H . Zhou J. Exp. Mar. Biol. Ecol. 256 2001 99 –121
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organic-free sediment with different levels of added leaf litter were inserted into the mangrove sediment and retrieved at different times to determine rates of meiofaunal
colonization at varying levels of leaf litter enrichment. Hypotheses to be tested were:
• H Meiofaunal colonization of sediment within the mangrove study area is unaffected
by the presence of decaying leaves; •
H Meiofaunal colonization and community structure within the mangrove study area is unaffected by the quantity of decaying organic matter in the sediment;
• H All meiofaunal taxa within the mangrove study area respond in the same way to
the presence of decaying organic matter in the sediment.
2. Materials and methods
2.1. Study site The Ting Kok mangrove 228289N, 1148139E, with an area of 8.8 hectares, is located
in the New Territories of Hong Kong Fig. 1. Although it is under the influence of some
Fig. 1. A map of Hong Kong showing the location of the Ting Kok mangrove.
102 H
. Zhou J. Exp. Mar. Biol. Ecol. 256 2001 99 –121 Table 1
Environmental variables measured within the study area of the Ting Kok mangrove forest in 1999 Annual average range
Surface sediment temperature 8C 24.8 14.0–34.5
a
Salinity ‰ 26.2 7.0–34.0
a
pH 6.8 6.5–7.2
a
Dissolved oxygen mg l 3.9 0.9–6.1
Total organic matter dry wt. 3.2 2.7–3.4
Mean6S.D. n 5 5 Silt clay ,63 mm, dry wt.
18.761.6 Sorting coefficient
w 3.260.1
Md w
1.260.1
a
Measured from surface pore water.
streams and surface pore water salinity is occasionally diluted to as low as 7.0‰ by monsoon rainfall during summer, salinity is generally oceanic and the forest bottom is of
sandy-mud silt clay , 20 with a standing organic matter content of 3.2 Table 1. Unlike an estuarine soft bottom muddy mangal in which litter input is from domestic
allochthonous as well as autochthonous sources Lee, 1988, the Ting Kok mangal receives the bulk of its litter from autochthonous sources, mainly leaves and roots. Ting
Kok is dominated by Kandelia candel L. Druce, which though occurring only within the Indo-Malaysian region Duke, 1992, is widely distributed along the coast of
Southern China and is the most dominant pioneer mangrove species in Hong Kong Morton and Morton, 1983. In Hong Kong, tides are mixed semi-diurnal with an annual
average tidal range of 1 m. The experiment was conducted in the eastern part of the mangrove around mid-water, where the substratum appeared more uniform without
patches of cobbles and spatial variations in the strength and frequency of tidal inundation upon the mangrove were relatively small. This site was chosen for the
experiment also because some preliminary work has been done in terms of meiofaunal composition and their spatial distribution unpublished data and this basic knowledge
will help to interpret the pattern observed in the present study.
2.2. Experimental sediment and leaf litter preparation Large volumes of surface sediment 8 cm in depth were collected near the study area.
After collection, they were combusted in a laboratory muffle furnace at 5008C for 6 h in order to obtain azoic and organic-free sediment for use in the colonization experiment.
Ready-to-fall, senescent Kandelia candel leaves were picked at the study site. After air-drying for 2 weeks, the leaves were powdered into | 0.7 mm grains using an electric
grinder.
2.3. Estimation of organic content Total organic matter TOM of natural mangrove sediment in the experimental area
H . Zhou J. Exp. Mar. Biol. Ecol. 256 2001 99 –121
103
was determined by incinerating five replicate dried 608C for 24 h sediment samples of known weight | 10 g at 5008C for 6 h and calculating the percentage loss of dry
weight Buchanan, 1984. TOM of the prepared leaf detritus was estimated in the same way. These estimates acted as a standard for calculating leaf litter additions to the
experimental sediment.
2.4. Experimental design A two-factor experimental design was adopted. Experimental treatments were
represented by different additions of leaf litter to tubes of azoic sediment, corresponding to 0 3 , 0.5 3 , 1 3 and 2 3 the TOM of the mangrove sediment and identified as C, L,
M and H, respectively. Meiofaunal colonization rates at different stages of leaf litter decomposition were determined by sampling over the time intervals of 1, 10, 30 and 60
days post-placement. Each treatment was replicated four times for each sampling date so that the overall design was 4 3 4 3 4, i.e. four replicates with each combination of four
levels of treatment and four levels of exposure time.
Appropriate amounts of leaf litter corresponding to different doses were homogenized with the azoic and organic-free sediment by stirring using a glass rod. The tubular
barrels of 50 cc medical syringes 2.8 cm in diameter and 8.0 cm in depth were used as experimental tubes, each with two meshed 2 mm rectangular windows 2 cm 3 3 cm
on opposite sides Fig. 2. Tubes filled with azoic experimental sediment, which had been enriched to varying degrees of TOM with leaf litter, were embedded in the natural
sediment of the study area. This design enabled the experimental sediment to exchange
Fig. 2. Diagram showing an experimental tube in the mangrove substratum.
104 H
. Zhou J. Exp. Mar. Biol. Ecol. 256 2001 99 –121
water and, thus, salts and gases with the surroundings and allowed the meiofauna to settle in the experimental sediment either from the overlying seawater or by migration
from the surrounding substratum. A total of 64 tubes filled with experimental sediment were labeled and placed
2
haphazardly in an area of | 8 m around the mid-tide level 1 1.5 m C.D. in the Ting Kok mangrove during a low tide on 12 July 1999. Tubes were inserted into the sediment
immediately after a similar size core had been removed so that disturbance to the surrounding meiofaunal community during placement was reduced to a minimum. Four
replicate tubes of each treatment were retrieved, again haphazardly, on days 1, 10, 30 and 60 post-placement, respectively. After retrieval, each tube was put into a plastic bag
and sealed. On each sampling date, four replicate field control cores were also taken haphazardly from within the experimental area to 8 cm sediment depth with a 50 cc
syringe corer to provide baseline information on the resident meiofaunal community.
2.5. Sample processing in the laboratory Upon arrival in the laboratory, each meiofaunal core was fixed immediately with 5
seawater buffered formalin. Faunal extraction procedures followed Somerfield and Warwick 1996 and Warwick et al. 1998. This was a combination of decanting
through 1000 and 63 mm sieves with tap water 10 times, flotation in diluted Ludox-TM 50 at a specific gravity of 1.15 and centrifugation three times at 3000 rpm
for 3 min each. Major meiofaunal taxa were identified and enumerated under a dissecting microscope. Nematodes were transferred to a glycerine–ethanol mixture pure
glycerine 70 ethanol, 1:9 in v v which was evaporated to pure glycerine, and mounted onto slides for further identification to putative species Gerlach and Riemann,
1973; Platt and Warwick, 1983, 1988; Warwick et al., 1998 as well as assignment to trophic groups Wieser, 1953.
2.6. Data analysis
22
Experimental effects on the densities ind. per core 5 ind. 6.15 cm of major
meiofaunal taxa and nematode species numbers were examined using univariate analysis based on a two-way ANOVA linear model: y 5
m 1 Time 1 Treatment 1 Time 3 Treatment SPSS for Windows 9.0, 1998. If an interaction of main effects, i.e. time and
treatment, was detected, a one-way ANOVA was conducted to examine treatment effects at a specific stage of the experiment. Prior to the analysis of variance, data were
lnx 1 1 transformed to conform to an approximate normal distribution and make the variances constant Zar, 1999 and then Levene’s test was employed to check the
assumption of homogeneity. When the assumption of equal variances was rejected, a non-parametric Kruskal–Wallis test was used instead of the one-way ANOVA. Factors
detected to be significant by ANOVAs were further analyzed using a Bonferroni multiple comparison procedure which adjusts the observed significance level by multiplying it by
ˇ the number of comparisons being made Norusis, 1999.
Treatment effects at each specific stage of the experiment on nematode species diversity were checked by plotting K-dominance curves Lambshead et al., 1983; Clarke
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105
and Warwick, 1994. Changes in nematode community structure were examined using non-parametric multivariate techniques contained in the PRIMER Plymouth Routines in
Multivariate Ecological Research package. MDS non-metric Multidimensional Scaling Ordination was based on the Bray–Curtis similarity of either single square root
transformed or untransformed, nematode species abundance data. Untransformed data weight more contributions on common rather than rare species compared with the single
square root transformation Clarke and Warwick, 1994. The ANOSIM Analysis of Similarity technique was used to test treatment and time effects on the nematode
community succession process. A SIMPER Similarity Percentages program was then employed to identify those species contributing to differences between field and
experimental samples and between treatments observed in the MDS and ANOSIM analysis Clarke and Warwick, 1994.
For both the univariate and multivariate analyses, a significance level of P 0.05 was used as the rejection value. Field control samples F were also analyzed with
experimental samples C, L, M and H whenever appropriate.
3. Results
