Biodiversity Data Journal :
Data Paper (Biosciences)
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Corresponding author: Wai Lok So (henrysowailok@yahoo.com.hk), Michelle Man Suet Law (michellelaw@cuhk.edu.hk), Derrick Yuk Fo Lai (dyflai@cuhk.edu.hk), Jerome Ho Lam Hui (hui.jerome@gmail.com)
Academic editor: Pedro Cardoso
Received: 20 Feb 2022 | Accepted: 16 Aug 2022 | Published: 04 Oct 2022
© 2022 Wai Lok So, Ka Wai Ting, Sheung Yee Lai, Elaine Yi Ying Huang, Yue Ma, Tze Kiu Chong, Ho Yin Yip, Hoi Ting Lee, Billy Chun Ting Cheung, Man Ka Chan, Hong Kong Soil Biodiversity Consortium, Wenyan Nong, Michelle Man Suet Law, Derrick Yuk Fo Lai, Jerome Ho Lam Hui
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
So WL, Ting KW, Lai SY, Huang EYY, Ma Y, Chong TK, Yip HY, Lee HT, Cheung BCT, Chan MK, Consortium HKSB, Nong W, Law MMS, Lai DYF, Hui JHL (2022) Revealing the millipede and other soil-macrofaunal biodiversity in Hong Kong using a citizen science approach. Biodiversity Data Journal 10: e82518. https://doi.org/10.3897/BDJ.10.e82518
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Soil biodiversity plays important roles in nutrient recycling in both the environment and agriculture. However, they are generally understudied worldwide. To reveal the diversity of soil macrofauna in Hong Kong, here we initiated a citizen science project involving university, non-governmental organisations and secondary school students and teachers. It is envisioned that the citizen science approach used in this study could be used as a demonstration to future biodiversity sampling and monitoring studies.
Throughout a year of monitoring and species sampling across different localities in Hong Kong, 150 soil macrofaunal morphospecies were collected. Eighty five of them were further identified by morphology and DNA barcoding was assigned to each identified morphospecies, yielding a total of 646 DNA barcodes, with new millipede sequences deposited to the GenBank. The soil macrofauna morphospecies in Hong Kong found in this study are mainly dominated by millipedes (23 out of 150) and oligochaetes (15 out of 150). Amongst the twenty three identified millipedes, two polyxenid millipedes, Monographis queenslandica Huynh & Veenstra, 2013 and Alloproctoides remyi Marquet and Condé, 1950 are first recorded in Hong Kong. Information has been curated on an online platform and database (http://biodiversity.sls.cuhk.edu.hk/millipedes). A postcard summarising the findings of millipedes in Hong Kong has also been made as a souvenir and distributed to citizen participants. The identified macrofauna morphospecies and their 646 DNA barcodes in this study established a solid foundation for further research in soil biodiversity.
Hong Kong, citizen science, millipedes, macrofauna, soil biodiversity, DNA barcoding
Training younger generation citizens to learn about biodiversity is of utmost importance and crucial to conservation engagement. In recent years, the level and scope of citizen science has been fast-growing and improving worldwide and volunteers begin to participate in various aspects of environmental assessments (
Located on the south-eastern coast of China and experiencing a subtropical climate, Hong Kong has a relatively rich biodiversity nurturing more than 5,500 species of animals and plants (
Previous literature has identified a number of local millipedes, including Anaulaciulus tonginus Karsch, 1881 (
This current study aims to provide a new framework for carrying out citizen science projects in Hong Kong that contain research elements to provide information for the animal biodiversity status of the poorly studied habitat. Working between the university academics, taxonomists and non-governmental organisation members, secondary school students were recruited to collect soil macrofauna in the vicinity of their schools throughout a year and reveal the poorly studied soil animal biodiversity in Hong Kong using a citizen science approach.
Revealing the millipede and other soil-macrofaunal biodiversity in Hong Kong using a citizen science approach
Wai Lok So, Ka Wai Ting, Sheung Yee Lai, Elaine Yi Ying Huang, Yue Ma, Tze Kiu Chong, Ho Yin Yip, Hoi Ting Lee, Billy Chun Ting Cheung, Man Ka Chan, Hong Kong Soil Biodiversity Consortium, Wenyan Nong, Michelle Man Suet Law, Derrick Yuk Fo Lai, Jerome Ho Lam Hui
Selected collection sites in Hong Kong, China (Fig.
Location code | School location | Area | Site 1 | Site 2 | Site 3 |
A | Immaculate Heart of Mary College | N.T. & KLN |
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B | St. Stephen's Church College | HK Island |
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C | St. Stephen's College | HK Island |
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22.2166,114.2143 | |
D | Ling Liang Church E Wun Secondary School | Islands |
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E | Yan Oi Tong Tin Ka Ping Secondary School | N.T. & KLN |
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F | Ho Lap College | N.T. & KLN |
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G | New Asia Middle School | N.T. & KLN |
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H | De La Salle Secondary School | N.T. & KLN |
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I | G.T. (Ellen Yeung) College | N.T. & KLN |
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J | SKH Kei Hau Secondary School | N.T. & KLN |
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K | Mu Kuang English School | N.T. & KLN |
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L | CNEC Lau Wing Sang Secondary School | HK Island |
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M | Pui Kiu Middle School | HK Island |
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N | Fanling Rhenish Church Secondary School | N.T. & KLN |
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O | Buddhist Tai Kwong Chi Hong College | N.T. & KLN |
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P | SALEM-Immanuel Lutheran College | N.T. & KLN |
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Q | Queen Elizabeth School Old Students' Association Secondary School | N.T. & KLN |
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R | NT Heung Yee Kuk Yuen Long District Secondary School | N.T. & KLN |
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S | CCC Kei Long College | N.T. & KLN |
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T | Caritas Chan Chun Ha Field Studies Centre | Islands |
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U | The Chinese University of Hong Kong | N.T. & KLN |
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This project aims to provide species and DNA barcode data of the soil macrofauna in Hong Kong. Twenty one schools/institutions were involved and a total of 36 sampling sites were selected for species collection, based on their geographical distributions (Fig.
This project has been funded by the Environment and Conservation Fund (ECF 2018-82), Hong Kong Research Grant Council General Research Fund (14100919) and Collaborative Research Fund (C4015-20EF) and The Chinese University of Hong Kong Direct Grant (4053433, 4053489).
Specimen sampling was carried out on a biweekly basis between October and December in 2019 and monthly from January to October in 2020 due to the COVID-19 pandemic. Sampling techniques and requirements were introduced to secondary school teachers and students prior to sampling and the geographic coordinates were recorded using the online application Google Maps. Soil macrofaunal samples were either hand-picked or collected using a 1-millimetre pore sieve with a radius of 10 cm. On each site, at least 10 people were involved in hand-picking the observed soil organisms by searching over the soil. In addition, around 2 to 3 people were involved in sieving the soil mass from deeper soil and isolating fast-moving and smaller soil animals. Approximately 60 minutes were allocated for sampling each site each time and collected specimens were preserved into ethanol. For better preservation of specimens without the effect of ethanol decolouration and structure distortion before photo-taking in the laboratory, 95% ethanol were used for storing fresh samples, except for earthworms and millipedes, for which 10% ethanol was used instead for short-term storage. In addition, the soil mass from each location was collected twice from December 2019 to June 2020. Three replicates of soil mass distanced with 2 metres from each sample collection site with 1 kilogram each were collected in plastic bags. On-site preserved animals and soil masses were transferred to the laboratories at The Chinese University of Hong Kong within 12 hours after collection and stored at -20ºC and 4ºC respectively until subsequent analyses. After documentation of each specimen, each sample was replaced by 95% ethanol for long-term storage at -20°C.
The selected collection sites in Hong Kong, China (Fig.
22.2053 to 22.5018; 113.9451 to 114.2502.
Rank | Scientific Name |
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phylum | Arthropoda |
subphylum | Chelicerata |
class | Arachnida |
order | Araneae |
family | Lycosidae |
family | Linyphiidae |
family | Theridiidae |
family | Oonopidae |
family | Sparassidae |
subphylum | Myriapoda |
class | Symphyla |
class | Chilopoda |
order | Scolopendromorpha |
family | Cryptopidae |
family | Scolopendridae |
order | Geophilomorpha |
family | Mecistocephalidae |
class | Diplopoda |
order | Polyxenida |
family | Lophoproctidae |
family | Polyxenidae |
family | Paradoxosomatidae |
order | Glomerida |
family | Glomeridae |
order | Sphaerotheriida |
family | Zephroniidae |
order | Polyzoniida |
family | Siphonotidae |
order | Polydesmida |
family | Pyrgodesmidae |
family | Haplodesmidae |
order | Spirobolida |
family | Pachybolidae |
family | Spirobolellidae |
family | Pseudospirobolellidae |
order | Julida |
family | Julidae |
order | Spirostreptida |
family | Cambalopsidae |
subphylum | Hexapoda |
class | Insecta |
order | Blattodea |
family | Blaberidae |
family | Termitidae |
family | Blaberidae |
family | Ectobiidae |
order | Hymenoptera |
family | Formicidae |
order | Coleoptera |
family | Coccinellidae |
order | Crassiclitellata |
order | Dermaptera |
family | Anisolabididae |
order | Hemiptera |
family | Pentatomidae |
family | Cydnidae |
family | Aradidae |
order | Orthoptera |
order | Lepidoptera |
family | Erebidae |
family | Noctuidae |
order | Mantodea |
class | Malacostraca |
order | Isopoda |
family | Philosciidae |
family | Platyarthridae |
phylum | Annelida |
class | Clitellata |
order | Haplotaxida |
order | Opisthopora |
family | Megascolecidae |
family | Glossoscolecidae |
order | Systellommatophora |
family | Bradybaenidae |
phylum | Mollusca |
class | Gastropoda |
order | Stylommatophora |
family | Veronicellidae |
The dataset includes the soil biodiversity of Hong Kong during Oct 2019 - Oct 2020. The information includes the taxonomy of sampled organisms, geographic coordinates, collection location, date of collection and DNA barcodes (with NCBI Accession number).
Column label | Column description |
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Record identifier | record code for each entry. |
Phylum | Taxonomic rank (phylum) of collected sample. |
Class | Taxonomic rank (class) of collected sample. |
Order | Taxonomic rank (order) of collected sample . |
Family | Taxonomic rank (family) of collected sample. |
Species name | Scientific name of the collected sample. |
Latitude | Latitude. |
Longitude | Longitude. |
Datum | Global datum reference. |
Location code | Sample collection site. |
Country | Sample collection country. |
District | Sample collection district. |
Date | Date of sample collection. |
Season | Sample collection season. |
NCBI accession number | Accession number granted by NCBI. |
Barcoding gene | DNA barcoding gene. |
Morphological identification: Each collected soil specimen was mainly examined under a Nikon SMZ745T stereomicroscope and photo-documented with an adapted Canon DS126761 camera. Features of larger specimens were documented with an Olympus TG-4 camera. For the polyxenid millipedes, specimens were prepared, stained, mounted and observed under high magnifications of a compound microscope as previously described (
DNA extraction and barcoding: After photo-documentation of specimens, tissues from either the head or body were dissected and blotted on tissue paper to remove excess ethanol. Genomic DNA from these tissues was isolated by a spin-column based extraction method using the PureLink™ Genomic DNA Mini Kit (Invitrogen, USA), following the manufacturer's instructions. The remaining body parts of the specimens were transferred back to 95% ethanol and stored at -20ºC. The extracted DNA was subjected to quality and quantity control by 1% gel electrophoresis and One/Onec Microvolume UV-Vis Spectrophotometer (Thermo Scientific, NanoDrop, USA). The qualified genomic DNA was subjected to polymerase chain reaction (PCR) using a pair of universal primers (LCO1490: 5'-GGTCAACAAATCATAAAGATATTGG-3' & HCO2198: 5'-TAAACTTCAGGGTGACCAAAAAATCA-3') for amplifying mitochondrial cytochrome c oxidase subunit I (COI) gene of all collected samples (
Molecular identification and sequence analysis: The chromatogram of each barcode was examined base by base on the software SnapGene Viewer. Manual deletion of primer sequences at the 5' and 3' ends was performed for each barcoded sequence. The resultant sequence was submitted to NCBI GenBank for homology sequence search. If there were existing barcodes that matched at least 99% of the input sequence, the species identity was assigned to the specimen. If the match score was below the threshold (99%), it meant that we have produced the first sequence for the species that was not previously barcoded. All the barcodes were then submitted to NCBI and the accession numbers provided are listed in Suppl. material
Soil Macrofauna Composition: A total of 3,588 individual samples were collected from October 2019 to October 2020, including 150 different morphospecies. (Fig.
Amongst the 1,440 collected millipede samples, 23 millipede morphospecies were identified (Fig.
Soil analysis: The pH value and electrical conductivity of each soil slurry sample (soil:distilled water = 1:2.5 w/v) were measured by a pH meter (EA940, Orion Research Inc., USA) and conductivity meter (ION6+, Oakton Instruments, USA), respectively. After extraction of soil samples using 1 M of ammonium acetate (pH 7), the concentration of exchangeable cations (Na, K, Mg, Ca) were then determined by either atomic absorption spectroscopy (SpectrAA-200, Varian, USA) or inductively coupled plasma optical emission spectroscopy (5800 ICP-OES, Agilent, USA). For the organic matter, soil samples were gravimetrically combusted at 550ºC for 4 hours according to the loss-on-ignition method, whereas the total Kjeldahl nitrogen and total phosphorus content were determined by semi-micro Kjeldahl digestion and acid digestion, while the latter was then measured by UV-visible spectroscopy (UV-1800, Shimadzu, Japan).
Soil properties in winter and summer: when averaged across all the sampling sites, the mean soil pH was 5.64 ± 0.08 (1 standard error) in winter and 5.82 ± 0.08 in summer, which was acidic and typical of the soil acidity in the local environment. The mean soil electrical conductivity was 297.77 ± 20.30 µS/cm in winter and 316.92 ± 25.88 µS/cm in summer, which was indicative of a general salt-free environment. The mean soil organic matter content was 6.99% ± 0.26% in winter and 9.27% ± 0.78% in summer, which was in the moderate range and considered suitable for plant growth. The average percentages of clay, silt and sand particles were 15.0%, 16.0% and 69.0%, respectively in winter and 14.53%, 12.28% and 73.19%, respectively in summer, leading to a sandy loam texture. The mean total Kjeldahl nitrogen concentration was 0.24% ± 0.01% in winter and 0.34% ± 0.04% in summer, which fell within the medium range for plant growth. The total phosphorus concentration in soils was 754 ± 70.35 mg/kg in winter and 1545.85 ± 361.30 mg/kg in summer, which was higher than the values of 300-500 mg/kg reported in some natural subtropical Chinese forests. This also implied a great seasonal variation in total phosphorus concentration (Suppl. material
Before the beginning of this project, the understanding of soil biodiversity in Hong Kong, including the understanding of its contained millipede species, was inadequate. Previous studies have identified native millipede species, including Trigoniulus corallinus Gervais, 1847, Anaulaciulus tonginus Karsch, 1881, Hyleoglomeris bicolor Wood, 1865, Zephronia profuga Attems, 1936, Glyphiulus granulatus Gervais, 1847, Glyphiulus formosus Pocock, 1985 and Helicorthomorpha holstii Pocock 1895, to be present in local soil ecosystems (
Amongst the collected soil morphospecies, millipedes and earthworms are dominant. After reviewing the collected samples, we agree that there might be some collection bias throughout the survey process. Since the collection were mainly done by citizens through hand-picking methods, there is a tendency for the non-experts to collect those soil animals that are relatively large, conspicuous and slow-moving. Thus, this somehow explains a slightly biased collection effort within particular groups, neglecting some of the other soil macrofauna, including spiders and centipedes. In addition, the time (60 minutes) for on-site soil surveying that was adopted in the current study might not have provided enough time for isolation and collection of soil organisms. In the future, a mass of soil body should be collected and adequate time should be spent on isolating soil-dwelling animals in the laboratory. Furthermore, we recognise that the unequal number of sampling sites between the three regions (four localities in Hong Kong Island, two in the islands and fifteen in New Territories/Kowloon) might have also contributed to biased sampling effort. An expansion of collection sites in these other areas should be conducted in the future to generate a more complete and thorough survey. There is no doubt that the approach adopted in this study still has some technical flaws, but this study has raised public awareness and potentially opens up opportunities for the general public to engage in scientific research in the future.
In this study, we have only collected and identified 23 morphospecies. Comparing to the mentioned local public forum, HKWildlife.Net, some orders were not covered in this study, including Platydesmida, Siphonophorida and Chordeumatida. In addition, Cawjeekelia pallida (
Another interesting finding in this study is an unexpected discovery of Monographis queenslandica and Alloproctoides remyi in Hong Kong, which are the only two polyxenids identified locally. From the biogeographic perspective, it seems puzzling that with seven species of Monographis identified to date from Vietnam and two others in southern China, that the Monographis species identified in Hong Kong is the one that is found in Australia. We propose that there might be possibilities that the species was introduced to Hong Kong many decades ago in soil products or goods from Queensland, Australia or vice versa. The other species, A. remyi, was first identified from Reunion and more recently from Mauritius. A number of polyxenids from the family Lophoproctidae are also quite widespread in their distributions, so this might also be the case for A. remyi.
This study differs from most conventional scientific studies being carried out in Hong Kong, which were mainly carried out by either government, non-governmental organisations or academics in universities alone. Utilising a citizen science approach through creating a "big community" (a consortium; Suppl. material
In terms of dissemination of the study, in addition to traditional academic output including this manuscript, sharing sessions by secondary school students were also carried out (online due to the COVID-19 pandemic). We have also summarised the findings and made online videos to a publicly-available online database/platform (http://biodiversity.sls.cuhk.edu.hk/millipedes) and designed a postcard (Fig.
This study has demonstrated a clear success in surveying the soil biodiversity in Hong Kong and opens up a possibility to carry out similar large-scale surveying via citizen science, together with taxonomic experts and researchers from universities. It is envisioned that the framework established in this study can also be adopted to reveal the biodiversity in other habitats in this region.
The authors would like to show our gratitude to Dr. Sergi Golovatch for his help on the identification of collected millipede samples. The appreciation also extends to Dr. Cuong Huynh and Dr. Megan Short for their identification of the Polyxenida. The authors would also like to acknowledge Dr. Annette Yin Pui Wong and Dr. Zhe Qu for the funding acquisition of the study. Finally, the authors would like to thank our reviewer, Dr. Carlos A. Martínez-Muñoz, for his thoughtful comments and efforts towards improving our manuscript.
JHLH conceived the study. JHLH, DYFL, WLS and MMSL supervised the study. WLS, KWT, SYL and EYYH contributed equally to this paper as co-first authors. WLS, KWT, SYL, TKC, HYY, HTL, BCTC, MKC and HKSBC carried out sample collection and DNA barcoding. WN managed the barcoding data and the development of the website. YM carried out soil sample analysis. WLS designed the Figures. SYL designed the postcard. WLS, HYY, WN, MMSL, DYFL and JHLH contributed to funding application. WLS, KWT, SYL, EYYH and JHLH wrote the first draft of manuscript.
The dataset includes the soil biodiversity of Hong Kong during Oct 2019 - Oct 2020. The information includes the taxonomy of sampled organisms, geographic coordinates, collection location, date of collection and DNA barcodes (with NCBI Accession number).