Freshwater Zooplankton Sample Processing Protocols

​​​Many protocols can be applied, depending upon the study goals. For all protocols, the level of identification outlined above is applied (i.e., generally to species). A rare-species scan can be added to any protocol if a full-species list is required. In this case, the remaining sample is examined and all the species not included in the count are identified and enumerated within the Comments section of the bench sheet. Due to the added processing effort, rare-species scans do incur an added cost.

 

1.  Dorset protocol (pages 74–77). This protocol was developed by the Dorset Environmental Science Centre of the Ontario Ministry of the Environment, Conservation and Parks. It is the "default" protocol, unless the client has more specific project goals. The aim is to minimize counts of immature Copepoda and also prevent any single species from dominating the count by ensuring that no species accounts for more than 20% of the count. Every individual included in the count is also measured.

• Count and measure 15–35 of each type of nauplii (Cyclopoida and Calanoida). 

• Count and measure 15–35 of each type of copepodid (Cyclopoida and Calanoida).

• The remaining number of individuals, up to the minimum of 240, is divided among Cladocera and adult Copepoda. Count and measure 45–60 individuals for a dominant species in a sub-sample.

• Identify and enumerate any zebra mussel veligers, Chaoborus and Bythotrephes encountered during the count, and note in the Comments. No measurements need to be taken and the entire sample does not need to be scanned for these species.

• In the Comments section, the genera of the most common Rotifera encountered during the count are noted. A rotifer index between 0 and 5 (0 meaning none present to 5 meaning extreme abundance) is assigned for each sample. (Note that Rotifera counts are generally not included in any protocol and are a supplementary community indicator only.)

​​​

2. Simcoe Bythotrephes protocol. The Ontario Ministry of the Environment, Conservation and Parks Lake Simcoe Monitoring Branch has a specific protocol that targets the invasive species Bythotrephes cederstroemii (i.e., spiny water flea).

• Measure every Bythotrephes in the sample and record its attributes (sex, instar, stage of fecundity, clutch size, and tail spine morphology) in ZEBRA2.  

• Measure every Mysis and record its sex in ZEBRA2.

• All other larger taxa (e.g., Leptodora, mites, etc.) are enumerated in a separate Excel spreadsheet.

​​

The Ontario Ministry of the Environment, Conservation and Parks compiled their own length-weight regressions which are incorporated into their custom Python script. This applies to count protocols 1 and 2.

​​

3.  Bowen protocol (PDF). Another extensively used protocol is that developed by the Department of Fisheries and Oceans, Great Lakes Laboratory for Fisheries and Aquatic Sciences in Burlington, Ontario (GLLFAS). Unlike the above protocols designed by the MECP, only a subset of the count is measured, then the average weight is applied to the entire per-species count. 

• Remove Bythotrephes, Mysis, and Cercopagis before splitting the sample with the syringe method. If present, count them ALL but only measure adults and count eggs in 30/species. If a sample is very dense with Bythotrephes, these are removed to a separate gridded tray and only 20% are included in the count (30 measured with eggs also counted).

• Minimum count set at 400 (200 if only 1 species is present) OR a maximum of 20% of the starting working volume.

• Count is evenly divided between four taxonomic groups: Cladocera, mature Copepoda, immature Calanoida, and immature Cyclopoida. Veligers encountered during the count are also included.

• Measure 30 each for the zebra mussel veligers, Calanoida nauplii, Cyclopoida nauplii, Calanoida copepodids, and Cyclopoida copepodids, then 50 each for the species of Cladocera and the species of mature Copepoda.

• Record if the measured animals have eggs (if so, note the number of eggs/animal).

• For loose eggs in the sample, track a maximum of 100 eggs and identify as belonging to Cladocera (neonates included), Cyclopoida, or Calanoida.

​​

The GLLFAS compiled their own length-weight regressions (PDF, page 15) which are incorporated into their custom Python script.

​

4.  Cultus protocol. The Department of Fisheries and Oceans, Cultus Lake Salmon Research Laboratory in BC developed this protocol. Similar to the Bowen protocol, the count is divided between categories but in this case, the target groups are based upon the measurements outlined below, not taxonomic divisions. Also, every individual included in the count is measured.

• Minimum count set at 400, with four size class targets as outlined below:

  • 50 maximum Copepoda nauplii < 0.6 mm

  • 100 minimum zooplankton < 0.6 mm 

  • 150 minimum zooplankton >= 0.6 mm and < 1.2 mm

  • 100 minimum zooplankton >= 1.2 mm

• A whole-sample rare-species scan for larger taxa is also required.

 

The Cultus Lab required integration with their long-term zooplankton database. A custom Python script populates Excel spreadsheets that were tailored to the exact specifications of the Cultus Lab with the raw ZEBRA2 measurement and species data. Cultus also has their own set of species codes which were associated to ZEBRA2's species codes. Both sets of codes are shown in their Excel spreadsheets.

​

5.  Size class protocol. Identical to the above Cultus protocol, minus the elements specific to Cultus's database (i.e., Cultus species codes and database formatting). Therefore, data are sorted into the four size classes outlined under Cultus.

​

6.  MFFP/UQAM protocol (Ministère des Forêts, de la Faune et des Parcs in association with l'Université du Québec à Montréal, QC). Only a subset is measured, then the average weights are applied to the entire count/species.

• Count up to 200 individuals of the same species, or 

• Count to 1000 individuals as the stopping point, or 

• Count the entire sample if criteria 1 or 2 are not met.

• Only 10 individuals/species or immature Copepoda group are measured.

• A whole-sample rare-species scan for larger taxa is also required.

• In the Comments section, the genera of the most common Rotifera encountered during the count are listed.