crawling ciliate Aspidisca. The E. moebiusi appeared in samples from this site when the operator began to return supernatant from a final lagoon, back to the head of the activated sludge plant. I guess algae were on the menu!
100x magnification 1 scale div = 10 µm
400x magnification 1 scale div = 2.5 µm
Spathidium spathula - Shovelhead (no not a HD) !
Seeing this predatory unusual shovel headed ciliate in an activated sludge sample from an extended aeration STP certainly got my motor running! (I only ever saw them in this single sample, out of all the hundreds of mixed liquors I have examined).
Spathidium can be described as voracious carnivores of other free-swimming ciliate microorganisms. They are special in that they can reproduce either asexually (by splitting themselves into two), for many generations, at least 1000 generations have been observed of this “solitary behaviour”; or sexually through conjugation with another Spathidium (which would have to be more fun!!).
It has been noted that Spathidium is more likely to engage in conjugation if living conditions are becoming less favourable for it.(No comment!)
Additionally they have the freakish ability to fully regenerate themselves from small fragments of their heads or tail ends.
Their significance in an activated sludge population is really only that there must be significant amounts of other free-swimming ciliate protozoa present, to be preyed upon by the Spathidium. The plant in question was running a 18 – 22 day sludge age and was extremely alkalinity limited (final effluent alkalinity < 50 mg/L as CaCO3 frequently). The operators were playing with the aeration around this period (in the middle of winter) and trying to restrict nitrification (and hence DO demand and alkalinity consumption) by reducing DO to levels in the range 0.5 – 1.0 ppm.
Don’t try this at home folks (at that time they had no nitrogen limits in their discharge consent).
Aspidisca: Mr. Hoover!
Aspidisca is one of the most commonly observed protozoa in activated sludge populations, and certainly the most common crawling ciliate seen. Aspidisca looks superficially similar to the much larger Euplotes. Aspidisca species range from approx. 25 – 45 µm in diameter, while Euplotes is typically 50-100 µm in diameter.
You can see the size comparison in the photo (100 x magnification, 1 scale division = 10 µm) at left.
A variety of Aspidisca species may be seen. These can be differentiated by looking at their backs. They range from the smooth backed Aspidisca lynceus; to Aspidisca turrita, who has a single large horn (which looks like a rose thorn) on his back; to the most commonly seen Aspidisca costata, distinguished by the multiple ridges on his back.
You can see these in the 400x magnification side view:
And back view:
I scale division = 2.5 µm.
Aspidisca are a crawling ciliate who use their clusters of hardened cilia, called cirri, to crawl over the surface of flocs, swimming and to brush bacteria (their food) off the surfaces they are travelling over.
You will see they have a little beak-like mouth that they stuff bacteria into! Aspidisca in decent numbers are a very positive sign in an activated sludge sample for two reasons.
Firstly, the presence of Aspidisca indicates that the surfaces of the flocs (aggregated bacterial biomass) are getting a good “hoovering”, exposing healthy, viable bacteria to consume the surrounding nutrients. Secondly, in my experience, when I see lots of healthy Aspidisca, the plant is usually nitrifying really well (excellent biological oxidation of ammonia).
Want to see some “live” action from Aspidisca? You’ll find a video below from Liz’s “Bug Lady” days in New Zealand.
Spirochaetes (a.k.a. Spirochetes, both are correct) and Spirillum – Twisted sisters
The term Spirochaete came into use in the late 19th century: from spiro- ‘in a spiral’ + Greek khaitē ‘long hair’. That reflects these squiggling little guys pretty accurately. Sometimes they can be confused with another wavy looking organism, the Spirillum.
These two groups of organisms reflect the saying same, same, but different… really well!!
The spirochaetes tend to be a little bit skinnier, often they are longer than their Spirillum counterparts and it is fairly common to see one end of the “strand” cross-over “itself”. They are very like a writhing snake in the way they move. They can be almost impossible to see at 100x magnification, you will just see a sort of “shimmer”, particularly along the edges of flocs. They often are visible at 400X & 1000x, like in the two photos above. You may have to watch a “field of view” for a while without moving around the slide, and then all of a sudden you can see them!
Spirillum are more of a rigid spiral (rather than wriggly curves) and they may seem to shimmer, teeter or “vibrate” as they move around the flocs. You are more likely to see Spirillum appear to be trying to “drill in” to the edge of a floc cluster, than you would to see a spirochete show similar behaviour.
As a diagnostic tool for the operator, either of these organisms is useful as a signal of low dissolved oxygen or septic conditions.
A sudden appearance or increase in of spirochetes or Spirillum in your mixed liquor may be an indication of:
Spirillum are defined as micro-aerophiles, which means they are favoured by low dissolved oxygen but must have some dissolved oxygen present to survive (typically 1 – 9 % DO preferred). They cannot survive without a little free oxygen (and cannot denitrify nitrate).
Spirochetes may be strictly anaerobic (conditions with no oxygen of any kind available) or facultatively anaerobic. They really like septic conditions.
Our experience has been as the population transitions from Spirillum to spirochetes, the DO is getting lower, the spirochetes are liking completely anaerobic conditions which the spirillum cannot handle. This makes it quite good if you are trying to get on top of an aeration issue, because if you can see the spirochete population being replaced by Spirillum, you know things are going back up in the right direction.