Eusideroxylon Zwageri)

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1 IMPENDING REGENERATION FAILURE OF THE IUCN VULNERABLE BORNEO IRONWOOD

2 (EUSIDEROXYLON ZWAGERI)

3 Lan Qie1, Alexander D. Elsy1, Ashley Stumvoll1, Magdalena Kwasnicka1, Anna L. Peel1, Joseph A. 4 Sullivan1, Maisie S. Ettinger1, Alasdair J. Robertson1, Jeanelle K. Brisbane1, Amber L. Sawyer1, 5 Yan N. Lui1, Siew Ngim Ow1, Matteo Sebastianelli1, Bartosz Majcher1, Muying Duan1, Hannah 6 Vigus1, Grace Pounsin2, Reuben Nilus3, and Robert Ewers1

7 1Department of Life Sciences, Silwood Park Campus, Imperial College London, Ascot SL5 7PY, UK

8 2Maliau Basin Conservation Area, Yayasan Sabah, 88817 Kota Kinabalu, Sabah, Malaysia

9 3Sabah Forestry Department Forest Research Centre, Mile 14 Jl. Sepilok, 90000 Sandakan, 10 Malaysia.

12 Abstract

13 The regeneration of many climax species in tropical forest critically depends on adequate seed

14 dispersal and seedling establishment. Here we report the decreased abundance and increased

15 spatial aggregation of younger trees of the Borneo ironwood (Eusideroxylon zwageri) in a

16 protected forest in Sabah Malaysia. We observed a high level of seedling herbivory with strong

17 density dependence, likely exacerbated by local aggregation and contributing to the

18 progressively shrinking size-distribution. We also note the largely undocumented selective

19 herbivory by sambar deer on E. zwageri seedlings. This study highlights the combined impact of

20 altered megafauna community on a tree population through interlinked ecological processes

21 and the need for targeted conservation intervention for this iconic tropical tree species.

23 Keywords

24 Seedling survival, neighborhood density, dispersal limitation, spatial clustering, threatened

25 species, wildings

26 INTRODUCTION

27 The loss of megafauna due to climate change, habitat alteration and hunting has a strong

28 impact on plants that are dependent on large-bodied animals for seed dispersal, and this effect

29 is particularly strong in tropical forests (Corlett 2010, Galetti et al. 2017). Many tree species

30 with large fruits have lost the animal dispersers they coevolved with, and consequently suffer

31 reduced seed dispersal and increased spatial aggregation of seedlings, leading to lower survival

32 and reduced gene flow (Harrison et al. 2013, Galetti et al. 2017). Such trees are typically late-

33 succession to canopy species (Harrison et al. 2016), and their regeneration is crucial to the

34 future of intact tropical forests and restoration efforts to accelerate succession in disturbed

35 forests.

36 Borneo ironwood (Eusideroxylon zwageri) is a classic example of rainforest climax tree species

37 facing regeneration challenges. E. zwageri is distributed in eastern and southern Sumatra,

38 Bangka, Belitung, Borneo, the Sulu archipelago and Palawan. In Borneo, E. zwageri was

39 formerly a common component of the main middle story in the mixed dipterocarp forest. It is a

40 prized timber species, renowned for its extraordinary strength, durability and rot resistance,

41 and it is a cultural keystone species to the indigenous people of the region (Franco et al. 2014).

42 Despite being long-living (>1000 years), due to over-exploitation (Peluso 1992) and slow natural

43 regeneration (typically requires >100 years to reach 30 cm diameter) (Irawan 2005), E. zwageri

44 has become scarce across its distributional range and is classified as Vulnerable in the IUCN Red

45 List (1998) . The species produces large drupaceous fruits that measure 10-18 cm x 5-10 cm,

46 and contain a single large seed, measuring 7-15 cm x 4-7 cm, with a very hard seed coat (Irawan

47 2005). Water dispersal for the heavy seeds of E. zwageri is possible but limited as the species

48 often occur away from rivers (Irawan 2005). The large fruit with edible fleshy mesocarp and nut

49 are strong indications that E. zwageri is dispersed by animals, although evidence of this is

50 anecdotal and mainly refers to porcupines (Kostermans et al. 1994, Phillipps & Phillipps 2016).

51 Only megafauna (>44 kg in body mass) would be capable of long-distance dispersal of these

52 large seeds (Galetti et al. 2017). It has been speculated that E. zwageri seeds were dispersed by

53 the Sumatran rhinoceros (Dicerorhinus sumatrensis) (Phillipps & Phillipps 2016), formerly

54 occurring throughout Borneo but now virtually extinct in the wild in Sabah, and down to the

55 last few individuals, if any, still surviving in Borneo (Hance 2015). Despite the value and

56 threatened status of E. zwageri, we still have very limited understanding of its ecology and, in

57 particular, its natural regeneration from seeds.

58 Post germination, the seedling stage represents another bottleneck for the regeneration of a

59 tree species under survival pressures such as pathogens and seedling herbivory, which are

60 typically density dependent processes (Wang & Smith 2002). There have been anecdotal

61 reports that one of the common herbivores in Bornean forests, the sambar deer (Rusa

62 unicolor), preferentially browsed on E. zwageri seedlings. This would potentially exacerbate the

63 regeneration challenge already faced by E. zwageri. Once stems survive the dynamic stage and

64 reach 10 cm diameter the degree of spatial aggregation is expected to be stable across size

65 classes (Condit et al. 2000). Therefore in a long-living E. zwageri population (oldest trees may

66 be > 1000 years), spatial aggregation in younger versus older trees should reflect long-term

67 change in the strength of seed dispersal. We conducted a survey of E. zwageri across all life

68 stages from seedlings to adult trees in an old growth Bornean lowland forest and tested three

69 hypotheses: 1) reduced regeneration in this local population will be reflected in its tree size

70 distribution curve with a flat gradient at small size classes, or a unimodal shape (Halpin &

71 Lorimer 2017), 2) there is increased spatial aggregation in the younger sub-population

72 compared to the older sub-population of adult trees (≥ 10 cm diameter), and 3) seedlings

73 herbivory damage has density dependence.

75 MATERIALS AND METHODS

76 We conducted this study in January 2018 at Maliau Basin Conservation Area (4.74° N, 116.97° E,

77 256 m a.s.l.), Sabah, Malaysia. The survey area was in old growth forest on the south bank of

78 the Maliau River near the edge of the conservation area, in the vicinity of a large buffer zone

79 consisting of selectively logged and secondary forests. A 4-ha square plot was set up, within

80 which a grid of 10 by 10 m subplots was established for tree surveys. All E. zwageri trees with

81 DBH ≥ 1 cm were mapped and recorded to diameter classes in 10 cm intervals.

82 There were abundant E. zwageri seedlings in the area from the January 2017 fruiting event.

83 Many of these had noticeable herbivory damage with young leaves missing from the top,

84 characteristic of that by mammalian herbivores, and often resulting in complete defoliation. We

85 surveyed seedlings (all stages with DBH < 1 cm) in a 1.35 ha area in the centre of the 4-ha plot,

86 using 5 by 5 m subplots within the 10 by 10 m grid. In each seedling subplot, we counted E.

87 zwageri seedlings in two categories: “surviving” if the apical meristem was intact, and “fatally

88 damaged” if the apical meristem was eaten, including dead E. zwageri seedlings with evidence

89 of herbivory damage. Leafless and dead E. zwageri seedlings were identified by their

90 characteristic straight and reddish bare stem, often with the stony seed coat still attached at

91 root. For each subplot we also recorded ground vegetation cover as a proxy for interspecific

92 competition, estimated into four classes, 0-25%, 25-50%, 50-75% and 75-100%.

93 We quantified spatial aggregation of E. zwageri adult trees (DBH ≥ 10 cm) using the relative

94 neighborhood density Ωx, defined as the average density of neighbours in the annulus at

95 distance x for each focal tree, standardized by mean population density (Condit et al. 2000). We

96 calculated Ωx for the 0-100m distance range in 10 m steps, and compared this metric for the

97 older and younger sub-populations, categorized as those above or below the median tree size

98 respectively.

99 For E. zwageri seedlings we used a generalized linear model (GLM) with a binomial error

100 distribution and a logit link function to test the effects of conspecific seedling density and

101 vegetation cover on the proportion of fatal herbivory damage. All analyses were conducted in

102 the R statistical computing environment (R Core Team, 2016).

103

104 RESULTS

105 We recorded 90 E. zwageri trees (DBH ≥ 1 cm) in the 4 ha plot, among which 80 were adult

106 trees (DBH ≥ 10 cm). As predicted from the reduced regeneration hypothesis the adult tree size

107 distribution had a unimodal shape peaking at the 70 cm class (median) and decreasing towards

108 smaller classes (Fig. 1a). All adult trees were aggregated at the 0-10m scale (Ωx > 1), and the

109 degree of aggregation was significantly higher in the younger sub-population, consistent with

110 the expectation of increased dispersal limitation over time (Fig 1b).

111 A total of 495 E. zwageri seedlings were recorded in 197 of the 540 5 by 5 m subplots surveyed,

112 with an overall mean density of 367 seedlings ha-1. Nearly all appeared to be less than one year

113 old and resulting from the fruiting event in January 2017. Among these, 53.3% had fatal

114 herbivory damage, already dead or highly unlikely to survive. The proportion of fatal herbivory

115 damage increased rapidly with conspecific seedling density at subplot level (p < 0.001),

116 predicted to reach 0.97 at 1 seedling m-2 (Fig. 2). Fatal herbivory was not significantly associated

117 with vegetation cover (p = 0.09).

118

119 DISCUSSION

120 The remarkably low abundance of young E. zwageri trees indicates that the population of this

121 species is not sustainable in this forest stand (Halpin & Lorimer 2017). E. zwageri adult trees

122 were aggregated at the 0-10 m spatial scale, characteristic of tropical trees (Condit et al. 2000);

123 a similar aggregation pattern at this scale was observed at genotypical level in a recent study

124 (Shao et al. 2017). However, the increased spatial aggregation of E. zwageri in younger trees

125 supports the hypothesis of an increasing seed dispersal limitation over time due to the lack of

126 seed dispersers, as shown in another Bornean forest (Harrison et al. 2013). Most strikingly, E.

127 zwageri seedlings more than one year old (height > 1 m) were almost absent, suggesting

128 complete failure of seedling establishment in recent years. This can be explained largely by the

129 high rate of fatal seedling herbivory, increasing with local aggregation (Fig. 2). These patterns

130 highlight the effect of dispersal limitation and negative density-dependent seedling survival,

131 operating together to influence species spatial distribution (Shao et al. 2017).

132 A survey of 13 other Bornean canopy species found a seedling survival rate of approximately

133 50% over ten years (Delissio et al. 2002). In contrast, we observed an overall survival rate of

134 46.7% in E. zwageri seedlings in less than one year since germination. The actual survival rate

135 may be even lower as we only accounted for dead stems with detectable remains. Using our

136 model as a conservative approximation of the density dependent annual mortality rate and

137 applying it to the surviving E. zwageri seedlings at subplot level, we predict that after six years

138 seedling density would drop to merely 3 individuals ha-1, leading to a progressively shrinking

139 demography (Fig. 1a). Seedling height may not reach 2 m in this time (Irawan 2005), and thus

140 they remain susceptible to ground level herbivory.

141 The heavy herbivory observed here was apparently associated with a single mammalian

142 herbivore, the sambar deer, based on characteristics of the stem damage and reports by local

143 rangers. Adapted to surviving shade conditions for relatively long period, E. zwageri seedlings

144 allocate a high proportion of resources towards defense in the form of concentrated tannin (CT)

145 and lignin (Kurokawa et al. 2004). This is an evolved mechanism against invertebrate herbivores

146 and pathogens, but mammalian herbivores such as sambar deer have co-evolved to produce a

147 tannin-binding protein in their saliva, and in fact show a preference towards plant species high

148 in CT and lignin (Semiadil et al. 1995).

149 It is possible that at local scale, browsing pressure from sambar deer can influence E. zwageri

150 regeneration. A generalist grazer and browser, the sambar deer appears to benefit from

151 intermediate forest disturbance, preferring forest edges and selectively logged forest due to

152 the increased availability of phytomass on the ground (Corlett 2007, Meijaard & Sheil 2007). A

153 recent study in our research area found that mammalian herbivores showed a 19% increase in

154 abundance in logged forest compared to unlogged forest and, in particular, the abundance of

155 sambar deer more than doubled (Wearn et al. 2017). There are few large predators in Borneo,

156 and predation pressure on this species here predominantly comes from hunting (Bennett et al.

157 2000). It is likely that in protected areas where hunting is prohibited the sambar deer has

158 enjoyed predator release. Our study site is in a protected area adjacent to a large buffer zone

159 consisting of selectively logged forests. The sambar deer population here therefore have

160 probably benefited from both favorable habitat and the absence of hunting. As an interesting

161 contrast, sambar deer population was low in another Sabah forest (Kabili-Sepilok) (Ross et al.

162 2013), and greater densities of young E. zwageri trees were recorded, on average 49 stems with

163 DBH 5 – 10 cm per 4-ha (Qie and Nilus, unpublished data).

164 Our results shine a spotlight on an iconic tree species E. zwageri. We show the intensified

165 interaction between this native tree and a native mammalian herbivore resulting from changes

166 in the megafauna community and forest landscape in Borneo. Anthropogenic forces on the

167 forest ecosystem in recent centuries seem to have eradicated its long-distance seed dispersers

168 and in some places favored its seedling herbivores. Where these impacts co-occur they may

169 lead to an impending regeneration failure of E. zwageri at local scale, as we are starting to see

170 in this population.

171 E. zwageri requires active conservation intervention and efforts should focus on its most

172 vulnerable seedling stage. Once survived through this recruitment bottleneck, the tall saplings

173 and trees of E. zwageri are known to be exceptionally resilient, including being drought- and

174 fire- tolerant (Delmy 2001, Van Nieuwstadt & Sheil 2005). Ex situ propagation of this species

175 has only been carried out on small scales partly due to inadequate supply of seeds and

176 seedlings (Azani et al. 2001, Irawan 2012), yet we show that a great number of E. zwageri

177 seedlings in natural forests are being lost to high mortality. Our results suggest that at local

178 density of < 0.2 m-2 E. zwageri seedlings may have optimal survival. Therefore, moderate

179 harvest of E. zwageri seeds and seedlings in natural forests (i.e. wildlings) for ex situ cultivation

180 and re-introduction once they reach sapling stage should be feasible. This approach may

181 simultaneously increase E. zwageri regeneration and population gene flow, providing a cost-

182 effective way of assisted forest regeneration.

183

184 ACKNOWLEDGEMENT

185 This study was supported by the Imperial College London MRes Tropical Forest Ecology field

186 course program. We thank the Sabah Biodiversity Centre for access license and the excellent

187 support of staff members at the Maliau Basin Studies Centre.

188

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254

255

256 FIGURE 1. Demographic and spatial distribution of E. zwageri adult trees (DBH ≥ 10 cm) in the

257 4-ha plot. a) Population size distribution with a unimodal shape and a mode at the 70 cm class.

258 b) Spatial aggregation measured as the mean relative neighborhood density in 10m annuli each

259 focal tree, standardized by mean population density. The population was divided into the older

260 (DBH ≥70cm) and younger (DBH < 70 cm) sub-populations. Bars represent 95% bootstrapped

261 confidence intervals.

262

263 FIGURE 2. Relationship between the rate of fatal herbivory of E. zwageri seedlings and

264 conspecific seedling density at 5x5m subplot level. Line was fitted using a GLM model. Shading

265 areas represent confidence intervals.

266