Only a few filamentous bacteria (Beggiatoa and Flexibacter) are able to move freely. This occurs by means of ‘gliding movement’ resembling the manner in which a worm propels itself.
If it is thought that a certain filament appears to move, then it should be verified that there is no liquid flow on the slide owing to evaporation of the water along the edges.
The absence or presence of branching is an important characteristic for identifying filamentous bacteria in activated sludge. Real and false branches are distinguished.
Actinomycetes and fungi show real branching. The side shoot actually sprouts from the main branch. With real branching, the cell in fact grows in two directions.
Sheath forming bacteria, principally Sphaerotilus natans and Type 1701, form false branches. These arise because ‘swarming cells’ attach themselves to the sheath around the filament and subsequently, through cell division, develop into side shoots (including a sheath).
Swarming cells are cells which have split away from the free end of the filament and temporarily, i.e. as long as they have not yet become attached, are present in the sludge as free living cells. A side branch can also be formed at a spot where damage to the sheath has occurred.
In the case of false branching, a very small space is often visible between the cells in the main branch and those in the side shoot. This space is lacking in the case of real branching. False branches are always ‘V’ shaped, while real branches often stand ‘dead straight’ on the main branch (but not in the case of the actinomycete Skermania piniformis). Actinomycetes always form numerous real branches. A number of branched filaments gathered together clearly resemble a bunch of sticks. Fungi are easily distinguishable from the sheath forming bacteria and the actinomycetes on account of their much more robust filaments.
Attachment of single celled bacteria to a filament (= attached growth) is sometimes mistaken for branching.
Occasionally in the sludge filaments are observed which are attached to each other at their bases. This is known as a rosette of filaments. These bacteria secrete substances with which they attach themselves. Thiothrix and Leucothrix strains mainly form rosettes.
Three groups of filament shapes are distinguished:
The word straight has been placed between inverted commas, as only Haliscomenobacter hydrossis forms really dead straight filaments. Straight examples are indeed somewhat bent, especially where relatively long filaments are concerned.
During microscopic investigation, a distinction should be made between (1) straight to slightly bent and (2) bent to coiled filaments.
The surface (= the outside wall) of filamentous micro-organisms is usually ‘clean’. Other cells or particles of floc are sometimes attached to the surface, however, and partially cover it. This is known as ‘attached growth’. The filaments of bacteria which have a sheath around the cells are often surrounded by attached growth.
When identifying filaments, a distinction must be made between (1) little or no attached growth and (2) much attached growth.
Filamentous micro-organisms are divided into three groups on the basis of their diameters:
· diameter < 1 mm
· diameter 1-2.5 mm
· diameter > 2.5 µm
For some filaments (Type 021N and Thiothrix strains), the cell diameter sometimes gradually decreases towards the tip of the filament.
Transverse walls or septa are the walls between consecutive cells of a filament. Septa are not always clearly visible with a light microscope. When identifying filamentous organisms, a distinction should be made between (1) clearly visible and (2) poorly visible/invisible. Where filaments possess much attached growth, attention should be paid to their clean extremities for assessing this feature.
The following cell shapes can be distinguished in free living bacterial cells:
· spherical or coccus
· rod shaped
· spiral shaped
Filamentous micro-organisms comprise coccus or rod shaped cells or other cell shapes derived from these shapes. This means that the cells in a filament are square or rectangular, if there are no constrictions of the communal outside wall at the position of the transverse wall.
The cell type can actually only be determined if the septa are clearly visible. The length of the cells in a given filament is often highly variable. The cell length of some filamentous species (Type 021N, S. natans and Thiothrix) can increase considerably towards the tip of the filament.
The cells of certain filamentous species are surrounded by a transparent cover known as a sheath, which is not often visible with a light microscope. The sheaths of some species are indeed visible following Gram staining. The sheath is sometimes more easily visible if the sludge sample is stored for a few days in a refrigerator. A lot of attached growth often indicates that a sheath is present.
In addition, it should be noted that empty cells in a filament are sometimes mistaken for sheaths. The transparent empty cells are generally very short. This means that long, transparent covers must be visible in or on the extremities of the filaments before it can be definitely concluded that a sheath is present. As this characteristic is often not properly established, it is of subsidiary importance for the identification of filamentous bacteria in activated sludge.
Granules are globules of storage material in the cells. Three types of granules are distinguished with microscopic investigation:
· polyphosphate granules. Without Neisser staining these granules are poorly or not visible at all;
· sulphur granules. These globules are of a dark shade when still small. On the other hand, larger examples are strongly refractive and are consequently clearly visible;
storage materials. This usually involves poly-b-hydroxybutyrate
(PHB). PHB granules are hardly ever observed in filamentous
domestic treatment plants. Like the sulphur granules, the PHB granules
For information on carrying out these staining methods, follow this link