First record of abnormal body coloration in a rockfish Sebastes koreanus(Scorpaenoidei:Sebastidae)from coastal water of China based on morphological characteristics and DNA barcoding

The first record of abnormal body coloration in Sebastes koreanus Kim and Lee,1994,from the Yellow Sea of China,was documented based on morphological characteristics and DNA barcoding.The two rockfish specimens were collected from the coastal waters of Qingdao,China,and the whole body and all fins of them were red.Of the two red-colored rockfish,there were tiny deep red spots on each fin,2 red radial stripes behind and below the eyes and 1 large deep red blotch on the opercula,while the similar stripe and spot patterns are also present in the S.koreanus specimens with normal body coloration.The countable characteristics of the two specimens are in the range of the morphometry of S.koreanus.To further clarify the species identity and taxonomic status of the two specimens,DNA barcode analysis was carried out.The genetic distance between the red-colored rockfish and S.koreanus was 0,and the minimum net genetic distances between the red-colored rockfish and other Sebastes species except for S.koreanus were 3.0%,which exceeds the threshold of species delimitation.The phylogenetic analysis showed that the DNA barcoding sequences of the two red-colored rockfish clustered with the S.koreanus sequences.The above results of DNA barcode analysis also support that the two red-colored rockfish could be identified as the species of S.koreanus.The mechanism of color variation in S.koreanus is desirable for further research and the species could be an ideal model to study the color-driven speciation of the rockfishes.

DNA Barcoding of Insects and Its Direct Application for Plant Protection

The introduction of invasive insect pests across national borders has become a major concern in crop production. Accordingly, national plant protection organizations are challenge to reinforce their monitoring strategies, which are hampered by the weight and size of inspection equipment, as well as the taxonomic extensiveness of interrupted species. Moreover, some insect pests that impede farmer productivity and profitability are difficult for researchers to address on time due to a lack of appropriate plant protection measures. Farmers’ reliance on synthetic pesticides and biocontrol agents has resulted in major economic and environmental ramifications. DNA barcoding is a novel technology that has the potential to improve Integrated Pest Management decision-making, which is dependent on the ability to correctly identify pest and beneficial organisms. This is due to some natural traits such as phenology or pesticide susceptibility browbeaten by IPM strategies to avert pest establishment. Specifically, Deoxyribonucleic acid (DNA) sequence information was applied effectively for the identification of some micro-organisms. This technology, DNA barcoding, allows for the identification of insect species by using short, standardized gene sequences. DNA barcoding is basically based on repeatable and accessible technique that allows for the mechanisation or automation of species discrimination. This technique bridges the taxonomic bio-security gap and meets the International Plant Protection Convention diagnostic standards for insect identification. This review therefore discusses DNA barcoding as a technique for insect pests’ identification and its potential application for crop protection.

Analysis of the Authenticity of Stone Medicinal Plants by DNA Barcoding

The genus Pyrrosia belongs to the family Polypodiaceae and are medium-sized epiphytic ferns,where the dried leaves of Pyrrosia lingua,Pyrrosia sheareri,Pyrrosia lanceolata,and Pyrrosia calvata are commonly used in medicinal practice.In this study,the authenticity of the collected medicinal plant samples of Shiwei was identified with the help of DNA barcoding technology using the internal transcribed spacer 2(ITS2)as the identifying sequence.The experimental samples were analyzed using the basic local alignment search tool(BLAST)and the authenticity of the samples was further verified with the results of similarity comparison.The results proved that the sequences of the experimentally collected samples of Pyrrosia lingua,Pyrrosia sheareri,Pyrrosia lanceolata,and Pyrrosia calvata had a similarity of more than 97%when compared with the corresponding sequences that were uploaded on the Internet.

DNA barcoding of fishes from Zhoushan coastal waters using mitochondrial COI and 12S rRNA genes

Accurate species identification is a key component of biodiversity research.DNA barcoding is an effective molecular method used for fish species identification.We aimed to study the DNA barcoding of fish in Zhoushan coastal waters,explore the differences and applicability of two gene fragments(12S rRNA and COI)of DNA barcoding in fish species identification,and established a comprehensive fish barcoding reference database.Two hundred and eighty-seven captured fish samples from Zhoushan coastal waters were identified using morphological characteristics and DNA barcoding.A total of 26412S rRNA sequences(belonging to eight orders,31 families,55 genera,and 66 species)and 188 COI sequences(belonging to seven orders,30 families,48 genera,and 58 species)were obtained.The lengths of the 12S rRNA sequences ranged from 165 to 178 bp,and the guanine-cytosine(GC)content was 45.37%.The average 12S rRNA interspecific and intraspecific genetic distances(K2P)were 0.10%and 26.66%,respectively.The length of the COI sequence ranged 574–655 bp,and the content of GC was 45.97%.The average 12S rRNA interspecific and intraspecific genetic distances(K2P)were 0.16%and 27.45%,respectively.The minimum interspecific genetic distances of 12S rRNA and COI(1.23%and 1.86%)were both greater than their maximum intraspecific genetic distances(2.42%and 8.66%).Three molecular analyses(NJ tree,ABGD,and GMYC)were performed to accurately identify and delineate species.Clustering errors occurred when the 12S rRNA sequences were delimited using the NJ tree method,and the delimitation results of ABGD and GMYC are consistent with the final species identification results.Our results demonstrate that DNA barcoding based on 12S rRNA and COI can be used as an effective tool for fish species identification,and 12S rRNA has good application prospects in the environmental DNA(eDNA)metabarcoding of marine fish.

DNA Barcoding and Mini-DNA Barcoding Reveal Mislabeling of Salmonids in Different Distribution Channels in the Qingdao Area

There is an increasing demand for salmonid authentication due to the globalization of the salmonid trade.DNA barcoding and mini-DNA barcoding are widely used for identifying fish species based on a fragment of the mitochondrial cytochrome c oxidase subunit I(COI)sequence.In this study,rainbow trout(Oncorhynchus mykiss),steelhead trout(O.mykiss),and Atlantic salmon(Salmo salar)collected from two salmonid aquaculture bases in China were authenticated by DNA barcoding(about 650 bp)and mini-DNA barcoding(127 bp)to evaluate the accuracy of the two methods in the identification of different salmonid species.The results revealed that both methods could effectively distinguish O.mykiss and S.salar with 100%accuracy.However,the two methods failed to separate rainbow trout(O.mykiss)and steelhead trout(O.mykiss),which are the same species but cultured in different water environments.Moreover,salmonid samples from three main distribution channels in the Qingdao area(traditional supermarkets,online supermarkets,and sushi bars)were identified by the two methods.Substitution of S.salar with O.mykiss was discovered,and the 27.78%overall substitution rate of salmonids in the Qingdao area was higher than those in other regions reported in previous studies.In addition,the mislabeling rates of salmonids from traditional supermarkets,online supermarkets,and sushi bars were compared in this study.The mislabeling rate was significantly greater in sushi bars(50%)than in the other two channels(16.67%),suggesting that stronger monitoring and enforcement measures are necessary for the aquatic food catering industry.

Identification of Kalidium species(Chenopodiaceae)by DNA barcoding

DNA barcoding is an increasingly prevalent molecular biological technology which uses a short and conserved DNA fragment to facilitate rapid and accurate species identification. Kalidium species are distributed in saline soil habitat throughout Southeast Europe and Northwest Asia, and used mainly as forage grass in China. The discrimination of Kalidium species was based only on morphology-based identification systems and limited to recognized species. Here, we tested four DNA candidate loci, one nuclear locus(ITS, internal transcribed spacer) and three plastid loci(rbc L, mat K and ycf1b), to select potential DNA barcodes for identifying different Kalidium species. Results showed that the best DNA barcode was ITS locus, which displayed the highest species discrimination rate(100%), followed by mat K(33.3%), ycf1b(16.7%), and rbc L(16.7%). Meanwhile, four loci clearly identified the variant species, Kalidium cuspidatum(Ung.-Sternb.) Grub.var. sinicum A. J. Li, as a single species in Kalidium.

Morphological characteristics and DNA barcoding of Pampus echinogaster(Basilewsky, 1855)

The morphological similarities of Pampus fishes have led to considerable confusion in species-level identification,and no accurate information on neotype or DNA barcoding of Pampus echinogaster is available. Two hundred and seven specimens of P. echinogaster were collected from the coastal waters of Dandong, Dongying, Qingdao,Nantong, Zhoushan, Wenzhou, Changle, Taiwan, and Wakayama(Japan), from June 2010 to April 2013. The diagnostic characteristics of P. echinogaster are as follows: dorsal fin VIII-XI-43–51, anal fin V-VIII-43–49, pectoral fin 22–27, caudal fin 19–22, pelvic fin absent; first gill rakers sparse, slender(pointed), 3–4+12–16=15–20; vertebrae39–41; transverse occipital canal on top of head moderately small, wavy ridges not reaching upper origin of pectoral fin; ventral branch of lateral line canal spare, shorter than dorsal branch of lateral line canal. By combining congener sequences of the cytochrome oxidase I(COI) gene from Gen Bank, two absolute groups were detected among all specimens, which further indicated that two valid species were present based on genetic differences in amino acid sequences and the distance between the groups. The sequences of Group 1 can be regarded as DNA barcoding of P. echinogaster. The correct morphological redescription and DNA barcoding of P.echinogaster are presented here to provide a guarantee for efficient and accurate studies, a theoretical basis for classification, and enable appropriate fishery management and conservation strategies for the genus Pampus in the future.

Molecular Identification of Dried Shellfish Products Sold on the Market Using DNA Barcoding

The dried shellfish products with rich nutrients and low-calorie content are favorite food in China,especially in coastal areas.However,the species of dried shellfish products in the market are usually unknown,as the taxonomic features were removed during the production process.This study described the application of DNA barcoding technique to the identification of 100 dried shellfish(scallop,squid,octopus and cuttlefish)products in markets.Samples were authenticated by comparing mitochondrial cytochrome oxidase subunit I(COI)gene and 16S ribosomal RNA(16S rRNA)gene sequences with public reference taxonomic databases.The results showed that all the 100 products can be identified at species level.Sixty four scallop adductor products were processed using the bay scallop,Argopecten irradians,and one was from Portuguese oyster,Crassostrea angulata.All the 27 squid,2 cuttlefish and 6 octopus products were produced by the Jumbo flying squid,Dosidicus gigas.The neighbour-joining tree is in agreement with the results of DNA barcoding analysis.The 64 scallop samples formed one A.irradians cluster,leaving Sca65 clustered with the reference oyster sequence C.angulata(MH997922).All the 35 cephalopod(squid,octopus and cuttlefish)samples formed a D.gigas cluster.This investigation revealed a low variety of dried shellfish products sold on the market,and highlighted the high rate of mislabeling and species substitution.Our present work provides a practical method for tracing and authenticating shellfish products.

Identification and DNA Barcoding of a New Sillago Species in Beihai and Zhanjiang,China,with a Key to Related Species

A new Sillago species,Sillago parasihama sp.n.,is identified based on 127 specimens collected from the southern coast of China.We compared the morphological characters between Sillago parasihama and all other 11 Sillago species with two posterior extensions on the swim bladder.The new species is like S.sihama in the countable characters and color pattern,but is different from the latter by the distinct swim bladders.The swim bladder of S.parasihama is without lateral process.The posterior sub-extensions of anterolateral extensions are unique with some dendritic or sometimes stunted blind tubule,which are unilateral and outward,ex-tending along the abdominal,and are about one-third to half of the body of swim bladder in length.But the swim bladder of S.si-hama with 8-10 lateral processes,the posterior sub-extensions of anterolateral extensions are kinky,long and complicated,extend-ing along the abdominal wall below the peritoneum to the base of posterior extensions.S.parasihama can be distinguished from other species in this group by color pattern,meristic,and morphometric characters.Moreover,the results of genetic analysis using sequences of the mitochondrial cytochrome c oxidase subunit I(COI)fragment show significant interspecific-level genetic distances(0.159-0.231)between S.parasihama and 8 congeners in the group,which also support the validity of new species.We also provide a distribution map and a key of the related species.

Discovery of Cladonema multiramosum sp.nov.(Cnidaria:Hydrozoa:Cladonematidae)using DNA barcoding and life cycle analyses

In contrast to typical planktonic hydromedusae,Cladonema medusae are mostly benthic,with specialised adhesive branches to adhere to the substrate.In this study,a Cladonema species discovered in a laboratory aquarium in Fuzhou,China,was confirmed as a new species,based on morphological and molecular analyses.The species was named Cladonema multiramosum sp.nov.Its medusa is distinct from that of congeners possessing substantially more adhesive branches(8-24,rarely 5-7),and tiny branches on the upper radial canals.The validity of C.multiramosumum sp.nov.was also supported by molecular phylogenetic analyses based on the mitochondrial 16S rDNA sequence.However,C.multiramosum sp.nov.medusa also displayed considerable phenotypic plasticity with respect to its radial canals,tentacles,stinging branches per tentacle,oral tentacles,manubrium,and gonads,hindering species identification based solely on morphology.Although some morphological characteristics of hydroids(filiform tentacles and medusa buds)and nematocysts could also be used as diagnostic characters in the genus Cladonema,this information is unavailable for some Cladonema species.Thus,the taxonomy within the genus Cladonema was re-evaluated based mainly on the morphological characteristics of the medusa.Further revision of the genus Cladonema should be made when supplementary information on the life cycle and DNA barcoding are updated.

DNA barcoding and molecular phylogeny of Dumasia(Fabaceae:Phaseoleae)reveals a cryptic lineage

Dumasia taxonomy and classification have long been problematic.Species within this genus have few morphological differences and plants without flowers or fruits are difficult to accurately identify.In this study,we evaluated the ability of six DNA barcoding sequences,one nuclear(ITS)and five chloroplast regions(trnH-psbA,matK,rbcL,trnL-trnF,psbB-psbF),to efficiently identify Dumasia species.Most single markers or their combinations identify obvious barcoding gaps between intraspecific and interspecific genetic variation.Most combined analyses including ITS showed good species resolution and identification efficiency.We therefore suggest that ITS alone or a combination of ITS with any cpDNA marker are most suitable for DNA barcoding of Dumasia.The phylogenetic analyses clearly indicated that Dumasia yunnanensis is not monophyletic and is separated as two independent branches,which may result from cryptic differentiation.Our results demonstrate that molecular data can deepen the comprehension of taxonomy of Dumasia and provide an efficient approach for identification of the species.

DNA Barcoding of Nigeria’s Forest Species Listed in CITES and Other Endangered Plant Species of National Interest

Illegal trade is considered one of the greatest threats to the loss of biodiversity of endangered plants. Some of these plant species are often trafficked in processed forms, making it extremely difficult for taxonomic experts to identify them. In the past, illegal traders of endangered species have been arrested and prosecuted but eventually cleared due to a lack of conclusive evidence. DNA barcoding is a veritable tool to protect these endangered species from illegal trade. It identifies all stages of the species’ life forms including processed products (milled or powdered animal and plant parts). The study utilised the rbcL gene as a single barcode region in the identification/authentication of 19 Nigeria’s endangered forest species legislated under the CITES and other endangered species of national interest. The generated sequence barcodes were used to query NCBI-GenBank and BOLD databases. 57.89% of the samples were identified down to species level and 42.11% to genus level. Amongst the 19 samples, sample (S7) yielded a high-quality sequence for a single sequencing read (forward), sufficient to identify the sample with a 99.81% identity match on NCBI-GenBank and BOLD. The results reveal that the rbcL single barcode efficiently identified most of the sampled plants;this supports the potential utilisation of DNA barcoding in the accurate detection and conservation of CITES-listed plants in Nigeria. The study documented the CITES-listed plants and other essential plants endangered or threatened plants in Nigeria and provided the first chloroplast DNA reference dataset to support the utilisation of DNA barcoding to identify CITES-listed plant species in Nigeria, which is significant for future studies.

DNA barcoding for the identification of Limonia crane flies(Diptera:Limoniidae)from China,including a new species and a newly recorded species

The genus Limonia Meigen,1803 is globally distributed with 216 known species/subspecies,of which 30 are recorded from China.In this study,we firstly use DNA barcodes(mt COI)to identify specimens of Limonia from China,result in a discovery of three species.Considering the morphological data,these species were identified as L.albiterminalis Lü,Ren&Zhang,sp.nov.,L.juvenca Alexander,1935(new record in China),and L.macrostigma(Schummel,1829),respectively.Six COI sequences of the three species were provided,analyzed with other 24 COI sequences from Limonia species.The result indicates that intraspecific distances in the genus are generally less than 2.2%,interspecific distances range from 7.9%to 17.2%,and there is no overlap between intra-and interspecific distances.Redescription and illustrations of L.juvenca and illustration of male hypopygium of L.macrostigma,as well as the key to the Chinese Limonia crane flies,are all provided.

Relationship between DNA Barcoding and Chemical Classification of Salvia Medicinal Herbs

Objective To make the identification of medicinal herbs in Salvia L.quickly and accurately.Methods In this work,DNA barcoding and chemical fingerprint were compared for the identification of herbs in Salvia L.First,the nucleotide sequences of the internal transcribed spacer region two amplified from 48 medicinal plants in Salvia L.,and three other groups of medicinal plants in Lamiaceae were sequenced.A molecular phylogeny was constructed using the minimum evolution and maximum parsimony methods according to their sequence diversity.Second,the water-solution bioactive components and lipid soluble components were tested by HPLC.Then a chemical phylogeny was built using HPLC fingerprint data.Comparing the molecular and chemical phylogenetic trees revealed many similarities.Results DNA barcoding was sequencing based and could therefore provide more accurate results within a shorter time especially in large-scale studies.Conclusion The results show that ITS2 region is a novel DNA barcode for the authentication of the species in Salvia L.This is the first work to show the relationship between DNA barcoding and chemical components.

Identification of Common Edible and Medicinal Mushrooms by DNA Barcoding

Objective:To identify common edible and medicinal mushrooms using the internal transcribed spacer(ITS) region.Method:A total of eighty-five samples belonging to forty-three species were used in this study.ITS regions were amplified and sequenced.All ITS regions were analyzed using BLAST and MEGA 5.10 methods.Results:Forty species were identified correctly via BLAST method whereas forty-one species were distinguished via Neighbor joining(NJ)tree.All the species could be distinguished by BLAST combined with NJ tree and ITS sequence characteristics.Conclusion:Forty-three common species of edible and medicinal mushrooms were effectively identified using DNA barcoding technology based on ITS region.

DNA-Barcoding of Some Medicinal Plant Species in Saudi Arabia Using rbcL and matK Genes

In the Kingdom of Saudi Arabia(KSA),thousands of plants are considered to have therapeutic value.The ambiguous use of identification mainly morphological characteristics of many plants has resulted in the adulteration and displacement of plant products which undermine their therapeutic value and weak documentation of plant resources.The aims of this study were therefore to evaluate genetic variability and explore the phylogeographic architecture for Saudi medicinal plant samples using rbcL and matK genes as barcodes for genomic identification.The matK and rbcL sequences collected for these samples were used as key markers for examining the relationship between Saudi medicinal plant species based on genetic diversity.During our study we were successful in identifying and documenting 4 different species(Foeniculum vulgare,Nitraria retusa,Dodonaea viscosa,and Rumex nervosus)located in Saudi Arabia using DNA barcoding technique.A total number of 8 sequences were obtained with a total sequence length of 6176 bp,where it ranged from 617 bp to 878 bp with an aver-age length of 772 bp.The total number of rbcL sequences length is 2801 bp,where it ranges from 617 bp to 807 bp with an average length of 700.2 bp.Out of the 4 plant samples used,only three samples were identified correctly on the species level with an identity percentage higher than 95%using rbcL gene.Additionally,4 matK sequences have been retrieved belong to 4 species.The total number of matK sequences length is 3375 bp,where it ranges from 819 bp to 878 bp with an average length of 843.8 bp.Out of the 4 plant samples used,only two samples were identified correctly on the species level with an identity percentage higher than 98% using matK gene.Both rbcL and matK have been able to identify most of our collected plant samples by genus,and some by species.Using only one DNA-barcoding technique was not reliable for plant identification,where matK and rbcL must be used as a dual DNA-barcoding procedure.

Testing complete plastomes and nuclear ribosomal DNA sequences for species identification in a taxonomically difficult bamboo genus Fargesia

Fargesia,the largest genus within the temperate bamboo tribe Arundinarieae,has more than 90 species mainly distributed in the mountains of Southwest China.The Fargesia bamboos are important components of the subalpine forest ecosystems that provide food and habitat for many endangered animals,including the giant panda.However,species-level identification of Fargesia is difficult.Moreover,the rapid radiation and slow molecular evolutionary rate of Fargesia pose a significant challenge to using DNA barcoding with standard plant barcodes(rbcL,matK,and ITS) in bamboos.With progress in the sequencing technologies,complete plastid genomes(plastomes) and nuclear ribosomal DNA(nrDNA)sequences have been proposed as organelle barcodes for species identification;however,these have not been tested in bamboos.We collected 196 individuals representing 62 species of Fargesia to comprehensively evaluate the discriminatory power of plastomes and nrDNA sequences compared to standard barcodes.Our analysis indicates that complete plastomes have substantially higher discriminatory power(28.6%) than standard barcodes(5.7%),whereas nrDNA sequences show a moderate improvement(65.4%) compared to ITS(47.2%).We also found that nuclear markers performed better than plastid markers,and ITS alone had higher discriminatory power than complete plastomes.The study also demonstrated that plastomes and nrDNA sequences can contribute to intrageneric phylogenetic resolution in Fargesia.However,neither of these sequences were able to discriminate all the sampled species,and therefore,more nuclear markers need to be identified.

Pharmacognostical s tudies o n Spermacoce p usilla Wallich

Background:Spermacoce pusilla Wallich(S.pusilla)is widely used in Guangdong province of China.It has valuable medicinal value in treating trauma,fractures,carbuncle,swelling,and poisonous snake bites.Method:The present study was carried out to provide a scientific basis for the identification and authenticity of S.pusilla with the help of pharmacognostical parameters,which has not been done before.In this study,a series of related identification information such as source,character,microscopic observation,physical and chemical reaction,and molecular markers were employed to establish accurate,comprehensive pharmacognostic identification information of S.pusilla.Results:The study provided some basic data regarding the genuine crude drug.The identification efficiency of ITS sequences obtained in this study is high,the psbA-trnH sequence was obtained from S.pusilla and sequenced for the first time in this study.Conclusion:In this study,traditional pharmacognosy identification and molecular marker identification of S.pusilla in the Guangdong region were carried out,and a systematic and comprehensive pharmacognosy identification information system was established.

Dietary analysis of the House Swift(Apus nipalensis)in Hong Kong using prey DNA in faecal samples

Background:To understand the dietary composition of the highly aerial swift(Apodidae),ecologists conventionally depend on the morphological identification of prey items from food boluses or stomach contents,but these techniques are often invasive,require expertise in identification,and often cannot produce accurate identifications at the species level.Methods:DNA barcoding was used to analyse the dietary composition of House Swifts(Apus nipalensis)in Hong Kong,China.Faecal samples from five different colonial nest sites were collected between 2019 and 2020.We used universal primers to amplify a region of the cytochrome C oxidase gene from prey DNA in the faecal samples for identification purposes.Results:Ten different orders and 44 families from three different classes of Arthropoda were identified in the collected faecal samples.Hymenoptera,Hemiptera and Diptera were the most prevalent groups of prey found in the samples.Differences in the dietary composition of House Swifts during the breeding(April to September)and nonbreeding(October to March)season were also found.Hymenoptera,particularly ants(Formicidae),were predominant in the diet during the breeding season,whereas Diptera and Hemiptera were predominant during the non-breeding season.Conclusion:The prey groups identified in this study were similar to those identified in a previous study of the diet of House Swift,which also suggests a possible role of House Swifts in reducing the numbers of local insect pests.This study demonstrates the usefulness of applying molecular tools for the dietary analysis of aerial feeders.Conserving local forested areas may be crucial for the maintenance of House Swift population.

Screening of DNA Barcode of Quarantine Phytophthora

Phytophtkora is genus of plant-damaging Oomycetes,whose member species are capable of causing enormous economic losses on crops worldwide.In the present study,four candidate genes ITS,CO1,EF-1α and β-tubulin were tested using 123 strains of 80 species of Phytophthora to investigate the feasibility of serving as DNA barcoding markers.The results showed that among the four candidate genes,ITS and CO1 had the highest success rate of PCR amplification and sequencing,up to 100%and 96.7%.There were obvious barcode gaps in ITS,CO1 and β-tubulin,but their frequency distributions of intra-and interspecific genetic distances were slightly overlapped.Wilcoxon rank sum test on intraspecific genetic distances of the four genes showed ITS = CO1 = β-tubulin = EF-1α,indicating they had the same effect on intraspecific discrimination,while the test on interspecific genetic distances of the four genes showed ITS > CO1 > β-tubulin > EF-1α.In summary,ITS and CO1 should be used in combination as the primary barcodes,β-tubulin as the complementary barcode for the identification of 11 quarantine Phytophthora species.