Investigation of insecticidal activity of rye α-amylase inhibitor gene expressed in transgenic tobacco (Nicotiana tabacum) toward cotton boll weevil (Anthonomus grandis)

Innumerable proteinaceous α-amylase inhibitors have been isolated and identified from different plant species. Among them, an α-amylase inhibitor gene with bioinsecticidal potential toward Anthonomus grandis (cotton boll weevil) was previously identified in rye seeds (Secale cereale). This cereal inhibitor was expressed in tobacco plants (Nicotiana tabacum) under control of phytohemaglutinin promoter by using Agrobacterium tumefasciens - mediated transformation. Presence of αBIII-rye gene and further protein expression were confirmed by PCR and Western blot analysis, respectively. Immunological assays indicated that the recombinant inhibitor was expressed in concentration range from 0.1% to 0.28% (w:w) of the total protein in tobacco seeds of R₀ plants. From 14 independent transformants, five plants with expression levels between 0.20% and 0.28% in seeds were in vitro assayed against A. grandis amylolytic enzymes causing clear inhibition. Moreover, bioassays using transgenic seed flour mixture for artificial diet produced 74% mortality in A. grandis first larval instar. These data suggest that rye inhibitor could be a promising biotechnological tool for produce transgenic cotton plants with an increased resistance to cotton boll weevil. Moreover, αBIII-rye gene should be considered a potential compound for a pyramiding strategy aiming to delay insect-resistance.     

Resistance to <Emphasis Type="Italic">Meloidogyne incognita</Emphasis> expresses a hypersensitive-like response in <Emphasis Type="Italic">Coffea arabica</Emphasis>

Root-knot nematodes (RKN) are obligate parasite species of the genus Meloidogyne that cause great losses in Arabica coffee (Coffea arabica L.) plantations. Identification of resistant genotypes would facilitate the improvement of coffee varieties aiming at an environmental friendly and costless nematode control. In this work, the C. arabica genotype ‘UFV 408-28’ was found to be resistant to the most destructive RKN species M. incognita. Pathogenicity assays indicated that the highly aggressive populations of M. incognita races 1, 2 and 3 were not able to successfully reproduce on ‘UFV 408-28’ roots and displayed a low gall index (GI = 2). An average reduction of 87% reduction of the M. incognita population was observed on ‘UFV 408-28’ when compared to the susceptible cultivar ‘IAC 15’. By contrast, ‘UFV 408-28’ was susceptible to the related species M. exigua and M. paranaensis (GI = 5 and 4, respectively). Histological observations performed on sections of UFV408-28 roots infected with M. incognita race 1 showed that nematode infection could be blocked right after penetration or during migration and establishment stages, at 6 days, 7 days and 8 days after infection (DAI). Fluorescence and bright field microscopy observations showed that root cells surrounding the nematodes exhibited HR-like features such as accumulation of phenolic compounds and a necrotic cell aspect. In the susceptible ‘IAC 15’ roots, 6 DAI, feeding sites contained giant cells with a dense cytoplasm. Necrotic cells were never observed throughout the entire infection cycle. The HR-like phenotype observed in the ‘UFV 408-28’—M. incognita interaction suggests that the coffee resistance may be mediated by a R-gene based immunity system and may therefore provide new insights for understanding the molecular basis of RKN resistance in perennial crops.     

Fine structure of phloematic trypanosomatid–coconut tree interaction

Phytomonas wilt or Hartrot is a fatal disease of palm (Arecaceae) species including Cocos nucifera (coconut) and is caused by a phloematic trypanosomatid, a promastigote parasite that inhabits phloem sieve elements of disease palms. In the present work, we described the morphology of the interaction between a phloematic trypanosomatid (Phytomonas staheli) and C. nucifera. Two varieties and one ecotype of the adult coconut palm from northeast and southeast Brazil were analyzed, totaling 34,000 plants. Coconut palm losses due to Hartrot varied according to the variety or ecotype and geographic area. Occurrence of Hartrot was insignificant in Rio de Janeiro state (southeast), but in Bahia state (northeast) losses were substantial when appropriate cultural practices were not applied. Symptomatic and healthy palm tissues were analyzed by light and electron microscopy. Laboratory diagnoses revealed the twisted promastigote form of phloematic trypanosomatids in the extracts of shoot apex, leaves, stems and inflorescence in diseased plants, but not in the healthy ones. No parasites were found in the roots. Although the general anatomy of healthy and diseased palms was similar, callose deposition in the sieve plates was revealed by histochemistry and immunocytochemistry in the diseased tissue. Plugging by the P-protein and plastid alterations was also observed. Our observations strongly suggest that parasite traffic between sieve elements took place, although their cell bodies were larger than the sieve pores. Phloematic trypanosomatid proliferation in the sieve tube elements might interrupt the transport of phloem or/and consume plant nutrients. In addition, an association between the percentage of sieve elements colonized by pathogen in palm tissues and disease severity was established.     

Potential for increasing the zinc content in common bean using genetic improvement

A large proportion of the world’s population is at risk of developing symptoms of zinc deficiency due to its low levels in food. The objective of this research was to estimate the heritability in early hybrid generations for the zinc content character in seeds of common bean (Phaseolus vulgaris L.), to investigate possible maternal effects on zinc contents and to evaluate the potential for genetic improvement. Reciprocal F₁, and F₂ generations as well as backcross (RCP₁ and RCP₂) populations were produced by crosses of three cultivars. The seed zinc content was measured by the nitric-perchloric digestion method and using the atomic absorption spectrophotometer. The range of variation in zinc content among tested progenies was from 21.76 mg kg⁻¹ dry matter to 53.48 mg kg⁻¹ dry matter, and no significant maternal effect was discovered. Narrow sense heritability was of an intermediate 57.5% to high 77.84% value and transgressive segregation was observed. These results suggest that breeding for increased zinc content in the seeds of common bean can be quite successful; an increase of 37.3% was obtained using just the three parents tested in this study.     

甘薯茎线虫乙酰胆碱酯酶基因Dd-ace-1全长cDNA的克隆和序列分析

利用反转录聚合酶链式反应,结合快速扩增cDNA末端(RACE)技术,克隆了甘薯茎线虫乙酰胆碱酯酶基因cDNA Dd-ace-1.该cDNA序列5′端存在反式剪接引导序列SL1,全长2196bp,包含1个1908bp的开放阅读框(GenBank登录号EF583059);在推导出的635个氨基酸残基的前体蛋白中,N端的前24个氨基酸残基为信号肽,随后的611个氨基酸残基是成熟的乙酰胆碱酯酶序列,其预测的相对分子质量为70144.12.在一级结构中,形成催化活性中心的3个氨基酸残基(Ser223、Glu360和His483)、胆碱结合位点Trp(106)以及在亚基内形成二硫键的6个半胱氨酸完全保守;在电鳐(Torpedo californica)乙酰胆碱酯酶分子的催化功能域中存在14个保守的芳香族氨基酸残基,其中10个在甘薯茎线虫乙酰胆碱酯酶中完全保守.该酶的氨基酸序列与南方根节线虫(Meloidogyne incognita)、胎生网尾线虫(Dictyocaulus viviparous)和秀丽小杆线虫(Caenorhabditis elegans)ACE-1的同源性为68.6%、67.2%和66.3%.与其他线虫乙酰胆碱酯酶的聚类分析显示,该乙酰胆碱酯酶(Dd-ACE-1)与其他线虫乙酰胆碱酯酶ACE-1同属一个支系.     

Co-existing squamous cell carcinoma and hemangioma on the ocular surface of a cat

A 14-year-old spayed female domestic short-haired cat was presented for evaluation of a mass in the right eye. Ophthalmic examination revealed a blind right eye and presence of two distinct masses: a pink and a red-to-brown mass, the latter occupying most of the cornea and part of the conjunctiva. Exenteration was performed under general anesthesia, and the ocular tissues were processed routinely for histopathology. Upon microscopic examination, a malignant epithelial neoplasm and a benign vascular neoplasm were present in the cornea. The conjunctiva and the third eyelid were also affected. Upon immunohistochemistry, the epithelial tumor was positive for cytokeratin and negative for vimentin and the endothelial tumor was negative for cytokeratin and positive for vimentin. A diagnosis of squamous cell carcinoma (SCC) and hemangioma was made. The SCC was affecting the cornea, bulbar conjunctiva (lateral and inferior) and the base of the third eyelid, whereas the hemangioma was affecting the cornea and medial limbus. To the authors' knowledge, this is the first report of concomitant SCC and hemangioma affecting the ocular surface in a cat.