Journal of Integrative Agriculture

The breakfast imperative: The changing context of global food security

YE Li-ming, Jean-Paul Malingreau, TANG Hua-jun, Eric Van Ranst

2016, 15(06): 1179-1185. DOI: 10.1016/S2095-3119(15)61296-5

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The power of microsatellite markers and AFLPs in revealing the genetic diversity of Hashemi aromatic rice from Iran

Alireza Tarang, Anahita Bakhshizadeh Gashti

2016, 15(06): 1186-1197. DOI: 10.1016/S2095-3119(15)61221-7

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Hashemi, a popular aromatic rice among Iranians, is famous for its fragrance and taste. Such features are major reasons for its higher price compared to non-aromatic varieties available in Iran. Therefore, the knowledge of genetic diversity of this profitable crop is a fundamental ineterst for plant breeders in future breeding programs. In the present research, genetic diversity among 35 genotypes of Hashemi aromatic rice (Oryza sativa L.) from Guilan and Mazandaran provinces of Iran was estimated using simple sequence repeat (SSR) and amplified fragment length polymorphism (AFLP) markers. Out of 21 SSR and 12 EcoRI-MseI AFLP marker combinations, only 16 SSRs and 10 AFLPs exhibited polymorphic patterns while others were monomorphic. The 10 AFLP primer combinations produced a total of 142 of bands and 20 were polymorphic (14.08%). Moreover, 40 out of 47 bands amplified with 16 SSR markers showed polymorphism (85.1%). The average number of alleles identified by SSR primers was 2.56 alleles per locus with a range of 2 to 4. The average value of polymorphic information content (PIC) was 0.393 and 0.468 for AFLP and SSR markers, respectively. However, the genetic similarity values ranged from 0.26 to 1 for SSRs and 0.21 to 1 for AFLPs. Later, a unweighted pair group method with arithmetic mean (UPGMA) dendrogram was generated and genotypes were clustered into four groups with SSRs at similarity coefficient of 0.55 while AFLPs clustered them into six groups at similarity coefficient of 0.41. Cluster analysis revealed a narrow genetic diversity and low correlation between geographical differentiation and genetic distance within cultivars.

Polymorphism and association analysis of a drought-resistant gene TaLTP-s in wheat

LI Qian, WANG Jing-yi, Nadia Khan, CHANG Xiao-ping, LIU Hui-min, JING Rui-lian

2016, 15(06): 1198-1206. DOI: 10.1016/S2095-3119(15)61189-3

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Lipid transfer protein (LTP) is a kind of small molecular protein, which is named for its ability to transfer lipid between cell membranes. It has been proved that the protein is involved in the responding to abiotic stresses. In this study, TaLTP-s, a genomic sequence of TaLTP was isolated from A genome of wheat (Triticum aestivum L). Sequencing analysis exhibited that there was no diversity in the coding region of TaLTP-s, but seven single nucleotide polymorphisms (SNPs) and 1 bp insertion/deletion (InDel) were detected in the promoter regions of different wheat accessions. Nucleotide diversity (π) in the region was 0.00033, and linkage disequilibrium (LD) extended over almost the entire TaLTP-s region in wheat. The dCAPS markers based on sequence variations in the promoter regions (SNP-207 and SNP-1696) were developed, and three haplotypes were identified based on those markers. Association analysis between the haplotypes and agronomic traits of natural population consisted of 262 accessions showed that three haplotypes of TaLTP-s were significantly associated with plant height (PH). Among the three haplotypes, HapIII is considered as the superior haplotype for increasing plant height in the drought stress environments. The G variance at the position of 207 bp could be a superior allele that significantly increased number of spikes per plant (NSP). The functional marker of TaLTP-s provide a tool for marker-assisted selection regarding to plant height and number of spikelet per plant in wheat.

De novo assembly of Zea nicaraguensis root transcriptome identified 5 261 full-length transcripts

JIANG Wei, LIU Hai-lan, WU Yuan-qi, ZHANG Su-zhi, LIU Jian, LU Yan-li, TANG Qi-lin, RONG Ting-zhao

2016, 15(06): 1207-1217. DOI: 10.1016/S2095-3119(15)61153-4

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Zea nicaraguensis, a wild relative of cultivated maize (Zea mays subsp. mays), is considered to be a valuable germplasm to improve the waterlogging tolerance of cultivated maize. Use of reverse genetic-based gene cloning and function verification to discover waterlogging tolerance genes in Z. nicaraguensis is currently impractical, because little gene sequence information for Z. nicaraguensis is available in public databases. In this study, Z. nicaraguensis seedlings were subjected to simulated waterlogging stress and total RNAs were isolated from roots stressed and non-stressed controls. In total, 80 mol L^–1 Illumina 100-bp paired-end reads were generated. De novo assembly of the reads generated 81 002 final non-redundant contigs, from which 5 261 full-length transcripts were identified. Among these full-length transcripts, 3 169 had at least one Gene Ontology (GO) annotation, 2 354 received cluster of orthologous groups (COG) terms, and 1 992 were assigned a Kyoto encyclopedia of genes and genomes (KEGG) Orthology number. These sequence data represent a valuable resource for identification of Z. nicaraguensis genes involved in waterlogging response.

GGE biplot analysis of yield stability and test location representativeness in proso millet (Panicum miliaceum L.) genotypes

ZHANG Pan-pan, SONG Hui, KE Xi-wang, JIN Xi-jun, YIN Li-hua, LIU Yang, QU Yang, SU Wang, FENG Nai-jie, ZHENG Dian-feng, FENG Bai-li

2016, 15(06): 1218-1227. DOI: 10.1016/S2095-3119(15)61157-1

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The experiments were conducted for three consecutive years across 14 locations using 9 non-waxy proso millet genotypes and 16 locations using 7 waxy proso millet genotypes in China. The objectives of this study were to analyze yield stability and adaptability of proso millets and to evaluate the discrimination and representativeness of locations by analysis of variance (ANOVA) and genotype and genotype by environment interaction (GGE) biplot methods. Grain yields of proso millet genotypes were significantly influenced by environment (E), genotype (G) and their interaction (G×E) (P<0.1%). G×E interaction effect was six times higher than G effect in non-waxy group and seven times in waxy group. N04-339 in non-waxy and Neimi 6 (NM6) in waxy showed higher grain yields and stability compared with other genotypes. Also, Neimi 9 (NM9, a non-waxy cultivar) and 90322-2-33 (a waxy cultivar) showed higher adaptability in 7 and in 11 locations, respectively. For non-waxy, Dalat, Inner Mongolia (E2) and Wuzhai, Shanxi (E5) were the best sites among all the locations for maximizing the variance among candidate cultivars, and Yanchi, Ningxia (E10) had the best representativeness. Wuzhai, Shanxi (e9) and Yanchi, Ningxia (e14) were the best representative locations, and Baicheng, Jilin (e2) was better discriminating location than others for waxy genotypes. Based on our results, E10 and e14 have enhanced efficiency and accuracy for non-waxy genotypes and waxy genotypes selection, respectively in national regional test of proso millet varieties.

Analysis of cytosine methylation in early generations of resynthesized Brassica napus

RAN Li-ping, FANG Ting-ting, RONG Hao, JIANG Jin-jin, FANG Yu-jie, WANG You-ping

2016, 15(06): 1228-1238. DOI: 10.1016/S2095-3119(15)61277-1

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DNA methylation, an important epigenetic modification, serves as a key function in the polyploidization of numerous crops. In this study, early generations of resynthesized Brassica napus (F₁, S₁–S₃), ancestral parents B. rapa and B. oleracea were analyzed to characterize their DNA methylation status during polyploidization, applying DNA methylation-sensitive amplification polymorphism (MSAP) and high-performance liquid chromatography methods. In F₁, 53.4% fragments were inherited from both A- and C-genomes. Besides, 5.04 and 8.87% fragments in F₁were inherited from A- and C- genome, respectively. 5.85 and 0.8% fragments were newly appeared and disappeared in resynthesized B. napus, respectively. 13.1% of these gene sites were identified with methylation changes in F₁, namely, hypermethylation (7.86%) and hypomethylation (5.24%). The lowest methylation status was detected in F₁ (38.7%) compared with in S₁–S₃. In S₃, 40.32% genes were methylated according to MSAP analysis. Sequencing of methylated fragments indicated that genes involved in multiple biological processes were modified, including transcription factors, protein modification, and transporters. Expression ananlysis of DNA methyltransferase 1 and DNA methyltransferase chromomethylase 3 in different materials was consistent to the DNA methylation status. These results can generally facilitate dissection of how DNA methylation contributes to genetic stability and improvement of B. napus during polyploidization.

Phenylalanine ammonia-lyase gene families in cucurbit species: Structure, evolution, and expression

DONG Chun-juan, CAO Ning, ZHANG Zhi-gang, SHANG Qing-mao

2016, 15(06): 1239-1255. DOI: 10.1016/S2095-3119(16)61329-1

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Phenylalanine ammonia-lyase (PAL), the first enzyme of phenylpropanoid pathway, is always encoded by multigene families in plants. In this study, using genome-wide searches, 13 PAL genes in cucumber (CsPAL1–13) and 13 PALs in melon (CmPAL1–13) were identified. In the corresponding genomes, ten of these PAL genes were located in tandem in two clusters, while the others were widely dispersed in different chromosomes as a single copy. The protein sequences of CsPALs and CmPALs shared an overall high identity to each other. In our previous report, 12 PAL genes were identified in watermelon (ClPAL1–12). Thereby, a total of 38 cucurbit PAL members were included. Here, a comprehensive comparison of PAL gene families was performed among three cucurbit plants. The phylogenetic and syntenic analyses placed the cucurbit PALs as 11 CsPAL-CmPAL-ClPAL triples, of which ten triples were clustered into the dicot group, and the remaining one, CsPAL1-CmPAL8-ClPAL2, was grouped with gymnosperm PALs and might serve as an ancestor of cucurbit PALs. By comparing the syntenic relationships and gene structure of these PAL genes, the expansion of cucurbit PAL families might arise from a series of segmental and tandem duplications and intron insertion events. Furthermore, the expression profiling in different tissues suggested that different cucurbit PALs displayed divergent but overlapping expression profiles, and the CsPAL-CmPAL-ClPAL orthologs showed correlative expression patterns among three cucurbit plants. Taken together, this study provided an extensive description on the evolution and expression of cucurbit PAL gene families and might facilitate the further studies for elucidating the functions of PALs in cucurbit plants.

Combining gas exchange and chlorophyll a fluorescence measurements to analyze the photosynthetic activity of drip-irrigated cotton under different soil water deficits

LUO Hong-hai, Tsimilli-michael Merope, ZHANG Ya-li, ZHANG Wang-feng

2016, 15(06): 1256-1266. DOI: 10.1016/S2095-3119(15)61270-9

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Gas exchange and chlorophyll a fluorescence were measured to study the effects of soil water deficit (75, 60 and 45% of field capacity, FC) on the photosynthetic activity of drip-irrigated cotton under field conditions. At light intensities above 1 200 µmol m⁻² s⁻¹, leaf net photosynthetic rate (P_n) at 60 and 45% FC was 0.75 and 0.45 times respectively than that of 75% FC. The chlorophyll content, leaf water potential and yield decreased as soil water deficit decreased. Fiber length was significantly lower at 45% FC than at 75% FC. The actual quantum yield of the photosystem II (PSII) primary photochemistry and the photochemical quenching were significantly greater at 60% FC than at 75% FC. The electron transport rate and non-photochemical quenching at 45% FC were 0.91 and 1.29 times than those at 75% FC, respectively. The amplitudes of the K- and L-bands were higher at 45% FC than at 60% FC. As soil water content decreased, active PSII reaction centers per chlorophyll decreased, functional PSII antenna size increased, and energetic connectivity between PSII units decreased. Electron flow from plastoquinol to the PSI end electron acceptors was significantly lower at 45% FC than at 75% FC. Similar to the effect on leaf P_n, water deficit reduced the performance index (PI_{ABS, total}) in the dark-adapted state. These results suggest that (i) the effect of mild water deficit on photosystem activity was mainly related to processes between plastoquinol and the PSI end electron acceptors, (ii) PSI end electron acceptors were only affected at moderate water deficit, and (iii) PI_{ABS, total} can reliably indicate the effect of water deficit on the energy supply for cotton metabolism.

Accumulated chilling hours during endodormancy impact blooming and fruit shape development in peach (Prunus persica L.)

LI Yong, FANG Wei-chao, ZHU Geng-rui, CAO Ke, CHEN Chang-wen, WANG Xin-wei, WANG Li-rong

2016, 15(06): 1267-1274. DOI: 10.1016/S2095-3119(16)61374-6

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Winter chill is essential for the growth and development of deciduous species. To understand the relationship between accumulated chilling hours during endodormancy and blooming and fruit shape development, we controlled chilling hours and investigated their effects on blooming date and fruit shape of peaches. The results showed that the number of days to full bloom date and the heat requirement for blooming were negatively correlated with accumulated chilling hours. Accumulated chilling hours were significantly negatively correlated with fruit shape index and fruit tip lengths, suggesting that the number of chilling hours affect the fruit shape development. Fewer accumulated chilling hours may be the major reason for longer fruit shape and protruding fruit tips. In conclusion, our results indicate specifically that decreased winter chilling hours can delay the bloom date and may lead to aberrant fruit shape development in peaches. Our study provides preliminary insights into the response of temperate fruit species to global climate change.

Proteome analysis of alfalfa roots in response to water deficit stress

Rahman Md Atikur, Kim Yong-Goo, Alam Iftekhar, LIU Gong-she, Lee Hyoshin, Lee Jeung Joo, Lee Byung-Hyun

2016, 15(06): 1275-1285. DOI: 10.1016/S2095-3119(15)61255-2

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To evaluate the response of alfalfa to water deficit (WD) stress, WD-induced candidates were investigated through a proteomic approach. Alfalfa seedlings were exposed to WD stress for 12 and 15 days respectively, followed by 3 days re-watering. Water deficit increased H₂O₂ content, lipid peroxidation, DPPH (1,1-diphenyl-2-picrylhydrazyl)-radical scavenging activity, and the free proline level in alfalfa roots. Root proteins were extracted and separated by two-dimentional polyacrylamide gel electrophoresis (2-DE). A total of 49 WD-responsive proteins were identified in alfalfa roots; 25 proteins were reproducibly found to be up-regulated and 24 were down-regulated. Two proteins, namely cytosolic ascorbate peroxidase (APx2) and putative F-box protein were newly detected on 2-DE maps of WD-treated plants. We identified several proteins including agamous-like 65, albumin b-32, inward rectifying potassium channel, and auxin-independent growth promoter. The identified proteins are involved in a variety of cellular functions including calcium signaling, abacisic acid (ABA) biosynthesis, reactive oxygen species (ROS) regulation, transcription/ translation, antioxidant/detoxification/stress defense, energy metabolism, signal transduction, and storage. These results indicate the potential candidates were responsible for adaptive response in alfalfa roots.

The influence of potassium to mineral fertilizers on the maize health

Jan Bocianowski, Piotr Szulc, Anna Tratwal, Kamila Nowosad, Dariusz Piesik

2016, 15(06): 1286-1292. DOI: 10.1016/S2095-3119(15)61194-7

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Field experiments (2009–2011) were conducted at the Department of Agronomy at Poznań University of Life Sciences on the fields of the Research Institute in Swadzim. We evaluated the health of maize plants of two types, depending on the variations in mineral fertilization. The conducted research recorded the occurrence of pests such as oscinella frit (Oscinella frit L.) and the European corn borer (Pyrausta nubilalis Hbn.). Diseases recorded during the research included two pathogenes: Fusarium (Fusarium ssp.) and corn smut (Ustilago maydis Corda). It was shown that the meteorological conditions during the maize vegetation had a significant influence on the occurrence of pests. Adding potassium to mineral fertilizers increased the maize resistance to Fusarium. Cultivation of “stay-green” cultivar shall be considered as an element of integrated maize protection. The occurrence of oscinella frit was correlated with the occurrence of Fusarium as well as the occurrence of the European corn borer for both examined cultivars.

An improved method for RNA extraction from urediniospores of and wheat leaves infected by an obligate fungal pathogen, Puccinia striiformis f. sp. tritici

MA Li-Jie, QIAO Jia-xing, KONG Xin-yu, WANG Jun-juan, XU Xiang-ming, HU Xiao-ping

2016, 15(06): 1293-1303. DOI: 10.1016/S2095-3119(15)61250-3

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Stripe rust, caused by Puccinia striiformis f. sp. tritici, is an important wheat disease in China, seriously threatening wheat production. Understanding the winter survival of the fungus is a key for predicting the spring epidemics of the disease, which determines the crop loss. Estimation of P. striiformis f. sp. tritici winter survival requires processing a large number of samples for sensitive detection of the pathogen in wheat leaf tissue using real-time quantitative reverse transcription PCR (qRT-PCR). A bottleneck for the analysis is the acquisition of a good yield of high quality RNA suitable for qRT-PCR to distinguish dead and alive fungal hyphae inside leaves. Although several methods have been described in the literatures and commercial kits are available for RNA extraction, these methods are mostly too complicated, expensive and inefficient. Thus, we modified three previously reported RNA extraction methods with common and low-cost reagents (LiCl, SDS and NaCl) to solve the problems and selected the best to obtain high quality and quantity RNA for use in qRT-PCR. In the three improved methods, the NaCl method was proven to be the best for extracting RNA from urediniospores of and wheat leaves infected by P. striiformis f. sp. tritici, although the modified LiCl and SDS methods also increased yield of RNA compared to the previous methods. The improved NaCl method has the following advantages: 1) Complete transfer of urediniospores of P. striiformis f. sp. tritici from the mortar and pestle can ensure the initial amount of RNA for the qRT-PCR analysis; 2) the use of low-cost NaCl to replace more expensive Trizol can reduce the cost; 3) the yield and quality of RNA can be increased; 4) the improved method is more suitable for a large number and high quantity of samples from fields. Using the improved NaCl method, the amount of RNA was increased three times from urediniospores of P. striiformis f. sp. tritici compared from the extraction kit. Approximately, 10.11 μg total RNA of high quality was obtained from 100 mg of infected leaves, which was 8.8, 6.5, 3.4 and 2.1 folds of the amounts obtained from the previous LiCl, SDS, NaCl and traditional Trizol methods, respectively. The method could be used to study the overwintering rates of P. striiformis f. sp. tritici over a large region of wheat production for predicting epidemic levels by determining pathogen survival levels after winter. The method can also be used in any studies which need a large number of high quality RNA samples.

The EPSPS Pro106Ser substitution solely accounts for glyphosate resistance in a goosegrass (Eleusine indica) population from Tennessee, United States

Janel L Huffman, Chance W Riggins, Lawrence E Steckel, Patrick J Tranel

2016, 15(06): 1304-1312. DOI: 10.1016/S2095-3119(15)61220-5

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Previous studies have documented the occurrence of glyphosate-resistant (GR) goosegrass (Eleusine indica (L.) Gaertn.) and, in at least some cases, resistance is due to an altered target site. Research was performed to determine if an altered target site was responsible for GR in a Tennessee, United States goosegrass population (TennGR). DNA sequencing revealed a mutation in TennGR plants conferring the Pro106Ser 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) substitution previously identified in other GR populations. F₂ populations were derived from TennGR plants crossed with plants from a glyphosate-susceptible population (TennGS) and analyzed for their response to glyphosate and genotyped at the EPSPS locus. Plants from the F₂ populations segregated 1:2:1 sensitive:intermediate:resistant in response to a selective dose of glyphosate, and these responses co-segregated with the EPSPS genotypes (PP106, PS106, and SS106). To separately investigate the effect of the Pro106Ser substitution on GR, glyphosate dose-response curves and 50% effective dose (ED₅₀) values were compared among the three genotypes and the two parental populations. The SS106 genotype was 3.4-fold resistant relative to the PP106 genotype, identical to the resistance level obtained when comparing the resistant and susceptible parental populations. We conclude that the mutation conferring a Pro106Ser EPSPS mutation is solely responsible for GR in the TennGR goosegrass population.

The production relationship of destruxins and blastospores of Metarhizium anisopliae with virulence against Plutella xylostella

DONG Ting-yan, ZHANG Bo-wen, WENG Qun-fang, HU Qiong-bo

2016, 15(06): 1313-1320. DOI: 10.1016/S2095-3119(15)61322-3

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Metarhizium anisopliae as an essential entomopathogenic fungus has been known to produce destruxins (a kind of cyclo-peptidic mycotoxins) and blastospores in submerged culture. Blastospores and destruxins are candidates for insecticides, but the relations of both productions and the impact factors are unclear yet. In this study, we investigated the effects of inoculums, rotation, dissolved oxygen (DO) on the productions of blastospores and destruxins A and B (DA and DB) in submerged culture of M. anisopliae strain MaQ01. The results indicated that DO levels were regulated by inoculum amounts and rotation speeds, meanwhile, the productions of DA, DB and blastospores were also closely influenced by those factors. Totally, when DO value was more than 40%, the higher productions of destruxins and blastospores were achieved, by contrast, lower than 40% of DO values resulted in lower productions. The regression analysis suggested that the productions of DA, DB and blastospores were positively correlated with the DO levels. Meanwhile, the positive correlations between the productions of DA or DB and blastospores were also found. Briefly, when the rotation is 150 r min–1 and the inoculum is 1.0×10⁶ spore mL^–1, the DA, DB and blastospores achieved the best production of 61.81 mg mL^–1, 24.74 mg mL^–1 and 5.73×10⁶ spore mL^–1, respectively. In addition, the pathogenicities of blastospores and conidia against Plutella xylostella were bioassayed. The higher mortalities of P. xylostella were totally recorded in blastospore treatments than in conidia treatments, especially in lower dosages and earlier periods. Our research will give some new insights to production of destruxins and blastospores by using M. anisopliae.

Porcine LEM domain-containing 3: Molecular cloning, functional characterization, and polymorphism associated with ear size

LIANG Jing, LI Na, ZHANG Long-chao, WANG Li-gang, LIU Xin, ZHAO Ke-bin, YAN Hua, PU Lei, ZHANG Yue-bo, SHI Hui-bi, ZHANG Qin, WANG Li-xian

2016, 15(06): 1321-1229. DOI: 10.1016/S2095-3119(15)61173-X

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Ear size exhibits remarkable diversity in pig breeds. LEM domain-containing 3 (LEMD3) on chromosome 5 is considered as an important candidate for porcine ear size. This is the first study on cloning and characterization of LEMD3 cDNA. The complete cDNA contains 4 843 bp, including a 2 736-bp open reading frame (ORF), a 37-bp 5´-untranslated region (UTR) and a 2 070-bp 3´-UTR. The complete LEMD3 gene is 126 241-bp and contains 13 exons and 12 introns. The ORF encodes a deduced LEMD3 protein of 911 amino acids, which shares 82–94% nucleic acid and 51–96% amino acid identity with other species. A phylogenetic tree constructed based on the amino acid sequences revealed that the porcine LEMD3 protein was closely related with cattle LEMD3. Resequencing of the ORF and promoter of LEMD3 from Minzhu pig and Large White revealed three single nucleotide polymorphisms (SNPs): L964C>A in the complete coding region, L4625A>G in the 3´ UTR, and L-394T>C in the promoter region. Genome-wide association study (GWAS) revealed that all of SNPs were shown significant association with ear size in Large White×Minzhu pig intercross population. With conditional GWAS, –log₁₀(P-value) decreased by more than 80% when each of three SNPs was included as a fixed effect. These results suggested direct involvement of LEMD3 or close linkage to the causative mutation for ear size. The findings of this study might form the basis for understanding the genetic mechanism of ear size variation in pigs and provide potential molecular markers for screening ear size diversity in pig breeds.

Modified Bfat-1 gene and its biological verification in mice by hydrodynamic tail vein injection

GAO Xue, DU Xin-hua, ZHANG Lu-pei, CHEN Yan, GAO Hui-jiang, XU Shang-zhong, LI Jun-ya

2016, 15(06): 1330-1337. DOI: 10.1016/S2095-3119(15)61218-7

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Omega-3 polyunsaturated fatty acids (ω-3 PUFAs) are essential components required for normal cellular function and have been shown to have important therapeutic and nutritional benefits in humans. But humans or mammals cannot naturally produce ω-3 PUFAs, due to the lack of the ω-3 fatty acid desaturase gene (fat-1 gene). Previously, fat-1 gene has been cloned from Caenorhabditis elegans and transferred into mice, pigs and sheep, but not yet into beef cattle. We attempt to transfer it into beef cattle. The object of this paper is to edit the fat-1 gene from C. elegans to express more efficiently in beef cattle and verify its biological function in mice model. As a result, the fat-1 gene from C. elegans was modified by synonymous codon usage and named it Bfat-1. We have demonstrated that degree of codon bias of Bfat-1 gene was increased in beef cattle. Moreover, Bfat-1 gene could be transiently expressed in mouse liver and muscle, the ω-6/ω-3 PUFAs ratio of 18 and 20 carbon was decreased significantly in liver (P<0.05), and the ratio of 20 carbon decreased significantly in muscle 24 and 72 h after injection (P<0.05). This confirms that the Bfat-1 gene modification was successful, and the protein encoded was able to catalyze the conversion of ω-6 PUFAs to ω-3 PUFAs.

Inducible overexpression of porcine homeobox A10 in the endometrium of transgenic mice

LIN Rui-yi, WU Di, ZHAO Chang-zhi, CHEN Shang-shang, XIAO Qian, LI Xin-yun, ZHAO Shu-hong

2016, 15(06): 1338-1344. DOI: 10.1016/S2095-3119(15)61169-8

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Homeobox A10 (HOXA10) is a well-known transcription factor that plays an important role in directing endometrial differentiation and establishing the conditions required for implantation. Interestingly, the expression level of HOXA10 may be associated with litter size. To study the effects of the porcine HOXA10 promoter fragment on the expression of HOXA10 gene in vivo, we generated a transgenic mouse model using pronuclear microinjection, and measured the expression of HOXA10 in the endometrium. There was no difference in the expression level of HOXA10 between transgenic and wild-type mice in the absence of hormone stimulation. However, following treatment with progesterone and estradiol benzoate, the expression level of HOXA10 was significantly increased in transgenic mice compared with that of wild-type mice. Furthermore, the litter size of transgenic females was larger than that of wild-type females (7.02±1.73 vs. 6.48±1.85; P=0.14). Moreover, the difference of litter size was greater in the later parities (7.33±1.62 vs. 6.37±2.02; P=0.08) compared with the first parity (6.76±1.81 vs. 6.61±1.67; P=0.77) between transgenic and wild-type mice. Therefore, our transgenic mouse model provides exciting insights regarding the actions of HOXA10 and its hormone-inducible promoter in vivo. The present study offers valuable proof of principle to develop transgenic pigs with a hormone-inducible promoter regulating HOXA10 to alter litter size.

Genomic characterization and antimicrobial susceptibility of bovine intrauterine Escherichia coli and its relationship with postpartum uterine infections

YANG Li-ming, WANG Yi-hao, PENG Yu, MIN Jiang-tao, HANG Su-qin, ZHU Wei-yun

2016, 15(06): 1345-1354. DOI: 10.1016/S2095-3119(15)61170-4

Abstract ( ) PDF in ScienceDirect

To investigate the roles of Escherichia coli in the pathogenesis of postpartum uterine diseases in dairy cows, a total of 145 E. coli isolates were recovered from 18 healthy cows (61 isolates) and 25 cows with clinical endometritis (84 isolates) at 25–35 days after parturition. Genomic characteristics including phylogenetic grouping, genetic diversity and virulence genes of E. coli isolates were screened to profile the characteristics related to uterine infections. The susceptibility of the bacteria against 23 antibiotics was also evaluated to support prevention and treatment of clinical cases. Genetic diversity of E. coli identified by random amplification of polymorphic DNA (RAPD) revealed 103 clonal types, including 3 common types to unaffected cows and endometritis cows, 39 types specific to healthy cows and 61 types in endometritis subjects. In addition, the isolates from endometritis uteri showed more genetic variability compared with that of healthy cows. According to the findings of phylogenetic grouping, the E. coli isolates were assigned to group A (35.9%), B1 (59.3%) and D (4.8%). The expression of 10 of 20 virulence gens were detected positively, and only fimH gene was revealed significantly (P<0.05) associated with endometritis. From antimicrobial susceptibility test, E. coli was found highly resistant to tetracycline, ampicillin, carbenicillin and amoxicillin, but sensitive to amikacin, netilmicin, tobramycin, cefepime and ceftazidime. In conclusion, E. coli were extensively observed in both healthy and endometritis cows, and presented a large clonal types, however, fimH was the only gene observed associated with clinical endometritis. Our results suggest that the drugs like amikacin, netilmicin, tobramycin and cefepime could be considered for preventing and treating clinical endometritis in the practical management of dairy cow.

Effect of feeding transgenic cry1Ab/cry1Ac rice on indices of immune function in broilers

LIU Ran-ran, ZHAO Gui-ping, ZHENG Mai-qing, LIU Jie, ZHANG Jing-jing, LI Peng, LI Qing-he, FENG Jing-hai, ZHANG Min-hong, WEN Jie

2016, 15(06): 1355-1363. DOI: 10.1016/S2095-3119(15)61281-3

Abstract ( ) PDF in ScienceDirect

The safety of feeding transgenic cry1Ab/cry1Ac rice (a genetically modified (GM) rice variety) to broilers was examined from an immunological perspective. Hatchling Arbor Acres chickens (240) were assigned to two dietary treatments (diets containing GM or non-GM rice) with 12 replicates per group and 10 birds per replicate. Traits were measured on one randomly selected bird from each replicate at d 21 and 42. The 42-d feeding trial revealed that cry1Ab/cry1Ac rice had no significant effect relative to non-GM rice on body weight and the immune organ indices. No significant pathological lesion in the spleen and bursa of Fabricius was found in the GM rice group. There were no significant differences in serum concentrations of immunoglobulin Y (IgY), IgM, interleukin 4 (IL-4) and IL-6 between the two groups at d 21 or 42, except for IL-6 being higher (P<0.05) in the GM-fed chickens at d 42. There were no differences in the T and B lymphocyte transformation rate and CD4+/CD8+ ratio between the two groups at d 42. Additionally, there was no significant difference between the two diets in expression of relevant genes viz. the major histocompatibility complex class II beta chain (BLB2), interferon beta 1 (IFNβ), tumour necrosis factor alpha-like (TNFα) and toll-like receptor 4 (TLR4) in the spleen and bursa of Fabricius. All the data demonstrated that transgenic cry1Ab/cry1Ac rice had no adverse effect on these aspects of immune function of broilers during 42-d feeding trial. Transgenic rice was therefore indistinguishable from non-GM rice in terms of short-term feeding in chickens.

Impact of agricultural intensification on soil organic carbon: A study using DNDC in Huantai County, Shandong Province, China

LIAO Yan, WU Wen-liang, MENG Fan-qiao, LI Hu

2016, 15(06): 1364-1375. DOI: 10.1016/S2095-3119(15)61269-2

Abstract ( ) PDF in ScienceDirect

Using the biogeochemical model denitrification/decomposition (DNDC), the dynamic changes of soil organic carbon (SOC) of farmland from the 1980s to 2030s were investigated in Huantai County, a typical intensive agricultural region in the Huang-Huai-Hai Plain of China. Prior to modelling, validation of the DNDC model against field data sets of SOC from Quzhou Experimental Station in the Huang-Huai-Hai Plain was conducted at the site scale. We compared the simulated results with the observed SOC in Huantai County during 1982–2011 under two different classification methods of simulation unit (the first method integrated soil type and land use of Huantai County to form the overlapped modeling units; the second selected the 11 administrative towns as the modeling units), and achieved a high accuracy in the model simulation with the improvement of the model parameters. Regional SOC (0–20 cm) density and stocks for Huantai County in the years 2012–2031 were predicted under different scenarios of farming management. Compared with current management practices, optimized fertilization (20% decrease of mineral N), crop straw incorporation (90%) and appropriate animal manure input (40 kg N ha^–1 yr^–1) could achieve the highest level of SOC density (56.8% higher than 2011) in the period of 2012–2031. The research highlighted the importance of crop straw incorporation, optimized N fertilization and integration of crop production with animal husbandry on the farmland carbon sequestration for maintaining a high land productivity in the Huang-Huai-Hai Plain.

Responses of soil microbial respiration to plantations depend on soil properties in subtropical China

ZHANG Yan-jie, YAN Yue, FU Xiang-ping, YANG Jie, ZHANG Su-yan, XU Shan, TANG Zheng, LI Zhong-fang, LU Shun-bao

2016, 15(06): 1376-1384. DOI: 10.1016/S2095-3119(15)61222-9

Abstract ( ) PDF in ScienceDirect

Assessing the impact of plantation on microbial respiration (MR) is vitally important to understand the interactions between belowground metabolism and land use change. In this study, cumulative MR was determined by alkali absorption method in 1, 3, 7, 14, 21, 28, 35, 42, 49, and 56 days from the soil in a representative plantations in the subtropical region of China. The treatment of plantations contained no plant (CK), orange trees (Citrus reticulata)+Bahia grass (Paspalum notatum) (GB), orange trees (C. reticulata)+Bahia grass (P. notatum)+soybean (Giycine max (L.) Merrill) (GBH). Results showed that plantation had significant effects on microbial respiration and the responses of microbial respiration to plantation from different soil layers and topographies were different: in 0–20 cm in uphill: GB>GBH>CK; in 20–40 cm in uphill: GBH>CK>GB; in 0–20 cm in downhill: GBH>CK>GB; in 20–40 cm in downhill: GB>CK>GBH. Furthermore, plantation also altered the relationships between MR and soil properties. In CK, microbial respiration was positively correlated with NH₄⁺ and soil total N, and negatively correlated with soil moisture, pH, NO₃^–, and microbial biomass carbon (MBC). In GB, microbial respiration under GB significantly negatively correlated with dissolved organic carbon (DOC). In GBH, microbial respiration under GBH was positively correlated with NH₄⁺, MBC, total soil carbon (TC), and total soil nitrogen (TN), and negatively correlated with soil moisture (SM), pH, NO₃^–, and DOC. The underlying mechanisms could be attributed to soil heterogeneity and the effects of plantation on soil properties. Our results also showed that plantation significantly increased soil C storage, which suggested plantation is a key measure to enhance soil C sequestration and mitigate global CO₂ emission, especially for the soil with low initial soil carbon content or bared soil.

Rice (Oryza sativa L.) nutrient management using mycorrhizal fungi and endophytic Herbaspirillum seropedicae

H Hoseinzade, M R Ardakani, A Shahdi, H Asadi Rahmani, G Noormohammadi, M Miransari

2016, 15(06): 1385-1394. DOI: 10.1016/S2095-3119(15)61241-2

Abstract ( ) PDF in ScienceDirect

Integrated nutrient management with biological and chemical fertilizers can improve rice (Oryza sativa L.) productivity, bio-fortification, soil health and fertility. Accordingly, this study was planned to evaluate the combined effects of biological fertilizers including arbuscular mycorrhizal (AM) fungi (Glomus mosseae) and free-living nitrogen-fixing bacteria (Herbaspirillum seropedicae), as well as chemical fertilizers on the yield and nutrient contents of wetland rice under field conditions. Seedlings were inoculated with AM fungi and the bacteria in the nursery and were then transplanted to the field. The experiment was carried out as a split factorial design with three replicates. Treatments included three rates of nitrogen (N1, N2 and N3) and phosphorous (P1, P2 and P3) fertilizers (100, 75 and 50% of the optimum level) in the main plots and mycorrhizal and bacterial treatments in the sub plots. The total of urea (g) used per plot was equal to N1=200, N2=150 and N3=100 at three different growth stages (seeding, tillering and heading) and the total of P (g) per plot used once at seeding using triple super phosphate including P1=16, P2=13 and P3=10. Plant growth and yield as well as the concentration of nitrogen (N), phosphorous (P), potassium (K), iron (Fe), and zinc (Zn) were measured in the soil, straw and grains. N-fertilizer and biological fertilizers had significant effects on root, shoot and grain yield of rice, however, P-fertilizer just significantly affected root and shoot dry weights. Interestingly, analyses of variance indicated that biological fertilization significantly affected all the experimental treatments except straw N. AM fungi, N1 and P1 resulted in the highest rate of rice growth and yield. The interactions of chemical and biological fertilization resulted in significant effects on grain Zn, Fe, P, and N as well as soil Fe, K and N. The highest rate of grain nutrient uptake was resulted by the combined use of biological fertilization and the medium level of chemical fertilization. Interestingly, with decreasing the rate of chemical N fertilization, rice nutrient use efficiency increased indicating how biological fertilization can be efficient in providing plants with its essential nutrients such as N. However, the highest rate of soil and straw nutrient concentration was related to the combined use of biological fertilization and the highest rate of chemical fertilization. We conclude that biological fertilizer, (mycorrhizal fungi and H. seropedicae) can significantly improve wetland rice growth and yield (resulting in the decreased rate of chemical fertilizer), especially if combined with appropriate rate of chemical fertilization, by enhancing nutrient uptake (fortification) and root growth.

Comparative short-term effects of sewage sludge and its biochar on soil properties, maize growth and uptake of nutrients on a tropical clay soil in Zimbabwe

Willis Gwenzi, Moreblessing Muzava, Farai Mapanda, Tonny P Tauro

2016, 15(06): 1395-1406. DOI: 10.1016/S2095-3119(15)61154-6

Abstract ( ) PDF in ScienceDirect

Soil application of biochar from sewage could potentially enhance carbon sequestration and close urban nutrient balances. In sub-Saharan Africa, comparative studies investigating plant growth effect and nutrients uptake on tropical soils amended with sewage sludge and its biochar are very limited. A pot experiment was conducted to investigate the effects of sewage sludge and its biochar on soil chemical properties, maize nutrient and heavy metal uptake, growth and biomass partitioning on a tropical clayey soil. The study compared three organic amendments; sewage sludge (SS), sludge biochar (SB) and their combination (SS+SB) to the unamended control and inorganic fertilizers. Organic amendments were applied at a rate of 15 t ha^–1 for SS and SB, and 7.5 t ha^–1 each for SS and SB. Maize growth, biomass production and nutrient uptake were significantly improved in biochar and sewage sludge amendments compared to the unamended control. Comparable results were observed with F, SS and SS+SB on maize growth at 49 d of sowing. Maize growth for SB, SS, SS+SB and F increased by 42, 53, 47, and 49%, respectively compared to the unamended control. Total biomass for SB, SS, SS+SB, and F increased by 270, 428, 329, and 429%, respectively compared with the unamended control. Biochar amendments reduced Pb, Cu and Zn uptakes by about 22% compared with sludge alone treatment in maize plants. However, there is need for future research based on the current pot experiment to determine whether the same results can be produced under field conditions.

Effect of maize sowing area changes on agricultural water consumption from 2000 to 2010 in the West Liaohe Plain, China

YANG Ling, YANG Yan-zhao, FENG Zhi-ming, ZHENG Ya-nan

2016, 15(06): 1407-1416. DOI: 10.1016/S2095-3119(15)61185-6

Abstract ( ) PDF in ScienceDirect

The West Liaohe Plain is located in the eastern Inner Mongolia, known as the golden maize belt in China, where maize acreage has continued to rise in recent years. Water is the main limiting factor for maize production in the region, therefore, this study calculated the effect of maize sowing area changes on agricultural water consumption in the West Liaohe Plain in 2000, 2005 and 2010, based on remote sensing and meteorological data. Maize remote sensing classification was constructed based on moderate-resolution imaging spectroradiometer, normalized difference vegatation index (MODIS NDVI) data. Then the maize sown area and water requirement and irrigation water resources were investigated. Finally, the effect of the maize sowing area changes on agricultural water consumption in the West Liaohe Plain was systematically analyzed in 2000, 2005, and 2010. The results showed that maize sown areas rose from 2000 to 2010 and were concentrated in the center of the West Liaohe Plain. Average per unit maize water deficit amount also increased in an uneven distribution, increasing from the south, east and north to the center and west of the West Liaohe Plain. The per unit area maize water deficit increased from 2000 to 2010, and reached 266 mm in 2000, 272 mm in 2005 and 273 mm in 2010, respectively. And the study concluded that water deficit during the whole growth period of maize in the West Liaohe Plain was defined by a single peak curve. The maize water requirements increased with maize sowing area changes from 2000 to 2010, and the maize water requirements increased from 0.89 billion m³ in 2000 to 1.19 billion m³ in 2005, and 1.21 billion m³ in 2010.

In vitro establishment of a highly effective method of castor bean (Ricinus communis L.) regeneration using shoot explants

ZHANG Ji-xing, WANG Xiao-yu, FENG Zi-zhou, GENG Xue-jun, MU Sha-moli, HUO Hong-yan, TONG Huan, LI Meng-zhu, LI Yi, CHI Yue, CHEN Yong-sheng

2016, 15(06): 1417-1422. DOI: 10.1016/S1671-2927(00)10558

Abstract ( ) PDF in ScienceDirect

An efficient plant regeneration protocol was established for castor bean (Ricinus communis L.), in which 0.3 mg L^–1 thidiazuron (TDZ) induced shoot clusters and increased the number of adventitious shoots from hypocotyl tissue. Our results showed that treatment under dark conditions significantly promoted the average number of shoots per explant to 37.36±4.54 (with a 6-d treatment). Modified 1/2 Murashige and Skoog (MS) basal medium supplemented with 440 mg L^–1 Ca²⁺, 0.2 mg L^–1 gibberellic acid and 0.1 mg L^–1 TDZ significantly increased shoot elongation rates and lowered vitrification rates. Furthermore, 1/2 MS media supplemented with 0.2 mg L^–1 1-naphthaleneacetic acid induced a higher rooting rate compared with other culture conditions.

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