Long-term cleaner fish presence affects growth of a coral reef fish

Clague et al. 2011

Electronic Supplementary Material

Detailed Methods

Cleaner removal

Following the initial cleaner removal in September 2000, juvenile bluestreak cleaner wrasse Labroides dimidiatus that had recruited onto the removal reefs were removed using hand-nets. All cleaner wrasse were measured [total length (TL), mm] before being relocated onto reefs >0.5 km from the original reef. Adults were classified by coloration, with adults having a black lateral stripe against a blue and yellow body, and juveniles having a lateral neon blue stripe against a black body (Randall et al., 1997). Over the past eight years, and on any one survey, removal reefs had between 0 and 5 adults (mean ± SE: 0.16 ± 0.04) and between 0 and 8 juveniles (0.44 ± 0.07) removed from September 2000 to October 2008 (Table S1). Control reefs were surveyed for cleaners, and had between 0 and 5 adults (2.15 ± 0.10), and 0 and 8 juveniles (0.85 ± 0.10) (Table S1).

Other cleaners, Periclimines or Urocaridella shrimp, were observed on reef 11 (removal) and 15 (control), respectively, but not removed; their densities were low and so their presence unlikely to have a significant impact on parasite load of client damselfish Pomacentrus moluccensis.

Client fish collection

Client damselfish Pomacentrus moluccensis were collected (n=10 per reef) by a scuba diver between 9-26 October 2008, using localized sprays of a clove oil solution (1:4:5 clove oil: 100% ethanol: seawater) and hand-nets. Individual fish were selected haphazardly. The density for P. moluccensis, adjusted for reef area, for removal reefs was 208.0 ± 29.4 (least square mean ± SE), and control reefs was 265.3 ± 25.7 (Waldie et al. in press). Fish were placed individually into click-seal bags to retain parasites and euthanized with an overdose of clove oil (2-3 squirts of the clove oil solution into bag). On the boat, bags were placed on ice during transport to the laboratory at Lizard Island Research Station, where they were immediately processed. The liquid in each bag was sieved through a 62µm sieve, and particles in the sieve washed into vials using 70% ethanol (EtOH). 80% EtOH was injected into the gut cavity with a syringe and 22-gauge needle to preserve any internal parasites for another study. Fish were placed in vials with the corresponding particulates and preserved in 70% EtOH. After 2, 5 and 10 days the ethanol was sieved again and the 70% EtOH replaced to ensure adequate fixation of internal parasites.

Client size estimation

Prior to estimating the TL of cleaner fish L. dimidiatus, client P. moluccensis, and conspecifics, estimating fish size was practiced underwater using models of fish. Outlines of fish 12-74 mm TL, painted on metal disks, were placed at varying distances on the reef. Sizes were estimated and these compared to the actual length of the model. Differences between estimated and actual length were calculated (mean ± SE: 1.4 ±1.1 mm).

Statistical analyses

To test for an effect of cleaner fish presence on P. moluccensis growth, a full linear mixed-effects (LME) model was employed, using the restricted maximum likelihood (REML) in JMP v8.0.1 (S.A.S. Institute Inc. 2009). The response variable was fish SL, fixed factors were collection site (Casuarina Beach and the Lagoon, two environments of interest at Lizard Island) and L. dimidiatus presence, the covariate was fish age, and reef was included as a random effect (representing a subsample of the larger population at each site). Data were log10transformed to satisfy the assumptions of the analysis. Two outliers were omitted (see Fig. 1), although this did not affect the outcome.

The total number of parasitic copepods (see Figure S1 for a photograph of a representative juvenile copepod) was summed across the 10 fish sampled per reef to reduce the number of zeros in the data, and fish standard length (SL) was averaged per reef. A generalized linear model (GLM) was used to test for difference between reefs with and without cleaner fish, with total numbers of copepods per reef as the response variable, treatment (i.e. cleaner presence) and collection site as fixed factors, and mean SLper reef as a covariate; a quasi-Poisson distribution with a loglink function was used to model these data.

A LME model with a REML procedure was performed to test for a difference between the estimated size (TL) of P. moluccensis that were cleaned and conspecifics within a 1 m radius that were not cleaned, using two fixed factors (cleaner presence and site), cleaner size as a covariate and reef as a random factor. REML was used to account for the unbalanced nature of the designs. These analyses were run in R v2.11.1. The assumptions of the statistical analyses were verified by examining the residuals and with quantile-quantiles plots.


Randall J.E., Allen G.R., & Steene R.C. (1997) Fishes of the Great Barrier Reef and Coral Sea. Honolulu, HI: University of Hawaii Press; 557 pp.

Waldie P., Blomberg S.P., Cheney K.L., Goldizen A.W., & Grutter A.S. (in press) Long-term effects of the cleaner fish Labroides dimidiatus on coral reef fish communities. Plos One