Standard Operating Procedures for Fish Larvae
Version 1.0
Contact Details
For Contractual and Administrative details:
Ms Mariann Fee
Chief Executive Officer
UoM Commercial Ltd
The University of Melbourne
442 Auburn Road
Hawthorn VIC 3122
Phone: +61 3 9810 3254
Fax:
+61 3 9810 3149
Email: mfee@unimelb.edu.au
For Technical Details:
Mr Wayne Koster
Arthur Rylah Institute for Environmental Research
Department of Environment and Primary Industries
Heidelberg VIC 3084
Phone: +61 3 94508766
Mobile: +61 400 558 261
Email: wayne.koster@depi.vic.gov.au
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CONTENTS
Introduction .......................................................................................................................................... 2
Objectives and hypotheses ..................................................................................................................... 2
Indicators .............................................................................................................................................. 2
Locations for monitoring ........................................................................................................................ 3
Timing and frequency of sampling .......................................................................................................... 3
Responsibilities – identifying key staff .................................................................................................... 3
Field program ................................................................................................................................................. 3
Laboratory requirements (if any)................................................................................................................... 3
Procedure for transferring knowledge to new team members ..................................................................... 3
Monitoring methods .............................................................................................................................. 4
Field methods ........................................................................................................................................ 4
Equipment ...................................................................................................................................................... 4
Sample placement within sites ...................................................................................................................... 4
Sampling protocol .......................................................................................................................................... 5
Laboratory methods ...................................................................................................................................... 5
Data analysis and reporting ........................................................................................................................... 6
Quality Assurance/Quality Control ................................................................................................................ 6
References............................................................................................................................................. 6
Attachment 1
LTIM Data Standard requirements ................................................................................ 8
Introduction
The Standard Operating Procedure (SOP) for Fish Larvae describes the purpose of the Fish Larvae
assessment, how the monitoring will be conducted, who is responsible for specific tasks and how the
collected data will be analysed and reported. The document is intended to be taken in the field during any
Fish Larvae surveys for the LTIM and should be updated throughout the life of the Long Term Intervention
Monitoring Program to reflect any agreed changes to method or procedure.
Objectives and hypotheses
This monitoring protocol for Fish Larvae addresses the following Basin scale evaluation questions:
Long-term (five year) questions:
 What did Commonwealth environmental water contribute to native fish populations?
 What did Commonwealth environmental water contribute to species diversity?
Short-term (one year) questions:
 What did Commonwealth environmental water contribute to fish community resilience?
 What did Commonwealth environmental water contribute to native fish survival?
 What did Commonwealth environmental water contribute to native fish reproduction?
The Area Specific evaluation questions to consider are:
Long-term (five year) questions:
 What did Commonwealth environmental water contribute to the recruitment of Golden Perch in the
adult population in the lower Goulburn River?
Short-term (one year) questions:
 What did Commonwealth environmental water contribute to Golden Perch spawning and in
particular what magnitude, timing and duration of flow is required to trigger spawning?
 What did Commonwealth environmental water contribute to the survival of Golden Perch larvae in
the lower Goulburn River?
We expect that spring-summer freshes will trigger spawning, particularly for Golden Perch. This may result in
a broader range of age classes in the population (currently the Golden Perch population comprises only
large, old fish) and an increase in abundance. Increased baseflows in late summer/early autumn might also
promote subsequent larval survival and recruitment into the adult Golden Perch population. A range of other
species might also be expected to benefit from spring-summer freshes and increased baseflows.
Indicators
Fish Larvae is a Category II monitoring indicator for the LTIM Program in the lower Goulburn River. It
requires annual surveys of larval fish and eggs at 3 sites in each nominated zone. The standard method
stipulates 6 larval surveys to be carried out each summer, but we feel this is insufficient to adequately
capture spawning events. Instead, we will undertake 10 surveys per season. The surveys will use a
combination of drift nets and light traps to catch fish eggs and larvae from each site. Fish collected will be
identified and counted to assess relative abundance and links to flow events.
This indicator will also be measured in the Edward-Wakool, Lachlan, Murrumbidgee and Gwydir Selected
Areas. The monitoring method has been specified by the M&E Advisor. The Fish River surveys will
complement this indicator because they will measure successful recruitment (young-of-year fish) following
any fish spawning. The Fish Movement component will also complement this indicator. In particular, it will
provide critical information on whether fish need to migrate to specific areas to spawn and under what
environmental conditions (e.g. freshes).
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Locations for monitoring
Monitoring will be conducted at 3 sites in Zone 2 (Shepparton to Murray junction), and at 1 site in Zone 1
(Goulburn Weir to Shepparton), noting that the Zone 1 monitoring will not be compliant with the standard
method (i.e. not being done at 3 sites).
Zone 2 was identified as a priority for monitoring because it is where we most expect to detect a response
(e.g. spawning) to flow events, as previous monitoring has shown that spawning activity by Golden Perch is
concentrated in this zone (Koster unpublished data). The monitoring sites are shown in Table 1. The sites
are subject to change depending on access.
Table 1: Proposed fish larval monitoring sites for Zones 1 and 2. Existing ARI larval
monitoring sites are shown.
Zone
Lat
Long
Existing ARI site
1
Site
Pyke Road
-36.427
145.357

2
Loch Garry
-36.242
145.287
2
McCoys Bridge
-36.177
145.123
2
Yambuna
-36.131
145.003

Timing and frequency of sampling
Larval sampling will generally occur weekly during the spawning season, with the specific timing to be
tailored to best measure the response of flow events in each year (e.g. before, during and after spring
freshes). At present flow events to trigger spawning are scheduled to occur between October and
December. This period coincides with planned environmental flow release and the peak spawning period for
Golden Perch. Sampling will be conducted every year at each site over the five year program.
Responsibilities – identifying key staff
Field program
Wayne Koster and David Dawson from the Arthur Rylah Institute will be responsible for overseeing the
planned monitoring, conducting the field surveys and any relevant training, and preparing safety plans. Other
experienced staff from ARI may also participate in the surveys at times where needed. Angus Webb from
Melbourne University will conduct data analysis. Wayne Koster and David Dawson will be responsible for
collating, checking and uploading collected data.
Laboratory requirements (if any)

Samples collected will be preserved in 90% ethanol and transported via car to ARI and sorted in the ARI
fish laboratory under a dissecting microscope, and identified using a guide (Serafini and Humphries
2004). All samples will be stored in ethanol in the ARI fish laboratory for the duration of the project.
Procedure for transferring knowledge to new team members
The discipline lead (Wayne Koster) will be responsible for ensuring new team members have a sound
understanding of the program and are able to competently undertake required tasks. The discipline lead will
undertake the following steps to facilitate this process
1.
2.
3.
4.
5.
Discuss the overall program to the new staff member and introduce to team members.
Outline and document the roles and responsibilities of the new staff member
Explain and provide access to relevant program documents
Explain and discuss the standard operating procedure
Explain and discuss the project risk assessment and the required safety measures
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6. Demonstrate sampling methods to staff in the field and supervise staff undertaking these methods
until satisfactory competency is demonstrated.
7. Explain and demonstrate data collation, analysis, uploading procedures and assist staff in
performing these tasks as required
No formal accreditation is required to undertake netting or light trap surveys. However, any new
inexperienced staff involved in surveys will only work with other staff who have at least 2-3 years’
experience. Staff must hold a valid Victorian drivers licences and boating licence. A Coxswains certificate will
also be required from mid-2015. Staff must also have completed accredited 4WD training and First Aid
(Level 2) training. David Dawson will be the senior field technical officer within the field team. David will liaise
regularly with Wayne Koster and will train any new staff required throughout the duration of the project.
Wayne Koster will convene (face to face) regular debriefs with the field team after field trips to identify any
issues of concern. Scheduled debriefings will occur after each field trip for the duration of the project. Any
significant matters will be dealt with immediately rather than waiting for a scheduled debrief.
Monitoring methods
Field methods
Equipment
Small boat
Fine mesh larval nets, floats and ropes
Light traps
GPS
Datasheets and/or field computer
Sample containers
Turbidity meter
Velocity meter
Sample placement within sites

Ten light traps should be set in randomly selected slackwater habitats within each site. Select the set of
10 random PS waypoints (see Randomisation approach below), then find the closest slackwater to that
waypoint. If no slackwater is available within 20 m either side of the waypoint select another random
waypoint.
 Light traps will be used for larval assemblage composition and potentially for relative abundance
comparisons/contrasts among areas. Their efficacy is heavily dependent on turbidity, so any
comparisons of relative abundance among areas will be dependent on the inter-area differences in
turbidity levels (which will be measured on each sampling occasion).
 Larval density is measured using stationary drift nets for higher current areas.
 Three Drift nets per site (total of nine per zone, per sampling event) should be positioned in water with a
moderate velocity, preferably where the discharge is concentrated through a narrow section of the river
(a funnel effect). Ideally, drift nets should not be closer than 100 m to each other.
Randomisation approach
 Each 800m site is subdivided by fixed transects spaced 50 m apart. Points of intersection between the
transects and the river bank define the sampling grid.
 The sample design specified defines two key sampling locations: electrofishing (EF) units (16 in total),
and passive-gear sample (PS) waypoints (34 in total) (see Figure 1). The passive sampling waypoints
will be used for setting the position of larval light traps.
 To establish the PS grid, providers should save each PS waypoint in a GPS. A GPS can be used to
locate each PS waypoint in subsequent visits throughout the LTIM.
 A random number generator should be used to randomly select a subset of 10 PS waypoints from the
total of 34. A waypoint encompasses a total of 40 m of bank (20 m either side of specific waypoint), so
providers should endeavour to find the point on the bank as close to the exact waypoint as possible. The
purpose of this system is to ensure sampling is random with respect to the environment, so providers
should not simply ignore a waypoint if setting of a net/trap is not as leisurely as they would like.
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Zone
River channel
Site 1
Site 2
Site 3
50 m
PS2
Site
Direction
of flow
PS4
EF
Unit
1
PS1
PS6
EF
Unit
2
PS3
PS8
EF
Unit
4
EF
Unit
3
PS5
PS10
PS7
PS12
EF
Unit
5
PS9
PS14
EF
Unit
6
PS11
PS16
EF
Unit
7
PS13
PS18
EF
Unit
8
PS15
PS20
EF
Unit
9
PS17
PS22
EF
Unit
10
PS19
PS24
EF
Unit
11
PS21
PS26
EF
Unit
12
PS23
PS28
EF
Unit
13
PS25
PS30
EF
Unit
14
PS27
PS32
EF
Unit
15
PS29
PS34
EF
Unit
16
PS31
PS33
Transects
Individual passive-gear sample (PS) waypoint
20 m
20 m
Acceptable passive gear positioning zone
(20 m either side of transect waypoint)
Figure 1: Diagram of hierarchical sample design illustrating zones, sites and sample
locations.
Sampling protocol
Light traps





Modified quatrefoil light traps should be used. Mesh should be fitted around the light traps to eliminate
larger fish from entering the trap, and eating the sample (3 mm knot-to-knot).
The ten light traps set within each of the three sites should be set in the afternoon and retrieved the
following morning.
Set and retrieval times should be recorded, so that relative abundance can be expressed as catch-perunit-effort (CPUE).
Each light trap should be ‘baited’ with a yellow Cyalume 12 h light stick (or equivalent manufacturer,
but yellow in colour).
Turbidity will be measured at each light trap using a TPS meter.
Drift nets


Drift nets should be constructed of 500 μm mesh, have an opening diameter of 50 cm, tapering over 1.5
m to an opening of 9 cm, to which a reducing bottle should be fitted.
Volume through the net should be estimated so that larval abundances in drift nets can be expressed as
a density: number of individuals per m3. The water volume passed through the net is determined as
follows: number of revolutions (T2-T1) x 0.3 (impeller pitch) x net opening area (m 2) x 1000 = water
volume
Laboratory methods

Samples collected will be preserved in 90% ethanol and transported via car to ARI and sorted in the ARI
fish laboratory under a dissecting microscope, and identified using a guide (Serafini and Humphries
2004). All samples will be stored in ethanol in the ARI fish laboratory for the duration of the project.
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Data analysis and reporting
Turbidity

Turbidity measures should be reported as mean turbidity per site per sampling event and matched to
Light trap abundance data.
Relative abundance


Light-trap abundances should be expressed as ‘catch-per-unit-effort’ (CPUE), where the units are
number of individuals for each species per trap per hour of deployment.
Drift net abundances will be expressed as densities; number of individuals for each species per cubic
metre of water filtered.
Community data



CPUE data should be reported at the level of the site (species by site abundance matrices).
Abundance data is reported for each species as the mean CPUE for the site.
Data should be provided separately for each sampling method.
A report will be produced that will include an assessment of relationships between the timing and abundance
of eggs/larvae and flows. The report will focus on the magnitude, timing and duration of flow required to
trigger spawning by Golden Perch because we expect spawning of this species be associated with high flow
events. Where relevant the report will include modifications or revisions to existing environmental water
releases in the lower Goulburn River to enhance fish spawning.
Detailed statistical analysis for the Selected Area evaluation will be conducted by or under the direction of Dr
Angus Webb at the University of Melbourne.
Quality Assurance/Quality Control







The boat and motor will also be serviced annually by Barry Lawrence Marine. All service certificates will
be stored in a filing cabinet in the maintenance department at ARI.
Larval nets and light traps will be checked for holes or cracks prior to every field trip, and also upon
completion of sampling at each site. Any net with a hole will be repaired or replaced immediately using a
repair kit that is taken on site.
The monitoring will be conducted under an existing Victorian Flora and Fauna Guarantee Permit
(expires Aug 2014 but is renewed annually) and Fisheries Victoria Research Permit (expires Aug 2014
but is renewed annually) and ethics permit 13-10 (Arthur Rylah Institute Animal Ethics Committee)
(expiry 31/12/2017 but will be renewed prior to expiry). Permits will be carried while sampling. Prior
notification to Fisheries for any sampling will be made as per permit conditions.
All nets and traps will be clearly marked with the permit holders name and permit number. Permit reports
will be lodged annually with the relevant body as per permit conditions.
All data provided for this indicator must conform to the data structure defined in the LTIM Data Standard
(Brooks and Wealands 2013) and included in Attachment 1.1. The data standard provides a means of
collating consistent data that can be managed within the LTIM Monitoring Data Management System
(MDMS).
Data will be entered into an excel spreadsheet that follows the prescribed template provided in
Attachment 1.1. Each cell of data will be cross-checked visually against data sheets for accuracy by
David Dawson. Data sheets will be stored in a filing cabinet in David Dawson’s office at ARI. All data
sheets will be photocopied and stored in a separate filing cabinet in Wayne Koster’s office at ARI.
Data will be stored electronically ARI on a computer network that is backed-up daily. All data is backed
up to an external site. Data will be sent by David Dawson via email to Melbourne University to be loaded
onto a central database.
References
Brooks, S. and Wealands, S. (2013) LTIM Data Standard v0.2. Draft Report prepared for the Commonwealth
Environmental Water Office by The Murray-Darling Freshwater Research Centre, MDFRC Publication
29.3/2013 November, 29pp.
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Serafini, L.G. and Humphries, P. (2004) Preliminary guide to the identification of larvae of fish, with a
bibliography, from the Murray-Darling Basin. Cooperative Research Centre for Freshwater Ecology
Identification and Ecology Guide No. 48. Cooperative Research Centre for Freshwater Ecology, MurrayDarling Freshwater Research Centre, Albury and Monash University, Clayton, 55 pp.
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Attachment 1
LTIM Data Standard requirements
Attachment 1.1
to the CEWO.
Standard Data format for larval fish survey results to be submitted
Variable
Description
Type
Req
assessmentUnitId
The site, which may be a length of stream
of an area of wetland(s) that meets the
criteria defined in the standard method
string
Y
dateStart
Start date (inclusive) that these measures
were observed
dateTime
Y
dateEnd
End date (exclusive) that these measures
were observed
dateTime
Y
speciesName
Latin name for species of fish
string
Y
lightTrapCatch
Number of individual per unit effort
(number of individuals per trap per hour of
deployment)
number
(8 decimals)
Y
driftNetCatch
Number of individual per unit effort
(number of individuals per cubic metre of
water filtered)
number
(8 decimals)
N
trawlNetCatch
Number of individual per unit effort
(number of individuals per cubic metre of
water filtered)
number
(8 decimals)
N
sampleType
Sampling method
category
Y
LightTrap
DriftNet
Trawl
turbidity
Turbidity of water at the time and site of
sampling. NTU
integer
N
[0,+]
qualityTurbidity
Quality code as per Water Quality standard
method
integer
N
[1,5]
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